QUATERNARY AMMONIUM SALTS AS INHIBITORS OF TRIMETHYLAMINE PRODUCTION

Abstract

Provided are compounds that can inhibit pathogenic, bacterial metabolite production and conjugates of the same. Also provided are pharmaceutical compositions comprising the same and methods of using the same.

Claims

1. A compound consisting of a cation and a pharmaceutically acceptable counterion, wherein the cation is chosen from: ##STR00375## ##STR00376## ##STR00377## ##STR00378## ##STR00379## ##STR00380## ##STR00381## ##STR00382## ##STR00383## ##STR00384## ##STR00385## ##STR00386## ##STR00387## ##STR00388## ##STR00389## ##STR00390## ##STR00391## ##STR00392## ##STR00393## ##STR00394## ##STR00395## ##STR00396## ##STR00397##

2. The compound according to claim 1, wherein the cation is chosen from: ##STR00398## ##STR00399## ##STR00400##

3. A compound consisting of a cation and a pharmaceutically acceptable counterion, wherein the cation is chosen from: ##STR00401## ##STR00402## ##STR00403## ##STR00404## ##STR00405## ##STR00406## ##STR00407## ##STR00408## ##STR00409## ##STR00410## ##STR00411##

4. A compound consisting of a cation and a pharmaceutically acceptable counterion, wherein the cation is a structure of formula (I): ##STR00412## wherein R.sup.1 is C.sub.2-6 alkyl substituted with —O-(acylated sugar) or isosorbide, wherein said C.sub.2-6 alkyl is further optionally substituted with oxo and/or methene; C.sub.4 cycloalkyl optionally substituted with hydroxyl, ethynyl, or —O-(acylated sugar); or C.sub.3 cycloalkyl substituted with C.sub.1-6 alkyl, hydroxyl, ethynyl, or —O-(acylated sugar) C.sub.3-4 cycloalkyl C.sub.1-2 alkyl; R.sup.2 is C.sub.2-6 alkyl optionally substituted with one or two hydroxyl, oxo, and —O-(acylated sugar); or R.sup.1 and R.sup.2, together with the nitrogen atom to which both are attached, combine to form a 4- or 5-membered heterocyclic ring optionally substituted with ethynyl or —(CH.sub.2).sub.n—OR.sup.5 or an acylated sugar, wherein n is 0 or 1, R.sup.5 is hydrogen or an acylated sugar; R.sup.3 is C.sub.1-6 alkyl optionally substituted with a halogen or hydroxyl; and R.sup.4 is C.sub.1-6 alkyl or propargyl.

5. The compound of claim 4, wherein R.sup.1 is C.sub.2-6 alkyl substituted with —O-(acylated sugar) and is optionally further substituted with oxo.

6. The compound of claim 4 or 5, wherein R.sup.2 is methyl.

7. The compound of any one of claims 4, 5, and 6, wherein R.sup.3 is C.sub.1-6 alkyl.

8. The compound of any one of claims 4-7, wherein R.sup.4 is propargyl.

9. The compound of claim 4, wherein R.sup.1 is C.sub.2-6 alkyl substituted with —O-(acylated sugar) and is optionally further substituted with oxo; R.sup.2 is methyl; R.sup.3 is C.sub.1-6 alkyl; and R.sup.4 is propargyl.

10. The compound of claim 4, wherein R.sup.1 is C.sub.2-6 alkyl substituted with isosorbide and is optionally further substituted with oxo and/or methene; R.sup.2 is methyl; R.sup.3 is C.sub.1-6 alkyl; and R.sup.4 is propargyl.

11. The compound of claim 4, wherein R.sup.1 is chosen from C.sub.2 alkyl substituted with —O-(acylated sugar) and optionally further substituted with oxo and C.sub.4 alkyl substituted with —O-(acylated sugar) and optionally further substituted with oxo; wherein the acylated sugar is chosen from groups of formula (A), groups of formula (B), and groups of formula (C): ##STR00413## wherein each R.sup.A is independently H or fatty acid acyl; and R.sup.B is H, —CH.sub.3, —CH.sub.2—OR.sup.A, —OCH.sub.3, —COOCH.sub.3, or —COOH; R.sup.2 is methyl; R.sup.3 is methyl; and R.sup.4 is propargyl.

12. The compound of claim 4, wherein R.sup.1 is chosen from C.sub.2 alkyl substituted with isosorbide and optionally further substituted with oxo and C.sub.3 alkyl substituted with isosorbide and optionally further substituted with oxo and methene; wherein isosorbide is chosen from groups of formula (C): ##STR00414## R.sup.2 is methyl; R.sup.3 is methyl; and R.sup.4 is propargyl.

13. The compound of claim 4, wherein R.sup.1 is C.sub.3-4 cycloalkyl C.sub.1-2 alkyl; R.sup.2 is C.sub.2-6 alkyl optionally substituted with one or two hydroxyl, oxo, and —O-(acylated sugar); or R.sup.1 and R.sup.2, together with the nitrogen atom to which both are attached, combine to form a 4- or 5-membered heterocyclic ring optionally substituted with ethynyl or —(CH.sub.2).sub.n—OR.sup.s or an acylated sugar, wherein n is 0 or 1, R.sup.s is hydrogen or an acylated sugar; R.sup.3 is C.sub.1-6 alkyl optionally substituted with a halogen or hydroxyl; and R.sup.4 is C.sub.1-6 alkyl or propargyl.

14. The compound of claim 13, wherein R.sup.1 is C.sub.3-4 cycloalkyl C.sub.1 alkyl.

15. The compound of claim 13 or 14, wherein R.sup.2 is C.sub.2 alkyl optionally substituted with one or two hydroxyl groups.

16. The compound of claim 13, wherein R.sup.1 and R.sup.2, together with the nitrogen atom to which both are attached, combine to form a 5-membered heterocyclic ring optionally substituted with ethynyl, —OH, or —CH.sub.2OH.

17. The compound of any one of claims 13-16, wherein R.sup.3 is C.sub.1 alkyl optionally substituted with a halogen.

18. The compound of any one of claims 13-17, wherein R.sup.4 is C.sub.1 alkyl or propargyl.

19. A compound consisting of a cation and a pharmaceutically acceptable counterion, wherein the cation is a structure of formula (II): ##STR00415## wherein R.sup.1 is C.sub.1-6 alkyl optionally substituted with one or more oxo, hydroxyl, halogen, cyano, —COOMe, amino, methene, ethenyl, ethynyl, hydroxyphenyl, C.sub.3-4 cycloalkyl, —OCH.sub.2CH.sub.2OH, —HNC(O)OCMe.sub.3, —SMe, —OMe, —HNS(O).sub.2Me, —SO.sub.3H, B(OH).sub.2, PO.sub.3H.sub.2, PO.sub.2H.sub.2, —P(O)(OCH.sub.2CH.sub.3).sub.2, —P(O)(OH)(OCH.sub.2CH.sub.3), heteroaryl ring; phenyl; benzyl; C.sub.3-4 heterocyclyl optionally substituted with C.sub.1 alkyl C.sub.4 heterocyclylalkyl C.sub.1 alkyl; or C.sub.4 cycloalkyl; R.sup.2 is C.sub.1-6 alkyl optionally substituted with one or two hydroxyl, oxo, methene, cyano, ethynyl, —HNC(O)H, or —C≡C—CH.sub.2OH; or —HN(CH.sub.2).sub.3C(O)OH; or R.sup.1 and R.sup.2, together with the nitrogen atom to which both are attached and optionally one or more additional heteroatoms, combine to form a 4-8 membered mono- or bi-cyclic heterocycle optionally substituted with ethynyl, trifluoromethyl, —CH.sub.2Ph, —OH, or —(CH.sub.2)OH; R.sup.3 is methyl or propargyl; and R.sup.4 is methyl or propargyl, with the proviso that (1) when two of R.sup.1, R.sup.2, R.sup.3, and R.sup.4 are methyl and one of the other two of R.sup.1, R.sup.2, R.sup.3, and R.sup.4 is propargyl, then the other of R.sup.1, R.sup.2, R.sup.3, and R.sup.4 is not methyl, a monohalomethyl, —CH.sub.2CH.sub.2OH, —COOH, —(CH.sub.2).sub.4C(O)OH, or —CH.sub.2C(CH.sub.2)CN; and (2) when two of R.sup.1, R.sup.2, R.sup.3, and R.sup.4 are methyl and one of the other two of R.sup.1, R.sup.2, R.sup.3, and R.sup.4 is —CH.sub.2CH.sub.2OH or —CH.sub.2C(O)OH, then the other of R.sup.1, R.sup.2, R.sup.3, and R.sup.4 is not a monohalomethyl.

20. A compound consisting of a cation and a pharmaceutically acceptable counterion, wherein the cation is a structure of formula (III): ##STR00416## wherein R.sup.1 is C.sub.1-6 alkyl optionally substituted with one or more oxo, hydroxyl, halogen, cyano, —COOMe, —COOH, methene, ethenyl, ethynyl, C.sub.3-4 cycloalkyl, —OCH.sub.2CH.sub.2OH, —SMe, —OMe, —HNS(O).sub.2Me, —P(O)(OCH.sub.2CH.sub.3).sub.2, —P(O)(OH)(OCH.sub.2CH.sub.3), or 5-membered heteroaryl ring; R.sup.2 is C.sub.1-6 alkyl optionally substituted with one or two hydroxyl, oxo, methene, cyano, ethynyl, —HNC(O)H, or —C≡C—CH.sub.2OH; or —HN(CH.sub.2).sub.3C(O)OH; or R.sup.1 and R.sup.2, together with the nitrogen atom to which both are attached and optionally one or more additional heteroatoms, combine to form a 4-8 membered mono- or bi-cyclic heterocycle optionally substituted with ethynyl, —OH, or —(CH.sub.2)OH; R.sup.3 is C.sub.1-6 alkyl optionally substituted with a halogen, hydroxyl, or ethynyl; and R.sup.4 is C.sub.1-6 alkyl or propargyl; with the proviso that (1) when two of R.sup.1, R.sup.2, R.sup.3, and R.sup.4 are methyl and one of the other two of R.sup.1, R.sup.2, R.sup.3, and R.sup.4 is propargyl, then the other of R.sup.1, R.sup.2, R.sup.3, and R.sup.4 is not methyl, a monohalomethyl, —CH.sub.2CH.sub.2OH, —COOH, —(CH.sub.2).sub.4C(O)OH, or —CH.sub.2C(CH.sub.2)CN; and (2) when two of R.sup.1, R.sup.2, R.sup.3, and R.sup.4 are methyl and one of the other two of R.sup.1, R.sup.2, R.sup.3, and R.sup.4 is —CH.sub.2CH.sub.2OH or —CH.sub.2C(O)OH, then the other of R.sup.1, R.sup.2, R.sup.3, and R.sup.4 is not a monohalomethyl.

21. A pharmaceutical composition comprising a pharmaceutically acceptable excipient and the compound of any one of claims 1 to 20.

22. A method of modulating a trimethylaminuria marker in a subject in need thereof, the method comprising administering to the subject in need thereof a therapeutically effective amount of the compound of any one of claims 1 to 20 or the pharmaceutical composition of claim 21.

23. The method of claim 22, wherein the trimethylaminuria marker is the trimethylamine and/or trimethylamine oxide levels in the subject's blood, plasma, serum, or urine.

24. A method of treating or preventing a disease associated with elevated levels of trimethylamine (TMA), a decreased rate of conversion of TMA to trimethylamine oxide (TMAO), or a high ratio of TMA to TMAO in a subject in need thereof, the method comprising administering a therapeutically effective amount of the compound of any one of claims 1 to 20 or the pharmaceutical composition of claim 21 to the subject.

25. The method of claim 24, further comprising detecting the presence of one or more genetic variants of the FMO3 gene of the subject in need before the administering step.

26. The method of claim 24, wherein the disease associated with elevated levels of trimethylamine (TMA) or trimethylamine N-oxide (TMAO), a decreased rate of conversion of TMA to TMAO, or a high ratio of TMA to TMAO is a cardiovascular disease, reduced or impaired kidney function, kidney disease, chronic kidney disease, end-stage renal disease, or diabetes mellitus.

27. A method of inhibiting a CutC choline lyase-mediated conversion of choline to trimethylamine, the method comprising contacting the compound of any one of claims 1 to 20 with the CutC choline lyase.

28. A method of inhibiting a CntA carnitine monooxygenase-mediated conversion of carnitine to trimethylamine, the method comprising contacting the compound of any one of claims 1 to 20 with the CntA carnitine monooxygenase.

29. A method of treating a subject in need of treatment for trimethylaminuria comprising contacting bacteria in vivo with a therapeutically effective amount of the compound of any one of claims 1 to 20 or the pharmaceutical composition of claim 21 to the subject.

30. The method of claim 29, wherein the bacteria are localized in the colon of the subject.

31. A method of identifying a subject suffering from trimethylaminuria, or predicting a predisposition for developing trimethylaminura in a subject, comprising: (i) analyzing a sample from the patient to detect the presence of at least one FMO3 genetic variant in the patient, and (ii) identifying a subject suffering from trimethylaminuria or predicting a predisposition for developing trimethylaminura in a subject.

32. The method of claim 31, further comprising (iii) administering a therapeutically effective amount of the compound of any one of claims 1 to 20 or the pharmaceutical composition of claim 21.

33. The method of claim 31 or 32, wherein the FMO3 genetic variant is chosen from g.-2092 to 10145del, g.94G>A+A29A2:A30, g.110T>C, g.11145A>G, g.11148G>T, g.11166G>A, g.11177A>G, g.11185delA, g.11192G>T, g.11239T>C, g.15036A>G, g.15123T>A, g.15137G>T, g.15153C>T, g.15526_15527delTG, g.15531T>A, g.15533T>C, g.15539C>A, g.18177G>A, g.18225G>C, g.21429G>T, g.21460G>T, g.21680G>T, g.21684G>A, g.21702delG, g.23580delG, g.24486G>A, g.24592C>T, g.24608G>A, g.24658C>T, and g.24682C>T.

Description

EXAMPLES

Example 1: Preparation of Exemplary Compounds

[0343] In the following examples, the title compounds may exist as a trifluoroacetate salt after prep-HPLC but one of ordinary skill in the art would understand that the salt may be changed by conventional methods of salt formation.

##STR00110##

Compound 1: N,N-dimethyl-N-(2-(((2R,3R,4S,5R,6R)-3,4,5-tris(butyryloxy)-6-((butyryloxy)methyl)tetrahydro-2H-pyran-2-yl)oxy)ethyl)prop-2-yn-1-aminium iodide

[0344] Step 1:

[0345] A mixture of (3R,4S,5S,6R)-6-(hydroxymethyl)tetrahydropyran-2,3,4,5-tetrol (20 g, 111.01 mmol, 1 eq) and butanoyl butanoate (96.59 g, 610.58 mmol, 99.89 mL, 5.5 eq) in Pyridine (200 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 20° C. for 12 h. TLC indicated the starting material was consumed completely and one new spot formed. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=1 to 20:1). [(2R,3R,4S,5R)-3,4,5,6-tetra(butanoyloxy)tetrahydropyran-2-yl]methyl butanoate (100 g, crude) was obtained as colorless oil.

[0346] Step 2:

[0347] To a mixture of [(2R,3R,4S,5R)-3,4,5,6-tetra(butanoyloxy)tetrahydropyran-2-yl]methyl butanoate (10 g, 18.85 mmol, 1 eq) in THF (100 mL) was added MeNH.sub.2 aq. solution (2.4 g, 30.91 mmol, 40% purity, 1.64 eq) in one portion at 20° C. under N2. The mixture was stirred at 20° C. for 12 h. TLC indicated [(2R,3R,4S,5R)-3,4,5,6-tetra(butanoyloxy)tetrahydropyran-2-yl]methyl butanoate was consumed completely and one new spot formed. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=1/0 to 20:1). Compound [(2R,3R,4S,5R)-3,4,5-tri(butanoyloxy)-6-hydroxy-tetrahydropyran-2-yl]methyl butanoate (5 g, 10.86 mmol, 57.6% yield) was obtained as colorless oil.

[0348] Step 3:

[0349] To a mixture of [(2R,3R,4S,5R)-3,4,5-tri(butanoyloxy)-6-hydroxy-tetrahydropyran-2-yl]methyl butanoate (2 g, 4.34 mmol, 1 eq) and DBU (330.58 mg, 2.17 mmol, 327.31 uL, 0.5 eq) in DCM (20 mL) was added 2,2,2-trichloroacetonitrile (6.27 g, 43.43 mmol, 4.35 mL, 10 eq) in one portion at 20° C. The mixture was stirred at 20° C. for 12 h. TLC showed [(2R,3R,4S,5R)-3,4,5-tri(butanoyloxy)-6-hydroxy-tetrahydropyran-2-yl]methyl butanoate was consumed completely and one new spot formed. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=30/1 to 20:1). Compound [(2R,3R,4S,5R)-3,4,5-tri(butanoyloxy)-6-(2,2,2-trichloroethanimidoyl)oxy-tetrahydropyran-2-yl]methyl butanoate (1.8 g, 2.98 mmol, 68.5% yield) was obtained as colorless oil.

[0350] Step 4:

[0351] To a solution of [(2R,3R,4S,5R)-3,4,5-tri(butanoyloxy)-6-(2,2,2-trichloroethanimidoyl) oxy-tetrahydropyran-2-yl]methyl butanoate (1 g, 1.65 mmol, 1 eq) and 2-bromoethanol (247.90 mg, 1.98 mmol, 140.85 uL, 1.2 eq) in DCM (10 mL) was added BF.sub.3.Et.sub.2O (1.17 g, 8.27 mmol, 1.02 mL, 5 eq). The mixture was stirred at 0° C. for 3 h and then warmed to 20° C. stirred for 12 h. TLC indicated [(2R,3R,4S,5R)-3,4,5-tri(butanoyloxy)-6-(2,2,2-trichloroethanimidoyl)oxy-tetrahydropyran-2-yl]methyl butanoate was consumed completely. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=30/1 to 20:1). Compound [(2R,3R,4S,5R)-6-(2-bromoethoxy)-3,4,5-tri(butanoyloxy)tetrahydropyran-2-yl]methyl butanoate (0.8 g, 1.41 mmol, 85.3% yield) was obtained as colorless oil.

[0352] Step 5:

[0353] A mixture of [(2R,3R,4S,5R)-6-(2-bromoethoxy)-3,4,5-tri(butanoyloxy)tetrahydropyran-2-yl] methyl butanoate (0.1 g, 176.22 umol, 1 eq), N-methylprop-2-yn-1-amine (36.53 mg, 528.67 umol, 44.02 uL, 3 eq) in THF (3 mL) was degassed and purged with N.sub.2 for 3 times, and then the mixture was stirred at 20° C. for 2 hr under N.sub.2 atmosphere, then warmed to 70° C. and stirred for 12 h. TLC indicated [(2R,3R,4S,5R)-6-(2-bromoethoxy)-3,4,5-tri(butanoyloxy)tetrahydropyran-2-yl]methyl butanoate was consumed completely and one new spot formed. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by prep-TLC (SiO.sub.2, Petroleum ether/Ethyl acetate=2:1). Compound [(2R,3R,4S,5R)-3,4,5-tri(butanoyloxy)-6-[2-[methyl(prop-2-ynyl)amino]ethoxy]tetrahydropyran-2-yl]methyl butanoate (0.03 g, 54 umol, 30.6% yield) was obtained as colorless oil.

[0354] Step 6:

[0355] A mixture of [(2R,3R,4S,5R)-3,4,5-tri(butanoyloxy)-6-[2-[methyl(prop-2-ynyl)amino]ethoxy] tetrahydropyran-2-yl]methyl butanoate (0.06 g, 107.98 umol, 1 eq), MeI (30.65 mg, 215.96 umol, 13.44 uL, 2 eq) in THF (3 mL) was degassed and purged with N.sub.2 for 3 times, and then the mixture was stirred at 20° C. for 2 hr under N.sub.2 atmosphere. LC-MS showed [(2R,3R,4S,5R)-3,4,5-tri(butanoyloxy)-6-[2-[methyl(prop-2-ynyl)amino]ethoxy]tetrahydropyran-2-yl]methyl butanoate was consumed. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC [water (0.1% TFA)-ACN]. The title compound (0.007 g, 12.1 umol, 11% yield) was obtained as colorless oil. LCMS (M.sup.+): 570.3 .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 5.33 (t, J=9.5 Hz, 1H), 5.04-4.81 (m, 3H), 4.37 (d, J=2.5 Hz, 2H), 4.22-4.00 (m, 6H), 3.62 (t, J=4.9 Hz, 2H), 3.09 (s, 6H), 2.33-2.11 (m, 8H), 1.61-1.38 (m, 8H), 0.93-0.78 (m, 12H

##STR00111##

Compound 2 and Compound 51: N,N-dimethyl-N-(2-(((3R,4S,5R)-3,4,5-tris(butyryloxy)tetrahydro-2H-pyran-2-yl)oxy)ethyl)prop-2-yn-1-aminium Iodide

[0356] This compound may be synthesized according to the experimental procedure described for Compound 1.

##STR00112##

Compound 3: N,N-dimethyl-N-(2-(((3R,4R,5R)-3,4,5-tris(butyryloxy)tetrahydro-2H-pyran-2-yl)oxy)ethyl)prop-2-yn-1-aminium Iodide

[0357] This compound may be synthesized according to the experimental procedure described for Compound 1.

##STR00113##

Compound 4 and Compound 53 and Compound 84: N,N-dimethyl-N-(2-(((3R,4S,5S)-3,4,5-tris(butyryloxy)tetrahydro-2H-pyran-2-yl)oxy)ethyl)prop-2-yn-1-aminium iodide

[0358] This compound may be synthesized according to the experimental procedure described for Compound 1.

##STR00114##

Compound 5: N,N-dimethyl-N-(2-(((3R,4R,5S,6S)-3,4,5-tris(butyryloxy)-6-methyltetrahydro-2H-pyran-2-yl)oxy)ethyl)prop-2-yn-1-aminium Iodide

[0359] This compound may be synthesized according to the experimental procedure described for Compound 1.

##STR00115##

Compound 6: N,N-dimethyl-N-(2-(((3S,4R,5R,6S)-3,4,5-tris(butyryloxy)-6-methyltetrahydro-2H-pyran-2-yl)oxy)ethyl)prop-2-yn-1-aminium Iodide

[0360] This compound may be synthesized according to the experimental procedure described for Compound 1.

##STR00116##

Compound 7: N,N-dimethyl-N-(2-(((2R,3R,4S,5R,6R)-3,4,5-triacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)ethyl)prop-2-yn-1-aminium Iodide

[0361] Step 1:

[0362] Compound β-D-Glucose pentaacetate (20 g, 51.2 mmol, 1 eq) was dissolved in DCM (100 mL) and cooled to 0° C. under N.sub.2, followed by addition of 2-bromoethanol (5.5 mL, 77.6 mmol, 1.5 eq) and boron trifluoride diethyl etherate (32 mL, 259 mmol, 5 eq). The ice bath was removed and the reaction was stirred at room temperature overnight. Silica was added to the reaction mixture and solution was concentrated to dryness to dry-load onto silica before purification by column chromatography (gradient: 0-100% EtOAc in hexanes, product eluted around 40-50% EtOAc in hexanes). Fractions containing product were concentrated and recrystallized by dissolution in ethyl acetate, followed by slow addition of hexanes until crystals started to form, at which point it was placed in the fridge for further crystallization. The white crystals were filtered and washed with hexanes to yield 1-(2-bromoethoxy)-2,3,4,6-tetra-O-acetyl-beta-D-glucopyranose (8.9 g, 19.5 mmol, 38%).

[0363] Step 2:

[0364] Compound 1-(2-bromoethoxy)-2,3,4,6-tetra-O-acetyl-beta-D-glucopyranose (1.7 g, 3.7 mmol, 1 eq) was stirred with sodium iodide (0.614 g, 4.1 mmol, 1.1 eq) for a few minutes, followed by addition of 3-dimethylamino-1-propyne (1.2 mL, 11.1 mmol, 3 eq). The reaction mixture was stirred overnight at 40° C. filtered, then cooled to 4° C. in the fridge for recrystallization. The crystals were washed with cold acetone to yield the title compound as a white crystalline solid (300 mg, 0.51 mmol, 14%, iodide salt-confirmed by ion chromatography). The remaining filtrate was concentrated and washed with cold acetone and ethyl acetate to yield the title compound as an amorphous powder (1.2 g, 2.05 mmol, 55%, iodide salt-confirmed by ion chromatography). LCMS (M+): 458.1 .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 5.26 (t, J=9.5 Hz, 1H), 4.98-4.77 (m, 3H), 4.36 (d, J=2.5 Hz, 2H), 4.24-3.97 (m, 6H), 3.67-3.54 (m, 2H), 3.08 (s, 6H), 2.05-1.89 (m, 12H).

##STR00117##

Compound 8: N,N-dimethyl-N-(2-(((3R,4S,5R)-3,4,5-triacetoxytetrahydro-2H-pyran-2-yl)oxy)ethyl)prop-2-yn-1-aminium iodideiodideiodideiodideiodide

[0365] Step 1:

[0366] To a solution of (3R,4S,5R)-tetrahydropyran-2,3,4,5-tetrol (20 g, 133.22 mmol, 1 eq) in Pyridine (200 mL) was added acetyl acetate (108.80 g, 1.07 mol, 99.82 mL, 8 eq) at 15° C. The mixture was stirred at 15° C. for 12 hr. TLC indicated new spots formed. The reaction mixture was concentrated under reduced pressure to give yellow oil. The oil was diluted with ethyl acetate 100 mL and washed with H.sub.2O (100 mL*3). The combined organic layers were washed with brine 100 mL, dried over anhydrous sodium sulfate, filtered and the filtrate was concentrated under reduced pressure to give yellow oil. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=1:0). Compound [(3R,4S,5R)-4,5,6-triacetoxytetrahydropyran-3-yl] acetate (42 g, crude) was obtained as a yellow oil.

[0367] Step 2:

[0368] To a solution of [(3R,4S,5R)-4,5,6-triacetoxytetrahydropyran-3-yl] acetate (21 g, 65.98 mmol, 1 eq) in THF (120 mL) was added aq. MeNH.sub.2 (7.51 g, 72.58 mmol, 30% purity, 1.1 eq) at 15° C. The mixture was stirred at 15° C. for 12 hr. TLC indicated new spots formed. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=1:1 to 0:1). Compound [(3R,4S,5R)-4,5-diacetoxy-6-hydroxy-tetrahydropyran-3-yl] acetate (9 g, 32.58 mmol, 49.38% yield) was obtained as a yellow solid.

[0369] Step 3:

[0370] To a solution of [(3R,4S,5R)-4,5-diacetoxy-6-hydroxy-tetrahydropyran-3-yl]acetate (5 g, 18.10 mmol, 1 eq) in DCM (20 mL) was added DBU (1.38 g, 9.05 mmol, 1.36 mL, 0.5 eq) and 2,2,2-trichloroacetonitrile (26.13 g, 181.00 mmol, 18.15 mL, 10 eq) at 15° C. The mixture was stirred at 15° C. for 1 hr. TLC indicated a new spot formed. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=1:1). Compound [(3R,4S,5R)-4,5-diacetoxy-6-(2,2,2-trichloroethanimidoyl)oxy-tetrahydropyran-3-yl] acetate (6 g, 14.26 mmol, 78.81% yield) was obtained as a yellow oil.

[0371] Step 4:

[0372] To a solution of [(3R,4S,5R)-4,5-diacetoxy-6-(2,2,2-trichloroethanimidoyl)oxy-tetrahydropyran-3-yl] acetate (3 g, 7.13 mmol, 1 eq) and 2-bromoethanol (891.27 mg, 7.13 mmol, 506.41 uL, 1 eq) in DCM (30 mL) was added BF.sub.3.Et.sub.2O (5.06 g, 35.66 mmol, 4.40 mL, 5 eq) at 0° C. The mixture was stirred at 0° C. for 2 hr. TLC indicated new spots formed. The reaction mixture was quenched by addition H.sub.2O (30 mL) at 0° C., and extracted with ethyl acetate (50 mL*3). The combined organic layers were washed with brine 30 mL, dried over anhydrous sodium sulfate, filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=10:1 to 1:1). Compound [(3R,4S,5R)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate (1.3 g, 3.39 mmol, 47.57% yield) was obtained as a white solid.

[0373] Step 5:

[0374] To a solution of [(3R,4S,5R)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate (1.3 g, 3.39 mmol, 1 eq) in acetone (20 mL) was added N,N-dimethylprop-2-yn-1-amine (564.06 mg, 6.79 mmol, 719.46 uL, 2 eq) and NaI (1.02 g, 6.79 mmol, 2 eq) at 15° C. The mixture was stirred at 80° C. for 4 hr. LCMS showed the desired compound was detected. TLC indicated 50% of [(3R,4S,5R)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate (1.3 g, 3.39 mmol, 1 eq) remained. The mixture was stirred at 80° C. for 8 hr. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=1:1 to Ethyl acetate:Methanol=1:1) The title compound (530 mg, 1.03 mmol, 30.30% yield) was obtained as a white solid. LCMS: (M+H.sup.+): 386.1 .sup.1H NMR (400 MHz, Deuterium Oxide) δ 5.10 (t, J=7.6 Hz, 1H), 4.93-4.79 (m, 2H), 4.71 (d, J=6.1 Hz, 1H), 4.20-4.14 (m, 3H), 4.06 (dd, J=12.3, 4.7 Hz, 1H), 3.98-3.90 (m, 1H), 3.70-3.59 (m, 2H), 3.50 (dd, J=12.3, 7.9 Hz, 1H), 3.19-3.13 (m, 1H), 3.11 (s, 6H), 2.02-1.92 (m, 9H).

##STR00118##

Compound 9: N,N-dimethyl-N-(2-(((3R,4R,5R)-3,4,5-triacetoxytetrahydro-2H-pyran-2-yl)oxy)ethyl)prop-2-yn-1-aminium bromide

[0375] This compound may be synthesized according to the experimental procedure described for Compound 7.

##STR00119##

Compound 10 and Compound 56: N,N-dimethyl-N-(2-(((3R,4S,5S)-3,4,5-triacetoxytetrahydro-2H-pyran-2-yl)oxy)ethyl)prop-2-yn-1-aminium Iodide

[0376] Step 1:

[0377] To a mixture of (2R,3S,4S)-2,3,4,5-tetrahydroxypentanal (10 g, 66.61 mmol, 1 eq) in pyridine (100 mL) was added Ac.sub.2O (40.80 g, 399.65 mmol, 37.43 mL, 6 eq) in one portion at 25° C. under N.sub.2. The mixture was stirred at 25° C. for 12 hours. TLC indicated one new spot formed. The reaction mixture was concentrated under reduced pressure to remove pyridine. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=30/1 to 0/1). Compound [(3S,4S,5R,6S)-4,5,6-triacetoxytetrahydropyran-3-yl] acetate (20 g, crude) was obtained as yellow oil.

[0378] Step 2:

[0379] To a mixture of [(3S,4S,5R,6S)-4,5,6-triacetoxytetrahydropyran-3-yl] acetate (10 g, 31.42 mmol, 1 eq) in DCM (100 mL) was added 2-bromoethanol (5.89 g, 47.13 mmol, 3.35 mL, 1.5 eq) and BF.sub.3.Et.sub.2O (22.30 g, 157.10 mmol, 19.39 mL, 5 eq) in one portion at 0° C. under N.sub.2. The mixture was heated to 25° C. and stirred for 12 hours. TLC indicated [(3S,4S,5R,6S)-4,5,6-triacetoxytetrahydropyran-3-yl] acetate was consumed and one new spot formed. The reaction mixture was quenched by addition of H.sub.2O (50 mL), and then diluted with H.sub.2O (100 mL) and extracted with DCM (100 mL*2). The combined organic layers were dried over Na.sub.2SO.sub.4, filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=20/1 to 0/1). Then the residue was purified by prep-HPLC [water (0.10% TFA)-ACN]. .sup.1HNMR indicated the compound [(3S,4S,5R,6S)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate (1.2 g, 3.13 mmol, 9.97% yield) was obtained as yellow oil and compound[(3S,4S,5R,6R)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate (2.3 g, 6.00 mmol, 19.10% yield) was obtained as yellow oil.

[0380] Step 3:

[0381] To a mixture of [(3S,4S,5R,6S)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate (1 g, 2.61 mmol, 1 eq) in acetone (10 mL) was added NaI (430.29 mg, 2.87 mmol, 1.1 eq) in one portion at 25° C. under N.sub.2. Then N,N-dimethylprop-2-yn-1-amine (1.08 g, 13.05 mmol, 1.38 mL, 5 eq) was added. The mixture was heated to 90° C. and stirred for 2 hours. TLC indicated [(3S,4S,5R,6S)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate was consumed and one new spot formed. The reaction mixture was filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=10/1 Ethyl acetate:Methanol=3/1).dimethyl-prop-2-ynyl-[2-[(2R,3R,4S,5S)-3,4,5-triacetoxytetrahydropyran-2-yl]oxyethyl]ammonium (318 mg, 617.64 umol, 23.67% yield, 99.7% purity, I) was obtained as white solid. LCMS: (M+): 386.2 1H NMR (400 MHz, Chloroform-d) δ 5.26-5.20 (m, 1H), 5.08 (dd, J=10.0, 7.4 Hz, 1H), 4.99 (dd, J=10.0, 3.5 Hz, 1H), 4.75 (t, J=3.0 Hz, 2H), 4.48 (d, J=7.4 Hz, 1H), 4.37-4.26 (m, 1H), 4.17-4.06 (m, 2H), 4.09-4.00 (m, 1H), 3.95 (dd, J=13.3, 2.5 Hz, 1H), 3.66 (dd, J=13.4, 1.5 Hz, 1H), 3.46 (s, 6H), 2.82 (t, J=2.5 Hz, 1H), 2.10 (s, 3H), 2.04 (s, 3H), 1.95 (s, 3H)))).).

##STR00120##

Compound 11: N,N-dimethyl-N-(2-(((3R,4R,5S,6S)-3,4,5-triacetoxy-6-methyltetrahydro-2H-pyran-2-yl)oxy)ethyl)prop-2-yn-1-aminium bromide

[0382] This compound may be synthesized according to the experimental procedure described for Compound 7.

##STR00121##

Compound 12: N,N-dimethyl-N-(2-(((3S,4R,5R,6S)-3,4,5-triacetoxy-6-methyltetrahydro-2H-pyran-2-yl)oxy)ethyl)prop-2-yn-1-aminium bromide

[0383] This compound may be synthesized according to the experimental procedure described for Compound 7.

##STR00122##

Compound 13: N,N-dimethyl-N-(2-oxo-2-(((3R,4S,5R,6R)-3,4,5-tris(butyryloxy)-6-((butyryloxy)methyl)tetrahydro-2H-pyran-2-yl)oxy)ethyl)prop-2-yn-1-aminium chloride

[0384] Step 1:

[0385] To a mixture of [(2R,3R,4S,5R)-3,4,5-tri(butanoyloxy)-6-hydroxy-tetrahydropyran-2-yl]methyl butanoate (1 g, 2.17 mmol, 1 eq) and 2-chloroacetyl chloride (735.76 mg, 6.51 mmol, 518.14 uL, 3 eq) in DCM (10 mL) was added pyridine (858.82 mg, 10.86 mmol, 876.35 uL, 5 eq) in one portion at 20° C. The mixture was stirred at 20° C. for 12 h. TLC showed [(2R,3R,4S,5R)-3,4,5-tri(butanoyloxy)-6-hydroxy-tetrahydropyran-2-yl]methyl butanoate was consumed completely and one new spot formed. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=1/0 to 20:1). Compound [(2R,3R,4S,5R)-3,4,5-tri(butanoyloxy)-6-(2-chloroacetyl)oxy-tetrahydropyran-2-yl]methyl butanoate (0.8 g, crude) was obtained as colorless oil.

[0386] Step 2:

[0387] A mixture of [(2R,3R,4S,5R)-3,4,5-tri(butanoyloxy)-6-(2-chloroacetyl)oxy-tetrahydropyran-2-yl] methyl butanoate (0.2 g, 372.44 umol, 1 eq) and N,N-dimethylprop-2-yn-1-amine (92.89 mg, 1.12 mmol, 118.48 uL, 3 eq) in acetone (3 mL) was stirred at 90° C. for 0.5 h. LCMS showed [(2R,3R,4S,5R)-3,4,5-tri(butanoyloxy)-6-(2-chloroacetyl)oxy-tetrahydropyran-2-yl]methyl butanoate was consumed completely. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC [water (0.1% TFA)-ACN]. The title compound (0.049 g, 67 umol, 18% yield) was obtained as colorless oil. LCMS (M.sup.+): 584.3 .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 6.30 (d, J=3.4 Hz, 1H), 5.46-5.35 (m, 1H), 5.25-5.11 (m, 2H), 4.80-4.63 (m, 2H), 4.61-4.46 (m, 2H), 4.30-3.98 (m, 4H), 3.30-3.16 (m, 6H), 2.37-2.10 (m, 8H), 1.63-1.41 (m, 8H), 0.95-0.75 (m, 12H).

##STR00123##

Compound 14 and Compound 52: N,N-dimethyl-N-(2-oxo-2-(((3R,4S,5R)-3,4,5-tris(butyryloxy)tetrahydro-2H-pyran-2-yl)oxy)ethyl)prop-2-yn-1-aminium chloride

[0388] This compound may be synthesized according to the experimental procedure described for Compound 13.

##STR00124##

Compound 15: N,N-dimethyl-N-(2-oxo-2-(((3R,4R,5R)-3,4,5-tris(butyryloxy)tetrahydro-2H-pyran-2-yl)oxy)ethyl)prop-2-yn-1-aminium chloride

[0389] This compound may be synthesized according to the experimental procedure described for Compound 13.

##STR00125##

Compound 16 and Compound 54: N,N-dimethyl-N-(2-oxo-2-(((3R,4S,5S)-3,4,5-tris(butyryloxy)tetrahydro-2H-pyran-2-yl)oxy)ethyl)prop-2-yn-1-aminium chloride

[0390] This compound may be synthesized according to the experimental procedure described for Compound 13.

##STR00126##

Compound 17 and Compound 60: N,N-dimethyl-N-(2-oxo-2-(((3R,4R,5S,6S)-3,4,5-tris(butyryloxy)-6-methyltetrahydro-2H-pyran-2-yl)oxy)ethyl)prop-2-yn-1-aminium chloride

[0391] This compound may be synthesized according to the experimental procedure described for Compound 13.

##STR00127##

Compound 18: N,N-dimethyl-N-(2-oxo-2-(((3S,4R,5R,6S)-3,4,5-tris(butyryloxy)-6-methyltetrahydro-2H-pyran-2-yl)oxy)ethyl)prop-2-yn-1-aminium chloride

[0392] This compound may be synthesized according to the experimental procedure described for Compound 13.

##STR00128##

Compound 19: N,N-dimethyl-N-(2-oxo-2-(((2R,3R,4S,5R,6R)-3,4,5-triacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)ethyl)prop-2-yn-1-aminium trifluoroacetate

[0393] Step 1

[0394] Compound 2,3,4,6-tetra-O-acetyl-D-glucopyranose (Carbosynth, 2 g, 5.7 mmol, 1 eq) was dissolved in DCM (10 mL), followed by addition of 2-chloroacetyl chloride (1.4 mL, 17.6 mmol, 3 eq), DMAP (0.6 g, 4.9 mmol, 0.9 eq), and DIPEA (3 mL, 17.2 mmol, 3 eq). The reaction mixture was stirred at room temperature overnight, followed by filtration and multiple purifications by column chromatography (0-100% EtOAc in hexanes) to yield 1-(2-chloroacetoxy)-2,3,4,6-tetra-O-acetyl-D-glucopyranose (0.8 g, 1.9 mmol, mixture of anomers, 33% yield).

[0395] Step 2

[0396] Compound 1-(2-chloroacetoxy)-2,3,4,6-tetra-O-acetyl-D-glucopyranose (0.6 g, 1.4 mmol, 1 eq) was dissolved in acetone, followed by addition of sodium iodide (0.6 g, 4.0 mmol, 3 eq). The reaction was stirred for a few minutes, and dimethylamino-1-propyne (0.5 mL, 4.6 mmol, 3.3 eq) was added. The reaction was stirred at room temperature overnight, then filtered. The filtrate was concentrated, diluted with DMSO and water, then purified by reverse phase C.sub.18 column chromatography (0.1% TFA in 95% water/5% MeCN to 100% MeCN). Fractions containing product were lyophilized to yield an off-white powder (230 mg, 0.39 mmol, 28% yield, TFA salt). LCMS (M+): 472.4 .sup.1H NMR (400 MHz, DMSO-d6) δ 6.27 (d, J=3.6 Hz, 1H), 5.35 (t, J=9.9 Hz, 1H), 5.17-5.08 (m, 2H), 4.76-4.62 (m, 2H), 4.53 (d, J=2.5 Hz, 2H), 4.30-3.97 (m, 4H), 3.26 (s, 6H), 2.08-1.91 (m, 12H).

##STR00129##

Compound 20 and Compound 55: N,N-dimethyl-N-(2-oxo-2-(((3R,4S,5R)-3,4,5-triacetoxytetrahydro-2H-pyran-2-yl)oxy)ethyl)prop-2-yn-1-aminium chloride

[0397] This compound may be synthesized according to the experimental procedure described for Compound 19.

##STR00130##

Compound 21: N,N-dimethyl-N-(2-oxo-2-(((3R,4R,5R)-3,4,5-triacetoxytetrahydro-2H-pyran-2-yl)oxy)ethyl)prop-2-yn-1-aminium chloride

[0398] This compound may be synthesized according to the experimental procedure described for Compound 19.

##STR00131##

Compound 22 and Compound 57: N,N-dimethyl-N-(2-oxo-2-(((3R,4S,5S)-3,4,5-triacetoxytetrahydro-2H-pyran-2-yl)oxy)ethyl)prop-2-yn-1-aminium chloride

[0399] This compound may be synthesized according to the experimental procedure described for Compound 19.

##STR00132##

Compound 23: N,N-dimethyl-N-(2-oxo-2-(((3R,4R,5S,6S)-3,4,5-triacetoxy-6-methyltetrahydro-2H-pyran-2-yl)oxy)ethyl)prop-2-yn-1-aminium chloride

[0400] This compound may be synthesized according to the experimental procedure described for Compound 19.

##STR00133##

Compound 24: N,N-dimethyl-N-(2-oxo-2-(((3S,4R,5R,6S)-3,4,5-triacetoxy-6-methyltetrahydro-2H-pyran-2-yl)oxy)ethyl)prop-2-yn-1-aminium chloride

[0401] This compound may be synthesized according to the experimental procedure described for Compound 19.

##STR00134##

Compound 25: N,N-dimethyl-N-(prop-2-yn-1-yl)-4-(((3R,4S,5R,6R)-3,4,5-tris(butyryloxy)-6-((butyryloxy)methyl)tetrahydro-2H-pyran-2-yl)oxy)butan-1-aminium bromide

[0402] Step 1:

[0403] To a solution of [(2R,3R,4S,5R)-3,4,5-tri(butanoyloxy)-6-(2,2,2-trichloroethanimidoyl)oxy-tetrahydropyran-2-yl]methyl butanoate (1.5 g, 2.48 mmol, 1 eq) and 4-bromobutan-1-ol (455.33 mg, 2.98 mmol, 1.2 eq) in DCM (15 mL) was added BF.sub.3.Et.sub.2O (1.76 g, 12.40 mmol, 1.53 mL, 5 eq). The mixture was stirred at 0° C. for 3 h and then warmed to 20° C. and stirred for 12 h. LC-MS showed[(2R,3R,4S,5R)-3,4,5-tri(butanoyloxy)-6-(2,2,2-trichloroethanimidoyl)oxy-tetrahydropyran-2-yl]methyl butanoate was consumed completely. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=20/1 to 3:1). Desired compound [(2R,3R,4S,5R)-6-(4-bromobutoxy)-3,4,5-tri(butanoyloxy)tetrahydropyran-2-yl]methyl butanoate (1 g, crude) was obtained as colorless oil.

[0404] Step 2:

[0405] To a mixture of [(2R,3R,4S,5R)-6-(4-bromobutoxy)-3,4,5-tri(butanoyloxy)tetrahydropyran-2-yl]methyl butanoate (0.2 g, 335.84 umol, 1 eq) and N,N-dimethylprop-2-yn-1-amine (83.76 mg, 1.01 mmol, 106.83 uL, 3 eq) in Acetone (3 mL) stirred at 90° C. for 2 h under N.sub.2. LCMS showed [(2R,3R,4S,5R)-6-(4-bromobutoxy)-3,4,5-tri(butanoyloxy)tetrahydropyran-2-yl]methyl butanoate was consumed completely. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC [water (0.10% TFA)-ACN]. The title compound (0.019 g, 28 μmol, 8.3% yield, mixture of anomers, Br) was obtained as colorless oil. LCMS (M.sup.−): 598.4 .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 5.41-5.27 (m, 1H), 5.11-4.77 (m, 3H), 4.35 (dd, J=10.3, 2.6 Hz, 2H), 4.23-3.97 (m, 4H), 3.79-3.65 (m, 1H), 3.59-3.44 (m, 1H), 3.11-3.04 (m, 6H), 2.34-2.11 (m, 8H), 1.80-1.39 (m, 12H), 0.96-0.78 (m, 12H).

##STR00135##

Compound 26: N,N-dimethyl-N-(prop-2-yn-1-yl)-4-(((3R,4S,5R)-3,4,5-tris(butyryloxy)tetrahydro-2H-pyran-2-yl)oxy)butan-1-aminium bromide

[0406] Step 1

[0407] To a solution of (3R,4S,5R)-tetrahydropyran-2,3,4,5-tetrol (10 g, 66.61 mmol, 1 eq) in Pyridine (100 mL) was added butanoyl butanoate (63.22 g, 399.66 mmol, 65.38 mL, 6 eq). The mixture was stirred at 25° C. for 10 h. TLC showed the starting reactant was consumed. The mixture was concentrated. The crude product [(3R,4S,5R)-4,5,6-tri(butanoyloxy)tetrahydropyran-3-yl] butanoate (64.89 g, crude) was obtained as yellow oil.

[0408] Step 2

[0409] To a solution of [(3R,4S,5R)-4,5,6-tri(butanoyloxy)tetrahydropyran-3-yl]butanoate (10 g, 23.23 mmol, 1 eq) in THF (100 mL) was added MeNH.sub.2 aq. solution (4.33 g, 41.81 mmol, 30% purity, 1.8 eq) in H.sub.2O. The mixture was stirred at 25° C. for 10 h. TLC showed the starting reactant was consumed. The mixture reaction was concentrated. The residue was purified by flash silica gel chromatography (PE:EA=5:1). [(3R,4S,5R)-4,5-di(butanoyloxy)-6-hydroxy-tetrahydropyran-3-yl] butanoate (2.65 g, 6.61 mmol, 28.4% yield, 90% purity) was obtained as yellow oil.

[0410] Step 3

[0411] To a solution of [(3R,4S,5R)-4,5-di(butanoyloxy)-6-hydroxy-tetrahydropyran-3-yl] butanoate (500 mg, 1.39 mmol, 1 eq) in DCM (10 mL) was added 2,2,2-trichloroacetonitrile (2.00 g, 13.87 mmol, 1.39 mL, 10 eq) and DBU (105.61 mg, 693.68 umol, 104.56 uL, 0.5 eq). The mixture was stirred at 25° C. for 10 h. TLC showed the starting reactant was consumed. The mixture was concentrated. The residue was purified by flash silica gel chromatography (PE:EA=5:1). Compound [(3R,4S,5R)-4,5-di(butanoyloxy)-6-(2,2,2-trichloroethanimidoyl)oxy-tetrahydropyran-3-yl] butanoate (0.5 g, 792.42 umol, 57.1% yield, 80% purity) was obtained as a white solid.

[0412] Step 4

[0413] To a solution of [(3R,4S,5R)-4,5-di(butanoyloxy)-6-(2,2,2-trichloroethanimidoyl)oxy-tetrahy-dropyran-3-yl] butanoate (400 mg, 792.42 umol, 1 eq) and 4-bromobutan-1-ol (181.88 mg, 1.19 mmol, 1.5 eq) in DCM (40 mL) was added BF.sub.3.Et.sub.2O (562.34 mg, 3.96 mmol, 489 uL, 5 eq) under dark at 0° C. and stirred for 3 h. Then the mixture was stirred at 25° C. for 12 h. TLC showed the starting reactant was consumed. The mixture was concentrated. The residue was purified by silica gel chromatography (PE:EA=5:1). [(3R,4S,5R)-6-(4-bromobutoxy)-4,5-di(butanoyloxy) tetrahydropyran-3-yl] butanoate (160 mg, crude) was obtained as yellow oil.

[0414] Step 5

[0415] A solution of [(3R,4S,5R)-6-(4-bromobutoxy)-4,5-di(butanoyloxy)tetrahydropyran-3-yl] butanoate (643 mg, 1.30 mmol, 1 eq) and N,N-dimethylprop-2-yn-1-amine (323.70 mg, 3.89 mmol, 412.88 uL, 3 eq) in acetone (10 mL) was stirred at 90° C. for 3 h. LCMS showed the product formed. The mixture was concentrated. The residue was purified by prep-HPLC (column: Xbridge 150*30 mm*10 um; mobile phase: [water (0.1% TFA)-ACN]; B %: 25%-55%, 10 min). The title compound (10 mg, 16.9 umol, 1.3% yield, mixture of anomers, Br) was obtained as yellow oil. LCMS (M.sup.+): 498.3 .sup.1H NMR (400 MHz, Chloroform-d) δ 5.45-5.12 (m, 1H), 4.95-4.74 (m, 3H), 4.46-4.32 (m, 2H), 3.80-3.67 (m, 2H), 3.58-3.35 (m, 4H), 3.31-3.18 (m, 6H), 2.91-2.79 (m, 1H), 2.32-2.10 (m, 6H), 1.87-1.45 (m, 10H), 0.93-0.79 (m, 9H).

##STR00136##

Compound 27 and Compound 67: N,N-dimethyl-N-(prop-2-yn-1-yl)-4-(((3R,4R,5R)-3,4,5-tris(butyryloxy)tetrahydro-2H-pyran-2-yl)oxy)butan-1-aminium bromide

[0416] This compound may be synthesized according to the experimental procedure described for Compound 26.

##STR00137##

Compound 28 and Compound 59: N,N-dimethyl-N-(prop-2-yn-1-yl)-4-(((2S,3R,4S,5S)-3,4,5-tris(butyryloxy)tetrahydro-2H-pyran-2-yl)oxy)butan-1-aminium bromide

[0417] Step 1:

[0418] To a solution of (3R,4S,5S)-tetrahydropyran-2,3,4,5-tetrol (5 g, 33.30 mmol, 1 eq) in pyridine (50 mL) was added butanoyl butanoate (31.61 g, 199.83 mmol, 32.69 mL, 6 eq). The mixture was stirred at 25° C. for 12 h. TLC showed the starting reactant was consumed. The mixture was concentrated. [(3S,4S,5R)-4,5,6-tri(butanoyloxy)tetrahydropyran-3-yl] butanoate (15 g, crude) was obtained as yellow oil.

[0419] Step 2

[0420] To a solution of [(3S,4S,5R)-4,5,6-tri(butanoyloxy)tetrahydropyran-3-yl]butanoate (15 g, 34.84 mmol, 1 eq) in THF (50 mL) was added MeNH.sub.2 aq. solution (5.41 g, 52.27 mmol, 30% purity, 1.5 eq) and stirred at 25° C. for 12 h. TLC showed the starting reactant was consumed. The mixture reaction was concentrated. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=10/1 to 5:1). [(3S,4S,5R)-4,5-di(butanoyloxy)-6-hydroxy-tetrahydropyran-3-yl] butanoate (4 g, 8.88 mmol, 25.5% yield, 80% purity) was obtained as yellow oil.

[0421] Step 3

[0422] To a solution of [(3S,4S,5R)-4,5-di(butanoyloxy)-6-hydroxy-tetrahydropyran-3-yl] butanoate (2 g, 5.55 mmol, 1 eq) and 2,2,2-trichloroacetonitrile (8.01 g, 55.49 mmol, 5.56 mL, 10 eq) in DCM (20 mL) was added DBU (422.42 mg, 2.77 mmol, 418.24 uL, 0.5 eq) at 25° C. and stirred for 12 h. TLC showed the starting reactant was consumed. The mixture was concentrated. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=10/1 to 3:1). [(3S,4S,5R)-4,5-di(butanoyloxy)-6-(2,2,2-trichloroethanimidoyl)oxy-tetrahydropyran-3-yl] butanoate (1.5 g, 2.67 mmol, 48.2% yield, 90% purity) was obtained as yellow oil.

[0423] Step 4

[0424] [(3S,4S,5R)-4,5-di(butanoyloxy)-6-(2,2,2-trichloroethanimidoyl)oxy-tetrahydropyran-3-yl]butanoate (1.5 g, 2.97 mmol, 1 eq) and 4-bromobutan-1-ol (682.05 mg, 4.46 mmol, 1.5 eq) was dissolved with DCM (20 mL) and stirred at 0° C. for 3 h in dark. BF.sub.3.Et.sub.2O (2.11 g, 14.86 mmol, 1.83 mL, 5 eq) was added into the mixture at 0° C. and stirred for 12 h at 25° C. TLC showed the starting reactant was consumed. The mixture was concentrated. The residue was purified by flash silica gel chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=5/1). Compound [(3S,4S,5R)-6-(4-bromobutoxy)-4,5-di(butanoyloxy)tetrahydropyran-3-yl] butanoate (1.6 g, crude) was obtained as brown oil.

[0425] Step 5

[0426] To a solution of [(3S,4S,5R)-6-(4-bromobutoxy)-4,5-di(butanoyloxy)tetrahydropyran-3-yl] butanoate (1.6 g, 3.23 mmol, 1 eq) in acetone (10 mL) was added N,N-dimethylprop-2-yn-1-amine (805.47 mg, 9.69 mmol, 1.03 mL, 3 eq) and stirred at 90° C. for 2 h. LCMS showed the product formed. The mixture was concentrated. The residue was purified by prep-HPLC (column: Luna C18 100*30 5u; mobile phase: [water (0.1% TFA)-ACN]; B %: 25%-50%, 12 min). The title compound (42 mg, 69 μmol, 2.1% yield, Br) as a yellow oil was obtained. The anomer (Compound 29) was also isolated. LCMS (M.sup.+): 498.3 .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 5.30-5.24 (m, 1H), 5.22 (dd, J=9.9, 3.6 Hz, 1H), 5.06-4.97 (m, 2H), 4.33 (d, J=2.6 Hz, 2H), 4.04 (t, J=2.4 Hz, 1H), 3.92 (dd, J=13.4, 1.6 Hz, 1H), 3.66-3.57 (m, 2H), 3.50-3.28 (m, 3H), 3.05 (s, 6H), 2.37-2.29 (m, 2H), 2.25 (td, J=7.2, 2.3 Hz, 2H), 2.14 (td, J=7.2, 1.8 Hz, 2H), 1.78-1.66 (m, 2H), 1.62-1.39 (m, 8H), 0.94-0.77 (m, 9H).

##STR00138##

Compound 29 and Compound 66: N,N-dimethyl-N-(prop-2-yn-1-yl)-4-(((2R,3R,4S,5S)-3,4,5-tris(butyryloxy)tetrahydro-2H-pyran-2-yl)oxy)butan-1-aminium bromide

[0427] This compound was synthesized according to the experimental procedure described for Compound 28. The title compound (21 mg, 34.5 μmol, 1.1% yield, Br) was isolated as a yellow oil. LCMS (M.sup.+): 498.2 .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 5.20-5.14 (m, 1H), 5.11 (dd, J=10.1, 3.6 Hz, 1H), 4.95 (dd, J=10.1, 7.7 Hz, 1H), 4.57 (d, J=7.7 Hz, 1H), 4.30 (d, J=2.6 Hz, 2H), 4.06-4.00 (m, 1H), 3.85-3.76 (m, 2H), 3.76-3.66 (m, 1H), 3.56-3.42 (m, 1H), 3.03 (s, 6H), 2.31 (t, J=7.2 Hz, 2H), 2.24 (td, J=7.2, 3.5 Hz, 2H), 2.17-2.09 (m, 2H), 1.73-1.62 (m, 2H), 1.60-1.38 (m, 8H), 0.95-0.76 (m, 9H).

##STR00139##

Compound 30: N,N-dimethyl-N-(prop-2-yn-1-yl)-4-(((2R,3R,4R,5S,6S)-3,4,5-tris(butyryloxy)-6-methyltetrahydro-2H-pyran-2-yl)oxy)butan-1-aminium bromide

[0428] Step 1:

[0429] To a solution of (3R,4R,5R,6S)-6-methyltetrahydro-2H-pyran-2,3,4,5-tetraol (10 g, 60.92 mmol, 1 eq) in Pyridine (100 mL) was added butanoyl butanoate (57.82 g, 365.51 mmol, 59.79 mL, 6 eq) at 25° C. The mixture was stirred at 25° C. for 12 h. TLC indicated new spots formed. The reaction mixture was concentrated under reduced pressure to give a residue. The reaction mixture was washed with saturated sodium bicarbonate solution 300 mL (100 mL*3) and extracted with ethyl acetate 100 mL. The organic layer were washed with brine 50 mL, dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to give (3R,4R,5S,6S)-6-methyltetrahydro-2H-pyran-2,3,4,5-tetrayl tetrabutyrate (34 g, crude) as yellow oil.

