Compound for treating clostridium difficile

Abstract

The invention relates to compounds, compositions and polymers comprising a first component adapted to promote germination of Clostridium difficile (C. diff) and a second component which acts as an antimicrobial agent. Said compounds, compositions and polymers are useful for destroying C. diff where conventional antimicrobial agents are unsuccessful. The compositions can be formulated as coating or materials which actively destroy C. diff which come into contact with it. The germination promotion is induced by bile salts. The invention also relates to the use of such materials as a treatment for C. diff associated diseases and toxic megacolon.

Claims

1. A compound according to general Formula 1
X-Z-M  Formula 1 wherein: X is a bile salt selected from the group consisting of formula 2 and formula 3: ##STR00126## wherein R.sup.1 is a hydroxy group; and R.sup.2 and R.sup.3 are each independently selected from: hydrogen, hydroxy, alkoxy, acyloxy, aryloxy, acrylate or methacrylate groups; Z is a covalent linker; and M has a structure selected from the group consisting of: ##STR00127## ##STR00128##

2. A compound according to claim 1, wherein R.sup.2 and R.sup.3 are each independently selected from hydrogen and hydroxy groups.

3. A compound according to claim 1, wherein Z comprises a first linkage ‘A’ and a second linkage ‘L’.

4. A compound according to claim 3, wherein ‘A’ is selected from: an amino acid, conjugated amino acid or —NR—, wherein R is selected from hydrogen, alkyl, aryl, acyl, alkenyl or alkynyl.

5. A compound according to claim 4, wherein ‘A’ is —NR—, wherein R is hydrogen, alkyl, aryl, acyl, alkenyl or alkynyl.

6. A compound according to claim 5, wherein R is selected from hydrogen or alkyl.

7. A compound according to claim 6, wherein R is a C.sub.1-C.sub.10 alkyl group.

8. A compound according to claim 6, wherein R is a linear alkyl group.

9. A compound according to claim 3, wherein ‘L’ is a covalent linker.

10. A compound according to claim 9, wherein ‘L’ is selected from: alkyl, aryl, acyl, alkenyl, alkynyl, ethers, polyether, esters, polyesters, amides, polyamides, amines or combinations thereof.

11. A compound according to claim 3, wherein ‘L’ is ethylene glycol.

12. A compound according to claim 1, wherein M is an antibiotic for treating gram-negative bacteria or gram-positive bacteria.

13. A composition comprising the compound according to claim 1 and one or more additives.

14. A composition according to claim 13, wherein the additives are amino acids, conjugated amino acid, bile salts and/or antimicrobial agents.

15. A composition according to claim 14, wherein the amino acids are selected from glycine, glutamine, leucine, isoleucine, alanine and taurine.

16. A composition according to claim 15, wherein the amino acid is taurine.

17. A composition according to claim 14, wherein the antimicrobial agent is an antibiotic for treating gram-negative bacteria or an antibiotic for treating gram-positive bacteria.

18. A composition according to claim 14, wherein the antimicrobial agent is a quaternary ammonium salt.

19. A composition according to claim 14, wherein the antimicrobial agent is a compound comprising a group according to general formula 5 ##STR00129## wherein each of R.sup.4, R.sup.5 and R.sup.6 are independently selected from: alkyl, aryl, alkenyl, alkynyl.

20. A composition according to claim 19, wherein at least two of R.sup.4, R.sup.5 and R.sup.6 are covalently bonded together.

21. A composition according to claim 19, wherein two of R.sup.4, R.sup.5 and R.sup.6 are covalently bonded together.

22. A coating composition comprising the compound according to claim 1, further comprising one or more dispersants, solvents or surfactants.

23. A compound according to claim 1, selected from the group consisting of: 3-cholanamidopropyl-benzyl-dimethyl-ammonium iodide; 3-cholanamidopropyl-dimethyl-(2-phenylethyl)ammonium bromide; N-[2-[1-[(4-vinylphenyl)methyl]pyrrolidin-1-ium-1-yl]ethyl]cholanamide chloride; 3-[[(4R)-4-[(3R,10S,13R,17R)-3-hydroxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]pentanoyl]amino]propyl-dimethyl-[(4-vinylphenyl)methyl]ammonium chloride; (4R)—N-[2-(1-benzylpyrrolidin-1-ium-1-yl)ethyl]-4-[(3R,10S,13R,17R)-3-hydroxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]pentanamide bromide; and (4R)-4-[(3R,10S,13R,17R)-3-hydroxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]-N-[2-[1-[(4-vinylphenyl)methyl]pyrrolidin-1-ium-1-yl]ethyl]pentanamide chloride.

24. A compound according to claim 1, selected from the group consisting of: 2-[[(4R)-4-[(3R,10S,13R,17R)-3-hydroxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]pentanoyl]amino]ethyl-dimethyl-[(4-vinylphenyl)methyl]ammonium chloride; N-[2-[1-(3-phenylpropyl)pyrrolidin-1-ium-1-yl]ethyl]cholanamide bromide; and N-[2-(1-benzylpyrrolidin-1-ium-1-yl)ethyl]cholanamide bromide.

25. A method of treating a surface containing C. difficile spores, comprising exposing the surface to a compound of claim 1.

Description

BRIEF DESCRIPTION OF FIGURES

(1) FIG. 1 shows germination activity of HDPE (high density polyethylene)/“T101” (radical initiator 2,5-dimethyl-2,5-bis-(tert-butyl peroxy)hexane) polymer films prepared with and without (control) C109 (compound 129) at 3% (at 1650 and 170° C.) and at 6% (at 160° C.).

(2) FIG. 2 shows germination and antimicrobial efficacy of C109 (compound 129) reactively processed in HDPE (high density polyethylene).

EXAMPLES

Clostridium Difficile Spore Germination Potential of Compound 129

(3) HDPE (high density polyethylene) samples containing the modified bile salt compound 129, reactively processed under different compression moulding conditions, were tested to determine the germination activity. Polymer film samples containing 3% of compound 129 prepared (at molar ratio of 0.005 of peroxide to bile salt) both at 165° C. and 170° C. demonstrated an equal level of germination activity at 2 hours, resulting in a 2.2 Log reduction compared to the HDPE/peroxide T101 control (FIG. 1). However, after 24 and 46 hours, samples processed at the lower temperature demonstrated a slightly higher level of germination activity suggesting that higher temperatures, used under these conditions, give rise to a reduction in the germination activity of the modified bile salt compound 129. Samples prepared similarly but with a higher amount of compound 129 (6%) did not perform well. Table 1 shows that, following subsequent exposure to ethanol, compound 129 prepared at 165° C./170° C. yields a 99.2% reduction in C. Difficile spores. This confirms the germination activity of compound 129 when incorporated into HDPE by reactive processing.

(4) TABLE 1 Germination Activity of Compound 129 Reactively Processed with HDPE after Subsequent Exposure to Ethanol

(5) TABLE-US-00001 Dual Activity of Compound 219 (Germination of C. Difficile Spore and Subsequent Elimination) Log Reduction % Reduction 2 24 96 2 24 96 Polymer hours hours hours hours hours hours compound 129 2.165 0.971 0.802 99.2 87.8 82.0 3% T101 170° C. compound 129 2.165 1.598 1.154 99.2 97.1 92.0 3% T101 165° C. compound 129 0.752 0.546 0.577 79.8 67.6 69.8 3% T101 160° C.

(6) Reactively processed HDPE polymer containing compound 129 is shown to have the ability to both germinate the spores of C. Difficile and reduce the subsequent metabolically active bio-load by 96.9%, see FIG. 2 and Table 2. This exemplifies, therefore, the dual action of compound 129 and its efficacy both as a germinant and an antimicrobial. Reactive processing the polymer at 170° C. changes markedly the dual activity of compound 129 giving a spore reduction of 79.6%.

(7) TABLE 2 Germination and Antimicrobial Efficiency of Compound 129 in HDPE

(8) TABLE-US-00002 Materials and Methods Log Reduction % Reduction 2 24 96 2 24 96 Polymer hours hours hours hours hours hours compound 129 0.747 0.666 0.998 79.6 75.4 88.5 3% T101 170° C. compound 129 1.563 0.940 0.714 96.9 86.9 78.0 3% T101 165° C. compound 129 0.057 0.315 0.936 0.0 44.8 86.8 6% T101 160° C.

(9) Proton NMR spectra were obtained on a Bruker AC 250 instrument operating at 250 MHz as solutions in CDCl.sub.3 and referenced from ΔCDCl.sub.3=7.26 ppm unless otherwise stated. Carbon NMR spectra were obtained on a Bruker AC 250 instrument operating at 63 MHz as solutions in CDCl.sub.3 and referenced from ΔCDCl.sub.3=7.26 ppm unless otherwise stated. Infrared spectra were recorded as KBr discs on a Mattson 3000 FTIR spectrophotometer. Electrospray mass spectrometry was carried out on a Waters LCT Premier ToF (Time of flight) mass spectrometer. Electrospray mass spectrometry and accurate mass spectrometry was also carried out by the EPSRC National Mass Spectrometry Facility in Swansea with a MAT95 magnetic sector. Melting points were obtained using a Reichert-Jung Thermo Galan hot stage microscope and are corrected. All chemicals were purchased from Sigma Aldrich.

Synthesis of methyl cholate (1)

(10) ##STR00009##

(11) Cholic acid (5.0 g, 12.2 mmol) was added to dry methanol (20 mL). Acetylchloride (0.5 mL, 0.04 mmol) was added under argon. The solution was heated at reflux for 45 minutes, then left to cool. Once at room temperature, the solution was cooled further on ice whereupon crystals appeared. The solid material was collected by filtration and washed with small amounts of methanol to give methyl cholate as a white solid. It was dried at room temperature under vacuum.

(12) Yield=3.548 g (68.9%).

(13) Melting point=110.8-111.9° C.

(14) .sup.1H NMR (CDCl.sub.3) δ ppm: 0.67 (s, 3H, Me-18), 0.88 (s, 3H, Me-19), 0.99 (d, 3H, Me-21), 1.0-2.43 (m steroid structure) 2.49 (s, 1H, H—C—C═O), 3.47 (s, 1H, H—C—OH), 3.66 (s, 1H, CH-3), 3.84 (d, 1H, CH-7), 3.95 (s, 1H, CH-12) ppm.

(15) .sup.13C NMR (CDCl.sub.3) δ 174.77 (C═O), 73.26 (C12), 68.66 (C7), 52.67 (C3), 51.50 (CH.sub.3), 50.33, (47.05, 46.44, 41.82, 39.47, 39.01, 35.24, 34.70, 31.06, 29.86, 28.17, 27.45, 26.60, 23.17 steroid structure) 22.44 (C19), 17.31 (C21), 12.49 (C18).

(16) MS (+APCI) m/z=482 (Mt).

(17) IR ν=3399 (OH), 2921, 2869, 1739 (C═O), 1449 cm.sup.−1.

Synthesis of cholic acid benzyl amide (2)

(18) ##STR00010##

(19) Methyl cholate (1 g, 2.3 mmol) was placed into a stainless steel pressure vessel along with benzyl amine (1.3 mL, 12.2 mmol) and toluene (5 mL). The pressure vessel was placed into an oil bath and heated to 150° C. for 48 hours. The pressure vessel was left to cool before dismantling it. The solution was poured into a round bottomed flask and placed on ice. The solid product was then collected by filtration and dried at room temperature under vacuum. The material was recrystallised from ethanol/water to give a white, powdery solid which was dried at room temperature under vacuum under vacuum.

(20) Yield=0.378 g (37.8%).

(21) Melting point=114.1-114.7° C.

(22) .sup.1H NMR (Methanol-d.sub.4) δ ppm: 0.69 (s, 3H, Me-18), 0.91 (s, 3H, Me-19), 1.02 (d, J=6.2 Hz, 3H, Me-21), 1.0-2.439 (m, steroid structure) 3.36 (m, 1H, CH-3), 3.78 (s, 1H, H—C-7), 3.94 (s, 1H, CH-12), 4.35 (d, 2H, J=1.9 Hz PH—CH.sub.2), 7.29 (m, J=6.2 Hz, 6H, aromatic ring).

(23) .sup.13C NMR (CDCl.sub.3) δ 219.45, 218.76, 173.58 (C═O), 154.31 (aromatic ring), 128.69 (aromatic ring), 127.85 (aromatic ring), 127.48 (aromatic ring), 125.07 (aromatic ring), 76.51 (C12), (43.61, 41.92, 39.51, 35.26, 34.69 steroid structure), 26.58 (C19), 17.44 (C21), 12.54 (C18).

(24) MS (+APCI) m/z=Found 498.3578; calculated for C.sub.31H.sub.48N.sub.1O.sub.4 497.71; −1.4 ppm.

(25) IR ν=3411, 2917, 1644 (C═O), 1540, 1457 cm.sup.−1

Synthesis of N-(4-aminobutyl) cholanamide (3)

(26) ##STR00011##

(27) Methyl cholate (1 g, 2.3 mmol) along with 1,4-diaminobutane (1,4-DAB) (10 mL) was put into a stainless steel pressure vessel and tightly secured. It was placed into an oil bath and heated to 150° C. for 48 hours. The pressure vessel was left to cool to room temperature before dismantling it. 30 mL chloroform was added to dissolve the product and excess 1,4-DAB which was then poured into a round bottomed flask. The chloroform was taken off using a rotary evaporator and the 1,4-DAB was taken off under reduced pressure rotary evaporation. Dichloromethane (90 mL) was added to precipitate the product. The yellow solid was collected by filtration and dried at room temperature under vacuum.

(28) Yield: 0.3 g, (30.14%).

(29) Melting point: 116.4-124.5° C.

(30) .sup.1H NMR (CDCl.sub.3) δ ppm: 0.68 (s, 3H, Me-18) 0.89 (s, 3H, Me-19) 1.00 (d, J=6.3 Hz, 3H, Me-21) 1.0-2.43 (m, steroid structure) 1.22 (dd, J-14.1, 8.5 Hz, CH) 2.71 (s, CH.sub.2) 3.28 (m, 1H, CH-3) 3.86 (s, 1H, CH-7) 3.98 (s, 1H, CH-12).

(31) .sup.13C NMR (DMSO) δ 172.43 (C═O), 107.85, 70.97 (C12), 66.20 (C7), (46.08, 45.70, 41.35, 40.17, 39.84, 37.97, 35.65, 35.13, 34.87, 34.36, 28.55, 27.51, 27.25, 26.42, 26.20, 22.81, 22.60 steroid ring), 41.71 (CH.sub.2), 40.51 (CH.sub.2), 30.38 (CH.sub.2), 20.75 (C19), 19.52 (C21), 17.10, 12.31 (C18).

(32) MS (+APCI) m/z=Found 479.3849; calculated for C.sub.28H.sub.50N.sub.2O.sub.4 478.3771; −1.0 ppm.

(33) IR (KBr) ν=3274, 2935, 2898, 2865, 2831, 1736, 1669 (C═O), 1548 cm.sup.−1.

Synthesis of N-[2-(2-aminomethylamino)ethyl]cholanamide (4)

(34) ##STR00012##

(35) Methyl cholate (0.5 g, 1.15 mmol) along with diethylenetriamine (2 mL) was put into a round bottomed flask under argon. It was placed into an oil bath and heated to 95° C. for 48 hours. The flask was left to cool to room temperature. Aceonitrile (15 mL) was added to precipitate the product which was then collected by filtration. Solvent extraction between water and chloroform to carried out to purify the product. The chloroform was evaporated under reduced pressure. The solid was dried at room temperature under vacuum at room temperature under vacuum.

(36) Yield=0.07 g (14%).

(37) Melting point: 205.6-206.8° C.

(38) .sup.1H NMR (CDCl.sub.3) δ ppm: 0.66 (s, 3H, Me-18) 0.87 (s, 3H, Me-19) 0.9 (d, J=6.0 Hz, 3H, Me-21) 1.0-2.44 (m, steroid structure) 2.69 (d, J=5.9 Hz, CH.sub.2) 2.75-2.88 (m) 3.35 (s) 3.47 (m, 1H, CH-3) 3.57 3.69 3.72 3.83 (s, 1H, CH-7) 3.94 (s, 1H, CH-12) 7.01 (s, 1H, NH).

(39) .sup.13C NMR (DMSO) δ ppm: 172.59 (C═O), 70.98 (C12), 70.41 (C3), 66.22 (C7), (45.70, 41.50, 41.32, 40.49, 40.16, 35.28, 34.35, 32.44, 31.57, 30.38, 28.53, 27.28, 26.18 steroid ring), 22.58 (C19), 17.08 (C21), 12.32 (C18).

(40) MS (+APCI) m/z=Found 494.3958; calculated for C.sub.29H.sub.53N.sub.3O.sub.4 493.3880; 0.6 ppm.

(41) IR (KBr) ν=3253, 2929, 2865, 1751, 1648 (C═O), 1557 cm.sup.−1.

Synthesis of N-[3-(cyclohexylamino)propyl] cholanamide (5)

(42) ##STR00013##

(43) Methyl cholate (2 g, 4.6 mmol) along with cyclohexyl-1,3-propanediamine (4.08 mL) was added to a round bottomed flask under nitrogen. The flask placed in a oil bath at 95° C. for 48 hours before raising the temperature to 120° C. for 12 hours. The flask was allowed to cool before chloroform (30 mL) was added to dissolve the product. Solvent extraction between chloroform (30 mL) and water (30 mL) removed any excess amine. This was repeated three times. The chloroform layer was dried over magnesium sulphate and evaporated under reduced pressure to give a solid that was recrystallized from chloroform-petrol 60-80.

(44) Yield=1.2 g (60%).

(45) Melting point: 104.5-105.8° C.