[0430] Step 2:

[0431] To a solution of (3R,4R,5S,6S)-6-methyltetrahydro-2H-pyran-2,3,4,5-tetrayl tetrabutyrate (34 g, 76.49 mmol, 1 eq) in THF (20 mL) was added MeNH.sub.2 aq. solution (10.69 g, 137.68 mmol, 40% purity, 1.8 eq) at 25° C. The mixture was stirred at 25° C. for 12 hr. TLC indicated new spots formed. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=10/1 to 1:1) to give (3R,4R,5S,6S)-2-hydroxy-6-methyltetrahydro-2H-pyran-3,4,5-triyl tributyrate (18 g, 48.1 mmol, 62.9% yield) as yellow oil.

[0432] Step 3:

[0433] To a solution of (3R,4R,5S,6S)-2-hydroxy-6-methyltetrahydro-2H-pyran-3,4,5-triyl tributyrate (5 g, 13.35 mmol, 1 eq) in DCM (50 mL) was added DBU (1.02 g, 6.68 mmol, 1.01 mL, 0.5 eq) and 2,2,2-trichloroacetonitrile (19.28 g, 133.54 mmol, 13.39 mL, 10 eq) at 25° C. The mixture was stirred at 25° C. for 2 h. TLC indicated new spots formed. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=10/1 to 1:1) to give (2S,3S,4R,5R)-2-methyl-6-(2,2,2-trichloro-1-iminoethoxy)tetrahydro-2H-pyran-3,4,5-triyl tributyrate (4 g, 7.7 mmol, 57.7% yield) as yellow oil.

[0434] Step 4:

[0435] To a solution of 4-bromobutan-1-ol (766.84 mg, 5.01 mmol, 1 eq) and (2S,3S,4R,5R)-2-methyl-6-(2,2,2-trichloro-1-iminoethoxy)tetrahydro-2H-pyran-3,4,5-triyl tributyrate (2.6 g, 5.01 mmol, 1 eq) in DCM (30 mL) was added BF.sub.3.Et.sub.2O (3.56 g, 25.06 mmol, 3.09 mL, 5 eq) at 0° C. The mixture was stirred at 0° C. for 3 hr. The mixture was stirred at 25° C. for 12 hr. TLC indicated new spots formed. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=20:1) to give (3R,4R,5S,6S)-2-(4-bromobutoxy)-6-methyltetrahydro-2H-pyran-3,4,5-triyl tributyrate (600 mg, 1.2 mmol, 23.5% yield) as yellow oil.

[0436] Step 5:

[0437] To a solution of (3R,4R,5S,6S)-2-(4-bromobutoxy)-6-methyltetrahydro-2H-pyran-3,4,5-triyl tributyrate (600 mg, 1.18 mmol, 1 eq) in Acetone (5 mL) was added N,N-dimethylprop-2-yn-1-amine (293.73 mg, 3.53 mmol, 374.66 uL, 3 eq) at 25° C. The mixture was stirred at 90° C. for 2 h. LCMS showed the desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (column: Luna C18 100*30 5u; mobile phase: [water (0.1% TFA)-ACN]; B %: 25%-55%, 12 min) to give the title compound (150 mg, 253 μmol, 21.5% yield) as a yellow oil. LCMS (M.sup.+): 512.3 .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 5.16-5.06 (m, 2H), 4.91 (t, J=9.9 Hz, 1H), 4.78 (d, J=1.6 Hz, 1H), 4.34 (d, J=2.5 Hz, 2H), 4.00 (t, J=2.5 Hz, 1H), 3.87-3.76 (m, 1H), 3.65 (dt, J=9.9, 6.1 Hz, 1H), 3.50-3.40 (m, 1H), 3.40-3.31 (m, 2H), 3.06 (s, 6H), 2.38-2.08 (m, 6H), 1.81-1.68 (m, 2H), 1.62-1.39 (m, 8H), 1.10 (d, J=6.2 Hz, 3H), 0.90 (t, J=7.4 Hz, 3H), 0.87-0.75 (m, 6H).

##STR00140##

Compound 31 and Compound 58: N,N-dimethyl-N-(prop-2-yn-1-yl)-4-(((3S,4R,5R,6S)-3,4,5-tris(butyryloxy)-6-methyltetrahydro-2H-pyran-2-yl)oxy)butan-1-aminium bromide

[0438] This compound may be synthesized according to the experimental procedure described for Compound 30.

##STR00141##

Compound 32: N,N-dimethyl-N-(prop-2-yn-1-yl)-4-(((3R,4S,5R,6R)-3,4,5-triacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)butan-1-aminium bromide

[0439] Step 1:

[0440] To a mixture of [(2R,3R,4S,5R)-3,4,5-triacetoxy-6-(2,2,2-trichloroethanimidoyl)oxy-tetrahydropyran-2-yl]methyl acetate (1 g, 2.03 mmol, 1 eq) and 4-bromobutan-1-ol (372.69 mg, 2.44 mmol, 1.2 eq) in DCM (10 mL) was added BF.sub.3.Et.sub.2O (1.44 g, 10.15 mmol, 1.25 mL, 5 eq) in one portion at 0° C. under N.sub.2. The mixture was stirred at 0° C. for 3 h and then heated to 20° C. and stirred for 12 h. TLC indicated the starting material was consumed completely. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=5/1 to 2:1). [(2R,3R,4S,5R)-3,4,5-triacetoxy-6-(4-bromobutoxy)tetrahydropyran-2-yl]methyl acetate (1 g, crude) was obtained as colorless oil.

[0441] Step 2:

[0442] To a mixture of [(2R,3R,4S,5R)-3,4,5-triacetoxy-6-(4-bromobutoxy)tetrahydropyran-2-yl]methyl acetate (0.2 g, 413.82 umol, 1 eq) and N,N-dimethylprop-2-yn-1-amine (103.20 mg, 1.24 mmol, 131.64 uL, 3 eq) in acetone (3 mL) was stirred at 90° C. for 2 h. LC-MS showed [(2R,3R,4S,5R)-3,4,5-triacetoxy-6-(4-bromobutoxy)tetrahydropyran-2-yl]methyl acetate was consumed completely. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC [water (0.1% TFA)-ACN]. The title compound (0.006 g, 10.5 μmol, 2.5% yield, mixture of anomers) was obtained as colorless oil. LCMS (M.sup.+): 486.2 .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 5.44-5.19 (m, 1H), 5.15-4.65 (m, 3H), 4.39-4.32 (m, 2H), 4.22-3.94 (m, 4H), 3.75-3.65 (m, 1H), 3.59-3.35 (m, 3H), 3.13-3.04 (m, 6H), 2.15-1.89 (m, 12H), 1.79-1.74 (m, 2H), 1.66-1.58 (m, 2H).

##STR00142##

Compound 33: N,N-dimethyl-N-(prop-2-yn-1-yl)-4-(((3R,4S,5R)-3,4,5-triacetoxytetrahydro-2H-pyran-2-yl)oxy)butan-1-aminium bromide

[0443] Step 1

[0444] To a solution of (3R,4S,5R)-tetrahydropyran-2,3,4,5-tetrol (10 g, 66.61 mmol, 1 eq) in Pyridine (100 mL) was added acetyl acetate (40.80 g, 399.65 mmol, 37.43 mL, 6 eq). The mixture was stirred at 25° C. for 10 h. TLC showed the starting reactant was consumed. The mixture was concentrated. Compound [(3R,4S,5R)-4,5,6-triacetoxytetrahydropyran-3-yl] acetate (10.6 g, crude) was obtained as a white solid.

[0445] Step 2

[0446] To a solution of [(3R,4S,5R)-4,5,6-triacetoxytetrahydropyran-3-yl] acetate (5 g, 15.71 mmol, 1 eq) in THF (50 mL) was added MeNH.sub.2 aq. solution (2.93 g, 28.28 mmol, 30% purity, 1.8 eq). The mixture was stirred at 25° C. for 10 h. TLC showed the starting reactant was consumed. The mixture reaction was concentrated. The residue was purified by flash silica gel chromatography (PE:EA=2:1). [(3R,4S,5R)-4,5-diacetoxy-6-hydroxy-tetrahydropyran-3-yl] acetate (1.53 g, 4.98 mmol, 31.7% yield, 90% purity) was obtained as a yellow solid.

[0447] Step 3

[0448] To a solution of [(3R,4S,5R)-4,5-diacetoxy-6-hydroxy-tetrahydropyran-3-yl]acetate (1 g, 3.62 mmol, 1 eq) in DCM (20 mL) was added 2,2,2-trichloroacetonitrile (5.23 g, 36.20 mmol, 3.63 mL, 10 eq) and DBU (275.55 mg, 1.81 mmol, 272.82 uL, 0.5 eq). The mixture was stirred at 25° C. for 10 h. TLC showed the starting reactant was consumed. The mixture was concentrated. The residue was purified by flash silica gel chromatography (PE:EA=5:1). Compound [(3R,4S,5R)-4,5-diacetoxy-6-(2,2,2-trichloroethanimidoyl)oxy-tetrahydropyran-3-yl] acetate (1.09 g, 2.07 mmol, 57.3% yield, 80% purity) was obtained as a white solid.

[0449] Step 4

[0450] To a solution of [(3R,4S,5R)-4,5-diacetoxy-6-(2,2,2-trichloroethanimidoyl)oxy-tetrahydropyran-3-yl] acetate (1.09 g, 2.59 mmol, 1 eq) and 4-bromobutan-1-ol (594.79 mg, 3.89 mmol, 1.5 eq) in DCM (10 mL) was added BF.sub.3.Et.sub.2O (1.84 g, 12.96 mmol, 1.60 mL, 5 eq) in dark and stirred at 0° C. for 3 h. Then the mixture was stirred at 25° C. for 12 h. TLC showed the starting reactant was consumed. The mixture was concentrated and purified by flash silica gel chromatography (PE:EA=5:1). [(3R,4S,5R)-4,5-diacetoxy-6-(4-bromobutoxy)tetrahydropyran-3-yl] acetate (429 mg, 835 umol, 32.2% yield, 80% purity) was obtained as yellow oil.

[0451] Step 5

[0452] The solution of [(3R,4S,5R)-4,5-diacetoxy-6-(4-bromobutoxy)tetrahydropyran-3-yl] acetate (429 mg, 1.04 mmol, 1 eq) and N,N-dimethylprop-2-yn-1-amine (260.16 mg, 3.13 mmol, 331.84 uL, 3 eq) in acetone (10 mL) was stirred at 90° C. for 3 h. LCMS showed the product formed. The mixture was concentrated. The residue was purified by prep-HPLC (column: Xbridge 150*30 mm*10 um; mobile phase: [water (0.1% TFA)-ACN]; B %: 25%-55%, 10 min). The title compound (10 mg, 23.2 μmol, 2.2% yield, mixture) was obtained as a yellow oil. LCMS (M.sup.+) 414.2 .sup.1H NMR (400 MHz, Chloroform-d) δ 5.5-5.1 (m, 1H), 5.0-4.8 (m, 3H), 4.6-4.4 (m, 2H), 3.9-3.7 (m, 2H), 3.7-3.4 (m, 4H), 3.4-3.2 (m, 6H), 3.0-2.6 (m, 1H), 2.4-2.0 (m, 9H), 2.0-1.6 (m, 4H).

##STR00143##

Compound 34: N,N-dimethyl-N-(prop-2-yn-1-yl)-4-(((3R,4R,5R)-3,4,5-triacetoxytetrahydro-2H-pyran-2-yl)oxy)butan-1-aminium bromide

[0453] This compound may be synthesized according to the experimental procedure described for Compound 33.

##STR00144##

Compound 35 and compound 62: N,N-dimethyl-N-(prop-2-yn-1-yl)-4-(((2S,3R,4S,5S)-3,4,5-triacetoxytetrahydro-2H-pyran-2-yl)oxy)butan-1-aminium bromide

[0454] Step 1

[0455] To a solution of (3R,4S,5S)-tetrahydropyran-2,3,4,5-tetrol (3 g, 19.98 mmol, 1 eq) in Pyridine (50 mL) was added acetyl acetate (12.24 g, 119.90 mmol, 11.23 mL, 6 eq) and then the mixture was stirred at 25° C. for 12 h. TLC showed the starting reactant was consumed. To the mixture was added H.sub.2O (250 mL) at 0° C., extracted with EtOAc (300 mL) and the organic layer was concentrated. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=10/1 to 4:1). [(3S,4S,5R)-4,5,6-triacetoxytetrahydropyran-3-yl] acetate (5 g, crude) was obtained as yellow solid.

[0456] Step 2

[0457] To a solution of [(3S,4S,5R)-4,5,6-triacetoxytetrahydropyran-3-yl] acetate (5 g, 15.71 mmol, 1 eq) in THF (50 mL) was added MeNH.sub.2 aq. solution (2.44 g, 23.56 mmol, 30% purity, 1.5 eq) and stirred at 25° C. for 12 h. TLC showed the starting reactant was consumed. The mixture reaction was concentrated. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=10/1 to 5:1). [(3S,4S,5R)-4,5-diacetoxy-6-hydroxy-tetrahydropyran-3-yl] acetate (2 g, 6.5 mmol, 41.5% yield, 90% purity) was obtained as yellow solid.

[0458] Step 3

[0459] To a solution of [(3S,4S,5R)-4,5-diacetoxy-6-hydroxy-tetrahydropyran-3-yl]acetate (2 g, 7.24 mmol, 1 eq) and 2,2,2-trichloroacetonitrile (10.45 g, 72.40 mmol, 7.26 mL, 10 eq) in DCM (10 mL) was added DBU (551.11 mg, 3.62 mmol, 545.65 uL, 0.5 eq) at 25° C. and stirred for 12 h. TLC showed the starting reactant was consumed. The mixture was combined work-up with another batch in Et16408-556. The combined mixture was concentrated. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=20/1 to 3:1). [(3S,4S,5R)-4,5-diacetoxy-6-(2,2,2-trichloroethanimidoyl)oxy-tetrahydropyran-3-yl] acetate (1.5 g in total) was obtained as white solid.

[0460] Step 4

[0461] [(3S,4S,5R)-4,5-diacetoxy-6-(2,2,2-trichloroethanimidoyl)oxy-tetrahydropyran-3-yl] acetate (500 mg, 1.19 mmol, 1 eq) and 4-bromobutan-1-ol (236.46 mg, 1.55 mmol, 1.3 eq) was dissolved with DCM (5 mL) and stirred at 0° C. for 3 h in dark. BF.sub.3.Et.sub.2O (843.55 mg, 5.94 mmol, 733.52 uL, 5 eq) was added into the mixture at 0° C. and stirred for 12 h at 25° C. TLC showed the starting reactant was consumed. The mixture was concentrated. The residue was purified by flash silica gel chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=5/1). [(3S,4S,5R)-4,5-diacetoxy-6-(4-bromobutoxy)tetrahydropyran-3-yl] acetate (500 mg, crude) was obtained as brown oil.

[0462] Step 5

[0463] To a solution of [(3S,4S,5R)-4,5-diacetoxy-6-(4-bromobutoxy)tetrahydropyran-3-yl] acetate (500 mg, 1.22 mmol, 1 eq) in acetone (5 mL) was added N,N-dimethylprop-2-yn-1-amine (303.22 mg, 3.65 mmol, 386.76 uL, 3 eq) and stirred at 90° C. for 2 h. LCMS showed the product formed. The reaction mixture was concentrated. The residue was purified by prep-HPLC (column: Luna C18 100*30 5u; mobile phase: [water (0.1% TFA)-ACN]; B %: 1%-30%, 12 min). The title compound (120 mg, 290 umol, 23.8% yield) was obtained as colorless oil. LCMS (M.sup.+): 414.2 .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 5.30-5.25 (m, 1H), 5.22 (dd, J=9.9, 3.6 Hz, 1H), 5.06-4.97 (m, 2H), 4.36 (d, J=2.6 Hz, 2H), 4.07 (t, J=2.5 Hz, 1H), 3.93 (dd, J=13.4, 1.5 Hz, 1H), 3.71-3.61 (m, 2H), 3.51-3.34 (m, 3H), 3.08 (s, 6H), 2.13-2.01 (m, 6H), 1.96 (s, 3H), 1.82-1.72 (m, 2H), 1.61 (q, J=6.7 Hz, 2H).

##STR00145##

Compound 36: N,N-dimethyl-N-(prop-2-yn-1-yl)-4-(((3R,4R,5S,6S)-3,4,5-triacetoxy-6-methyltetrahydro-2H-pyran-2-yl)oxy)butan-1-aminium bromide

[0464] Step 1

[0465] To a solution of (3R,4R,5R,6S)-6-methyltetrahydropyran-2,3,4,5-tetrol (10 g, 60.92 mmol, 1 eq) in Pyridine (100 mL) was added acetyl acetate (37.31 g, 365.51 mmol, 34.23 mL, 6 eq) at 25° C. The mixture was stirred at 25° C. for 12 hr. TLC indicated new spots formed. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=10:1 to 1:1) to give [(2S,3S,4R,5R)-4,5,6-triacetoxy-2-methyl-tetrahydropyran-3-yl] acetate (16 g, 48 mmol, 79% yield) as a colorless oil.

[0466] Step 2

[0467] To a solution of [(2S,3S,4R,5R)-4,5,6-triacetoxy-2-methyl-tetrahydropyran-3-yl] acetate (16 g, 48.15 mmol, 1 eq) in THF (160 mL) was added MeNH.sub.2 aq. (6.73 g, 86.67 mmol, 40% purity, 1.8 eq) at 25° C. The mixture was stirred at 25° C. for 12 hr. TLC indicated new spots formed. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=10/1 to 0:1) to give [(2S,3S,4R,5R)-4,5-diacetoxy-6-hydroxy-2-methyl-tetrahydropyran-3-yl] acetate (8 g, 28 mmol, 57% yield) as a white solid.

[0468] Step 3

[0469] To a solution of [(2S,3S,4R,5R)-4,5-diacetoxy-6-hydroxy-2-methyl-tetrahydropyran-3-yl] acetate (3 g, 10.34 mmol, 1 eq) in DCM (30 mL) was added DBU (786.71 mg, 5.17 mmol, 778.92 uL, 0.5 eq) and 2,2,2-trichloroacetonitrile (14.92 g, 103.35 mmol, 10.36 mL, 10 eq) at 25° C. The mixture was stirred at 25° C. for 2 hr. TLC indicated new spots formed. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=10:1 to 9:1) to give [(2S,3S,4R,5R)-4,5-diacetoxy-2-methyl-6-(2,2,2-trichloroethanimidoyl)oxy-tetrahydropyran-3-yl] acetate (3 g, 6.9 mmol, 66.8% yield) as a yellow oil.

[0470] Step 4

[0471] To a solution of [(2S,3S,4R,5R)-4,5-diacetoxy-2-methyl-6-(2,2,2-trichloroethanimidoyl)oxy-tetrahydropyran-3-yl] acetate (1.5 g, 3.45 mmol, 1 eq) and 4-bromobutan-1-ol (686.49 mg, 4.49 mmol, 1.3 eq) in DCM (15 mL) was added BF.sub.3.Et.sub.2O (2.45 g, 17.26 mmol, 2.13 mL, 5 eq) at 0° C. The mixture was stirred at 0° C. for 3 hr. The mixture was stirred at 25° C. for 12 hr. TLC indicated new spots formed. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=10:1 to 1:1) to give [(2S,3S,4R,5R)-4,5-diacetoxy-6-(4-bromobutoxy)-2-methyl-tetrahydropyran-3-yl]acetate (300 mg, 705 umol, 20% yield) as a yellow oil.

[0472] Step 5

[0473] To a solution of [(2S,3S,4R,5R)-4,5-diacetoxy-6-(4-bromobutoxy)-2-methyl-tetrahydropyran-3-yl] acetate (250 mg, 587.9 umol, 1 eq) in acetone (5 mL) was added N,N-dimethylprop-2-yn-1-amine (146.61 mg, 1.76 mmol, 187.00 uL, 3 eq) at 25° C. The mixture was stirred at 90° C. for 4 hr. LCMS showed the desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (column: Luna C18 100*30 5u; mobile phase: [water (water (0.1% TFA)-ACN]; B %: 10%-35%, 12 min) to give the title compound (47 mg, 90.6 umol, 15.4% yield, mixture of anomers) as a yellow solid. LCMS (M.sup.+): 428.2 .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 5.12-5.03 (m, 2H), 4.87 (t, J=9.8 Hz, 1H), 4.79 (d, J=1.6 Hz, 1H), 4.34 (d, J=2.6 Hz, 2H), 4.01 (t, J=2.5 Hz, 1H), 3.86-3.75 (m, 1H), 3.70-3.60 (m, 1H), 3.50-3.36 (m, 1H), 3.39-3.31 (m, 2H), 3.06 (s, 6H), 2.09 (s, 3H), 2.02 (s, 3H), 1.92 (s, 3H), 1.79-1.68 (m, 2H), 1.64-1.46 (m, 2H), 1.11 (d, J=6.2 Hz, 3H).

##STR00146##

Compound 37 and Compound 37: N,N-dimethyl-N-(prop-2-yn-1-yl)-4-(((3S,4R,5R,6S)-3,4,5-triacetoxy-6-methyltetrahydro-2H-pyran-2-yl)oxy)butan-1-aminium bromide

[0474] This compound may be synthesized according to the experimental procedure described for Compound 36.

##STR00147##

Compound 38: N,N-dimethyl-4-oxo-N-(prop-2-yn-1-yl)-4-(((3R,4S,5R,6R)-3,4,5-tris(butyryloxy)-6-((butyryloxy)methyl)tetrahydro-2H-pyran-2-yl)oxy)butan-1-aminium bromide

[0475] Step 1:

[0476] To a mixture of [(2R,3R,4S,5R)-3,4,5-tri(butanoyloxy)-6-hydroxy-tetrahydropyran-2-yl]methyl butanoate (1 g, 2.17 mmol, 1 eq) in DCM (10 mL) was added 4-bromobutanoyl chloride (1.21 g, 6.51 mmol, 755.05 uL, 3 eq) and Pyridine (858.82 mg, 10.86 mmol, 876.35 uL, 5 eq) in one portion at 20° C. under N.sub.2. The mixture was stirred at 20° C. for 12 h. TLC showed indicated[(2R,3R,4S,5R)-3,4,5-tri(butanoyloxy)-6-hydroxy-tetrahydropyran-2-yl]methyl butanoate was consumed completely. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=10/1 to 5:1). Compound [(3R,4S,5R,6R)-3,4,5-tri(butanoyloxy)-6-(butanoyloxymethyl)tetrahydropyran-2-yl]4-bromobutanoate (1.5 g, crude) was obtained as colorless oil.

[0477] Step 2:

[0478] To a mixture of [(3R,4S,5R,6R)-3,4,5-tri(butanoyloxy)-6-(butanoyloxymethyl)tetrahydropyran-2-yl] 4-bromobutanoate (0.2 g, 328.14 umol, 1 eq) and N,N-dimethylprop-2-yn-1-amine (81.84 mg, 984.41 umol, 104.38 uL, 3 eq) in Acetone (3 mL) was stirred at 90° C. for 2 h. LCMS showed [(3R,4S,5R,6R)-3,4,5-tri(butanoyloxy)-6-(butanoyloxymethyl)tetrahydropyran-2-yl]4-bromobutanoate (0.2 g, 328.14 umol, 1 eq) was consumed completely. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC [water (0.1% TFA)-ACN]. The title compound (0.028 g, 40.4 μmol, 12.3% yield, mixture of anomers, Br) was obtained as a colorless oil. LCMS (M.sup.+): 612.3 .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 6.28-5.95 (m, 1H), 5.56-5.34 (m, 1H), 5.20-4.94 (m, 2H), 4.44-4.35 (m, 2H), 4.30-4.12 (m, 2H), 4.10-3.98 (m, 2H), 3.45-3.30 (m, 2H), 3.14-3.06 (m, 6H), 2.78-2.40 (m, 2H), 2.34-2.13 (m, 8H), 2.06-1.88 (m, 2H), 1.61-1.40 (m, 8H), 0.93-0.79 (m, 12H).

##STR00148##

Compound 39: N,N-dimethyl-4-oxo-N-(prop-2-yn-1-yl)-4-(((3R,4S,5R)-3,4,5-tris(butyryloxy)tetrahydro-2H-pyran-2-yl)oxy)butan-1-aminium bromide

[0479] This compound may be synthesized according to the experimental procedure described for Compound 38.

##STR00149##

Compound 40 and Compound 68: N,N-dimethyl-4-oxo-N-(prop-2-yn-1-yl)-4-(((3R,4R,5R)-3,4,5-tris(butyryloxy)tetrahydro-2H-pyran-2-yl)oxy)butan-1-aminium bromide

[0480] This compound may be synthesized according to the experimental procedure described for Compound 38.

##STR00150##

Compound 41A: N,N-dimethyl-4-oxo-N-(prop-2-yn-1-yl)-4-(((2R,3R,4S,5S)-3,4,5-tris(butyryloxy)tetrahydro-2H-pyran-2-yl)oxy)butan-1-aminium bromide

[0481] Step 1

[0482] [(3S,4S,5R)-4,5-di(butanoyloxy)-6-hydroxy-tetrahydropyran-3-yl] butanoate (500 mg, 1.39 mmol, 1 eq) and 4-bromobutanoyl chloride (771.84 mg, 4.16 mmol, 482.40 uL, 3 eq) was dissolved with DCM (5 mL). Then Pyridine (548.70 mg, 6.94 mmol, 559.90 uL, 5 eq) was added into the mixture and stirred for 12 h at 25° C. TLC showed the starting reactant was consumed. The reaction mixture was concentrated. [(3R,4S,5S)-3,4,5-tri(butanoyloxy)tetrahydropyran-2-yl] 4-bromobutanoate (0.5 g, crude) was obtained as colorless oil.

[0483] Step 2

[0484] To a solution of [(3R,4S,5S)-3,4,5-tri(butanoyloxy)tetrahydropyran-2-yl] 4-bromobutanoate (500 mg, 981.58 umol, 1 eq) in Acetone (5 mL) was added N,N-dimethylprop-2-yn-1-amine (244.80 mg, 2.94 mmol, 312.24 uL, 3 eq) and stirred at 90° C. for 2 h. LCMS showed the product formed. The mixture was concentrated. The residue was purified by prep-HPLC (column: Luna C18 100*30 Su; mobile phase: [water (0.1% TFA)-ACN]; B %: 25%-50%, 12 min). The title compound (67 mg, 107 umol, 11% yield, Br) as a yellow oil was obtained. The anomer (Compound 42) was also isolated. LCMS (M.sup.+): 512.3 .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 6.23 (d, J=3.6 Hz, 1H), 5.41-5.35 (m, 1H), 5.32 (dd, J=10.7, 3.4 Hz, 1H), 5.18 (dd, J=10.7, 3.6 Hz, 1H), 4.39 (d, J=2.6 Hz, 2H), 4.17-4.06 (m, 2H), 3.80 (dd, J=13.4, 2.0 Hz, 1H), 3.41-3.34 (m, 2H), 3.10 (s, 6H), 2.70-2.58 (m, 2H), 2.43-2.34 (m, 2H), 2.34-2.14 (m, 4H), 2.05-1.90 (m, 2H), 1.66-1.44 (m, 6H), 0.97-0.81 (m, 9H).

##STR00151##

Compound 41B and Compound 65: N,N-dimethyl-4-oxo-N-(prop-2-yn-1-yl)-4-(((2S,3R,4S,5S)-3,4,5-tris(butyryloxy)tetrahydro-2H-pyran-2-yl)oxy)butan-1-aminium bromide

[0485] This compound was synthesized according to the experimental procedure described for Compound 41. The title compound (30 mg, 48.1 μmol, 4.9% yield, Br) was obtained as a yellow oil. LCMS (M.sup.+): 512.3 .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 5.81 (d, J=7.8 Hz, 1H), 5.40-5.28 (m, 1H), 5.27-5.08 (m, 2H), 4.43-4.35 (m, 2H), 4.15-3.84 (m, 4H), 3.40-3.31 (m, 2H), 3.09 (s, 6H), 2.49-2.14 (m, 8H), 1.97-1.90 (m, 2H), 1.66-1.42 (m, 6H), 0.93 (t, J=7.4 Hz, 3H), 0.92-0.80 (m, 6H).

##STR00152##

Compound 42: N,N-dimethyl-4-oxo-N-(prop-2-yn-1-yl)-4-(((3R,4R,5S,6S)-3,4,5-tris(butyryloxy)-6-methyltetrahydro-2H-pyran-2-yl)oxy)butan-1-aminium bromide

[0486] This compound may be synthesized according to the experimental procedure described for Compound 38.

##STR00153##

Compound 43: N,N-dimethyl-4-oxo-N-(prop-2-yn-1-yl)-4-(((2S,3S,4R,5R,6S)-3,4,5-tris(butyryloxy)-6-methyltetrahydro-2H-pyran-2-yl)oxy)butan-1-aminium bromide

[0487] Step 1:

[0488] (3S,4R,5S,6S)-6-methyltetrahydro-2H-pyran-2,3,4,5-tetraol (5 g, 30.46 mmol, 1 eq) was dissolved in Pyridine (50 mL) and butanoyl butanoate (28.91 g, 182.75 mmol, 29.90 mL, 6 eq) was added. The mixture was stirred at 25° C. for 12 h. TLC indicated the starting material was consumed completely. The reaction mixture was concentrated under reduced pressure to give a residue. (3S,4R,5R,6S)-6-methyltetrahydro-2H-pyran-2,3,4,5-tetrayl tetrabutyrate (12.8 g, crude) was obtained as yellow oil.

[0489] Step 2:

[0490] To a solution of (3S,4R,5R,6S)-6-methyltetrahydro-2H-pyran-2,3,4,5-tetrayl tetrabutyrate (12.5 g, 28.12 mmol, 1 eq) in THF (100 mL) was added MeNH.sub.2 aq. solution (5.24 g, 50.62 mmol, 30% purity, 1.8 eq). The mixture was stirred at 25° C. for 12 h. TLC indicated the starting material was consumed completely. The reaction mixture was concentrated under reduced pressure to give a residue as colorless oil. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=10/1 to 3:1). (3S,4R,5R,6S)-2-hydroxy-6-methyltetrahydro-2H-pyran-3,4,5-triyl tributyrate (4.0 g, crude) was obtained as a colorless oil.

[0491] Step 3:

[0492] To a solution of (3S,4R,5R,6S)-2-hydroxy-6-methyltetrahydro-2H-pyran-3,4,5-triyl tributyrate (1 g, 2.67 mmol, 1 eq) in DCM (10 mL) was added 4-bromobutanoyl chloride (1.49 g, 8.01 mmol, 928.66 uL, 3 eq) and Pyridine (1.06 g, 13.35 mmol, 1.08 mL, 5 eq). The mixture was stirred at 25° C. for 12 h. TLC indicated the reaction was completed. The reaction mixture was concentrated under reduced pressure to give a residue as yellow oil. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=15:1 to 5:1). (3S,4R,5R,6S)-2-((4-bromobutanoyl)oxy)-6-methyltetrahydro-2H-pyran-3,4,5-triyl tributyrate (600 mg, crude) was obtained as a yellow oil.

[0493] Step 4:

[0494] To a solution of (3S,4R,5R,6S)-2-((4-bromobutanoyl)oxy)-6-methyltetrahydro-2H-pyran-3,4,5-triyl tributyrate (0.3 g, 573.16 umol, 1 eq) in acetone (10 mL) was added N,N-dimethylprop-2-yn-1-amine (142.94 mg, 1.72 mmol, 182.33 uL, 3 eq). The mixture was stirred at 90° C. for 2 h. LCMS showed the desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue as brown liquid. The residue was purified by prep-HPLC (column: Xbridge 150*30 mm*10 um; mobile phase: [water (0.1% TFA)-ACN]) to give the crude product (0.12 g, 227.86 umol, 39.75% yield) as a brown liquid. Then the crude product was purified twice by prep-HPLC (column: Luna C18 100*30 5u; mobile phase: [water (0.1% TFA)-ACN]) to afford the title compound (9.5 mg, 16 μmol, 2.7% yield) and the anomer (Compound 45) as yellow oils. LCMS (M.sup.+): 526.3 .sup.1H NMR (400 MHz, Chloroform-d) δ 5.60 (d, J=8.3 Hz, 1H), 5.28-5.18 (m, 2H), 5.05 (dd, J=10.4, 3.5 Hz, 1H), 4.43 (s, 2H), 3.90 (q, J=6.4 Hz, 1H), 3.57-3.53 (m, 2H), 3.25 (s, 6H), 2.81 (s, 1H), 2.52-2.46 (m, 2H), 2.36 (td, J=7.3, 2.2 Hz, 2H), 2.23-2.09 (m, 4H), 2.02 (s, 2H), 1.71-1.59 (m, 2H), 1.57-1.44 (m, 4H), 1.15 (d, J=6.4 Hz, 3H), 0.93 (t, J=7.4 Hz, 3H), 0.84 (t, J=7.4 Hz, 6H).

##STR00154##

Compound 44: N,N-dimethyl-4-oxo-N-(prop-2-yn-1-yl)-4-(((2R,3S,4R,5R,6S)-3,4,5-tris(butyryloxy)-6-methyltetrahydro-2H-pyran-2-yl)oxy)butan-1-aminium bromide

[0495] This compound was synthesized according to the experimental procedure described for Compound 44. The title compound (19 mg, 31 μmol, 5.5% yield) was isolated as a yellow oil. LCMS (M.sup.+): 526.3 .sup.1H NMR (400 MHz, Chloroform-d) δ 6.38 (s, 1H), 5.41-5.35 (m, 3H), 4.54-4.50 (m, 2H), 4.33-4.26 (m, 1H), 3.75-3.71 (m, 2H), 3.35 (s, 6H), 2.89 (s, 1H), 2.67-2.62 (m, 2H), 2.44 (t, J=7.3 Hz, 2H), 2.21 (dd, J=24.3, 16.5 Hz, 6H), 1.80-1.66 (m, 2H), 1.68-1.57 (m, 4H), 1.18 (d, J=6.1 Hz, 3H), 1.02 (t, J=7.4 Hz, 3H), 0.93 (t, J=7.3 Hz, 6H).

##STR00155##

Compound 45: N,N-dimethyl-4-oxo-N-(prop-2-yn-1-yl)-4-(((3R,4S,5R,6R)-3,4,5-triacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)butan-1-aminium bromide

[0496] Step 1:

[0497] To a mixture of [(2R,3R,4S,5R)-3,4,5-triacetoxy-6-hydroxy-tetrahydropyran-2-yl]methyl acetate (2 g, 5.74 mmol, 1 eq) and Pyridine (2.27 g, 28.71 mmol, 2.32 mL, 5 eq) in DCM (10 mL) was added 4-bromobutanoyl chloride (3.19 g, 17.23 mmol, 2.00 mL, 3 eq) in one portion at 25° C. under N.sub.2. The mixture was stirred at 25° C. for 12 hours. TLC indicated SM 1 was consumed completely and one new spot formed. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=5/1 to 2:1). Desired compound [(3R,4S,5R,6R)-3,4,5-triacetoxy-6-(acetoxymethyl)tetrahydropyran-2-yl] 4-bromobutanoate (1.5 g, crude) was obtained as colorless oil.

[0498] Step 2:

[0499] To a mixture of [(3R,4S,5R,6R)-3,4,5-triacetoxy-6-(acetoxymethyl)tetrahydropyran-2-yl] 4-bromobutanoate (0.2 g, 402.18 umol, 1 eq) and N,N-dimethylprop-2-yn-1-amine (100.30 mg, 1.21 mmol, 127.94 uL, 3 eq) in Acetone (3 mL) was stirred at 90° C. for 2 hours. LC-MS showed the reaction was completed. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC [water (0.1% TFA)-ACN]. The title compound (0.039 g, 67.2 umol, 16.7% yield, mixture of anomers, Br) was obtained as colorless oil. LCMS (M.sup.+): 500.2 .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 6.30-5.92 (m, 1H), 5.52-5.21 (m, 1H), 5.17-4.90 (m, 2H), 4.43-4.36 (m, 2H), 4.28-4.10 (m, 2H), 4.09-3.97 (m, 2H), 3.45-3.32 (m, 2H), 3.14-3.07 (m, 6H), 2.75-2.43 (m, 2H), 2.08-1.89 (m, 14H).

##STR00156##

Compound 46 and Compound 53: N,N-dimethyl-4-oxo-N-(prop-2-yn-1-yl)-4-(((3R,4S,5R)-3,4,5-triacetoxytetrahydro-2H-pyran-2-yl)oxy)butan-1-aminium bromide

[0500] This compound may be synthesized according to the experimental procedure described for Compound 45.

##STR00157##

Compound 47: N,N-dimethyl-4-oxo-N-(prop-2-yn-1-yl)-4-(((3R,4R,5R)-3,4,5-triacetoxytetrahydro-2H-pyran-2-yl)oxy)butan-1-aminium bromide

[0501] This compound may be synthesized according to the experimental procedure described for Compound 45.

##STR00158##

Compound 48 and Compound 64: N,N-dimethyl-4-oxo-N-(prop-2-yn-1-yl)-4-(((3R,4S,5S)-3,4,5-triacetoxytetrahydro-2H-pyran-2-yl)oxy)butan-1-aminium bromide

[0502] Step 1

[0503] [(3S,4S,5R)-4,5-diacetoxy-6-hydroxy-tetrahydropyran-3-yl] acetate (200 mg, 724.01 umol, 1 eq) and 4-bromobutanoyl chloride (402.80 mg, 2.17 mmol, 251.75 uL, 3 eq) was dissolved with DCM (2 mL). Then Pyridine (286.35 mg, 3.62 mmol, 292.19 uL, 5 eq) was added into the mixture and stirred for 12 h at 25° C. TLC showed the starting reactant was consumed. The reaction mixture was concentrated. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=10/1). [(3R,4S,5S)-3,4,5-triacetoxytetrahydropyran-2-yl] 4-bromobutanoate (200 mg, 400 umol, 55% yield, 85% purity) was obtained as a yellow oil.

[0504] Step 2

[0505] To a solution of [(3R,4S,5S)-3,4,5-triacetoxytetrahydropyran-2-yl] 4-bromobutanoate (200 mg, 470.34 umol, 1 eq) in acetone (5 mL) was added N,N-dimethylprop-2-yn-1-amine (117.30 mg, 1.41 mmol, 149.62 uL, 3 eq) and stirred at 90° C. for 2 h. LCMS showed the product formed and the starting reactant was consumed. The mixture reaction was concentrated. The residue was purified by prep-HPLC (column: Luna C18 100*30 5u; mobile phase: [water (0.1% TFA)-ACN]; B %: 1%-30%, 12 min). The title compound (30 mg, 63.0 μmol, 13.4% yield, mixture of anomers) was obtained as a colorless oil. LCMS (M.sup.+): 428.1 .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 6.26-5.75 (m, 1H), 5.37-5.23 (m, 2H), 5.22-5.04 (m, 1H), 4.42-4.35 (m, 2H), 4.16-3.77 (m, 3H), 3.43-3.30 (m, 2H), 3.12-3.06 (m, 6H), 2.66-2.57 (m, 2H), 2.19-1.94 (m, 11H).

##STR00159##

Compound 49: N,N-dimethyl-4-oxo-N-(prop-2-yn-1-yl)-4-(((3R,4R,5S,6S)-3,4,5-triacetoxy-6-methyltetrahydro-2H-pyran-2-yl)oxy)butan-1-aminium bromide

[0506] Step 1

[0507] To a solution of [(2S,3S,4R,5R)-4,5-diacetoxy-6-hydroxy-2-methyl-tetrahydropyran-3-yl] acetate (1 g, 3.45 mmol, 1 eq) in DCM (10 mL) was added pyridine (1.36 g, 17.23 mmol, 1.39 mL, 5 eq) and 4-bromobutanoyl chloride (1.92 g, 10.34 mmol, 1.20 mL, 3 eq) at 25° C. The mixture was stirred at 25° C. for 12 hr. TLC indicated new spots formed. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=15:1 to 5:1) to give [(3R,4R,5S,6S)-3,4,5-triacetoxy-6-methyl-tetrahydropyran-2-yl] 4-bromobutanoate (1.2 g, 2.73 mmol, 79.3% yield) as a yellow oil.

[0508] Step 2

[0509] To a solution of [(3R,4R,5S,6S)-3,4,5-triacetoxy-6-methyl-tetrahydropyran-2-yl] 4-bromobutanoate (200 mg, 455.32 umol, 1 eq) in acetone (5 mL) was added N,N-dimethylprop-2-yn-1-amine (113.55 mg, 1.37 mmol, 144.84 uL, 3 eq) at 25° C. The mixture was stirred at 90° C. for 4 hr. LCMS showed the desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (column: Luna C18 100*30 5u; mobile phase: [water (0.1% TFA)-ACN]; B %: 5%-35%, 12 min) to afford the title compound (71 mg, 132 μmol, 30.0% yield, mixture of anomers) as a yellow solid. LCMS (M.sup.+): 442.2 .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 5.97-5.74 (m, 1H), 5.18-4.71 (m, 3H), 4.39-4.32 (m, 2H), 4.07-3.88 (m, 2H), 3.40-3.29 (m, 2H), 3.10-3.03 (m, 6H), 2.67-2.50 (m, 2H), 2.19-1.86 (m, 11H), 1.13-0.98 (m, 3H).

##STR00160##

Compound 50: N,N-dimethyl-4-oxo-N-(prop-2-yn-1-yl)-4-(((3S,4R,5R,6S)-3,4,5-triacetoxy-6-methyltetrahydro-2H-pyran-2-yl)oxy)butan-1-aminium bromide

[0510] Step 1

[0511] (3S,4R,5S,6S)-6-methyltetrahydropyran-2,3,4,5-tetrol (5 g, 30.46 mmol, 1 eq) was dissolved with Pyridine (50 mL) and acetyl acetate (18.66 g, 182.76 mmol, 17.12 mL, 6 eq) was added to the mixture. The mixture was stirred at 25° C. for 12 h. TLC showed the reaction was completed. The mixture was concentrated. [(2S,3R,4R,5S)-4,5,6-triacetoxy-2-methyl-tetrahydropyran-3-yl] acetate (10 g, crude) was obtained as yellow oil.

[0512] Step 2

[0513] To a solution of [(2S,3R,4R,5S)-4,5,6-triacetoxy-2-methyl-tetrahydropyran-3-yl] acetate (10 g, 30.09 mmol, 1 eq) in THF (50 mL) was added MeNH.sub.2 aq. solution (5.61 g, 54.17 mmol, 30% purity, 1.8 eq) and stirred at 25° C. for 12 h. TLC showed the starting reactant consumed. The mixture reaction was concentrated. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=10/1 to 5:1). [(2S,3R,4R,5S)-4,5-diacetoxy-6-hydroxy-2-methyl-tetrahydropyran-3-yl] acetate (6 g, 18.60 mmol, 61.8% yield, 90% purity) was obtained as yellow oil.

[0514] Step 3

[0515] To a solution of [(2S,3R,4R,5S)-4,5-diacetoxy-6-hydroxy-2-methyl-tetrahydropyran-3-yl] acetate (1 g, 3.45 mmol, 1 eq) and 4-bromobutanoyl chloride (1.66 g, 8.96 mmol, 1.04 mL, 2.6 eq) in DCM (20 mL) was added Pyridine (1.36 g, 17.23 mmol, 1.39 mL, 5 eq). The mixture was stirred at 25° C. for 12 h. TLC showed the starting reactant consumed. The mixture was concentrated. The residue was purified by flash silica gel chromatography (PE:EA=5:1). [(3S,4R,5R,6S)-3,4,5-triacetoxy-6-methyl-tetrahydropyran-2-yl] 4-bromobutanoate (1.93 g, crude) was obtained as yellow oil.

[0516] Step 4

[0517] To a solution of [(3S,4R,5R,6S)-3,4,5-triacetoxy-6-methyl-tetrahydropyran-2-yl] 4-bromobutanoate (1.83 g, 4.17 mmol, 1 eq) in acetone (20 mL) was added N,N-dimethylprop-2-yn-1-amine (1.04 g, 12.50 mmol, 1.33 mL, 3 eq). The mixture was stirred at 90° C. for 12 h. LCMS showed the starting reactant was consumed. The mixture was concentrated. The residue was purified by prep-HPLC (column: Xbridge 150*30 mm*10 um; mobile phase: [water (0.1% TFA)-ACN]; B %: 1%-60%, 10 min). The title compound (100.9 mg, 183.5 μmol, 4.4% yield, mixture of anomers, Br) was obtained as a colorless oil. LCMS (M.sup.+): 442.2 .sup.1H NMR (400 MHz, Chloroform-d) δ 6.4-6.3 (m, 1H), 5.4-5.3 (m, 3H), 4.6-4.4 (m, 2H), 4.3-4.2 (m, 1H), 3.8-3.6 (m, 2H), 3.34 (s, 6H), 2.9 (s, 1H), 2.7-2.5 (m, 2H), 2.2-2.0 (m, 11H), 1.16 (d, 3H).

##STR00161##

Compound 51: dimethyl(prop-2-yn-1-yl)(2-{[(2R,3R,4S,5R)-3,4,5-tris(butanoyloxy)oxan-2-yl]oxy}ethyl)azanium

[0518] This compound may be synthesized according to the experimental procedure described for Compound 1.

##STR00162##

Compound 69: N,N′-((((3R,3aR,6S,6aR)-hexahydrofuro[3,2-b]furan-3,6-diyl)bis(oxy))bis(2-oxoethane-2,1-diyl))bis(N,N-dimethylprop-2-yn-1-aminium) bistrifluoroacetate

[0519] Step 1:

[0520] To a solution of (3R,3aR,6S,6aR)-2,3,3a,5,6,6a-hexahydrofuro[3,2-b]furan-3,6-diol (500 mg, 3.42 mmol, 1 eq) in CHCl3 (5 mL) was added pyridine (1.62 g, 20.53 mmol, 1.66 mL, 6 eq) and (2-chloroacetyl) 2-chloroacetate (2.34 g, 13.69 mmol, 4 eq) at 0° C. The mixture was stirred at 15° C. for 12 h. TLC indicated new spots formed. The reaction mixture was quenched by addition aq. HCl (1M, 10 mL) and extracted with ethyl acetate (20 mL*3). The combined organic layers were washed with brine (20 mL), dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=20/1 to 5:1). Compound [(3R,3aR,6S,6aR)-6-(2-chloroacetyl)oxy-2,3,3a,5,6,6a-hexahydrofuro[3,2-b]furan-3-yl] 2-chloroacetate (750 mg, 2.51 mmol, 73.29% yield) was obtained as a yellow oil.

[0521] Step 2:

[0522] To a solution of [(3R,3aR,6S,6aR)-6-(2-chloroacetyl)oxy-2,3,3a,5,6,6a-hexahydrofuro[3,2-b] furan-3-yl]2-chloroacetate (550 mg, 1.84 mmol, 1 eq) in ACN (3 mL) was added N,N-dimethylprop-2-yn-1-amine (336.30 mg, 4.05 mmol, 428.95 uL, 2.2 eq) at 15° C. The mixture was stirred at 80° C. for 4 h. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (column: Xbridge 150*30 mm*10 um; mobile phase: [water (0.1% TFA)-ACN]; B %:1%-20%, 10 min). The title compound (190 mg, 305.22 umol, 16.60% yield, 2CF.sub.3COO—)—) was obtained as a colorless oil. LCMS: (M+/2): 197.1 .sup.1H NMR (400 MHz, Methanol-d4) δ 5.39-5.33 (m, 2H), 4.93 (t, J=5.4 Hz, 1H), 4.62-4.45 (m, 8H), 4.16-3.88 (m, 4H), 3.64 (t, J=2.5 Hz, 2H), 3.38 (d, J=2.8 Hz, 12H).

##STR00163##

Compound 70: N-(2-(((3S,3aR,6R,6aR)-6-hydroxyhexahydrofuro[3,2-b]furan-3-yl)oxy)-2-oxoethyl)-N,N-dimethylprop-2-yn-1-aminium trifluoroacetate

[0523] Step 1:

[0524] To a solution of (3R,3aR,6S,6aR)-2,3,3a,5,6,6a-hexahydrofuro[3,2-b]furan-3,6-diol (300 mg, 2.05 mmol, 1 eq) in ACN (8 mL) was added K.sub.2CO.sub.3 (283.71 mg, 2.05 mmol, 1 eq) and 2-chloroacetyl chloride (231.85 mg, 2.05 mmol, 163.27 uL, 1 eq) at 0° C. Then the mixture was stirred at 15° C. for 2 h under N.sub.2. TLC indicated one major new spot was detected. Compound [(3S,3aR,6R,6aR)-3-hydroxy-2,3,3a,5,6,6a-hexahydrofuro[3,2-b]furan-6-yl] 2-chloroacetate was obtained as liquid (˜2.05 mmol, 8 mL in ACN). The crude product was used into the next step without further purification.

[0525] Step 2:

[0526] To a solution of [(3S,3aR,6R,6aR)-3-hydroxy-2,3,3a,5,6,6a-hexahydrofuro[3,2-b]furan-6-yl] 2-chloroacetate (2.05 mmol, 8 mL in ACN, 1 eq) was added N,N-dimethylprop-2-yn-1-amine (170.65 mg, 2.05 mmol, 217.67 uL, 1 eq). The mixture was stirred at 70° C. for 10 h under N.sub.2. LCMS showed expected mass was detected. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (column: Xbridge 150*30 mm*10 um; mobile phase: [water (0.1% TFA)-ACN]; B %: 1%-10%, 10 min) to give N-(2-(((3S,3aR,6R,6aR)-6-hydroxyhexahydrofuro[3,2-b]furan-3-yl)oxy)-2-oxoethyl)-N,N-dimethylprop-2-yn-1-aminium trifluoroacetate (34 mg, 107.86 umol, 5.25% yield, CF.sub.3COO—) as colorless oil. LCMS: (M+) 270.1 .sup.1H NMR (400 MHz, Methanol-d4) δ 5.33 (t, J=2.3 Hz, 1H), 4.59 (t, J=2.7 Hz, 4H), 4.53 (s, 2H), 4.41-4.22 (m, 1H), 4.08 (d, J=2.3 Hz, 2H), 3.89 (dd, J=8.9, 6.3 Hz, 1H), 3.68-3.62 (m, 1H), 3.55 (dd, J=8.9, 7.0 Hz, 1H), 3.41 (s, 6H).

##STR00164##

Compound 71: N,N′-((((3R,3aR,6S,6aR)-hexahydrofuro[3,2-b]furan-3,6-diyl)bis(oxy))bis(ethane-2,1-diyl))bis(N,N-dimethylprop-2-yn-1-aminium) bistrifluoroacetate

[0527] Step 1:

[0528] To a solution of (3R,3aR,6S,6aR)-2,3,3a,5,6,6a-hexahydrofuro[3,2-b]furan-3,6-diol (4 g, 27.37 mmol, 1 eq) in DMF (40 mL) was added NaH (3.28 g, 82.11 mmol, 60% purity, 3 eq) at 0° C. for 0.5 h. Then to the mixture was added 2-bromoethoxy-tert-butyl-dimethyl-silane (19.64 g, 82.11 mmol, 3 eq). The mixture was stirred at 15° C. for 11.5 h. TLC showed the starting reactant was consumed and three new spots formed. The reaction mixture was quenched by addition of H.sub.2O (30 mL) at 0° C., and extracted with EtOAc (30 mL*3). Then the organic layer was dried over Na.sub.2SO.sub.4 and concentrated under reduced pressure. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=50:1 to 20:1). Compound 2-[[(3R,3aR,6S,6aR)-3-[2-[tert-butyl(dimethyl)silyl]oxyethoxy]-2,3,3a,5,6,6a-hexahydrofuro[3,2-b]furan-6-yl]oxy]ethoxy-tert-butyl-dimethyl-silane (3.35 g, 7.24 mmol, 26.45% yield) was obtained as yellow oil.

[0529] Step 2:

[0530] To a solution of 2-[[(3R,3aR,6S,6aR)-3-[2-[tert-butyl(dimethyl)silyl]oxyethoxy]-2,3,3a,5,6,6a-hexahydrofuro[3,2-b]furan-6-yl]oxy]ethoxy-tert-butyl-dimethyl-silane (2.3 g, 4.97 mmol, 1 eq) in THF (20 mL) was added pyridine.hydrofluoride (2.81 g, 19.88 mmol, 2.56 mL, 70%, 4 eq). The mixture was stirred at 15° C. for 12 h. TLC showed the starting reactant was consumed and one new spot formed. The mixture was concentrated under reduced pressure. The residue was purified by column chromatography (SiO.sub.2, Petroleum Ether:Ethyl Aacetate=0:1 to EA:EtOH=20:1). Compound 2-[[(3R,3aR,6S,6aR)-6-(2-hydroxyethoxy)-2,3,3a,5,6,6a-hexahydrofuro[3,2-b]furan-3-yl]oxy]ethanol (800 mg, 3.42 mmol, 68.72% yield) was obtained as yellow oil.

[0531] Step 3:

[0532] To a solution of 2-[[(3R,3aR,6S,6aR)-6-(2-hydroxyethoxy)-2,3,3a,5,6,6a-hexahydrofuro[3,2-b] furan-3-yl]oxy]ethanol (1 g, 4.27 mmol, 1 eq) in THF (10 mL) was added imidazole (1.16 g, 17.08 mmol, 4 eq) and PPh3 (4.48 g, 17.08 mmol, 4 eq). Then to the mixture was added iodine (4.33 g, 17.08 mmol, 3.44 mL, 4 eq). The mixture was stirred at 15° C. for 2 h. LCMS showed the starting reactant was consumed. The reaction mixture was quenched by addition 15% aq. Na.sub.2S2O.sub.4 (10 mL) at 0° C., and extracted with EtOAc (10 mL*3). Then the organic layer was dried over Na.sub.2SO.sub.4 and concentrated reduced pressure. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=50/1 to 20:1). Compound (3R,3aR,6S,6aR)-3,6-bis(2-iodoethoxy)-2,3,3a,5,6,6a-hexahydrofuro[3,2-b]furan (1.2 g, 2.64 mmol, 61.91% yield) was obtained as yellow oil.

[0533] Step 4:

[0534] To a solution of N,N-dimethylprop-2-yn-1-amine (1.10 g, 13.21 mmol, 1.40 mL, 6 eq) in acetone (10 mL) was added (3R,3aR,6S,6aR)-3,6-bis(2-iodoethoxy)-2,3,3a,5,6,6a-hexahydrofuro[3,2-b] furan (1 g, 2.20 mmol, 1 eq). The mixture was stirred at 90° C. for 4 h. LCMS showed the starting reactant was consumed. The mixture was concentrated under reduced pressure. The residue was purified by prep-HPLC (column: Xbridge 150*30 mm*10 um; mobile phase: [water (0.1% TFA)-ACN]; B %: 1%-60%, 10 min). N,N′-((((3R,3aR,6S,6aR)-hexahydrofuro[3,2-b]furan-3,6-diyl)bis(oxy))bis(ethane-2,1-diyl))bis(N,N-dimethylprop-2-yn-1-aminium) bistrifluoroacetate (470 mg, 793.22 umol, 36.02% yield, 100% purity, 2CF3CO2-) was obtained as colorless oil. LCMS: (M.sup.2+/2): 183.2 .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 4.63 (t, J=4.6 Hz, 1H), 4.51 (d, J=4.5 Hz, 1H), 4.39 (dd, J=11.9, 2.5 Hz, 4H), 4.13-4.01 (m, 4H), 4.01-3.83 (m, 5H), 3.81 (dd, J=8.9, 6.5 Hz, 1H), 3.72 (dd, J=10.3, 3.6 Hz, 1H), 3.68-3.51 (m, 4H), 3.45 (dd, J=8.9, 7.1 Hz, 1H), 3.14-3.07 (m, 12H).

##STR00165##

Compound 72: 343: N-(2-(((3R,3aR,6S,6aR)-6-hydroxyhexahydrofuro[3,2-b]furan-3-yl)oxy)ethyl)-N,N-dimethylprop-2-yn-1-aminium trifluoroacetate

[0535] Step 1:

[0536] To a solution of (3R,3aR,6S,6aR)-6-benzyloxy-2,3,3a,5,6,6a-hexahydrofuro[3,2-b]furan-3-ol (1.9 g, 8.04 mmol, 1 eq) in DMF (30 mL) was added NaH (482.47 mg, 12.06 mmol, 60% purity, 1.5 eq) at 0° C. and stirred for 0.5 h. Then 2-bromoethoxy-tert-butyl-dimethyl-silane (2.89 g, 12.06 mmol, 1.5 eq) was added to the mixture and stirred at 15° C. for 9.5 h. TLC indicated three new spots formed. The reaction mixture was quenched by addition of H.sub.2O (40 mL) at 0° C., and then diluted with EtOAc (40 mL) and extracted with EtOAc (40 mL*3). The combined organic layers were dried over Na.sub.2SO.sub.4, filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=20/1 to 0:1) to give 2-[[(3R,3aR,6S,6aR)-6-benzyloxy-2,3,3a,5,6,6a-hexahydrofuro[3,2-b] furan-3-yl]oxy]ethoxy-tert-butyl-dimethyl-silane (1.9 g, 4.82 mmol, 59.88% yield) as yellow oil.

[0537] Step 2:

[0538] To a solution of 2-[[(3R,3aR,6S,6aR)-6-benzyloxy-2,3,3a,5,6,6a-hexahydrofuro[3,2-b] furan-3-yl]oxy]ethoxy-tert-butyl-dimethyl-silane (1.9 g, 4.82 mmol, 1 eq) in THF (20 mL) was added pyridine; hydrofluoride (2.05 g, 14.45 mmol, 1.86 mL, 70% purity, 3 eq). The mixture was stirred at 15° C. for 10 hr under N.sub.2. TLC indicated one new spot formed. The reaction mixture was filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=10/1 to 0/1) to give 2-[[(3R,3aR,6S,6aR)-6-benzyloxy-2,3,3a,5,6,6a-hexahydrofuro[3,2-b]furan-3-yl]oxy]ethanol (1.1 g, 3.92 mmol, 81.49% yield) as yellow oil.

[0539] Step 3:

[0540] To a solution of 2-[[(3R,3aR,6S,6aR)-6-benzyloxy-2,3,3a,5,6,6a-hexahydrofuro[3,2-b] furan-3-yl]oxy]ethanol (1.1 g, 3.92 mmol, 1 eq) in THF (50 mL) was added PPh3 (2.32 g, 8.83 mmol, 2.25 eq) and imidazole (801.44 mg, 11.77 mmol, 3 eq). Then 12 (2.49 g, 9.81 mmol, 1.98 mL, 2.5 eq) was added to the mixture and stirred at 15° C. for 2 hr. TLC indicated one new spot was detected. The reaction mixture was filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=20/1 to 0/1) to give (3R,3aR,6S,6aR)-6-benzyloxy-3-(2-iodoethoxy)-2,3,3a,5,6,6a-hexahydrofuro[3,2-b]furan (1.5 g, 3.84 mmol, 97.96% yield) as yellow oil.

[0541] Step 4:

[0542] A mixture of (3R,3aR,6S,6aR)-6-benzyloxy-3-(2-iodoethoxy)-2,3,3a,5,6,6a-hexahydrofuro [3,2-b]furan (1 g, 2.56 mmol, 1 eq) in THF (10 mL) was dropped to dimethylamine (2 M, 10.00 mL, 7.80 eq, THF) and degassed and purged with N.sub.2 3 times, and then the mixture was stirred at 15° C. for 10 hr under N.sub.2 atmosphere. TLC indicated one new spot was detected. LCMS showed desired compound was detected. The reaction mixture was quenched by addition of H.sub.2O (10 mL) at 15° C., and then diluted with EtOAc (10 mL) and extracted with EtOAc (10 mL*3). The combined organic layers were dried over Na.sub.2SO.sub.4, filtered and the filtrate was concentrated under reduced pressure to give 2-[[(3R,3aR,6S,6aR)-6-benzyloxy-2,3,3a,5,6,6a-hexahydrofuro[3,2-b]furan-3-yl]oxy]-N,N-dimethyl-ethanamine (0.8 g, crude) as yellow oil. LCMS: (M+H+) 308.1

[0543] Step 5:

[0544] To a solution of 2-[[(3R,3aR,6S,6aR)-6-benzyloxy-2,3,3a,5,6,6a-hexahydrofuro[3,2-b]furan-3-yl]oxy]-N,N-dimethyl-ethanamine (0.9 g, 2.93 mmol, 1 eq) in EtOH (100 mL) was added Pd(OH)2/C (0.9 g, 20% purity). The mixture was stirred at 80° C. for 12 hr under H2 (50 Psi). TLC indicated one major new spot was detected. The reaction mixture was filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=5/1 to 0/1) to give (3S,3aR,6R,6aR)-6-[2-(dimethylamino)ethoxy]-2,3,3a,5,6,6a-hexahydrofuro [3,2-b]furan-3-ol (0.5 g, 2.30 mmol, 78.60% yield) as yellow oil.

[0545] Step 6:

[0546] To a solution of (3S,3aR,6R,6aR)-6-[2-(dimethylamino)ethoxy]-2,3,3a,5,6,6a-hexahydrofuro [3,2-b]furan-3-ol (500 mg, 2.30 mmol, 1 eq) in ACN (10 mL) was added 3-bromoprop-1-yne (301.15 mg, 2.53 mmol, 218.22 uL, 1.1 eq). The mixture was stirred at 15° C. for 5 hr under N.sub.2. LCMS showed desired compound was detected. The reaction mixture was filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (column: Waters Xbridge Prep OBD C18 100*19 mm*5 um; mobile phase: [water (0.1% TFA)-ACN]; B %: 1%-10%, 10 min) to N-(2-(((3R,3aR,6S,6aR)-6-hydroxyhexahydrofuro[3,2-b]furan-3-yl)oxy)ethyl)-N,N-dimethylprop-2-yn-1-aminium trifluoroacetate (290 mg, 785.20 umol, 34.12% yield, CF.sub.3COO—) as colorless oil. LCMS: (M+) 256.1 1H NMR (400 MHz, Methanol-d4) δ 4.80 (t, J=4.4 Hz, 1H), 4.48 (d, J=2.6 Hz, 2H), 4.44 (dd, J=4.2, 1.2 Hz, 1H), 4.27-4.21 (m, 1H), 4.23-4.13 (m, 2H), 3.99-3.93 (m, 2H), 3.96-3.86 (m, 2H), 3.79 (ddd, J=14.2, 8.8, 2.4 Hz, 1H), 3.67 (ddd, J=14.2, 5.0, 2.2 Hz, 1H), 3.62-3.53 (m, 2H), 3.30 (d, J=1.8 Hz 6H).

##STR00166##

Compound 73: N-(2-(((3R,3aR,6S,6aR)-6-hydroxyhexahydrofuro[3,2-b]furan-3-yl)oxy)-2-oxoethyl)-N,N-dimethylprop-2-yn-1-aminium bromide

[0547] Step 1:

[0548] To a solution of (3R,3aR,6S,6aR)-6-benzyloxy-2,3,3a,5,6,6a-hexahydrofuro[3,2-b]furan-3-ol (500 mg, 2.12 mmol, 1 eq) in DCM (5 mL) was added TEA (428.29 mg, 4.23 mmol, 589.13 uL, 2 eq). Then 2-chloroacetyl chloride (262.92 mg, 2.33 mmol, 185.16 uL, 1.1 eq) was dropped to the mixture at 0° C. and stirred at 15° C. for 10 hr. TLC indicated (3R,3aR,6S,6aR)-6-benzyloxy-2,3,3a,5,6,6a-hexahydrofuro[3,2-b]furan-3-ol was consumed completely and one new spot formed. The reaction mixture was quenched by addition of H.sub.2O (5 mL) at 15° C., and then diluted with EtOAc (5 mL) and extracted with EtOAc (5 mL*3). The combined organic layers were dried over Na.sub.2SO.sub.4, filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=0/1 to 1/1) to give [(3S,3aR,6R,6aR)-3-benzyloxy-2,3,3a,5,6,6a-hexahydrofuro[3,2-b]furan-6-yl] 2-chloroacetate (230 mg, 735.42 umol, 34.75% yield) as yellow oil.

[0549] Step 2:

[0550] A mixture of [(3S,3aR,6R,6aR)-3-benzyloxy-2,3,3a,5,6,6a-hexahydrofuro[3,2-b]furan-6-yl] 2-chloroacetate (230 mg, 735.42 umol, 1 eq) in THF (5 mL) was dropped to dimethylamine (2 mL, 2M, in THF) and degassed and purged with N.sub.2 3 times. And then the mixture was stirred at 15° C. for 10 hr under N.sub.2 atmosphere. TLC indicated [(3S,3aR,6R,6aR)-3-benzyloxy-2,3,3a,5,6,6a-hexahydrofuro[3,2-b]furan-6-yl] 2-chloroacetate was consumed completely and one new spot formed. LCMS showed desired mass was detected. The reaction mixture was quenched by addition of H.sub.2O (5 mL) at 15° C., and then diluted with EtOAc (5 mL) and extracted with EtOAc (5 mL*3). The combined organic layers were dried over Na.sub.2SO.sub.4, filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by prep-TLC (SiO.sub.2, DCM:MeOH=10:1) to give [(3S,3aR,6R,6aR)-3-benzyloxy-2,3,3a,5,6,6a-hexahydrofuro[3,2-b]furan-6-yl] 2-(dimethylamino)acetate (180 mg, 503.03 umol, 68.40% yield, HCl) as yellow oil. LCMS: (M+H+) 322.1

[0551] Step 3:

[0552] A mixture of [(3S,3aR,6R,6aR)-3-benzyloxy-2,3,3a,5,6,6a-hexahydrofuro[3,2-b]furan-6-yl] 2-(dimethylamino)acetate (180 mg, 503.03 umol, 1 eq, HCl) and Pd(OH).sub.2/C (0.18 g, 20% purity) in EtOH (20 mL) was degassed and purged with H.sub.2 3 times. And then the mixture was stirred at 60° C. for 10 hr under H.sub.2 (15 Psi) atmosphere. LCMS showed desired compound was detected. The reaction mixture was filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by prep-TLC (SiO.sub.2, DCM:MeOH=10:1, 5% NH.sub.3H.sub.2O) to give compound [(3S,3aR,6R,6aR)-3-hydroxy-2,3,3a,5,6,6a-hexahydrofuro[3,2-b]furan-6-yl] 2-(dimethylamino)acetate (100 mg, 432.44 umol, 85.97% yield) as colorless oil. LCMS: (M+H+) 232.1

[0553] Step 4:

[0554] To a solution of [(3S,3aR,6R,6aR)-3-hydroxy-2,3,3a,5,6,6a-hexahydrofuro[3,2-b]furan-6-yl] 2-(dimethylamino)acetate (100 mg, 432.44 umol, 1 eq) in ACN (5 mL) was added 3-bromoprop-1-yne (56.59 mg, 475.69 umol, 41.01 uL, 1.1 eq). The mixture was stirred at 15° C. for 10 hr. LCMS showed desired mass was detected. The reaction mixture was filtered and the filter cake was concentrated under reduced pressure to give N-(2-(((3R,3aR,6S,6aR)-6-hydroxyhexahydrofuro[3,2-b]furan-3-yl)oxy)-2-oxoethyl)-N,N-dimethylprop-2-yn-1-aminium bromide (60 mg, 131.92 umol, 30.51% yield, 77% purity, Br—) as a white solid. LCMS: (M+) 270.1 .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 5.26-5.16 (m, 2H), 4.75 (t, J=5.3 Hz, 1H), 4.57-4.40 (m, 4H), 4.21 (d, J=4.8 Hz, 1H), 4.17-4.11 (m, 1H), 4.08 (t, J=3.5 Hz, 1H), 3.77 (d, J=4.2 Hz, 2H), 3.71 (d, J=9.4 Hz, 1H), 3.64 (dd, J=9.4, 3.2 Hz, 1H), 3.24 (s, 6H).

##STR00167##

Compound 74: N-(2-(((3S,3aR,6R,6aR)-6-hydroxyhexahydrofuro[3,2-b]furan-3-yl)oxy)ethyl)-N,N-dimethylprop-2-yn-1-aminium trifluoroacetate

[0555] Step 1:

[0556] To a solution of (3S,3aR,6R,6aR)-6-benzyloxy-2,3,3a,5,6,6a-hexahydrofuro[3,2-b]furan-3-ol (7 g, 29.63 mmol, 1 eq) in DMF (70 mL) was added NaH (1.78 g, 44.44 mmol, 60% purity, 1.5 eq) at 0° C. and stirred for 0.5 h. Then 2-bromoethoxy-tert-butyl-dimethyl-silane (10.63 g, 44.44 mmol, 1.5 eq) was added to the mixture and stirred at 15° C. for 11.5 h. TLC indicated (3S,3aR,6R,6aR)-6-benzyloxy-2,3,3a,5,6,6a-hexahydrofuro[3,2-b]furan-3-ol was consumed completely and two new spots formed. The reaction mixture was quenched by addition of H.sub.2O (80 mL) at 0° C., diluted with EtOAc (80 mL) and extracted with EtOAc (100 mL*4). The combined organic layers were dried over Na.sub.2SO.sub.4, filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=0/1 to 2/1) to give 2-[[(3S,3aR,6R,6aR)-6-benzyloxy-2,3,3a,5,6,6a-hexahydrofuro[3,2-b]furan-3-yl]oxy]ethoxy-tert-butyl-dimethyl-silane (4.2 g, 10.64 mmol, 35.93% yield) as yellow oil.

[0557] Step 2:

[0558] To a solution of 2-[[(3S,3aR,6R,6aR)-6-benzyloxy-2,3,3a,5,6,6a-hexahydrofuro[3,2-b] furan-3-yl]oxy]ethoxy-tert-butyl-dimethyl-silane (4.2 g, 10.64 mmol, 1 eq) in THF (40 mL) was added HF.Py (4.52 g, 31.93 mmol, 4.11 mL, 70%, 3 eq). The mixture was stirred at 15° C. for 12 hr under N.sub.2. LCMS showed desired mass was detected. TLC indicated 2-[[(3S,3aR,6R,6aR)-6-benzyloxy-2,3,3a,5,6,6a-hexahydrofuro[3,2-b]furan-3-yl]oxy]ethoxy-tert-butyl-dimethyl-silane was consumed completely and one new spot formed. The reaction mixture was filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Ethyl acetate/Methanol=1/0 to 0/1) to give 2-[[(3S,3aR,6R,6aR)-6-benzyloxy-2,3,3a,5,6,6a-hexahydrofuro[3,2-b]furan-3-yl]oxy]ethanol (1.67 g, 5.96 mmol, 55.97% yield) as yellow oil. LCMS: (M+H+) 281.1

[0559] Step 3:

[0560] To a solution of 2-[[(3S,3aR,6R,6aR)-6-benzyloxy-2,3,3a,5,6,6a-hexahydrofuro[3,2-b]furan-3-yl] oxy]ethanol (1.67 g, 5.96 mmol, 1 eq) in THF (17 mL) was added PPh3 (3.52 g, 13.40 mmol, 2.25 eq) and imidazole (1.22 g, 17.87 mmol, 3 eq). Then 12 (3.78 g, 14.89 mmol, 3.00 mL, 2.5 eq) was added to the mixture and stirred at 15° C. for 12 hr. TLC indicated two new spots formed. The reaction mixture was filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=50/1 to 2/1) to give (3S,3aR,6R,6aR)-6-benzyloxy-3-(2-iodoethoxy)-2,3,3a,5,6,6a-hexahydrofuro[3,2-b]furan (2 g, 5.13 mmol, 86.03% yield) as yellow oil.

[0561] Step 4:

[0562] A mixture of (3S,3aR,6R,6aR)-6-benzyloxy-3-(2-iodoethoxy)-2,3,3a,5,6,6a-hexahydrofuro [3,2-b]furan (2 g, 5.13 mmol, 1 eq) in THF (20 mL) was dropped to dimethylamine (20 mL, 2 M, in THF) and degassed and purged with N.sub.2 3 times, and then the mixture was stirred at 15° C. for 10 hr under N.sub.2 atmosphere. TLC indicated one new spot formed. LCMS showed desired compound was detected. The reaction mixture was quenched by addition of H.sub.2O (30 mL) at 15° C., and then diluted with EtOAc (30 mL) and extracted with EtOAc (30 mL*4). The combined organic layers were dried over Na.sub.2SO.sub.4, filtered, and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Ethyl acetate/Methanol=1/0 to 0/1) to give 2-[[(3S,3aR,6R,6aR)-6-benzyloxy-2,3,3a,5,6,6a-hexahydrofuro[3,2-b]furan-3-yl]oxy]-N,N-dimethyl-ethanamine (1.9 g, 4.36 mmol, 85.16% yield, HI) as yellow oil. LCMS: (M+H+) 308.1

[0563] Step 5:

[0564] To a solution of 2-[[(3S,3aR,6R,6aR)-6-benzyloxy-2,3,3a,5,6,6a-hexahydrofuro[3,2-b] furan-3-yl]oxy]-N,N-dimethyl-ethanamine (1.9 g, 4.36 mmol, 1 eq, HI) in EtOH (40 mL) was added Pd(OH).sub.2/C (1.4 g, 20% purity). The mixture was stirred at 80° C. for 10 hr under H.sub.2 (15 Psi). TLC indicated one new spot formed. The reaction mixture was filtered and the filtrate was concentrated under reduced pressure to give (3R,3aR,6S,6aR)-6-[2-(dimethylamino)ethoxy]-2,3,3a,5,6,6a-hexahydrofuro[3,2-b]furan-3-ol as yellow oil without further purification.

[0565] Step 6:

[0566] To a solution of (3R,3aR,6S,6aR)-6-[2-(dimethylamino)ethoxy]-2,3,3a,5,6,6a-hexahydrofuro [3,2-b]furan-3-ol (0.88 g, 4.05 mmol, 1 eq) in ACN (10 mL) was added 3-bromoprop-1-yne (530.02 mg, 4.46 mmol, 384.07 uL, 1.1 eq). The mixture was stirred at 15° C. for 10 hr. LCMS showed desired mass was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (column: Waters Xbridge BEH C18 100*30 mm*10 um; mobile phase: [water (0.1% TFA)-ACN]; B %: 1%-10%, 10 min) to give N-(2-(((3S,3aR,6R,6aR)-6-hydroxyhexahydrofuro[3,2-b]furan-3-yl)oxy)ethyl)-N,N-dimethylprop-2-yn-1-aminium trifluoroacetate (671 mg, 1.82 mmol, 44.85% yield, 100.0% purity, CF3CO2-) as yellow oil. LCMS: (M+) 256.1 1H NMR (400 MHz, Methanol-d4) δ 4.56-4.46 (m, 2H), 4.38 (d, J=2.6 Hz, 2H), 4.31-4.22 (m, 1H), 4.15-3.96 (m, 4H), 3.93-3.80 (m, 2H), 3.75-3.65 (m, 2H), 3.56 (t, J=2.5 Hz, 1H), 3.48 (dd, J=8.8, 7.2 Hz, 1H), 3.23 (s, 6H).

##STR00168##

Compound 75: (3-{[(3S,3aR,6R,6aR)-6-hydroxy-hexahydrofuro[3,2-oxopropyl)dimethyl(prop-2-yn-1-yl)azanium

[0567] This compound may be synthesized according to the experimental procedure described for Compound 344.

##STR00169##

Compound 76: (3-{[(3R,3aR,6S,6aR)-6-hydroxy-hexahydrofuro[3,2-b]furan-3-yl]oxy}-2-methylidene-3-oxopropyl)dimethyl(prop-2-yn-1-yl)azanium

[0568] This compound may be synthesized according to the experimental procedure described for Compound 344.

##STR00170##

Compound 102: N,N-dimethyl-N-(2-oxo-2-(((2S,3R,4S,5R,6R)-2,3,5-triacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-4-yl)oxy)ethyl)prop-2-yn-1-aminium trifluoroacetate

[0569] Step 1:

[0570] Tetrahydrofuran (5 mL) was added to 60% NaH (0.2 g, 5.0 mmol, 1.3 eq) in mineral oil under nitrogen, followed by addition of TBAI (71 mg, 0.22 mmol, 0.05 eq), then cooled to 0° C. Compound 1,2:5,6-Di-O-isopropylidene-α-D-glucofuranose (Sigma Aldrich, CAS: 582-52-5, 1 g, 3.8 mmol, 1 eq) in THF (3 mL) was added dropwise, followed by dropwise addition of benzyl bromide (0.55 mL, 4.6 mmol, 1.2 eq). The ice bath was removed and the reaction was warmed to 55° C., and stirred overnight. The reaction was quenched with methanol (1 mL), then filtered and loaded onto silica. Purification by column chromatography (100% hexane to 100% ethyl acetate) yielded 3-benzyl-1,2:5,6-Di-O-isopropylidene-α-D-glucofuranose as a viscous oil (877 mg, 2.5 mmol 65% yield).

[0571] Step 2:

[0572] Compound 3-benzyl-1,2:5,6-Di-O-isopropylidene-α-D-glucofuranose (877 mg, 2.5 mmol, 1 eq) was dissolved in TFA (1 mL) followed by addition of water (1 mL), then stirred overnight. The reaction solution was concentrated by rotary evaporation, and the solids were precipitated out in ethyl acetate and washed with diethyl ether. Solids were dried in vacuo to 3-benzyl-beta-D-glucopyranose as a white solid (400 mg, 1.48 mmol, 59% yield).

[0573] Step 3:

[0574] Compound 3-benzyl-beta-D-glucopyranose (400 mg, 1.48 mmol, 1 eq) was stirred in acetic anhydride (1.6 mL, 16.9 mmol, 11.4 eq) and pyridine (1.6 mL, 19.9 mmol, 13.4 eq) overnight at room temperature. The reaction was concentrated by rotary evaporation and purified by column chromatography (100% hexanes to 100% ethyl acetate) to yield 3-benzyl-1,2,4,6-tetra-O-acetyl-beta-D-glucopyranose as a waxy solid (436 mg, 0.99 mmol, 67% yield).

[0575] Step 4:

[0576] Compound 3-benzyl-1,2,4,6-tetra-O-acetyl-beta-D-glucopyranose (500 mg, 1.14 mmol, 1 eq) was dissolved in methanol (approximately 10 mL), and flowed through an H-cube hydrogenation flow system with a 10% Pd/C catalyst cartridge at 0.4 mL/min and hydrogen pressure at 30 bar. Reaction was continuously fed through the H-cube until completed as monitored by LCMS. If necessary, the cartridge was replaced. The solution was concentrated by rotary evaporation and dried in vacuo to yield 1,2,4,6-tetra-O-acetyl-beta-D-glucopyranose (170 mg, 0.49 mmol, 43% yield).

[0577] Step 5:

[0578] Compound 3-benzyl-1,2,4,6-tetra-O-acetyl-beta-D-glucopyranose (170 mg, 0.49 mmol, 1 eq) was dissolved in DCM (2 mL), followed by addition of chloroacetyl chloride (0.11 mL, 1.38 mmol, 2.8 eq) and pyridine (0.12 mL, 1.49 mmol, 3 eq). The reaction was stirred overnight, filtered, concentrated, and purified by column chromatography (100% hexanes to 100% ethyl acetate) to yield 3-(2-chloroacetoxy)-1,2,4,6-tetra-O-acetyl-beta-D-glucopyranose as a yellow viscous oil (0.19 g, 0.45 mmol, 91% yield).

[0579] Step 6:

[0580] Compound 3-(2-chloroacetoxy)-1,2,4,6-tetra-O-acetyl-beta-D-glucopyranose (0.19 g, 0.45 mmol, 1 eq) was dissolved in acetone (4 mL), followed by addition of sodium iodide (0.1 g, 0.68 mmol, 1.5 eq). The reaction was stirred for a few minutes, and 3-dimethylamino-1-propyne (0.144 mL, 1.3 mmol, 3 eq) was added. The reaction was stirred at room temperature overnight, then filtered. The filtrate was concentrated, diluted with DMSO and water, and injected onto reverse phase C18 column chromatography (0.1% TFA in 95% water/5% MeCN to 100% MeCN). Fractions containing product were lyophilized to yield an off-white powder (113 mg, 0.19 mmol, 43% yield). LCMS (M+): 472.1 .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 6.00 (d, J=8.2 Hz, 1H), 5.64 (t, J=9.5 Hz, 1H), 5.11-4.98 (m, 2H), 4.58-4.44 (m, 4H), 4.29-4.10 (m, 3H), 4.00 (dd, J=12.5, 2.3 Hz, 1H), 3.21 (s, 6H), 2.12-1.93 (m, 12H).

##STR00171##

Compound 77: (fluoromethyl)(methyl)(prop-2-yn-1-yl)(2-{[(2R,3R,4S,5R,6R)-3,4,5-tris(acetyloxy)-6-[(acetyloxy)methyl]oxan-2-yl]oxy}ethyl)azanium

[0581] Step 1:

[0582] To a mixture of [(2R,3R,4S,5R,6S)-3,4,5,6-tetraacetoxytetrahydropyran-2-yl]methyl acetate (20 g, 51.24 mmol, 1 eq) in DCM (300 mL) was added 2-bromoethanol (9.60 g, 76.86 mmol, 5.46 mL, 1.5 eq) and BF.sub.3.Et.sub.2O (36.36 g, 256.19 mmol, 31.62 mL, 5 eq) in one portion at 0° C. under N.sub.2. The mixture stirred at 25° C. for 12 hours. TLC indicated the starting material was consumed and one new spot formed. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=20/1 to 0/1). Compound [(2R,3R,4S,5R,6R)-3,4,5-triacetoxy-6-(2-bromoethoxy)tetrahydropyran-2-yl]methyl acetate (10 g, 21.97 mmol, 42.87% yield) was obtained as white solid.

[0583] Step 2:

[0584] To a mixture of [(2R,3R,4S,5R,6R)-3,4,5-triacetoxy-6-(2-bromoethoxy)tetrahydropyran-2-yl]methyl acetate (2 g, 4.39 mmol, 1 eq) and N-methylprop-2-yn-1-amine (1.67 g, 24.16 mmol, 2.01 mL, 5.5 eq) in THF (5 mL) was added Na.sub.2CO.sub.3 (931.26 mg, 8.79 mmol, 2 eq) and NaI (658.51 mg, 4.39 mmol, 1 eq) in one portion at 25° C. under at 25° C. under N.sub.2. Then mixture was heated to 70° C. and stirred for 12 hours. TLC indicated the stating material was consumed and one new spot formed. LCMS showed the expected mass was detected. The reaction mixture was filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=20/1 to 0/1). Compound[(2R,3R,4S,5R,6R)-3,4,5-triacetoxy-6-[2-[methyl(prop-2-ynyl)amino]ethoxy]tetrahydropyran-2-yl]methyl acetate (1.6 g, crude) was obtained as a yellow oil. LCMS: (M+H+): 444.3@ 0.125 min

[0585] Step 3:

[0586] To a mixture of [(2R,3R,4S,5R,6R)-3,4,5-triacetoxy-6-[2-[methyl(prop-2-ynyl)amino]ethoxy]tetrahydropyran-2-yl]methyl acetate (1 g, 2.26 mmol, 1 eq) in acetone (10 mL) was added fluoro(iodo)methane (1.80 g, 11.28 mmol, 5 eq) in one portion at 25° C. under N.sub.2. The mixture was heated to 90° C. and stirred for 2 hours. TLC indicated the starting material was consumed and one new spot formed. LCMS showed one main peak with expected mass was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=10/1 to Ethyl acetate/Methanol=3/1). Then the residue was purified by prep-HPLC [water (0.1% TFA)-ACN]. The title compound (266 mg, 441.46 umol, 19.58% yield, 98% purity, TFA) was obtained as colorless oil. LCMS: (M+): 476.2 .sup.1H NMR (400 MHz, Chloroform-d) δ 5.72 (d, J=48.1 Hz, 2H), 5.24 (t, J=9.5 Hz, 1H), 5.08 (t, J=9.4 Hz, 1H), 4.99 (t, J=8.8 Hz, 1H), 4.65-4.58 (m, 3H), 4.40 (s, 1H), 4.32-4.18 (m, 5H), 3.82-3.73 (m, 1H), 3.42 (s, 3H), 2.91 (s, 1H), 2.07 (dd, J=21.9, 12.1 Hz, 12H).

##STR00172##

Compound 78: N-(chloromethyl)-N,N-dimethyl-2-(((2S,3S,4R,5S,6S)-3,4,5-triacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)ethan-1-aminium Iodide

[0587] Step 1:

[0588] To a solution of (2S,3R,4S,5R,6R)-6-(acetoxymethyl)tetrahydro-2H-pyran-2,3,4,5-tetrayl tetraacetate (60.0 g, 1 Eq, 154 mmol) in anhydrous DCM (300 mL) at 0° C. under nitrogen, 2-bromoethan-1-ol (28.8 g, 16.3 mL, 1.5 Eq, 231 mmol) was added followed by BF.sub.3OEt.sub.2 (109 g, 97.4 mL, 5 Eq, 769 mmol). The reaction was stirred at room temperature for 18 hrs. Reaction was quenched slowly with 300 mL of H.sub.2O in an ice bath and 50 mL of saturated sodium bicarbonate was added. The layers were separated. The water layer was washed 2× with 200 mL DCM. Organic layers were combined, concentrated and residue was purified by normal phase flash chromatography (0-40% ethyl acetate in hexanes). Filtrate was concentrated and recrystallized by dissolving in minimal ethyl acetate and adding hexanes until crystals began forming, then solution was cooled to 4° C. overnight. Crystals were collected and washed with cold 1:2 ethyl acetate/hexanes followed by cold hexanes. (2R,3R,4S,5R,6R)-2-(acetoxymethyl)-6-(2-bromoethoxy)tetrahydro-2H-pyran-3,4,5-triyl triacetate (39.41 g, 56.3%) was obtained as a white crystalline solid.

[0589] Step 2:

[0590] To a stirred solution of (2S,3S,4R,5S,6S)-2-(acetoxymethyl)-6-(2-bromoethoxy)tetrahydro-2H-pyran-3,4,5-triyl triacetate (3.60 g, 1 Eq, 7.90 mmol) in 16 mL anhydrous THF, 2 M dimethylamine in THF (16.0 mL, 4.0 Eq, 32.0 mmol) was added followed by sodium iodide (1.2 g, 1 Eq, 7.9 mmol). Reaction was stirred under nitrogen at room temperature for 18 hours, filtered, and concentrated. Residue was purified by normal phase flash chromatography (0-10% methanol in DCM) to give (2S,3S,4R,5S,6S)-2-(acetoxymethyl)-6-(2-(dimethylamino)ethoxy)tetrahydro-2H-pyran-3,4,5-triyl triacetate, Iodide (1.6 g, 37%) as a yellow oil that crystallized upon standing.

[0591] Step 3:

[0592] To a stirred solution of (2S,3S,4R,5S,6S)-2-(acetoxymethyl)-6-(2-(dimethylamino)ethoxy)tetrahydro-2H-pyran-3,4,5-triyl triacetate, iodide (1.15 g, 1 Eq, 2.10 mmol) in 10 mL anhydrous DCM at room temperature, chloroiodomethane (766 μL, 5 Eq, 10.5 mmol) was added followed by DIEA (0.40 mL, 1.1 Eq, 2.32 mmol). Vial was sealed and flushed with nitrogen. Reaction was heated to 40° C. and stirred for 2.5 hours. Reaction was allowed to come to room temperature, concentrated by rotary evaporation, and purified by normal phase HPLC (0-10% methanol in DCM) to give N-(chloromethyl)-N,N-dimethyl-2-(((2S,3S,4R,5S,6S)-3,4,5-triacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)ethan-1-aminium, Iodide (250 mg, 19.9%) as a slightly yellow solid. LCMS (M.sup.+): 468.3. .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 5.33 (s, 2H), 5.28 (t, 1H), 4.99-4.81 (m, 3H), 4.19 (dd, 1H), 4.16-4.01 (m, 4H), 3.72-3.67 (m, 2H), 3.15 (s, 6H), 2.03 (s, 3H), 2.02 (s, 3H), 1.99 (s, 3H), 1.94 (s, 3H).

##STR00173##

Compound 79: N-(fluoromethyl)-N,N-dimethyl-2-(((2S,3S,4R,5S,6S)-3,4,5-triacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)ethan-1-aminium Iodide

[0593] Step 1:

[0594] To a solution of (2S,3R,4S,5R,6R)-6-(acetoxymethyl)tetrahydro-2H-pyran-2,3,4,5-tetrayl tetraacetate (60.0 g, 1 Eq, 154 mmol) in anhydrous DCM (300 mL) at 0° C. under nitrogen, 2-bromoethan-1-ol (28.8 g, 16.3 mL, 1.5 Eq, 231 mmol) was added followed by BF.sub.3OEt.sub.2 (109 g, 97.4 mL, 5 Eq, 769 mmol). The reaction was stirred at room temperature for 18 hrs. Reaction was quenched slowly with 300 mL of H.sub.2O in an ice bath and 50 mL of saturated sodium bicarbonate was added. The layers were separated. The water layer was washed 2× with 200 mL DCM. Organic layers were combined, concentrated and residue was purified by normal phase flash chromatography (0-40% ethyl acetate in hexanes). Filtrate was concentrated and recrystallized by dissolving in minimal ethyl acetate and adding hexanes until crystals began forming, then solution was cooled to 4° C. overnight. Crystals were collected and washed with cold 1:2 ethyl acetate/hexanes followed by cold hexanes. Yielded (2R,3R,4S,5R,6R)-2-(acetoxymethyl)-6-(2-bromoethoxy)tetrahydro-2H-pyran-3,4,5-triyl triacetate (39.41 g, 56.3%) as a white crystalline solid.

[0595] Step 2:

[0596] To a stirred solution of (2S,3S,4R,5S,6S)-2-(acetoxymethyl)-6-(2-bromoethoxy)tetrahydro-2H-pyran-3,4,5-triyl triacetate (2.00 g, 1 Eq, 4.39 mmol) in 8.8 mL anhydrous THF, dimethylamine (792 mg, 8.8 mL, 4.0 Eq, 17.6 mmol) was added followed by sodium iodide (658 mg, 1 Eq, 4.39 mmol). Reaction was stirred under nitrogen at room temperature for 18 hours, filtered, and concentrated. Residue was purified by normal phase flash chromatography (0-10% methanol in DCM) to give (2S,3S,4R,5S,6S)-2-(acetoxymethyl)-6-(2-(dimethylamino)ethoxy)tetrahydro-2H-pyran-3,4,5-triyl triacetate, Iodide (1.0 g, 42%) as a yellow oil that crystallized upon standing.

[0597] Step 3:

[0598] To a stirred solution of (2S,3S,4R,5S,6S)-2-(acetoxymethyl)-6-(2-(dimethylamino)ethoxy)tetrahydro-2H-pyran-3,4,5-triyl triacetate, Iodide (517 mg, 1 Eq, 946 μmol) in 15 mL anhydrous dichloromethane at room temperature, fluoroiodomethane (757 mg, 320 μL, 5 Eq, 4.73 mmol) was added followed by DIEA (135 mg, 0.18 mL, 1.1 Eq, 1.04 mmol). Reaction vessel was flushed with nitrogen, then reaction was stirred for 2 hours at 40° C. Reaction was then allowed to come to room temperature, concentrated, and purified by normal phase flash HPLC 0-10% MeOH in DCM to give N-(fluoromethyl)-N,N-dimethyl-2-(((2S,3S,4R,5S,6S)-3,4,5-triacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)ethan-1-aminium, Iodide (231 mg, 42.1%) as a light yellow solid. LC-MS (M.sup.+): 452.3. .sup.1H NMR (400 MHz, Chloroform-d) δ 5.75 (s, 1H), 5.63 (s, 1H), 5.22 (t, 1H), 5.06 (t, 1H), 4.95 (dd, 1H), 4.75-4.68 (m, 1H), 4.49-4.37 (m, 1H), 4.33-4.14 (m, 5H), 3.85 (ddd, 1H), 3.51 (t, 6H), 2.12 (s, 3H), 2.09 (d, 3H), 2.04 (d, 3H), 2.00 (s, 3H).

##STR00174##

Compound 80: N,N-dimethyl-N-(2-{[(3R,4S,5R,6R)-3,4,5-tris(acetyloxy)-6-[(acetyloxy)methyl]oxan-2-yl]oxy}ethyl)cyclopropanaminium Iodide

[0599] Step 1:

[0600] To a mixture of [(2R,3R,4S,5R,6S)-3,4,5,6-tetraacetoxytetrahydropyran-2-yl]methyl acetate (20 g, 51.24 mmol, 1 eq) in DCM (300 mL) was added 2-bromoethanol (9.60 g, 76.86 mmol, 5.46 mL, 1.5 eq) and BF.sub.3.Et.sub.2O (36.36 g, 256.19 mmol, 31.62 mL, 5 eq) in one portion at 0° C. under N.sub.2. The mixture stirred at 25° C. for 12 hours. TLC indicated the starting material was consumed and one new spot formed. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=20/1 to 0/1). Compound [(2R,3R,4S,5R,6R)-3,4,5-triacetoxy-6-(2-bromoethoxy)tetrahydropyran-2-yl]methyl acetate (10 g, 21.97 mmol, 42.87% yield) was obtained as white solid.

[0601] Step 2:

[0602] To a mixture of [(2R,3R,4S,5R,6R)-3,4,5-triacetoxy-6-(2-bromoethoxy)tetrahydropyran-2-yl]methyl acetate (2 g, 4.39 mmol, 1 eq) in THF (20 mL) was added NaI (658.51 mg, 4.39 mmol, 1 eq) and Na.sub.2CO.sub.3 (931.26 mg, 8.79 mmol, 2 eq) in one portion at 25° C. under N.sub.2. Then N-methylcyclopropanamine (2.60 g, 24.16 mmol, 5.5 eq, HCl) was added. The mixture was heated to 70° C. and stirred for 12 hours. TLC indicated [(2R,3R,4S,5R,6R)-3,4,5-triacetoxy-6-(2-bromoethoxy)tetrahydropyran-2-yl]methyl acetate was consumed and one new spot formed. The reaction mixture was filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=20/1 to 0/1). Compound[(2R,3R,4S,5R)-3,4,5-triacetoxy-6-[2-[cyclopropyl(methyl)amino]ethoxy]tetrahydropyran-2-yl]methyl acetate (1.4 g, 3.14 mmol, 71.54% yield) was obtained as yellow oil. LCMS: (M+H+): 446.3

[0603] Step 3:

[0604] To a mixture of [(2R,3R,4S,5R)-3,4,5-triacetoxy-6-[2-[cyclopropyl(methyl)amino]ethoxy]tetrahydropyran-2-yl]methyl acetate (1.4 g, 3.14 mmol, 1 eq) in Acetone (15 mL) was added MeI (2.23 g, 15.71 mmol, 978.26 uL, 5 eq) in one portion at 25° C. under N.sub.2. Then mixture was heated to 90° C. and stirred for 2 hours. TLC indicated the starting material was consumed and one new spot formed. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=10/1 to Ethyl acetate:Methanol=3/1). Compound N,N-dimethyl-N-(2-{[(3R,4S,5R,6R)-3,4,5-tris(acetyloxy)-6-[(acetyloxy)methyl]oxan-2-yl]oxy}ethyl)cyclopropanaminium iodide (1 g, 1.70 mmol, 54.17% yield, 100% purity, I) was obtained as yellow solid. LCMS: (M+): 460.2 .sup.1H NMR (400 MHz, Chloroform-d) δ 5.21 (t, J=9.6 Hz, 1H), 5.04 (t, J=9.7 Hz, 1H), 4.94 (dd, J=9.7, 8.0 Hz, 1H), 4.67 (d, J=8.0 Hz, 1H), 4.51-4.42 (m, 1H), 4.36-4.06 (m, 6H), 3.83 (dt, J=10.2, 3.5 Hz, 1H), 3.70-3.60 (m, 1H), 3.20 (d, J=9.6 Hz, 6H), 2.16-1.97 (m, 12H), 1.29-1.21 (m, 2H), 1.09-0.96 (m, 2H).

##STR00175##

Compound 81: dimethyl(prop-2-yn-1-yl)(2-{[(2R,3R,4S,5R,6R)-3,4,5-tris(acetyloxy)-6-({[(2S,3R,4S,5S,6R)-3,4,5-tris(acetyloxy)-6-[(acetyloxy)methyl]oxan-2-yl]oxy}methyl)oxan-2-yl]oxy}ethyl)azanium Iodide

[0605] Step 1:

[0606] Preparation of solution 1: Ac.sub.2O (64.31 g, 629.94 mmol, 59 mL, 21.56 eq) and NaOAc (4.21 g, 51.32 mmol, 1.76 eq) was stirred at reflux under N.sub.2. (3R,4S,5S,6R)-6-[[(2S,3R,4S,5R,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydropyran-2-yl]oxymethyl]tetrahydropyran-2,3,4,5-tetrol (10 g, 29.21 mmol, 1 eq) in toluene (150 mL) was stirred at 100° C. Then the mixture was added to solution 1 and stirred at 100° C. for 1 hour. TLC indicated one new spot formed. The reaction mixture was poured into ice water (400 mL), extracted with ethyl acetate (200 mL*3). The combined organic layers were washed with NaHCO.sub.3 (200 mL*2), dried over Na.sub.2SO.sub.4, filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=20/1 to 0/1). Compound [(2R,3S,4S,5R,6S)-3,4,5-triacetoxy-6-[[(2R,3R,4S,5R,6S)-3,4,5,6-tetraacetoxytetrahydropyran-2-yl]methoxy]tetrahydropyran-2-yl]methyl acetate (5.8 g, 8.55 mmol, 29.26% yield) was obtained as white solid.

[0607] Step 2:

[0608] To a mixture of [(2R,3S,4S,5R,6S)-3,4,5-triacetoxy-6-[[(2R,3R,4S,5R,6S)-3,4,5,6-tetraacetoxytetrahydropyran-2-yl]methoxy]tetrahydropyran-2-yl]methyl acetate (5.8 g, 8.55 mmol, 1 eq) in DCM (50 mL) was added 2-bromoethanol (1.60 g, 12.82 mmol, 910.31 uL, 1.5 eq) and BF.sub.3.Et.sub.2O (6.07 g, 42.74 mmol, 5.27 mL, 5 eq) in one portion at 0° C. under N.sub.2. Then the mixture was heated to 25° C. and stirred for 12 hours. TLC indicated [(2R,3S,4S,5R,6S)-3,4,5-triacetoxy-6-[[(2R,3R,4S,5R,6S)-3,4,5,6-tetraacetoxytetrahydropyran-2-yl]methoxy]tetrahydropyran-2-yl]methyl acetate was consumed and one new spot formed. The reaction mixture was quenched by addition of H.sub.2O (50 mL), diluted with H.sub.2O (50 mL) and extracted with ethyl acetate (50 mL*3). The combined organic layers were dried over Na.sub.2SO.sub.4, filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=20/1 to 0/1). [(2R,3S,4S,5R,6S)-3,4,5-triacetoxy-6-[[(2R,3R,4S,5R)-3,4,5-triacetoxy-6-(2-bromoethoxy)tetrahydropyran-2-yl]methoxy]tetrahydropyran-2-yl]methyl acetate (2 g, crude) was obtained as yellow oil.

[0609] Step 3:

[0610] To a mixture of [(2R,3S,4S,5R,6S)-3,4,5-triacetoxy-6-[[(2R,3R,4S,5R)-3,4,5-triacetoxy-6-(2-bromoethoxy)tetrahydropyran-2-yl]methoxy]tetrahydropyran-2-yl]methyl acetate (2.00 g, 2.69 mmol, 1 eq) in Acetone (20 mL) was added NaI (443.53 mg, 2.96 mmol, 1.1 eq) and N,N-dimethylprop-2-yn-1-amine (1.12 g, 13.45 mmol, 1.43 mL, 5 eq) in one portion at 25° C. under N.sub.2. Then the mixture was heated to 80° C. and stirred for 10 hours. TLC indicated [(2R,3S,4S,5R,6S)-3,4,5-triacetoxy-6-[[(2R,3R,4S,5R)-3,4,5-triacetoxy-6-(2-bromoethoxy)tetrahydropyran-2-yl]methoxy]tetrahydropyran-2-yl]methyl acetate was consumed and one new spot formed. LCMS showed desired m/z was detected. The reaction mixture was filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC [water (0.1% TFA)-ACN]. The title compound (136 mg, 155.67 umol, 5.6% yield, 100% purity, I—) was obtained as white solid. LCMS: (M+): 746.3 .sup.1H NMR (400 MHz, Methanol-d4) δ 5.49-5.43 (m, 1H), 5.37-5.27 (m, 3H), 5.24-5.02 (m, 3H), 4.95 (dd, J=9.7, 8.0 Hz, 1H), 4.79 (d, J=8.0 Hz, 1H), 4.51-4.39 (m, 2H), 4.38-4.26 (m, 2H), 4.24-4.16 (m, 1H), 4.14-4.05 (m, 2H), 3.95-3.87 (m, 1H), 3.84-3.72 (m, 3H), 3.65-3.58 (m, 1H), 3.28 (s, 6H), 2.20-1.95 (m, 21H).

##STR00176##

Compound 82: dimethyl(prop-2-yn-1-yl)(2-{[(2R,3R,4S,5S,6S)-3,4,5-tris(acetyloxy)-6-carboxyoxan-2-yl]oxy}ethyl)azanium trifluoroacetate

[0611] Step 1:

[0612] To a solution of (2S,3S,4S,5R)-3,4,5,6-tetrahydroxytetrahydropyran-2-carboxylic acid (10 g, 51.51 mmol, 1 eq) in DMF (100 mL) was added DBU (8.63 g, 56.66 mmol, 8.54 mL, 1.1 eq), stirred for 15 min at 25° C., and then 3-bromoprop-1-ene (7.48 g, 61.81 mmol, 1.2 eq) was added to the mixture at 0° C. The mixture was stirred for 10 h at 25° C. under N.sub.2. TLC showed the starting reactant was consumed and one new spot formed. The mixture was concentrated. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=30/1 to 0/1). Allyl (2S,3S,4S,5R)-3,4,5,6-tetrahydroxytetrahydropyran-2-carboxylate (10 g, 38.43 mmol, 74.60% yield, 90% purity) was obtained as a white solid.

[0613] Step 2:

[0614] To a solution of allyl (2S,3S,4S,5R)-3,4,5,6-tetrahydroxytetrahydropyran-2-carboxylate (10 g, 42.70 mmol, 1 eq) in Pyridine (100 mL) was added Ac.sub.2O (43.59 g, 426.98 mmol, 39.99 mL, 10 eq). The mixture was stirred at 25° C. for 12 h. TLC showed the starting reactant was consumed completely and one new spot formed. The reaction was clean according to TLC. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=20/1 to 2/1). Allyl (2S,3S,4S,5R)-3,4,5,6-tetraacetoxytetrahydropyran-2-carboxylate (13 g, 29.08 mmol, 68.10% yield, 90% purity) was obtained as a yellow solid.

[0615] Step 3:

[0616] To a mixture of allyl (2S,3S,4S,5R)-3,4,5,6-tetraacetoxytetrahydropyran-2-carboxylate (5 g, 12.43 mmol, 1 eq), 2-bromoethanol (4.66 g, 37.28 mmol, 2.65 mL, 3 eq) in DCM (50 mL) at 0° C. was added BF.sub.3.Et.sub.2O (17.64 g, 124.27 mmol, 15.34 mL, 10 eq), purged with N.sub.2 3 times, and then the mixture was stirred at 25° C. for 12 h under N.sub.2 atmosphere. To the mixture was added H.sub.2O (30 mL), the mixture was extracted with EtOAc (10 mL*3), the organic layer was dried over Na.sub.2SO.sub.4, filtered, and the filtrate was concentrated. The residue was purified by prep-HPLC [water (0.10% TFA)-ACN]. Allyl (2S,3S,4S,5R)-3,4,5-triacetoxy-6-(2-bromoethoxy)tetrahydropyran-2-carboxylate (660 mg, 1.27 mmol, 10.23% yield, 90% purity) was obtained as yellow oil.

[0617] Step 4:

[0618] To a solution of allyl (2S,3S,4S,5R)-3,4,5-triacetoxy-6-(2-bromoethoxy)tetrahydropyran-2-carboxylate (280 mg, 599.24 umol, 1 eq) in ACN (10 mL) was added pyridine (56.00 mg, 787.39 umol, 65.73 uL, 1.31 eq) and Pd(PPh3).sub.4 (84.00 mg, 72.69 umol, 1.21 eq) at 0° C. The mixture was stirred at 0° C. for 1 h. TLC showed the starting reactant was consumed. The reaction mixture was concentrated. The residue was purified by prep-HPLC [water (0.1% TFA)-ACN]. Compound (2S,3S,4S,5R)-3,4,5-triacetoxy-6-(2-bromoethoxy)tetrahydropyran-2-carboxylic acid (156 mg, 346.91 umol, 28.95% yield, 95% purity) was obtained as a yellow solid. LCMS: (M+H+): 427.0.

[0619] Step 5:

[0620] To a solution of (2S,3S,4S,5R)-3,4,5-triacetoxy-6-(2-bromoethoxy)tetrahydropyran-2-carboxylic acid (67 mg, 156.84 umol, 1 eq) in acetone (5 mL) was added N,N-dimethylprop-2-yn-1-amine (65.19 mg, 784.18 umol, 83.15 uL, 5 eq). The mixture was stirred at 80° C. for 12 h. LCMS showed the reactant was consumed completely. The reaction mixture was concentrated. The residue was purified by prep-HPLC [water (0.1% TFA)-ACN]. The title compound (6 mg, 12.84 umol, 8.19% yield, 90% purity) was obtained as a yellow solid. LCMS: (M): 430.2 .sup.1H NMR (400 MHz, Deuterium Oxide) δ 5.35 (t, J=9.6 Hz, 1H), 5.13-4.97 (m, 3H), 4.31-4.15 (m, 3H), 4.12-4.04 (m, 1H), 3.98-3.88 (m, 1H), 3.84-3.74 (m, 1H), 3.58 (dd, J=14.5, 4.2 Hz, 1H), 3.20-3.11 (m, 7H), 1.98-1.87 (m, 9H).

##STR00177##

Compound 83: (2-{[(2R,3R,4R,5S,6R)-4,5-bis(acetyloxy)-6-[(acetyloxy)methyl]-3-acetamidooxan-2-yl]oxy}ethyl)dimethyl(prop-2-yn-1-yl)azanium

[0621] This compound may be synthesized according to the experimental procedure described for Compound 7.

##STR00178##

Compound 85: dimethyl(prop-2-yn-1-yl)(2-{[(2R,3R,4S,5S,6S)-3,4,5-tris(acetyloxy)-6-(methoxycarbonyl)oxan-2-yl]oxy}ethyl)azanium bromide

[0622] Step 1:

[0623] To a solution of methyl (2S,3S,4S,5R,6S)-3,4,5,6-tetraacetoxytetrahydropyran-2-carboxylate (5 g, 13.29 mmol, 1 eq) and 2-bromoethanol (2.49 g, 19.93 mmol, 1.42 mL, 1.5 eq) in DCM (50 mL) was added dropwise BF.sub.3.Et.sub.2O (9.43 g, 66.43 mmol, 8.20 mL, 5 eq) at 0° C., stirred at 0° C. for 1 h, then stirred at 25° C. for 11 h. TLC showed the starting reactant was consumed. The mixture was poured into H.sub.2O (50 mL) at 0° C., and extracted with DCM (20 mL*3). The combined organic layer was dried over Na.sub.2SO.sub.4, filtered, and the filtrate concentrated. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=30/1 to 3/1). Methyl (2S,3S,4S,5R,6R)-3,4,5-triacetoxy-6-(2-bromoethoxy)tetrahydropyran-2-carboxylate (3 g, 5.44 mmol, 40.94% yield, 80% purity) was obtained as colorless oil.

[0624] Step 2:

[0625] To a solution of methyl (2S,3S,4S,5R,6R)-3,4,5-triacetoxy-6-(2-bromoethoxy)tetrahydropyran-2-carboxylate (260 mg, 589.27 umol, 1 eq) in acetone (10 mL) was added N,N-dimethylprop-2-yn-1-amine (244.94 mg, 2.95 mmol, 312.42 uL, 5 eq), stirred at 90° C. for 2 h. LCMS showed the starting reactant was consumed. The mixture was concentrated. The residue was purified by prep-HPLC (column: Phenomenex Luna C18 100*30 mm*5 um; mobile phase: [water (0.1% TFA)-ACN]; B %: 1%-28%, 12 min). The title compound (50 mg, 82.01 umol, 13.92% yield, 86% purity, Br—) was obtained as white solid.

[0626] LCMS: (M+): 444.1 .sup.1H NMR (400 MHz, Deuterium Oxide) δ 5.5 (m, 1H), 5.2-5.1 (m, 4H), 4.5 (m, 1H), 4.4-4.3 (m, 2H), 4.2 (m, 1H), 4.1 (m, 1H), 3.9 (m, 1H), 3.7 (m, 3H), 3.3-3.2 (m, 7H), 2.1-2.0 (m, 9H).

##STR00179##

Compound 86: N-(fluoromethyl)-N-methyl-N-(2-{[(2R,3R,4S,5R,6R)-3,4,5-tris(acetyloxy)-6-[(acetyloxy)methyl]oxan-2-yl]oxy}ethyl)cyclopropanaminium Iodide

[0627] Step 1:

[0628] To a mixture of [(2R,3R,4S,5R,6R)-3,4,5-triacetoxy-6-(2-bromoethoxy)tetrahydropyran-2-yl]methyl acetate (2 g, 4.39 mmol, 1 eq) in THF (20 mL) was added NaI (658.51 mg, 4.39 mmol, 1 eq), Na.sub.2CO.sub.3 (931.26 mg, 8.79 mmol, 2 eq) and N-methylcyclopropanamine (2.60 g, 24.16 mmol, 5.5 eq, HCl) in one portion at 25° C. under N.sub.2. The mixture was heated to 70° C. and stirred for 12 hours. TLC indicated [(2R,3R,4S,5R,6R)-3,4,5-triacetoxy-6-(2-bromoethoxy)tetrahydropyran-2-yl]methyl acetate was consumed and one new spot formed. LCMS showed one main peak with expected mass was detected. The reaction mixture was filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=50/1 to 0/1). 1.2 g crude product was obtained. 200 mg of the crude product was further purified by prep-HPLC [water (0.10% TFA)-ACN] to get 82 mg of product (99% purity, TFA) as white solid. The crude compound [(2R,3R,4S,5R,6R)-3,4,5-triacetoxy-6-[2-[cyclopropyl(methyl)amino]ethoxy]tetrahydropyran-2-yl]methyl acetate (1 g, 40.28% yield, crude) as white solid was used in the next step directly. LCMS: (M+H)+: 446.2

[0629] Step 2:

[0630] To a mixture of [(2R,3R,4S,5R,6R)-3,4,5-triacetoxy-6-[2-[cyclopropyl(methyl)amino]ethoxy]tetrahydropyran-2-yl]methyl acetate (1.78 g, 2.20 mmol, 1 eq) in acetone (20 mL) was added fluoro(iodo)methane (1.76 g, 11.00 mmol, 5 eq) in one portion at 25° C. under N.sub.2. The mixture was heated to 90° C. and stirred for 2 hours. TLC indicated [(2R,3R,4S,5R,6R)-3,4,5-triacetoxy-6-[2-[cyclopropyl(methyl)amino]ethoxy]tetrahydropyran-2-yl]methyl acetate was consumed and one new spot formed. LCMS showed one main peak with expected mass was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Ethyl acetate:Methanol=1/0 to 3/1). The title compound (1.11 g, 58% yield 99% purity, I—) was obtained as yellow solid. LCMS: (M+): 478.2 .sup.1H NMR (400 MHz, Methanol-d4) δ 5.58 (s, 1H), 5.46 (s, 1H), 5.32 (t, J=9.5 Hz, 1H), 5.08 (t, J=9.8 Hz, 1H), 4.98 (dd, J=9.7, 8.0 Hz, 1H), 4.86-4.81 (m, 1H), 4.50-4.35 (m, 1H), 4.30 (d, J=3.5 Hz, 2H), 4.27-4.10 (m, 1H), 4.02-3.93 (m, 1H), 3.92-3.82 (m, 2H), 2.97 (dd, J=6.0, 2.2 Hz, 3H), 2.14-2.02 (m, 9H), 2.00 (s, 3H), 1.47-1.30 (m, 2H), 1.12-0.95 (m, 2H).

##STR00180##

Compound 88: (chloromethyl)dimethyl(2-{[(2R,3R,4S,5R)-3,4,5-tris(acetyloxy)oxan-2-yl]oxy}ethyl)azanium Iodide

[0631] Step 1:

[0632] To a mixture of (2R,3R,4S,5R)-tetrahydropyran-2,3,4,5-tetrol (50 g, 333.04 mmol, 1 eq) in pyridine (300 mL) was added acetyl acetate (204.00 g, 2.00 mol, 187.16 mL, 6 eq) in one portion at 0° C. under N.sub.2. The mixture was heated to 25° C. and stirred for 12 hours. TLC indicated one new spot formed. The reaction was clean according to TLC. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=50/1 to 0/1). Compound [(3R,4S,5R)-4,5,6-triacetoxytetrahydropyran-3-yl]acetate (80 g, 251.35 mmol, 75.47% yield) was obtained as yellow oil.

[0633] Step 2:

[0634] To a mixture of [(3R,4S,5R)-4,5,6-triacetoxytetrahydropyran-3-yl] acetate (20 g, 62.84 mmol, 1 eq) in DCM (200 mL) was added 2-bromoethanol (11.78 g, 94.26 mmol, 6.69 mL, 1.5 eq) and BF.sub.3.Et.sub.2O (9.10 g, 62.84 mmol, 7.91 mL, 98% purity, 1 eq) in one portion at 0° C. under N.sub.2. The mixture was heated to 25° C. and stirred for 12 hours. TLC indicated [(3R,4S,5R)-4,5,6-triacetoxytetrahydropyran-3-yl] acetate was consumed and one new spot formed. The reaction mixture was quenched by addition of H.sub.2O (100 mL), and then diluted with H.sub.2O (200 mL) and extracted with ethyl acetate (200 mL*3). The combined organic layers were dried over Na.sub.2SO.sub.4, filtered and the filtrate was concentrated under reduced pressure to give a residue. The crude product was purified by re-crystallization from ethyl acetate:Petroleum ether (40 ml:30 mL) at 0° C. for 12h. The mixture was filtered and the filter cake was concentrated under reduced pressure to give a residue. The filtrate was purified by prep-HPLC [water (0.1% TFA)-ACN].[(3R,4S,5R,6S)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate (3.4 g, 8.87 mmol, 14.12% yield) was obtained as a white solid. Compound [(3R,4S,5R,6R)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate (3.5 g, 9.13 mmol, 14.54% yield) was obtained as yellow oil.