(46) .sup.1H NMR (CDCl.sub.3) δ ppm: 0.67 (s, 2H, Me-18) 0.89 (s, 2H, Me-19) 0.98 (d, 3H, Me-21) 1.0-2.43 (m, steroid structure) 2.61-2.79 (m, ring) 3.26 (m, 1H, CH-3) 3.43 (m, 1H, NH) 3.84 (s, 1H, CH-7) 3.97 (s, 1H, CH-12).

(47) .sup.13C NMR (CDCl.sub.3) δ ppm: 173.94 (C═O), 71.85 (C12), 68.27 (C7), 56.87 (CH), (46.51, 41.49, 40.08, 39.58, 38.83, 35.38, 31.74, 31.62, 30.41, 29.08, 28.17, 27.61, 26.43 (CH), 26.10 steroid ring), 45.30 (CH.sub.2), 40.68 (CH.sub.2), 25.11 (CH), 34.80 (CH), 33.12 (CH), 23.31, 22.69, 22.51 (C19), 17.57 (C21), 14.17, 12.50 (C18), 11.48.

(48) MS (+APCI) m/z=Found 547.4475; calculated for C.sub.33H.sub.58N.sub.2O.sub.4 546.4397; 0.4 ppm.

(49) IR (KBr) ν=3265, 3068, 2919, 2853, 1648 (C═O), 1554 cm.sup.−1.

Coupling of Cholic Acid with Ethylchloroformate and Various Amines

(50) General procedure for the coupling of cholic acid to primary and secondary amines Cholic acid (0.5 g, 1.2 mmol) was dissolved in THF (30 mL) along with triethylamine (2.9 mL, 0.3 mmol). The solution was put on ice for 10 minutes before ethylchloroformate (0.13 mL, 0.013 mmol) was dripped in over 10 minutes. The solution was allowed to react for two hours at room temperature. The required amine (1.2 mmol) was added and left to react for 3 hours. The reaction was quenched with water (30 mL). The mixture was washed with water (3×30 mL). The organic layer was dried over magnesium sulphate and the solvent was evaporated under reduced pressure. Solvent extraction between water and ethyl acetate was preformed 3 times before the organic layer was removed on the rotary evaporator. The product was dried at room temperature under vacuum.

Synthesis of N-[3-(cyclohexylamino)propyl] cholanamide (6)

(51) ##STR00014##

(52) The procedure was followed as the above with the exception of the removal of the two hour wait before adding the amine. Cyclohexylpropanediamine (0.24 mL, 1.56 mmol) added in one portion. The crude material was purified by flash chromatography eluting with methanol.

(53) Yield=0.36 g (72%).

(54) Melting point: 104.5-105.9° C.

(55) .sup.1H NMR (CDCl.sub.3) δ ppm: 0.67 (s, 2H, Me-18) 0.89 (s, 2H, Me-19) 0.98 (d, 3H, Me-21) 1.0-2.439 (m, steroid structure) 2.61-2.79 (m, ring) 3.26 (m, 1H, CH-3) 3.43 (m, 1H, NH) 3.84 (s, 1H, CH-7) 3.97 (s, 1H, CH-12).

(56) .sup.13C NMR (CDCl.sub.3) δ ppm: 173.94 (C═O), 73.08 (12), 71.85 (C3), 68.27 (C7), 56.87 (CH), (46.51, 41.49, 40.08, 39.58, 38.83, 35.38, 31.74, 31.62, 30.41, 29.08, 28.17, 27.61, 26.10 steroid ring), 45.30 (CH.sub.2), 40.68 (CH.sub.2), 34.80 (CH.sub.2), 33.12 (CH.sub.2), 26.43 (CH), 25.11 (CH), 23.31, 22.69, 22.51 (C19), 17.57 (C21), 14.17, 12.50 (C18), 11.48.

(57) MS (+APCI) m/z=Found 547.4461; calculated for C.sub.33H.sub.59N.sub.2O.sub.4 547.4469; −1.5 ppm.

(58) IR (KBr) ν=3265, 3068, 2919, 2853, 1648 (C═O), 1554 cm.sup.−1.

Synthesis of N-[3-(dibutylamino)propyl] cholanamide (7)

(59) ##STR00015##

(60) The procedure was followed as above with 3-dibutylaminopropylamine (0.22 mL, 1.18 mmol). Purification of the crude product was done dissolving the product in ethyl acetate (10 mL), removing solid impurities by filtration followed by removing the organic solvent. The product was dried at room temperature under vacuum.

(61) Yield=0.42 g (84%).

(62) Melting point: 145.1-145.9° C.

(63) .sup.1H NMR (CDCl.sub.3) δ ppm: 0.68 (s, 3H, Me-18) 0.89 (s, 3H, Me-19) 0.92 (d, J=6.3 Hz 3H, Me-21) 1.0-2.439 (m, steroid structure) 2.44 (m, butyl chain) 2.55 (m, butyl chain) 3.33 (m, propyl chain) 3.44 (m, 1H, CH-3) 3.84 (s, 1H, CH-7) 3.97 (s, 1H, CH-12) 7.43 (m, 1H, NH).

(64) Carbon=N/A.

(65) MS (+APCI) m/z=577.4931.

(66) IR (KBr) ν=3299, 3089, 2919, 2853, 2358, 1727, 1642 (C═O), 1545, 1463 cm.sup.−1.

Synthesis of N-(2-pyrrolidin-1-ylethyl) cholanamide (10)

(67) ##STR00016##

(68) The procedure was followed as above with 1-(2-aminoethyl) pyrrolidine (0.21 mL, 1.8 mmol).

(69) Yield=0.74 g (74%).

(70) Melting point: 96-97.1° C.

(71) .sup.1H NMR (CDCl.sub.3) δ ppm: 0.68 (s, 3H, Me-18) 0.89 (s, 3H, Me-19) 1.00 (d, J=6.2 Hz, 3H, Me-21) 1.0-2.43 (m, steroid structure) 2.23 (d, J=12.1 Hz, CH.sub.2) 2.64 (s 2H, CH.sub.2) 3.44 (m 1H, CH-3) 3.84 (s, 1H, CH-7) 3.96 (s, 1H, CH-12) 6.40 (s, 1H, NH).

(72) .sup.13C NMR (DMSO) δ ppm: 172.45 (C═O), 79.14, 70.98 (C12), 66.21 (C7), 55.02 (CH.sub.2), 53.58 (CH.sub.2 ring), (46.12, 45.70, 41.49, 37.81, 35.13, 34.35, 32.51, 31.70, 26.18 steroid ring), 41.34 (CH.sub.2) 23.08 (CH.sub.2 ring), 22.59 (C19), 17.07 (C21), 12.30 (C18).

(73) MS (+APCI) m/z=Found 505.3993; calculated for C.sub.30H.sub.53N.sub.2O.sub.4 505.4000; −1.4 ppm.

(74) IR (KBr) ν=3287, 2929, 2862, 2155, 1642 (C═O) cm.sup.−1.

Synthesis of N-[3-(dimethylamino)propyl] cholanamide (12)

(75) ##STR00017##

(76) The procedure was followed as above with 3-(dimethylamino)-1-propylamine (0.18 mL, 1.76 mmol).

(77) Yield=0.70 g (70%).

(78) Melting point: 163.2-164.0° C.

(79) .sup.1H NMR (CDCl.sub.3) δ ppm: 0.67 (s, 2H, Me-18) 0.87 (s, 2H, Me-19) 0.97 (d, J=6.2 Hz, 3H, Me-21) 1.0-2.43 (m, steroid structure) 2.21 (m, CH.sub.2) 2.33 (s, 6H, 2CH.sub.3) 3.30 (m, J=5.7 Hz, 1H) 3.82 (s, 1H, CH-7) 3.96 (s, 1H, CH-12) 6.97 (s, 1H, NH).

(80) .sup.13C NMR (DMSO) δ ppm: 176.07 (C═O), 70.99 (C12), 70.41 (C3), 66.23 (C7), 56.42 (CH.sub.3), (46.25, 44.96, 41.50, 41.31, 40.14, 35.25, 34.84, 34.36, 32.61, 30.38, 28.51, 27.30, 26.77, 26.17 steroid ring), 45.72 (CH.sub.3), 40.47 (CH.sub.2), 31.52 (CH.sub.2), 22.80, 22.58 (C19), 17.04 (C21), 12.34 (C18).

(81) MS (+APCI) m/z=Found 493.3994; calculated for C.sub.29H.sub.53N.sub.2O.sub.4 493.4000; −1.2 ppm.

(82) IR (KBr) ν=3387, 2932, 2862, 2209, 1991 cm.sup.−1.

Synthesis of N-(1-phenyl-4-piperidyl) cholanamide (13)

(83) ##STR00018##

(84) The procedure was followed as above with 4-amino-1-benzylpiperidine (0.34 mL, 1.79 mmol).

(85) Yield=0.88 g (88%).

(86) Melting point: 79.9-86.5° C.

(87) .sup.1H NMR (CDCl.sub.3) δ ppm: 0.67 (s, 3H, Me-18) 0.89 (s 3H, Me-19) 0.99 (d, J=6.1 Hz, 3H, Me-21) 1.0-2.43 (m, steroid structure) 2.83 (d, J=11.9 Hz) 3.43 (m, 1H, CH-3) 3.48 (d, J=3.2 Hz) 3.83 (s, 1H, CH-7) 3.96 (s, 1H, CH-12) 5.72 (d, J=8.0 Hz, 1H) 7.317 (s, 1H, NH).

(88) .sup.13C NMR (DMSO) δ ppm: 171.78 (C═O), 138.72, 138.63, (128.65, 128.09, 126.77 aromatic ring), 70.98 (C12), 70.41 (C3), 66.21 (C7), 62.15, 51.97, 51.87, (46.12, 45.71, 41.50, 41.33, 40.51, 40.32, 40.17, 35.29, 35.11, 34.85, 34.36, 32.59, 31.73, 31.62, 30.39, 28.53, 22.77 steroid ring), 27.27 (CH.sub.2 ring), 26.19 (CH.sub.2 ring) 22.59 (C19), 17.12 (C21), 14.63, 12.31 (C18).

(89) MS (+APCI) m/z=Found 519.4138; calculated for C.sub.36H.sub.57N.sub.2O.sub.4 581.4313. −61=benzene ring.

(90) IR (KBr) ν=3279, 3006, 2925, 2810, 2358, 2162, 1782, 1646 (C═O), 1513 cm.sup.−1.

Synthesis of 1-(3-phenylimidazolidin-1-yl) cholanone (14)

(91) ##STR00019##

(92) The procedure was followed as above with 1-(4-pyridyl)piperazine (0.24 g, 1.5 mmol).

(93) Yield=0.523 g (53%).

(94) Melting point: 139.6-141.2° C.

(95) .sup.1H NMR (CDCl.sub.3) δ ppm: 0.68 (s, 3H, Me-18) 0.89 (s, 3H, Me-19) 0.98 (d, 3H, Me-21) 1.0-2.43 (m steroid structure) 2.26 (m 1H) 3.63 (m 1H CH-3) 3.85 (m 1H CH-7) 3.98 (s 1H CH-12) 4.12 (d, J=6.95 Hz, 2H) 6.55-6.81 (m, 2H NH) 8.12-8.45 (m, 2H NH).

(96) .sup.13C NMR (DMSO) δ ppm: 173.30 (C═O), 171.33, 154.17, 149.80, 108.29, 99.48, 71.02 (C12), 70.41 (C3), 66.21 (C7), 59.58, (45.74, 45.74, 45.12, 40.47, 40.14, 35.27, 34.96, 34.35, 30.70, 30.37, 28.49, 26.17 steroid rings), 27.27 (CH.sub.2 ring) 22.77, 22.59 (C19), 17.17, 16.87 (C21), 14.11, 12.28 (C18).

(97) MS (+APCI) m/z=Found 454.3528; calculated for C.sub.33H.sub.51N.sub.3O.sub.4 553.387. MI-N(Ph)-NCH.sub.2CH.sub.2.

(98) IR (KBr) ν=3396, 3250, 2929, 2859, 2158, 1724, 1593, 1515 cm.sup.−1.

Synthesis of 1-[4-(2-hydroxyethyl)piperazin-1-yl] cholanone (15)

(99) ##STR00020##

(100) The procedure was followed as above with 1-(2-hydroxyethyl)piperazine (0.2 mL, 1.5 mmol). Proton NMR analysis showed excess of amine present.

(101) Yield=0.4 g (80%).

(102) Melting point: 167.8-168.1° C.

(103) .sup.1H NMR (CDCl.sub.3) δ ppm: 0.70 (s, 2H, Me-18) 0.90 (s, 2H, Me-19) 1.10 (d, J=5.3 Hz, 3H, Me-21) 1.0-2.43 (m, steroid structure) 2.50-2.63 (m, CH—N) 3.48 (m, J=4.9 1 Hz, CH-3 and CH) 3.64 (d, J=5.3 Hz, CH—N) 3.86 (s, 1H, CH-7) 3.98 (s, 1H, CH-12).

(104) .sup.13C NMR (DMSO) δ ppm: 170.97 (C═O), 154.56, 71.00 (C12), 70.40 (C3), 66.22 (C7), 60.61, 60.12 (CH.sub.2), 58.43 (CH.sub.2), 52.93 (CH.sub.2 ring), 52.80, (46.08, 43.29, 41.48, 35.26, 35.21, 34.83, 34.35, 31.19, 30.39, 29.48, 28.47, 27.29, 26.17 steroid ring), 45.74 (CH.sub.2 ring), 22.81, 22.59 (C19), 17.14 (C21), 14.54, 12.32 (C18).

(105) MS (+APCI) m/z=521.3943.

(106) IR (KBr) ν=3617, 3414, 2916, 2853, 2810, 1687 (C═O), 1624 cm.sup.−1.

Synthesis of N-octadecylcholanamide (16)

(107) ##STR00021##

(108) The procedure was followed as above with octadecylamine (0.8 g, 2.9 mmol). Further purification carried out using solvent extraction (×3) between THF/water (30 mL) and chloroform (30 mL). The organic layer was dried over magnesium sulphate and the solvent was evaporated under reduced pressure. The product was dried at room temperature under vacuum.

(109) Yield=0.922 g (90%).

(110) Melting point: 83.6-84.4° C.

(111) .sup.1H NMR (CDCl.sub.3) δ ppm: 0.66 (s, 3H, Me-18) 0.87 (d, J=3.4 Hz, 3H, Me-19) 0.97 (d, J=5.9 Hz, 3H, Me-21) 1.0-2.43 (m, steroid structure) 1.23 (s, 37H, CH.sub.2) 3.2 (q, J=7.3, 6.8 Hz) 3.43 (m, 1H, CH-3) 3.83 (s, 1H, CH-7) 3.96 (s, 1H, CH-12) 5.56 (s, 1H, NH).

(112) .sup.13C NMR (DMSO) δ ppm: 172.90 (C═O), 172.43, 70.98 (C12), 70.40 (C3), 66.21 (C7), 56.68, (46.15, 45.68, 45.08, 41.48, 36.68, 35.12, 34.35, 31.75, 31.26, 30.34, 29.02, 28.67, 27.13, 26.18 steroid ring), 22.79 (CH.sub.2), 22.59 (CH.sub.2), 22.57 (C19), 22.06, 17.08 (C21), 13.92 (CH.sub.2), 12.31 (C18).

(113) MS (+APCI) m/z=Found 660.5923; calculated for C.sub.42H.sub.77N.sub.1O.sub.4 660.5925; −0.4 ppm.

(114) IR (KBr) ν=3299, 2916, 2847, 1648 (C═O), 1642 cm.sup.−1.

Synthesis of N-benzylcholanamide (17)

(115) ##STR00022##

(116) The procedure was followed as above with benzylamine (0.12 mL, 1.1 mmol). The product was recrystallised in dichoromethane.

(117) Yield=0.282 g (56.4%).

(118) Melting point=114.1-114.7° C.

(119) .sup.1H NMR (Methanol-d.sub.4) δ ppm: 0.69 (s, 2H, Me-18), 0.91 (s, 2H, Me-19), 1.02 (d, J=6.2 Hz, 3H, Me-21), 1.0-2.43 (m, steroid structure) 3.36 (m, 1H, CH-3), 3.78 (s, 1H, H—C-7), 3.94 (s, 1H, CH-12), 4.35 (d), 7.29 (m, J=6.2 Hz, 6H).

(120) .sup.13C NMR (DMSO) δ ppm: 172.55 (C═O), (139.77, 128.17, 127.06, 126.60 aromatic ring), 70.97 (C12), 70.41 (C3), 66.20 (C7), (46.15, 45.71, 41.89, 40.49, 40.16, 35.28, 35.09, 34.86, 34.36, 34.36, 32.49, 31.77, 30.38, 28.53, 27.30, 26.18 steroid ring), 22.61 (C19), 17.06 (C21), 12.31 (C18).

(121) MS (+APCI) m/z=Found 498.3569; calculated for C.sub.31H.sub.47N.sub.1O.sub.4 498.3578; −1.8 ppm.

(122) IR ν=3411, 2917, 1644, 1540, 1457 cm.sup.−1.

Synthesis of 2-cholanamidobenzamide (18)

(123) ##STR00023##

(124) The procedure was followed as above with 2-aminobenzomide (0.5 g, 3.7 mmol). Further separation of the crude product to purify it was carried out with sodium hydrogen carbonate (30 mL) and 2M hydrochloric acid (30 mL). The organic layer was dried over magnesium sulphate and the solvent was evaporated under reduced pressure. The product was dried at room temperature under vacuum.

(125) Melting point: 103-104° C.

(126) .sup.1H NMR (CDCl.sub.3) δ ppm: 0.69 (s, 3H, Me-18) 0.89 (s, 3H, Me-19) 1.03 (d, J=6.0 Hz, 3H, Me-21) 1.0-2.43 (m steroid structure) 2.10 (s) 3.47 (m, J=7.9 Hz, 1H, CH-3) 3.84 (s, 1H, CH-7) 3.98 (s, 1H, CH-12) 6.25 7.07 (m, ring) 7.52 (d, J=8.3 Hz, ring) 8.62 (d, J=8.4 Hz, 1H, NH) 11.10 (s).