[0635] Step 3:

[0636] To a mixture of [(3R,4S,5R,6S)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate (3.4 g, 8.87 mmol, 1 eq) in THF (30 mL) was added NaI (1.33 g, 8.87 mmol, 1 eq) and Na.sub.2CO.sub.3 (1.88 g, 17.75 mmol, 2 eq) in one portion at 25° C. under N.sub.2. N-methylmethanamine (2 M, 24.40 mL, 5.5 eq) was added. The mixture was heated to 70° C. and stirred for 12 hours. TLC indicated [(3R,4S,5R,6S)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate was consumed incompletely and one new spot formed. LCMS showed [(3R,4S,5R,6S)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate was consumed and one main peak with expected mass was detected. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Ethyl acetate:Methanol=1/0 to 1/1). The 200 mg crude product was purified by prep-HPLC [water (0.1% TFA)-ACN]. Compound [(3R,4S,5R,6R)-4,5-diacetoxy-6-[2-(dimethylamino)ethoxy]tetrahydropyran-3-yl] acetate (3.1 g, crude) was obtained as yellow oil. Compound [(3R,4S,5R,6R)-4,5-diacetoxy-6-[2-(dimethylamino)ethoxy]tetrahydropyran-3-yl] acetate (14.7 mg, 98% purity) was obtained as white solid. LCMS: (M+H).sup.+: 348.1

[0637] Step 4:

[0638] To a mixture of [(3R,4S,5R,6R)-4,5-diacetoxy-6-[2-(dimethylamino)ethoxy]tetrahydropyran-3-yl] acetate (3 g, 8.64 mmol, 1 eq) in acetone (30 mL) was added chloro(iodo)methane (7.62 g, 43.18 mmol, 3.13 mL, 5 eq) in one portion at 25° C. under N.sub.2. The mixture was heated to 90° C. and stirred for 2 hours. LCMS showed [(3R,4S,5R,6R)-4,5-diacetoxy-6-[2-(dimethylamino)ethoxy]tetrahydropyran-3-yl]acetate was consumed and one main peak with expected mass was detected. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC [water (0.1% TFA)-ACN]. Compound chloromethyl-dimethyl-[2-[(2R,3R,4S,5R)-3,4,5-triacetoxytetrahydropyran-2-yl]oxyethyl]ammonium (315 mg, 589.41 umol, 6.82% yield, 98% purity, I—) was obtained as yellow oil. LCMS: (M+).sup.+: 396.1

[0639] Step 5:

[0640] To a mixture of [(3R,4S,5R,6R)-4,5-diacetoxy-6-[2-(dimethylamino)ethoxy]tetrahydropyran-3-yl] acetate (2.8 g, 8.06 mmol, 1 eq) in DCM (20 mL) was added chloro(iodo)methane (7.11 g, 40.30 mmol, 2.93 mL, 5 eq) and DIPEA (1.06 g, 8.06 mmol, 1.43 mL, 98% purity, 1 eq) in one portion at 25° C. under N.sub.2. The mixture was stirred at 40° C. for 12 hours in a 50 mL sealed tube. TLC indicated [(3R,4S,5R,6R)-4,5-diacetoxy-6-[2-(dimethylamino)ethoxy]tetrahydropyran-3-yl] acetate was consumed incompletely and one new spot formed. LCMS showed one main peak with expected mass was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Ethyl acetate:Methanol=1/0 to 2/1). The residue was then purified by prep-HPLC [water (0.1% TFA)-ACN]. Compound chloromethyl-dimethyl-[2-[(2R,3R,4S,5R)-3,4,5-triacetoxytetrahydropyran-2-yl]oxyethyl]ammonium (617 mg, 1.18 mmol, 14.6% yield, 100% purity, I—) was obtained as yellow solid. LCMS: (M+).sup.+: 396.1 .sup.1H NMR (400 MHz, Methanol-d4) δ 5.31 (s, 2H), 5.32-5.22 (m, 1H), 5.06-4.91 (m, 2H), 4.73 (d, J=7.4 Hz, 1H), 4.35-4.26 (m, 1H), 4.19-4.02 (m, 2H), 3.87-3.73 (m, 2H), 3.52 (dd, J=11.7, 9.6 Hz, 1H), 3.29 (s, 6H), 2.15-1.99 (m, 9H).

##STR00181##

Compound 89: (chloromethyl)dimethyl(2-{[(3R,4S,5S)-3,4,5-tris(acetyloxy)oxan-2-yl]oxy}ethyl)azanium

[0641] This compound may be synthesized according to the experimental procedure described for Compound 325.

##STR00182##

Compound 90: (fluoromethyl)dimethyl(2-{[(2R,3R,4S,5R)-3,4,5-tris(acetyloxy)oxan-2-yl]oxy}ethyl)azanium Iodide

[0642] Step 1:

[0643] To a solution of (2R,3R,4S,5R)-tetrahydropyran-2,3,4,5-tetrol (50 g, 333.04 mmol, 1 eq) in pyridine (500 mL) was added Ac.sub.2O (204.00 g, 2.00 mol, 187.16 mL, 6 eq). The mixture was stirred at 25° C. for 12 hr. TLC indicated (2R,3R,4S,5R)-tetrahydropyran-2,3,4,5-tetrol was consumed completely and one new spot formed. The reaction mixture was concentrated under reduced pressure. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=80/1 to 0/1). Compound [(3R,4S,5R)-4,5,6-triacetoxytetrahydropyran-3-yl] acetate (100 g, 314.19 mmol, 94.34% yield) was obtained as a yellow oil.

[0644] Step 2:

[0645] To a solution of [(3R,4S,5R)-4,5,6-triacetoxytetrahydropyran-3-yl] acetate (20 g, 62.84 mmol, 1 eq) in DCM (150 mL) was added dropwise 2-bromoethanol (11.78 g, 94.26 mmol, 6.69 mL, 1.5 eq) and BF.sub.3.Et.sub.2O (45.50 g, 314.19 mmol, 39.57 mL, 98% purity, 5 eq) at 0° C. The mixture was warmed to 25° C. and was stirred at 25° C. for 12 hr. TLC indicated [(3R,4S,5R)-4,5,6-triacetoxytetrahydropyran-3-yl] acetate was consumed completely and new spots formed. 5 batches were combined for work-up. The reaction mixture was quenched by addition of H.sub.2O (500 mL) at 25° C. and extracted with EtOAc (900 mL, 300 mL*3). The combined organic layers were concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=20/1 to 0/1). The crude product was purified by re-crystallization from ethyl acetate:Petroleum ether (40 ml:30 mL) at 0° C. for 12h. Compound [(3R,4S,5R,6S)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate (3 g, 7.83 mmol, 2.49% yield) was obtained as a white solid. The filtrate was purified by prep-HPLC [water (0.1% TFA)-ACN]. Compound [(3R,4S,5R,6R)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate (7 g, 18.27 mmol, 5.81% yield) was obtained as a colorless oil.

[0646] Step 3:

[0647] To a solution of [(3R,4S,5R,6S)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate (1 g, 2.61 mmol, 1 eq) in THF (10 mL) was added Me.sub.2NH.THF (2 M, 7.18 mL, 5.50 eq) and DIEA (674.57 mg, 5.22 mmol, 909.12 uL, 2 eq). The mixture was stirred at 70° C. for 12 hr. LCMS showed [(3R,4S,5R,6S)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate was consumed completely and the expected mass was detected. The reaction mixture was concentrated under reduced pressure. The residue was purified by column chromatography (SiO.sub.2, Ethyl acetate/Methanol=100/1 to 0/1). Compound [(3R,4S,5R,6R)-4,5-diacetoxy-6-[2-(dimethylamino)ethoxy]tetrahydropyran-3-yl] acetate (500 mg, 1.44 mmol, 55.16% yield) was obtained as a yellow solid.

[0648] Step 4:

[0649] To a solution of [(3R,4S,5R,6R)-4,5-diacetoxy-6-[2-(dimethylamino)ethoxy]tetrahydropyran-3-yl] acetate (500 mg, 1.44 mmol, 1 eq) in DCM (5 mL) was added fluoro(iodo)methane (1.15 g, 7.20 mmol, 5 eq) and DIEA (186.04 mg, 1.44 mmol, 250.72 uL, 1 eq). The mixture was stirred at 40° C. for 3 hr. LCMS showed the reaction was completed and one main peak with expected mass was detected. The mixture was filtered and the filter cake was concentrated under reduced pressure to give a residue. The crude product was washed with ethyl acetate (10 mL). The mixture was filtered and the filter cake was concentrated under reduced pressure to give a residue. The title compound (397 mg, 782.59 umol, 54.37% yield, 100% purity, I—) was obtained as a white solid. LCMS: (M+): 380.1 .sup.1H NMR (400 MHz, Deuterium Oxide) δ 5.35 (s, 1H), 5.24 (s, 1H), 5.14 (t, J=7.6 Hz, 1H), 4.96-4.83 (m, 2H), 4.75 (d, J=6.1 Hz, 1H), 4.21 (d, J=13.8 Hz, 1H), 4.10 (dd, J=12.3, 4.7 Hz, 1H), 3.99 (d, J=13.6 Hz, 1H), 3.70-3.63 (m, 2H), 3.54 (dd, J=12.3, 7.9 Hz, 1H), 3.13 (d, J=2.0 Hz, 6H), 2.06-1.97 (m, 9H).

##STR00183##

Compound 91 and Compound 97: (fluoromethyl)dimethyl(2-{[(3R,4S,5S)-3,4,5-tris(acetyloxy)oxan-2-yl]oxy}ethyl)azanium

[0650] This compound may be synthesized according to the experimental procedure described for Compound 311.

##STR00184##

Compound 92: (fluoromethyl)(methyl)(prop-2-yn-1-yl)(2-{[(2R,3R,4S,5R)-3,4,5-tris(acetyloxy)oxan-2-yl]oxy}ethyl)azanium Iodide

[0651] Step 1:

[0652] To a mixture of (2R,3R,4S,5R)-tetrahydropyran-2,3,4,5-tetrol (50 g, 333.05 mmol, 1 eq) in pyridine (300 mL) was added acetyl acetate (204.00 g, 2.00 mol, 187.16 mL, 6 eq) in one portion at 0° C. under N.sub.2. The mixture was heated to 25° C. and stirred for 12 hours. TLC indicated one new spot formed. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=50/1 to 0/1). Compound [(3R,4S,5R)-4,5,6-triacetoxytetrahydropyran-3-yl] acetate (100 g, 314.19 mmol, 94.34% yield) was obtained as yellow oil.

[0653] Step 2:

[0654] To a solution of [of [(3R,4S,5R)-4,5,6-triacetoxytetrahydropyran-3-yl] acetate (20 g, 62.84 mmol, 1 eq) in DCM (150 mL) was added dropwise 2-bromoethanol (11.78 g, 94.26 mmol, 6.69 mL, 1.5 eq) and BF.sub.3.Et.sub.2O (45.50 g, 314.19 mmol, 39.57 mL, 98% purity, 5 eq) at 0° C. The mixture was warmed to 25° C. and stirred at 25° C. for 12 hr. TLC indicated [(3R,4S,5R)-4,5,6-triacetoxytetrahydropyran-3-yl] acetate was consumed completely and two new spots formed. The reaction mixture was quenched by addition of H.sub.2O (500 mL) and extracted with EtOAc (900 mL, 300 mL*3). The combined organic layers were concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=20/1 to 0/1). Two batches were obtained successively. The batch 1 was further purified by recrystallization from ethyl acetate:Petroleum etherat 0° C. for 12h. Compound [(3R,4S,5R,6S)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate (3 g, 2.5% yield) was obtained as a white solid. The batch 2 was further purified by prep-HPLC [water (0.1% TFA)-ACN]. Compound [(3R,4S,5R,6R)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate (7 g, 5.8% yield) was obtained as a colorless oil.

[0655] Step 3:

[0656] To a solution of [(3R,4S,5R,6S)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate (1.8 g, 4.70 mmol, 1 eq) in THF (20 mL) was added N-methylprop-2-yn-1-amine (1.79 g, 25.84 mmol, 2.15 mL, 5.5 eq) and DIPEA (1.21 g, 9.39 mmol, 1.64 mL, 2 eq). The mixture was stirred at 70° C. for 12 hr. LCMS showed [(3R,4S,5R,6S)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate was consumed completely and one main peak with expected mass was detected. The reaction mixture was concentrated under reduced pressure. The crude product (300 mg) was purified by prep-HPLC [water (0.1% TFA)-ACN]. Compound [(3R,4S,5R,6R)-4,5-diacetoxy-6-[2-[methyl(prop-2-ynyl)amino]ethoxy]tetrahydropyran-3-yl] acetate (97 mg, 100% purity, TFA) was obtained as white solid. The other crude product[(3R,4S,5R,6R)-4,5-diacetoxy-6-[2-[methyl(prop-2-ynyl)amino]ethoxy]tetrahydropyran-3-yl] acetate (1.4 g) was used into the next step without further purification. LCMS: (M+H).sup.+: 372.1

[0657] Step 4:

[0658] To a solution of [(3R,4S,5R,6R)-4,5-diacetoxy-6-[2-[methyl(prop-2-ynyl)amino]ethoxy]tetrahydropyran-3-yl] acetate (800 mg, 2.15 mmol, 1 eq) in DCM (8 mL) was added fluoro(iodo)methane (1.72 g, 10.77 mmol, 5 eq) and DIPEA (278.40 mg, 2.15 mmol, 375.21 uL, 1 eq). The mixture was stirred at 40° C. for 3 hr. LCMS showed [(3R,4S,5R,6R)-4,5-diacetoxy-6-[2-[methyl(prop-2-ynyl)amino]ethoxy]tetrahydropyran-3-yl] acetate was consumed completely and expected mass was detected. The reaction mixture was concentrated under reduced pressure. The residue was purified by prep-HPLC [water (0.10% TFA)-ACN]. The title compound (234 mg, 20.5% yield, 100% purity, I—) was obtained as yellow oil. LCMS:(M+): 404.1 δ 1H NMR (400 MHz, Methanol-d4) δ 5.65 (s, 1H), 5.54 (s, 1H), 5.26 (t, J=9.1 Hz, 1H), 5.03-4.91 (m, 2H), 4.73 (d, J=7.3, 0.7 Hz, 1H), 4.57-4.51 (m, 2H), 4.38-4.27 (m, 1H), 4.19-4.05 (m, 2H), 3.93-3.79 (m, 2H), 3.66 (t, J=2.6 Hz, 1H), 3.52 (dd, J=11.8, 9.6 Hz, 1H), 3.30 (d, J=2.1 Hz, 3H), 2.12-2.01 (m, 9H).

##STR00185##

Compound 93: (fluoromethyl)(methyl)(prop-2-yn-1-yl)(2-{[(3R,4S,5S)-3,4,5-tris(acetyloxy)oxan-2-yl]oxy}ethyl)azanium

[0659] This compound may be synthesized according to the experimental procedure described for Compound 309.

##STR00186##

##STR00187##

Compound 94: (chloromethyl)dimethyl(2-{[(2S,3R,4S,5R)-3,4,5-tris(acetyloxy)oxan-2-yl]oxy}ethyl)azanium Iodide

[0660] Step 1:

[0661] To a mixture of (2R,3R,4S,5R)-tetrahydropyran-2,3,4,5-tetrol (50 g, 333.04 mmol, 1 eq) in pyridine (300 mL) was added acetyl acetate (204.00 g, 2.00 mol, 187.16 mL, 6 eq) in one portion at 0° C. under N.sub.2. The mixture was heated to 25° C. and stirred for 12 hours. TLC indicated one new spot. The reaction was clean according to TLC. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=50/1 to 0/1). Compound [(3R,4S,5R)-4,5,6-triacetoxytetrahydropyran-3-yl] acetate (80 g, 251.35 mmol, 75.47% yield) was obtained as yellow oil.

[0662] Step 2:

[0663] To a mixture of [(3R,4S,5R)-4,5,6-triacetoxytetrahydropyran-3-yl] acetate (20 g, 62.84 mmol, 1 eq) in DCM (200 mL) was added 2-bromoethanol (11.78 g, 94.26 mmol, 6.69 mL, 1.5 eq) and BF.sub.3.Et.sub.2O (9.10 g, 62.84 mmol, 7.91 mL, 98% purity, 1 eq) in one portion at 0° C. under N.sub.2. The mixture was heated to 25° C. and stirred for 12 hours. TLC indicated [(3R,4S,5R)-4,5,6-triacetoxytetrahydropyran-3-yl] acetate was consumed and one new spot formed. The reaction mixture was quenched by addition of H.sub.2O (100 mL), and then diluted with H.sub.2O (200 mL) and extracted with ethyl acetate (200 mL*3). The combined organic layers were dried over Na.sub.2SO.sub.4, filtered, and the filtrate was concentrated under reduced pressure to give a residue. The crude product was purified by re-crystallization from ethyl acetate: Petroleum ether (40 ml:30 mL) at 0° C. for 12h. The mixture was filtered and the filter cake was concentrated under reduced pressure to give a residue. The filtrate was purified by prep-HPLC [water (0.1% TFA)-ACN]. [(3R,4S,5R,6S)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate (3.4 g, 8.87 mmol, 14.12% yield) was obtained as a white solid. Compound [(3R,4S,5R,6R)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate (3.5 g, 9.13 mmol, 14.54% yield) was obtained as yellow oil.

[0664] Step 3:

[0665] To a mixture of [(3R,4S,5R,6S)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate (3.4 g, 8.87 mmol, 1 eq) in THF (30 mL) was added NaI (1.33 g, 8.87 mmol, 1 eq) and Na.sub.2CO.sub.3 (1.88 g, 17.75 mmol, 2 eq) in one portion at 25° C. under N.sub.2. Then N-methylmethanamine (2 M, 24.40 mL, 5.5 eq) was added. The mixture was heated to 70° C. and stirred for 12 hours. TLC indicated [(3R,4S,5R,6S)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate was consumed incompletely and one new spot formed. LCMS showed [(3R,4S,5R,6S)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate was consumed and one main peak with desired m/z was detected. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Ethyl acetate:Methanol=1/0 to 1/1). The 200 mg crude product was purified by prep-HPLC [water (0.1% TFA)-ACN]. Compound [(3R,4S,5R,6R)-4,5-diacetoxy-6-[2-(dimethylamino)ethoxy]tetrahydropyran-3-yl] acetate (3.1 g, crude) was obtained as yellow oil. Compound [(3R,4S,5R,6R)-4,5-diacetoxy-6-[2-(dimethylamino)ethoxy]tetrahydropyran-3-yl] acetate (14.7 mg, 98% purity) was obtained as white solid. LCMS: (M+H).sup.+: 348.1

[0666] Step 4:

[0667] To a mixture of [(3R,4S,5R,6R)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate (3.5 g, 9.13 mmol, 1 eq) in THF (30 mL) was added NaI (1.37 g, 9.13 mmol, 1 eq), Na.sub.2CO.sub.3 (1.94 g, 18.27 mmol, 2 eq) and N-methylmethanamine (2 M, 25.12 mL, 5.5 eq) in one portion at 25° C. under N.sub.2. The mixture was heated to 70° C. and stirred for 12 hours. LCMS showed [(3R,4S,5R,6R)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate was consumed and one main peak with desired m/z was detected. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The 500 mg crude product was purified by prep-HPLC [water (0.10% TFA)-ACN]. Compound [(3R,4S,5R,6S)-4,5-diacetoxy-6-[2-(dimethylamino)ethoxy]tetrahydropyran-3-yl] acetate (34 mg, 73.69 umol, 5.12% yield, 100% purity, TFA) was obtained as colorless oil. Compound [(3R,4S,5R,6S)-4,5-diacetoxy-6-[2-(dimethylamino)ethoxy]tetrahydropyran-3-yl] acetate (4.2 g, crude) was obtained as yellow oil and the crude product was used into the next step without further purification. LCMS: (M+H).sup.+: 348.1

[0668] Step 5:

[0669] To a mixture of [(3R,4S,5R,6S)-4,5-diacetoxy-6-[2-(dimethylamino)ethoxy]tetrahydropyran-3-yl] acetate (4.2 g, 12.09 mmol, 1 eq) in acetone (40 mL) was added chloro(iodo)methane (10.66 g, 60.46 mmol, 4.39 mL, 5 eq) in one portion at 25° C. under N.sub.2. The mixture was heated to 90° C. and stirred for 2 hours. TLC indicated [(3R,4S,5R,6S)-4,5-diacetoxy-6-[2-(dimethylamino)ethoxy]tetrahydropyran-3-yl] acetate was consumed and one new spot formed. LCMS showed one main peak with desired m/z. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Ethyl acetate: Methanol=1/0 to 2/1). Then the residue was purified by prep-HPLC [water (0.1% TFA)-ACN]. The title compound (26 mg, 49.64 umol, 9.85% yield, I—) was obtained as yellow oil. LCMS: (M.sup.+).sup.+: 396.0 .sup.1H NMR (400 MHz, Methanol-d4) δ 5.45-5.25 (m, 4H), 5.18 (s, 1H), 4.42-4.31 (m, 2H), 4.30-4.15 (m, 2H), 4.07-3.97 (m, 1H), 3.82 (t, J=4.7 Hz, 2H), 3.32 (s, 6H), 2.16-2.04 (m, 9H).

##STR00188##

Compound 95: (fluoromethyl)dimethyl(2-{[(3R,4S,5S,6R)-3,4,5-tris(acetyloxy)-6-methyloxan-2-yl]oxy}ethyl)azanium

[0670] This compound may be synthesized according to the experimental procedure described for Compound 311.

##STR00189##

Compound 96: (fluoromethyl)(methyl)(prop-2-yn-1-yl)(2-{[(2S,3R,4S,5R)-3,4,5-tris(acetyloxy)oxan-2-yl]oxy}ethyl)azanium Iodide

[0671] Step 1:

[0672] To a mixture of (2R,3R,4S,5R)-tetrahydropyran-2,3,4,5-tetrol (50 g, 333.05 mmol, 1 eq) in pyridine (300 mL) was added acetyl acetate (204.00 g, 2.00 mol, 187.16 mL, 6 eq) in one portion at 0° C. under N.sub.2. The mixture was heated to 25° C. and stirred for 12 hours. TLC indicated one new spot formed. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=50/1 to 0/1). Compound [(3R,4S,5R)-4,5,6-triacetoxytetrahydropyran-3-yl] acetate (100 g, 314.19 mmol, 94.34% yield) was obtained as yellow oil.

[0673] Step 2:

[0674] To a solution of [of [(3R,4S,5R)-4,5,6-triacetoxytetrahydropyran-3-yl] acetate (20 g, 62.84 mmol, 1 eq) in DCM (150 mL) was added dropwise 2-bromoethanol (11.78 g, 94.26 mmol, 6.69 mL, 1.5 eq) and BF.sub.3.Et.sub.2O (45.50 g, 314.19 mmol, 39.57 mL, 98% purity, 5 eq) at 0° C. The mixture was warmed to 25° C. and stirred at 25° C. for 12 hr. TLC indicated [(3R,4S,5R)-4,5,6-triacetoxytetrahydropyran-3-yl] acetate was consumed completely and two new spots formed. The reaction mixture was quenched by addition of H.sub.2O (500 mL) and extracted with EtOAc (900 mL, 300 mL*3). The combined organic layers were concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=20/1 to 0/1). Two batches were obtained successively. The batch 1 was further purified by re-crystallization from ethyl acetate:Petroleum ether at 0° C. for 12h. Compound [(3R,4S,5R,6S)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate (3 g, 2.5% yield) was obtained as a white solid. The batch 2 was further purified by prep-HPLC [water (0.10% TFA)-ACN]. The title compound (7 g, 5.8% yield) was obtained as colorless oil.

[0675] Step 3:

[0676] To a solution of [(3R,4S,5R,6R)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate (1.50 g, 3.91 mmol, 1 eq) in THF (15 mL) was added N-methylprop-2-yn-1-amine (1.49 g, 21.53 mmol, 1.79 mL, 5.5 eq) and DIPEA (1.01 g, 7.83 mmol, 1.36 mL, 2 eq). The mixture was stirred at 70° C. for 12 hr. LCMS showed [(3R,4S,5R,6R)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate was consumed completely and one main peak with expected mass was detected. The reaction mixture was concentrated under reduced pressure. The crude product (300 mg) was purified by prep-HPLC [water (0.1% TFA)-ACN]. Compound[(3R,4S,5R,6S)-4,5-diacetoxy-6-[2-[methyl(prop-2-ynyl)amino]ethoxy]tetrahydropyran-3-yl] acetate (110 mg, 100% purity, TFA) was obtained as yellow oil. The other crude product [(3R,4S,5R,6S)-4,5-diacetoxy-6-[2-[methyl(prop-2-ynyl)amino]ethoxy]tetrahydropyran-3-yl] acetate (600 mg, crude) was used in the next step without further purification. LCMS: (M+H).sup.+: 372.1

[0677] Step 4:

[0678] To a solution of [(3R,4S,5R,6S)-4,5-diacetoxy-6-[2-[methyl(prop-2-ynyl)amino]ethoxy]tetrahydropyran-3-yl] acetate (300 mg, 807.80 umol, 1 eq) in DCM (3 mL) was added fluoro(iodo)methane (645.95 mg, 4.04 mmol, 5 eq) and DIPEA (104.40 mg, 807.80 umol, 140.70 uL, 1 eq). The mixture was stirred at 40° C. for 3 hr. LCMS showed [(3R,4S,5R,6S)-4,5-diacetoxy-6-[2-[methyl(prop-2-ynyl)amino]ethoxy]tetrahydropyran-3-yl] acetate was consumed completely and the expected mass was detected. The reaction mixture was concentrated under reduced pressure. The residue was purified by prep-HPLC [water (0.10% TFA)-ACN]. Compound fluoromethyl-methyl-prop-2-ynyl-[2-[(2R,3R,4S,5R)-3,4,5-triacetoxytetrahydropyran-2-yl]oxyethyl]ammonium (95 mg, 29.1% yield, 100% purity, I—) was obtained as yellow oil. LCMS: (M+): 404.1 .sup.1H NMR (400 MHz, Methanol-d4) δ 5.72 (d, J=2.1 Hz, 1H), 5.61 (d, J=2.1 Hz, 1H), 5.41 (td, J=9.8, 2.5 Hz, 1H), 5.11-5.05 (m, 1H), 5.05-4.94 (m, 2H), 4.61 (dd, J=2.8, 1.4 Hz, 2H), 4.19 (d, J=12.4 Hz, 1H), 4.07-3.91 (m, 2H), 3.91-3.83 (m, 2H), 3.77-3.61 (m, 2H), 3.34 (d, J=2.1 Hz, 3H), 2.14-1.97 (m, 9H).

Compound 97: (fluoromethyl)dimethyl(2-{[(2S,3R,4S,5R)-3,4,5-tris(acetyloxy)oxan-2-yl]oxy}ethyl)azanium Iodide

[0679] Step 1:

[0680] To a solution of (2R,3R,4S,5R)-tetrahydropyran-2,3,4,5-tetrol (50 g, 333.04 mmol, 1 eq) in pyridine (500 mL) was added Ac.sub.2O (204.00 g, 2.00 mol, 187.16 mL, 6 eq). The mixture was stirred at 25° C. for 12 hr. TLC indicated (2R,3R,4S,5R)-tetrahydropyran-2,3,4,5-tetrol was consumed completely and one new spot formed. The reaction mixture was concentrated under reduced pressure. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=80/1 to 0/1). Compound [(3R,4S,5R)-4,5,6-triacetoxytetrahydropyran-3-yl] acetate (100 g, 314.19 mmol, 94.34% yield) was obtained as a yellow oil.

[0681] Step 2:

[0682] To a solution of [(3R,4S,5R)-4,5,6-triacetoxytetrahydropyran-3-yl] acetate (20 g, 62.84 mmol, 1 eq) in DCM (150 mL) was added dropwise 2-bromoethanol (11.78 g, 94.26 mmol, 6.69 mL, 1.5 eq) and BF.sub.3.Et.sub.2O (45.50 g, 314.19 mmol, 39.57 mL, 98% purity, 5 eq) at 0° C. The mixture was warmed to 25° C. and was stirred at 25° C. for 12 hr. TLC indicated [(3R,4S,5R)-4,5,6-triacetoxytetrahydropyran-3-yl] acetate was consumed completely and new spots formed. 5 batches were combined for work-up. The reaction mixture was quenched by addition of H.sub.2O (500 mL) at 25° C. and extracted with EtOAc (900 mL, 300 mL*3). The combined organic layers were concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=20/1 to 0/1). The crude product was purified by re-crystallization from ethyl acetate:Petroleum ether (40 ml:30 mL) at 0° C. for 12h. Compound [(3R,4S,5R,6S)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate (3 g, 7.83 mmol, 2.49% yield) was obtained as a white solid. The filtrate was purified by prep-HPLC [water (0.1% TFA)-ACN]. Compound [(3R,4S,5R,6R)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate (7 g, 18.27 mmol, 5.81% yield) was obtained as a colorless oil.

[0683] Step 3:

[0684] To a mixture of [(3R,4S,5R,6R)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate (0.5 g, 1.30 mmol, 1 eq) in THF (5 mL) was added MeNH.sub.2 (810.49 mg, 5.22 mmol, 20% purity, 4 eq) and NaI (195.59 mg, 1.30 mmol, 1 eq) in one portion at 25° C. under N.sub.2. The mixture was stirred at 25° C. for 12 hours. LCMS showed the expected mass was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Ethyl acetate/MeOH=0/1 to 10/1). The crude product [(3R,4S,5R,6S)-4,5-diacetoxy-6-[2-(dimethylamino)ethoxy]tetrahydropyran-3-yl] acetate (0.38 g, 1.09 mmol, 83.84% yield) was obtained as yellow oil, and used into the next step without further purification.

[0685] Step 4:

[0686] To a mixture of [(3R,4S,5R,6S)-4,5-diacetoxy-6-[2-(dimethylamino)ethoxy]tetrahydropyran-3-yl] acetate (370 mg, 1.07 mmol, 1 eq) in DCM (5 mL) was added fluoro(iodo)methane (851.76 mg, 5.33 mmol, 5 eq). The mixture was stirred at 40° C. for 3 hours. LCMS showed desired mass was detected. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC [water (0.10% TFA)-ACN]. The title compound (14 mg, 36.07 umol, 3.39% yield, 98% purity) was obtained as colorless oil. (M+): 380.1 .sup.1H NMR (400 MHz, Methanol-d4) δ 5.58 (s, 1H), 5.47 (s, 1H), 5.42 (t, J=9.7 Hz, 1H), 5.11 (d, J=3.7 Hz, 1H), 5.07-4.96 (m, 2H), 4.20 (d, J=13.2 Hz, 1H), 4.02 (t, J=11.0 Hz, 1H), 3.94-3.82 (m, 2H), 3.82-3.74 (m, 1H), 3.68 (t, J=10.8 Hz, 1H), 3.31 (d, J=2.1 Hz, 6H), 3.04-2.94 (m, 1H), 2.09-2.01 (m, 9H).

##STR00190##

Compound 98: (fluoromethyl)dimethyl(2-{[(2R,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)oxan-2-yl]oxy}ethyl)azanium Chloride

[0687] N-(fluoromethyl)-N,N-dimethyl-2-(((3R,4S,5R,6R)-3,4,5-triacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)ethan-1-aminium iodide (1.00 g, 1 Eq, 1.73 mmol) is dissolved in methanol (3 mL), followed by addition of 0.5M NaOMe in methanol (9.32 mg, 0.35 mL, 0.1 Eq, 173 μmol). The reaction was stirred at room temperature for a few hours, then concentrated by rotary evaporation. The sample was redissolved in MeOH, then purified by column chromatography (C18AQ flash column, elutes at 100% Water with 0.1% TFA). The pure fractions were lyophilized and collected. The product was redissolved in water and stirred with Amberlyst IRA-400 Cl-resin (300 mg resin/100 mg of TFA salt), then lyophilized to yield the title compound (0.400 g, 72.5%)). LCMS M+: 284.2. .sup.1H NMR (400 MHz, Deuterium Oxide) δ 5.39 (s, 1H), 5.28 (s, 1H), 4.41 (dd, J=8.0, 0.9 Hz, 1H), 4.35-4.25 (m, 1H), 4.09-3.99 (m, 1H), 3.87-3.79 (m, 1H), 3.74-3.59 (m, 3H), 3.45-3.33 (m, 2H), 3.33-3.27 (m, 1H), 3.31-3.19 (m, 1H), 3.16 (d, J=2.1 Hz, 6H).

##STR00191##

Compound 99: (chloromethyl)dimethyl(2-{[(2R,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)oxan-2-yl]oxy}ethyl)azanium Chloride

[0688] N,N-dimethyl-N-(2-(((3R,4S,5R,6R)-3,4,5-triacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)ethyl)cyclopropanaminium iodide (1.00 g, 1 Eq, 1.70 mmol, as synthesized in Compound 310) is dissolved in methanol (3 mL), followed by addition of 0.5M NaOMe in methanol (18.4 mg, 0.68 mL, 0.2 Eq, 340 μmol). The reaction was stirred at room temperature for a few hours, then concentrated by rotary evaporation. The sample was redissolved in MeOH, then purified by column chromatography (C18AQ flash column, elutes at 100% Water with 0.1% TFA). The pure fractions were lyophilized and collected. The product was redissolved in water and stirred with Amberlyst IRA-400 Cl-resin (300 mg resin/100 mg of TFA salt), then lyophilized to yield the title compound (0.200 g, 35.8%). LCMS M+: 300.2. .sup.1H NMR (400 MHz, Deuterium Oxide) δ 5.15 (s, 2H), 4.41 (d, J=7.9 Hz, 1H), 4.35-4.25 (m, 1H), 4.09-3.99 (m, 1H), 3.83 (dd, J=12.3, 2.2 Hz, 1H), 3.79-3.69 (m, 2H), 3.63 (dd, J=12.3, 5.8 Hz, 1H), 3.44-3.33 (m, 2H), 3.33-3.17 (m, 2H), 3.20 (s, 6H).

##STR00192##

Compound 100: N,N-dimethyl-N-(2-{[(2R,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)oxan-2-yl]oxy}ethyl)cyclopropanaminium Chloride

[0689] N,N-dimethyl-N-(2-(((3R,4S,5R,6R)-3,4,5-triacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)ethyl)cyclopropanaminium iodide (1.00 g, 1 Eq, 1.70 mmol, as synthesized in Compound 312) is dissolved in methanol (3 mL), followed by addition of 0.5M NaOMe in methanol (18.4 mg, 0.68 mL, 0.2 Eq, 340 μmol). The reaction was stirred at room temperature for a few hours, then concentrated by rotary evaporation. The sample was redissolved in MeOH, then purified by column chromatography (C18AQ flash column, elutes at 100% Water with 0.1% TFA). The pure fractions were lyophilized and collected. The product was redissolved in water and stirred with Amberlyst IRA-400 Cl-resin (300 mg resin/100 mg of TFA salt), then lyophilized to yield the title compound (0.200 g, 35.8%). LCMS M+: 292.3. .sup.1H NMR (400 MHz, Deuterium Oxide) δ 4.40 (dd, J=8.0, 1.0 Hz, 1H), 4.37-4.28 (m, 1H), 4.12-4.02 (m, 1H), 3.82 (dd, J=12.4, 2.1 Hz, 1H), 3.69-3.57 (m, 3H), 3.44-3.33 (m, 2H), 3.33-3.24 (m, 1H), 3.24-3.16 (m, 1H), 3.16-3.08 (m, 1H), 2.93 (d, J=2.0 Hz, 6H), 1.18-1.09 (m, 2H), 0.91-0.80 (m, 2H).

##STR00193##

Compound 101: N,N-dimethyl-N-(2-{[(2S,3R,4S,5R,6R)-3,4,5-tris(acetyloxy)-6-[(acetyloxy)methyl]oxan-2-yl]oxy}ethyl)cyclopropanaminium

[0690] This compound may be synthesized according to the experimental procedure described for Compound 312.

##STR00194##

Compound 103: N,N-dimethyl-N-(prop-2-yn-1-yl)-2-((((2R,3R,4S,5R)-3,4,5-triacetoxy-6-methoxytetrahydro-2H-pyran-2-yl)methoxy)carbonyl)prop-2-en-1-aminium 2,2,2-trifluoroacetate

[0691] Compound 2-carboxy-N,N-dimethyl-N-(prop-2-yn-1-yl)prop-2-en-1-aminium (315 mg, 2 Eq, 1.87 mmol) and EDC (359 mg, 2 Eq, 1.87 mmol) were stirred in DMF (3 mL). Next (2R,3R,4S,5R)-2-(hydroxymethyl)-6-methoxytetrahydro-2H-pyran-3,4,5-triyl triacetate (300 mg, 1 Eq, 937 μmol) was added to the solution and the reaction was stirred at room temperature overnight. The reaction mixture was injected directly onto reverse phase C18 column chromatography (5% ACN in water with 0.1% TFA to 100% ACN), and product containing fractions were lyophilized to yield N,N-dimethyl-N-(prop-2-yn-1-yl)-2-((((2R,3R,4S,5R)-3,4,5-triacetoxy-6-methoxytetrahydro-2H-pyran-2-yl)methoxy)carbonyl)prop-2-en-1-aminium 2,2,2-trifluoroacetate (18 mg, 3.3%). LCMS M+: 470.1. .sup.1H NMR (400 MHz, DMSO-d6) δ 6.90 (s, 1H), 6.52 (s, 1H), 5.31 (t, J=9.7 Hz, 1H), 5.07 (t, J=9.9 Hz, 1H), 4.96-4.83 (m, 2H), 4.37 (d, J=2.5 Hz, 2H), 4.33-4.21 (m, 4H), 4.11-4.05 (m, 1H), 4.00 (dt, J=10.3, 3.3 Hz, 1H), 3.34 (s, 3H), 3.03 (s, 6H), 2.02-1.93 (m, 9H).

##STR00195##

Compound 104: N,N-dimethyl-N-(prop-2-yn-1-yl)-2-((((2R,3R,4S,5R)-3,4,5,6-tetraacetoxytetrahydro-2H-pyran-2-yl)methoxy)carbonyl)prop-2-en-1-aminium2,2,2-trifluoroacetate

[0692] 2-carboxy-N,N-dimethyl-N-(prop-2-yn-1-yl)prop-2-en-1-aminium (193 mg, 2 Eq, 1.15 mmol) and EDC (220 mg, 2 Eq, 1.15 mmol) were stirred in DMF (3 mL). Next (3R,4S,5R,6R)-6-(hydroxymethyl)tetrahydro-2H-pyran-2,3,4,5-tetrayltetraacetate (200 mg, 1 Eq, 574 mol) was added to the solution and the reaction was stirred at room temperature overnight. The reaction mixture was injected directly onto reverse phase C18 column chromatography (5% ACN in water with 0.1% TFA to 100% ACN; two column purifications were performed), and product containing fractions were lyophilized to yield N,N-dimethyl-N-(prop-2-yn-1-yl)-2-((((2R,3R,4S,5R)-3,4,5,6-tetraacetoxytetrahydro-2H-pyran-2-yl)methoxy)carbonyl)prop-2-en-1-aminium2,2,2-trifluoroacetate (33 mg, 9.4%). LCMS M+: 498.1. .sup.1HNMR (400 MHz, DMSO-d6) δ 6.88 (s, 1H), 6.51 (s, 1H), 5.94 (d, J=8.3 Hz, 1H), 5.45 (t, J=9.6 Hz, 1H), 5.08 (t, J=9.4 Hz, 1H), 5.00 (dd, J=9.7, 8.3 Hz, 1H), 4.36 (d, J=2.5 Hz, 2H), 4.32-4.16 (m, 5H), 4.08 (t, J=2.4 Hz, 1H), 3.02 (s, 6H), 2.11-1.90 (m, 12H).

##STR00196##

Compound 105: N,N-dimethyl-N-(2-(((3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)ethyl)prop-2-yn-1-aminium 2,2,2-trifluoroacetate

[0693] Step 1:

[0694] To a solution of compound 1 (200 g, 512 mmol, 1.00 eq) in DCM (1000 mL) was added 4A molecular sieves (100 g). 2-iodoethanol (88.1 g, 512 mmol, 40.0 mL, 1.00 eq) was added to the suspension drop-wise at 0° C. and the resulting suspension was stirred at 0° C. for 30 mins. BF.sub.3.Et.sub.2O (218 g, 1.54 mol, 190 mL, 3.00 eq) was added to the suspension drop-wise at 0° C. under N.sub.2. After addition, the suspension was stirred at 20° C. for 18 hrs. TLC (Petroleum ether:Ethyl acetate=2:1, R.sub.f of material=0.4) showed the reaction was completed. K.sub.2CO.sub.3 (280 g) was added to the suspension in one portion, and the suspension was stirred for 30 mins. The suspension was filtered. The filtrate was washed by water (1500 mL×2) followed by brine (1000 mL). The organic layer was dried over Na.sub.2SO.sub.4, filtered and concentrated under vacuum to give a crude product. The crude product was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=100/1 to 5/1) to give a solid. The solid was re-crystallized from MTBE/Petroleum ether (3:1, 300 mL) to give compound 2 (195 g, 388 mmol, 37.9% yield) as a white solid.

[0695] Step 2:

[0696] To a solution of compound 2 (80 g, 159 mmol, 1.00 eq) in acetone (500 mL) was added compound 2A (19.9 g, 239 mmol, 25.3 mL, 1.50 eq). After addition, the solution was stirred at 40° C. for 16 hrs. TLC (Petroleum ether:Ethyl acetate=2:1, Rf of material=0.5) showed the reaction was completed. The solution was cooled to 0° C. and solid precipitated out. The solid was filtered, collected and concentrated under vacuum to give 3 (100 g, 171 mmol, 53.6% yield) as a white solid.

[0697] Step 3:

[0698] N,N-dimethyl-N-(2-(((3R,4S,5R,6R)-3,4,5-triacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)ethyl)prop-2-yn-1-aminium iodide (3.00 g, 1 Eq, 5.12 mmol) is dissolved in methanol (3 mL), followed by addition of 0.5M NaOMe in methanol (sodium methoxide (277 mg, 10 mL, 1 Eq, 5.12 mmol)). The reaction was stirred at room temperature for a few hours, then concentrated by rotary evaporation. The sample was dissolved in water, then purified by column chromatography (C18AQ flash column, elutes at 100% Water with 0.1% TFA). The pure fractions were lyophilized to yield product N,N-dimethyl-N-(2-(((3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)ethyl)prop-2-yn-1-aminium 2,2,2-trifluoroacetate (590 mg, 28.5%)).

[0699] Step 4:

[0700] Compound N,N-dimethyl-N-(2-(((2R,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)ethyl)prop-2-yn-1-aminium (0.300 g, 1 Eq, 1.03 mmol) was stirred in acetic-2,2,2-d3 anhydride (1.12 g, 977 μL, 10 Eq, 10.3 mmol) and pyridine (817 mg, 836 μL, 10 Eq, 10.3 mmol). The reaction was diluted in dichloromethane then purified by column chromatography (100% DCM to 10% MeOH in DCM; product elutes around 8% MeOH in DCM). Fractions containing product were dried in vacuo to yield N,N-dimethyl-N-(2-(((2R,3R,4S,5R,6R)-3,4,5-tris(acetoxy-d3)-6-((acetoxy-d3)methyl)tetrahydro-2H-pyran-2-yl)oxy)ethyl)prop-2-yn-1-aminium 2,2,2-trifluoroacetate (75 mg, 12%). LCMS M+: 470.4. .sup.1H NMR (400 MHz, DMSO-d6) δ 5.26 (t, J=9.5 Hz, 1H), 4.99-4.74 (m, 3H), 4.36 (d, J=2.5 Hz, 2H), 4.24-3.94 (m, 6H), 3.61 (t, J=5.0 Hz, 2H), 3.08 (s, 6H).

##STR00197##

Compound 106: N,N-dimethyl-N-(2-(((3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)ethyl)prop-2-yn-1-aminium 2,2,2-trifluoroacetate

[0701] Step 1:

[0702] To a solution of compound 1 (200 g, 512 mmol, 1.00 eq) in DCM (1000 mL) was added 4A molecular sieves (100 g). 2-iodoethanol (88.1 g, 512 mmol, 40.0 mL, 1.00 eq) was added to the suspension drop-wise at 0° C. and the resulting suspension was stirred at 0° C. for 30 mins. BF.sub.3.Et.sub.2O (218 g, 1.54 mol, 190 mL, 3.00 eq) was added to the suspension drop-wise at 0° C. under N.sub.2. After addition, the suspension was stirred at 20° C. for 18 hrs. TLC (Petroleum ether:Ethyl acetate=2:1, Rf of material=0.4) showed the reaction was completed. K.sub.2CO.sub.3 (280 g) was added to the suspension in one portion, and the suspension was stirred for 30 mins. The suspension was filtered. The filtrate was washed by water (1500 mL×2) followed by brine (1000 mL). The organic layer was dried over Na.sub.2SO.sub.4, filtered and concentrated under vacuum to give a crude product. The crude product was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=100/1 to 5/1) to give a solid. The solid was re-crystallized from MTBE/Petroleum ether (3:1, 300 mL) to give compound 2 (195 g, 388 mmol, 37.9% yield) as a white solid.

[0703] Step 2:

[0704] To a solution of compound 2 (80 g, 159 mmol, 1.00 eq) in acetone (500 mL) was added compound 2A (19.9 g, 239 mmol, 25.3 mL, 1.50 eq). After addition, the solution was stirred at 40° C. for 16 hrs. TLC (Petroleum ether:Ethyl acetate=2:1, Rf of material=0.5) showed the reaction was completed. The solution was cooled to 0° C. and lots of solid precipitated out. The solid was filtered, collected and concentrated under vacuum to give 3 (100 g, 171 mmol, 53.6% yield) as a white solid.

[0705] Step 3:

[0706] N,N-dimethyl-N-(2-(((3R,4S,5R,6R)-3,4,5-triacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)ethyl)prop-2-yn-1-aminium iodide (3.00 g, 1 Eq, 5.12 mmol) is dissolved in methanol (3 mL), followed by addition of 0.5M NaOMe in methanol (sodium methoxide (277 mg, 10 mL, 1 Eq, 5.12 mmol)). The reaction was stirred at room temperature for a few hours, then concentrated by rotary evaporation. The sample was dissolved in water, then purified by column chromatography (C18AQ flash column, elutes at 100% Water with 0.1% TFA). The pure fractions were lyophilized to yield product N,N-dimethyl-N-(2-(((3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)ethyl)prop-2-yn-1-aminium 2,2,2-trifluoroacetate (590 mg, 28.5%)). LCMS M+: 290.2. .sup.1H NMR (400 MHz, Deuterium Oxide) δ 4.43 (d, J=7.9 Hz, 1H), 4.36-4.23 (m, 3H), 4.11-4.00 (m, 1H), 3.84 (dd, J=12.4, 2.2 Hz, 1H), 3.73-3.61 (m, 3H), 3.46-3.35 (m, 2H), 3.34-3.18 (m, 3H), 3.18 (s, 6H).

##STR00198##

Compound 107 and Compound 131: N-(cyclopropylmethyl)-2-hydroxy-N,N-dimethylethan-1-aminium Bromide

[0707] To a solution of 2-(dimethylamino) ethanol (1 g, 11.22 mmol, 1.13 mL, 1 eq) in THF (10 mL) was added (bromomethyl) cyclopropane (1.67 g, 12.34 mmol, 1.18 mL, 1.1 eq) at 20° C. The mixture was stirred at 20° C. for 12 h. The reaction mixture was filtered to give a residue. The residue was diluted with H.sub.2O (10 mL) and extracted with ethyl acetate (10 mL). The water layer was freeze-drying to give N-(cyclopropylmethyl)-2-hydroxy-N, N-dimethylethanaminium bromide (170 mg, 758.46 umol, 6.76% yield) as a white solid. MS: M.sup.+=144.1 .sup.1H NMR (400 MHz, D2O) δ 4.0 (m, 2H), 3.5 (m, 2H), 3.2 (d, 2H), 3.1 (s, 6H), 1.2 (m, 1H), 0.8 (m, 2H), 0.4 (m, 2H) ppm.

##STR00199##

Compound 108: (2S)-2-(hydroxymethyl)-1-methyl-1-(prop-2-yn-1-yl)pyrrolidin-1-ium bromide

[0708] To a solution of [(2S)-1-methylpyrrolidin-2-yl] methanol (200 mg, 1.74 mmol, 206.19 uL, 1 eq) in DCM (5 mL) was added 3-bromoprop-1-yne (247.89 mg, 2.08 mmol, 179.63 uL, 1.2 eq). The mixture was stirred at 25° C. for 5 h. Some solid precipitate out. The mixture was filtered and the filter cake was washed with DCM (5 mL*2) and concentrated to dryness. Compound (2S)-2-(hydroxymethyl)-1-methyl-1-(prop-2-yn-1-yl) pyrrolidin-1-ium bromide (346 mg, crude) was obtained as a white solid. .sup.1H NMR (400 MHz, DMSO) δ 5.5 (m, 1H), 4.3 (m, 2H), 3.8 (m, 5H), 3.3 (s 6H), 2.0 (m, 4H) ppm.

##STR00200##

Compound 109: (2S)-1-(fluoromethyl)-2-(hydroxymethyl)-1-methylpyrrolidin-1-ium iodide

[0709] To a solution of (3S)-1-methylpyrrolidin-3-ol (200 mg, 1.98 mmol, 201.41 uL, 1 eq) in DCM (5 mL) was added fluoro (iodo) methane (379.48 mg, 2.37 mmol, 1.2 eq). The mixture was stirred at 25° C. for 5 h. MS showed desired compound was detected. The reaction mixture was concentrated under reduced pressure to remove solvent. The residue was dissolved in H.sub.2O (1 mL), washed with EtOAc (2 mL*2) and lyophilized the aqueous phase to give a residue. Compound (2S)-1-(fluoromethyl)-2-(hydroxymethyl)-1-methylpyrrolidin-1-ium iodide (111 mg, crude) was obtained as a white solid. .sup.1H NMR (400 MHz, DMSO) δ 5.6 (m, 3H), 3.8 (m, 4H), 3.5 (m, 1H), 3.1 (s, 3H), 2.0 (m, 4H) ppm

##STR00201##

Compound 110: 2-(hydroxymethyl)-1,1-dimethylpyrrolidin-1-ium Iodide

[0710] To a solution of (1-methylpyrrolidin-2-yl) methanol (300 mg, 2.60 mmol, 1 eq) and DMAP (159.11 mg, 1.30 mmol, 0.5 eq) in THF (5 mL) was added (Boc).sub.2O (1.42 g, 6.51 mmol, 1.50 mL, 2.5 eq) at 25° C. The mixture was stirred at 25° C. for 12 h. LCMS showed the desired compound was detected. The reaction mixture was filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=3:1 to 1:1) to give tert-butyl ((1-methylpyrrolidin-2-yl) methyl) carbonate (190 mg, 882.54 umol, 33.88% yield) as a yellow oil.

[0711] To a solution of tert-butyl ((1-methylpyrrolidin-2-yl) methyl) carbonate (190 mg, 882.54 umol, 1 eq) in THF (5 mL) was added MeI (250.53 mg, 1.77 mmol, 109.88 uL, 2 eq) at 25° C. The mixture was stirred at 25° C. for 5 h. LCMS showed the desired compound was detected. The reaction mixture was diluted with H.sub.2O (10 mL) and extracted with ethyl acetate (10 mL). The water layer was freeze-drying to give 2-(((tert-butoxycarbonyl)oxy)methyl)-1,1-dimethylpyrrolidin-1-ium iodide (200 mg, 559.87 umol, 63.44% yield) as a yellow solid.

[0712] To a solution of 2-(((tert-butoxycarbonyl)oxy)methyl)-1,1-dimethylpyrrolidin-1-ium iodide (180 mg, 503.88 umol, 1 eq) in HCl/dioxane (0.5 mL, 4M) and dioxane (1 mL) was stirred at 25° C. for 2 h. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was diluted with H.sub.2O (10 mL) and extracted with ethyl acetate (10 mL). The water layer was freeze-drying to give 2-(hydroxymethyl)-1,1-dimethylpyrrolidin-1-ium iodide (70 mg, 272.26 umol, 54.03% yield) as a yellow solid. MS: M.sup.+=130.1 .sup.1H NMR (400 MHz, DMSO) δ 3.9 (m, 2H), 3.7 (m, 1H), 3.5 (m, 2H), 3.2 (s, 3H), 3.0 (s, 3H), 2.3 m, 1H0, 2.1 (m, 2H), 1.8 (m, 1H) ppm.

##STR00202##

Compound 111: N-(2-hydroxyethyl)-N,N-dimethylcyclopropanaminium Iodide

[0713] To a solution of N-methylcyclopropanamine (150 mg, 2.11 mmol, 1.68 eq) and (2-bromoethoxy)(tert-butyl) dimethylsilane (300 mg, 1.25 mmol, 1 eq) in ACN (5 mL) was added K.sub.2CO.sub.3 (173.32 mg, 1.25 mmol, 1 eq) at 25° C. The mixture was stirred at 50° C. for 12 h. TLC indicated new spot formed. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=5:1 to 1:1) to give N-(2-((tert-butyldimethylsilyl)oxy)ethyl)-N-methylcyclopropanamine (120 mg, 523.03 umol, 41.71% yield) as colorless oil.

[0714] To a solution of N-(2-((tert-butyldimethylsilyl) oxy) ethyl)-N-methylcyclopropanamine (70 mg, 305.10 umol, 1 eq) in THF (2 mL) was added MeI (86.61 mg, 610.20 umol, 37.99 uL, 2 eq) at 25° C. The mixture was stirred at 25° C. for 12 h. The solid was filtered and the filter cake was concentrated to give N-(2-((tert-butyldimethylsilyl) oxy) ethyl)-N, N-dimethylcyclopropanaminium iodide (50 mg, 134.64 umol, 44.13% yield) as a white solid.

[0715] To a solution of N-(2-((tert-butyldimethylsilyl) oxy) ethyl)-N, N-dimethylcyclopropanaminium iodide (20 mg, 53.85 umol, 1 eq) in H.sub.2O (2 mL) was added KF (31.29 mg, 538.54 umol, 12.62 uL, 10 eq) at 25° C. The mixture was stirred at 25° C. for 12 h. The precipitation was separated out by filtration. And the filtrate was diluted with H.sub.2O (10 mL) and extracted with ethyl acetate (10 mL). The water layer was concentrated under reduced pressure to give a residue. The residue was washed by acetonitrile (2 mL). The acetonitrile layer was filtered and the filtrate was concentrated under reduced pressure to give N-(2-hydroxyethyl)-N, N-dimethylcyclopropanaminium iodide (10 mg, 38.89 umol, 72.22% yield) as a white solid. .sup.1H NMR (400 MHz, D2O) δ 4.1 (m, 2H), 3.6 (m, 2H), 3.2 (m, 1H), 3.0 (s, 6H), 1.2 (m, 2H), 0.9 (m, 2H) ppm.

##STR00203##

Compound 112: (3S)-3-hydroxy-1-methyl-1-(prop-2-yn-1-yl)pyrrolidin-1-ium bromide

[0716] To a solution of (3S)-1-methylpyrrolidin-3-ol (200 mg, 1.98 mmol, 201.41 uL, 1 eq) in DCM (5 mL) was added 3-bromoprop-1-yne (282.27 mg, 2.37 mmol, 204.54 uL, 1.2 eq). The mixture was stirred at 25° C. for 5 h. MS showed desired compound was detected. The reaction mixture was concentrated under reduced pressure to remove solvent. The residue was dissolved in H.sub.2O (1 mL), washed with EtOAc (2 mL*2) and lyophilized the aqueous phase to give a residue. Compound (3S)-3-hydroxy-1-methyl-1-(prop-2-yn-1-yl) pyrrolidin-1-ium bromide (311 mg, crude) was obtained as yellow oil. .sup.1H NMR (400 MHz, DMSO) δ 5.6 (m, 1H), 4.4 (m, 3H), 4.0 (m, 1H), 3.6 (m, 4H), 3.1 (m, 3H), 2.4 (m, 1H), 1.9 (m 1H) ppm.

##STR00204##

Compound 113: (3R)-3-hydroxy-1-methyl-1-(prop-2-yn-1-yl)pyrrolidin-1-ium bromide

[0717] To a solution of (3R)-1-methylpyrrolidin-3-ol (200 mg, 1.98 mmol, 217.16 uL, 1 eq) in DCM (5 mL) was added 3-bromoprop-1-yne (282.27 mg, 2.37 mmol, 204.54 uL, 1.2 eq). The mixture was stirred at 25° C. for 5 h. MS showed desired compound was detected. The reaction mixture was concentrated under reduced pressure to remove solvent. The residue was dissolved in H.sub.2O (1 mL), washed with EtOAc (2 mL*2) and lyophilized the aqueous phase to give a residue. Compound (3R)-3-hydroxy-1-methyl-1-(prop-2-yn-1-yl) pyrrolidin-1-ium bromide (368 mg, crude) was obtained as yellow oil. .sup.1H NMR (400 MHz, DMSO) δ 5.6 (m, 1H), 4.4 (m, 3H), 4.0 (m, 1H), 3.6 (m, 4H), 3.1 (m, 3H), 2.4 (m, 1H), 1.9 (m 1H) ppm.

##STR00205##

Compound 114: (3S)-1-(fluoromethyl)-3-hydroxy-1-methylpyrrolidin-1-ium Iodide

[0718] To a solution of (3S)-1-methylpyrrolidin-3-ol (200 mg, 1.98 mmol, 201.41 uL, 1 eq) in DCM (5 mL) was added fluoro (iodo) methane (379.48 mg, 2.37 mmol, 1.2 eq). The mixture was stirred at 25° C. for 5 h. MS showed desired compound was detected. The reaction mixture was concentrated under reduced pressure to remove solvent. The residue was dissolved in H.sub.2O (1 mL), washed with EtOAc (2 mL*2) and lyophilized the aqueous phase to give a residue. Compound (3S)-1-(fluoromethyl)-3-hydroxy-1-methylpyrrolidin-1-ium iodide (111 mg, crude) was obtained as a white solid. .sup.1H NMR (400 MHz, DMSO) δ 5.7 (m, 1H), 5.4 (m, 2H), 4.5 (m, 1H), 3.6 (m, 4H), 3.2 (s, 3H), 2.4 (m, 1H), 1.9 (m, 1H) ppm.

##STR00206##

Compound 115: (3R)-1-(fluoromethyl)-3-hydroxy-1-methylpyrrolidin-1-ium Iodide

[0719] To a solution of (3R)-1-methylpyrrolidin-3-ol (0.2 g, 1.98 mmol, 217.16 uL, 1 eq) in DCM (5 mL) was added fluoro (iodo) methane (379.48 mg, 2.37 mmol, 1.2 eq). The mixture was stirred at 25° C. for 5 h. MS showed desired compound was detected. The reaction mixture was concentrated under reduced pressure to remove solvent. The residue was dissolved in H.sub.2O (1 mL), washed with EtOAc (2 mL*2) and lyophilized the aqueous phase to give a residue. Compound (3R)-1-(fluoromethyl)-3-hydroxy-1-methylpyrrolidin-1-ium iodide (358 mg, crude) was obtained as yellow oil. .sup.1H NMR (400 MHz, DMSO) δ 5.7 (m, 1H), 5.4 (m, 2H), 4.5 (m, 1H), 3.6 (m, 4H), 3.2 (s, 3H), 2.4 (m, 1H), 1.9 (m, 1H) ppm.

##STR00207##

Compound 116: (R)-3-hydroxy-1,1-dimethylpyrrolidin-1-ium Iodide

[0720] To a solution of (3R)-pyrrolidin-3-ol (4 g, 32.37 mmol, 3.81 mL, 1 eq, HCl) in THF (50 mL) was added NaOH (1.29 g, 32.37 mmol, 1 eq) at 25° C. After addition, the mixture was stirred at this temperature for 20 min. Then paraformaldehyde (1.17 g, 38.84 mmol, 1.2 eq) and HCOOH (4.82 g, 100.34 mmol, 3.1 eq) were added 25° C. The resulting mixture was stirred at 60° C. for 5 h 40 min. TLC indicated the reaction was completed. The reaction mixture was cooled to 0° C., quenched by addition 10N NaOH to pH about 10, and extracted with EtOAc (30 mL*3). The combined organic layers were washed with H.sub.2O (100 mL*1), dried over Na.sub.2SO.sub.4, filtered and concentrated under reduced pressure to give a residue. The crude product (3R)-1-methylpyrrolidin-3-ol (0.7 g, crude) as yellow oil was used into the next step without further purification.

[0721] To a solution of (3R)-1-methylpyrrolidin-3-ol (0.2 g, 1.98 mmol, 217.16 uL, 1 eq) in DCM (10 mL) was added MeI (561.32 mg, 3.95 mmol, 246.19 uL, 2 eq). The mixture was stirred at 25° C. for 12 h. Some solid precipitate out and the solid was filtered. The filter cake was washed with DCM (5 mL*2) and concentrated to dryness. Compound (R)-3-hydroxy-1,1-dimethylpyrrolidin-1-ium iodide (333 mg, crude) was obtained as a white solid. .sup.1H NMR (400 MHz, DMSO) δ 5.5 (s, 1H), 4.5 (m, 1H), 3.6 (4H), 3.2 (s, 3H), 3.1 (s, 3H), 2.4 (m, 1H), 1.9 (m, 1H) ppm.

##STR00208##

Compound 117: (S)-3-hydroxy-1,1-dimethylpyrrolidin-1-ium Iodide

[0722] To a solution of (3S)-pyrrolidin-3-ol (4 g, HCl salt, 32.37 mmol, 3.70 mL, 1 eq) in THF (50 mL) was added NaOH (1.29 g, 32.37 mmol, 1 eq) at 25° C. After addition, the mixture was stirred at this temperature for 20 min. Then Paraformaldehyde (1.13 g, 38.84 mmol, 1.2 eq) and HCOOH (4.82 g, 100.34 mmol, 3.1 eq) were added at 25° C. The resulting mixture was stirred at 60° C. for 5 h 40 min. TLC indicated the reaction was completed. The reaction mixture was cooled to 0° C. and quenched by addition 10N NaOH to pH about 10, and extracted with EtOAc (30 mL*3). The combined organic layers were washed with H.sub.2O (100 mL*1), dried over Na.sub.2SO.sub.4, filtered and concentrated under reduced pressure to give a residue. The crude product (3S)-1-methylpyrrolidin-3-ol (0.7 g, crude) as yellow oil was used into the next step without further purification.

[0723] To a solution of (3S)-1-methylpyrrolidin-3-ol (0.2 g, 1.98 mmol, 201.41 uL, 1 eq) in DCM (10 mL) was added MeI (561.32 mg, 3.95 mmol, 246.19 uL, 2 eq). The mixture was stirred at 25° C. for 12 h. Some solid precipitate out and the solid was filtered. The filter cake was washed with DCM (5 mL*2) and concentrated to dryness. Compound (S)-3-hydroxy-1,1-dimethylpyrrolidin-1-ium iodide (272 mg, crude) was obtained as a white solid. .sup.1H NMR (400 MHz, DMSO) δ 5.5 (s, 1H), 4.5 (m, 1H), 3.6 (m, 4H), 3.2 (s, 3H), 3.1 (s, 3H), 2.4 (m, 1H), 1.0 (m, 1H) ppm.

##STR00209##

Compound 118: (1R,2S)-2-hydroxy-N,N,N-trimethylcyclopropan-1-aminium Iodide

[0724] (1R, 2S), 2-dimethylamino cyclopropanol is dissolved in THF followed by the dropwise addition of methyl iodide. The resulting precipitate is collected and dried and recrystallized to afford the title compound.

##STR00210##

Compound 119: (1S,2R)-2-hydroxy-N,N,N-trimethylcyclopropan-1-aminium Iodide

[0725] (1S, 2R), 2-dimethylamino cyclopropanol is dissolved in THF followed by the dropwise addition of methyl iodide. The resulting precipitate is collected and dried and recrystallized to afford the title compound.

##STR00211##

Compound 120: (1S,2R)-2-hydroxy-N,N-dimethyl-N-(prop-2-yn-1-yl)cyclopropan-1-aminium Iodide

[0726] (1S, 2R), 2-dimethylamino cyclopropanol is dissolved in THF followed by the dropwise addition of propargyl iodide. The resulting precipitate is collected and dried and recrystallized to afford the title compound.

##STR00212##

Compound 121: (1R,2S)-2-hydroxy-N,N-dimethyl-N-(prop-2-yn-1-yl)cyclopropan-1-aminium Iodide

[0727] (1R, 2S), 2-dimethylamino cyclopropanol is dissolved in THF followed by the dropwise addition of propargyl iodide. The resulting precipitate is collected and dried and recrystallized to afford the title compound.

##STR00213##

Compound 122: (2R)-2-ethynyl-1-(2-hydroxyethyl)-1-methylazetidin-1-ium Iodide

[0728] (2R) 2-ethynyl azetidine is dissolved in THF water and 2-hydroxyacetaldehyde is added to the mixture. The resulting mixture is stirred at room temperature and then sodium cyanoborohydide is added portion wise to the mixture. The mixture is stirred for 1 hour then quenched by the addition of water ethyl acetate. The resulting material is taken up in ether, and treated with dropwise addition of methyl iodide. The resulting precipitate is collected and dried and recrystallized to afford the title compound.

##STR00214##

Compound 123: (1S,2R)-2-hydroxy-N,N,N-trimethylcyclobutan-1-aminium Iodide

[0729] (1S,2R)-2-hydroxy dimethylamino cyclobutanol is dissolved in THF and then treated with the dropwise addition of methyl iodide. The resulting precipitate is collected and dried and recrystallized to afford the title compound.

##STR00215##

Compound 124: (2S)-2-ethynyl-1-(2-hydroxyethyl)-1-methylazetidin-1-ium Iodide

[0730] (2S) 2-ethynyl azetidine is dissolved in THF water and 2-hydroxyacetaldehyde is added to the mixture. The resulting mixture is stirred at room temperature and then sodium cyanoborahydide is added portion wise to the mixture. The mixture is stirred for 1 hour then quenched by the addition of water ethyl acetate. The resulting material is taken up in ether, and treated with dropwise addition of methyl iodide. The resulting precipitate is collected and dried and recrystallized to afford the title compound.

##STR00216##

Compound 125: 1-methyl-1-(prop-2-yn-1-yl)pyrazolidin-1-ium Iodide

[0731] Step 1:

[0732] To a solution of tert-butyl pyrazolidine-1-carboxylate (200 mg, 1.16 mmol, 1 eq) in THF (10 mL) was added K2CO3 (176.54 mg, 1.28 mmol, 1.1 eq) at 15° C. and stirred at 15° C. for 10 min. Then 3-bromoprop-1-yne (207.22 mg, 1.74 mmol, 150.16 uL, 1.5 eq) was added to the mixture and the mixture was stirred at 15° C. for 12 h. TLC indicated reactant was consumed completely. The reaction mixture was filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by prep-TLC (SiO.sub.2, PE:EtOAc=1:1). Compound tert-butyl 2-prop-2-ynylpyrazolidine-1-carboxylate (170 mg, 808.48 umol, 69.62% yield) was obtained as a white solid.

[0733] Step 2:

[0734] To a solution of tert-butyl 2-prop-2-ynylpyrazolidine-1-carboxylate (170 mg, 808.48 umol, 1 eq) in THF (5 mL) was added MeI (688.52 mg, 4.85 mmol, 301.98 uL, 6 eq). The mixture was stirred at 15° C. for 12 h. MS showed the desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The crude product tert-butyl 2-methyl-2-prop-2-ynyl-pyrazolidin-2-ium-1-carboxylate (200 mg, crude, I—) as a yellow solid was used into the next step without further purification. LCMS: (M+): 225.2

[0735] Step 3:

[0736] To a solution of tert-butyl 2-methyl-2-prop-2-ynyl-pyrazolidin-2-ium-1-carboxylate (200 mg, 887.68 umol, 1 eq) in HCl/dioxane (4 M, 5 mL, 22.53 eq). The mixture was stirred at 15° C. for 4 h. MS showed the desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC ([water (0.1% TFA)-ACN]). Compound 1-methyl-1-(prop-2-yn-1-yl)pyrazolidin-1-ium iodide (26 mg, 103.14 umol, 11.6% yield) was obtained as a colorless oil. LCMS: (M+): 125.1 .sup.1H NMR (400 MHz, Deuterium Oxide) δ 4.42-4.28 (m, 2H), 3.91-3.80 (m, 1H), 3.75-3.64 (m, 1H), 3.44-3.23 (m, 5H), 3.16-3.11 (m, 1H), 2.37 (p, J=7.4 Hz, 2H).

##STR00217##

Compound 126: (1S,3S)-3-hydroxy-N,N-dimethyl-N-(prop-2-yn-1-yl-)cyclobutan-1-aminium Iodide

[0737] Cis-3-dimethylamine cyclobutanol is taken up in THF and treated with drop wise addition of propargyl iodide. The resulting precipitate is collected and dried and recrystallized to afford the title compound.

##STR00218##

Compound 127: 3-carboxy-N,N,N-trimethylpropan-1-aminium Iodide

[0738] 4-(dimethylamino)butanoic acid is taken up in THF and treated with drop wise addition of methyl iodide. The resulting precipitate is collected and dried and recrystallized to afford the title compound.

##STR00219##

Compound 129: (1R,2R)-2-(hydroxymethyl)-1-methyl-1-(prop-2-yn-1-yl)pyrrolidin-1-ium Bromide

[0739] To a solution of [(2R)-1-methylpyrrolidin-2-yl]methanol (200 mg, 1.74 mmol, 1 eq) in THF (10 mL) was added 3-bromoprop-1-yne (826.30 mg, 6.95 mmol, 598.77 uL, 4 eq). The mixture was stirred at 15° C. for 24 hr. MS showed the desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC ([water (0.10% TFA)-ACN]). The title compound (60 mg, 389.03 umol, 22.40% yield, Br) was obtained as a colorless oil. LCMS:(M+) 154.1 .sup.1H NMR (400 MHz, Deuterium Oxide) δ 4.46-4.23 (m, 1H), 4.15 (d, J=2.5 Hz, 1H), 4.04-3.69 (m, 4H), 3.63-3.39 (m, 1H), 3.24 (s, 1H), 3.14-3.01 (m, 3H), 2.33-2.02 (m, 3H), 1.99-1.77 (m, 1H).

##STR00220##

Compound 132: 4-hydroxy-N,N,N-trimethylbut-2-yn-1-aminium Chloride

[0740] To a solution of 4-chlorobut-2-yn-1-ol (88.17 mg, 843.45 umol, 1 eq) in THE (5 mL) was added N, N-dimethylmethanamine (3 M, 562.30 uL, 2 eq, in THF). The mixture was stirred at 25° C. for 12 h. The reaction mixture was diluted with H.sub.2O (10 mL) and extracted with ethyl acetate (10 mL). The water layer was freeze-drying to give 4-hydroxy-N, N, N-trimethylbut-2-yn-1-aminium chloride (55 mg, 336.09 umol, 39.85% yield) as colorless oil. MS: M.sup.+=128.1 .sup.1H NMR (400 MHz, DMSO) δ 4.3 (m, 4H), 3.1 (s, 9H) ppm.

##STR00221##

Compound 134: 2-[(benzyloxy)methyl]-1,1-dimethylpyrrolidin-1-ium Iodide

[0741] The title compound may be synthesized by reacting Compound 57 with benzyl bromide, followed by purification by recrystallization.

##STR00222##

Compound 135: 1-(carboxymethyl)-1-(prop-2-yn-1-yl)azetidin-1-ium

[0742] This compound may be synthesized according to the experimental procedure described for Compound 133.

##STR00223##

Compound 137: 3-ethynyl-1,1-dimethylpyrrolidin-1-ium Iodide

[0743] This compound may be synthesized according to the experimental procedure described for Compound 262.

##STR00224##

Compound 138: N-(3-hydroxypropyl)-N,N-dimethylprop-2-yn-1-aminium Trifluoroacetate

[0744] The mixture of N,N-dimethylprop-2-yn-1-amine (500 mg, 6.01 mmol, 637.76 uL, 1 eq) and 3-bromopropan-1-ol (1.00 g, 7.22 mmol, 651.40 uL, 1.2 eq) in EtOH (5 mL) was stirred at 80° C. under N.sub.2 2 hours. LCMS showed desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. Then the residue was purified by prep-HPLC (column: Xbridge 150*30 mm*10 um; mobile phase: [water (0.10% TFA)-ACN]; B %: 1%-10%, 10 min) to give N-(3-hydroxypropyl)-N,N-dimethylprop-2-yn-1-aminium trifluoroacetate (107 mg, 409.35 umol, 6.81% yield, TFA) as a colorless oil. LCMS: (M+) 142.2 .sup.1H NMR (400 MHz, Deuterium Oxide) δ 4.08 (d, J=2.6 Hz, 2H), 3.53 (t, J=5.9 Hz, 2H), 3.40-3.32 (m, 2H), 3.07 (t, J=2.5 Hz, 1H), 3.01 (s, 6H), 1.91-1.79 (m, 2H).

##STR00225##

Compound 139: 5-hydroxy-N,N-dimethyl-N-(prop-2-yn-1-yl)pentan-1-aminium Bromide

[0745] A mixture of N,N-dimethylprop-2-yn-1-amine (500 mg, 6.01 mmol, 637.76 uL, 1 eq) and 5-bromopentan-1-ol (1.05 g, 6.31 mmol, 1.05 eq) in EtOH (5 mL) was degassed and purged with N.sub.2 3 times, and then the mixture was stirred at 80° C. for 10 hr under N.sub.2 atmosphere. LCMS showed desired mass was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was diluted with H.sub.2O 20 mL and washed with EtOAc (20 mL*3). Then the water layers were lyophilized to afford 5-hydroxy-N,N-dimethyl-N-(prop-2-yn-1-yl)pentan-1-aminium bromide (625 mg, 1.98 mmol, 32.90% yield) as a yellow oil. LCMS: (M+) 170.1 .sup.1H NMR (400 MHz, Deuterium Oxide) δ 4.17 (s, 2H), 3.55 (t, J=6.5 Hz, 2H), 3.42-3.33 (m, 2H), 3.19-3.14 (m, 1H), 3.10 (s, 6H), 1.81-1.69 (m, 2H), 1.61-1.49 (m, 2H), 1.42-1.30 (m, 2H).

##STR00226##

Compound 140: (methanesulfinylmethyl)trimethylazanium Chloride

[0746] Step 1:

[0747] To a solution of chloro(methylsulfanyl)methane (200 mg, 2.07 mmol, 173.91 uL, 1 eq) in THF (5 mL) was added N,N-dimethylmethanamine (1 M, 4.14 mL, 2 eq, in THF) at 15° C. The mixture was stirred at 70° C. for 12 hr. The precipitation was found. The reaction mixture was filtered to give a residue. Compound methylsulfanyl-[N,N,N-(trimethyl)-azanyl]methane (100 mg, crude) was obtained as a white solid.

[0748] Step 2:

[0749] A mixture of methylsulfanyl-[N,N,N (trimethyl)-azanyl]methane (50 mg, 321.15 umol, 1 eq) in H.sub.2O.sub.2 (2 mL, 30%) and H.sub.2O (2 mL) was stirred at 80° C. for 12 hr. LCMS showed the desired compound was detected. The reaction mixture was quenched by saturated sodium sulfite (30 mL), and the aqueous phase was freeze-dried. The residue was diluted with methanol (50 mL) and filtered. The filtrate was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (column: Waters Xbridge BEH C18 100*30 mm*10 um; mobile phase: [water (0.1% TFA)-ACN]; B %: 1%-20%, 10 min). The residue was purified by prep-HPLC (column: YMC-Actus Triart Diol-Hilic 100*30 mm Sum; mobile phase: [water (0.1% TFA)-ACN]; B %: 99%-85%, 12 min). The title compound (5 mg, 28.46 umol, Cl-salt, 24.43% yield, 97.73% purity) was obtained as a colorless oil. LCMS: (M+): 136.1 1H NMR (400 MHz, Deuterium Oxide) δ 4.62-4.53 (m, 2H), 3.25 (s, 9H), 2.75 (s, 3H).

##STR00227##

Compound 140A: 3-(trimethylammonio)propane-1-sulfonate

[0750] To a solution of oxathiolane 2, 2-dioxide (200 mg, 1.64 mmol, 143.88 uL, 1 eq) in acetone (5 mL) was added N,N-dimethylmethanamine (322.63 mg, 1.64 mmol, 377.34 uL, 1 eq) at 25° C. The mixture was stirred at 25° C. for 12 h. The precipitate was separated from the reaction mixture by filtration and the solid was concentrated under reduced pressure to dryness. Compound 3-(trimethylammonio) propane-1-sulfonate (157 mg, 0.87 mmol, 53% yield) was obtained as a white solid. LCMS (M+H.sup.+): 182.0 .sup.1H NMR (400 MHz, Deuterium Oxide) δ 3.47-3.43 (m, 2H), 3.11 (s, 9H), 2.95 (t, 2H), 2.24-2.18 (m, 2H).2.95 (t, 2H), 2.24-2.18 (m, 2H).

##STR00228##

Compound 141: 4-hydroxy-N,N-dimethyl-N-(prop-2-yn-1-yl)butan-1-aminium Trifluoroacetate

[0751] A mixture of N,N-dimethylprop-2-yn-1-amine (500 mg, 6.01 mmol, 637.76 uL, 1 eq) and 4-bromobutan-1-ol (966.35 mg, 6.32 mmol, 1.05 eq) in EtOH (5 mL) was degassed and purged with N.sub.2 3 times. Then the mixture was stirred at 80° C. for 10 hr under N.sub.2 atmosphere. LCMS showed expected mass was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (column: Xbridge 150*30 mm*10 um; mobile phase: [water (0.1% TFA)-ACN]; B %: 1%-5%, 10 min) to give 4-hydroxy-N,N-dimethyl-N-(prop-2-yn-1-yl)butan-1-aminium trifluoroacetate (150 mg, 529.64 umol, 8.81% yield) as colorless oil. LCMS: (M+) 156.3 1H NMR (400 MHz, Deuterium Oxide) δ 4.1 (d, 2H), 3.6-3.5 (m, 2H), 3.4-3.3 (m, 2H), 3.1-3.0 (m, 1H), 3.0 (s, 6H), 1.8-1.7 (m, 2H), 1.5-1.4 (m, 2H).

##STR00229##

Compound 142: N-(2-carboxyethyl)-N,N-dimethylprop-2-yn-1-aminium Bromide

[0752] A mixture of N,N-dimethylprop-2-yn-1-amine (500 mg, 6.01 mmol, 637.76 uL, 1 eq) 3-bromopropanoic acid (966.08 mg, 6.32 mmol, 652.76 uL, 1.05 eq) in EtOH (5 mL) was degassed and purged with N.sub.2 3 times. And then the mixture was stirred at 80° C. for 10 hr under N.sub.2 atmosphere. LCMS showed desired mass was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (column: Xbridge 150*30 mm*10 um; mobile phase: [water (0.04% NH.sub.3H.sub.2O)-ACN]; B %: 1%-10%, 10 min) to give N-(2-carboxyethyl)-N,N-dimethylprop-2-yn-1-aminium bromide (95 mg, 345.71 umol, 5.75% yield) as colorless oil. LCMS: (M+) 156.1 .sup.1H NMR (400 MHz, Deuterium Oxide) δ 4.2 (d, 2H), 3.8-3.6 (m, 2H), 3.2-3.1 (m, 1H), 3.1 (s, 6H), 2.7-2.6 (m, 2H).

##STR00230##

Compound 143: N,N,N-trimethyl-3-phosphonopropan-1-aminium Trifluoroacetatetrifluoroacetatetrifluoroacetatetrifluoroacetatetrifluoroacetate

[0753] To a solution of 1-bromo-3-diethoxyphosphoryl-propane (500 mg, 1.93 mmol, 370.37 uL, 1 eq) in ACN (5 mL) was added 1-bromo-3-diethoxyphosphoryl-propane (500 mg, 1.93 mmol, 370.37 uL, 1 eq.). The mixture was stirred at 15° C. for 48 h. Then the reaction mixture was concentrated under reduced pressure to give a residue. Then HCl aq. (5 mL, 12 M) was added to the residue at 15° C. and the mixture was stirred at 80° C. for 12 h. LC-MS showed the desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC ([water (0.10% TFA)-ACN]). The title compound (225 mg, 1.22 mmol, 63.36% yield) was obtained as a white solid. LCMS: (M+): 182.2 .sup.1H NMR (400 MHz, Deuterium Oxide) δ 3.36-3.26 (m, 2H), 3.06-3.00 (m, 9H), 2.02-1.92 (m, 2H), 1.78-1.65 (m, 2H).

##STR00231##

Compound 144: N-(3-carboxypropyl)-N,N-dimethylprop-2-yn-1-aminium Iodide

[0754] 4-(dimethylamino)butanoic acid is taken up in THF and treated with drop wise addition of propargyl iodide. The resulting precipitate is collected and dried and recrystallized to afford the title compound.

Compound 147: 3-(hydroxyhydrophosphoryl)-N,N,N-trimethylpropan-1-aminium Chloridechloridechloridechloridechloride

[0755] Step 1:

[0756] To a solution of 1-[ethoxy(ethoxyphosphonoyl)methoxy]ethane (1 g, 5.10 mmol, 1 eq) in THF (40 mL) was added slowly NaH (224.26 mg, 5.61 mmol, 60% purity, 1.1 eq) at 0° C. under N.sub.2. Then the mixture was stirred at 15° C. for 1 h under N.sub.2 atmosphere. Then the mixture was dropwise added to the solution of 1,3-dibromopropane (2.79 g, 13.81 mmol, 1.41 mL, 2.71 eq) in THF (20 mL) at 15° C. Then the mixture was stirred at 15° C. for 9 h under N.sub.2 atmosphere. TLC indicated 1-[ethoxy(ethoxyphosphonoyl)methoxy]ethane was consumed completely and one new spot formed. The reaction mixture was quenched by addition of H.sub.2O (100 mL) at 0° C. and extracted with EtOAc (20 mL*3). The combined organic layers were washed with brine (30 mL). The combined organic layer was dried over Na.sub.2SO.sub.4, filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=1/1). Compound 1-bromo-3-[diethoxymethyl(ethoxy)phosphoryl] propane (400 mg, 1.26 mmol, 24.74% yield) was obtained as yellow oil.

[0757] Step 2:

[0758] A mixture of 1-bromo-3-[diethoxymethyl(ethoxy)phosphoryl]propane (400 mg, 1.26 mmol, 1 eq), trimethylamine (1.49 g, 7.57 mmol, 30% purity, 6 eq., in EtOH) in ACN (5 mL) was degassed and purged with N.sub.2 3 times, and then the mixture was stirred at 15° C. for 96 hr under N.sub.2 atmosphere. TLC indicated 20% of 1-bromo-3-[diethoxymethyl(ethoxy)phos-phoryl]propane (400 mg, 1.26 mmol, 1 eq) remained and one major new spot with larger polarity was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (TFA condition; column: Luna C18 100*30 5u; mobile phase: [water (0.1% TFA)-ACN]; B %: 1%-25%, 12 min). Compound 3-[diethoxymethyl(ethoxy)phosphoryl] propyl-trimethyl-ammonium (150 mg, 366.41 umol, 29.05% yield, CF3CO2-) was obtained as colorless oil.

[0759] Step 3:

[0760] A mixture of 3-[diethoxymethyl(ethoxy)phosphoryl]propyl-trimethyl-ammonium (150 mg, 366.41 umol, 1 eq, CF.sub.3COO—), in conc. HCl (10 mL) was degassed and purged with N.sub.2 3 times, and then the mixture was stirred at 110° C. for 10 hr under N.sub.2 atmosphere. LC-MS showed 100% desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a colorless oil. Compound 3-(hydroxyhydrophosphoryl)-N,N,N-trimethylpropan-1-aminium chloride (120 mg, quantitative yield, Cl—) was obtained as colorless oil. LCMS: (M+): 166.1 .sup.1H NMR (400 MHz, Deuterium Oxide) δ 6.95 (dt, J=546.5, 1.8 Hz, 1H), 3.31-3.22 (m, 2H), 2.99 (s, 9H), 1.98-1.83 (m, 2H), 1.71-1.58 (m, 2H).)

##STR00232##

Compound 148: (1s,4s)-3-hydroxy-1-methylquinuclidin-1-ium

[0761] (S)-(+)-3-quinuclidinol (1 equiv) is stirred with methyl iodide (1.1 equiv) to yield the title compound.

##STR00233##

Compound 149: 4-hydroxy-1-methylquinuclidin-1-ium

[0762] 1-azabicyclo[2.2.2]octan-4-ol (1 equiv) is stirred with methyl iodide (1.1 equiv) to yield the title compound.

##STR00234##

Compound 151: (2R)-1-(fluoromethyl)-2-(hydroxymethyl)-1-methylpyrrolidin-1-ium Iodide

[0763] A mixture of [(2R)-1-methylpyrrolidin-2-yl]methanol (251 mg, 2.18 mmol, 1 eq), fluoro(iodo)methane (348.54 mg, 2.18 mmol, 1 eq) in THF (5 mL) was degassed and purged with N.sub.2 3 times, and then the mixture was stirred at 15° C. for 2 hr under N.sub.2 atmosphere. TLC indicated [(2R)-1-methylpyrrolidin-2-yl]methanol was consumed completely and one new spot formed. The reaction mixture was filtered and the filter cake was concentrated under reduced pressure to give a residue. Then the crude product was purified by p-HPLC (column: Phenomenex Luna C18 100*30 mm*5 um; mobile phase: [water (0.1% TFA)-ACN]; B %: 1%-3%, 4 min) to get the desired product (2R)-1-(fluoromethyl)-2-(hydroxymethyl)-1-methylpyrrolidin-1-ium iodide (150 mg, 545.25 umol, 25.02% yield) as a yellow solid. LCMS (M+): 148.1 .sup.1H NMR (400 MHz, Deuterium Oxide) δ 5.47-5.31 (m, 1H), 5.34-5.22 (m, 1H), 3.93-3.60 (m, 4H), 3.54-3.36 (m, 1H), 3.15-2.94 (m, 3H), 2.21-1.71 (m, 4H).

Compound 152: (S)-1-carboxy-N,N,N-trimethylbut-3-yn-1-aminium Iodide

[0764] To a solution of (2S)-2-aminopent-4-ynoic acid (1 g, 8.84 mmol, 1 eq) in MeOH (10 mL) was added NaOH (800.06 mg, 20.00 mmol, 2.26 eq) in EtOH (2 mL) and MeI (3.76 g, 26.52 mmol, 1.65 mL, 3 eq) at 15° C. The mixture was stirred at 15° C. for 12 hr. LCMS showed the desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (column: Phenomenex Luna C18 100*30 mm*5 um; mobile phase: [water (0.2% FA)-ACN]; B %: 1%-1%, 4 min). The mixture was freeze-dried. The residue was re-purified by prep-HPLC (column: Phenomenex Luna C18 100*30 mm*5 um; mobile phase: [water (0.2% FA)-ACN]; B %: 1%-1%, 4 min). The title compound (48 mg, 158.73 umol, I-salt, 1.80% yield) was obtained as a white solid. LCMS: (M+): 156.1 .sup.1H NMR (400 MHz, Deuterium Oxide) δ 3.82 (ddd, J=8.9, 5.1, 0.9 Hz, 1H), 3.16 (s, 9H), 2.96-2.77 (m, 2H), 2.49-2.40 (m, 1H).

##STR00235##

Compound 153: N-(fluoromethyl)-N-(2-hydroxyethyl)-N-methylprop-2-yn-1-aminium Iodide

[0765] To a solution of 2-[methyl(prop-2-ynyl)amino]ethanol (400 mg, 3.53 mmol, 1 eq) in THF (5 mL) was added fluoro(iodo)methane (565.33 mg, 3.53 mmol, 1 eq) at 15° C. The mixture was stirred at 15° C. for 12 hr. TLC indicated 2-[methyl(prop-2-ynyl)amino]ethanol (400 mg, 3.53 mmol, 1 eq) was consumed completely. The reaction mixture was diluted with H2O (10 mL) and extracted with ethyl acetate 40 mL. The aqueous phase is freeze-dried. Compound N-(fluoromethyl)-N-(2-hydroxyethyl)-N-methylprop-2-yn-1-aminium iodide (247 mg, 882.41 umol, 24.96% yield) was obtained as a white solid. LCMS (M+): 146.1 .sup.1H NMR (400 MHz, Deuterium Oxide) δ 5.56-5.37 (m, 2H), 4.35 (s, 2H), 3.95 (s, 2H), 3.63-3.56 (m, 2H), 3.19-3.14 (m, 4H).

##STR00236##

Compound 154: 1-(2-hydroxyethyl)-1-(prop-2-yn-1-yl)piperidin-1-ium Bromide

[0766] To a solution of 2-(1-piperidyl)ethanol (300 mg, 2.32 mmol, 308.32 uL, 1 eq) in THF (5 mL) was added 3-bromoprop-1-yne (303.85 mg, 2.55 mmol, 220.18 uL, 1.1 eq). The mixture was stirred at 15° C. for 12 hr. The precipitation was found. The reaction mixture was filtered to give a residue. The residue was washed with ethyl acetate 20 mL. Compound 1-(2-hydroxyethyl)-1-(prop-2-yn-1-yl)piperidin-1-ium bromide (554 mg, 2.18 mmol, 94% yield) was obtained as a white solid. LCMS: (M+): 168.1 .sup.1H NMR (400 MHz, DMSO-d6) δ 5.36 (t, J=4.9 Hz, 1H), 4.53 (d, J=2.5 Hz, 2H), 4.04 (t, J=2.5 Hz, 1H), 3.86 (q, J=5.1 Hz, 2H), 3.59-3.49 (m, 4H), 3.49-3.38 (m, 2H), 1.83 (p, J=5.9 Hz, 4H), 1.65-1.45 (m, 2H).

##STR00237##

Compound 155: 2-carboxy-N,N,N-trimethylprop-2-en-1-aminium Bromide

[0767] To a solution of 2-(bromomethyl)prop-2-enoic acid (200 mg, 1.21 mmol, 1 eq) in THF (3 mL) was added N,N-dimethylmethanamine (1 M, 1.21 mL, 1 eq., in THF) at 15° C. The mixture was stirred at 15° C. for 12 hr. The precipitation was found. The reaction mixture was filtered to give a residue. The residue was washed by ethyl acetate 10 mL. Compound 2-carboxy-N,N,N-trimethylprop-2-en-1-aminium bromide (131 mg, 561.19 umol, 46.29% yield) was obtained as a white solid. LCMS: (M+): 144.1 .sup.1H NMR (400 MHz, Deuterium Oxide) δ 6.77 (s, 1H), 6.23 (s, 1H), 4.09 (s, 2H), 2.96 (s, 9H).

##STR00238##

Compound 156: 2-carboxy-N-(2-carboxyallyl)-N,N-dimethylprop-2-en-1-aminium Bromide

[0768] To a solution of 2-(bromomethyl)prop-2-enoic acid (600 mg, 3.64 mmol, 3 eq) in acetone (5 mL) was added N-methylmethanamine (2 M, 606.12 uL, 1 eq., in THF) at 15° C. The mixture was stirred at 15° C. for 12 hr, and a precipitate formed. The reaction mixture was filtered to give a residue. The residue was washed by acetone 10 mL. Compound 2-carboxy-N-(2-carboxyallyl)-N,N-dimethylprop-2-en-1-aminium bromide (100 mg, 310.22 umol, 25.59% yield) was obtained as a white solid. LCMS: (M+): 214.1 .sup.1H NMR (400 MHz, Deuterium Oxide) δ 6.9 (s, 2H), 6.4 (s, 2H), 4.2 (s, 4H), 2.9 (s, 6H).

##STR00239##

Compound 157: 2-hydroxy-N-(2-hydroxyethyl)-N,N-dimethylethan-1-aminium Iodide

[0769] To a solution of 2-(dimethylamino)ethanol (100 mg, 1.12 mmol, 112.61 uL, 1 eq) in THF (5 mL) was added 2-iodoethanol (578.77 mg, 3.37 mmol, 263.08 uL, 3 eq) at 15° C. The mixture was stirred at 50° C. for 12 hr. The precipitation was found. The reaction mixture was filtered to give a residue. The residue was washed by ethyl acetate 10 mL. Compound 2-hydroxy-N-(2-hydroxyethyl)-N,N-dimethylethan-1-aminium iodide (230 mg, 872.08 umol, 77.73% yield) was obtained as a white solid. LCMS: (M+): 134.2 .sup.1H NMR (400 MHz, Deuterium Oxide) δ 3.95-3.87 (m, 4H), 3.48-3.40 (m, 4H), 3.07 (s, 6H).

##STR00240##

Compound 158: 1,1-dimethylpyrazolidin-1-ium Iodide

[0770] Step 1

[0771] To a solution of NaH (600.78 mg, 15.02 mmol, 60% purity, 2 eq.) in DMF (20 mL) at 0° C. was added tert-butyl N-(benzyloxycarbonylamino)carbamate (2 g, 7.51 mmol, 1 eq) at 0° C. and stirred at 0° C. for 20 min. Then 1,3-dibromopropane (1.52 g, 7.51 mmol, 765.80 uL, 1 eq) was added to the mixture at 0° C. The mixture was stirred at 15° C. for 12 h. TLC indicated reactant was consumed completely. The reaction mixture was quenched by addition of H.sub.2O 100 mL at 0° C., and then diluted with H.sub.2O 100 mL and extracted with EtOAc (200 mL*2). The combined organic layers were washed with saturate brine 300 mL (150 mL*2), dried over, filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=20/1 to 4/1). Compound 02-benzyl 01-tert-butyl pyrazolidine-1,2-dicarboxylate (1.5 g, 4.90 mmol, 65.19% yield) was obtained as a colorless oil.

[0772] Step 2

[0773] To a solution of 02-benzyl 01-tert-butyl pyrazolidine-1,2-dicarboxylate (800 mg, 2.61 mmol, 1 eq) in THF (15 mL) was added Pd/C (0.5 g, 2.61 mmol, 10% purity). The suspension was degassed and purged with H.sub.2 3 times. The mixture was stirred under H.sub.2 (15 Psi.) at 15° C. for 3 h. TLC indicated reactant was consumed completely. The reaction mixture was filtered and the filtrate was concentrated under reduced pressure to give a residue. Compound tert-butyl pyrazolidine-1-carboxylate (400 mg, 2.32 mmol, 88.94% yield) was obtained as a colorless oil and used into the next step without further purification.

[0774] Step 3

[0775] To a solution of tert-butyl pyrazolidine-1-carboxylate (200 mg, 1.16 mmol, 1 eq) in THF (10 mL) was added MeI (494.49 mg, 3.48 mmol, 216.88 uL, 3 eq). The mixture was stirred at 15° C. for 12 h. LC-MS showed the desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. Compound tert-butyl 2,2-dimethylpyrazolidin-2-ium-1-carboxylate (150 mg, crude, I—) was obtained as a yellow solid was used into the next step without further purification. LCMS: (M+): 201.2.

[0776] Step 4

[0777] A solution of tert-butyl 2,2-dimethylpyrazolidin-2-ium-1-carboxylate (150 mg, 745.21 umol, 1 eq) in HCl/dioxane (4 M, 10 mL, 53.68 eq) was stirred at 15° C. for 1 h. MS showed the desired mass was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC ([water (0.1% TFA)-ACN]). Compound 1,1-dimethylpyrazolidin-1-ium iodide (10 mg, 43.85 umol, 5.88% yield, I—) was obtained as a colorless oil. MS: (M+): 101.1. .sup.1H NMR (400 MHz, Chloroform-d) δ 3.77 (t, J=7.7 Hz, 2H), 3.42 (t, J=7.5 Hz, 2H), 3.36 (s, 6H), 2.43 (p, J=7.6 Hz, 2H).

##STR00241##

Compound 159: 1-(carboxymethyl)-1-(prop-2-yn-1-yl)pyrrolidin-1-ium Bromide

[0778] Step 1:

[0779] A mixture of pyrrolidine (5 g, 70.30 mmol, 5.87 mL, 1 eq), benzyl 2-bromoacetate (14.49 g, 63.27 mmol, 9.93 mL, 0.9 eq), Na.sub.2CO.sub.3 (7.45 g, 70.30 mmol, 1 eq) in THF (100 mL) was degassed and purged with N.sub.2 3 times, and then the mixture was stirred at 15° C. for 10 hr under N.sub.2 atmosphere. TLC indicated reactant was consumed completely and one new spot formed. The reaction was clean according to TLC. The reaction mixture was filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=10/1 to 1:1). Compound benzyl 2-pyrrolidin-1-ylacetate (8 g, 36.48 mmol, 51.89% yield) was obtained as yellow oil.

[0780] Step 2:

[0781] A mixture of benzyl 2-pyrrolidin-1-ylacetate (5 g, 22.80 mmol, 1 eq), Pd/C (2 g, 22.80 mmol, 10% purity) in EtOH (100 mL) was degassed and purged with H.sub.2 for 3 times, and then the mixture was stirred at 15° C. for 10 hr under H.sub.2 atmosphere at 15 psi. TLC indicated benzyl 2-pyrrolidin-1-ylacetate was consumed completely and one new spot formed. The reaction mixture was filtered and the filtrate was concentrated to give the crude product. Compound 2-pyrrolidin-1-ylacetic acid (2 g, 15.49 mmol, 67.91% yield) was obtained as a white solid. The crude was used directly in next step.

[0782] Step 3:

[0783] A mixture of 2-pyrrolidin-1-ylacetic acid (300 mg, 2.32 mmol, 1 eq), 3-bromoprop-1-yne (303.95 mg, 2.56 mmol, 220.25 uL, 1.1 eq) in DMF (5 mL) was stirred at 15° C. for 10 hrs under N.sub.2. LCMS indicated desired mass was detected. The reaction mixture was concentrated under reduced pressure to give the crude product. The residue was purified by prep-HPLC (column: Waters Xbridge BEH C18 100*30 mm*10 um; mobile phase: [water (0.10% TFA)-ACN]; B %: 1%). Compound 1-(carboxymethyl)-1-(prop-2-yn-1-yl)pyrrolidin-1-ium bromide (48 mg, 193.03 umol, 8.31% yield) was obtained as a white solid. LCMS: (M+): 168.1 .sup.1H NMR (400 MHz, Deuterium Oxide) δ 4.40 (d, J=2.6 Hz, 2H), 4.11 (s, 2H), 3.77-3.69 (m, 2H), 3.61-3.54 (m, 2H), 3.03 (d, J=2.6 Hz, 1H), 2.12-2.06 (m, 4H).

##STR00242##

Compound 160: 1-(carboxymethyl)-1-(prop-2-yn-1-yl)piperidin-1-ium bromide

[0784] Step 1:

[0785] A mixture of piperidine (5 g, 58.72 mmol, 5.80 mL, 1 eq), benzyl 2-bromoacetate (13.45 g, 58.72 mmol, 9.21 mL, 1 eq), Na2CO3 (6.22 g, 58.72 mmol, 1 eq), in THF (100 mL) was degassed and purged with N.sub.2 3 times, and then the mixture was stirred at 15° C. for 10 hr under N.sub.2 atmosphere. LC-MS showed 15.64% of desired compound was detected. The reaction mixture was filtered. Then the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=1/1). Compound benzyl 2-(1-piperidyl)acetate (10 g, 42.86 mmol, 72.99% yield) was obtained as a yellow oil. LCMS: (M+H.sup.+): 234.2

[0786] Step 2:

[0787] A mixture of benzyl 2-(1-piperidyl)acetate (10 g, 42.86 mmol, 1 eq), Pd/C (2 g, 10% purity), in EtOH (100 mL) was degassed and purged with H2 for 3 times, and then the mixture was stirred at 15° C. for 10 hr under H2 atmosphere at 15 psi. TLC indicated benzyl 2-(1-piperidyl)acetate (10 g, 42.86 mmol, 1 eq) was consumed completely and one new spot formed. LC-MS showed desired compound was detected. The reaction mixture was filtered and the filtrate was concentrated under reduced pressure to give a residue. Compound 2-(1-piperidyl)acetic acid (2.5 g, 17.46 mmol, 40.74% yield) was obtained as a white solid. The crude was used directly for next step. LCMS: (M+H.sup.+): 144.1

[0788] Step 3:

[0789] A mixture of 2-(1-piperidyl)acetic acid (300 mg, 2.10 mmol, 1 eq), 3-bromoprop-1-yne (288.60 mg, 2.30 mmol, 209.13 uL, 1.1 eq), in DMF (20 mL) was stirred at 15° C. for 10 hr under N.sub.2 atmosphere. LC-MS showed desired mass was detected. The reaction mixture was concentrated under reduced pressure to give the crude product. The residue was purified by prep-HPLC (column: Waters Xbridge BEH C18 100*30 mm*10 um; mobile phase: [water (0.1% TFA)-ACN]; B %:1%-10%, 10 min). Compound 2-(1-prop-2-ynylpiperidin-1-ium-1-yl)acetic acid (18 mg, 68.52 umol, 3.27% yield, 99.79% purity, Br—)—) was obtained as white solid. Compound 1-(carboxymethyl)-1-(prop-2-yn-1-yl)piperidin-1-ium bromide (19 mg, 64.35 umol, 3.07% yield) was obtained as white solid. LCMS: (M+): 182 .sup.1H NMR (400 MHz, Deuterium Oxide) δ 4.49 (s, 2H), 4.04-4.00 (m, 2H), 3.63-3.55 (m, 2H), 3.51-3.43 (m, 2H), 3.06 (s, 1H), 1.81-1.74 (m, 4H), 1.60-1.50 (m, 2H).

##STR00243##

Compound 161: (2R)-1-(fluoromethyl)-2-(hydroxymethyl)-1-methylpiperidin-1-ium Iodide

[0790] To a solution of [(2R)-1-methyl-2-piperidyl]methanol (80 mg, 619.20 umol, 1 eq) in THF (5 mL) was added fluoro(iodo)methane (148.54 mg, 928.79 umol, 1.5 eq). The mixture was stirred at 15° C. for 12 hr. LCMS showed the desired compound was detected. The precipitation was found. The reaction mixture was filtered to give a residue. The residue was washed by acetone 20 mL. The residue was purified by prep-HPLC (column: Waters Xbridge BEH C18 100*30 mm*10 um; mobile phase: [water (0.1% TFA)-ACN]; B %: 1%-10%, 10 min). Compound (2R)-1-(fluoromethyl)-2-(hydroxymethyl)-1-methylpiperidin-1-ium iodide (40 mg, 138.14 umol, 22.31% yield) was obtained as a yellow oil. LCMS: (M+): 162.2 .sup.1H NMR (400 MHz, Deuterium Oxide) δ 5.56 (d, J=6.1 Hz, 0.5H), 5.51-5.38 (m, 1H), 5.30 (d, J=6.0 Hz, 0.5H), 3.84-3.70 (m, 2H), 3.70-3.58 (m, 1H), 3.49-3.38 (m, 2H), 3.19-2.96 (m, 3H), 1.81-1.73 (m, 5H), 1.57-1.35 (m, 1H).

##STR00244##

Compound 162: (R)—N-(2-carboxypropyl)-N,N-dimethylprop-2-yn-1-aminium Chloride

[0791] Step 1:

[0792] To a solution of methyl (2S)-3-bromo-2-methyl-propanoate (300 mg, 1.66 mmol, 1.3 eq) in ACN (5 mL) was added N,N-dimethylprop-2-yn-1-amine (105.97 mg, 1.27 mmol, 135.17 uL, 1 eq) at 15° C. The mixture was stirred at 80° C. for 12 hr. LCMS showed the desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (column: Waters Xbridge BEH C18 100*30 mm*10 um; mobile phase: [water (0.1% TFA)-ACN]; B %: 1%-10%, 10 min). Compound [(2R)-3-methoxy-2-methyl-3-oxo-propyl]-dimethyl-prop-2-ynyl-ammonium (15 mg, 48.50 umol, 3.80% yield, CF.sub.3COO—) was obtained as a colorless oil.

[0793] Step 2:

[0794] To a solution of [(2R)-3-methoxy-2-methyl-3-oxo-propyl]-dimethyl-prop-2-ynyl-ammonium (10 mg, 33.64 umol, 1 eq, CF.sub.3COO—) in dioxane (2 mL) and aq. HCl (8 mL, 6 M) was stirred at 50° C. for 4 hr. LCMS showed the desired compound was detected and 50% of [(2R)-3-methoxy-2-methyl-3-oxo-propyl]-dimethyl-prop-2-ynyl-ammonium (10 mg, 33.64 umol, 1 eq, CF.sub.3COO—) remained. The mixture was stirred at 50° C. for 12 hr. LCMS showed the desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (column: Waters Xbridge BEH C18 100*30 mm*10 um; mobile phase: [water (0.10% TFA)-ACN]; B %: 1%-10%, 10 min). Compound (2R)-3-(N,N-dimethyl-prop-2-ynyl-azanyl)-2-methyl-propanoic acid (Cl-salt) was obtained as a yellow oil. LCMS: (M+): 170.2 .sup.1H NMR (400 MHz, Deuterium Oxide) δ 4.20 (d, J=2.6 Hz, 2H), 3.91 (dd, J=13.8, 8.9 Hz, 1H), 3.36 (dd, J=13.9, 2.2 Hz, 1H), 3.20 (t, J=2.6 Hz, 1H), 3.10 (s, 6H), 3.03 (ddd, J=9.3, 7.2, 2.1 Hz, 1H), 1.25 (d, J=7.3 Hz, 3H).

##STR00245##

Compound 164: 1-(fluoromethyl)-1-(prop-2-yn-1-yl)piperidin-1-ium Iodide

[0795] To a solution of 1-prop-2-ynylpiperidine (100 mg, 811.72 umol, 1 eq) in THE (5 mL) was added fluoro(iodo)methane (389.45 mg, 2.44 mmol, 3 eq) at 15° C. The mixture was stirred at 15° C. for 12 hr. The precipitation was found. The reaction mixture was diluted with H2O6060 mL and extracted with ethyl acetate 60 mL. The aqueous phase is freeze-dried to give the residue. The residue was purified by prep-HPLC (column: Waters Xbridge BEH C18 100*30 mm*10 um; mobile phase: [water (0.1% TFA)-ACN]; B %: 1%-10%, 10 min). Compound 1-(fluoromethyl)-1-(prop-2-yn-1-yl)piperidin-1-ium iodide (60 mg, 211.92 umol, 41.10% yield) was obtained as a colorless oil. LCMS (M+): 156.2 .sup.1H NMR (400 MHz, Deuterium Oxide) δ 5.51 (d, J=44.9 Hz, 2H), 4.39 (d, J=1.8 Hz, 2H), 3.61-3.33 (m, 5H), 1.89 (p, J=6.3 Hz, 4H), 1.77-1.58 (m, 2H).

##STR00246##

Compound 165: N,N-dimethyl-N-(prop-2-yn-1-yl)prop-2-yn-1-aminium Bromide

[0796] To a solution of N,N-dimethylprop-2-yn-1-amine (100 mg, 1.20 mmol, 127.55 uL, 1 eq) in THF (5 mL) was added 3-bromoprop-1-yne (143.10 mg, 1.20 mmol, 103.69 uL, 1 eq) at 15° C. The mixture was stirred at 15° C. for 12 hr. The precipitation was found. The reaction mixture was filtered to give a residue. The residue was purified by prep-HPLC (column: Waters Xbridge BEH C18 100*30 mm*10 um; mobile phase: [water (0.1% TFA)-ACN]; B %: 1%-3%, 10 min). Compound N,N-dimethyl-N-(prop-2-yn-1-yl)prop-2-yn-1-aminium bromide (88 mg, 435.45 umol, 56.77% yield) was obtained as a white solid. LCMS: (M+): 122.1 .sup.1H NMR (400 MHz, DMSO-d6) δ 4.4 (s, 4H), 4.1 (s, 2H), 3.1 (s, 6H).

##STR00247##

Compound 166: (S)—N-(2-carboxypropyl)-N,N-dimethylprop-2-yn-1-aminium Chloride

[0797] Step 1:

[0798] To a solution of methyl (2R)-3-bromo-2-methyl-propanoate (500 mg, 2.76 mmol, 1 eq) in ACN (10 mL) was added N,N-dimethylprop-2-yn-1-amine (160.73 mg, 1.93 mmol, 205.01 uL, 0.7 eq) at 15° C. The mixture was stirred at 80° C. for 12 hr. TLC indicated most of N,N-dimethylprop-2-yn-1-amine remained. The mixture was stirred at 80° C. for 24 hr. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (column: Waters Xbridge BEH C18 100*30 mm*10 um; mobile phase: [water (0.1% TFA)-ACN]; B %: 1%-20%, 10 min). Compound methyl (2S)-3-(N,N-dimethyl-prop-2-ynyl-azanyl)-2-methyl-propanoate (10 mg, 37.86 umol, Br-salt, 3.33% yield) was obtained as a colorless oil.

[0799] Step 2:

[0800] A solution of methyl (2S)-3-(N,N-dimethyl-prop-2-ynyl-azanyl)-2-methyl-propanoate (10 mg, 37.86 umol, 1 eq) in dioxane (2 mL) and HCl (8 mL) (6 M, H2O) was stirred at 50° C. for 16 hr. LCMS showed the desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (column: Waters XbridgeBEH C18 100*30 mm*10 um; mobile phase: [water (0.1% TFA)-ACN]; B %: 1%-10%, 10 min). The residue was further purified by prep-HPLC (column: Waters Xbridge BEH C18 100*30 mm*10 um; mobile phase: [water (0.1% TFA)-ACN]; B %: 1%-10%, 10 min) to give the title compound (6 mg, 29.17 umol, Cl-salt, 60.00% yield) as a yellow oil. LCMS: (M+): 170.2 .sup.1H NMR (400 MHz, Deuterium Oxide) δ 4.15 (d, J=2.6 Hz, 2H), 3.85 (dd, J=13.8, 8.9 Hz, 1H), 3.30 (dd, J=13.8, 2.2 Hz, 1H), 3.14 (t, J=2.5 Hz, 1H), 3.04 (s, 6H), 3.01-2.90 (m, 1H), 1.18 (d, J=7.2 Hz, 3H).

##STR00248##

Compound 167: (3-hydroxy-2-methylidenepropyl)trimethylazanium trifluoroacetate

[0801] Step 1:

[0802] To a solution of 2-methylenepropane-1,3-diol (1 g, 11.35 mmol, 925.93 uL, 1 eq) in DCM (10 mL) was added SOCl2 (2.03 g, 17.03 mmol, 1.24 mL, 1.5 eq) at −78° C. and the mixture was stirred at 15° C. for 1.5 h. TLC indicated Reactant was consumed completely. The reaction mixture was concentrated under reduced pressure to give a residue. The crude product 5-methylene-1,3,2-dioxathiane 2-oxide (1.5 g, crude) as a brown liquid was used into the next step without further purification.

[0803] Step 2:

[0804] To a solution of 5-methylene-1,3,2-dioxathiane 2-oxide (600 mg, 4.47 mmol, 1 eq) in THF (10 mL) was added N,N-dimethylmethanamine (1 M, 4.92 mL, 1.1 eq. in THF) at 15° C. and the mixture was stirred at 70° C. for 12 h. MS showed the desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC ([water (0.10% TFA)-ACN]) to give the title compound (21 mg, 85.82 umol, 1.92% yield, 99.4% purity, CF3CO2-) was obtained as a colorless oil. MS: (M+) 130.1 .sup.1H NMR (400 MHz, Methanol-d4) δ 5.84 (s, 1H), 5.62 (s, 1H), 4.23 (s, 2H), 4.04 (s, 2H), 3.17 (s, 9H).

##STR00249##

Compound 168: (cyanomethyl)dimethyl(prop-2-yn-1-yl)azanium Bromide

[0805] A mixture of N,N-dimethylprop-2-yn-1-amine (500 mg, 6.01 mmol, 637.76 uL, 1 eq), 2-bromoacetonitrile (721.43 mg, 6.01 mmol, 400.80 uL, 1 eq) in THF (5 mL) degassed and purged with N.sub.2 3 times, and then the mixture was stirred at 15° C. for 10 hr under N.sub.2 atmosphere. LC-MS showed 80.4% desired compound was detected. The reaction mixture was filtered and the filter cake was concentrated under reduced pressure to give a residue. Then the residue was washed with THF (4 mL) to give the title compound (1.0 g, 4.15 mmol, 69.07% yield, 84.37% purity, Br—) as a white solid. LCMS: (M+) 123.1 1H NMR (400 MHz, DMSO-d.sub.6) δ 4.98 (s, 2H), 4.61 (d, J=2.6 Hz, 2H), 4.23 (t, J=2.5 Hz, 1H), 3.28 (s, 6H).

##STR00250##

Compound 169: (cyanomethyl)trimethylazanium Iodide

[0806] A mixture of N,N-dimethylmethanamine (1 M, 8.46 mL, 1 eq), 2-bromoacetonitrile (1.01 g, 8.46 mmol, 563.68 uL, 1 eq) in THF (5 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 15° C. for 10 hr under N2 atmosphere. TLC indicated N,N-dimethylmethanamine was consumed completely and one new spot larger polarity was formed. The reaction mixture was filtered and the filter cake was concentrated under reduced pressure to give a residue. Then the residue was washed with THF (4 mL) to give a residue as compound cyanomethyl(trimethyl)azanium (1.38 g, 7.71 mmol, 91.11% yield, 100% purity, I—) was obtained as a white solid. LCMS: (M+) 99.1 1H NMR (400 MHz, DMSO-d6) δ 4.98 (s, 2H), 3.26 (s, 9H).

##STR00251##

Compound 170: N-(3-methoxy-2-methylidene-3-oxopropyl)-N,N-dimethylcyclopropanaminium Iodide

[0807] Step 1:

[0808] To a solution of N-methylcyclopropanamine (300 mg, 4.22 mmol, 1 eq) in THF (10 mL) was added methyl 2-(bromomethyl)prop-2-enoate (755.43 mg, 4.22 mmol, 1 eq) and K2CO3 (874.84 mg, 6.33 mmol, 1.5 eq) at 25° C. The mixture was stirred at 70° C. for 12 hr. LCMS showed the desired compound was detected. The reaction mixture was filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=10:1 to 1:1). Compound methyl 2-[[cyclopropyl(methyl)amino]methyl]prop-2-enoate (400 mg, 2.36 mmol, 56.01% yield) was obtained as a colorless liquid.