(127) .sup.13C NMR (CDCl.sub.3) δ ppm: 176.74, 176.03 (C═O), (145.05, 137.37, 133.74, 124.55 aromatic ring), 71.47 (C12), 51.28, 51.00, (46.75, 46.58, 45.75, 45.42, 45.09, 44.75, 44.42, 44.09, 43.75, 40.29, 39.61, 31.43, 27.84 steroid ring), 22.27 (C19), 17.56 (C21).

(128) MS (+APCI) m/z=Found 527.3473; calculated for C.sub.31H.sub.46N.sub.5O 527.3479; −1.2 ppm.

(129) IR (KBr) ν=3350, 3220, 2932, 2865, 1721, 1660 (c=C═O), 1612, 1581 cm.sup.−1.

Synthesis of N-(4-benzoylphenyl)cholanamide (19)

(130) ##STR00024##

(131) The procedure was followed as above with 4-aminobenzophenone (0.23 g, 1.17 mmol).

(132) Yield=0.1270 g (25.4%).

(133) Melting point: 125.8-127.6° C.

(134) .sup.1H NMR (CDCl.sub.3) δ ppm: 0.70 (s, 3H, Me-18) 0.90 (s, 2H, Me-19) 1.02 (d, J=6.1 Hz, 3H, Me-21) 1.0-2.43 (m, steroid structure) 3.48 (m, 1H, CH-3) 3.86 (s, 1H, CH-7) 3.99 (s, 1H, CH-12) 7.37-7.65 (m, 6H, ring) 7.65-7.91 (m, 6H, ring) 8.25 (s, 1N, NH).

(135) .sup.13C NMR (DMSO) δ ppm: 194.48 (C═O), 172.42 (C═O), (143.55, 137.59, 132.14, 131.11, 130.99, 129.30, 128.43, 118.16 aromatic rings), 66.21 (C7), (45.72, 40.50, 40.17, 39.84, 39.50, 39.17, 38.84, 38.50, 35.27, 35.12, 34.36, 34.36, 33.56, 31.29, 30.39, 28.54, 27.31, 26.20 steroid ring), 22.60 (C19), 17.13 (C21), 12.34 (C18).

(136) MS (+APCI) m/z=Found 588.3684; calculated for C.sub.37H.sub.50N.sub.1O.sub.5 587.36; −0.6 ppm.

(137) IR (KBr) ν=3317, 3098, 2932, 2865, 1967, 1645, 1584, 1521 cm.sup.−1.

Synthesis of N-(4-vinylphenyl)acetamide (20)

(138) ##STR00025##

(139) The procedure was followed as above with 4-vinyl aniline (0.42 mL, 3.5 mmol). Further purification with washing product dissolved in ethyl acetate (30 mL) with 2M HCl (30 mL) four times. The organic layer was dried over magnesium sulphate and the solvent was evaporated under reduced pressure. The product was dried at room temperature under vacuum. Proton NMR analysis showed the product contained excess 4-vinyl aniline.

(140) Melting point: N/A.

(141) .sup.1H NMR (CDCl.sub.3) δ ppm: 0.69 (s, 3H, Me-18) 0.90 (s, 3H, Me-19) 1.02 (d, J=6.0 Hz, 3H, Me-21) 1.0-2.43 (m, steroid structure) 2.36 (s) 2.62 (s) 3.47 (m, 1H, CH-3OH) 3.87 (s, 1H, CH-7) 3.99 (s, 1H, CH-12) 5.18 (d, J=11.5 Hz, 2H, CH.sub.2) 5.67 (d, J=18.0 Hz, 2H, CH.sub.2) 6.61 (dd) 7.36 (d, J=5.4 Hz, aromatic) 7.52 (d, J=8.5 Hz, aromatic).

(142) .sup.13C NMR (DMSO) δ 171.74 (C═O), (153.40, 139.14, 138.93, 131.73, 131.22, 128.96, 128.86, 128.17, 126.58, 126.47, 125.28, 118.85, 117.95, 112.28 aromatic ring, with excess 4-vinyl aniline), (137.30, 136.16, 112.46 C═C), 70.95 (C12), 70.40 (C3), 66.19 (C7), 60.14, (46.06, 45.69, 41.46, 41.34, 40.42, 40.09, 35.27, 35.16, 34.85, 34.35, 33.43, 31.43, 30.35, 28.53, 27.28, 26.16 steroid ring), 22.78, 22.59 (C19), 21.02, 17.11 (C21), 14.48, 12.33 (C18), 10.94.

(143) MS (+APCI) m/z=Found 510.3570; calculated for C.sub.32H.sub.48N.sub.1O.sub.4 510.3578; −1.5 ppm.

(144) IR (KBr) ν=3429, 3296, 3101, 3044, 2929, 2868, 2364, 1672 (C═O), 1587, 1521, 1508, 1460 cm.sup.1.

Synthesis of N-(2-dimethylaminoethyl)cholamide (21)

(145) ##STR00026##

(146) The procedure was followed as above with cholic acid (5 g, 12.2 mmol), triethylamine (2.9 mL, 28.7 mmol), ethylchloroformate (1.3 mL, 12 mmol) and dimethylethylenediamine (1.3 mL, 14.7 mmol).

(147) Yield=3.2185 g (64%).

(148) Melting point: 181-182.2° C.

(149) .sup.1H NMR (CDCl.sub.3) δ ppm: 0.66 (s, 3H, Me-18) 0.87 (s, 3H, Me-19) 0.98 (d, J=6.0 Hz, 3H, Me-21) 1.0-2.43 (m steroid structure) 2.37 (s, 4H, 2CH.sub.2) 2.62 (t, 2H, CH.sub.2) 3.39 (d, J=5.7 Hz, CH.sub.2 plus CH-3) 3.81 (s, 1H, CH-7) 3.94 (s, 1H, CH-12) 7.17 (s, 1H, NH).

(150) .sup.13C NMR (CDCl.sub.3) δ ppm: 218.02, 178.14, 174.59 (C═O), 73.01, 71.80 (C12), 68.37 (C7), 57.82 (CH.sub.2), (46.54, 46.49, 44.98, 44.63, 41.69, 39.73, 39.59, 35.62, 35.49, 34.87, 33.15, 31.75, 30.62, 28.30, 27.70, 26.42 steroid ring), 36.26 (CH.sub.3), 23.39, 23.32, 22.58 (C19), 17.55 (C21), 14.73, 12.53 (C18).

(151) MS (+APCI) m/z=Found 479.3835; calculated for C.sub.28H.sub.50N.sub.2O.sub.4 479.3843-1.7 ppm.

(152) IR (KBr) ν=3484, 3250, 3074, 2925, 2865, 1715, 1633, 1551 cm.sup.−1.

(153) Each cholic acid derivative was dissolved in either chloroform or dichloromethane along with a tenfold excess of the alkylating agent. The mixtures were left at room temperature for 4-48 hours. Upon the precipitation of the product, the solid was collected by filtration and washed with solvent, dried and purified if necessary.

Synthesis of 3-cholanamidopropyl(trimethyl)ammonium iodide (24)

(154) ##STR00027##

(155) N-[3-(dimethylamino)propyl] cholanamide (12) (0.5 g, 1.0 mmol) was dissolved in DCM (25 mL) along with methyl iodide (1.4 mL, 4.3 mmol). The reaction was left for 48 hours where upon a yellow solid formed. The solid was collected by filtration and washed with chloroform (10 mL) before drying at room temperature under vacuum.

(156) Yield=0.18 g (36%).

(157) Melting point: 134.6-135.4° C.

(158) .sup.1H NMR (D.sub.2O) δ ppm: 0.70 (s, 3H, Me-18) 0.90 (s, 3H, Me-19) 0.96 (d, J=Hz, 3H, Me-21) 1.0-2.43 (m, steroid structure) 3.11 (s, 9H, 3CH.sub.3) 3.30 (ddt, J=17.8 Hz) 3.48 (dd, J=10.4, 5.3 Hz, 1H, CH-3) 3.89 (s, 1H, CH-7) 4.05 (s, 1H, CH-12) 7.65 (s, 1H, NH).

(159) .sup.13C NMR (DMSO) δ ppm: 212.03, 172.84 (C═O), 79.97, 70.95 (C12), 70.37 (C3), 66.20 (C7), 52.27 (CH.sub.3), 52.18, 45.68 (CH.sub.2), 40.52 (CH.sub.2), (40.43, 40.18, 40.10, 35.27, 34.35, 32.33, 29.95 steroid ring), 22.97, 22.60 (19), 17.11 (C21), 12.31 (C18), 6.83.

(160) MS (+APCI) m/z=Found 507.4150; calculated for C.sub.30H.sub.55N.sub.2O.sub.4 507.4156; −1.3 ppm.

(161) IR (KBr) ν=3387, 2929, 2862, 1700, 1642 cm.sup.−1.

Synthesis of 3-cholanamidopropyl-ethyl-dimethyl-ammonium iodide (25)

(162) ##STR00028##

(163) N-[3-(dimethylamino)propyl] cholanamide (12) (0.5 g, 1.0 mmol) was dissolved in DCM (20 mL) along with ethyl iodide (0.4 mL, 1.3 mmol) and methanol (1 mL). The reaction was left for 1 week before the solvent was removed under reduced pressure. The solid was washed with ether (10 mL) and chloroform (10 mL) before drying. The product was a yellow solid.

(164) Yield=0.267 g (53%).

(165) Melting point: 122.5-123.1° C.

(166) .sup.1H NMR (D.sub.2O) δ ppm: 0.72 (s, 3H, Me-18) 0.92 (s, 3H, Me-19) 0.97 (d, 3H, Me-21) 1.0-2.43 (m, steroid structure) 3.04 (s, 6H, CH.sub.2) 3.21-3.34 (m, 4H, 2CH.sub.2) 3.38 (q, J=7.3 Hz, 2H, CH.sub.2) 3.44-3.62 (m, 1H, CH-3) 3.89 (s, 1H, CH-7) 4.06 (s, 1H, CH-12) 7.67 (s, 1H, NH).

(167) .sup.13C NMR (DMSO) δ ppm: 197.83, 172.88 (C═O), 70.96 (C12), 70.37 (C3), 66.20 (C7), 60.46, 58.59 (CH.sub.3), 49.59, 49.52, (45.96, 41.46, 41.38, 40.51, 40.17, 39.84, 39.51, 39.42, 39.17, 39.01, 38.84, 38.51, 35.47, 35.25, 35.15, 34.87, 34.34, 32.35, 30.35, 28.55, 27.26, 26.20 steroid ring), 45.68 (CH.sub.2), 31.52 (CH.sub.2), 22.77, 22.59 (C19), 22.52 (CH.sub.2), 17.10 (C21), 12.31 (C18), 7.76.

(168) MS (+APCI) m/z=Found 521.4305; calculated for C.sub.31H.sub.57N.sub.2O.sub.4 521.4313; −1.5 ppm.

(169) IR (KBr) ν=3420, 3256, 2913, 2856, 2243, 1639 (C═O) cm.sup.−1.

Synthesis of 3-cholanamidopropyl-propyl-dimethyl-ammonium iodide (26)

(170) ##STR00029##

(171) N-[3-(dimethylamino)propyl] cholanamide (12) (0.5 g, 1.0 mmol) was dissolved in DCM (20 mL) along with 1-iodopropane (1.5 mL, 5.0 mmol) and methanol (1 mL). The reaction was left for 1 week before solvent extraction (×3) between dichloromethane (20 mL) and water (20 mL). The organic layer was dried with magnesium sulphate before being removed under reduced pressure. The product was a white solid which was dried at room temperature under vacuum.

(172) Yield=0.233 g (46%).

(173) Melting point: 108.1-108.9° C.

(174) .sup.1H NMR (DMSO) δ ppm: 0.57 (s, 3H, Me-18) 0.79 (s, 3H, Me-19) 0.86-0.94 (m, 37H, Me-21 plus 2CH.sub.2) 1.0-2.43 (m, steroid structure) 2.94-3.28 (m, 14H) 3.61 (s, 1H, 7CH) 3.77 (s, 1H, 12CH) 3.99 (d, J=, 1H 3OH) 4.10 (d, J=, 1H, 70H) 4.32 (d, J=4.0 Hz, 1H, 120H) 7.88 (t, J=5.7 Hz, NH).

(175) .sup.13C NMR (DMSO) δ ppm: 172.92 (C═O), 70.97 (C12), 70.37 (C3), 66.20 (C7), 64.37, 61.10, 50.20, 50.16, (45.97, 35.45, 35.15, 34.86, 34.34, 32.37, 30.34, 28.53, 26.20 steroid ring), 45.68 (CH.sub.3), 31.54 (CH.sub.2), 27.26 (CH.sub.2), 22.58 (C19), 17.10 (C21), 15.32 (CH.sub.3), 12.31 (C18), 10.45 (CH.sub.2).

(176) MS (+APCI) m/z=Found 535.4461; calculated for C.sub.32H.sub.59N.sub.2O.sub.4 535.4469; −1.6 ppm.

(177) IR (KBr) ν=3396, 2935, 2865, 2246, 2124, 1706, 1645 (C═O), 1533 cm.sup.−1.

Synthesis of 3-cholanamidopropyl-butyl-dimethyl-ammonium iodide (27)

(178) ##STR00030##

(179) N-[3-(dimethylamino)propyl] cholanamide (12) (0.5 g, 1.0 mmol) was dissolved in DCM (20 mL) along with 1-iodobutane (1.2 mL, 4 mmol) and methanol (1 mL). The reaction was left for 1 week before solvent extraction (×3) between dichloromethane (20 mL) and water (20 mL). The organic layer was dried with magnesium sulphate before being removed under reduced pressure. The product was a white solid which was dried at room temperature under vacuum.

(180) Yield=0.3115 g (62%).

(181) Melting point: 125.8-126.4° C.

(182) .sup.1H NMR (D.sub.2O) δ ppm: 0.68 (s, 3H, Me-18) 0.88 (s, H, Me-19) 0.84-1.00 (m, 5H, Me-21 plus CH.sub.2) 1.0-2.43 (m, steroid structure) 3.02 (s, 6H, CH.sub.2) 3.14-3.32 (m, 6H, CH.sub.2) 3.47 (m, 1H, 3CH) 3.86 (s, 1H, 7CH) 4.02 (s, 1H, 12CH).

(183) MS (+APCI) m/z=Found 549.4619; calculated for C.sub.33H.sub.61N.sub.2O.sub.4 549.4626; −1.2 ppm.

(184) IR (KBr) ν=3378, 2932, 2865, 2358, 2337, 2155, 2009, 1976, 1651, 1633 (C═O), 1539 cm.sup.−1.

Synthesis of 3-cholanamidopropyl-pentyl-dimethyl-ammonium iodide (28)

(185) ##STR00031##

(186) N-[3-(dimethylamino)propyl] cholanamide (12) (0.5 g, 1 mmol) was dissolved in chloroform (75 mL) along with 1-iodopentane (1.5 mL) and methanol (1 mL). The reaction was left for 1 week before solvent extraction (×3) between chloroform (75 mL) and water (70 mL). The organic layer was dried with magnesium sulphate before being removed under reduced pressure. The product was a white solid which was dried at room temperature under vacuum.

(187) Yield=0.077 g (77%).

(188) Melting point: 107.9-108.3° C.

(189) .sup.1H NMR (Methanol-d.sub.4) δ ppm: 0.67 (s 3H Me-18) 0.83-1.05 (m 10H Me-21 plus CH.sub.2) 1.0-2.439 (m steroid structure) 3.04 (s 6H CH.sub.2) 3.47 (m 1H 3CH) 3.76 (s 1H 7CH) 3.92 (d J=3.3 Hz 1H 12CH)

(190) .sup.13C NMR (DMSO) δ ppm: 170.89 (C═O), 154.50, 71.01 (C12), 70.42 (C3), 66.22 (C7), 60.58, 57.47, 57.41 (CH.sub.3), 53.14, 52.55, 52.46, 48.55, 46.07, 45.02, 43.30, 41.52, 41.33, 40.93, 35.30, 35.22, 34.85, 29.49, 28.47, 28.35, 27.30, 26.17 steroid ring), 45.74 (CH.sub.3), 40.50 (CH.sub.2), 30.37 (CH.sub.2), 34.34 (CH.sub.2), 31.17 (CH.sub.2), 22.79 (CH.sub.2), 22.57 (C19), 20.03, 17.12 (C21), 14.51, 13.84 (CH.sub.2), 12.29 (C18).

(191) MS (+APCI) m/z=563.4767

(192) IR (KBr) ν=3381 2929 2871 1645 (C═O) 1533 cm.sup.−1.

Synthesis of 3-cholanamidopropyl-hexyl-dimethyl-ammonium iodide (29)

(193) ##STR00032##

(194) N-[3-(dimethylamino)propyl] cholanamide (12) (0.5 g, 1 mmol) was dissolved in chloroform (15 mL) along with 1-iodohexane (2.1 mL, 6.8 mmol) and methanol (1 mL). The reaction was left for 5 days before solvent was removed under reduced vacuum. The product was purified by washing with diethyl ether was collected by filtration and dried at room temperature under vacuum. The product was an orange solid.

(195) Yield=0.34 g (68%).

(196) Melting point: 101.7-102.3° C.

(197) .sup.1H NMR (Methanol-d.sub.4) δ ppm: 0.68 (s, 3H, Me-18) 0.90 (d, J=5.6 Hz, 6H) 1.01 (d, J=5.6 Hz, 3H, Me-21) 1.0-2.43 (m, steroid structure) 1.15 (td, J=7.0, 0.8 Hz) 3.04 (s, 6H, CH.sub.2) 3.47 (m, 1H, 3CH) 3.77 (s, 1H, 7CH) 3.92 (s, 1H, 12CH).