[0809] Step 2:

[0810] To a solution of methyl 2-[[cyclopropyl(methyl)amino]methyl]prop-2-enoate (100 mg, 590.95 umol, 1 eq) in THF (10 mL) was added MeI (503.27 mg, 3.55 mmol, 220.73 uL, 6 eq) at 25° C. The mixture was stirred at 25° C. for 12 hr. The precipitation was found. The reaction mixture was filtered to give a residue. The title compound (106 mg, 340.66 umol, I-salt, 57.65% yield, 100% purity) was obtained as a white solid. LCMS: (M+): 184.1 1H NMR (400 MHz, Deuterium Oxide) δ 6.79 (s, 1H), 6.28 (s, 1H), 4.24 (s, 2H), 3.71 (s, 3H), 3.05-2.98 (m, 1H), 2.81 (s, 6H), 1.07-0.99 (m, 2H), 0.78-0.68 (m, 2H).

##STR00252##

Compound 171: (2-carboxy-2-methylideneethyl)(fluoromethyl)dimethylazanium Iodide

[0811] To a solution of 2-[(dimethylamino)methyl]prop-2-enoic acid (30 mg, 232.28 umol, 1 eq) in ACN (5 mL) was added fluoro added fluoro(iodo)methane (185.74 mg, 1.16 mmol, 5 eq) at 15° C. The mixture was stirred at 15° C. for 24 hr. LCMS showed the LCMS showed the desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (column: Waters Xbridge BEH C18 100*30 mm*10 um; mobile phase: [water (0.1% TFA)-ACN]; B %: 1%-10%, 10 min). The title compound (3 mg, 8.05 umol, I-salt, 3.47% yield, 77.59% purity) was obtained as a colorless oil. LCMS: (M+): 162.1 1H NMR (400 MHz, Deuterium Oxide) δ 6.76 (s, 1H), 6.31 (s, 1H), 5.78 (s, 1H), 5.65 (s, 1H), 3.92 (s, 2H), 2.75 (s, 6H).

##STR00253##

Compound 172: N-(fluoromethyl)-N-(2-hydroxyethyl)-N-methylcyclopropanaminium Iodide

[0812] Step 1:

[0813] To a solution of N-methylcyclopropanamine (1 g, 14.06 mmol, 1.68 eq) and 2-bromoethoxy-tert-butyl-dimethyl-silane (2.00 g, 8.37 mmol, 1 eq) in acetonitrile (20 mL) was added K2CO3 (1.16 g, 8.37 mmol, 1 eq) at 25° C. The mixture was stirred at 50° C. for 12 hr. TLC indicated new spots formed. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=10:1 to 1:1). Compound N-[2-[tert-butyl(dimethyl)silyl]oxyethyl]-N-methyl-cyclopropanamine (3.2 g, 13.95 mmol, 83.32% yield) was obtained as a colorless oil.

[0814] Step 2:

[0815] To a solution of N-[2-[tert-butyl(dimethyl)silyl]oxyethyl]-N-methyl-cyclopropanamine (200 mg, 871.71 umol, 1 eq) in THF (10 mL) was added fluoro(iodo)methane (418.24 mg, 2.62 mmol, 3 eq) at 25° C. The mixture was stirred at 25° C. for 12 hr. LCMS showed 50% of N-[2-[tert-butyl(dimethyl)silyl]oxyethyl]-N-methyl-cyclopropanamine remained. Fluoro(iodo)methane (418.24 mg, 2.62 mmol, 3 eq) was added. The mixture was stirred at 25° C. for 12 hr. The reaction mixture was concentrated under reduced pressure to give a residue. Compound tert-butyl-[2-[N-cyclopropyl-(fluoromethyl)-methyl-azanyl]ethoxy]-dimethyl-silane (200 mg, 513.66 umol, 58.93% yield) was obtained as a white solid.

[0816] Step 3:

[0817] To a solution of tert-butyl-[2-[N-cyclopropyl-(fluoromethyl)-methyl-azanyl]ethoxy]-dimethyl-silane (200 mg, 513.66 umol, 1 eq) in H.sub.2O (10 mL) was added KF (298.42 mg, 5.14 mmol, 120.33 uL, 10 eq) at 25° C. The mixture was stirred at 25° C. for 12 hr. LCMS showed the desired compound was detected. The reaction mixture was extracted with ethyl acetate (50 mL). The aqueous phase is freeze-dried. The residue was washed with washed by acetonitrile (100 mL). The acetonitrile layer was filtered and the filtrate was concentrated under reduced pressure to give a residue. The solid was diluted with H.sub.2O (3 mL) and extracted with ethyl acetate (5 mL*4). The aqueous phase was freeze-dried. The title compound (36 mg, 130.86 umol, I-salt, 85.71% yield) was obtained as a white solid. LCMS: (M+): 148.1 1H NMR (400 MHz, Deuterium Oxide) δ 5.39 (q, J=5.8 Hz, 1H), 5.28 (q, J=5.8 Hz, 1H), 4.02-3.97 (m, 2H), 3.58-3.51 (m, 2H), 3.16-3.12 (m, 1H), 2.78 (d, J=2.2 Hz, 3H), 1.18-1.14 (m, 2H), 0.96-0.79 (m, 2H).

##STR00254##

Compound 172A: (cyanomethyl)(2-hydroxyethyl)dimethylazanium Bromide

[0818] A mixture of 2-(dimethylamino)ethanol (500 mg, 5.61 mmol, 563.06 uL, 1 eq) in THF (5 mL) was added slowly 2-bromoacetonitrile (672.84 mg, 5.61 mmol, 373.80 uL, 1 eq) at 15° C., and then the mixture was stirred at 15° C. for 10 hr under N2 atmosphere. TLC indicated 2-(dimethylamino) ethanol was consumed completely and one new spot formed. The reaction was clean according to TLC. The reaction mixture was filtered and the filter cake was concentrated under reduced pressure to give a residue. The crude product was washed with THE (4 mL). The title compound (200 mg, 956.55 umol, 17.05% yield, 100% purity, Br—) was obtained as a white solid. LCMS: (M+) 129.1 1H NMR (400 MHz, DMSO-d6) δ 5.45 (t, J=4.8 Hz, 1H), 4.91 (s, 2H), 3.91-3.82 (m, 2H), 3.63-3.53 (m, 2H), 3.26 (s, 6H).

##STR00255##

Compound 173: benzyl(2-hydroxyethyl)dimethylazanium

[0819] A mixture of 2-(dimethylamino)ethanol (500 mg, 5.61 mmol, 563.06 uL, 1 eq) and bromomethylbenzene (959.40 mg, 5.61 mmol, 666.25 uL, 1 eq) in THF (5 mL) was degassed and purged with N.sub.2 for 3 times. And then the mixture was stirred at 15° C. for 10 hr under N.sub.2 atmosphere. TLC indicated 2-(dimethylamino)ethanol was consumed completely and one new spot larger polarity formed. The reaction mixture was filtered and the filter cake was concentrated under reduced pressure to give a residue. The crude product was triturated with THF (30 mL) at 15° C. for 30 min. Then the reaction mixture was filtered and the filter cake was concentrated under reduced pressure to give a residue. The title compound (200 mg, 699.54 umol, 12.47% yield, 91% purity, Br—) was obtained as a white solid. LCMS: (M+) 180.1 1H NMR (400 MHz, Methanol-d4) δ 7.64-7.48 (m, 5H), 4.63 (s, 2H), 4.11-4.03 (m, 2H), 3.52-3.45 (m, 2H), 3.12 (s, 6H).

##STR00256##

Compound 174: [(2R)-3-methoxy-2-methyl-3-oxopropyl]dimethyl(prop-2-yn-1-yl)azanium

[0820] To a solution of methyl (2S)-3-bromo-2-methyl-propanoate (300 mg, 1.66 mmol, 2 eq) in ACN (10 mL) was added N,N-dimethylprop-2-yn-1-amine (68.88 mg, 828.60 umol, 87.86 uL, 1 eq) at 15° C. The mixture was stirred at 80° C. for 12 hr. LCMS showed most of N,N-dimethylprop-2-yn-1-amine remained. Then the mixture Then the mixture was stirred at 80° C. for 24 hr. LCMS showed the desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (column: Waters Xbridge BEH C18 100*30 mm*10 um; mobile phase: [water (0.1% TFA)-ACN]; B %: 1%-20%, 10 min). The title compound (7 mg, 26.50 umol, Br-salt, 3.20% yield, 100% purity) was obtained as a yellow oil. LCMS: (M+): 184.1 1H NMR (400 MHz, Deuterium Oxide) δ 4.18 (d, J=2.5 Hz, 2H), 3.94 (dd, J=13.9, 8.7 Hz, 1H), 3.69 (s, 3H), 3.37 (dd, J=13.9, 2.2 Hz, 1H), 3.26-3.14 (m, 1H), 3.17-3.08 (m, 1H), 3.07 (s, 6H), 1.23 (d, J=7.2 Hz, 3H).

##STR00257##

Compound 175: [(2S)-3-methoxy-2-methyl-3-oxopropyl]dimethyl(prop-2-yn-1-yl)azanium

[0821] To a solution of methyl (2R)-3-bromo-2-methyl-propanoate (300 mg, 1.66 mmol, 1 eq) in ACN (10 mL) was added N,N-dimethylprop-2-yn-1-amine (68.88 mg, 828.60 umol, 87.86 uL, 0.5 eq) at 15° C. The mixture was stirred at 80° C. for 24 hr. LCMS showed the desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (column: Waters Xbridge BEH C18 100*30 mm*10 um; mobile phase: [water (0.1% TFA)-ACN]; B %: 1%-20%, 10 min). The title compound (5 mg, 18.93 umol, Br-salt, 1.14% yield, 100% purity) was obtained as a yellow oil. LCMS: (M+): 184.1 1H NMR (400 MHz, Deuterium Oxide) δ 4.17 (d, J=2.5 Hz, 2H), 3.93 (ddd, J=13.9, 8.7, 1.3 Hz, 1H), 3.67 (d, J=1.3 Hz, 3H), 3.35 (dd, J=13.9, 2.1 Hz, 1H), 3.18 (t, J=2.5 Hz, 1H), 3.12 (dd, J=8.9, 7.0 Hz, 1H), 3.05 (s, 6H), 1.22 (dd, J=7.2, 1.3 Hz, 3H).

##STR00258##

Compound 176: trimethyl[(3-methyloxiran-2-yl)methyl]azanium

[0822] Step 1:

[0823] To a solution of mCPBA (14.29 g, 66.26 mmol, 1.2 eq) (80% purity) in CHCl3 (100 mL) was added (E)-1-chlorobut-2-ene (5 g, 55.22 mmol, 1 eq) at 0° C. The mixture was stirred at 25° C. for 12 h. TLC indicated the reactant was consumed completely. The reaction mixture was quenched by addition Na.sub.2SO.sub.3 aqueous solution (500 ml), and then the mixture was partitioned and the organic phase was separated, dried over Na.sub.2SO.sub.4, then filtered and the crude solution as 0.55 mol/L of 2-(chloromethyl)-3-methyl-oxirane (crude) in CHCl3 (100 ml) was used in the next step without further purification.

[0824] Step 2:

[0825] A solution of N,N-dimethylmethanamine (1 M, 57.86 mL, 1.23 eq) (THF) and 0.55 mol/L of 2-(chloromethyl)-3-methyl-oxirane (55 mmol, 100 mL, 1 eq) in CHCl.sub.3 (100 ml) was stirred at 25° C. for 12 h. MS showed the desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC ([water (0.10% TFA)-ACN]). The title compound (5 mg, 19.12 umol, 93.00% yield, 93% purity, TFA-) was obtained as colorless oil. LCMS: (M+): 130.2. 1H NMR (400 MHz, Methanol-d4) δ 3.89 (dd, J=13.6, 1.8 Hz, 1H), 3.29-3.21 (m, 10H), 3.15 (dd, J=13.6, 9.1 Hz, 1H), 3.00 (qd, J=5.2, 1.9 Hz, 1H), 1.38 (d, J=5.2 Hz, 3H).

##STR00259##

Compound 177: dimethyl(prop-2-yn-1-yl)[(1H-1,2,3,4-tetrazol-5-yl)methyl]azanium

[0826] A mixture of 5-(chloromethyl)-1H-tetrazole (500 mg, 4.22 mmol, 1 eq) and N,N-dimethylprop-2-yn-1-amine (350.69 mg, 4.22 mmol, 447.31 uL, 1 eq) in THF (5 mL) was degassed and purged with N.sub.2 3 times, and then the mixture was stirred at 15° C. for 10 hr under N2 atmosphere. LCMS showed desired compound was detected. The reaction mixture was filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (column: Waters Xbridge BEH C18 100*30 mm*10 um; mobile phase: [water (0.04% NH3H2O)-ACN]; B %: 1%-3%, 10 min). The residue was further purified by SFC (column: DAICEL CHIRALPAK AD (250 mm*30 mm, 10 um); mobile phase: [0.1% NH3H2O EtOH]; B %: 20%-20%, 20 min). The title compound (6 mg, 29.75 umol, 60.00% yield, 100% purity) was obtained as a white solid. LCMS: (M+): 166.1 1H NMR (400 MHz, DMSO-d6) δ 4.68 (s, 2H), 4.36 (d, J=2.5 Hz, 2H), 4.04 (t, J=2.4 Hz, 1H), 3.00 (s, 6H).

##STR00260##

Compound 178: (cyclopropylmethyl)trimethylazanium Iodide

[0827] To a solution of cyclopropylmethanamine (1 g, 9.30 mmol, 1 eq, HCl) in DCM (10 mL) was added dropwise MeI (26.39 g, 185.91 mmol, 11.57 mL, 20 eq) at 0° C. After addition, the mixture was stirred at 25° C. for 12h. TLC indicated reactant 1 was consumed completely. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was dissolved in MeOH (7 mL), then to the solution was added MTBE (70 mL) and precipitation formed, was filtered and the filter cake was dissolved in MeOH (7 mL), then MTBE (70 mL) was added and precipitation formed, was filtered and the filter cake was concentrated under reduce pressure to give the target. The title compound (676 mg, 2.78 mmol, 29.86% yield, 99% purity, I—) was obtained as a white solid. LCMS: (M+): 114.2 1H NMR (400 MHz, Deuterium Oxide) δ 3.00 (d, J=7.3 Hz, 2H), 2.91 (s, 9H), 1.01-0.86 (m, 1H), 0.62-0.51 (m, 2H), 0.24-0.15 (m, 2H).

##STR00261##

Compound 179: (fluoromethyl)dimethyl(prop-2-yn-1-yl)azanium Iodide

[0828] To a mixture of N,N-dimethylprop-2-yn-1-amine (0.20 g, 2.39 mmol, 1 eq) in THF (5 mL) was added fluoro(iodo)methane (1.15 g, 7.18 mmol, 3 eq). The mixture was stirred at 25° C. for 10 h. The reaction mixture was filtered and the filter cake was dried under reduced pressure to give a crude product. The crude product was washed with EtOAc (10 mL) and then dried. The title compound (0.35 g, 1.44 mmol, 60.19% yield, 100% purity, I—) was obtained as a white solid. LCMS: (M+): 116.2 1H NMR (400 MHz, Chloroform-d) δ 6.29 (d, J=1.7 Hz, 1H), 5.79 (d, J=1.5 Hz, 1H), 3.79 (s, 3H), 3.39 (d, J=2.4 Hz, 2H), 2.35 (s, 3H), 2.25 (t, J=2.3 Hz, 1H).

##STR00262##

Compound 181: dimethyl[(3-methyloxiran-2-yl)methyl](prop-2-yn-1-yl)azanium

[0829] Step 1:

[0830] To a solution of m-CPBA (8.58 g, 39.76 mmol, 80% purity, 1.2 eq) in CHCl3 (100 mL) was added (E)-1-chlorobut-2-ene (3 g, 33.13 mmol, 1 eq) at 0° C. The mixture was stirred at 25° C. for 12 hr. TLC indicated (E)-1-chlorobut-2-ene was consumed completely. The reaction mixture was filtered to give organic layers. The combined organic layers were quenched by saturated sodium sulfite solution (200 mL), and then extracted to give organic layers. Compound 0.33 mol/L of 2-(chloromethyl)-3-methyl-oxirane (crude) in CHCl.sub.3 (100 mL) was obtained as a colorless liquid and used into the next step without further purification.

[0831] Step 2:

[0832] To a solution of 2-(chloromethyl)-3-methyl-oxirane (0.33 M, 100 mL, 1 eq) (CHCl.sub.3) was added N,N-dimethylprop-2-yn-1-amine (2.74 g, 33.00 mmol, 3.50 mL, 1 eq) at 25° C. The mixture was stirred at 25° C. for 12 hr. LCMS indicated desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a liquid. The liquid was purified liquid was purified by prep-HPLC (column: Waters Xbridge BEH C18 100*30 mm*10 um; mobile phase: [water (0.1% TFA)-ACN]; B %: 1%-10%, 10 min). Compound 2-[(N,N-dimethyl-prop-2-ynyl-azanyl)methyl]-3-methyl-oxirane (20 mg, 105.44 umol, 0.32% yield) was obtained as a colorless oil. The oil was further purified by prep-HPLC (column: Waters XbridgeBEH C.sub.18 100*30 mm*10 um; mobile phase: [water (0.1% TFA)-ACN]; B %: 1%-5%, 10 min). The title compound (7 mg, 36.90 umol, Cl-salt, 35.00% yield, 100% purity) was obtained as a colorless oil. LCMS: (M+): 154.2 .sup.1H NMR (400 MHz, Methanol-d4) δ 4.49-4.35 (m, 2H), 3.99 (d, J=12.3 Hz, 1H), 3.33-3.18 (m, 9H), 3.07-2.97 (m, 1H), 1.36 (d, J=5.2 Hz, 3H).

##STR00263##

Compound 182: (2-hydroxyethyl)dimethyl(prop-2-en-1-yl)azanium Bromide

[0833] To a solution of 2-(dimethylamino)ethanol (200 mg, 2.24 mmol, 225.23 uL, 1 eq) in THF (10 mL) was added 3-bromoprop-1-ene (271.44 mg, 2.24 mmol, 1 eq) at 0° C. The mixture was stirred at 25° C. for 12 hr. The precipitation was found. LCMS showed desired compound was detected. The reaction mixture was diluted with H.sub.2O (60 mL) and extracted with ethyl acetate (60 mL). The aqueous phase is freeze-dried. The residue was washed with ethyl acetate (50 mL) at 0° C. Compound (2-hydroxyethyl)dimethyl(prop-2-en-1-yl)azanium (160 mg, 761.50 umol, 33.94% yield, 100% purity) was obtained as a white solid. LCMS: (M+): 130. 1H NMR (400 MHz, Deuterium Oxide) δ 6.00-5.82 (m, 1H), 5.64-5.51 (m, 2H), 3.96-3.83 (m, 4H), 3.37-3.30 (m, 2H), 2.98 (s, 6H).

##STR00264##

Compound 183: (2-aminoethyl)dimethyl(prop-2-yn-1-yl)azanium

[0834] Step 1:

[0835] To a solution of tert-butyl N-(2-bromoethyl)carbamate (200 mg, 892.48 umol, 1 eq) in THF (10 mL) was added N,N-dimethylprop-2-yn-1-amine (74.19 mg, 892.48 umol, 94.63 uL, 1 eq) at 25° C. The mixture was stirred at 25° C. for 5 hr. LCMS showed LCMS showed most of tert-butyl N-(2-bromoethyl)carbamate was remained. The mixture was stirred at 70° C. for 12 hr. LCMS showed desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was diluted with H.sub.2O 60 mL and extracted with ethyl acetate 60 mL. The aqueous phase is freeze-dried. The residue was purified by prep-HPLC (column: Waters Xbridge BEH C18 100*30 mm*10 um; mobile phase: [water (0.1% TFA)-ACN]; B %: 1%-60%, 9 min). Compound tert-butyl N-[2-(N,N,N-dimethyl-prop-2-ynyl-azanyl)ethyl]carbamate (120 mg, 390.59 umol, Br-salt, 80.00% yield, 100% purity) was obtained as a colorless soil. LCMS: (M+): 227.2

[0836] Step 2:

[0837] To a solution of tert-butyl N-[2-(N,N-dimethyl-prop-2-ynyl-azanyl)ethyl]carbamate (50 mg, 162.75 umol, 1 eq) in DCM (3 mL) and TFA (0.3 mL) was stirred at 25° C. for 12 hr. LCMS showed tert-butyl N-[2-(N,N-dimethyl-prop-2-ynyl-azanyl)ethyl]carbamate was consumed completely and the desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The title compound (30 mg, 144.85 umol, Br-salt, 89.00% yield, 100% purity) was obtained as a colorless oil. LCMS: (M+): 127.1 1H NMR (400 MHz, Deuterium Oxide) δ 4.24 (d, J=2.5 Hz, 2H), 3.73-3.64 (m, 2H), 3.47-3.38 (m, 2H), 3.23 (t, J=2.4 Hz, 1H), 3.15 (s, 6H).

##STR00265##

Compound 184: (2-hydroxyethyl)dimethyl[(oxetan-3-yl)methyl]azanium Bromide

[0838] To a solution of 2-(dimethylamino)ethanol (100 mg, 1.12 mmol, 112.61 uL, 1 eq) in ACN (5 mL) was added 3-(bromomethyl)oxetane (169.41 mg, 1.12 mmol, 1 eq) at 25° C. The mixture was stirred at 70° C. for 12 hr. The reaction mixture was diluted with H.sub.2O 60 mL and extracted with ethyl acetate 60 mL. The aqueous phase is freeze-dried. Compound 2-[N,N-dimethyl-(oxetan-3-ylmethyl)-azanyl]ethanol (70 mg, 290.26 umol, 25.87% yield, Br-salt, 99.576% purity) was obtained as a white solid. LCMS: (M+): 160.1 1H NMR (400 MHz, Deuterium Oxide) δ 4.81-4.73 (m, 2H), 4.55-4.47 (m, 2H), 3.94-3.86 (m, 2H), 3.76-3.61 (m, 3H), 3.37-3.30 (m, 2H), 2.95 (s, 6H).

##STR00266##

Compound 185: (2-chloroprop-2-en-1-yl)dimethyl(prop-2-yn-1-yl)azanium Chloride

[0839] To a mixture of N of N,N-dimethylprop-2-yn-1-amine (200 mg, 2.41 mmol, 255.10 uL, 1 eq) in THF (3 mL) was added 2,3-dichloroprop-1-ene (533.95 mg, 4.81 mmol, 441.28 uL, 2 eq) in one portion at 25° C. under N.sub.2. Then the mixture was heated to 66° C. and stirred for 12 hours. LCMS showed one main peak with desired m/z was detected. The reaction mixture was diluted with H.sub.2O (55 mL) and extracted with ethyl acetate (9 mL, 3 mL*3). The combined aqueous layer was lyophilized. Compound 2-chloroallyl-dimethyl-prop-2-ynyl-ammonium (266 mg, 1.36 mmol, 56.67% yield, 100% purity, Cl—) was obtained as yellow solid. LCMS: (M+): 158.1 1H NMR (400 MHz, Deuterium Oxide) δ 5.89 (dd, J=20.2, 2.3 Hz, 2H), 4.27-4.20 (m, 4H), 3.19 (t, J=2.6 Hz, 1H), 3.15 (s, 6H).

##STR00267##

Compound 186: (2-fluoroethyl)(2-hydroxyethyl)dimethylazanium

[0840] To a solution of 2-(dimethylamino)ethanol (100 mg, 1.12 mmol, 112.61 uL, 1 eq) in THF (5 mL) was added 1-fluoro-2-iodo-ethane (195.16 mg, 1.12 mmol, 1 eq) at 25° C. The mixture was stirred at 25° C. for 5 hr. The mixture was stirred at 70° C. for 12 hr. The precipitate was washed with tetrahydrofuran (10 mL). The mixture was diluted with H.sub.2O 60 mL and extracted with ethyl acetate 100 mL. The aqueous phase is freeze-dried. The title compound (83 mg, 315.48 umol, I-salt, 28.12% yield, 100% purity) was obtained as a white solid. LCMS: (M+): 136.1 1H NMR (400 MHz, Deuterium Oxide) δ 4.93-4.86 (m, 1H), 4.82-4.74 (m, 1H), 3.97-3.89 (m, 2H), 3.79-3.73 (m, 1H), 3.72-3.66 (m, 1H), 3.51-3.44 (m, 2H), 3.11 (s, 6H).

##STR00268##

Compound 187: 1-(fluoromethyl)-1-(prop-2-yn-1-yl)pyrrolidin-1-ium

[0841] Step 1:

[0842] To a mixture of pyrrolidine (1 g, 14.06 mmol, 1.17 mL, 1 eq) and 3-bromoprop-1-yne (1.84 g, 15.47 mmol, 1.33 mL, 1.1 eq) in EtOH (10 mL) was added NaHCO.sub.3 (2.95 g, 35.15 mmol, 1.37 mL, 2.5 eq). The mixture was stirred at 25° C. for 5 h. LC-MS showed the desired product was detected. The residue was purified by prep-HPLC (column: Phenomenex Luna C18 100*30 mm*5 um; mobile phase: [water (0.04% HCl)-ACN]; B %: 1%-1%, 12 min). The solution of 1-prop-2-ynylpyrrolidine (after prep-HPLC) was adjusted to pH=8 by NaHCO.sub.3. The mixture was extracted with ethyl acetate (40 mL*2). The combined organic phase dried with anhydrous Na.sub.2SO.sub.4, filtered, and concentrated in vacuum. 1-prop-2-ynylpyrrolidine (in EtOAc (80 mL) was obtained. LCMS: (M+): 110.2

[0843] Step 2:

[0844] To the solution of 1-prop-2-ynylpyrrolidine (60 mL, 1 eq) in EtOAc (60 mL) was added fluoro(iodo)methane (0.8 g, 5.00 mmol, 1 eq) at 25° C. The mixture was stirred at 25° C. for 5 hr. TLC (Dichloromethane:Methanol=10:1, Rf=0.04) indicated reactant was consumed completely. The mixture was concentrated in reduced pressure. To the mixture was added H.sub.2O (10 mL). The aqueous phase was extracted with ethyl acetate (10 mL*2). The aqueous phase was lyophilized to give a yellow solid. 1-(fluoromethyl)-1-prop-2-ynyl-pyrrolidin-1-ium (70 mg, 254.78 umol, 97.942% purity, I—) was obtained as yellow solid. LCMS: (M+): 142.1 .sup.1H NMR (400 MHz, Methanol-d4) δ 5.55 (d, J=45.1 Hz, 2H), 4.50 (d, J=1.6 Hz, 2H), 3.82-3.71 (m, 4H), 3.55 (s, 1H), 2.30-2.23 (m, 4H).

##STR00269##

Compound 188: 1,1-bis(prop-2-yn-1-yl)pyrrolidin-1-ium Bromidebromide

[0845] To pyrrolidine (1 g, 14.06 mmol, 1.17 mL, 1 eq) was added 3-bromoprop-1-yne (836.32 mg, 7.03 mmol, 606.03 uL, 0.5 eq) at −10° C. under N.sub.2. The mixture was stirred at 25° C. for 12 h. LCMS showed the desired compound was detected. The mixture was purified directly without work-up. The residue was purified by prep-HPLC (column: Phenomenex Luna C18 100*30 mm*5 um; mobilephase: [water (0.04% HCl)-ACN]; B %: 1%-1%, 12 min). The title compound (150 mg, 98.62% purity) was obtained as yellow solid. LCMS: (M+): 148.2. .sup.1H NMR (400 MHz, Deuterium Oxide) δ 4.70-4.60 (m, 4H), 3.72 (t, 4H), 3.20-3.10 (m, 2H), 2.25-2.15 (m, 4H).

##STR00270##

Compound 189: N-(cyanomethyl)-2-(methoxycarbonyl)-N,N-dimethylprop-2-en-1-aminium Bromide

[0846] Methyl 2-(bromomethyl)acrylate (0.200 g, 1 Eq, 1.12 mmol) was dissolved in DCM (3 mL), followed by addition of 2-(dimethylamino)acetonitrile (94.0 mg, 109 μL, 1 Eq, 1.12 mmol). The reaction was stirred at 40° C. overnight. The solution was concentrated by rotary evaporation until product started to precipitate out. More DCM was added and material was triturated in DCM. The solids were further washed in DCM and dried in vacuo to yield product N-(cyanomethyl)-2-(methoxycarbonyl)-N,N-dimethylprop-2-en-1-aminiumbromide (0.24 g, 82%) as a white solid. LCMS M+: 183.3 .sup.1H NMR (400 MHz, DMSO-d6) δ 6.86 (s, 1H), 6.55 (s, 1H), 4.90 (s 2H), 4.41 (s, 2H), 3.76 (s, 3H), 3.20 (s, 6H).

##STR00271##

Compound 190: N-((1-(methoxycarbonyl)cyclopropyl)methyl)-N,N-dimethylprop-2-yn-1-aminium Bromide

[0847] Compound methyl 1-(bromomethyl)cyclopropane-1-carboxylate (0.100 g, 1 Eq, 518 μmol) was dissolved in DCM (3 mL), followed by addition of N,N-dimethylprop-2-yn-1-amine (45.2 mg, 58.6 μL, 1.05 Eq, 544 μmol). The reaction was stirred at 40° C. overnight, after which an immiscible viscous oil appeared. The oil was washed with DCM multiple times and dried in vacuo to yield product N-((1-(methoxycarbonyl)cyclopropyl)methyl)-N,N-dimethylprop-2-yn-1-aminium bromide (0.025 g, 17%) as a viscous glassy oil. LCMS M+: 196.3 .sup.1H NMR (400 MHz, DMSO-d6) δ 4.38 (d, J=2.5 Hz, 2H), 4.06 (t, J=2.5 Hz, 1H), 3.77 (s, 2H), 3.63 (s, 3H), 3.09 (s, 6H), 1.44 (q, J=4.5 Hz, 2H), 1.27 (q, J=4.4 Hz, 2H).

##STR00272##

Compound 191: [2-(2-hydroxyethoxy)ethyl]dimethyl(prop-2-yn-1-yl)azanium

[0848] To a solution of 2-(2-chloroethoxy)ethanol (299.68 mg, 2.41 mmol, 253.97 uL, 1 eq) in THF (10 mL) was added NaI (1.08 g, 7.22 mmol, 3 eq) at 25° C. The mixture was stirred at 70° C. for 0.5 hr. N,N-dimethylprop-2-yn-1-amine (200 mg, 2.41 mmol, 255.10 uL, 1 eq) was added and stirred at 70° C. for 12 hr. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (column: Waters Xbridge BEH C18 100*30 mm*10 um; mobile phase: [water (0.10% TFA)-ACN]; B %: 1%-5%, 9 min). The title compound (79 mg, 262.62 umol, I-salt, 10.92% yield, 99.447% purity) was obtained as yellow oil. LCMS: (M+): 172.1 1H NMR (400 MHz, Deuterium Oxide) δ 4.17 (d, J=2.6 Hz, 2H), 3.86-3.79 (m, 2H), 3.61-3.51 (m, 4H), 3.54-3.45 (m, 2H), 3.11-3.06 (m, 7H).

##STR00273##

Compound 192: tert-butyl N-{2-[dimethyl(prop-2-yn-1-yl)azaniumyl]ethyl}carbamate Bromide

[0849] To a solution of tert-butyl N-(2-bromoethyl)carbamate (200 mg, 892.48 umol, 1 eq) in THF (10 mL) was added N,N-dimethylprop-2-yn-1-amine (74.19 mg, 892.48 umol, 94.63 uL, 1 eq) at 25° C. The mixture was stirred at 25° C. for 5 hr. LCMS showed most of the tert-butyl N-(2-bromoethyl)carbamate remained. The mixture was stirred at 70° C. for 12 hr. LCMS showed desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was diluted with H.sub.2O 60 mL and extracted with ethyl acetate 60 mL. The aqueous phase is freeze-dried. The residue was purified by prep-HPLC (column: Waters Xbridge BEH C18 100*30 mm*10 um; mobile phase: [water (0.1% TFA)-ACN]; B %: 1%-60%, 9 min). The title compound (120 mg, 390.59 umol, Br-salt, 80.00% yield, 100% purity) was obtained as a colorless oil. LCMS: (M+): 227.2 1H NMR (400 MHz, Deuterium Oxide) δ 4.22 (d, J=2.5 Hz, 2H), 3.51 (s, 4H), 3.23 (s, 1H), 3.16 (s, 6H), 1.37 (s, 9H).

##STR00274##

Compound 193: (difluoromethyl)(2-hydroxyethyl)dimethylazanium

[0850] To a solution of 2-(dimethylamino)ethanol (500 mg, 5.61 mmol, 563.06 uL, 1 eq) in DCM (10 mL) was added MeOH (197.71 mg, 6.17 mmol, 249.70 uL, 1.1 eq) and 1-[[bromo(difluoro)methyl]-ethoxy-phosphoryl]oxyethane (4.49 g, 16.83 mmol, 3 eq) at 25° C. The mixture was stirred at 25° C. for 12 hr. The precipitation was formed. The reaction mixture was filtered to give a residue. To the residue was added solvent (ethyl acetate:petroleum ether=1:1, 1 mL). The mixture was diluted with H.sub.2O (1 mL). The aqueous phase was freeze-dried. Compound 2-[N-(difluoromethyl)-dimethyl-azanyl]ethanol (39 mg, 177.23 umol, Br-salt, 4.2% yield, 100% purity) was obtained n as a white solid. LCMS: (M+): 140.1 1H NMR (400 MHz, Deuterium Oxide) δ 7.02 (t, J=58.7 Hz, 1H), 3.96 (t, J=4.8 Hz, 2H), 3.64-3.57 (m, 2H), 3.18 (t, J=1.5 Hz, 6H).

##STR00275##

Compound 194: (cyclobutylmethyl)(2-hydroxyethyl)dimethylazanium Bromide

[0851] To a mixture of 2-(dimethylamino)ethanol (200 mg, 2.24 mmol, 225.23 uL, 1 eq) in THF (3 mL) was added bromomethylcyclobutane added bromomethylcyclobutane (668.77 mg, 4.49 mmol, 502.83 uL, 2 eq) in one portion at 25° C. under N.sub.2. Then the mixture was heated to 66° C. and stirred for 12 hours. LCMS showed one main peak with desired m/z was detected. The reaction mixture was diluted with H.sub.2O (5 mL) and extracted with ethyl acetate (9 mL, 3 mL*3). The aqueous layer was lyophilized. The residue was purified by prep-HPLC [water (0.10% TFA)-ACN]. The title compound (76 mg, 319.11 umol, 14.22% yield, 100% purity, Br—) was obtained as colorless oil. LCMS: (M+): 158.2 1H NMR (400 MHz, Deuterium Oxide) δ 3.90-3.83 (m, 2H), 3.33-3.24 (m, 4H), 2.92 (s, 6H), 2.74 (p, J=7.9 Hz, 1H), 2.06-1.95 (m, 2H), 1.93-1.61 (m, 4H).

##STR00276##

Compound 195: (2-hydroxyethyl)dimethyl[(methylsulfanyl)methyl]azanium

[0852] To a mixture of 2-(dimethylamino)ethanol (200 mg, 2.24 mmol, 225.23 uL, 1 eq) in THF (3 mL) was added chloro added chloro(methylsulfanyl)methane (433.40 mg, 4.49 mmol, 376.87 uL, 2 eq) in one portion at 25° C. under N.sub.2. Then the mixture was heated to 66° C. and stirred for 12 hours. LCMS showed one main peak with desired m/z was detected. The reaction mixture was diluted with H.sub.2O (5 mL) and extracted with ethyl acetate (9 acetate 9 mL, 3 mL*3). The aqueous layer was lyophilized was lyophilized. The residue was purified by prep-HPLC [water (0.10% TFA)-ACN]. Compound 2-hydroxyethyl-dimethyl-(methylsulfanylmethyl)azanium (72 mg, 387.69 umol, 17.28% yield, 100% purity, Cl—)—) was obtained as colorless oil. LCMS: (M+): 150.1 1H NMR (400 MHz, Deuterium Oxide) δ 4.47 (s, 2H), 3.91-3.83 (m, 2H), 3.43-3.36 (m, 2H), 2.99 (s, 6H) 2.23 (s, 3H).

##STR00277##

Compound 196: dimethyl[(methylsulfanyl)methyl](prop-2-yn-1-yl)azanium

[0853] To a mixture of N,N-dimethylprop-2-yn-1-amine (200 mg, 2.41 mmol, 255.10 uL, 1 eq) in THF (3 mL) was added chloro(methylsulfanyl)methane (464.71 mg, 4.81 mmol, 404.09 uL, 2 eq) in one portion at 25° C. under N.sub.2. Then the mixture was heated to 66° C. and stirred for 12 hours. LCMS showed one main peak with desired m/z was detected. The reaction mixture was diluted with H.sub.2O (5 mL) and extracted with ethyl acetate (9 mL, 3 mL*3). The aqueous layer was lyophilized. The residue was purified by prep-HPLC [water (0.10% TFA)-ACN]. Compound dimethyl[(methylsulfanyl)methyl](prop-2-yn-1-yl)azanium (93 mg, 507.15 umol, 21.08% yield, 98% purity, TFA) was obtained as colorless oil. LCMS: (M+): 144.1 1H NMR (400 MHz, Deuterium Oxide) δ 4.59 (s, 2H), 4.23 (d, J=2.6 Hz, 2H), 3.17 (t, J=2.6 Hz, 1H), 3.10 (s, 6H), 2.34 (s, 3H).

##STR00278##

Compound 197: dimethyl(2-oxopropyl)(prop-2-yn-1-yl)azanium Bromide

[0854] To a mixture of N,N-dimethylprop-2-yn-1-amine (200 mg, 2.41 mmol, 255.10 uL, 1 eq) in THF (3 mL) was added 1-bromopropan-2-one (659.07 mg, 4.81 mmol, 2 eq) in one portion at 25° C. under N.sub.2. Then the mixture the mixture was heated to 66° C. and stirred for 12 hours. LCMS showed one main peak with desired m/z was detected. The reaction mixture was diluted with H.sub.2O (5 mL) and extracted with ethyl acetate9 mL (3 mL*3). The aqueous layer was lyophilized. The residue was purified by prep-HPLC [water (0.10% TFA)-ACN]. The title compound (155 mg, 704.21 umol, 29.27% yield, 100% purity, Br—)—) was obtained as white solid. LCMS: (M+): 140.1 1H NMR (400 MHz, Deuterium Oxide) δ 4.55 (s, 2H), 4.38 (d, J=2.6 Hz, 2H), 3.24 (s, 6H), 3.20 (t, J=2.6 Hz, 1H), 2.18 (s, 3H).

##STR00279##

Compound 198: (2-chloroethyl)(2-hydroxyethyl)dimethylazanium Iodide

[0855] To a solution of 2-(dimethylamino)ethanol (200 mg, 2.24 mmol, 225.23 uL, 1 eq) in THF (10 mL) was added 1-chloro-2-iodo-ethane (427.24 mg, 2.24 mmol, 1 eq) at 25° C. The mixture was stirred at 25° C. for 12 hr. LCMS showed most of 2-(dimethylamino)ethanol was remained and a little of desired compound was detected. The mixture was stirred at 70° C. for 6 hr. The precipitation was found. The reaction mixture was diluted with H.sub.2O (60 mL) and extracted with ethyl acetate (100 mL). The aqueous phase is freeze-dried. The residue was purified by prep-HPLC (column: Phenomenex Luna C18 100*30 mm*5 um; mobile phase: [water (0.1% TFA)-ACN]; B %: 1%-1%, 12 min). The title compound (39 mg, 135.99 umol, I-salt, 9.50% yield, 97.478% purity) was obtained as a colorless oil. LCMS: (M+): 152.1 1H NMR (400 MHz, Deuterium Oxide) δ 3.95-3.82 (m, 4H), 3.68 (t, J=6.7 Hz, 2H), 3.47-3.40 (m, 2H), 3.08 (s, 6H).

##STR00280##

Compound 199: N-(2-hydroxyethyl)-N,N-dimethylcyclobutanaminium

[0856] Step 1:

[0857] To a mixture of 2 of 2-(methylamino)ethanol (500 mg, 6.66 mmol, 534.76 uL, 1 eq) in DCE (5 mL) was added HOAc (799.53 mg, 13.31 mmol, 761.46 uL, 2 eq) in one portion at 25° C. under N.sub.2. The mixture was stirred at 25° C. for 0.5 h, then cyclobutanone (933.17 mg, 13.31 mmol, 994.85 uL, 2 eq) and NaBH(OAc).sub.3 (2.82 g, 13.31 mmol, 2 eq) was added. Then the mixture was stirred at 25° C. for 11.5 hours. LCMS showed one main peak with expected mass was detected. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. Compound 2-[cyclobutyl(methyl)amino]ethanol (1 g, crude) was obtained as yellow oil. LCMS: (M+): 130.1

[0858] Step 2:

[0859] To a mixture of 2-[cyclobutyl(methyl)amino]ethanol (430 mg, 3.33 mmol, 1 eq) in THF (3 mL) was added MeI (2.36 g, 16.64 mmol, 1.04 mL, 5 eq) in one portion at 25° C. under N.sub.2. The mixture was stirred at 25° C. for 12 hours. LCMS showed one main peak with desired m/z was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC [water (0.10% TFA)-ACN] and by prep-HPLC [water (0.2% FA)-ACN]. Compound cyclobutyl-(2-hydroxyethyl)-dimethyl-ammonium (38 mg, 140.15 umol, 4.21% yield, I—)—) was obtained as colorless oil. LCMS: (M+): 144.1 1H NMR (400 MHz, Deuterium Oxide) δ 3.99 (p, J=8.9 Hz, 1H), 3.90-3.86 (m, 2H), 3.25-3.18 (m, 2H), 2.88 (s, 6H), 2.24 (p, J=10.3 Hz, 2H), 2.13-2.02 (m, 2H), 1.76-1.51 (m, 2H).

##STR00281##

Compound 200: (3R)-3-hydroxy-1,1-bis(prop-2-yn-1-yl)pyrrolidin-1-ium

[0860] To a mixture of (3R)-pyrrolidin-3-ol (0.5 g, 5.74 mmol, 476.19 uL, 1 eq) in THF (10 mL) and EtOH (10 mL) was added 3 added 3-bromoprop-1-yne (1.37 g, 11.48 mmol, 989.61 uL, 2 eq) in one portion at −5° C. under N.sub.2. The mixture was stirred at 25° C. for 5h. LCMS (product: RT=0.142 min) showed the starting material was consumed completely. The mixture was concentrated in reduced pressure at 45° C. The residue was poured into water (20 mL). The aqueous phase was extracted with ethyl acetate (10 mL*3). The aqueous phase was lyophilized to give yellow oil. The crude product was purified by prep-HPLC (column: PhenomenexLuna C18 100*30 mm*5 um; mobile phase: [water (0.1% TFA)-ACN]; B %: 1%-1%, 12 min). The product was added to ACN (10 mL). The mixture was stirred at 25° C. for 30 min. The mixture was filtered. The organic phase was concentrated in vacuum. The title compound (60 mg, 4.4% yield, Br—)—) was obtained as yellow oil. LCMS: (M+): 164.1 .sup.1H NMR (400 MHz, Methanol-d4) δ 4.67 (s, 1H), 4.61 (d, J=2.5 Hz, 2H), 4.55-4.42 (m, 2H), 4.03-3.91 (m, 1H), 3.91-3.81 (m, 1H), 3.83-3.72 (m, 2H), 3.54-3.47 (m, 2H), 2.59-2.45 (m, 1H), 2.25-2.17 (m, 1H).

##STR00282##

Compound 201: 1,1-bis(prop-2-yn-1-yl)pyrazolidin-1-ium Bromide

[0861] Step 1:

[0862] To a solution of tert-butyl pyrazolidine-1-carboxylate (2 g, 11.61 mmol, 1 eq) in THF (20 mL) was added K.sub.2CO.sub.3 (1.77 g, 12.77 mmol, 1.1 eq) and 3-bromoprop-1-yne (1.66 g, 13.94 mmol, 1.20 mL, 1.2 eq). The mixture was stirred at 25° C. for 12 hr. LC-MS showed the desired compound was detected. The reaction mixture was filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=10:1 to 1:1). Compound tert-butyl 2-prop-2-ynylpyrazolidine-1-carboxylate (1.8 g, 8.56 mmol, 73.71% yield) was obtained as a yellow solid. LCMS: (M-56): 155.1

[0863] Step 2:

[0864] To a solution of tert-butyl 2-prop-2-ynylpyrazolidine-1-carboxylate (50 mg, 237.79 umol, 1 eq) in THF (5 mL) was added 3-bromoprop-1-yne (113.15 mg, 951.15 umol, 81.99 uL, 4 eq) at 25° C. The mixture was stirred at 25° C. for 12 hr. LC-MS showed the desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was washed with tetrahydrofuran (20 mL). Compound tert-butyl-2,2-bis(prop-2-ynyl)-1,2-diazolidine-1-carboxylate (50 mg, 151.87 umol, Br-salt, 63.87% yield) was obtained as a white solid. LCMS: (M+): 249.2

[0865] Step 3:

[0866] A solution of tert-butyl 2,2-bis(prop-2-ynyl)-1,2-diazolidine-1-carboxylate (50 mg, 151.87 umol, 1 eq) in TFA (0.5 mL) and DCM (5 mL) was stirred at 25° C. for 12 hr. LCMS showed the desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. Compound 1,1-bis(prop-2-ynyl)-1,2-diazolidine (22 mg, 96.02 umol, Br-salt, 63.23% yield, 100% purity) was obtained as a yellow oil. LCMS: (M+H+): 149.1 1H NMR (400 MHz, Deuterium Oxide) δ 4.50-4.36 (m, 4H), 3.84 (t, J=7.6 Hz, 2H), 3.31 (t, J=7.0 Hz, 2H), 3.10 (t, J=2.5 Hz, 2H), 2.36-2.24 (m, 2H).

##STR00283##

Compound 202: diethyl {2-[dimethyl(prop-2-yn-1-yl)azaniumyl]ethyl}phosphonate Bromide

[0867] A mixture of N,N-dimethylprop-2-yn-1-amine (0.5 g, 6.01 mmol, 637.76 uL, 1 eq) and 1-bromo-2-diethoxyphosphoryl-ethane (1.77 g, 7.22 mmol, 1.2 eq) in THF (10 mL) was degassed and purged with N.sub.2 3 times, and then the mixture was stirred at 25 at 25° C. for 10 hr under N.sub.2 atmosphere. LC-MS showed N,N-dimethylprop-2-yn-1-amine was remained. Desired compound was detected by LCMS. Then crude product was added water added water (20 mL), and the aqueous phase is freeze-dried. The title compound (100 mg, 398.75 umol, 6.63% yield, 99% purity) was obtained as a white oil. LCMS (M+): 248.1. .sup.1H NMR (400 MHz, Methanol-d4) δ 4.40 (d, J=2.5 Hz, 2H), 4.24-4.11 (m, 4H), 3.75-3.64 (m, 2H), 3.62-3.56 (m, 1H), 3.21 (s, 6H), 2.53-2.39 (m, 2H), 1.36 (t, J=7.1 Hz, 6H).

##STR00284##

Compound 203: N-allyl-2-(methoxycarbonyl)-N,N-dimethylprop-2-en-1-aminium 2,2,2-trifluoroacetate

[0868] Methyl 2-(bromomethyl)acrylate (0.200 g, 1 Eq, 1.12 mmol) was dissolved in DCM (3 mL), followed by addition of N,N-dimethylprop-2-en-1-amine (95.1 mg, 132 μL, 1 Eq, 1.12 mmol). The reaction was stirred at 40° C. overnight, then concentrated by rotary evaporation. The reaction mixture was purified by reverse phase column chromatography on an C18AQ column (100% water with 0.1% TFA). Fractions containing product were lyophilized to yield N-allyl-2-(methoxycarbonyl)-N,N-dimethylprop-2-en-1-aminium 2,2,2-trifluoroacetate (150 mg, 45.2%) as a gel-like solid. LCMS M+: 184.3. .sup.1H NMR (400 MHz, Deuterium Oxide) δ 6.90 (s, 1H), 6.34 (s, 1H), 6.09-5.91 (m, 1H), 5.74-5.55 (m, 2H), 4.16 (s, 2H), 3.86 (d, J=7.4 Hz, 2H), 3.76 (s, 3H), 2.91 (s, 6H).

##STR00285##

Compound 204: dimethyl(prop-2-yn-1-yl)propylazanium Iodide

[0869] N,N-dimethylprop-2-yn-1-amine (500 mg, 6.01 mmol, 637.76 uL, 1 eq) and 1-iodopropane (2.04 g, 12.03 mmol, 1.18 mL, 2 eq) in THF (2 mL) was stirred at 25° C. for 10 h. LCMS showed the starting reactant consumed completely. The mixture was filtered and filter cake was washed by EtOAc (10 mL*3). The filter cake was concentrated. The title compound (250 mg, 4% yield, I—) was obtained as a white solid. LCMS: (M+): 126.2 1H NMR (400 MHz, Deuterium Oxide) δ 4.10 (s, 2H), 3.30-3.21 (m, 2H), 3.03 (s, 6H), 1.75-1.60 (m, 2H), 0.85 (t, J=7.3 Hz, 3H).

##STR00286##

Compound 205: (2S)-1-(fluoromethyl)-2-(hydroxymethyl)-1-(prop-2-yn-1-yl)pyrrolidin-1-ium Iodide

[0870] To a mixture of [of [(2S)-1-prop-2-ynylpyrrolidin-2-yl]methanol (100 mg, 718.42 umol, 1 eq) in THF (3 mL) was added fluoro added fluoro(iodo)methane (574.48 mg, 3.59 mmol, 5 eq) in one portion at 25° C. under N.sub.2. The mixture was stirred at 25° C. for 12 at 25° C. for 12 hours. LCMS showed one main peak with desired m/z was detected. The reaction mixture was diluted with H.sub.2O (3 mL). The aqueous layer was washed with ethyl acetate (3 mL*4). Then the aqueous layer was lyophilized. Compound [(2S)-1-(fluoromethyl)-1-prop-2-ynyl-pyrrolidin-1-ium-2-yl]methanol (115 mg, 384.46 umol, 53.51% yield, I—) was obtained as white solid. LCMS: (M+): 172.1 1H NMR (400 MHz, Deuterium Oxide) δ 5.65-5.39 (m, 2H), 4.49-4.40 (m, 1H), 4.40-4.30 (m, 1H), 4.20-3.99 (m, 1H), 3.99-3.91 (m, 1H), 3.90-3.64 (m, 3H), 3.12 (t, J=2.6 Hz, 1H), 2.28-2.18 (m, 1H), 2.18-2.03 (m, 2H), 2.06-1.87 (m, 1H).

##STR00287##

Compound 206: ethyldimethyl(prop-2-yn-1-yl)azanium Iodide

[0871] A mixture of N,N-dimethylprop-2-yn-1-amine (500 mg, 6.01 mmol, 637.76 uL, 1 eq) and iodoethane (1.88 g, 12.03 mmol, 962.11 uL, 2 eq) in THF (2 mL) was stirred at 25° C. for 10 h. TLC showed Reactant was consumed completely. The reaction mixture was concentrated under reduced pressure to remove THF. The residue was diluted with water (30 mL) and extracted with EtOAc (10 mL*3). The water layer was concentrated by lyophilization. The title compound (200 mg, 794.66 umol, 13.21% yield, 95% purity, I—) was obtained as a yellow solid. LCMS: (M+): 112.1 1H NMR (400 MHz, Deuterium Oxide) δ 4.09 (s, 2H), 3.39 (q, J=7.3 Hz, 2H), 3.03-2.99 (m, 6H), 1.25 (t, J=7.4 Hz, 3H).

##STR00288##

Compound 207: dimethyl[(1,3-oxazol-2-yl)methyl](prop-2-yn-1-yl)azanium Chloride

[0872] A mixture of N,N-dimethylprop-2-yn-1-amine (100 mg, 1.20 mmol, 127.55 uL, 1 eq) and 2-(chloromethyl)oxazole (141.38 mg, 1.20 mmol, 1 eq) in THF (2 mL) was stirred at 25° C. for 10 h. LCMS showed the starting reactant was consumed completely. The reaction mixture was concentrated. The residue was dissolved with H.sub.2O (10 mL) and extracted with EtOAc (10 mL*3), Water phase was freeze-dried. The title compound (95 mg, crude, Cl—) was obtained as yellow solid. LCMS: (M+): 165.1 1H NMR (400 MHz, Deuterium Oxide) δ 7.95 (d, J=0.9 Hz, 1H), 7.25 (d, J=0.9 Hz, 1H), 4.74 (s, 2H), 4.22 (d, J=2.5 Hz, 2H), 3.28-3.17 (m, 1H), 3.15 (s, 6H).

##STR00289##

Compound 208: N,N,2-trimethyl-N-(prop-2-yn-1-yl)prop-2-en-1-aminium 2,2,2-trifluoroacetate

[0873] N,N-dimethylprop-2-yn-1-amine (116 mg, 0.150 mL, 1 Eq, 1.39 mmol) and 3-bromo-2-methylprop-1-ene (188 mg, 1 Eq, 1.39 mmol) were added to a vial with DCM (2 mL), and the reaction was stirred at 40 C overnight. The mixture was concentrated and re-dissolved in water before purification by reverse phase column chromatography on a C18AQ column (100% water with 0.1% TFA). Fractions containing product were lyophilized to yield N,N,2-trimethyl-N-(prop-2-yn-1-yl)prop-2-en-1-aminium 2,2,2-trifluoroacetate (0.0290 g, 8.29%) as a clear viscous oil. LCMS M+: 138.2 1H NMR (400 MHz, Deuterium Oxide) δ 5.52 (s, 1H), 5.35 (s, 1H), 4.14 (d, J=2.6 Hz, 2H), 3.91 (s, 2H), 3.20 (t, J=2.6 Hz, 1H), 3.09 (s, 6H), 1.90 (s, 3H).

##STR00290##

Compound 209: 2-fluoro-N,N-dimethyl-N-(prop-2-yn-1-yl)prop-2-en-1-aminium 2,2,2-trifluoroacetate

[0874] N,N-dimethylprop-2-yn-1-amine (116 mg, 0.150 mL, 1 Eq, 1.39 mmol) and 3-bromo-2-fluoroprop-1-ene (194 mg, 1 Eq, 1.39 mmol) were added to a vial with DCM (2 mL), and the reaction was stirred at 40° C. overnight. The mixture was concentrated and re-dissolved in water before purification by reverse phase column chromatography on an C18AQ column (100% water with 0.1% TFA). Fractions containing product were lyophilized to yield 2-fluoro-N,N-dimethyl-N-(prop-2-yn-1-yl)prop-2-en-1-aminium 2,2,2-trifluoroacetate (262 mg, 73.7%) as a clear viscous oil. LCMS M+: 142.2. .sup.1H NMR (400 MHz, Deuterium Oxide) δ 5.31 (dd, J=15.8, 4.0 Hz, 1H), 5.12 (dd, J=47.9, 3.9 Hz, 1H), 4.31-4.17 m 4H 3.24 (t, J=2.6 Hz, 1H), 3.17 (s, 6H).

##STR00291##

Compound 210: {2-[dimethyl(prop-2-yn-1-yl)azaniumyl]ethyl}phosphonic Acid

[0875] A mixture/n of 3-[N-(2-diethoxyphosphorylethyl)-dimethyl-azanyl]prop-1-yne (300 mg, 914.13 umn/ol, 1 eq) in aq. HCl (20 mL, 12 N) was stirred at 100° C. for 15 hr. LC-MS showed the desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (column: Phenomenex Luna C18 100*30 mm*5 um; mobile phase: [water (0.10% TFA)-ACN]; B %: 1%-1%, 12 min). Compound 2-(N,N-dimethyl-prop-2-ynyl-azanyl)ethylphosphonic acid (44 mg, 161.72 umol, Br-salt, 17.69% yield, 100% purity) was obtained as a white solid. LCMS: (M+): 192.1. .sup.1H NMR (400 MHz, Deuterium Oxide) δ 4.12 (d, J=2.6 Hz, 2H), 3.54-3.44 (m, 2H), 3.12 (t, J=2.5 Hz, 1H), 3.04 (s, 6H), 2.05-1.91 (m, 2H).

##STR00292##

Compound 211: N,N-dimethyl-N-(prop-2-yn-1-yl)prop-2-en-1-aminium 2,2,2-trifluoroacetate

[0876] Compound 3-bromoprop-1-yne (200 mg, 0.150 mL, 1 Eq, 1.68 mmol) and N,N-dimethylprop-2-en-1-amine (143 mg, 199 μL, 1 Eq, 1.68 mmol) were added to a vial with DCM (2 mL), and the reaction was stirred at 40 C overnight. The mixture was concentrated and re-dissolved in water before purification by reverse phase column chromatography on a C18AQ column (100% water with 0.1% TFA). Fractions containing product were lyophilized to yield N,N-dimethyl-N-(prop-2-yn-1-yl)prop-2-en-1-aminium 2,2,2-trifluoroacetate (260 mg, 65.1%) as a clear viscous oil. LCMS M+: 124.2. .sup.1H NMR (400 MHz, Deuterium Oxide) δ 6.03-5.88 (m, 1H), 5.75-5.61 (m, 2H), 4.12 (d, J=2.4 Hz, 2H), 3.95 (d, J=7.4 Hz, 2H), 3.21-3.14 (m, 1H), 3.07 (s, 6H).