(198) .sup.13C NMR (DMSO) δ ppm: 172.86 (C═O), 70.96 (C12), 70.37 (C3), 66.20 (C7), 62.99, 60.98, 50.12, (45.98, 35.46, 35.13, 30.05, 29.48, 28.54, 27.26, 26.20 steroid ring), 45.68 (CH.sub.3), 34.34 (CH.sub.2), 30.65 (CH.sub.2), 31.55 (CH.sub.2), 25.40, 22.59 (C19), 21.87 (CH.sub.2), 21.59, 17.10 (C21), 13.81 (CH.sub.3), 12.32 (C18), 9.10 (CH.sub.2).

(199) MS (+APCI) m/z=Found 577.4949; calculated for C.sub.35H.sub.65N.sub.2O.sub.4 577.4939; 1.8 ppm.

(200) IR (KBr) ν=3390, 2925, 2859, 1654 (C═O) cm.sup.−1.

Synthesis of 3-cholanamidopropyl-benzyl-dimethyl-ammonium iodide (30)

(201) ##STR00033##

(202) N-[3-(dimethylamino)propyl] cholanamide (12) (0.5 g, 1 mmol) was dissolved in chloroform (20 mL) along with 2-phenylethyl bromide (1.36 mL, 5.4 mmol) and methanol (1 mL). The reaction was left for 24 hours before solvent was removed using a rotary evaporator. The resulting solid was washed with diethyl ether (10 mL) and dried under vacuum at room temperature. The product was a white solid.

(203) Melting point: 103.8-105° C.

(204) .sup.1H NMR (CDCl.sub.3) δ ppm: 0.58 (s, 2H, Me-18) 0.81 (s, 2H, Me-19) 0.92 (d, 3H, Me-21) 1.0-2.43 (m, steroid structure) 2.49 (p, J=1.9 Hz, 14H) 2.55-2.77 (m, 2H), 2.99 (s, 8H) 3.15-3.27 (m, 1H) 3.49 (t, J=6.6 Hz, 1H, 3CH) 3.60 (s, 1H, 7CH) 3.77 (s, 1H, 12CH) 4.05 (dd, J=22.2, 3.3 Hz, 1H, 3OH) 4.10 (d, 1H, 70H) 4.32 (d, J=4.0 Hz, 1H, 120H),

(205) .sup.13C NMR (DMSO) δ ppm: 172.89 (C═O), (140.23, 128.25, 126.21, 125.98 aromatic ring), 70.96 (C12), 70.38 (C3), 50.13, (45.68, 35.47, 34.85, 34.83, 34.35, 32.36, 31.65, 26.20 steroid ring), 23.54 (CH.sub.2), 22.54 (C19), 17.10 (C21), 12.32 (C18)

(206) MS (+APCI) m/z=583.4473

(207) IR (KBr) ν=3362, 3059, 3023, 2925, 2859, 1645 (C═O) cm.sup.−1.

Synthesis of 3-cholanamidopropyl-dimethyl-octadecyl-ammonium iodide (31)

(208) ##STR00034##

(209) N-[3-(dimethylamino)propyl] cholanamide (12) (0.5 g, 1 mmol) was dissolved in chloroform (15 mL) along with 1-iodo-octadecane (2.4 g, 5 mmol) and methanol (1 mL). The reaction was left for 72 hours before solvent was removed using a rotary evaporator. The resulting solid was washed with diethyl ether (10 mL) before being collected by filtration and dried under vacuum at room temperature. The product was a yellow solid.

(210) Yield=0.30 g (60%).

(211) Melting point: 115.3-116.4° C.

(212) .sup.1H NMR (Methanol-d.sub.4) δ ppm: 0.68 (s, 3H, Me-18) 0.84 (s, 3H, Me-19) 1.01 (d, 3H, Me-21) 1.0-2.43 (m, steroid structure) 1.26 (s, 9H) 2.23 (s, 7H) 3.05 (s) 3.15 (t) 3.27 (m) 3.44 (m, 1H, CH-3OH) 3.76 (s, 1H, CH-70H) 3.92 (s, 1H, CH-120H).

(213) .sup.13C NMR (DMSO) δ ppm: 172.30 (C═O), 70.98 (C12), 66.21 (C7), (46.21, 45.70, 34.85, 34.35, 31.80, 31.27, 30.34, 29.43, 28.77, 28.51, 27.26, 26.34, 26.17 steroid ring), 22.56 (C19), 22.06, 17.04 (C21), 13.91 (CH.sub.2), 12.29 (C18)

(214) MS (+APCI) m/z=754.6801

(215) IR (KBr) ν=3362, 2922, 2850, 1639 (C═O) cm.sup.−1.

Synthesis of 3-cholanamidopropyl-(2,2-dimethylpropanoyloxymethyl)-dimethyl-ammonium chloride (32)

(216) ##STR00035##

(217) N-[3-(dimethylamino)propyl] cholanamide (12) (0.5 g, 1 mmol) was dissolved in chloroform (20 mL) along with chloromethyl pivalate (1.44 g, 5 mmol) and methanol (1 mL). The reaction was left for 12 hours before solvent was removed using a rotary evaporator. The product was purified by washing with diethyl ether before being collected by filtraction and dried at room temperature under vacuum. The product was a white solid.

(218) Yield=0.335 g (66%).

(219) Melting point: 127.9-128.3° C.

(220) .sup.1H NMR (CDCl.sub.3) δ ppm: 0.57 (s, 3H, Me-18) 0.80 (s, 3H, Me-19) 1.08 (d, 3H, Me-21) 1.0-2.43 (m, steroid structure) 2.70 (s) 3.05 (s) 3.36 (dd, J=15.7, 8.7 Hz, CH-3) 3.60 (s, 1H, CH-7) 4.01 (d, J=3.3 Hz, 1H, 3OH) 4.10 (d, J=4.1 Hz, 1H, 70H) 4.34 (d, J=4.1 Hz, 1H, 120H) 5.25 (s, 2H) 5.85 (s, 1H) 7.93 (t, J=5.7 Hz, 1H, NH).

(221) .sup.13C NMR (DMSO) δ 179.30 (C═O), 172.96 (C═O), 70.96 (C12), 66.20 (C7), 54.42 (CH.sub.3), 47.85, (46.00, 45.70, 41.92, 34.86, 34.35, 27.24, 26.98, 26.43 steroid ring), 24.19, 22.59 (C19), 17.09 (C21), 12.31 (C18).

(222) MS (+APCI) m/z=607.4669

(223) IR (KBr) ν=3402, 3362, 2932, 2871, 2470, 1760, 1706, 1624 (C═O) cm.sup.−1.

Synthesis of 3-cholanamidopropyl-dimethyl-(2-phenylethyl) ammonium bromide (33)

(224) ##STR00036##

(225) N-[3-(dimethylamino)propyl] cholanamide (12) (0.5 g, 1 mmol) was dissolved in chloroform (20 mL) along with 1-bromo-3-phenylpropane (1.52 mL, 5 mmol) and methanol (1 mL). The reaction was left for 12 hours before solvent was removed using a rotary evaporator. The product was purified by washing with diethyl ether (10 mL) was collected by filtration and dried at room temperature under vacuum. The product was a white solid.

(226) Yield=0.4 g (80%).

(227) Melting point: 176.8-177.4° C.

(228) .sup.1H NMR (DMSO) δ ppm: 0.58 (s, 3H, Me-18) 0.82 (s, 3H, Me-19) 0.95 (d, J=5.6 Hz, 3H, Me-21) 1.10 (d, J=7.0 Hz) 3.12 (d, J=14.5 Hz) 3.61 (m) 3.78 (m) 4.00 (d, 1H, 3OH) 4.10 (d, 1H, 70H) 4.34 (d, 1H, 120H) 7.28 (m, 8H, aromatic) 7.93 (m, 1H, NH).

(229) .sup.13C NMR (DMSO) δ: 172.24 (C═O), (133.08, 128.86, 127.54 aromatic ring), 35.10, 34.35, 22.60 (C19), 17.12 (C21), 12.31 (18).

(230) MS (+APCI) m/z=611.4411

(231) IR (KBr) ν=3373, 3026, 2929, 2859, 1691 (C═O), 1645 cm.sup.−1.

Synthesis of 3-cholanamidopropyl-allyl-dimethyl-ammonium;bromide (34)

(232) ##STR00037##

(233) N-[3-(dimethylamino)propyl] cholanamide (12) (0.5 g, 1 mmol) was dissolved in chloroform (20 mL) along with allyl bromide (1.52 mL, 5 mmol) and methanol (1 mL). The reaction was left for 12 hours before solvent was removed using a rotary evaporator. The product was purified by washing with diethyl ether was collected by filtration and dried at room temperature under vacuum. The product was a white solid.

(234) Yield=0.19 g (40%).

(235) Melting point: 121.3-121.9° C.

(236) .sup.1H NMR (CDCl.sub.3) δ ppm: 0.68 (s, 3H, Me-18) 0.89 (s, 3H, Me-19) 0.85-1.61 (m, 29H) 1.00 (d, 3H, Me-21) 1.0-2.43 (m, steroid structure) 3.04 3.76 (s, 1H, CH-3) 3.87-4.13 (m, 6H) 5.61-5.76 (m, 3H) 5.93-6.17 (m, 2H), 7.88 (s, 1H, NH).

(237) .sup.13C NMR (DMSO) δ ppm: 172.88 (C═O), 127.60 (C═C), 125.76 (C═C), 70.96 (C12), 70.37 (C3), 66.20 (C7), 65.11, 61.12, 49.73, (45.97, 45.68, 35.26, 34.35, 28.53, 27.29, 26.20 steroid ring), 31.55 (CH.sub.2—Br), 25.04 (CH.sub.2), 22.56 (C19), 17.11 (C21), 12.31 (C18).

(238) MS (+APCI) m/z=Found 533.4307; calculated for C.sub.32H.sub.57N.sub.2O.sub.4 533.4313; −1.1 ppm.

(239) IR (KBr) ν=3368, 3077, 2925, 2862, 2710, 1733, 1651 (C═O), 1633, 1466 cm.sup.−1.

Synthesis of 3-cholanamidopropyl-cyclopentyl-dimethyl-ammonium bromide (35)

(240) ##STR00038##

(241) N-[3-(dimethylamino)propyl] cholanamide (12) (0.5 g, 1 mmol) was dissolved in DCM (10 mL) along with) of cyclopentyl bromide (0.9 mL, 10 mmol). The reaction was left for 72 hours where the product precipitated out. It was collected by filtration and dried at room temperature under vacuum. The product was a white solid.

(242) Melting point: 102.6-103° C.

(243) .sup.1H NMR (DMSO) δ ppm: 0.57 (d, J=1.4 Hz, 3H Me-18) 0.76-0.97 (m, 6H, Me-19 and Me-21) 1.0-2.43 (m, steroid structure) 2.13 (s, dimethyl) 3.14 (s, 3H) 3.4-3.55 (m, 1H, CH-3OH) 3.60 (s 1H CH-70H) 3.77 (s, 1H, CH-120H) 3.94-4.18 (m, 4H) 4.32 (t, J=3.9 Hz, 1H) 5.35 (s, 1H, NH).

(244) .sup.13C NMR (DMSO) δ ppm: 216.00, 138.38 (C═O), 70.92 (C19), 70.38 (C21), 66.17 (C18), 60.48 (CH—Br), (45.71, 43.19, 41.39, 40.51, 40.17, 36.53, 35.28, 34.89, 34.35, 30.64, 30.37, 30.35, 30.18, 28.51, 26.19 steroid ring), 25.66, 24.51 (CH.sub.2 ring), 22.59 (C19), 21.64, 16.91 (C21), 12.29 (C18).

(245) MS (+APCI) m/z=562.4448

(246) IR (KBr) ν=3368, 2932, 2865, 2355, 1715 (C═O), 1578, 1460 cm.sup.−1.

Synthesis of 3-cholanamidoethylyl(trimethyl)ammonium iodide (36)

(247) ##STR00039##

(248) N-[3-(dimethylamino)ethyl]cholanamide (21) (0.5 g, 1.0 mmol) was dissolved in DCM (20 mL) along with methyl iodide (0.3 mL, 2.1 mmol). The reaction was left for 12 hours where upon a solid formed and the solvent had evaporated. Diethyl ether (10 mL) was added to the flask and the solid was collected by filtration before being washed with chloroform (10 mL). The product was dried at room temperature under vacuum.

(249) Yield=0.314 g (63%).

(250) Melting point: 215.3-216.1° C.

(251) .sup.1H NMR (D.sub.2O) δ ppm: 0.70 (s, 3H, Me-18) 0.90 (s, 3H, Me-19) 0.96 (d, J=Hz, 3H, Me-21) 1.0-2.43 (m, steroid structure) 3.11 (s, 9H, 3 CH.sub.3) 3.30 (ddt, J=17.8 Hz) 3.48 (dd, J=10.4, 5.3 Hz, 1H, CH-3) 3.89 (s, 1H, CH-7) 4.05 (s, 1H, CH-12) 7.65 (s, 1H, NH).

(252) .sup.13C NMR (DMSO) δ ppm: 212.03, 172.84 (C═O), 79.97, 70.95 (C12), 70.37 (C3), 66.20 (C7), 52.27 (CH.sub.3), 52.18, 45.68 (CH.sub.2), 40.52 (CH.sub.2), (40.43, 40.18, 40.10, 35.27, 34.35, 32.33, 29.95 steroid ring), 22.97, 22.60 (19), 17.11 (C21), 12.31 (C18), 6.83.

(253) MS (+APCI) m/z=Found 493.3991; calculated for C.sub.29H.sub.53N.sub.2O.sub.4 493.4000; −1.8 ppm.

(254) IR (KBr) ν=3423, 3241, 2935, 2862, 2252, 2118, 1663, 1618, 1539 cm.sup.−1.

Synthesis of 3-cholanamidoethylyl-ethyl-dimethyl-ammonium iodide (37)

(255) ##STR00040##

(256) N-[3-(dimethylamino)ethyl]cholanamide (21) (0.5 g, 1.0 mmol) was dissolved in DCM (20 mL) along with ethyl iodide (0.4 mL, 1.3 mmol) and methanol (1 mL). The reaction was left for 12 hours before the solvent was removed under reduced pressure. The solid was washed with ether (10 mL) and chloroform (10 mL) before drying. The product was a yellow solid.

(257) Yield=0.280 g (56%).

(258) Melting point: 142.3-142.7° C.

(259) .sup.1H NMR (D.sub.2O) δ ppm: 0.72 (s, 3H, Me-18) 0.92 (s, 3H, Me-19) 0.97 (d, 3H, Me-21) 1.0-2.43 (m, steroid structure) 3.04 (s, 6H, CH.sub.2) 3.21-3.34 (m, 4H, 2 CH.sub.2) 3.38 (q, J=7.3 Hz, 2H, CH.sub.2) 3.44-3.62 (m, 1H, CH-3) 3.89 (s, 1H, CH-7) 4.06 (s, 1H, CH-12) 7.67 (s, 1H, NH).

(260) .sup.13C NMR (DMSO) δ ppm: 197.83, 172.88 (C═O), 70.96 (C12), 70.37 (C3), 66.20 (C7), 60.46, 58.59 (CH.sub.3), 49.59, 49.52, (45.96, 41.46, 41.38, 40.51, 40.17, 39.84, 39.51, 39.42, 39.17, 39.01, 38.84, 38.51, 35.47, 35.25, 35.15, 34.87, 34.34, 32.35, 30.35, 28.55, 27.26, 26.20 steroid ring), 45.68 (CH.sub.2), 31.52 (CH.sub.2), 22.77, 22.59 (C19), 22.52 (CH.sub.2), 17.10 (C21), 12.31 (C18), 7.76.

(261) MS (+APCI) m/z=Found 507.4147; calculated for C.sub.30H.sub.55N.sub.2O.sub.4 507.4156; −1.8 ppm.

(262) IR (KBr) ν=3387, 2932, 2862, 2689, 1706, 1648 (C═O), 1533 cm.sup.−1.

Synthesis of 3-cholanamidopropyl-propyl-dimethyl-ammonium iodide (38)

(263) ##STR00041##

(264) N-[3-(dimethylamino)ethyl]cholanamide (21) (0.5 g, 1.0 mmol) was dissolved in DCM (20 mL) along with 1-iodopropane (0.5 mL, 1.7 mmol) and methanol (1 mL). The reaction was left for 6 days before the solvent was removed under reduced pressure. The solid was washed with diethyl ether (10 mL) and collected by filtration. Further purification was carried out by washing the crude product with diethyl ether (2×10 mL) and chloroform (2×10 mL). The product was a white solid which was dried at room temperature under vacuum.

(265) Yield=0.283 g (56%).

(266) Melting point: 108.1-108.9° C.

(267) .sup.1H NMR (DMSO) δ ppm: 0.57 (s, 3H, Me-18) 0.79 (s, 3H, Me-19) 0.86-0.94 (m, 37H, Me-21 plus 2 CH.sub.2) 1.0-2.43 (m, steroid structure) 2.94-3.28 (m, 14H) 3.61 (s, 1H, 7CH) 3.77 (s, 1H, 12CH) 3.99 (d, J=, 1H 3OH) 4.10 (d, J=, 1H, 7OH) 4.32 (d, J=4.0 Hz, 1H, 120H) 7.88 (t, J=5.7 Hz, NH).

(268) .sup.13C NMR (DMSO) δ ppm: 172.92 (C═O), 70.97 (C12), 70.37 (C3), 66.20 (C7), 64.37, 61.10, 50.20, 50.16, (45.97, 35.45, 35.15, 34.86, 34.34, 32.37, 30.34, 28.53, 26.20 steroid ring), 45.68 (CH.sub.3), 31.54 (CH.sub.2), 27.26 (CH.sub.2), 22.58 (C19), 17.10 (C21), 15.32 (CH.sub.3), 12.31 (C18), 10.45 (CH.sub.2)

(269) MS (+APCI) m/z=Found 521.4303; calculated for C.sub.31H.sub.57N.sub.2O.sub.4 521.4313; −1.9 ppm.

(270) IR (KBr) ν=3390, 2935, 2862, 2243, 2121, 1651 (C═O), 1536 cm.sup.−1.

Synthesis of 3-cholanamidoethyl-butyl-dimethyl-ammonium iodide (39)

(271) ##STR00042##

(272) N-[3-(dimethylamino)propyl] cholanamide (21) (0.5 g, 1.0 mmol) was dissolved in DCM (20 mL) along with 1-iodobutane (0.6 mL, 2 mmol) and methanol (1 mL). The reaction was left for 3 days where a precipitate formed. The crude product was collected by filtration and washed with chloroform (15 mL). The product was a white solid which was dried at room temperature under vacuum.

(273) Yield=0.338 g (67%).

(274) Melting point: 186.4-187.4° C.

(275) .sup.1H NMR (D.sub.2O) b ppm: 0.68 (s, 3H, Me-18) 0.88 (s, H, Me-19) 0.84-1.00 (m, 5H, Me-21 plus CH.sub.2) 1.0-2.43 (m, steroid structure) 3.02 (s, 6H, CH.sub.2) 3.14-3.32 (m, 6H, CH.sub.2) 3.47 (m, 1H, 3CH) 3.86 (s, 1H, 7CH) 4.02 (s, 1H, 12CH).

(276) MS (+APCI) m/z=Found 535.4458; calculated for C.sub.32H.sub.59N.sub.2O.sub.4 535.4469; −2.1 ppm.

(277) IR (KBr) ν=3484, 3432, 3250, 3071, 2922, 2865, 1633 (C═O), 1551 cm.sup.−1.

Synthesis of 3-cholanamidoethyl-pentyl-dimethyl-ammonium iodide (40)

(278) ##STR00043##

(279) N-[3-(dimethylamino)ethyl]cholanamide (21) (0.5 g, 1 mmol) was dissolved in DCM (20 mL) along with 1-iodopentane (0.6 mL, 1.9 mmol) and methanol (1 mL). The reaction was left for 3 days where a precipitate formed. The product was collected by filtration, washed with chloroform (20 mL) and dried at room temperature under vacuum. The product was a white solid.

(280) Yield=0.396 q (79%).

(281) Melting point: 139.6-141.2° C.

(282) .sup.1H NMR (Methanol-d.sub.4) δ ppm: 0.67 (s 3H Me-18) 0.83-1.05 (m 10H Me-21 plus CH.sub.2) 1.0-2.439 (m steroid structure) 3.04 (s 6H CH.sub.2) 3.47 (m 1H 3CH) 3.76 (s 1H 7CH) 3.92 (d J=3.3 Hz 1H 12CH).

(283) .sup.13C NMR (DMSO) δ ppm: 170.89 (C═O), 154.50, 71.01 (C12), 70.42 (C3), 66.22 (C7), 60.58, 57.47, 57.41 (CH.sub.3), 53.14, 52.55, 52.46, 48.55, 46.07, 45.02, 43.30, 41.52, 41.33, 40.93, 35.30, 35.22, 34.85, 29.49, 28.47, 28.35, 27.30, 26.17 steroid ring), 45.74 (CH.sub.3), 40.50 (CH.sub.2), 30.37 (CH.sub.2), 34.34 (CH.sub.2), 31.17 (CH.sub.2), 22.79 (CH.sub.2), 22.57 (C19), 20.03, 17.12 (C21), 14.51, 13.84 (CH.sub.2), 12.29 (C18).

(284) MS (+APCI) m/z=Found 549.4618; calculated for C.sub.33H.sub.61N.sub.2O 4549.4626; −1.4 ppm.

(285) IR (KBr) ν=3484, 3250, 3071, 2925, 2868, 1627 (C═O), 1560 cm.sup.−1.

Synthesis of 3-cholanamidoethyl-hexyl-dimethyl-ammonium iodide (41)

(286) ##STR00044##

(287) N-[3-(dimethylamino)ethyl]cholanamide (21) (0.5 g, 1 mmol) was dissolved in DCM (10 mL) along with 1-iodohexane (0.7 mL, 2 mmol) and methanol (1 mL). The reaction was left for 3 days where a precipitate formed. The product was collected by filtration, washed with chloroform (20 mL) and dried at room temperature under vacuum. The product was a white solid.

(288) Yield=0.45 g (91%).

(289) Melting point: 104.2-104.5° C.

(290) .sup.1H NMR (Methanol-d.sub.4) δ ppm: 0.68 (s, 3H, Me-18) 0.90 (d, J=5.6 Hz, 6H) 1.01 (d, J=5.6 Hz, 3H, Me-21) 1.0-2.43 (m, steroid structure) 1.15 (td, J=7.0, 0.8 Hz) 3.04 (s, 6H, CH.sub.2) 3.47 (m, 1H, 3CH) 3.77 (s, 1H, 7CH) 3.92 (s, 1H, 12CH).

(291) .sup.13C NMR (DMSO) δ ppm: 172.86 (C═O), 70.96 (C12), 70.37 (C3), 66.20 (C7), 62.99, 60.98, 50.12, (45.98, 35.46, 35.13, 30.05, 29.48, 28.54, 27.26, 26.20 steroid ring), 45.68 (CH.sub.3), 34.34 (CH.sub.2), 30.65 (CH.sub.2), 31.55 (CH.sub.2), 25.40, 22.59 (C19), 21.87 (CH.sub.2), 21.59, 17.10 (C21), 13.81 (CH.sub.3), 12.32 (C18), 9.10 (CH.sub.2).

(292) MS (+APCI) m/z=Found 563.4771; calculated for C.sub.34H.sub.63N.sub.2O.sub.4 563.4782; −2.0 ppm.

(293) IR (KBr) ν=3393, 3238, 3056, 2925, 2853, 1703, 1651 (C═O), 1606 cm.sup.−1.

Synthesis of 3-cholanamidoethyl-allyl-dimethyl-ammonium bromide (42)

(294) ##STR00045##

(295) N-[3-(dimethylamino)ethyl]cholanamide (21) (0.5 g, 1 mmol) was dissolved in DCM (10 mL) along with allyl bromide (0.4 mL, 3.3 mmol) and methanol (1 mL). The reaction was left for 12 hours before solvent was removed using a rotary evaporator. The product was purified by washing with diethyl ether was collected by filtration and dried at room temperature under vacuum. The product was a white solid.

(296) Yield=0.129 g (26%).

(297) Melting point: 119.1-120.3° C.

(298) .sup.1H NMR (CDCl.sub.3) δ ppm: 0.68 (s, 3H, Me-18) 0.89 (s, 3H, Me-19) 0.85-1.61 (m, 29H) 1.00 (d, 3H, Me-21) 1.0-2.43 (m, steroid structure) 3.04 3.76 (s, 1H, CH-3) 3.87-4.13 (m, 6H) 5.61-5.76 (m, 3H) 5.93-6.17 (m, 2H), 7.88 (s, 1H, NH).

(299) .sup.13C NMR (DMSO) δ ppm: 172.88 (C═O), 127.60 (C═C), 125.76 (C═C), 70.96 (C12), 70.37 (C3), 66.20 (C7), 65.11, 61.12, 49.73, (45.97, 45.68, 35.26, 34.35, 28.53, 27.29, 26.20 steroid ring), 31.55 (CH.sub.2—Br), 25.04 (CH.sub.2), 22.56 (C19), 17.11 (C21), 12.31 (C18)

(300) MS (+APCI) m/z=Found 519.4146; calculated for C.sub.31H.sub.5N.sub.2O.sub.4 519.4156; −2.0 ppm.

(301) IR (KBr) ν=3362, 2929, 2862, 1700, 1645 (C═O), 1536 cm.sup.−1.

Synthesis of N-[2-(1-methylpyrrolidin-1-ium-1-yl)ethyl]cholanamide iodide (43)

(302) ##STR00046##

(303) N-(2-pyrrolidin-1-ylethyl)cholanamide (10) (0.5 g, 1 mmol) was dissolved in DCM (10 mL) along with of methyl iodide (1.4 mL, 10 mmol). The reaction was left for 60 hours where the product precipitated out. It was then collected by filtration, washed with chloroform (20 mL) and dried at room temperature under vacuum. The product was a white solid.

(304) Melting point: 140.7-141-4° C.

(305) .sup.1H NMR (DMSO) δ ppm: 0.58 (s, 3H, Me-18) 0.81 (s, 3H, Me-19) 0.93 (d, J=5.8 Hz, 3H, Me-21) 1.0-2.43 (m, steroid structure) 2.23 (s) 2.55 2.64 3.44 (1H, CH-3OH) 3.84 (s, 1H, CH-70H) 3.96 (s, 1H, CH-120H) 6.40 (s, 1H, NH).

(306) .sup.13C NMR (DMSO) δ: 173.30 (C═O), 70.96 (C12), 70.37 (C3), 66.19 (C7), 63.86 (CH.sub.2), 61.67 (CH.sub.2 ring), 47.44, (45.98, 45.68, 41.46, 35.09, 34.34, 33.58, 32.25, 31.40, 28.54, 27.26, 26.20 steroid ring), 41.39 (CH.sub.2), 22.76 (CH.sub.2 ring), 22.59 (C19), 20.91, 17.04 (C21), 12.31 (C18).

(307) MS (+APCI) m/z=Found 519.4152; calculated for C.sub.31H.sub.55N.sub.2O 4519.4156; −0.8 ppm.

(308) IR (KBr) ν=3390, 3274, 2922, 2862, 1651 (C═O), 1533, 1460 cm.sup.−1.

Synthesis of N-[2-(1-ethylpyrrolidin-1-ium-1-yl)ethyl]cholanamide iodide (44)

(309) ##STR00047##

(310) N-(2-pyrrolidin-1-ylethyl)cholanamide (10) (0.5 g, 1 mmol) was dissolved in DCM (10 mL) along with of ethyl iodide (0.4 mL, 10 mmol). The reaction was left for 60 hours where the product precipitated out. It was collected by filtration, washed with chloroform and dried at room temperature under vacuum. The product was a yellow solid.

(311) Melting point: 120.7-121.7° C.

(312) .sup.1H NMR (DMSO) δ ppm: 0.57 (s, 3H, Me-18) 0.80 (s, 3H, Me-19) 0.92 (d, J=6.2 Hz, 3H, Me-21) 1.0-2.43 (m, steroid structure) 2.23 (d, J=12.1 Hz) 2.55 2.64 3.44 (1H, CH-3OH) 3.60 (s, 1H, CH-70H) 3.77 (s, 1H, CH-120H) 5.75 8.08 (s, 1H, NH).

(313) Carbon N/A

(314) MS (+APCI) m/z=Found 533.4302; calculated for C.sub.32H.sub.57N.sub.2O.sub.4 533.4313; −2.0 ppm.

(315) IR (KBr) ν=3378, 2929, 2862, 2361, 2158, 2018, 1645 (C═O), 1527 cm.sup.−1.

Synthesis of N-[2-(1-propylpyrrolidin-1-ium-1-yl)ethyl]cholanamide;iodide (45)

(316) ##STR00048##

(317) N-(2-pyrrolidin-1-ylethyl)cholanamide (10) (0.5 g, 1 mmol) was dissolved in DCM (15 mL) along with of 1-iodopropane (3 mL, 10 mmol). The reaction was left for 12 hours where the product precipitated out. It was collected by filtration, washed with chloroform and dried. The product was a yellow solid.

(318) Yield=0.947 g

(319) Melting point: 203.8-204.1° C.

(320) .sup.1H NMR (D.sub.2O) δ ppm: 0.68 (s, 3H, Me-18) 0.80 (s, 3H, Me-19) 0.91 (d, 3H, Me-21) 1.0-2.43 (m, steroid structure) 3.18 (m) 3.49 (ddt, J=40.0, 13.4, 6.6 Hz, 8H, CH-30H, CH.sub.2) 3.87 (s, 1H, CH-70H) 4.03 (s, 1H, CH-120H) 7.64 (s, 1H, NH) Carbon=N/A

(321) MS (+APCI) m/z=Found 547.4463; calculated for C.sub.33H.sub.59N.sub.2O.sub.4 547.4469; −1.2 ppm.

(322) IR (KBr) ν=3566, 3353, 3217, 2913, 2856, 2246, 2121, 1642, 1527 cm.sup.−1.

Synthesis of N-[2-(1-hexylylpyrrolidin-1-ium-1-yl)ethyl]cholanamide iodide (48)

(323) ##STR00049##

(324) N-(2-pyrrolidin-1-ylethyl)cholanamide (10) (0.5 g, 1 mmol) was dissolved in DCM (15 mL) along with of 1-iodopentane (2 mL, 10 mmol). The reaction was left for 96 hours where the product precipitated out. It was collected by filtration, washed with chloroform (15 mL) and dried at room temperature under vacuum. The product was a yellow solid.

(325) Yield=0.49 g (98%).

(326) .sup.1H NMR (CDCl.sub.3) δ ppm: 0.58 (s, 3H, Me-18) 0.81 (s, 3H, Me-19) 0.91 (d, 3H, Me-21) 1.0-2.43 (m, steroid structure) 2.50 (p, J=1.8 HZ, 4H) 3.33 (m) 3.49 (m, 8H, CH-30H, CH.sub.2) 3.77 (s, 1H, CH-70H) 4.32 (s, 1H, CH-120H) 7.72 (s, 1H, NH), 8.08, 8.31

(327) .sup.13C NMR (DMSO) δ 173.38 (C═O), 70.96 (C12), 70.37 (C3), 70.30, 66.19 (C7), 62.48, 58.76, 56.74 (CH.sub.2 ring), (45.95, 45.69, 40.48, 40.15, 39.81, 39.48, 39.15, 38.81, 38.48, 35.09, 34.34, 32.22, 31.36, 30.71, 28.54, 27.25, 26.21 steroid ring), 25.43 (CH.sub.2 ring), 22.59 (C19), 21.91 (CH.sub.2), 21.15, 17.04 (C21), 13.82 (CH.sub.3), 12.29 (C18).

(328) MS (+APCI) m/z=589.4926

(329) IR (KBr) ν=3466, 3362, 3186, 3044, 2953, 2922, 1706, 1660, 1624 (C═O), 1530 cm.sup.−1.

Synthesis of N-[2-(1-allylpyrrolidin-1-ium-1-yl)ethyl]cholanamide bromide (49)

(330) ##STR00050##

(331) N-(2-pyrrolidin-1-ylethyl)cholanamide (10) (0.5 g, 1 mmol) was dissolved in chloroform (20 mL) along with allylbromide (1.2 mL, 10 mmol). The reaction was left for 96 hours where the product precipitated out. It was collected by filtration, washed with chloroform (20 mL) and dried at room temperature under vacuum. The product was a white solid.

(332) Yield=0.59 g (60%).

(333) Melting point: 186.5-186.9° C.

(334) .sup.1H NMR (DMSO) δ ppm: 0.59 (s, 3H, Me-18) 0.88 (s, 3H, Me-19) 0.94 (d, 3H, Me-21) 1.0-2.43 (m, steroid structure) 2.07 (m) 2.23 (d, J=12.1 Hz, CH.sub.2) 2.55 2.64 3.52 (m, 3H, CH-30H+CH.sub.2) 3.61 (s, 1H, CH-70H) 3.79 (s, 1H, CH-120H) 4.00 (d, 2H, CH.sub.2 allyl bromide) 5.66 (t, 2H, allyl bromide) 6.08 (m, 1H, allyl bromide) 8.13 (t, 1H, NH) 8.32 (s, 1H).

(335) .sup.13C NMR (DMSO) δ ppm: 173.37 (C═O), 127.23 (C═C), 126.28 (C═C), 79.17, 70.94 (C12), 70.36 (C3), 66.18 (C3), 61.74, 60.54, 57.71 (CH.sub.2), (45.94, 45.68, 40.48, 40.15, 40.07, 39.82, 39.73, 39.48, 39.15, 38.81, 38.48, 34.34, 33.11, 31.39, 30.35, 28.54, 28.53, 27.29, 26.20 steroid ring), 32.18 (CH.sub.2), 22.60 (C19), 21.16, 17.03 (C21), 12.30 (C18).

(336) MS (+APCI) m/z=Found 545.4306; calculated for C.sub.33H.sub.57N.sub.2O.sub.4 545.4313; −1.3 ppm.

(337) IR (KBr) ν=3472, 3365, 3211, 3050, 2922, 2859, 2458, 2042, 1630 (C═O), 1542 cm.sup.−1.

Synthesis of N-[2-(1-benzylpyrrolidin-1-ium-1-yl)ethyl]cholanamide bromide (50)

(338) ##STR00051##

(339) N-(2-pyrrolidin-1-ylethyl)cholanamide (10) (0.5 g, 1 mmol) was dissolved in DCM (15 mL0 along with benzyl bromide (1.2 mL, 10 mmol). The reaction was left for 48 hours where the product precipitated out. It was collected by filtration, washed with chloroform (15 mL) and dried at room temperature under vacuum. The product was a yellow solid.

(340) Yield=0.301 g (30%).

(341) Melting point: 165.7-165.9° C.