##STR00293##

Compound 212: (2-hydroxyethyl)(methoxymethyl)dimethylazanium trifluoroacetate

[0877] To a solution of 2-(dimethylamino)ethanol (500 mg, 5.61 mmol, 563.06 uL, 1 eq) in DCM (5 mL) was added MOMCl (677.45 mg, 8.41 mmol, 639.10 uL, 1.5 eq) at 0° C. The mixture was stirred at 25° C. for 5 h. LCMS showed Reactant was consumed completely. The mixture was quenched with water (10 mL) then extracted with EtOAc (10 mL*3). The aqueous phase was concentrated by lyophilization. The residue was purified by prep-HPLC (column: Phenomenex Luna C18 100*30 mm*5 um; mobile phase: [water (0.10% TFA)-ACN]. The title compound (135 mg, 905.39 umol, 16.14% yield, 90% purity, TFA salt) was obtained as yellow oil. LCMS: (M): 134.1 1H NMR (400 MHz, Deuterium Oxide) δ 4.50 (s, 2H), 3.91-3.82 (m, 2H), 3.52 (s, 3H), 3.35-3.28 (m, 2H), 2.94 (s, 6H).

##STR00294##

Compound 213: ethyl(fluoromethyl)(2-hydroxyethyl)(methyl)azanium Iodide

[0878] Step 1:

[0879] To a solution of 2-bromoethanol (1 g, 8.00 mmol, 568.18 uL, 1 eq) in THF (10 mL) was added K.sub.2CO.sub.3 (1.22 g, 8.80 mmol, 1.1 eq) and N-methylethanamine (520.31 mg, 8.80 mmol, 756.27 uL, 1.1 eq). The mixture was stirred at 25° C. for 12 hr. LCMS showed the desired compound was detected. The reaction mixture was filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=0:1 to Ethyl acetate:Methanol=1:1). Compound 2-[ethyl(methyl)amino]ethanol (120 mg, 1.14 mmol, 14.25% yield, 98% purity) was obtained as a yellow oil. LCMS: (M+H+): 104.1

[0880] Step 2:

[0881] To a solution of 2-[ethyl(methyl)amino]ethanol (100 mg, 969.35 umol, 1 eq) in THF (5 mL) was added fluoro(iodo)methane (775.13 mg, 4.85 mmol, 5 eq). The mixture was stirred at 25° C. for 12 hr. LCMS showed the desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was diluted with H.sub.2O (50 mL) and extracted with ethyl acetate (50 mL*3). The aqueous phase is freeze-dried. Compound 2-[N-ethyl-N-(fluoromethyl)-methyl-azanyl]ethanol (60 mg, 221.85 umol, 23.53% yield, I-salt, 97.28% purity) was obtained as a yellow solid. LCMS: (M+): 136.1 1H NMR (400 MHz, Deuterium Oxide) δ 5.46 (s, 1H), 5.35 (s, 1H), 4.01-3.95 (m, 2H), 3.61-3.46 (m, 4H), 3.10 (d, J=2.2 Hz, 3H), 1.37-1.22 (m, 3H).

##STR00295##

Compound 214: (3R)-1-(fluoromethyl)-3-hydroxy-1-(prop-2-yn-1-yl)pyrrolidin-1-ium

[0882] Fluoro(iodo)methane (1.28 g, 7.99 mmol, 5 eq) and (3R)-1-prop-2-ynylpyrrolidin-3-ol (200 mg, 1.60 mmol, 1 eq) in DCM (2 mL) was stirred at 25° C. for 12 h. LCMS showed the starting reactant was consumed completely. The reaction mixture was concentrated. The residue was dissolved by water (30 mL) and extracted with ethyl acetate (10 mL*3). Water phase was concentrated by lyophilization. The residue was purified by prep-HPLC [water (0.1% TFA)-ACN]. Compound (3R)-1-(fluoromethyl)-1-prop-2-ynyl-pyrrolidin-1-ium-3-ol (80 mg, 252.55 umol, 15.81% yield, 90% purity, CF3CO2-) was obtained as yellow oil. LCMS: (M+): 158. .sup.1H NMR (400 MHz, Deuterium Oxide) δ 5.56-5.48 (m, 1H), 5.46-5.37 (m, 1H), 4.30 (d, J=1.2 Hz, 2H), 3.87-3.74 (m, 3H), 3.78-3.61 (m, 3H), 2.45-2.26 (m, 1H), 2.15-2.10 (m, 1H).

Compound 215: [2-(methanesulfonylcarbamoyl)ethyl]dimethyl(prop-2-yn-1-yl)azanium

[0883] Step 1:

[0884] To a solution of 3-bromopropanoyl chloride (5 g, 29.17 mmol, 2.94 mL, 1 eq) in toluene (50 mL) was added methanesulfonamide (1.14 g, 11.98 mmol, 4.11e-1 eq). The mixture was stirred at 110° C. for 5 hr. LCMS showed 3-bromopropanoyl chloride was consumed completely and desired m/z was detected. The reaction mixture was cooled to 0-5° C. with ice, and the resulting solid was filtered. The filter cake was washed with cold toluene. Compound 3-bromo-N-methylsulfonyl-propanamide (2.8 g, crude) was obtained as white solid and used into the next step without further purification. LCMS: (M+H)+: 230.1

[0885] Step 2:

[0886] To a solution of 3-bromo-N-methylsulfonyl-propanamide (500 mg, 2.17 mmol, 1.2 eq) in ACN (5 mL) was added N,N-dimethylprop-2-yn-1-amine (150.55 mg, 1.81 mmol, 192.03 uL, 1 eq). The mixture was stirred at 25 at 25° C. for 5 hr. LCMS showed N,N-dimethylprop-2-yn-1-amine was consumed completely and desired m/z was detected. The reaction mixture was concentrated under reduced pressure. The residue was purified by prep-HPLC [water (0.1% TFA)-ACN]. The title compound (93 mg, 575.82 umol, 31.80% yield, TFA) was obtained as a white solid. Mass: (M+): 233.2 1H NMR (400 MHz, Methanol-d4) δ 4.37 (d, J=2.6 Hz, 2H), 3.80 (t, J=7.3 Hz, 2H), 3.61 (t, J=2.6 Hz, 1H), 3.25 (s, 3H), 3.19 (s, 6H), 2.99-2.90 (m, 2H).

##STR00296##

Compound 216: [(methanesulfonylcarbamoyl)methyl]dimethyl(prop-2-yn-1-yl)azanium

[0887] Step 1:

[0888] To a solution of methanesulfonamide (2 g, 21.03 mmol, 1 eq) in EtOAc (30 mL) was added 2-bromoacetyl bromide (4.24 g, 21.01 mmol, 1.83 mL, 1 eq). The mixture was stirred at 65° C. for 5 hr. LCMS showed methanesulfonamide was consumed completely and desired m/z was detected. The reaction was cooled, and a large amount of white solid precipitated. The mixture was filtered and the filter cake was concentrated under reduced pressure to give a residue. Compound 2-bromo-N-methylsulfonyl-acetamide (1.7 g, crude) was obtained as white solid and used in the next step without further purification. LCMS: (M+H)+: 216.0

[0889] Step 2:

[0890] To a solution of N,N-dimethylprop-2-yn-1-amine (160.32 mg, 1.93 mmol, 204.49 uL, 1 eq) in THF (4 mL) was added 2-bromo-N-methylsulfonyl-acetamide (500 mg, 2.31 mmol, 1.2 eq). The mixture was stirred at 25° C. for 5 hr. LCMS showed N,N-dimethylprop-2-yn-1-amine was consumed completely and one main peak with desired m/z was detected. The reaction mixture was concentrated under reduced pressure and the residue was purified by prep-HPLC [water (0.1% TFA)-ACN].[2-(methanesulfonamido)-2-oxo-ethyl]-dimethyl-prop-2-ynyl-ammonium (68 mg, 227.28 umol, 11.79% yield, 100% purity, TFA salt) was obtained as white solid. LCMS: (M+): 219.1 1H NMR (400 MHz, Methanol-d4) δ 4.61 (d, J=2.6 Hz, 2H), 4.31 (s, 2H), 3.64 (t, J=2.6 Hz, 1H), 3.41 (s, 6H), 3.26 (s, 3H).

##STR00297##

Compound 217: (2-bromoprop-2-en-1-yl)dimethyl(prop-2-yn-1-yl)azanium

[0891] To a solution of N,N-dimethylprop-2-yn-1-amine (200 mg, 2.41 mmol, 255.10 uL, 1 eq) in THF (4 mL) was added 2 added 2,3-dibromoprop-1-ene (528.94 mg, 2.65 mmol, 258.02 uL, 1.1 eq). The mixture was stirred at 25 at 25° C. for 5 hr. LCMS showed N,N-dimethylprop-2-yn-1-amine was consumed completely and one main peak with desired m/z was detected. The mixture was filtered and the filter cake was concentrated under reduced pressure to give a residue. The title compound (90 mg, 318.02 umol, 13.22% yield, 100% purity, Br—) was obtained as a yellow solid. LCMS: (M+): 202.0 1H NMR (400 MHz, Deuterium Oxide) δ 6.38 (d, J=2.5 Hz, 1H), 6.28 (d, J=2.6 Hz, 1H), 4.40 (s, 2H), 4.31 (d, J=2.1 Hz, 2H), 3.27 (t, J=2.5 Hz, 1H), 3.23 (s, 6H).

##STR00298##

Compound 218: 2-hydroxy-4,4-dimethyl-2-(trifluoromethyl)morpholin-4-ium trifluoroacetate

[0892] Step 1:

[0893] To a mixture of 2-(dimethylamino)ethanol (1 g, 11.22 mmol, 1.13 mL, 1 eq) in THF (10 mL) was added DMAP (137.06 mg, 1.12 mmol, 0.1 eq) and Boc2O (2.69 g, 12.34 mmol, 2.84 mL, 1.1 eq) in one portion at 25 at 25° C. under N.sub.2. The mixture was heated to 50° C. and stirred for 12 hours. TLC indicated 2-(dimethylamino) ethanol was consumed and one new spot formed. LCMS showed one main peak with desired m/z was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=50/1 to 0/1). Desired compound tert-butyl 2-(dimethylamino)ethyl carbonate (850 mg, 4.49 mmol, 40.03% yield) was obtained as colorless oil.

[0894] Step 2:

[0895] To a mixture of tert-butyl 2-(dimethylamino)ethyl carbonate (850 mg, 4.49 mmol, 1 eq) in THF (8 mL) was added 3 added 3-bromo-1,1,1-trifluoro-propan-2-one (1.72 g, 8.98 mmol, 932.19 uL, 2 eq) in one portion at 0° C. under N.sub.2. The mixture was stirred at 25° C. for 10 hours. LCMS showed desired compound was detected. The reaction mixture was concentrated under reduced pressure to remove THF. The residue was diluted with H.sub.2O (3 mL). The residue was purified by prep-HPLC [water (0.1% TFA)-ACN]. Compound 2-tert-butoxycarbonyloxyethyl-dimethyl-(3,3,3-trifluoro-2-oxo-propyl)ammonium (50 mg, 166.50 umol, 3.71% yield) was obtained as a white solid.

[0896] Step 3:

[0897] 2-tert-butoxycarbonyloxyethyl-dimethyl-(3,3,3-trifluoro-2-oxo-propyl)ammonium (0.05 g, 166.50 umol, 1 eq) in DCM (3 mL) and TFA (1 mL) was stirred at 25° C. for 3 hours. LCMS showed LCMS showed desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC[water (0.1% TFA)-ACN]. The title compound (0.029 g, 92.30 umol, 55.43% yield, TFA) was obtained as white solid.

[0898] MS: (M+): 200.1 1H NMR (400 MHz, Deuterium Oxide) δ 4.48-4.36 (m, 1H), 4.10 (d, J=14.1 Hz, 1H), 3.69 (d, J=13.4 Hz, 1H), 3.65-3.51 (m, 3H), 3.41 (s, 3H), 3.23 (s, 3H).

##STR00299##

Compound 219: N-(2-hydroxyethyl)-N,N-dimethyloxetan-3-aminium trifluoroacetate

[0899] Step 1:

[0900] To a mixture of 2-(methylamino)ethanol (1 g, 13.31 mmol, 1.07 mL, 1 eq) in DCM in DCM (5 mL) was added TBDPSCl (4.03 g, 14.65 mmol, 3.76 mL, 1.1 eq), TEA (1.48 g, 14.65 mmol, 2.04 mL, 1.1 eq) and DMAP (813.26 mg, 6.66 mmol, 0.5 eq) in one portion at 25° C. under N.sub.2, then heated to 40° C. and stirred for 12 hours. LCMS showed desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=30/1 to 1/1). .sup.1HNMR indicated the desired compound 2-[tert-butyl(diphenyl)silyl]oxy-N-methyl-ethanamine (1.2 g, crude) was obtained as colorless oil.

[0901] Step 2:

[0902] To a mixture of 2-[tert-butyl(diphenyl)silyl]oxy-N-methyl-ethanamine (1 g, 3.19 mmol, 1 eq) in DCE (10 mL) was added AcOH added AcOH (1.92 g, 31.90 mmol, 1.82 mL, 10 eq) in one portion at 25° C. under N.sub.2. The mixture was stirred at 25° C. for 0.5h, then oxetan-3-one (459.72 mg, 6.38 mmol, 2 eq) and NaBH(OAc).sub.3 (1.35 g, 6.38 mmol, 2 eq) were added, and the mixture stirred for 11.5 hours. LC-MS showed the desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=20/1 to 0/1). The crude product N-[2-[tert-butyl(diphenyl)silyl]oxyethyl]-N-methyl-oxetan-3-amine (0.72 g, crude) as white solid.

[0903] Step 3:

[0904] To a mixture of N-[2-[tert-butyl(diphenyl)silyl]oxyethyl]-N-methyl-oxetan-3-amine (0.72 g, 1.95 mmol, 1 eq) in THF (5 mL) was added MeI (1.66 g, 11.69 mmol, 727.70 uL, 6 eq) in one portion at 25° C. under N.sub.2. The mixture was stirred at 70° C. for 12 hours. LCMS showed the desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=20/1 to 1/1). The crude product 2-[tert-butyl(diphenyl)silyl]oxyethyl-dimethyl-(oxetan-3-yl)ammonium iodide (0.67 g, crude, I) was obtained as yellow solid.

[0905] Step 4:

[0906] To a mixture of 2-[tert-butyl(diphenyl)silyl]oxyethyl-dimethyl-(oxetan-3-yl)ammonium (300 mg, 586.50 umol, 1 eq, I) in THF (3 mL) was added pyridine hydrofluoride (174.38 mg, 1.76 mmol, 158.52 uL, 3 eq) in one portion at 5° C. under N.sub.2. The mixture was stirred at 25° C. for 12 hours. LCMS showed desired mass was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC [water (0.10% TFA)-ACN]. The title compound (38 mg, 146.02 umol, 24.90% yield, TFA) as obtained as white gum. LCMS: (M+): 146.1 .sup.1H NMR (400 MHz, Deuterium Oxide) δ 4.93-4.85 (m, 2H), 4.89-4.70 (m, 3H), 3.86 (dq, J=5.2, 2.6 Hz, 2H), 3.42-3.35 (m, 2H), 3.12 (s, 6H).

##STR00300##

Compound 220: 3-(hydroxymethyl)-4,4-dimethylmorpholin-4-ium

[0907] Step 1:

[0908] To a mixture of 2-(methylamino)ethanol (1 g, 13.31 mmol, 1.07 mL, 1 eq) in DCM (5 mL) was added TBDPSCI added TBDPSCl (4.03 g, 14.65 mmol, 3.76 mL, 1.1 eq), TEA (1.48 g, 14.65 mmol, 2.04 mL, 1.1 eq) and DMAP (813.26 mg, 6.66 mmol, 0.5 eq) in one portion at 25° C. under N2, then heated to 40° C. and stirred for 12 hours. LCMS showed LCMS showed desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=30/1 to 1/1). 1HNMR indicated the desired compound 2-[tert-butyl(diphenyl)silyl]oxy-N-methyl-ethanamine (1.2 g, crude) was obtained as colorless oil.

[0909] Step 2:

[0910] To a mixture of 2-[tert-butyl(diphenyl)silyl]oxy-N-methyl-ethanamine (1 g, 3.19 mmol, 1 eq) in DCE (10 mL) was added AcOH added AcOH (1.92 g, 31.90 mmol, 1.82 mL, 10 eq) in one portion at 25° C. under N.sub.2. The mixture was stirred at 25° C. for 0.5h, then oxetan-3-one (459.72 mg, 6.38 mmol, 2 eq) and NaBH(OAc).sub.3 (1.35 g, 6.38 mmol, 2 eq) were added, and the mixture stirred for 11.5 hours. LC-MS showed the desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=20/1 to 0/1). The crude product N-[2-[tert-butyl(diphenyl)silyl]oxyethyl]-N-methyl-oxetan-3-amine (0.72 g, crude) as white solid.

[0911] Step 3:

[0912] To a mixture of N-[2-[tert-butyl(diphenyl)silyl]oxyethyl]-N-methyl-oxetan-3-amine (0.72 g, 1.95 mmol, 1 eq) in THF (5 mL) was added MeI added MeI (1.66 g, 11.69 mmol, 727.70 uL, 6 eq) in one portion at 25° C. under N.sub.2. The mixture was stirred at 70° C. for 12 hours. LCMS showed the desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=20/1 to 1/1). The crude product 2-[tert-butyl(diphenyl)silyl]oxyethyl-dimethyl-(oxetan-3-yl)ammonium (0.67 g, crude, I) was obtained as yellow solid.

[0913] Step 4:

[0914] 2-[tert-butyl(diphenyl)silyl]oxyethyl-dimethyl-(oxetan-3-yl)ammonium (0.2 g, 520.01 umol, 1 eq) and KF (151.06 mg, 2.60 mmol, 60.91 uL, 5 eq) in THF (5 mL) was stirred at 25 at 25° C. for 12 hours. LC-MS showed desired mass was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC [water (0.1% TFA)-ACN]. The title compound (54 mg, 207.51 umol, 39.90% yield, TFA) was obtained as colorless oil. MS: (M+): 146.1 1H NMR (400 MHz, Deuterium Oxide) δ 3.99-3.77 (m, 6H), 3.57-3.43 (m, 2H), 3.42-3.33 (m, 1H), 3.19 (s, 3H), 3.08 (s, 3H).

##STR00301##

Compound 221: dimethyl(prop-2-yn-1-yl)(2-sulfoethyl)azanium Trifluoroacetate

[0915] To a solution of 2-(methylamino)ethanesulfonic acid (100 mg, 718.53 umol, 1 eq) in H.sub.2O (3 mL) and EtOH (3 mL) was added NaCO.sub.3 (178.91 mg, 2.16 mmol, 3 eq) and 3-bromoprop-1-yne (85.48 mg, 718.53 umol, 61.94 uL, 1 eq). The mixture was stirred at 25° C. for 12 hr. The mixture was added MeI (305.96 mg, 2.16 mmol, 134.19 uL, 3 eq) at 25° C. The mixture was stirred at 25° C. for 12 hr. LCMS showed desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC [water (0.1% TFA)-ACN]. Compound 2-[dimethyl(prop-2-ynyl)ammonio]ethanesulfonate (72 mg, 376.48 umol, 52.4% yield) was obtained as white solid. LCMS: (M+H)+: 192.1. .sup.1H NMR (400 MHz, Deuterium Oxide) δ 4.27-4.22 (m, 2H), 3.81-3.72 (m, 2H), 3.43-3.34 (m, 2H), 3.27-3.21 (m, 1H), 3.17 (s, 6H).

##STR00302##

Compound 222: {3-[dimethyl(prop-2-yn-1-yl)azaniumyl]propyl}boronic Acid

[0916] Step 1:

[0917] To 3-bromoprop-1-ene (500 mg, 4.13 mmol, 1 eq) was added 1,3,2-benzodioxaborole (495.61 mg, 4.13 mmol, 517.34 uL, 1 eq). The mixture was stirred at 100° C. for 4 hr. LCMS showed 3-bromoprop-1-ene was consumed completely and desired mass was detected. The reaction mixture was concentrated under reduced pressure. Compound 2-(3-bromopropyl)-1,3,2-benzodioxaborole (˜1 g, crude) as a yellow oil was obtained and the crude product was used in the next step without further purification.

[0918] Step 2:

[0919] To a solution of 2-(3-bromopropyl)-1,3,2-benzodioxaborole (1 g, 4.15 mmol, 1 eq) in THF (3 mL) was added N added N,N-dimethylprop-2-yn-1-amine (345.10 mg, 4.15 mmol, 440.18 uL, 1 eq). The mixture was stirred at 25° C. for 12 hr. LCMS showed 2-(3-bromopropyl)-1,3,2-benzodioxaborole was consumed completely and desired mass was detected. The reaction mixture was concentrated under reduced pressure. The residue was purified by prep-HPLC [water (0.10% TFA)-ACN]. The title compound (40 mg, 139.41 umol, 3.36% yield, 99% purity, TFA) was obtained as white solid. LCMS: (M+): 170.0 1H NMR (400 MHz, Deuterium Oxide) δ 4.14 (d, J=2.6 Hz, 2H), 3.35-3.27 (m, 2H), 3.16 (t, J=2.5 Hz, 1H), 3.08 (s, 6H), 1.84-1.71 (m, 2H), 0.74 (t, J=8.0 Hz, 2H).

##STR00303##

Compound 223: methylbis(prop-2-yn-1-yl)(2-sulfoethyl)azanium

[0920] To a solution of 2-(methylamino)ethanesulfonic acid (100 mg, 718.53 umol, 1 eq) in DMF (5 mL) was added NaH (57.48 mg, 1.44 mmol, 60% purity, 2 eq) at 0° C. The mixture was stirred at 0° C. for 0.5 hr. Then 3-bromoprop-1-yne (94.02 mg, 790.38 umol, 68.13 uL, 1.1 eq) in DMF (1 mL) was added slowly via syringe at 0° C. The mixture was stirred at 25° C. for 12 hr. LC-MS showed the desired compound was detected. The reaction mixture was quenched by addition of H.sub.2O (1 mL) at 0° C. and concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC [water (0.1% TFA)-ACN]. The title compound (13 mg, 43.89 umol, 6.11% yield, 100% purity) was obtained as a white solid. LCMS: (M+): 216.0 .sup.1H NMR (400 MHz, Deuterium Oxide) δ 4.38 (d, J=2.5 Hz, 4H), 3.92-3.84 (m, 2H), 3.45-3.37 (m, 2H), 3.29 (t, J=2.5 Hz, 2H), 3.24 (s, 3H).

##STR00304##

Compound 227: {2-[dimethyl(prop-2-yn-1-yl)azaniumyl]ethyl}phosphinic Acid

[0921] This compound may be synthesized according to the experimental procedure described for Compound 226.

##STR00305##

Compound 228: (fluoromethyl)(2-methoxyethyl)dimethylazanium Iodide

[0922] 2-methoxy-N,N-dimethyl-ethanamine (50 mg, 484.67 umol, 1 eq) and fluoro(iodo)methane (232.54 mg, 1.45 mmol, 3 eq) in THF (3 mL) was stirred at 25 was stirred at 25° C. for 12 hours. The reaction mixture was filtered and the filter cake was concentrated under reduced pressure to give a residue. The title compound (71 mg, 269.87 umol, 55.68% yield, I) was obtained as white solid. MS: (M+): 136.2 1H NMR (400 MHz, Deuterium Oxide) δ 5.40 (s, 1H), 5.29 (s, 1H), 3.90-3.81 (m, 2H), 3.67-3.60 (m, 2H), 3.35 (s, 3H), 3.17 (s, 6H).

##STR00306##

Compound 229: (fluoromethyl)dimethyl[2-(methylsulfanyl)ethyl]azanium

[0923] Step 1:

[0924] To a mixture of 1-chloro-2-methylsulfanyl-ethane (500 mg, 4.52 mmol, 446.43 uL, 1 eq) in THF (5 mL) was added NaI (1.36 g, 9.04 mmol, 2 eq) and N-methylmethanamine (2 M, 6.78 mL, 3 eq) in one portion at 25° C. under N.sub.2. The mixture was heated to 50° C. and stirred for 12 hours. LCMS showed one main peak with desired mass was detected. The crude product N,N-dimethyl-2-methylsulfanyl-ethanamine (500 mg, crude) was obtained as colorless oil and used into the next step without further purification. LCMS: (M+H+): 120.1

[0925] Step 2:

[0926] To a mixture of N,N-dimethyl-2-methylsulfanyl-ethanamine (200 mg, 1.68 mmol, 1 eq) in THF (5 mL) was added NaI added NaI (502.88 mg, 3.35 mmol, 2 eq) and fluoro(iodo)methane (804.82 mg, 5.03 mmol, 3 eq) in one portion at 25° C. under N.sub.2. The mixture was stirred at 25° C. for 12 hours. LCMS showed N,N-dimethyl-2-methylsulfanyl-ethanamine was consumed incompletely and one new peak with desired mass was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC [water (0.10% TFA)-ACN]. The title compound (62 mg, 222.10 umol, 13.24% yield, I—)—) was obtained as yellow oil. LCMS: (M+): 152.1 1H NMR (400 MHz, Deuterium Oxide) δ 5.27 (s, 1H), 5.16 (s, 1H), 3.54-3.45 (m, 2H), 3.02 (d, J=2.0 Hz, 6H), 2.83-2.70 (m, 2H), 1.99 (d, J=1.0 Hz, 3H).

##STR00307##

Compound 230: [(2E)-2-ethylidene-3-methoxy-3-oxopropyl]dimethyl(prop-2-yn-1-yl)azanium

[0927] This compound may be synthesized according to the experimental procedure described for Compound 52.

##STR00308##

Compound 231: (4-methoxy-3-methylidene-4-oxobutyl)dimethyl(prop-2-yn-1-yl)azanium

[0928] This compound may be synthesized according to the experimental procedure described for Compound 52.

##STR00309##

Compound 236: dimethyl[2-(methylcarbamoyl)-2-methylideneethyl](prop-2-yn-1-yl)azanium

[0929] This compound may be synthesized by reacting Compound 88 with methylamine, followed by concentration in vacuo and purification by recrystallization.

##STR00310##

Compound 237: [(1E)-3-methoxy-3-oxoprop-1-en-1-yl]dimethyl(prop-2-yn-1-yl)azanium

[0930] This compound may be synthesized according to the experimental procedure described for Compound 52.

##STR00311##

Compound 238: [(1Z)-3-methoxy-3-oxoprop-1-en-1-yl]dimethyl(prop-2-yn-1-yl)azanium

[0931] This compound may be synthesized according to the experimental procedure described for Compound 52.

##STR00312##

Compound 240: dimethyl[2-(1,3-oxazol-2-yl)ethyl](prop-2-yn-1-yl)azanium

[0932] This compound may be synthesized according to the experimental procedure described for Compound 239.

##STR00313##

Compound 241: dimethyl(prop-2-yn-1-yl)[2-(1H-1,2,4-triazol-5-yl)ethyl]azanium

[0933] This compound may be synthesized according to the experimental procedure described for Compound 239.

##STR00314##

Compound 242: dimethyl(prop-2-yn-1-yl)[(1H-1,2,4-triazol-5-yl)methyl]azanium

[0934] This compound may be synthesized according to the experimental procedure described for Compound 239.

##STR00315##

Compound 243: dimethyl[2-(1,2,4-oxadiazol-5-yl)ethyl](prop-2-yn-1-yl)azanium

[0935] This compound may be synthesized according to the experimental procedure described for Compound 239.

##STR00316##

Compound 246: {3-[dimethyl(prop-2-yn-1-yl)azaniumyl]prop-1-en-2-yl}phosphonic Acid

[0936] This compound may be synthesized according to the experimental procedure described for Compound 245, followed by hydrolysis of the phosphate esters with NaOH in water and purification by recrystallization.

##STR00317##

Compound 248: (2,2-dimethoxyethyl)(2-hydroxyethyl)dimethylazanium

[0937] This compound may be synthesized according to the experimental procedure described for Compound 247.

##STR00318##

Compound 249: (2-hydroxyethyl)dimethyl(3,3,3-trifluoro-2-oxopropyl)azanium

[0938] This compound may be synthesized according to the experimental procedure described for Compound 247.

##STR00319##

Compound 250: (2-hydroxyethyl)dimethyl[2-(methylsulfanyl)-2-oxoethyl]azanium

[0939] This compound may be synthesized according to the experimental procedure described for Compound 247.

##STR00320##

Compound 253: (2-hydroxyethyl)dimethyl(4-methylpenta-2,3-dien-1-yl)azanium

[0940] This compound may be synthesized according to the experimental procedure described for Compound 60.

##STR00321##

Compound 254: (2-hydroxyethyl)dimethyl(3-methylbuta-1,2-dien-1-yl)azanium

[0941] This compound may be synthesized according to the experimental procedure described for Compound 60.

##STR00322##

Compound 255: N-(2-hydroxyethyl)-N,N,2-trimethylcyclopropan-1-aminium

[0942] Step 1:

[0943] To a mixture of 2-methylcyclopropanamine (0.2 g, 1.86 mmol, 1 eq, HCl) in HCO2H (3 mL) was added HCHO (1.51 g, 18.59 mmol, 1.38 mL, 37% purity, 10 eq) in one portion at 25° C. under N.sub.2. The mixture was stirred at 90° C. for 12 hours. LCMS showed desired m/z was detected. The reaction mixture was concentrated under reduced pressure to remove HCHO and HCO2H. The residue was diluted with H.sub.2O (3 mL). The residue was purified by prep-HPLC [water (0.10% TFA)-ACN]. Compound N,2-trimethylcyclopropanamine (0.176 g, 825.53 umol, 44.41% yield, TFA) was obtained as colorless oil.

[0944] Step 2:

[0945] To a mixture of N,N,2-trimethylcyclopropanamine (0.1 g, 1.01 mmol, 1 eq) and 2-bromoethanol (252.01 mg, 2.02 mmol, 143.19 uL, 2 eq) in THF (3 mL) was added Na.sub.2CO.sub.3 (213.74 mg, 2.02 mmol, 2 eq) and NaI (151.14 mg, 1.01 mmol, 1 eq) in one portion at 25° C. under N.sub.2. The mixture was stirred at 70° C. for 12 hours. LCMS showed desired mass was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC [water (0.1% TFA)-ACN]). The title compound (72 mg, 265.55 umol, 26.34% yield, I) was obtained as colorless oil. MS: (M+): 144.2. .sup.1H NMR (400 MHz, Deuterium Oxide) δ 4.07-3.99 (m, 2H), 3.58-3.45 (m, 2H), 2.95 (d, J=6.4 Hz, 6H), 2.88-2.81 (m, 1H), 1.59-1.51 (m, 1H), 1.34-1.24 (m, 1H), 1.05 (d, J=6.1 Hz, 3H), 0.72 (q, J=7.3 Hz, 1H).

##STR00323##

Compound 256: 2-fluoro-N-(2-hydroxyethyl)-N,N-dimethylcyclopropan-1-aminium

[0946] This compound may be synthesized according to the experimental procedure described for Compound 255.

##STR00324##

Compound 257: 2,2-difluoro-N-(2-hydroxyethyl)-N,N-dimethylcyclopropan-1-aminium

[0947] This compound may be synthesized according to the experimental procedure described for Compound 255.

##STR00325##

Compound 258: N-(2-hydroxyethyl)-N,N,2,2-tetramethylcyclopropan-1-aminium

[0948] This compound may be synthesized according to the experimental procedure described for Compound 255.

##STR00326##

Compound 259: 1-fluoro-N-(2-hydroxyethyl)-N,N-dimethylcyclopropan-1-aminium

[0949] This compound may be synthesized according to the experimental procedure described for Compound 255.

##STR00327##

Compound 260: 1-methyl-1-(prop-2-yn-1-yl)-4,5-dihydro-1H-pyrazol-1-ium

[0950] This compound may be synthesized according to the experimental procedure described for Compound 208.

##STR00328##

Compound 261: 1-methyl-1-(prop-2-yn-1-yl)imidazolidin-1-ium

[0951] This compound may be synthesized according to the experimental procedure described for Compound 208 using boc-protected imidazolidine, followed by deprotection to yield the title compound.

##STR00329##

Compound 264: 3-bromo-1,1-dimethylpyrrolidin-1-ium Bromide

[0952] This compound may be synthesized according to the experimental procedure described for Compound 262.

##STR00330##

Compound 265: 1,1-dimethyl-2-(prop-2-yn-1-yl)pyrazolidin-1-ium

[0953] The title compound may be synthesized by reaction Compound 127 with propargyl bromide, followed by purification by recrystallization.

##STR00331##

Compound 266: (carbamothioylmethyl)(2-hydroxyethyl)dimethylazanium

[0954] This compound may be synthesized according to the experimental procedure described for Compound 247.

##STR00332##

Compound 267: 2-hydroxy-N,N,N-trimethylcyclopropan-1-aminium

[0955] This compound may be synthesized according to the experimental procedure described for Compound 184.

##STR00333##

Compound 271: (2-iodoprop-2-en-1-yl)dimethyl(prop-2-yn-1-yl)azanium

[0956] This compound may be synthesized according to the experimental procedure described for Compound 270.

##STR00334##

Compound 272: [2-(fluoromethyl)prop-2-en-1-yl]dimethyl(prop-2-yn-1-yl)azanium

[0957] This compound may be synthesized according to the experimental procedure described for Compound 270.

##STR00335##

Compound 273: (2-chloroprop-2-en-1-yl)(2-hydroxyethyl)dimethylazanium

[0958] This compound may be synthesized according to the experimental procedure described for Compound 270.

##STR00336##

Compound 274: N-cyclopropyl-N-(2-hydroxyethyl)-N-methylcyclopropanaminium

[0959] This compound may be synthesized according to the experimental procedure described for Compound 58.

##STR00337##

Compound 275: N,N-diethyl-N-(2-hydroxyethyl)cyclopropanaminium

[0960] This compound may be synthesized according to the experimental procedure described for Compound 58.

##STR00338##

Compound 276: (2-carboxyethyl)(fluoromethyl)dimethylazanium

[0961] This compound may be synthesized according to the experimental procedure described for Compound 111.

##STR00339##

Compound 277: (2-carboxyethyl)(chloromethyl)dimethylazanium

[0962] This compound may be synthesized according to the experimental procedure described for Compound 111.

##STR00340##

Compound 278: 1-(fluoromethyl)-1-methylpyrazolidin-1-ium

[0963] This compound may be synthesized according to the experimental procedure described for Compound 74.

##STR00341##

Compound 279: (4R)-4-hydroxy-1-methyl-1-(prop-2-yn-1-yl)pyrazolidin-1-ium

[0964] This compound may be synthesized according to the experimental procedure described for Compound 74, followed by SFC purification of stereoisomers.

##STR00342##

Compound 280: (4S)-4-hydroxy-1-methyl-1-(prop-2-yn-1-yl)pyrazolidin-1-ium

[0965] This compound may be synthesized according to the experimental procedure described for Compound 74, followed by SFC purification of stereoisomers.

##STR00343##

Compound 284: {2-[dimethyl(prop-2-yn-1-yl)azaniumyl]ethyl}boronic Acid

[0966] This compound may be synthesized according to the experimental procedure described for Compound 283.

##STR00344##

Compound 285: [2-(trimethylazaniumyl)ethyl]boronic Acid

[0967] This compound may be synthesized according to the experimental procedure described for

##STR00345##

Compound 289: (fluoromethyl)[(2R)-3-methoxy-2-methyl-3-oxopropyl]dimethylazanium

[0968] Step 1:

[0969] To a solution of (2R)-3-amino-2-methyl-propanoic acid (300 mg, 2.91 mmol, 1 eq) in HCOOH (5 mL) was added formaldehyde (2.36 g, 29.09 mmol, 2.17 mL, 37% purity, 10 eq). The mixture was stirred at 90° C. for 12 hr. LCMS showed the starting reactant was consumed and the product had the desired mass. The mixture was concentrated. The residue was purified by prep-HPLC [water (0.04% HCl)-ACN]. Compound (2R)-3-(dimethylamino)-2-methyl-propanoic acid (300 mg, 2.29 mmol, 78.61% yield) was obtained as a white solid.

[0970] Step 2:

[0971] To a solution of (2R)-3-(dimethylamino)-2-methyl-propanoic acid (250.00 mg, 1.91 mmol, 1 eq) in MeOH (5 mL) was added SOCl2 (453.49 mg, 3.81 mmol, 276.52 uL, 2 eq) at 0° C. The mixture was stirred at 15° C. for 12 hr. LCMS showed the starting reactant was consumed and have the desired mass. The mixture was concentrated. Compound methyl (2R)-3-(dimethylamino)-2-methyl-propanoate (313 mg, crude) was obtained as a white solid and used in the next step without further purification.

[0972] Step 3:

[0973] To a solution of methyl (2R)-3-(dimethylamino)-2-methyl-propanoate (100 mg, 688.71 umol, 1 eq) in THF (5 mL) was added Na.sub.2CO.sub.3 (73.00 mg, 688.71 umol, 1 eq) and fluoro(iodo)methane (330.43 mg, 2.07 mmol, 3 eq). The mixture was stirred at 15° C. for 10 h and stirred at 50° C. for 2 h. LCMS showed the starting reactant was consumed and the product had the desired mass. The mixture was filtered and the filtrate was concentrated. The residue was purified by prep-HPLC [water (0.04% HCl)-ACN]. Compound fluoromethyl-[(2R)-3-methoxy-2-methyl-3-oxo-propyl]-dimethyl-ammonium (19 mg, 62.27 umol, 9.04% yield, I—)—) was obtained as a white solid. MS: (M+): 178.1 .sup.1H NMR (400 MHz, Deuterium Oxide) δ 5.30 (q, J=6.0 Hz, 1H), 5.18 (q, J=6.0 Hz, 1H), 3.92 (dd, J=14.0, 8.9 Hz, 1H), 3.65 (s, 3H), 3.37-3.29 (m, 1H), 3.17-3.10 (m, 1H), 3.04 (dd, J=7.2, 2.1 Hz, 6H), 1.19 (d, J=7.2 Hz, 3H).

##STR00346##

Compound 290: (fluoromethyl)[(2S)-3-methoxy-2-methyl-3-oxopropyl]dimethylazanium

[0974] Step 1:

[0975] To a mixture of (2S)-3-amino-2-methyl-propanoic acid (0.3 g, 2.91 mmol, 1 eq) in HCOOH (5 mL) was added aq. HCHO (2.36 g, 29.09 mmol, 2.17 mL, 37% purity, 10 eq) in one portion at 25° C. under N.sub.2. The mixture was stirred at 90° C. for 12 hours. LCMS showed the desired mass was detected. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC [water (0.1% TFA)-ACN]; B %:1%-5%, 12 min). Compound (2S)-3-(dimethylamino)-2-methyl-propanoic acid (0.2 g, 815.68 umol, 28.04% yield, TFA) was obtained as colorless oil.

[0976] Step 2:

[0977] To a mixture of (2S)-3-(dimethylamino)-2-methyl-propanoic acid (0.2 g, 1.13 mmol, 1 eq) in MeOH (5 mL) was added SOCl.sub.2 (268.56 mg, 2.26 mmol, 163.76 uL, 2 eq) in one portion at 25° C. under N.sub.2. The mixture was stirred at 25° C. for 12 hours. LCMS showed desired mass was detected. The reaction mixture was concentrated under reduced pressure to give a residue. No further purification was performed. Compound methyl (2S)-3-(dimethylamino)-2-methyl-propanoate (150 mg, 1.03 mmol, 91.53% yield) was obtained as a white solid.

[0978] Step 3:

[0979] To a mixture of methyl (2S)-3-(dimethylamino)-2-methyl-propanoate (50 mg, 276.78 umol, 1 eq, C.sub.1) and fluoro(iodo)methane (132.79 mg, 830.33 umol, 3 eq) in THF (5 mL) was added Na2CO3 added Na2CO3 (88.01 mg, 830.33 umol, 3 eq) in one portion at 25° C. under N.sub.2. The mixture was stirred at 25° C. for 12 hours. LCMS showed desired mass was detected. The reaction mixture was filtered and concentrated under remove THF, then the residue was diluted with H.sub.2O (3 ml), washed by EtOAc 9 ml (3 ml*3). The water phase was lyophilized. The title compound (32 mg, 179.55 umol, 64.87% yield) was obtained as white solid. MS: (M+): 178.1 1H NMR (400 MHz, Deuterium Oxide) δ 5.28 (q, J=6.0 Hz, 1H), 5.17 (q, J=6.0 Hz, 1H), 3.91 (dd, J=14.0, 9.0 Hz, 1H), 3.64 (s, 3H), 3.36-3.28 (m, 1H), 3.16-3.08 (m, 1H), 3.03 (dd, J=7.4, 2.1 Hz, 6H), 1.18 (d, J=7.3 Hz, 3H).

##STR00347##

Compound 291: (fluoromethyl)[(2R)-2-hydroxypropyl]dimethylazanium

[0980] Step 1:

[0981] To a solution of methyl (2R)-2-hydroxypropanoate (10 g, 96.06 mmol, 9.17 mL, 1 eq) and imidazole (13.08 g, 192.12 mmol, 2.0 eq) in DCM (100 mL) was added tert added tert-butyl-chloro-dimethyl-silane (28.96 g, 192.12 mmol, 23.54 mL, 2 eq) at 0° C. The mixture was stirred at 25° C. for 12 hr. TLC indicated methyl (2R)-2-hydroxypropanoate was consumed completely and many new spots formed. The reaction mixture was diluted with H.sub.2O (150 mL) and extracted with ethyl acetate 450 mL (150 mL*3). The combined organic layers were dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=1:0). Compound methyl (2R)-2-[tert butyl(dimethyl)silyl]oxypropanoate (15 g, 68.69 mmol, 71.51% yield) was obtained as a colorless oil.

[0982] Step 2:

[0983] To a solution of methyl (2R)-2-[tert-butyl(dimethyl)silyl]oxypropanoate (5 g, 22.90 mmol, 1 eq) in THF (100 mL) was added DIBAL added DIBAL-H (1 M, 45.79 mL, 2 eq, in Tol.) at 0° C. The mixture was stirred at 25 at 25° C. for 12 hr. TLC indicated new spots formed. The reaction mixture was quenched by addition of H.sub.2O (100 mL) at 0° C., and then diluted with ethyl acetate 50 mL. The mixture was filtered. The filtrate was extracted with ethyl acetate 300 mL (100 mL*3). The combined organic layers were dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=1:0 to 10:1). Compound (2R)-2-[tert-butyl(dimethyl)silyl]oxypropan-1-ol (6 g, 36.77 mmol, 80.30% yield) was obtained as a colorless oil without further purification.

[0984] Step 3:

[0985] To a solution of (2R)-2-[tert-butyl(dimethyl)silyl]oxypropan-1-ol (6 g, 31.52 mmol, 1 eq) in DCM (60 mL) was added imidazole (10.73 g, 157.60 mmol, 5 eq), PPh3 (20.67 g, 78.80 mmol, 2.5 eq) and 12 (24.00 g, 94.56 mmol, 19.05 mL, 3 eq) at 25° C. The mixture was stirred at 25° C. for 12 hr. TLC indicated (2R)-2-[tert-butyl(dimethyl)silyl]oxypropan-1-ol was consumed completely. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=1:0). Compound tert-butyl-[(1R)-2-iodo-1-methyl-ethoxy]-dimethyl-silane (6 g, 19.98 mmol, 63.40% yield) was obtained as a colorless oil.

[0986] Step 4:

[0987] To a solution of tert-butyl-[(1R)-2-iodo-1-methyl-ethoxy]-dimethyl-silane (4 g, 13.32 mmol, 1 eq) in THF (50 mL) was added N-methylmethanamine (2 M, 33.31 mL, 5 eq)(in THF) at 25° C. The mixture was stirred at 25° C. for 12 hr. LCMS showed the desired compound was detected. TLC indicated most of tert-butyl-[(1R)-2-iodo-1-methyl-ethoxy]-dimethyl-silane remained. The mixture was stirred at 70° C. for 12 hr. The reaction mixture was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=1:0 to 0:1). Compound (2R)-2-[tert-butyl(dimethyl)silyl]oxy-N,N-dimethyl-propan-1-amine (3 g, 8.69 mmol, 65.21% yield, HI) was obtained as yellow oil.

[0988] Step 5:

[0989] To a solution of (2R)-2-[tert-butyl(dimethyl)silyl]oxy-N,N-dimethyl-propan-1-amine (3 g, 8.69 mmol, 1 eq, HI) in THF (30 mL) was added fluoro(iodo)methane (6.95 g, 43.44 mmol, 5 eq) at 25° C. The mixture was stirred at 25° C. for 12 hr. TLC indicated (2R)-2-[tert-butyl(dimethyl)silyl]oxy-N,N-dimethyl-propan-1-amine was remained. The mixture was added another batch of fluoro(iodo)methane (4.17 g, 26.06 mmol, 3 eq) at 25° C. The mixture was stirred at 50° C. for 3 hr. TLC indicated (2R)-2-[tert-butyl(dimethyl)silyl]oxy-N,N-dimethyl-propan-1-amine was consumed completely. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was diluted with H.sub.2O 100 mL and extracted with ethyl acetate 150 mL (50 mL*3). The aqueous phase was freeze-dried.

[0990] Compound tert-butyl-[(1R)-2-[N-(fluoromethyl)-dimethyl-azanyl]-1-methyl-ethoxy]-dimethyl-silane (900 mg, 2.39 mmol, 27.45% yield) was obtained as a white solid without further purification.

[0991] Step 6:

[0992] To a solution of tert-butyl-[(1R)-2-[N-(fluoromethyl)-dimethyl-azanyl]-1-methyl-ethoxy]-dimethyl-silane (500 mg, 1.33 mmol, 1 eq) in H.sub.2O (20 mL) was added KF (1.54 g, 26.50 mmol, 620.80 uL, 20 eq) at 25° C. The mixture was stirred at 50° C. for 12 hr. LC-MS showed the desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was diluted with H.sub.2O 5 mL and extracted with ethyl acetate 15 mL (5 mL*3). The residue was purified by prep-HPLC [water (0.1% TFA)-ACN]. Compound fluoromethyl-[(2R)-2-hydroxypropyl]-dimethyl-ammonium (98 mg, 372.49 umol, 28.11% yield, I—) was obtained as yellow oil. MS: (M+): 136.1 .sup.1H NMR (400 MHz, Deuterium Oxide) δ 5.48-5.37 (m, 1H), 5.37-5.26 (m, 1H), 4.37 (q, J=6.4 Hz, 1H), 3.42-3.36 (m, 2H), 3.20-3.14 (m, 6H), 1.18 (d, J=6.4 Hz, 3H).

##STR00348##

Compound 292: (fluoromethyl)[(2S)-2-hydroxypropyl]dimethylazanium

[0993] This compound may be synthesized according to the experimental procedure described for Compound 169.

##STR00349##

Compound 294: trimethyl(2-methylpent-4-yn-2-yl)azanium

[0994] This compound may be synthesized according to the experimental procedure described for Compound 174.

##STR00350##

Compound 295: (fluoromethyl)dimethyl[(pyrrolidin-2-yl)methyl]azanium Iodide

[0995] Step 1:

[0996] To a mixture of pyrrolidin-2-ylmethanol (5 g, 49.43 mmol, 1 eq) and ethyl 2,2,2-trifluoroacetate (10.53 g, 74.15 mmol, 10.23 mL, 1.5 eq) in MeOH (50 mL) was added TEA added TEA (5.50 g, 54.38 mmol, 7.57 mL, 1.1 eq) in one portion at 25° C. under N.sub.2. The mixture as heated to 70° C. and stirred for 12 hours. TLC indicated pyrrolidin-2-ylmethanol was consumed and one new spot formed. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=50/1 to 0/1). Compound 2,2,2-trifluoro-1-[2-(hydroxymethyl)pyrrolidin-1-yl]ethanone (5 g, 25.36 mmol, 51.30% yield) was obtained as yellow oil. LCMS: (M+H+): 198.0.

[0997] Step 2:

[0998] To a mixture of PPh3 (6.39 g, 24.35 mmol, 2.4 eq) in DCM (15 mL) was added imidazole (3.45 g, 50.72 mmol, 5 eq) and 12 (6.44 g, 25.36 mmol, 2.5 eq) in one portion at 25° C. under N.sub.2. The mixture was stirred at 25° C. for 6 min, then 2,2,2-trifluoro-1-[2-(hydroxymethyl)pyrrolidin-1-yl]ethanone (2 g, 10.14 mmol, 1 eq) was added. The mixture was stirred at 25° C. for 11.9 hours. TLC indicated 2,2,2-trifluoro-1-[2-(hydroxymethyl)pyrrolidin-1-yl]ethanone was consumed and many new spots formed. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=100/1 to 5/1). Compound 2,2,2-trifluoro-1-[2-(iodomethyl)pyrrolidin-1-yl]ethanone (2.7 g, crude) was obtained as colorless oil.

[0999] Step 3:

[1000] To a mixture of 2,2,2-trifluoro-1-[2-(iodomethyl)pyrrolidin-1-yl]ethanone (1 g, 3.26 mmol, 1 eq) in THF (5 mL) was added Na.sub.2CO.sub.3 (690.37 mg, 6.51 mmol, 2 eq) and N-methylmethanamine (2 M, 4.89 mL, 3 eq) in one portion at 25° C. under N.sub.2. The mixture was stirred at 25° C. for 12 hours. TLC indicated 2,2,2-trifluoro-1-[2-(iodomethyl)pyrrolidin-1-yl]ethanone was consumed and one new spot formed. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=80/1 to 0/1). Compound 1-[2-[(dimethylamino)methyl]pyrrolidin-1-yl]-2,2,2-trifluoro-ethanone (150 mg, crude) was obtained as yellow oil. LCMS: (M+H+): 225.1

[1001] Step 4:

[1002] To a mixture of 1-[2-[(dimethylamino)methyl]pyrrolidin-1-yl]-2,2,2-trifluoro-ethanone (100 mg, 445.98 umol, 1 eq) in THF (3 mL) was added NaI (133.70 mg, 891.97 umol, 2 eq), Na.sub.2CO.sub.3 (94.54 mg, 891.97 umol, 2 eq) and fluoro(iodo)methane (213.98 mg, 1.34 mmol, 3 eq) in one portion at 25° C. under N.sub.2. The mixture was stirred at 25° C. for 12 hours. LCMS showed desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC [water (0.1% TFA)-ACN]. The title compound (88 mg, 305.40 umol, 68.48% yield, I) was obtained as colorless oil. LCMS: (M+H+): 161.2 1H NMR (400 MHz, Deuterium Oxide) δ 5.42 (s, 1H), 5.31 (s, 1H), 4.12-4.03 (m, 1H), 3.96-3.81 (m, 2H), 3.31 (dd, J=8.5, 6.4 Hz, 2H), 3.17 (s, 6H), 2.40-2.30 (m, 1H), 2.05-2.00 (m, 1H), 1.98-1.84 (m, 1H), 1.84-1.69 (m, 1H).

##STR00351##

Compound 296: (fluoromethyl)dimethyl{[1-(2,2,2-trifluoroacetyl)pyrrolidin-2-yl]methyl}azanium

[1003] This compound may be synthesized by reacting Compound 295 with trifluoracetic anhydride, followed by purification by recrystallization.

##STR00352##

Compound 297: (fluoromethyl)(3-methoxy-2-methyl-3-oxopropyl)dimethylazanium Iodide

[1004] Step 1:

[1005] To a mixture of 3-amino-2-methyl-propanoic acid (300 mg, 2.91 mmol, 1 eq) in HCOOH (5 mL) was added HCHO (2.36 g, 29.09 mmol, 2.17 mL, 37% purity, 10 eq) in one portion at 25° C. under N.sub.2. The mixture was stirred at 90° C. for 12 hours. LCMS showed desired mass was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC [water (0.10% TFA)-ACN]. Compound 3-(dimethylamino)-2-methyl-propanoic acid (100 mg, 762.36 umol, 26.20% yield) was obtained as colorless oil.

[1006] Step 2:

[1007] To a mixture of 3-(dimethylamino)-2-methyl-propanoic acid (50 mg, 381.18 umol, 1 eq) in MeOH (3 mL) was added SOCl.sub.2 (90.70 mg, 762.35 umol, 55.30 uL, 2 eq) in one portion at 25° C. under N.sub.2. The mixture was stirred at 25° C. for 12 hours. LCMS showed desired mass was detected. The reaction mixture was concentrated under reduced pressure to give a residue. No further purification was performed. The crude product methyl 3-(dimethylamino)-2-methyl-propanoate (50 mg, crude) was obtained as colorless oil, and used in the next step without further purification.

[1008] Step 3:

[1009] To a mixture of methyl 3-(dimethylamino)-2-methyl-propanoate (50 mg, 344.35 umol, 1 eq) in THF (3 mL) was added Na.sub.2CO.sub.3 (73.00 mg, 688.71 umol, 2 eq) in one portion at 25° C. under N.sub.2, then fluoro(iodo)methane (165.22 mg, 1.03 mmol, 3 eq) was added. The mixture was stirred at 25° C. for 12 hours. LCMS showed desired compound was detected. The reaction mixture was filtered and concentrated under reduced pressure to remove THF, then 5 ml H.sub.2O was added, extracted with EtOAc (15 ml, 5 ml*3). The water phase was lyophilized. The title compound (39.4 mg, 221.07 umol, 64.20% yield) was obtained as white solid. LCMS: (M+H+): 178.1 .sup.1H NMR (400 MHz, Deuterium Oxide) δ 5.4 (m, 1H), 5.3 (m, 1H), 4.1-4.0 (m, 1H), 3.8 (s, 3H), 3.4 (m, 1H), 3.2 (m, 1H), 3.2-3.1 (m, 6H), 1.3 (m, 3H).

##STR00353##

Compound 298: [2-(benzyloxy)ethyl](chloromethyl)methyl(prop-2-yn-1-yl)azanium

[1010] Step 1:

[1011] To a mixture of 2-benzyloxyacetaldehyde (500 mg, 3.33 mmol, 467.29 uL, 1 eq) and N-methylprop-2-yn-1-amine (230.08 mg, 3.33 mmol, 277.21 uL, 1 eq) in THF (3 mL) was added AcOH (10.00 mg, 166.47 umol, 9.52 uL, 0.05 eq) in one portion at 0° C. under N.sub.2. The mixture was stirred at 0° C. for 30 min, then NaBH(OAc).sub.3 (1.41 g, 6.66 mmol, 2 eq) was added heated to 25° C. and stirred for 11.5 hours. LC-MS showed desired mass was detected. The reaction mixture was diluted with H.sub.2O (3 mL) and extracted with EtOAc 15 mL (5 mL*3), filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=20/1 to 1/1). compound N-(2-benzyloxyethyl)-N-methyl-prop-2-yn-1-amine (600 mg, 2.95 mmol, 88.65% yield) was obtained as yellow oil.

[1012] Step 2:

[1013] To a mixture of N-(2-benzyloxyethyl)-N-methyl-prop-2-yn-1-amine (20 mg, 98.39 umol, 1 eq) in acetone (0.5 mL) was added chloro(iodo)methane (86.77 mg, 491.93 umol, 35.71 uL, 5 eq). The mixture was stirred at 25° C. for 12 hours. LCMS showed desired mass was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (column: Waters Xbridge BEH C18 100*30 mm*10 um; mobile phase: [water (0.04% HCl)-ACN]; B %: 5%-35%, 12 min). The title compound (18 mg, 47.41 umol, 48.19% yield, I—) was obtained as colorless oil. LCMS: (M+): 252.1 .sup.1H NMR (400 MHz, Deuterium Oxide) δ 7.45-7.36 (m, 5H), 5.30 (s, 2H), 4.60 (s, 2H), 4.44 (d, J=2.5 Hz, 2H), 4.03-3.97 (m, 2H), 3.87-3.80 (m, 2H), 3.27 (s, 4H).

##STR00354##

Compound 299: [2-(benzyloxy)ethyl](fluoromethyl)methyl(prop-2-yn-1-yl)azanium

[1014] Step 1:

[1015] To a mixture of 2-benzyloxyacetaldehyde (500 mg, 3.33 mmol, 467.29 uL, 1 eq) and N-methylprop-2-yn-1-amine (230.08 mg, 3.33 mmol, 277.21 uL, 1 eq) in THF (3 mL) was added AcOH (10.00 mg, 166.47 umol, 9.52 uL, 0.05 eq) in one portion at 0° C. under N.sub.2. The mixture was stirred at 0° C. for 30 min, then NaBH(OAc).sub.3 (1.41 g, 6.66 mmol, 2 eq) was added heated to 25° C. and stirred for 11.5 hours. LC-MS showed desired mass was detected. The reaction mixture was diluted with H.sub.2O (3 mL) and extracted with EtOAc 15 mL (5 mL*3), filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=20/1 to 1/1). compound N-(2-benzyloxyethyl)-N-methyl-prop-2-yn-1-amine (600 mg, 2.95 mmol, 88.65% yield) was obtained as yellow oil.

[1016] Step 2:

[1017] To a solution of N-(2-benzyloxyethyl)-N-methyl-prop-2-yn-1-amine (100 mg, 491.93 umol, 1 eq) in THF (5 mL) was added fluoro(iodo)methane (236.02 mg, 1.48 mmol, 3 eq) at 25° C. The mixture was stirred at 25° C. for 12 hr. LCMS showed the desired compound was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was diluted with H.sub.2O 5 mL and extracted with ethyl acetate 15 mL (5 mL*3). The residue was purified by prep-HPLC [water (0.04% HCl)-ACN]. Desired compound 2-benzyloxyethyl-(fluoromethyl)-methyl-prop-2-ynyl-ammonium (58 mg, 159.69 umol, 32.46% yield, I—) was obtained as colorless oil. LCMS: (M+): 236.1 1H NMR (400 MHz, Methanol-d4) δ 7.41-7.27 (m, 5H), 5.70-5.62 (m, 1H), 5.59-5.51 (m, 1H), 4.60 (s, 2H), 4.57-4.51 (m, 2H), 3.99-3.94 (m, 2H), 3.86-3.79 (m, 2H), 3.62 (t, J=2.6 Hz, 1H), 3.28 (d, J=2.1 Hz, 3H).