(342) .sup.1H NMR (D.sub.2O) δ ppm: 0.56 (s, 3H, Me-18) 0.87 (s, 3H, Me-19) 0.92 (d, 3H, Me-21) 1.0-2.43 (m, steroid structure) 2.19 (s) 2.23 (d, J=12.1 Hz, CH.sub.2) 3.37-3.68 (m, 6H, CH-30H+CH.sub.2) 3.75 (s, 1H, CH-70H) 3.98 (s, 1H, CH-120H) 7.38 (s, 1H, NH) 7.54 (m, aromatic)

(343) .sup.13C NMR (DMSO) δ ppm: 173.43 (C═O), (132.61, 129.04, 128.41 aromatic ring), 60.89 (CH.sub.2), 60.86 (CH.sub.2), (45.69, 35.25, 35.10, 34.94, 34.67, 34.34 steroid ring), 22.59 (C19), 20.86, 17.03 (C21), 12.28 (C18).

(344) MS (+APCI) m/z=595.4461

(345) IR (KBr) ν=3368, 3053, 2922, 2862, 2361, 1788, 1706, 1648 (C═O), 1536 cm.sup.−1.

Synthesis of N-[2-[1-(3-phenylpropyl) pyrrolidin-1-ium-1-yl]ethyl]cholanamide bromide (51)

(346) ##STR00052##

(347) N-(2-pyrrolidin-1-ylethyl)cholanamide (10) (0.5 g, 1 mmol) was dissolved in DCM (15 mL) along with 1-bromo-3-phenylpropane (2 mL, 10 mmol). The reaction was left for 12 hours where the product precipitated out. It was collected by filtration, washed with chloroform (10 mL) and dried at room temperature under vacuum. The product was a yellow solid.

(348) Yield=0.986 g (98%).

(349) Melting point: 140.8-141.3° C.

(350) .sup.1H NMR (DMSO) δ ppm: 0.57 (s, 3H, Me-18) 0.77 (s, 3H, Me-19) 0.92 (d, J=5.9 Hz, 3H, Me-21) 1.0-2.43 (m, steroid structure) 2.68, (dt, J=10.9, 7.4 Hz, 2H, CH.sub.2) 3.16-3.58 (m, 9H, CH-30H, CH.sub.2) 3.60 (s, 1H, CH-70H) 3.77 (s, 1H, CH-120H) 7.12-7.36 (m, 4H, aromatic) 8.11 (t, J=5.5 Hz, 1H, NH).

(351) .sup.13C NMR (DMSO) δ ppm: 173.32 (C═O), (140.52, 128.37, 128.37, 128.33, 125.98 aromatic ring), 79.15, 70.97 (C12), 70.38 (C3), 66.20 (C7), 63.84 (CH.sub.2), 61.68 (CH.sub.2), 47.42, (45.98, 45.69, 45.68, 40.48, 40.14, 39.81, 39.48, 39.14, 38.81, 38.47, 35.10, 34.86, 34.35, 33.85, 33.58, 33.36, 32.25, 31.39, 30.34, 28.53, 27.27, 26.20 steroid ring), 34.34 (CH.sub.2), 22.59 (C19), 20.90, 17.04 (C21), 12.30 (C18).

(352) MS (+APCI) m/z=Found 519.4150; expected 623.478 (MI− CH.sub.2 CH.sub.2C.sub.6H.sub.6)

(353) IR (KBr) ν=3387, 3256, 3068, 3026, 2929, 2868, 1651, 1624 (C═O), 1533 cm.sup.−1.

Synthesis of [1-(2-cholanamidoethyl) pyrrolidin-1-ium-1-yl] methyl 2,2-dimethylpropanoate chloride (52)

(354) ##STR00053##

(355) N-(2-pyrrolidin-1-ylethyl)cholanamide (10) (0.5 g, 1 mmol) was dissolved in DCM (10 mL) along with chloromethyl pivalate (1.4 mL, 10 mmol). The reaction was left for 72 hours where the product precipitated out. It was collected by filtration, washed with chloroform and (15 mL) dried at room temperature under vacuum. The product was a yellow solid.

(356) Yield=0.335 g (34%).

(357) Melting point: 121.2-121.8° C.

(358) .sup.1H NMR (DMSO) δ ppm: 0.68 (s 3H Me-18) 0.80 (s 3H Me-19) 0.91 (d 3H Me-21) 1.0-2.439 (m steroid structure) 3.38 (t J=9.1 Hz 1H CH-3) 3.87 (s 1H CH-70H) 4.03 (s 1H CH-120H) 8.66 (s 1H NH) 11.39 (s 1H).

(359) .sup.11C NMR (DMSO) δ 206.43, 179.27, 175.20 (C═O), 173.36, 173.14, 77.70, 70.93 (C12), 70.37 (C3), 69.58, 66.19 (C7), 60.88, 57.80, 53.07 (CH.sub.2), 52.88 (CH.sub.2 ring), (45.71, 37.66, 35.16, 34.85, 34.34, 33.32, 33.29, 32.16, 31.29, 30.65, 30.65, 28.50, 26.97, 26.97, 26.88, 26.42, 26.31, 26.20 steroid ring), 22.57 (C19), 21.81 (CH.sub.2 ring), 17.07 (C21), 12.28 (C18)

(360) MS (+APCI) m/z=619.4647

(361) IR (KBr) ν=3368, 2929, 2874, 2607, 2476, 2249, 2118, 1754, 1709, 1651 (C═O), 1539 cm.sup.−1.

Synthesis of N-(1-methyl-1-phenyl-piperidin-1-ium-4-yl) cholanamide iodide (53)

(362) ##STR00054##

(363) N-(1-phenyl-4-piperidyl) cholanamide (13) (0.5 g, 1 mmol) was dissolved in DCM (15 mL) along with methyl iodide (1.4 mL, 10 mmol). The reaction was left for 72 hours where the product precipitated out. It was collected by filtration, washed with DCM (20 mL) and dried at room temperature under vacuum. The product was a yellow solid.

(364) Yield=0.7 g (70%).

(365) Melting point: 165.8-166.7° C.

(366) .sup.1H NMR (D.sub.2O) δ ppm: 0.58 (s, 3H, Me-18) 0.79 (s, 3H, Me-19) 0.92 (d, 3H, Me-21) 1.0-2.43 (m, steroid structure) 2.82-3.18 (m, 7H) 3.43 (m, 1H, CH-3) 3.60 (s, 1H, CH-7) 3.78 (s, 1H, CH-12) 4.38-4.54 (m, 2H, CH.sub.2) 4.60 (m, 2H, CH.sub.2) 7.53 (m, 5H, aromatic) 7.79-7.90 (m, 1H, NH).

(367) .sup.13C NMR (DMSO) δ ppm: 172.24 (C═O), (133.08, 128.86, 127.54 aromatic ring), 35.10, 34.35, 22.60 (C19), 17.12 (C21), 12.31 (C18).

(368) MS (+APCI) m/z=Found 595.4464; calculated for C.sub.36H.sub.57N.sub.2O.sub.4 595.4469; 0.84 ppm.

(369) IR (KBr) ν=3396, 2925, 2862, 2355, 1648 (C═O), 1533 cm.sup.−1.

Synthesis of N-(1-propyl-1-phenyl-piperidin-1-ium-4-yl) cholanamide iodide (54)

(370) ##STR00055##

(371) N-(1-phenyl-4-piperidyl) cholanamide (13) (0.5 g, 1 mmol) was dissolved in DCM (15 mL) along with methyl iodide (1.5 mL, 10 mmol). The reaction was left for 72 hours where the product precipitated out. It was collected by filtration, washed with DCM (15 mL) and dried at room temperature under vacuum. The product was a yellow solid.

(372) Yield=0.7 g (70%).

(373) Melting point: 113-113.8° C.

(374) .sup.1H NMR (DMSO) δ ppm: 0.58 (d J=4.1 Hz 3H Me-18) 0.80 (s 3H Me-19) 0.91 (m 9H Me-21+2 CH.sub.3) 1.0-2.439 (m steroid structure) 2.08 (s CH.sub.2) 3.01-3.33 (m 9H CH.sub.2) 3.61 (s 1H CH-70H) 3.78 (s 1H CH-120H) 3.92-4.11 (m 2H) 4.60 (d J=10.3 Hz 1H) 7.42-7.62 (m 6H aromatic ring)

(375) Carbon=N/A

(376) MS (+APCI) m/z=Found 651.5086; expected 609.4625 (MI+ 52)

(377) IR (KBr) ν=3393, 2932, 2862, 2361, 2155, 2018, 1642 (C═O), 1533 cm.sup.−1.

Synthesis of N-(1-hexyl-1-phenyl-piperidin-1-ium-4-yl)cholanamide iodide (57)

(378) ##STR00056##

(379) N-(1-phenyl-4-piperidyl) cholanamide (13) (0.5 g, 1 mmol) was dissolved in DCM (15 mL) along with 1-iodohexane (1.5 mL, 10 mmol). The reaction was left for 120 hours where the product did not precipitate. The solvent was removed under vacuum and the product was sonicated then washed with diethyl ether (10 mL). The product was dried at room temperature under vacuum and it a yellow solid.

(380) Yield=0.55 g (55%).

(381) Melting point: 126.4-127.8° C.

(382) .sup.1H NMR (CDCl.sub.3) δ ppm: 0.66 (s, 3H, Me-18) 0.89 (s, 3H, Me-19) 0.79 (d, Me-21) 1.0-2.43 (m, steroid structure) 2.04 (s) 2.83 (m) 3.47 (m, 1H, CH-3) 3.56 (m, CH.sub.2) 3.81 (s, 1H, CH-7) 3.94 (s, 1H, CH-12) 6.06 (d, J=7.8 Hz, 1H, NH) 7.32 (q, J=4.3, 3.7 Hz, 6H, aromatic)

(383) .sup.13C NMR (DMSO) δ ppm: 206.44, 171.82 (C═O), (128.73, 128.16, 126.95 aromatic ring), 70.97 (C12), 70.46 (C3), 66.20 (C7), 51.90 (CH.sub.2), (46.11, 45.69, 40.49, 40.16, 35.30, 34.86, 34.35, 34.35, 32.56, 31.72, 30.66, 29.68, 28.53, 27.30, 26.18 steroid ring), 22.59 (C19), 17.11 (C21), 12.31 (C18).

(384) MS (+APCI) m/z=Found 665.5257; calculated for C.sub.41H.sub.67N.sub.2O.sub.4 665.5252; 0.77 ppm.

(385) IR (KBr) ν=3372, 3296, 3056, 3032, 2932, 2862, 2364, 1642 (C═O), 1530 cm.sup.−1.

Synthesis of N-(1-benzyl-1-phenyl-piperidin-1-ium-4-yl)cholanamide (58)

(386) ##STR00057##

(387) N-(1-phenyl-4-piperidyl) cholanamide (13) (0.5 g, 1 mmol) was dissolved in DCM (15 mL) along with benzyl bromide (1.5 mL, 10 mmol). The reaction was left for 12 days where the product precipitated. The product washed with diethyl ether (10 mL). The product was dried at room temperature under vacuum and it a peach solid.

(388) Yield=0.1839 g (18%).

(389) Melting point: 150.3-150.9° C.

(390) .sup.1H NMR (MeOD) δ ppm: 0.68 (s, 3H, Me-18) 0.90 (s, 3H, Me-19) 0.98 (d, 3H, Me-21) 1.0-2.43 (m, steroid structure) 3.68 (m, 1H, CH-3) 3.78 (s, 1H, CH-7) 3.92 (s, 1H, CH-12) 4.49 (s, 2H, CH.sub.2) 7.19-7.67 (m, 11H, aromatics)

(391) .sup.13C NMR (DMSO) δ ppm: (133.56, 133.08, 130.33, 129.23, 127.46, 127.12, 126.37 aromatic ring), 70.96 (C12), 45.69, 34.35 (CH.sub.2), 27.31, 24.50, 22.59 (C19), 17.04 (C21), 12.30 (C18), 5.85, 1.58.

(392) MS (+APCI) m/z=Found 671.4770; expected 657.4625 (MI+CH.sub.3)

(393) IR (KBr) ν=3411, 3329, 3062, 2929, 2859, 2671, 2355, 1654 (C═O), 1536, 1493 cm.sup.−1.

Synthesis of (4-cholanamido-1-phenyl-piperidin-1-ium-1-yl)methyl 2,2-dimethylpropanoate chloride (59)

(394) ##STR00058##

(395) N-(1-phenyl-4-piperidyl) cholanamide (13) (0.5 g, 1 mmol) was dissolved in chloroform (15 mL) along with of chloromethyl pivalate (1.4 mL, 10 mmol). The reaction was left for 12 days where the product did not precipitate. The solvent was taken off under vacuum and the product sonicated and washed with diethyl ether (10 mL). The product was dried at room temperature under vacuum and it a yellow solid.

(396) Yield=0.5215 g (52%).

(397) Melting point: 135.9-137° C.

(398) .sup.1H NMR (MeOD) δ ppm: 0.70 (s, 3H, Me-18) 0.92 (s, 3H, Me-19) 1.02 (d, 3H, Me-21) 1.0-2.439 (m, steroid structure) 2.99 (d, J=11.4 Hz, 1H) 3.78 (s, 1H, CH-7) 3.97 (s, 1H, CH-12) 5.75 (s, 2H, O—CH.sub.2—C1) 7.24-7.45 (m, 2H, aromatic) 7.55 (m, 1H, NH).

(399) .sup.13C NMR (DMSO) δ ppm: 206.44 (C═O), 175.60 (C═O), 128.69, 69.59 (CH.sub.2),) 45.69, 35.12, 34.35, 30.66, 26.97, 26.32 steroid ring), 22.59 (C19), 17.11 (C21), 12.30 (C18).

(400) MS (+APCI) m/z=Found 695.4994; calculated for C.sub.41H.sub.65N.sub.2O 6695.4993; 0.143 ppm.

(401) IR (KBr) ν=3365, 3065, 2935, 2865, 2631, 2525, 2361, 1751, 1642 (C═O), 1542 cm.sup.−1.

Synthesis of N-[1-phenyl-1-(3-phenylpropyl) piperidin-1-ium-4-yl]cholanamide bromide (60)

(402) ##STR00059##

(403) N-(1-phenyl-4-piperidyl) cholanamide (13) (0.5 g, 1 mmol) was dissolved in chloroform (15 mL) along with 1-bromo-3-phenylpropane (1.3 mL, 10 mmol). The reaction was left for 12 days where the product did not precipitate. The solvent was taken off under vacuum and the product sonicated and washed with diethyl ether (10 mL). The product was dried at room temperature under vacuum and it a yellow solid.

(404) Yield=0.79 g (80%).

(405) Melting point: 126.1-126.7° C.

(406) .sup.1H NMR (MeOD) δ ppm: 0.69 (s, 3H, Me-18) 0.91 (s, 3H, Me-19) 0.98 (d, 3H, Me-21) 1.0-2.43 (m, steroid structure) 3.43 (m) 3.66 (m, 1H, CH-3) 3.79 (s, 1H, CH-7) 3.94 (s, 1H, CH-12) 7.09-7.41 (m, 7H, aromatic)

(407) .sup.13C NMR (DMSO) δ ppm: (140.52, 128.85, 128.43, 128.37, 128.33, 125.98 aromatic ring), 70.99 C12), 70.38 (C3), 66.20 (C7), (45.95, 45.69, 45.69, 40.49, 40.15, 35.27, 34.89, 33.36, 31.62, 30.38, 28.52, 27.31, 26.19 steroid ring), 34.61 (CH), 34.36 (CH.sub.2), 33.86 (CH.sub.2), 22.59 (C19), 17.12 (C21), 12.31 (C18).

(408) MS (+APCI) m/z=Found 699.5087; calculated for C.sub.44H.sub.65N.sub.2O.sub.4 695.5095; 1.143 ppm.

(409) IR (KBr) ν=3375, 3023, 2932, 2859, 1642 (C═O), 1536 cm.sup.−1.

Synthesis of N-(1-allyl-1-phenyl-piperidin-1-ium-4-yl) cholanamide bromide (61)

(410) ##STR00060##

(411) N-(1-phenyl-4-piperidyl) cholanamide (13) (0.5 g, 1 mmol) was dissolved in DCM (15 mL) along with allyl bromide (1.2 mL, 10 mmol). The reaction was left for 4 days where the product precipitated then washed with DCM (25 mL). The product was dried at room temperature under vacuum and to give an orange solid.

(412) Yield=0.242 g (24%).

(413) Melting point: 161.4-161.7° C.

(414) .sup.1H NMR (CDCl.sub.3) δ ppm: 0.58 (d, J=3.6 Hz, 2H, Me-18) 0.81 (s, 2H, Me-19) 0.93 (d, J=5.8 Hz 3H, Me-21) 1.0-2.43 (m steroid structure) 3.18, 3.61 (s, 1H, CH-3), 3.78 (s 1H CH-7), 3.95 (dd, J=16.7, 9.8 Hz 1H CH-12) 4.08 (s) 4.31 (s 2H CH.sub.2) 4.55-4.68 (m C═CH) 5.61-5.83 (m C═CH) 7.42-7.62 (m 6H aromatic ring)

(415) .sup.13C NMR (DMSO) δ ppm: 187.15 (C═O), 133.09 (C═C), (131.49, 130.30, 128.97, 127.16 aromatic ring), 92.56 (C═C), 70.96 (C12), 70.37 (C3), 66.19 (C7), 55.64 (CH.sub.2), (46.01, 41.48, 40.50, 40.16, 39.83, 39.49, 39.15, 38.82, 38.74, 38.50, 37.34, 35.27, 30.66, 28.56 steroid ring), 22.59 (C19), 17.13 (C21), 12.30 (C18).

(416) MS (+APCI) m/z=Found 621.4620; calculated for C.sub.38H.sub.59N.sub.2O.sub.4 621.4626; 0.96 ppm.

(417) IR (KBr) ν=3378, 3056, 2935, 2856, 2552, 2358, 1639 (C═O), 1533 cm.sup.−1.

Synthesis of 3α, 7, 12α-triacrylate cholic acid methyl ester (62)

(418) ##STR00061##

(419) Methyl cholate (4.22 g, 10 mmol) was dissolved in dry chloroform (30 mL). Triethylamine (2.23 mL, 16 mmol) was added and the flask was cooled to ice temperature. Acryloyl chloride (1.3 mL, 15 mmol) in chloroform (10 mL) was dripped in over 30 minutes. The mixture was protected from the light and left at room temperature overnight. More acryloyl chloride (1.3 mL, 15 mmol) in dry chloroform (10 mL) was dripped in over 30 minutes and the reaction was left for 6 hours at room temperature. The flask was maintained at −20° C. for 5 days. More acryoyl chloride (1.5 mL) was added. The reaction was left for 24 hours at room temperature. The solvent was removed using a rotary evaporator (the temperature was set to 30° C.) to produce a white solid. Ethyl acetate (50 mL) was added and the solid (salt) was collected by filtration. The ethyl acetate was removed on the rotary evaporator to leave a yellow oil. Flask column chromatography using 95% DCM/5% ethyl acetate was used to purify the product, increasing to 9/1 ratio then 3/1 ratio. The solvents were removed by rotary evaporation before the product was dried at room temperature under vacuum.

(420) Yield=0.2176 g (5%).

(421) .sup.1H NMR (CDCl.sub.3) δ ppm: 0.75 (s 2H Me-18) 0.83 (s 2H Me-19) 0.94 (d 3H Me-21) 1-2.43 (steroid structure) 2.67-2.91 (m 2H) 3.60-3.86 (m 1H) 4.63 (m 1H CH-3) 5.03 (s 1H CH-7) 5.19 (s 1H CH-12) 5.72-5.97 (m 2H C═CH) 5.99-6.52 (m, 3H C═CH).

(422) .sup.13C NMR (CDCl.sub.3) δ ppm: 174.48 (C═O), 169.70 (C═O), 169.38 (C═O), 169.20 (C═O), 165.67 (C═O excess), 165.46 (C═O excess), 165.38 (C═O excess), 165.29, 165.23, 136.42 (C═C), 130.55 (C═C), 130.42, 130.26, 130.18, 129.12, 129.00, 75.43, 74.62, 74.08, 73.92, 70.94 (C12), 70.85 (C3), 51.48, (47.44, 45.23, 45.12, 43.40, 43.32, 43.22, 40.84, 40.74, 39.32, 39.20, 39.14, 38.22, 38.00, 37.87, 34.75, 34.62, 34.55, 34.43, 34.35, 31.29, 30.91, 30.85, 30.74, 28.82, 28.60, 27.17, 26.74, 26.65 steroid ring), 25.46, 25.16, 22.84, 22.5 (C19), 22.34, 17.53 (C21), 17.45, 12.18 (C18), 12.08.

(423) MS (+APCI) m/z=Found 602.1935; expected 584.3349 (MI+CH.sub.3+3H).

(424) IR (KBr) ν=2947, 271, 1715, 1633 (C═O), 1612, 1469 cm.sup.−1.

Synthesis of 3α,12α diacroylate-7αhydroxycholic acid (63)

(425) ##STR00062##

(426) This product was collected by column chromatography of the previous product (3α,7α,12α-triacrylate cholic acid methyl ester).

(427) Yield=0.7744 g (18%).

(428) .sup.1H NMR (CDCl.sub.3) δ ppm: 0.68 (s 2H Me-18) 0.84 (s 2H Me-19) 0.95 (d 3H Me-21) 1-2.43 (Steroid structure) 2.66-2.93 (m 1H) 3.61-3.93 (s 3H CH.sub.3) 4.12 (q, J=7.1 Hz, 1H ethyl acetate), 4.64 (tt, J=10.7, 6.5 Hz, 1H CH-3) 4.99 (s 1H CH-7) 5.18 (s 1H CH-12) 5.34 (m 1H) 5.70-5.93 (m 1H C═CH) 5.98-6.52 (m 2H C═CH).

(429) .sup.13C NMR (CDCl.sub.3) δ ppm: 174.55 (C═O), (169.68, 169.57, 165.78, 165.71, 165.57, 163.18 C═O, including excess acyloyl chloride), (130.66, 130.54, 130.30, 130.19, 130.08, 129.12, 129.00, 128.87 C═C, including excess acyloyl chloride), 75.63, 74.40, 74.28, 74.17, 72.57, 71.79, 71.02 (C12), 67.93 (C7), 60.36, 51.46, 47.46, 4?. 11, (46.56, 45.20, 45.06, 43.55, 43.40, 42.08, 42.02, 41.18, 41.12, 40.90, 40.84, 39.23, 38.23, 38.10, 35.23, 35.16, 34.96, 34.73, 34.61, 34.52, 34.34, 31.31, 30.98, 30.81, 28.60, 28.17, 28.10, 27.69, 27.51, 27.24, 26.64 steroid ring), 25.51, 25.36, 22.90, 22.53 (C19), 17.41 (C21), 17.32, 12.50, 12.26 (C18), 12.19.

(430) MS (+APCI) m/z=Found 548.1638; expected 531.3322 (MI+CH.sub.3+2H).

(431) IR (KBr) ν=35 26 2944, 2871, 1715, 1639 (C═O), 1612, 1469 cm.sup.−1.

Synthesis of 3α acetate-7α,12α dihydroxy cholic acid (64)

(432) ##STR00063##

(433) This product was collected by column chromatography of the product 3α,7α,12α-triacrylate cholic acid methyl ester.

(434) Yield=0.0232 g

(435) .sup.1H NMR (CDCl.sub.3) δ ppm: 0.70 (s 3H Me-18) 0.82 (s 3H Me-19) 0.94 (d 3H Me-21) 1-2.43 (steroid structure) 3.67 (s 3H CH.sub.3) 3.87 (s 1H CH-7) 4.11 (s 1H CH-12) 5.78 (dd J=10.3, 1.7 Hz, 1H C═CH) 6.07 (dd J=17.3, 10.3 Hz 1H C═CH) 6.37 (dd J=17.3, 1.7 Hz, 1H).

(436) .sup.13C NMR (CDCl.sub.3) δ ppm: 217.85, 216.22, 215.83, 215.45, 214.86, 214.47, 173.94, 73.08, 71.85 (C12), 68.27 (C7), 56.87, (46.52, 45.30, 41.74, 41.49, 40.08, 39.58, 38.83, 35.37, 34.80, 33.50, 33.13, 31.74, 30.41, 29.10, 28.16, 27.62, 26.43, 26.10 steroid ring), 25.01, 23.32, 22.69, 22.51 (C19), 17.57 (C21), 14.17, 12.50 (C18), 11.48.

(437) MS (+APCI) m/z=Found 494.16; expected 476.3138 (MI+CH.sub.3+3H).

(438) IR (KBr) ν=3375, 2959, 2932, 2865, 1712, 1630 (C═O), 1618, 1533 cm.sup.−1.

Synthesis of 3α methacrylate 7α,12α dihydroxyl cholic acid methyl ester (65)

(439) ##STR00064##

(440) Methyl cholate (4.22 g, 10 mmol) was dissolved in dry chloroform (30 mL). Triethylamine (2.23 mL, 16 mmol) was added and the flask was put on ice. Methacryloyl chloride (1.45 mL, 15 mmol) in chloroform (10 mL) was dripped in over 30 minutes. The mixture was protected from the light and left at room temperature overnight. The solvent was removed using a rotary evaporator (the temperature was set to 30° C.) to produce a white solid. Ethyl acetate (50 mL) was added and the solid (salt) was collected by filtration. The ethyl acetate was removed on the rotary evaporator to leave a white solid. Flask column chromatography using 75% DCM/25% ethyl acetate was used to purify the product. The solvents were removed by rotary evaporation before the product was dried at room temperature under vacuum.

(441) .sup.1H NMR (CDCl.sub.3) δ ppm: 0.70 (s 3H Me-18) 0.83 (s 3H Me-19) 0.98 (d 3H Me-21) 1-2.43 (steroid structure) 3.77 (s 3H CH.sub.3) 3.82 (d, J=26.9 Hz 1H CH-7) 4.10 (s 1H CH-12) 4.63 (tt J=11.2, 4.4 Hz, 1H CH-3) 5.5 (m 1H C═CH) 5.83 (m 1H C═CH) 6.07 (s 1H) 6.25 (s 1H).

(442) .sup.13C NMR (CDCl.sub.3) δ ppm: 218.61, 174.68 (C═O), 167.10 (C═O), 136.92 (C), 124.92 (C═C), 74.54, 72.92 (C12), 68.26 (C7), 51.52, 47.26, (46.57, 42.15, 41.23, 39.57, 35.22, 35.13, 34.90, 34.71, 34.42, 31.05, 30.89, 28.44, 27.42, 26.83, 26.69 steroid ring), 23.14, 22.57 (C19), 18.35 (CH.sub.3), 17.37 (C21), 12.57 (C18).

(443) MS (+APCI) m/z=Found 535.3181; expected 476 (MI+CH.sub.3CN+H.sub.2O)

(444) IR (KBr) ν=3599, 3544, 2971, 2935, 2865, 1733, 1703, 1639 (C═O), 1469 cm.sup.−1.

Synthesis of N-[2-[1-[(4-vinylphenyl)methyl]pyrrolidin-1-ium-1-yl]ethyl]cholanamide chloride (69)

(445) ##STR00065##

(446) N-(2-pyrrolidin-1-ylethyl) cholanamide (10) (0.35 g, 0.7 mmol) was dissolved in chloroform (10 mL). Vinyl benzyl (0.5 mL, 3.5 mmol) chloride was added. The reaction was protected from the light and left to stir overnight. The reaction was heated under reflux for 24 hours. The solvent was removed under reduced pressure before the product was dried at room temperature under vacuum.

(447) .sup.1H NMR (CDCl.sub.3) δ ppm: 0.58 (s 3H Me-18) 0.84 (s 3H Me-19) 0.93 (d 3H Me-21) 1-2.43 (steroid ring) 3.11-3.24 (m, 8H 2 CH.sub.2), 3.60 (m, 1H CH-3), 3.77 (s, 1H CH-7), 3.95 (m, 1H CH-12), 4.35 (d, J=4.0 Hz, 2H CH.sub.2), 4.54 (s, 2H CH.sub.2), 5.38 (d, J=10.9 Hz, 1H C═CH), 5.96 (d, J=17.6 Hz, 1H C═CH), 6.80 (dd, J=17.7, 11.0 Hz, 1H), 8.23 (d, J=5.2 Hz, 1H NH).

(448) .sup.13C NMR (DMSO) δ ppm: 173.40 (C═O), (145.12, 138.80, 132.94, 127.83, 126.57, 104.54 aromatic ring), 135.74 (C═C), 116.15 (C═C), 79.52, 78.99, 78.46, 70.94 (C12), 70.39 (C3), 67.05, 66.21 (C7), 61.14, 60.78, 57.11 (CH.sub.2), 48.55, (45.95, 45.72, 41.48, 41.38, 40.50, 40.42, 40.17, 35.28, 35.13, 34.86, 34.34, 33.10, 32.23, 31.42, 28.51, 27.30, 26.19 steroid ring), 22.58 (C19), 20.86, 17.05 (C21), 14.56, 12.27 (C18).

(449) MS (+APCI) m/z=621.4614

(450) IR (KBr) ν=3350, 3059, 2932, 2862, 1703, 1651 (C═O), 1533 cm.sup.−1.

Synthesis of 3-acetamidopropyl-dimethyl-[(4-vinylphenyl)methyl]ammonium chloride (70)

(451) ##STR00066##

(452) N-[3-dimethylamino) propyl] cholanamide (12) (0.35 g, 0.7 mmol) was dissolved in chloroform (10 mL). Vinyl benzyl chloride (0.5 mL, 3.5 mmol) was added. The reaction was protected from the light and left to stir overnight. The reaction was heated under reflux for 24 hours. The product precipitated out, collected by filtration and washed with petrol 40-60 (30 mL). Product not pure.

(453) .sup.1H NMR (CDCl.sub.3) δ ppm: 0.65 (s 3H Me-18) 0.87 (s 3H Me-19) 0.95 (d 3H Me-21) 1-2.43 (steroid structure) 3.49 (s, 1H CH-12), 3.69 (d, J=19.8 Hz, 1H CH-7), 3.90 (s, 1H CH-3), 5.38 (dd, J=11.0, 5.9 Hz, 1H CH═CH), 5.83 (dd, J=17.7, 4.8 Hz, 1H CH═CH), 6.71 (dd, J=17.6, 10.9 Hz, 1H), 7.30-7.63 (m, 4H aromatic ring), 8.49 (s, 1H NH).

(454) .sup.13C NMR (DMSO) δ ppm: 172.97 (C═O), (138.80, 135.76, 133.21, 127.38, 126.40 aromatic ring), 116.09 (C═C), 79.52, 78.99, 78.46, 70.97 (C12), 70.39 (C3), 66.20 (C7), 65.85, 61.35, 59.67 (CH.sub.3), 49.22, 48.55, (45.92, 41.48, 41.36, 40.51, 40.18, 35.51, 35.27, 35.18, 34.87, 34.35, 32.31, 28.51, 27.28, 26.20 steroid ring), 45.70 (CH.sub.3), 31.50 (CH.sub.2), 22.79, 22.68, 22.58 (C19), 17.11 (C21), 14.61, 12.29 (C18).

(455) MS (+APCI) m/z=609.4619

(456) IR (KBr) ν=3347, 2929, 2862, 1775, 1694, 1627 (C═O), 1551 cm.sup.−1.

Synthesis of N-[1-phenyl-1-[(4-vinylphenyl)methyl]piperidin-1-ium-4-yl]cholanamide chloride (71)

(457) ##STR00067##

(458) N-(1-phenyl-4-piperidyl) cholanamide (13) (1.5 g, 3.5 mmol) was dissolved in DCM (20 mL). Vinyl benzyl chloride (1.5 mL, 10.5 mmol) was added. The reaction was protected from the light and left to stir overnight. The reaction was heated under reflux for 24 hours. The product precipitated out, which was then collected by filtration and washed with petrol 40-60 (30 mL). Further purification by solvent extraction (×3) between petrol 40-60 (10 mL) and methanol (10 mL) left a white solid.

(459) .sup.1H NMR (CDCl.sub.3) δ ppm: 0.57 (s32H Me-18) 0.88 (s 3H Me-19) 0.98 (d 3H Me-21) 1-2.43 (steroid structure) 3.18 (tt, J=14.7, 7.3 Hz, 7H CH.sub.2 ring), 3.61 (d, J=3.7 Hz, 1H CH-3), 3.77 (d, J=3.6 Hz, 1H CH-7), 4.04 (d, J=3.2 Hz, 1H CH-12), 4.14 (d, J=3.3 Hz, 1H C3-OH), 4.36 (d, J=4.0 Hz, 1H C7-OH), 4.53 (s, 1H C12-OH), 5.38 (d, J=10.9 Hz, 1H CH═CH), 5.95 (d, J=17.7 Hz, 1H CH═CH), 6.79 (dd, J=17.6, 11.0 Hz, 1H), 7.73-7.43 (m, 4H aromatic ring), 8.07 (t, J=5.7 Hz, 1H NH).

(460) .sup.13C NMR (DMSO) δ ppm: 172.94 (C═O), (138.80, 135.78, 133.22, 127.43, 126.42, aromatic ring), 116.15 (C═C), 70.95 (C12), 70.37 (C3), 66.19 (C7), 65.83, 61.33, 49.24, (45.94, 45.70, 41.48, 41.36, 40.51, 40.17, 35.50, 35.28, 35.19, 34.87, 34.35, 32.32, 31.51, 30.35, 28.54, 27.29, 26.20 steroid ring), 22.79, 22.67, 22.60 (C19), 17.12 (C21), 12.31 (C18).

(461) MS (+APCI) m/z=Found 609.4612; expected 683.4782 (MI−73) MI− (vbc+propane)

(462) IR (KBr) ν=3356, 2925, 2862, 2164, 1648 (C═O), 1548 cm.sup.−1.

Synthesis of N-Boc-1,4-butanediamine (72)

(463) ##STR00068##

(464) 1,4-diaminobutane (1 mL, 12.94 mmol) was dissolved in DCM (20 mL) and put on ice. Di-tert-butyl dicarbonate (0.3 g, 1.29 mmol) in DCM (5 mL) was dripped in over 50 minutes. The reaction was left at ice temperature for 12 hours. The reaction was washed with water (100 mL) followed by brine (100 mL), before being dried with magnesium sulphate. The solvent was removed under vacuum to leave a yellow solid.

(465) Yield=0.087 g (8.7%).

(466) .sup.1H NMR (CDCl.sub.3) δ ppm: 1.40-1.61 (m 12H) 3.13 (q J=6.1 Hz 2H) 4.55 (s 1H).

(467) .sup.13C NMR (DMSO) δ ppm: 155.57 (C═O), 77.26 (C), 28.36 (CH.sub.3), 28.21, 27.29, 26.93, 26.86.

(468) MS (+APCI) m/z=Found 289.2126; expected 188. Diamer produced.

(469) IR (KBr) ν=3372 2983 2941 2847 1681 (C═O) 1521 cm.sup.−1.

Polymerisation of N-(4-cholanamidobutyl)-2-methyl-prop-2-enamide

(470) N-(4-cholanamidobutyl)-2-methyl-prop-2-enamide (75) (0.6 g, 0.96 mmol) was dissolved in methanol (7 mL). Styrene (2.4 mL, 23.04 mmol) was added along with a small amount of AIBN. The boiling tube was purged with argon 3 times and put into an oil bath at 60° C. for 48 hours. The white solid precipitate was collected by filtration, dissolved into chloroform and dripped onto stirring methanol (50 mL) for purification. The polymer was collected by filtration, washed with methanol (30 mL) and dried at room temperature under vacuum.