##STR00355##

Compound 300: (fluoromethyl)(methyl)(prop-2-yn-1-yl)(2-{[(2S)-3,3,3-trifluoro-2-methoxy-2-phenylpropanoyl]oxy}ethyl)azanium

[1018] Step 1:

[1019] A mixture of (2S)-3,3,3-trifluoro-2-methoxy-2-phenyl-propanoic acid (0.6 g, 2.56 mmol, 461.54 uL, 1 eq), 2-[methyl(prop-2-ynyl)amino]ethanol (318.65 mg, 2.82 mmol, 1.1 eq), DCC (581.02 mg, 2.82 mmol, 569.62 uL, 1.1 eq), DMAP (156.38 mg, 1.28 mmol, 0.5 eq) in DCM (15 mL) was degassed and purged with N.sub.2 for 3 times, and then the mixture was stirred at 0° C. for 0.5 hr. LCMS showed desired mass was detected. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=90/1 to 30/1). Compound 2-[methyl(prop-2-ynyl)amino]ethyl (2S)-3,3,3-trifluoro-2-methoxy-2-phenyl-propanoate (100 mg, 303.66 umol, 11.86% yield) was obtained as colorless oil.

[1020] Step 2:

[1021] 2-[methyl(prop-2-ynyl)amino]ethyl (2S)-3,3,3-trifluoro-2-methoxy-2-phenyl-propanoate (200 mg, 607.32 umol, 1 eq) in fluoro(iodo)methane (1.94 g, 12.15 mmol, 20 eq) was stirred at 25° C. for 12 hours. LC-MS showed desired mass was detected. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC

[water (0.04% HCl)-ACN]. The title compound (47 mg, 96.07 umol, 15.82% yield, I—) was obtained as white solid. MS (M+): 362.0 1H NMR (400 MHz, Deuterium Oxide) δ 7.58-7.47 (m, 5H), 5.46-5.36 (m, 1H), 5.34-5.25 (m, 1H), 4.84 (s, 2H), 4.31-4.17 (m, 2H), 3.95 (s, 2H), 3.47 (s, 3H), 3.27-3.22 (m, 1H), 3.08 (s, 3H).

##STR00356##

Compound 301: 1-(fluoromethyl)-1-(2-hydroxyethyl)pyrrolidin-1-ium

[1022] To a mixture of 2-pyrrolidin-1-ylethanol (100 mg, 868.26 umol, 101.52 uL, 1 eq) in THF (3 mL) was added fluoro added fluoro(iodo)methane (416.58 mg, 2.60 mmol, 3 eq) in one portion at 25° C. under N.sub.2. The mixture was stirred at 25° C. for 12 hours. LCMS showed desired mass was detected. The reaction mixture was concentrated under reduced pressure to remove THF, and diluted with H.sub.2O 3 mL and extracted with EtOAc (15 mL, 5 mL*3). The water phase was lyophilized. The title compound (90 mg, 327.15 umol, 37.68% yield, I—) was obtained as yellow solid. MS (M+): 148.0 1H NMR (400 MHz, Deuterium Oxide) δ 5.47 (s, 1H), 5.35 (s, 1H), 4.03-3.96 (m, 2H), 3.78-3.68 (m, 2H), 3.70-3.59 (m, 4H), 2.24-2.05 (m, 4H).

##STR00357##

Compound 303: N,N-dimethyl-N-(2-oxo-2-(((2S,3R,4S,5R,6R)-3,4,5-triacetoxy-6-(acetoxymethyl)tetrahydro-2H-pyran-2-yl)oxy)ethyl)prop-2-yn-1-aminium Trifluoroacetate

[1023] Step 1

[1024] Compound 2,3,4,6-tetra-O-acetyl-D-glucopyranose (Carbosynth, 2 g, 5.7 mmol, 1 eq) was dissolved in DCM (10 mL), and cooled to 0° C. under N.sub.2. Then 2-chloroacetyl chloride (1.4 mL, 17.6 mmol, 3 eq) and pyridine (1.4 mL, 17.4 mmol, 3 eq) in DCM (10 mL) were added over ice, the reaction was stirred overnight. The reaction mixture was filtered and purified by column chromatography (0-100% EtOAc in hexanes) to yield 1-(2-chloroacetoxy)-2,3,4,6-tetra-O-acetyl-D-glucopyranose (1.18 g, 2.8 mmol, 48% yield).

[1025] Step 2

[1026] Compound 1-(2-chloroacetoxy)-2,3,4,6-tetra-O-acetyl-D-glucopyranose (1.18 g, 2.8 mmol, 1 eq) was dissolved in acetone, followed by addition of sodium iodide (0.625 g, 4.2 mmol, 1.5 eq). The reaction was stirred for a few minutes, followed by addition of dimethylamino-1-propyne (0.76 mL, 7.1 mmol, 2.5 eq). The reaction was stirred at room temperature overnight, then filtered. The filtrate was concentrated, diluted with DMSO and water, then purified by reverse phase C18 column chromatography (0.1% TFA in 95% water/5% MeCN to 100% MeCN). Fractions containing product were lyophilized to yield an off-white powder (420 mg, 0.72 mmol, 26% yield). LCMS (M+): 472.4 .sup.1H NMR (400 MHz, DMSO-d6) δ 6.13 (d, J=8.2 Hz, 1H), 5.48 (t, J=9.6 Hz, 1H), 5.08-4.94 (m, 2H), 4.75-4.46 (m, 4H), 4.35-3.95 (m, 4H), 3.28-3.15 (m, 6H), 2.07-1.86 (m, 12H).

##STR00358##

Compound 304: N,N-dimethyl-N-(2-oxo-2-(((2R,3R,4S,5R)-3,4,5,6-tetraacetoxytetrahydro-2H-pyran-2-yl)methoxy)ethyl)prop-2-yn-1-aminium Trifluoroacetate

[1027] Step 1:

[1028] Compound 1,2,3,4-tetra-O-acetyl-beta-D-glucopyranose (Alfa Aesar, CAS: 13100-46-4, 0.5 g, 1.44 mmol, 1 eq) was dissolved in DCM (5 mL) and cooled to 0° C. Chloroacetyl chloride (0.3425 mL, 4.3 mmol, 3 eq) was added, followed by dropwise addition of pyridine (0.35 mL, 4.3 mmol, 3 eq). The reaction was directly loaded onto silica, and purified by column chromatography (100% hexanes to 100% ethyl acetate) to yield 6-(2-chloroacetoxy)-1,2,3,4-tetra-O-acetyl-beta-D-glucopyranose as an off-white to yellow solid (0.5 g, 1.18 mmol, 82% yield).

[1029] Step 2:

[1030] Compound 6-(2-chloroacetoxy)-1,2,3,4-tetra-O-acetyl-beta-D-glucopyranose (0.5 g, 1.18 mmol, 1 eq) was dissolved in acetone, followed by addition of sodium iodide (0.35 g, 2.3 mmol, 2 eq). The reaction was stirred for a few minutes, followed by addition of 3-dimethylamino-1-propyne (0.25 mL, 2.3 mmol, 2 eq) was added. The reaction was stirred at room temperature overnight, then filtered. The filtrate was concentrated, diluted with DMSO and water and purified by reverse phase C18 column chromatography (0.1% TFA in water 95%/MeCN 5% to 100% MeCN). Fractions containing product was lyophilized to yield an off-white powder (476 mg, 0.81 mmol, 69% yield). LCMS (M+): 472.1 1H NMR (400 MHz, DMSO-d6) δ 5.98 (d, J=8.3 Hz, 1H), 5.47 (t, J=9.6 Hz, 1H), 5.10-4.84 (m, 2H), 4.50 (t, J=2.2 Hz, 4H), 4.41-4.17 (m, 3H), 4.16-4.07 (m, 1H), 3.23 (d, J=145.8 Hz, 6H), 2.15-1.88 (m, 12H).

Compound 306: dimethyl(prop-2-yn-1-yl)(2-{[(2S,3R,4S,5S)-3,4,5-tris(acetyloxy)oxan-2-yl]oxy}ethyl)azanium Iodide

[1031] Step 1:

[1032] To a mixture of (2R,3S,4S)-2,3,4,5-tetrahydroxypentanal (10 g, 66.61 mmol, 1 eq) in pyridine (100 mL) was added Ac.sub.2O (40.80 g, 399.65 mmol, 37.43 mL, 6 eq) in one portion at 25° C. under N.sub.2. The mixture was stirred at 25° C. for 12 hours. TLC indicated one new spot formed. The reaction mixture was concentrated under reduced pressure to remove pyridine. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=30/1 to 0/1). Compound [(3S,4S,5R,6S)-4,5,6-triacetoxytetrahydropyran-3-yl] acetate (20 g, crude) was obtained as yellow oil.

[1033] Step 2:

[1034] To a mixture of [(3S,4S,5R,6S)-4,5,6-triacetoxytetrahydropyran-3-yl] acetate (10 g, 31.42 mmol, 1 eq) in DCM (100 mL) was added 2-bromoethanol (5.89 g, 47.13 mmol, 3.35 mL, 1.5 eq) and BF.sub.3.Et.sub.2O (22.30 g, 157.10 mmol, 19.39 mL, 5 eq) in one portion at 0° C. under N.sub.2. The mixture was heated to 25° C. and stirred for 12 hours. TLC indicated [(3S,4S,5R,6S)-4,5,6-triacetoxytetrahydropyran-3-yl] acetate was consumed and one new spot formed. The reaction mixture was quenched by addition of H.sub.2O (50 mL), and then diluted with H.sub.2O (100 mL) and extracted with DCM (100 mL*2). The combined organic layers were dried over Na.sub.2SO.sub.4, filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=20/1 to 0/1). Then the residue was purified by prep-HPLC [water (0.1% TFA)-ACN]. .sup.1HNMR indicated compound [(3S,4S,5R,6S)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate (1.2 g, 3.13 mmol, 9.97% yield) was obtained as yellow oil and compound[(3S,4S,5R,6R)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate (2.3 g, 6.00 mmol, 19.10% yield) was obtained as yellow oil.

[1035] Step 3:

[1036] To a mixture of [(3S,4S,5R,6R)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate (1.00 g, 2.61 mmol, 1 eq) in acetone (10 mL) was added NaI (430.29 mg, 2.87 mmol, 1.1 eq) in one portion at 25° C. under N.sub.2. Then N,N-dimethylprop-2-yn-1-amine (1.08 g, 13.05 mmol, 1.38 mL, 5 eq) was added. The mixture was heated to 90° C. and stirred for 2 hours. TLC indicated [(3S,4S,5R,6R)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate was consumed and one new spot formed. The reaction mixture was filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=10/1 to Ethyl acetate:Methanol=3/1). The title compound (109 mg, 210.22 umol, 8.06% yield, 99% purity, I) was obtained as white solid. LCMS: (M+): 386.1 1H NMR (400 MHz, Methanol-d4) δ 5.21 (s, 1H), 5.12-5.06 (m, 2H), 4.48 (d, J=2.5 Hz, 2H), 4.43 (dd, J=11.6, 3.9 Hz, 1H), 4.38 (q, J=4.4 Hz, 1H), 4.29-4.16 (m, 2H), 4.06-3.96 (m, 1H), 3.86-3.74 (m, 2H), 3.61 (t, J=2.5 Hz, 1H), 3.31 (s, 6H), 2.20-1.95 (m, 9H).

##STR00359##

Compound 308: dimethyl(prop-2-yn-1-yl)(2-{[(3R,4R,5R)-3,4,5-tris(acetyloxy)oxan-2-yl]oxy}ethyl)azanium

[1037] Step 1:

[1038] To a mixture of (2R,3R,4R)-2,3,4,5-tetrahydroxypentanal (10 g, 66.61 mmol, 1 eq) in pyridine (100 mL) was added Ac.sub.2O (40.80 g, 399.65 mmol, 37.43 mL, 6 eq) in one portion at 25° C. under N.sub.2. The mixture was stirred at 25° C. for 12 hours. TLC indicated one new spot formed. The reaction mixture was concentrated under reduced pressure to remove the pyridine. The residue was purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=30/1 to 0/1). Compound [(3R,4R,5R)-4,5,6-triacetoxytetrahydropyran-3-yl] acetate (20 g, crude) was obtained as white solid.

[1039] Step 2:

[1040] To a mixture of [(3R,4R,5R)-4,5,6-triacetoxytetrahydropyran-3-yl] acetate (10 g, 31.42 mmol, 1 eq) in DCM (50 mL) was added 2-bromoethanol (5.89 g, 47.13 mmol, 3.35 mL, 1.5 eq) in one portion at 0° C. under N.sub.2. BF.sub.3.Et.sub.2O was added (22.30 g, 157.10 mmol, 19.39 mL, 5 eq) and stirred at 0° C. for 6 minutes, then heated to 25° C. and stirred for 11.9 hours. TLC indicated [(3R,4R,5R)-4,5,6-triacetoxytetrahydropyran-3-yl] acetate was consumed and one new spot formed. The reaction mixture was quenched by addition H.sub.2O (20 mL), then diluted with H.sub.2O (50 mL) and extracted with DCM (100 mL, 50 mL*2). The combined organic layers were dried over Na.sub.2SO.sub.4, filtered and the filtrate was concentrated under reduced pressure to give a residue. The residue purified by column chromatography (SiO.sub.2, Petroleum ether/Ethyl acetate=20/1 to 0/1). Then the residue was purified by prep-HPLC [water (0.1% TFA)-ACN]. Compound [(3R,4R,5R)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate (2.6 g, 6.79 mmol, 21.60% yield) was obtained as a yellow oil.

[1041] Step 3:

[1042] To a mixture of [(3R,4R,5R)-4,5-diacetoxy-6-(2-bromoethoxy)tetrahydropyran-3-yl] acetate (1.5 g, 3.91 mmol, 1 eq) in acetone (15 mL) was added N,N-dimethylprop-2-yn-1-amine (1.63 g, 19.57 mmol, 2.08 mL, 5 eq) in one portion at 25° C. under N.sub.2. Then the mixture was heated to 90° C. and stirred for 2 hours. LCMS showed one main peak with expected mass was detected. The reaction mixture was concentrated under reduced pressure to give a residue. Then the residue was purified by prep-HPLC [water (0.10% TFA)-ACN]. The title compound (619 mg, 1.33 mmol, 33.91% yield, 100% purity, Br—) was obtained as colorless oil. LCMS: (M+): 386.2 .sup.1H NMR (400 MHz, Methanol-d4) δ 5.49 (t, J=3.3 Hz, 1H), 5.11 (dt, J=6.8, 3.5 Hz, 1H), 5.01 (dd, J=5.2, 3.3 Hz, 1H), 4.96 (d, J=5.1 Hz, 1H), 4.42 (d, J=2.6 Hz, 2H), 4.26 (dt, J=13.5, 4.4 Hz, 1H), 4.13-3.99 (m, 2H), 3.86 (dd, J=12.1, 6.6 Hz, 1H), 3.78 (t, J=4.7 Hz, 2H), 3.59 (t, J=2.5 Hz, 1H), 3.27 (s, 6H), 2.16-1.99 (m, 9H).

##STR00360##

Compound 336: (2-{[(2R,3R,4R,5S,6R)-4,5-bis(acetyloxy)-6-[(acetyloxy)methyl]-3-acetamidooxan-2-yl]oxy}ethyl)(fluoromethyl)dimethylazanium

[1043] This compound may be synthesized according to the experimental procedure described for Compound 311.

##STR00361##

Compound 337: (2-{[(2R,3R,4R,5S,6R)-3-acetamido-4,5-dihydroxy-6-(hydroxymethyl)oxan-2-yl]oxy}ethyl)(fluoromethyl)dimethylazanium

[1044] This compound may be synthesized according to the experimental procedure described for Compound 317.

##STR00362##

Compound 338: (fluoromethyl)dimethyl(2-{[(2R,3R,4S,5S,6S)-3,4,5-tris(acetyloxy)-6-carboxyoxan-2-yl]oxy}ethyl)azanium

[1045] This compound may be synthesized according to the experimental procedure described for Compound 321.

##STR00363##

Compound 339: (2-{[(2R,3R,4S,5S,6S)-6-carboxy-3,4,5-trihydroxyoxan-2-yl]oxy}ethyl)(fluoromethyl)dimethylazanium

[1046] This compound may be synthesized according to the experimental procedure described for Compound 321, followed by deacetylation following procedure for Compound 317.

##STR00364##

Compound 159: N-(carboxymethyl)-N,N-dimethylprop-2-yn-1-aminium Bromide

[1047] To the mixture of N,N-dimethylprop-2-yn-1-amine (1 g, 12.03 mmol, 1.28 mL, 1 eq) in Acetone (20 mL) was added dropwise 2-bromoacetic acid (1.82 g, 13.11 mmol, 943.97 uL, 1.09 eq), and then the mixture was stirred at 15° C. for 20 min. TLC indicated N,N-dimethylprop-2-yn-1-amine was consumed completely and one new spot formed. The reaction mixture was filtered and filter cake was concentrated under reduced pressure to give a residue. Compound carboxymethyl-dimethyl-prop-2-ynyl-ammonium (700 mg, 3.15 mmol, 26.20% yield, Br—) was obtained as a white solid. .sup.1H NMR (400 MHz, DMSO) δ 4.5 (s, 2H), 4.0 (m, 3H), 3.2 (s, 6H) ppm.

##STR00365##

[1048] Compound J60: N-(2-hydroxyethyl)-N,N-dimethylprop-2-yn-1-aminium Iodide

[1049] To a solution of N,N-dimethylprop-2-yn-1-amine (1 g, 12.03 mmol, 1.28 mL, 1 eq) in THF (3 mL) was added dropwise 2-iodoethanol (2.17 g, 12.63 mmol, 987.35 uL, 1.05 eq) at 0° C. The mixture was warmed to 15° C. for 5 hr. TLC indicated N,N-dimethylprop-2-yn-1-amine was consumed completely and one new spot formed. The reaction was clean according to TLC. The reaction mixture was filtered and filter cake was concentrated under reduced pressure to give a residue. Compound N-(2-hydroxyethyl)-N,N-dimethyl-prop-2-ynyl-ammonium iodide (2 g, 7.84 mmol, 65.17% yield, I—) was obtained as a white solid. .sup.1H NMR (400 MHz, DMSO) δ 5.3 (m, 1H), 4.4 (s, 2H), 4.0 (s, 1H), 3.8 (m, 2H), 3.4 (m, 2H), 3.1 (s, 6H) ppm.

##STR00366##

Compound Z76: 1-(prop-2-yn-1-yl)-1H-imidazole

[1050] Imidazole is reacted with propargyl bromide in THF and diisopropyl ethyl amine. The resulting compound is purified to afford the title compound.

##STR00367##

Compound A51: 4-carboxy-N,N-dimethyl-N-(prop-2-yn-1-yl)butan-1-aminium Bromide

[1051] To a solution of N, N-dimethylprop-2-yn-1-amine (300 mg, 3.61 mmol, 382.65 uL, 1.08 eq) in acetone (5 mL) was added 5-bromopentanoic acid (604.89 mg, 3.34 mmol, 1 eq) at 15° C. The mixture was stirred at 60° C. for 16 h. The reaction mixture was concentrated under reduced pressure. The residue was diluted with H.sub.2O (10 mL) and extracted with ethyl acetate (10 mL*2). The combined water layer was freeze-dried to get 4-carboxy-N, N-dimethyl-N-(prop-2-yn-1-yl) butan-1-aminium bromide (89 mg, 336.92 umol, 10.08% yield) as a yellow solid. .sup.1H NMR (400 MHz, D2O) δ 4.22 brS, 2H), 3.4 (m, 2H), 3.1 s (1H), 3.05 (brs, 6H), 2.4 (t, 2H), 1.8 (m, 2H), 1.6 m, 2H).

##STR00368##

[1052] Compound B52: 2-carboxy-N,N-dimethyl-N-(prop-2-yn-1-yl)prop-2-en-1-aminium Bromide

[1053] To a solution of N,N-dimethylprop-2-yn-1-amine (200 mg, 2.41 mmol, 255.10 uL, 1 eq) in ACN (10 mL) was added dropwise 2-(bromomethyl)prop-2-enoic acid (417.49 mg, 2.53 mmol, 1.05 eq) at 15° C. After addition the mixture was stirred at 15° C. for 12 h. The white precipitate was formed and TLC showed the starting reactant was consumed. The mixture reaction was filtered to obtain white solid. Compound 2-carboxy-N,N-dimethyl-N-(prop-2-yn-1-yl)prop-2-en-1-aminium bromide (103 mg, 551.09 umol, 22.87% yield) was obtained as a white solid. LCMS: (M+): 168.1 1H NMR (400 MHz, Deuterium Oxide) δ 6.79 (s, 1H), 6.30 (s, 1H), 4.19 (s, 2H), 4.10 (d, J=2.6 Hz, 2H), 3.14 (t, J=2.6 Hz, 1H), 2.99 (s, 6H).

##STR00369##

Compound AA 77: (1-(prop-2-yn-1-yl)-1H-imidazol-2-yl)methanol

[1054] 1-(prop-2-yn-1-yl)-1H-imidazole is dissolved in THF and cooled to −78° C. n-Butyl lithium is added followed by paraformaldehyde. The resulting mixture is stirred at −78 then quenched with water followed by ethyl acetate. The material was purified to afford the title compound.

##STR00370##

Compound AB78: 3-methyl-1-(prop-2-yn-1-yl)-1H-imidazol-3-ium Iodide

[1055] Imidazole is reacted with propargyl bromide in THF and diisopropyl ethyl amine. The resulting compound is purified to afford 1-(prop-2-yn-1-yl)-1H-imidazole. This material is taken up in diethyl ether and treated with dropwise addition of methyl iodide. The resulting precipitate is collected and dried and recrystallized to afford the title compound.

##STR00371##

Compound AC79: (1-(prop-2-yn-1-yl)-1H-imidazol-5-yl)methanol

[1056] (1H-imidazol-4-yl)methanol is treated with propargyl bromide and diisopropyl ethyl amine in THF overnight. The resulting material is purified to afford the title compound.

##STR00372##

Compound AE81: N-(3-(hydroxyhydrophosphoryl)propyl)-N,N-dimethylprop-2-yn-1-aminium

[1057] (3-(dimethylamino)propyl)phosphinic acid is taken up in THF and treated with drop wise addition of propargyl iodide. The resulting precipitate is collected and dried and recrystallized to afford the title compound.

##STR00373##

Compound AG83: N,N-dimethyl-N-(3-phosphonopropyl)prop-2-yn-1-aminium Iodide

[1058] (3-(dimethylamino)propyl)phosphonic acid is taken up in THF and treated with drop wise addition of propargyl iodide. The resulting precipitate is collected and dried and recrystallized to afford the title compound.

##STR00374##

Compound AI85: N,N-dimethyl-N-(3-sulfopropyl)prop-2-yn-1-aminium Iodide

[1059] 3-(dimethylamino)propane-1-sulfonic acid is taken up in THF and treated with drop wise addition of propargyl iodide. The resulting precipitate is collected and dried and recrystallized to afford the title compound.

Example 2: Cell-Based Assay for TMA Production from a Native Producer

[1060] Trimethylamine lyase (CutC) from Clostridium sporogenes ATCC 15579 (protein EDU36695.1) and Proteus mirabilis strain HI4320 (protein WP_012368484.1) was identified as an enzyme that converts choline into trimethylamine. In each bacterium, CutC was modified into its activated form by a cognate activating enzyme, CutD (protein EDU36696.1 for C. sporogenes; protein WP_004249185.1 for P. mirabilis). We have also identified additional enzymes that convert carnitine into trimethylamine. Trimethylamine may subsequently be converted by human enzymes such as flavin monooxygenase 3 to trimethylamine N-oxide, which might be associated with negative health consequences in humans.

[1061] These examples of bacterial enzymes producing metabolites negatively associated with human health are not intended to be limiting.

[1062] A compound's ability to inhibit CutC from Proteus mirabilis and C. sporogenes from converting choline into trimethylamine was tested in cells as described. A BHI blood plate was inoculated with a frozen stock of Proteus mirabilis or Clostridium sporogenes under anaerobic conditions. A single colony was isolated from the plate and inoculated into 5 mL BHI supplemented with 1 mM choline under anaerobic conditions. The liquid cultures were grown overnight at 37° C.

[1063] To prepare each sample, 300 μL of the saturated culture was then inoculated into a 30 mL dilution blank under anaerobic conditions. 1 mM (trimethyl-d.sub.9)-choline and a sufficient amount of a compound to obtain a final concentration of 10 μM were added to the inoculated dilution blank. Optionally, the IC.sub.50 of an inhibitor may instead be determined by adding an inhibitor across a range of concentrations, for example, 0, 0.001, 0.01, 0.1, 1, and 10 μM. The mixture was incubated at 37° C. for 5 h. Chilled 85% acetonitrile, 15% methanol and 0.2% formic acid was added to the reaction and the mixture was centrifuged. After centrifugation, the samples were analyzed by LCMS to determine the amount of d.sub.9-TMA produced. Results are shown in Table 1.

TABLE-US-00001 TABLE 1 IC.sub.50 Standard Number Whole Deviation of assay Com- cell C. Average IC.sub.50 for IC.sub.50 in runs for pound sporogene Whole cell P. P. P. No. s (uM)* mirabilis (uM) mirabilis mirabilis 127 10.00 5.01 7.05 2 B52  0.01 0.03 0.04 3 Z76 10.00 4.04 5.26 3 128 10.00 10.00 N/A 1 159  0.76 3.51 5.63 3 159 10.00 0.28 0.37 2 129 10.00 10.00 0.00 2 J60  0.00 0.18 0.13 7 130 10.00 3.47 1.71 2 A51  0.93 1.10 1.39 2 107 10.00 10.00 0.00 2 132 10.00 10.00 N/A 1 133  0.44 0.66 N/A 1 134 10.00 10.00 N/A 1 110 10.00 10.00 N/A 1 111 — 0.14 0.12 2 137 10.00 10.00 N/A 1 117 10.00 10.00 N/A 1 116 10.00 10.00 N/A 1 109  0.42 0.62 0.58 6 108  2.46 2.19 N/A 1 113  1.25 0.70 N/A 1 114  1.63 2.08 N/A 1 115  9.85 2.07 N/A 1 116 10.00 10.00 N/A 1 138  5.35 1.05 0.28 3 112  0.28 0.55 0.30 2 140 10.00 10.00 N/A 1 141 10.00 5.25 4.20 3 111  0.22 0.27 N/A 1 143 10.00 10.00 0.00 2 144  0.70 0.58 N/A 1 145  1.38 10.00 0.00 2 146  1.50 10.00 N/A 1 147 10.00 0.00 2 148 10.00 N/A 1 149 10.00 N/A 1 Prodrug 7 3.16 N/A 1 Prodrug 19 1.730 N/A 1 Prodrug 8 10.00 N/A 1 Prodrug 69 10.00 N/A 1 Prodrug 73 10.00 N/A 1 Prodrug 70 10.00 N/A 1 Prodrug 71 10.00 N/A 1 Prodrug 72 10.00 N/A 1 Prodrug 73 10.00 N/A 1 Prodrug 74 10.00 N/A 1 Prodrug 106 10.00 N/A 1 Prodrug 78 10.00 N/A 1 Prodrug 78 10.00 N/A 1 Prodrug 77 10.00 N/A 1 Prodrug 79 10.00 N/A 1 Prodrug 79 10.00 N/A 1 Prodrug 80 10.00 N/A 1 Prodrug 86 10.00 N/A 1 Prodrug 98 0.00 N/A 1 Prodrug 99 0.00 N/A 1 151 0.89 1.16 2 152 10.00 N/A 1 153 0.02 0.01 2 159 10.00 N/A 1 160 10.00 N/A 1 161 10.00 N/A 1 162 0.29 0.23 2 125 0.19 0.02 2 164 10.00 N/A 1 165 2.13 N/A 1 J60 0.09 0.02 2 B52 0.04 N/A 1 166 1.26 N/A 1 167 10.00 N/A 1 168 0.87 N/A 1 169 10.00 N/A 1 171 10.00 N/A 1 172 10.00 N/A 1 173 10.00 N/A 1 174 0.17 N/A 1 175 0.17 N/A 1 154 10.00 N/A 1 155 2.58 N/A 1 156 10.00 N/A 1 157 10.00 N/A 1 176 10.00 N/A 1 177 7.57 N/A 1 178 10.00 N/A 1 179 0.11 N/A 1 180 >10 N/A 1 181 >10 N/A 1 182 3.150 N/A 1 183 >10 N/A 1 184 >10 N/A 1 185 0.235 N/A 1 186 >10 N/A 1 187 0.763 N/A 1 188 >10 N/A 1 189 >10 N/A 1 190 1.280 N/A 1 191 2.165 N/A 1 192 >10 N/A 1 193 1.375 N/A 1 194 >10 N/A 1 195 >10 N/A 1 196 7.780 N/A 1 197 4.970 N/A 1 198 >10 N/A 1 199 >10 N/A 1 200 2.740 N/A 1 201 >10 N/A 1 202 0.242 N/A 1 203 1.720 N/A 1 204 0.660 N/A 1 205 8.300 N/A 1 206 0.069 N/A 1 207 0.310 N/A 1 208 >10 N/A 1 209 6.800 N/A 1 210 >10 N/A 1 211 0.211 N/A 1 212 1.990 N/A 1 213 0.180 N/A 1 214 3.870 N/A 1 215 0.409 N/A 1 216 >10 N/A 1 217 0.771 N/A 1 218 >10 N/A 1 219 >10 N/A 1 220 >10 N/A 1 221 >10 N/A 1 222 >10 N/A 1 223 2.280 N/A 1 224 0.223 0.102 2 225 0.629 0.095 2 226 0.244 0.007 2 227 >10 N/A 1 228 >10 N/A 2 229 >100.000 N/A 1 IC.sub.50 values were given the value “10” when a best fit ICso line could not calculated or when the % TMA remaining at the highest concentration of inhibitor (10 μM) was higher than 15% * C. sporogenes was run once

Example 3: Cell-Based Assay with Bacterial Enzymes

[1064] A vector may be designed to express an enzyme, such as those identified in Example 2, in its wild-type form or a catalytically inactive version. The vector may be transformed into a heterologous host, for example an E. coli expression strain. Alternatively, a strain natively encoding one of the enzymes identified in Example 2 may be procured.

[1065] The heterologous host (containing the wild-type enzyme, a catalytically inert version, or the empty vector) or the native strain may be grown to a desirable level.

[1066] The native substrate or an appropriately-chosen substrate analog may be added to the bacterial growth or uninoculated medium. Optionally, an inhibitor might be added to prevent conversion of substrate to product. The mixture may be incubated for a designated period of time.

[1067] After incubation, the mixture may be rendered compatible with LCMS analysis via addition of organic solvent or lyophilization followed by dissolution in organic solvent. The amount of residual starting material and of the product may be quantified by LCMS analysis.

[1068] For example, the following cell-based assay for TMA production from a heterologous host is a representative example of the above-described assays. E. coli strains capable of co-expressing full-length WT CutC and CutD, the catalytically inert G821A CutC and CutD, or empty pET28 can be produced by transforming the aforementioned plasmids into E. coli BL21(DE3) by heat shocking the cells. Transformants may be selected by growing on LB plates supplemented with kanamycin (50 μg/mL).

[1069] A single colony may be isolated from the plate and inoculated into 10 mL LB supplemented with kanamycin (50 μg/mL). The liquid cultures may be grown overnight at 37° C. with rocking on a nutator.

[1070] 500 μL of the saturated overnight culture may be inoculated into 30 mL LB-Kan50 under anaerobic conditions. The cultures may be grown at 37° C. until the optical density at 600 nm reached 0.6 A.U. Then, IPTG may be added to a final concentration of 0.5 mM.

[1071] To 1 mL of induced culture or a media blank may be added 1 mM (trimethyl-d.sub.9)-choline. A compound to be tested for inhibition of CutC may also be added at this time at a fixed final concentration of, for example, 10 μM. Optionally, the IC.sub.50 of an inhibitor may instead be determined by adding an inhibitor across a range of concentrations, for example, 0, 0.001, 0.01, 0.1, 1, and 10 μM. The mixture may be incubated at 37° C. for 8 h. Chilled 85% acetonitrile, 15% methanol and 0.2% formic acid may be added to the reaction. After centrifugation, the samples may be analyzed by LCMS, for example, to determine the amount of d.sub.9-TMA that has been produced from the d.sub.9-choline.

[1072] Another representative example of the above-described assays involves assaying active and inactive versions of the proteins listed above and in Example 2 for their ability to react with their native substrate or an appropriately chosen substrate analog in a cell-based assay. In each instance, this incubation may be carried out in the presence of an inhibitor. LCMS may be used to quantify the amount of product that is produced from the native substrate or the isotope-labeled substrate.

Example 4: Determination of Inhibitor Effect on Bacterial Growth

[1073] To verify that the inhibition of d.sub.9-TMA (such as assayed by a method disclosed in Example 3 above) is due to inhibition of enzymatic activity and not cell death, the lag phase of the growth curve from either P. mirabilis or C. sporogenes was measured.

[1074] A single colony was isolated from the plate and inoculated into 5 mL BHI supplemented with 1 mM choline under anaerobic conditions. The liquid cultures were grown overnight at 37° C.

[1075] 300 μL of the saturated overnight culture was inoculated into 30 mL BHI under anaerobic conditions supplemented in the presence of 10 μM of compound to be tested. The growth of the bacteria was monitored with a spectrophotometer at an Optical Density of 600 nm for 24 hours. Results are shown in Table 2.

TABLE-US-00002 TABLE 2 Avg Lag Time Avg Lag Time (% compared (% compared to control) [C. Number of SD of C. to control) [P. Number of SD of P. sporogenes, C.sporogenes sporogenes mirabilis, P.mirabilis mirabilis Cmpd No. Conc: 10 μM] Replicates Lag Time Conc: 10 μM] Replicates Lag Time B52 102.55 2 17.04 100.40 2 10.32 Z76 100.20 1 N/A 92.30 1 N/A J60 98.30 2 2.26 101.93 3 7.29 H58 N/A 0 N/A 106.10 1 N/A 109 92.70 1 N/A 99.40 2 4.38 138 102.60 1 N/A 82.80 1 N/A 112 97.20 1 N/A 97.20 1 N/A 145 107.70 1 N/A 107.10 1 N/A 153 N/A 0 N/A 98.20 1 N/A X74 N/A 0 N/A 98.40 1 N/A 78 93.8 109.4 79 83.5 103.3 80 95.9 95.3 228 108.3 112.2 317 83 111.1 318 83.1 103 229 96.9 112.4

Example 5: Inhibition of Enzymatic Activity in Fecal Matter

[1076] Inhibition of enzymatic activity in fecal matter was determined by adding a substrate (d9-choline, 1 mM final concentration) and a compound in a range of concentrations in order to determine an IC.sub.50. After incubation for a designated 16 hours at 37° C. in anaerobic conditions, samples were be prepared for LCMS analysis, and the amount of d9-TMA produced was determined using LCMS analysis as in the Examples above. Results are shown in Table 3.

TABLE-US-00003 TABLE 3 Compound IC.sub.50 Number of Runs  78 0.39 6  79 1.17 6  80 0.38 5 317 0.43 3

Example 6: Acute Mouse Model

[1077] The following acute mouse model may be used to determine inhibition of the CutC/D enzyme and thus the inhibition of the formation of trimethylamine (TMA) from choline.

[1078] On day 1, male C57BL/6 mice were given a chemically defined diet (Tekland Global Rodent Diet 2018) containing 1.0% choline (g/g) for the duration of the experiment. Concurrently on day 1, mice were orally gavaged with 200 μL of compounds of interest (formulated in water). Mice were orally gavaged with the compounds and vehicle control again on day 2 and day 3 at a fixed time. Twenty hours after the last gavage, mice were sacrificed, and plasma was collected and prepared for detection of trimethylamine-N-oxide (TMAO) via liquid chromatography with on-line tandem mass spectrometry (LC-MS/MS). Food consumption and mouse weight were measured each day.

[1079] Plasma samples were prepared for LCMS as follows: An aliquot of 20 μL plasma sample was protein precipitated with 200 μL internal standard solution (100 ng/mL Labetalol & 100 ng/mL Tolbutamide & 100 ng/mL Diclofenac in acetonitrile, the mixture was vortex-mixed and centrifuged at 4000 rpm for 15 min, 4° C. An aliquot of 100 μL supernatant was transferred to the sample plate for LCMS injection. 1 μL of sample was injected onto an LC column for TMAO LCMS analysis.

[1080] Concentrations of TMAO were determined with a calibration curve ranging 0.05-100 μM for d9-TMAO in control plasma. Results are shown in Tables 3a-3e.

TABLE-US-00004 TABLE 4a Mean % change plasma from vehicle TMAO control + Diet + Compound (μM) choline diet Normal Diet + Vehicle Control  4.56 −86.75 Choline Diet + Vehicle Control 34.39   0.00 Choline diet + Compound I59 (100 mg/kg) 10.85 −68.44 Choline diet + Compound 19 (333 mg/kg)  5.40 −84.29 Choline diet + Compound J60 (10 mg/kg) 31.06 −28.46 Choline diet + Compound J60 (33 mg/kg) 17.20 −60.39 Choline diet + Compound J60 (100 mg/kg)  5.52 −87.29 Choline diet + Compound 7 (358 mg/kg)  6.90 −79.95

TABLE-US-00005 TABLE 4b Mean % change plasma from vehicle TMAO control + Diet + Compound (μM) choline diet normal diet + vehicle  2.55 −89.74 choline diet + vehicle 24.9    0.00 choline diet + Compound J60 (10 mg/kg) 24.7   −0.69 choline diet + Compound B52 (10 mg/kg) 30.0   20.76 choline diet + Compound 141 (10 mg/kg)  2.92 −88.23 choline diet + Compound J60 (10 mg/kg) 27.4   10.16

TABLE-US-00006 TABLE 4c Mean % change plasma from vehicle TMAO control + Diet + Compound (μM) choline diet normal diet +vehicle  2.17 −89.66 choline diet + vehicle 21.0    0.00 choline diet + Compound J60 (100 mg/kg)  5.41 −74.22 choline diet + Compound 7 (35.8 mg/kg) 20.4   −2.97 choline diet + Compound 7 (107 mg/kg) 15.8  −24.83 choline diet + Compound 7 (358 mg/kg)  6.41 −69.46

TABLE-US-00007 TABLE 4d Mean % change plasma from vehicle TMAO control + Diet + Compound (μM) choline diet normal diet + vehicle  2.49 −87 choline diet + vehicle 18.8    0 choline diet + Compound J60 (100 mg/kg)  6.00 −68 choline diet + Compound 58 (10 mg/kg) 10.1  −46 choline diet + Compound 310 (33.8 mg/kg) 2.3 −88 choline diet + Compound 312 (35.4 mg/kg) 11.69 −38 choline diet + Compound 309 (32.6 mg/kg) 1.9 −90 choline diet + Compound 311 (37.0 mg/kg) 1.8 −90 choline diet + Compound 8 (100 mg/kg) 12.5  −33 choline diet + Compound 10 (100 mg/kg) 14.9  −21 choline diet + Compound 9 (100 mg/kg) 33.91  80

TABLE-US-00008 TABLE 4e Mean % change plasma from vehicle TMAO control + Diet + Compound (μM) choline diet normal diet + vehicle  1.90 −93 choline diet + vehicle 26.4    0 choline diet + Compound J60 (100 mg/kg)  3.81 −86 choline diet + Compound 185 (123.7 mg/kg) 22.2  −16

TABLE-US-00009 TABLE 4f Mean % change plasma from vehicle TMAO control + Diet + Compound (μM) choline diet normal diet + vehicle  3.91 −88 choline diet + vehicle 32.3    0 choline diet + Compound J60 (100 mg/kg)  6.20 −81 choline diet + Compound 98 (36.6 mg/kg) 10.1  −69 choline diet + Compound 98 (11.0 mg/kg) 34.3    6 choline diet + Compound 98 (3.70 mg/kg) 29.7   −8 choline diet + Compound 309 (37.2 mg/kg)  5.45 −83 choline diet + Compound 309 (11.0 mg/kg) 15.4  −52 choline diet + Compound 309 (3.70 mg/kg) 35.2    9 choline diet + Compound 311 (35.3 mg/kg) 11.23 −65 choline diet + Compound 311 (10.6 mg/kg) 17.4  −46 choline diet + Compound 311(3.5 mg/kg) 20.22 −37

TABLE-US-00010 TABLE 4g Mean % change plasma from vehicle TMAO control + Diet + Compound (μM) choline diet normal diet + vehicle  3.04 −89 choline diet + vehicle 27.8    0 choline diet + Compound J60 (100 mg/kg) 13.1  −53 choline diet + Compound 125 (97.7 mg/kg) 32.8   18 choline diet + Compound 125 (9.8 mg/kg) 26.2   −6 choline diet + Compound 86 (112 mg/kg)  4.72 −83 choline diet + Compound 86 (37.3 mg/kg) 18.6  −33 choline diet + Compound 109 (116 mg/kg)  2.71 −90 choline diet + Compound 151 (116 mg/kg) 29.4    6

TABLE-US-00011 TABLE 4h Mean % change plasma from vehicle TMAO control + Diet + Compound (μM) choline diet normal diet + vehicle  3.07 −93 choline diet + vehicle 44.4    0 choline diet + Compound J60 (100 mg/kg)  8.72 −80 choline diet + Compound 315 (226.5 mg/kg)  8.96 −80 choline diet + Compound 315 (67.9 mg/kg) 26.6  −40

[1081] The data in Tables 4a-4h show that compounds of the present disclosure can inhibit the production of trimethyl amine and thus can be useful in treating diseases in which CutC and the production of trimethylamine are implications (e.g. treating trimethylaminuria).

Example 7: Compound Efficacy in a Preclinical Chronic Kidney Disease Mouse Model

[1082] The following preclinical model has been used to determine the effect of TMA/TMAO on chronic kidney disease (CKD) (Tang et al 2015; Circulation Research, 116(3): 448-455.). C57BL/6 mice may be fed a chemically defined diet with or without 1.0% choline (g/g) for 16 weeks. Earlier time points of compound intervention may define the potential to prevent the progression of kidney disease. Later time points of intervention may determine the ability of the compounds to treat an established kidney disease phenotype. The weight and food consumption of each mouse will be monitored at regular intervals throughout the study.

[1083] All mice may be sacrificed after 16 weeks of defined diet and compound intervention. As described in the acute mouse model, plasma may be collected and prepared for detection of TMA and TMAO via LCMS. To assess the effect of compounds on renal function, protein in urine may be quantified and blood urea nitrogen (BUN) levels may be measured in the blood. To assess the effect of compounds on kidney injury, kidneys may be collected, and levels of fibrosis may be quantified via histological examination as well as expression of markers of kidney damage and inflammation. Additionally, aortas will be collected and examined for the presence of vascular injury via protein markers of vascular damage.

[1084] For histological examination, kidneys may be fixed and embedded. Fibrosis may be quantified via collagen deposition using the Mason trichome staining procedure (Tang et al 2015, supra, and Sun et al 2017; Biochem. Biophys. Res. Commun. 493(2): 964-970).

[1085] Expression of kidney damage markers, such as pSMAD3, kidney injury molecule (KIM)-1, TNFα, TL-1β, neutrophil gelatinase-associated lipocalin (NGAL), plasma cystatin C, urine albumin, and NOX-4, may be determined via Western blot analysis of homogenized kidneys. Expression of vascular damage markers, such as ICAM, VCAM, TNFα, and IL-1β, may be determined via the MesoScale Discovery instrument.

Example 8: Release of Active Agents from Conjugates

[1086] Sprague-Dawley rats (three rats per compound) were treated with a single intravenous (IV) 1 mg/kg dose or orally (PO) a single 10 mg/kg dose. Plasma concentrations of the conjugates were determined by LCMS at 5 minutes, 15 minutes, 30 minutes, 1 hour, 2 hours, 4 hours, 8 and 24 hours post compound administration. For all conjugates, PO doses displayed substantially lower plasma exposure compared to IV administered rats and therefor all conjugates have a low bioavailability (% F) as shown in Table 5 below. In addition, the plasma exposure and bioavailability of the active agent of was lower when dosed as a conjugate instead of the active agent alone.

TABLE-US-00012 TABLE 5 Compound Compound IV C.sub.max PO C.sub.max Dosed Measured (nmol/L) (nmol/L) % F 78 78  8310 130 0.99 78 Active     6 70 1.91 agent of 78 Active Active 17200 970 4.20 agent of 78 agent of 78 79 79 13600 260 2.16 79 Active     1 50 1.60 agent of 79 Active Active 41700 2360 5.99 agent of 79 agent of 79 98 98 23600 306 0.72 Active Active   134 142 3.78 agent of 98 agent of 98 80 80  7680 100 0.68 80 Active     0 20 1.5  agent of 80 Active Active 36900 1000 2.16 agent of 80 agent of 80  7  7 19700 16.2 1.1 

Example 9. Targeted Delivery of Active Agents by Non-Systemic Conjugates

[1087] To confirm that the active agent component of the conjugate is released in the gastrointestinal tract, a pharmacokinetic study was conducted in male C57B3L/6 mice examining conjugate and active agent of concentrations in both the colon contents and plasma after oral dose of conjugate. Conjugate and active agent of were quantified by LCMS at 0 (pre-dose), 0.25, 0.5, 1, 2, 4, 6, 8, 12, and 24 hours post-dose of compound. The results are shown in Table 6 below. The active agents of the conjugate was at least 100,000-fold more concentrated in the colon versus the plasma, suggesting non-systemic, gut restrictive delivery of the active agent by the conjugate.

TABLE-US-00013 TABLE 6 Colon Plasma Compound Compound C.sub.max C.sub.max Dosed Measured (μmol/L) (μmol/L) 78 78 0.07 Not detected 78 Active agent 7077 0.09 of 78 79 79 Not 0.005 detected 79 Active agent 4081 0.16 of 79 98 98 21.83 1.3 98 Active agent 5522 0.40 of 98 80 80 0.13 Not detected 80 Active agent 4759 Not of 80 detected

Example 10: In Vitro Stability Studies of Conjugates

Assay 1. Stability of Conjugates in Simulated Gastric Fluid (SGF)

[1088] This assay was used to assess the stability of a conjugate in a stomach.

[1089] Medium was prepared by dissolving 2 g of sodium chloride in 0.6 L in ultrapure water (MilliQ®, Millipore Sigma, Darmstadt, Germany). The pH was adjusted to 1.6 with 1N hydrochloric acid, and the volume was then adjusted to 1 L with purified water.

[1090] 60 mg FaSSIF powder (Biorelevant™, London, UK) were dissolved in 500 mL buffer (above). Pepsin was added (0.1 mg/mL) (Millipore Sigma, Darmstadt, Germany), and the solution was stirred. The resulting SGF media were used fresh for each experiment.

[1091] Test compounds were dissolved in DMSO stock to 1 mM. An aliquot of the DMSO stock solution was removed and diluted in the SGF Media in 15 mL falcon tubes to generate a total compound concentration of 1 μM. A 1 mL aliquot was immediately removed and diluted once with 1 volume of acetonitrile for T0 timepoint. The mixture was sealed and mixed at 37° C. in an incubator. Aliquots (1 mL) were removed at regular intervals and immediately quenched by the addition of 1 volume of acetonitrile. The resulting samples were analyzed by LCMS to determine degradation rates in SGF. The results for percent of compound remaining at 1 hour are shown in Table 7.

Assay 2. Stability of Conjugates in Simulated Intestinal Fluid (SIF).

[1092] This assay was used to assess the stability of a conjugate in a small intestine.

[1093] Phosphate buffer was prepared by dissolving 0.42 g of sodium hydroxide pellets and 3.95 g of monobasic sodium phosphate monohydrate and 6.19 g of sodium chloride in ultrapure water (MilliQ®, Millipore Sigma, Darmstadt, Germany). The pH was adjusted to 6.7 using aq. HCl and aq. NaOH, as necessary, and the solution was diluted with ultrapure water to produce 1 L of the pH 6.7 buffer.

[1094] 112 mg FaSSIF powder (Biorelevant™, London, UK) was dissolved in 50 mL of the pH 6.7 buffer. 2 to 3 mL of the resulting solution were then added to 500 mg pancreatin (Millipore Sigma, Darmstadt, Germany). The resulting mixture was agitated by finger tapping the vessel containing the mixture until milky suspension formed. At this time, the remainder of the 50 mL FaSSiF/pH 6.7 buffer solution was added. The resulting suspension was flipped upside down 10 times to produce SIF, which was used fresh.

[1095] Test compounds were dissolved in DMSO stock to 1 mM. An aliquot of the DMSO stock solution was removed and diluted in the SIF media in 15 mL falcon tubes to produce a mixture with a tested compound concentration of 1 μM. A 1 mL aliquot was immediately removed and diluted once with 1 volume of acetonitrile for T0 timepoint. The mixture was sealed and agitated at 37° C. in an incubator. Aliquots (1 mL) were removed at regular intervals and immediately quenched by the addition of 1 volume of acetonitrile. The resulting samples were analyzed by LCMS to determine degradation rates. The results for percent of compound remaining at 4 hours are shown in Table 7.

Assay 3. In Vitro Colonic Material Stability Assay.

[1096] This assay was used to assess the stability of a conjugate in a large intestine.

[1097] All experiments were performed in an anaerobic chamber containing 90% nitrogen, 5% hydrogen and 5% carbon dioxide. Colonic material was resuspended as a slurry (15% w/v final concentration) in pre-reduced, anaerobically sterilized dilution blanks (Anaerobe Systems AS-908). The colonic material was then inoculated into 96 well plates containing YCFAC media (Anaerobe Systems AS-680, 6.7 μL slurry into 1 mL total media). A compound was added to each individual well to reach a final analyte concentration of 1 μM, and the material was mixed by pipetting. Each sample was removed after set timepoints (0, 120, 240, 480, 1440, 2880 minutes after initiation of the assay), quenched with acetonitrile containing internal standard, and analyzed by LCMS. Results are shown in Table 7.

TABLE-US-00014 TABLE 7 Assay 3- Colonic Assay 1- Assay 2- Material FaSSGF FaSSIF Stability Stability: Stability: Assay % % Remaining % Remaining Remaining Compound at 1 hr at 4 hr at 24 hr   1  84.79  30.79 80   7  85.06 121.01 56   7 100.35  93.47 51  19 101.36  11.06 2.5  19  85.52   1.52 0.18 303  60.49   0.03 0.16 304 114.28  13.44 0.94  70  98.36  28.06 47 305  92.73   0.25 0.04  72 102.71 — 110  73 107.49  62.73 44  74 107.3  121.56 99   8 105.93 113.28 17 315 109.96 120.35 68 310  97.99 115.11 55 309  97.94  94.72 16 311 104.43  97.27 33  80 145.32  99.34 25  86 — — 56 317  96.24  82.34

Example 11: Caco-2 Permeability Assay

[1098] Preparation of Caco-2 Cells: 50 μL and 25 mL of cell culture medium were added to each well of the Transwell insert and reservoir, respectively. The HTS transwell plates were incubated at 37° C., 5% CO.sub.2 for 1 hour before cell seeding. Caco-2 cells were diluted to 6.86×105 cells/mL with culture medium and 50 μL of cell suspension were dispensed into the filter well of the 96-well HTS Transwell plate. Cells were cultivated for 14-18 days in a cell culture incubator at 37° C., 5% CO.sub.2, 95% relative humidity. Cell culture medium was replaced every other day, beginning no later than 24 hours after initial plating.

[1099] Assessment of Cell Monolayer Integrity: Medium was removed from the reservoir and each Transwell insert and replaced with prewarmed fresh culture medium. Transepithelial electrical resistance (TEER) across the monolayer was measured using Millicell Epithelial Volt-Ohm measuring system (Millipore, USA). The Plate was returned to the incubator once the measurement was done. The TEER value was calculated according to the following equation:

TEER measurement (ohms)×Area of membrane (cm2)=TEER value (ohm.Math.cm2)

TEER value should be greater than 230 ohm.Math.cm2, which indicates the well-qualified Caco-2 monolayer.

[1100] Preparation of Solutions: 2 mM stock solutions in DMSO of control compounds were prepared and diluted with HBSS (10 mM HEPES, pH 7.4) to get 10 μM working solution. 0.2 mM stock solutions of test compounds in DMSO were prepared and diluted with HBSS (10 mM HEPES, pH 7.4 with 0.5% BSA) to get 1 μM working solution. Metoprolol, erythromycin and cimetidine were used as control compounds.

[1101] Performing the Drug Transport Assay.

[1102] The Caco-2 plate was removed from the incubator. The monolayer was washed twice with pre-warmed HBSS (10 mM HEPES, pH 7.4). The plate was incubated at 37° C. for 30 minutes. To determine the rate of drug transport in the apical to basolateral direction, 125 μL of the working solution was added to the Transwell insert (apical compartment). A 50 μL sample was transferred immediately from the apical compartment to 200 μL of acetonitrile containing IS (100 nM alprazolam, 200 nM Caffeine and 100 nM tolbutamide) in a new 96-well plate as the initial donor sample (A-B) and it was vortexed at 1000 rpm for 10 minutes. The wells in the receiver plate (basolateral compartment) were filled with 235 μL of transport buffer. To determine the rate of drug transport in the basolateral to apical direction, 285 μL of the working solution were added to the receiver plate wells (basolateral compartment). A 50 μL sample was transferred immediately from the basolateral compartment to 200 μL of acetonitrile containing IS (100 nM alprazolam, 200 nM Caffeine and 100 nM tolbutamide) in a new 96-well plate as the initial donor sample (B-A) and it was vortexed at 1000 rpm for 10 minutes. The Transwell insert (apical compartment) was filled with 75 μL of transport buffer. The apical to basolateral direction and the basolateral to apical direction need to be done at the same time. The plates were incubated at 37° C. for 2 hours. At the end of the incubation, 50 μL samples from donor sides (apical compartment for Ap.fwdarw.Bl flux, and basolateral compartment for B1.fwdarw.Ap) and receiver sides (basolateral compartment for Ap.fwdarw.Bl flux, and apical compartment for B1.fwdarw.Ap) were transferred to wells of a new 96-well plate, followed by the addition of 4 volume of acetonitrile containing IS (100 nM alprazolam, 200 nM Caffeine and 100 nM tolbutamide). Samples were vortexed for 10 minutes, 50 μL samples were transferred to wells of a new 96-well plate, followed by the addition of 50 μL Hepes and 200 μL IS. All samples were vortexed for 10 minutes, and then centrifuged at 3,220 g for 40 minutes. An aliquot of 150 μL of the supernatant was mixed with an appropriate volume of ultra-pure water before LC-MS/MS analysis.

[1103] Data Analysis

[1104] All calculations were carried out using Microsoft Excel. Peak areas were determined from extracted ion chromatograms. Lucifer yellow leakage of monolayer can be calculated using the following equation:

[00001]$\mathrm{LY}\mathrm{Leakage}=\left(\frac{I_{\mathrm{acceptor}}\times 0.3}{I_{\mathrm{acceptor}}\times 0.3+I_{\mathrm{donor}}\times 0.1}\right)\times 100\%$

where I.sub.acceptor is the fluorescence intensity in the acceptor well (0.3 mL), and I.sub.donor is the fluorescence intensity in the donor well (0.1 mL) and expressed as % leakage. Lucifer yellow percentage amount transported values should be less than 1.5%. However, if the lucifer yellow percentage amount transported value for a particular transwell is higher than 1.5 but the determined digoxin P.sub.app in that transwell is qualitatively similar to that determined in the replicate transwells then, based upon the scientific judgement of the responsible scientist, the monolayer is considered acceptable.

[1105] Apparent permeability (P.sub.app) can be calculated for drug transport assays using the following equation:

[00002]$P_{\mathrm{app}}=\frac{\mathrm{dQ}/\mathrm{dt}}{A\times D_o}$

where:

[1106] P.sub.app is apparent permeability (cm/s×10.sup.−6);

[1107] dQ/dt is the rate of drug transport (pmol/second);

[1108] A is the surface area of the membrane (cm.sup.2); and

[1109] D.sub.0 is the initial donor concentration (nM; pmol/cm.sup.3).

[1110] Efflux ratio can be determined using the following equation:

[00003]$\mathrm{Efflux}\mathrm{Ratio}=\frac{P_{\mathrm{app}\left(B-A\right)}}{P_{\mathrm{app}\left(A-B\right)}}$

where

[1111] P.sub.app (B-A) indicates the apparent permeability coefficient in basolateral to apical direction, and

[1112] P.sub.app (A-B) indicates the apparent permeability coefficient in apical to basolateral direction. The results are shown in Table 8 below.

TABLE-US-00015 TABLE 8 Compound Papp A-B Papp B-A No. (*10.sup.−6 cm/s) (*10.sup.−6 cm/s) 159 0.894 1.43   77 0.218 0.337  79 0.214 0.255  80 0.28  0.313 J60 11.07   6.23  107 26.8    31.1      7 0.1   0.15  111 0.8   0.8   109 0.6   0.5   315 0.3   0.3   125 0.466 0.482 310 0.206 0.23  

OTHER EMBODIMENTS

[1113] Various modifications and variations of the described disclosure will be apparent to those skilled in the art without departing from the scope and spirit of the disclosure. Although the disclosure has been described in connection with specific embodiments, it should be understood that the disclosure as claimed should not be unduly limited to such specific embodiments. Indeed, various modifications of the described modes for carrying out the disclosure that are obvious to those skilled in the art are intended to be within the scope of the disclosure.

[1114] Other embodiments are in the claims.