Polymerisation of 3α,12α diacroylate-7αhydroxycholic acid

(471) 3α,12α diacroylate-7αhydroxycholic acid (63) (0.175 g) was dissolved in toluene (2.5 mL). AIBN (0.1 g) was added, along with EGDMA (0.82 mL). 1000 of the solution was pipetted out into a 96 well plate and a PTFE-lined cover was clamped across the top of the plate to seal the wells. It was place in at oven set at 60° C. for 12 hours. The products were white discs.

Polymerisation of methacrylate 7α,12α dihydroxyl cholic acid

(472) Methacrylate 7α,12α dihydroxyl cholic acid (65) (0.2 g) was dissolved in toluene (3.5 mL). AIBN (0.1 g) was added, along with EGDMA (0.8 mL). 1000 of the solution was pipetted out into a 96 well plate and clamped. It was place in at oven set at 60° C. for 12 hours. The products were white discs.

Polymerisation of 3α hydroxyl 7α,12α diacetate methyl cholate

(473) 3α hydroxyl 7α,12α diacetate methyl cholate (77) (0.5 g) was added to a round bottomed flask along with bis[acetylacetonato]copper (0.02 g) and vinyl benzyl chloride (2 mL). The reaction was heated to 120° C. for 4.5 hours. The crude product was purified using column chromatography with petrol 60-80, slowly increasing the amount of ethyl acetate (0-100%). The solvents were removed under reduced pressure and the polymer was dried at room temperature under vacuum.

Polymerisation of 3-acetamidopropyl-dimethyl-[(4-vinylphenyl)methyl]ammonium chloride (70)

(474) 3-acetamidopropyl-dimethyl-[(4-vinylphenyl)methyl]ammonium chloride (70) (0.5 g) was dissolved in ethanol (4 mL). Styrene (3.9 mL) was added along with AIBN (100 mg). The reaction was degassed and heated to 65° C. for 12 hours where a white solid precipitated out. It was purified by dissolving in chloroform (20 mL) and dripping into stirring methanol (100 mL). The product was collected by filtration and dried at room temperature under vacuum to give a white solid.

(475) Proton NMR analysis shows very little incorporation of N-[3-(dimethylamino)propyl]cholanamide (12) into the polymer.

(476) 3-acetamidopropyl-dimethyl-[(4-vinylphenyl)methyl]ammonium chloride (70) (0.25 g) was dissolved in ethanol (4 mL) along with AIBN (100 mg). Tert-butyl methacrylate (0.23 mL) was added. The mixture was degassed which argon and heated to 60° C. for 4 days. The solution was purified by dripping onto ethyl acetate (75 mL) to give a white powdery solid which was dried at room temperature under vacuum.

(477) Proton NMR analysis showed that no polymerisation had taken place.

(478) 3-acetamidopropyl-dimethyl-[(4-vinylphenyl)methyl]ammonium chloride (70) (0.25 g) was suspended in propan-2-ol (6 mL) along with AIBN (100 mg). The mixture was degassed and heated to 70° C. for 5 days. The white, cloudy solution was collected by filtration to give a off white solid which was dried at room temperature under vacuum.

(479) No NMR analysis could be undertaken of this polymer due to its insolubility.

Polymerisation of Synthesis of N-[1-phenyl-1-[(4-vinylphenyl)methyl]piperidin-1-ium-4-yl]cholanamide chloride (71)

(480) Synthesis of N-[l-phenyl-1-[(4-vinylphenyl)methyl]piperidin-1-ium-4-yl]cholanamide chloride (70) (0.3 g) was dissolved in proan-2-ol (8 mL) along with AIBN (100 mg). The mixture was degassed with argon and heated to 70° C. for 6 days. An white solid precipitated out, which was collected by filtration and dried at room temperature under vacuum.

(481) Proton NMR analysis shows a very short chained polymer was synthesised.

Synthesis of 4-benzylbenzoyl chloride (96)

(482) ##STR00069##

(483) 4-benzyl benzoic acid (1 g, 4.42 mmol) was added to a dry flask along with thionyl chloride (3 mL), DMF (0.5 mL) and toluene (13 mL). The flask was heated under reflux for 5 days. The solvent was removed under reduced pressure and the product was re-dissolved in toluene (5 mL) twice and the solvent removed. The product was dried under vacuum at room temperature under vacuum to give a white solid.

(484) Yield=1.03 g

(485) .sup.1H NMR (CDCl.sub.3) δ ppm: 8.56, 8.54, 8.29, 8.24, 8.24, 8.23, 8.21, 8.20, 8.20, 8.19, 8.18, 7.93, 7.90, 7.89, 7.88, 7.87, 7.86, 7.86, 7.85, 7.84, 7.82, 7.81, 7.80, 7.80, 7.78, 7.78, 7.77, 7.67, 7.66, 7.65, 7.64, 7.63, 7.62, 7.62, 7.61, 7.60, 7.60, 7.59, 7.58, 7.54, 7.53, 7.52, 7.50, 7.50, 7.49, 7.48, 7.47, 7.47, 7.24, 6.82, 3.95, 3.47, 3.33, 2.15, 1.79, 0.217.83-7.73 (m, 2H), 7.96-7.80 (m, 2H), 8.30-8.13 (m, 2H),

(486) .sup.1H NMR (250 MHz, Chloroform-d) δ 7.69-7.56 (m, 1H), 7.50 (tt, J=6.6, 1.5 Hz, 2H).

(487) MS (+APCI) m/z=Found 245.0366; calculated for C.sub.14H.sub.10Cl.sub.1O.sub.2 245.0364; 0.9 ppm.

Synthesis of N-(4-benzoylphenyl)formamide cholate (98)

(488) ##STR00070##

(489) Cholic acid (0.5 g, 1.2 mmol) was dissolved in THE (30 mL) along with triethylamine (2.9 mL, 0.3 mmol). The solution was put on ice for 10 minutes before ethylchloroformate (0.13 mL, 0.013 mmol) was dripped in over 10 minutes. The solution was allowed to react for two hours at room temperature. 4-aminobenzophenone (0.23 g, 1.2 mmol) was added and left to react for 3 hours. The reaction was quenched with water (30 mL). The mixture was washed with water (3×30 mL). The organic layer was dried over magnesium sulphate and the solvent was evaporated under reduced pressure. Solvent extraction between water and ethyl acetate was preformed 3 times before the organic layer was removed under reduced pressure. The product was dried at room temperature under vacuum.

(490) MS (+APCI) m/z=Found 588.3680; calculated for C.sub.37H.sub.50N.sub.1O.sub.5 588.3684; −0.6 ppm.

Synthesis of methyl lithocholate (99)

(491) ##STR00071##

(492) Lithocholic acid (5.0 g, 0.01329 mol) was added to methanol (90 mL) to produce a suspension. Acetyl chloride (0.5 mL, 0.006 mol) was then added. The solution was heated and stirred at 80° C. for 40 minutes as a homogeneous solution, then allowed to cool overnight in an ice bath. This was added to water (150 mL) and the resulting precipitate was collected by filtration, washed with water (3×20 mL) and dried under vacuum.

(493) Yield; 5 g, 0.01328 mol, 96.5%.

(494) Melting point: 75-76° C.

(495) .sup.1H NMR (250 MHz) δ=0.66 (s, 3H, 18-CH.sub.3), 0.94 (s, 3H, 19-CH.sub.3), 3.65 (m, 1H, 3-CH), 3.69 (s, 3H, O—CH.sub.3) ppm.

(496) .sup.13C NMR (62.9 MHz) δ=12.0 (CH.sub.3, C18), 18.6 (CH.sub.3, C21), 20.8 (CH.sub.2, C11), 23.7 (CH.sub.3, C19), 24.2 (CH.sub.2, C15), 26.4 (CH.sub.2, C7), 27.2 (CH.sub.2, C6), 28.2 (CH.sub.2, C16), 30.5 (CH.sub.2, C2), 30.9 (CH.sub.2, C22), 31.0 (CH.sub.2, C23), 34.5 (C, C10), 35.3 (CH, C20), 35.4 (CH.sub.2, C1), 35.8 (CH, C8), 36.4 (CH.sub.2, C4), 40.1 (CH.sub.2, C12), 40.4 (CH, C9), 42.0 (CH, C5), 42.7 (C, C13), 51.5 (O—CH.sub.3) 55.9 (CH, C17), 56.5 (CH, C14), 71.9 (CH, C3), 174.8 (CO, C24) ppm.

(497) IR; 3347 (OH stretch), 2937 (C—H), 2859 (O—CH.sub.3), 1733 (C═O), 1640, 1436, 1206, 1043 (R.sup.2CH—OH) cm.sup.−1.

Synthesis of methyl deoxycholate (100)

(498) ##STR00072##

(499) Deoxycholic acid (5.0 g 0.01 mol) was dissolved in methanol (30 mL) and treated with acetyl chloride (0.5 mL, 0.006 mol). The solution was heated and stirred at 80° C. for 40 minutes then allowed to cool overnight in an ice bath. The resultant crystals were collected by vacuum filtration to produce a white crystalline powder. This was washed with water (2×20 mL) and dried under vacuum.

(500) Yield; 1.81 g, 0.004 mol, 35%.

(501) Melting point: 70-72° C.

(502) .sup.1H NMR (250 MHz) δ=0.68 (s, 3H, 18-CH.sub.3), 0.91 (s, 3H, 19-CH.sub.3), 3.60 (m, 1H, 3-CH), 3.67 (s, 3H, O—CH.sub.3), 3.99 (s, 1H, 12-CH) ppm.

(503) .sup.13C NMR (62.9 MHz) δ=12.8 (CH.sub.3, C18), 17.3 (CH.sub.3, C21), 23.2 (CH.sub.3, C19), 26.6 (CH.sub.2, C15), 26.1 (CH.sub.2, C7), 27.1 (CH.sub.2, C6), 27.4 (CH.sub.2, C16), 28.7 (CH.sub.2, C11), 30.5 (CH.sub.2, C2), 30.9 (CH.sub.2, C23), 31.1 (CH.sub.2, C22), 33.7 (CH, C9), 34.1 (C, C10), 35.1 (CH.sub.2, C1), 35.2 (CH, C20), 36.0 (CH.sub.2, C4), 36.4 (CH, C8), 42.0 (CH, C5), 46.5 (C, C13), 47.3 (CH, C17), 48.3 (CH, C14), 51.5 (O—CH.sub.3) 71.8 (CH, C3), 73.1 (CH.sub.2, C12), 174.7 (CO, C24) ppm.

(504) IR; 3474 (OH stretch) 2985 (C—H), 2852 (O—CH.sub.3), 1743 (C═O), 1450, 1380, 1040 (R.sub.2CH—OH) cm.sup.−1.

(505) .sup.13C NMR (62.9 MHz) δ=12.0 (CH.sub.3, C18), 18.2 (CH.sub.3, C21), 20.8 (CH.sub.2) 23.3 (CH.sub.3, C19), 24.2 (CH.sub.2, C15), 26.4 (CH.sub.2, C7), 27.1 (CH.sub.2, C6), 28.2 (CH.sub.2, C16), 30.5 (CH.sub.2, C1), 30.9 (CH.sub.2, C2), 31.1 (CH.sub.2, C23), 34.5 (CH, C9), 35.3 (C, C10), 35.8 (CH.sub.2, C1), 36.4 (CH, C20), 40.1 (CH, C5), 40.4 (C, C13), 55.9 (CH.sub.2), 56.5 (CH.sub.2), 61.3 (CH.sub.2) 65.9 (CH, C3), 71.8 (CH.sub.2, C12), 174.7 (CO, C24) ppm.

(506) IR=3509 (OH), 3309 (OH), 2925 (alkyl), 2857 (alkyl), 1718 (C═O), 1449, 1356, 1292, 1189.78 (C═O ester stretch), 1027.73 (R.sub.2CH—OH) cm.sup.−1.

(507) MS (+ESI) m/z=Found 454.3527 (M+H)+; calculated for C.sub.26H.sub.48NO.sub.5 454.3527; 0.0 ppm.

Synthesis of (4R)—N-(2-dimethylaminoethyl)-4-[(3R,10S,13R,17R)-3-hydroxy-10,13-dimeth-yl-2,3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenan-thren-17-yl]pentanamide (101)

(508) ##STR00073##

(509) A mixture of lithocholic acid (1.0 g 0.005 mol) and N, N′-dimethylethylenediamine (0.16 mL 0.001 mol) was dissolved in toluene (20 mL). The solution was heated at reflux for 24 hours. More N, N′-dimethylethylenediamine (0.32 mL 0.003 mol) of was added and the mixture was heated at reflux for 24 h. Water (100 mL) was added to the solution. The resulting precipitate was collected by vacuum filtration and the crude product was recrystallized from ethyl acetate to produce a white powder.

(510) Yield 0.85 g, 0.001 mol, 69%.

(511) .sup.1H NMR (250 MHz) CDCl.sub.3 5=0.64 (s, 3H, 18-CH.sub.3), 0.92 (s, 3H, 19-CH.sub.3), 2.21 (s, 6H, 2×CH.sub.3), 2.42 (t, 2H CH.sub.2 J=5.0) 3.34 (q, 2H, CH.sub.2, J=7.5), 3.63 (m, 1H, 3-CH), 6.20 (broad s, 1H, NH) ppm.

(512) .sup.13C NMR (62.9 MHz) δ=12.0 (CH.sub.3, C18), 18.3 (CH.sub.3, C21), 20.8 (CH.sub.2, C11), 23.3 (CH.sub.3, C19), 24.2 (CH.sub.2, C15), 26.4 (CH.sub.2, C7), 27.2 (CH.sub.2, C6), 28.2 (CH.sub.2, C16), 30.5 (CH.sub.2, C2), 31.7 (CH.sub.2, C22), 33.5 (CH.sub.2, C23), 34.5 (C, C10), 35.3 (CH, C20), 35.5 (CH.sub.2, C1), 35.8 (CH, C8), 36.4 (CH.sub.2, C4), 36.6 (CH.sub.2) 40.2 (CH.sub.2, C12), 40.4 (CH, C9), 42.1 (CH, C5), 42.7 (C, C13), 45.1 (CH.sub.3) 56.0 (CH, C17), 56.5 (CH, C14), 57.9 (CH.sub.2) 71.8 (CH, C3), 173.7 (CO, C24) ppm.

(513) IR; 3377 (OH), 3293 (NH) 2929 (C—H), 2870 (C—H), 1646 (C═O), 1543, 1445 cm.sup.1.

(514) MS (+ESI) m/z=Found 447.3943 (M+H)+; calculated for C.sub.28H.sub.51N.sub.2O.sub.2 447.3945; 0.5 ppm.

Synthesis of (4R)-4-[(3R,10S,12S,13R,17R)-3,12-dihydroxy-10,13-dimethyl-2,3,4,5,6,7,8,-9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]-N-(2-dimethylaminoethyl) pentanamide (102)

(515) ##STR00074##

(516) A mixture of deoxycholic acid (1.0 g 0.002 mol) and N, N′-dimethylethylenediamine (0.16 mL 0.001 mol) was dissolved in toluene (20 mL). The solution was heated at reflux for 24 hours. More N, N′-dimethylethylenediamine (0.16 mL 0.001 mol) was added and the reaction mixture was heated at reflux for a further 24 hours. The solvent was then evaporated under reduced pressure. The residue was triturated with hot water (˜10 mL) and washed with water (3×20 mL), then dried under vacuum.

(517) Yield 0.310 g, 0.0006 mol, 24%.

(518) .sup.1H NMR (CDCl.sub.3) (250 MHz) δ=0.68 (s, 3H, 18-CH.sub.3), 0.91 (s, 3H, 19-CH.sub.3), 1.01 (d, 3H, 21-CH.sub.3, J=6.5), 2.26 (s, 6H, 2×CH.sub.3), 2.46 (t, 2H, CH.sub.2, J=5.0), 3.34 (t, 2H, CH.sub.2, J=5.0), 3.61 (m, 1H, 3-CH), 3.97 (broad singlet, 1H, 12-CH), 6.44 (broad singlet, 1H, NH) ppm.

(519) .sup.13C NMR (62.9 MHz) δ=12.7 (CH.sub.3, C18), 17.5 (CH.sub.3, C21), 23.1 (CH.sub.3, C19), 23.6 (CH.sub.2, CIS), 26.1 (CH.sub.2, C7), 27.1 (CH.sub.2, C6), 27.8 (CH.sub.2, C16), 28.5 (CH.sub.2, C11), 30.5 (CH.sub.2, C2), 31.6 (CH.sub.2, C23), 33.3 (CH.sub.2, C22), 33.6 (CH, C9), 34.1 (C, C10), 35.2 (CH.sub.2, C1), 36.0 (CH, C20), 36.4 (CH.sub.2, C4), 42.0 (CH, C5), 44.9 (C, C13), 46.5 (CH.sub.3) 47.3 (CH, C17), 48.2 (CH, C14), 57.9 (CH.sub.3) 71.7 (CH, C3), 73.0 (CH.sub.2, C12), 172.7 (CO, C24) ppm.

(520) IR=3305, (OH), 2929 (alkyl), 2861 (alkyl), 1720 (C═O), 1044 (R.sup.2CH—OH) cm.sup.−1.

(521) MS (+ESI) m/z=Found 463.3888 (M+H)+; calculated for C.sub.28H.sub.51N.sub.2O.sub.3 463.3894; 1.3 ppm.

Synthesis of (4R)—N-[3-(dimethylamino)propyl]-4-[(3R,10S,13R,17R)-3-hydroxy-10,13-dim-ethyl-2,3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phe-nanthren-17-yl]pentanamide (103)

(522) ##STR00075##

(523) A mixture of methyl lithocholate (0.5 g 0.001 mol) and 3-dimethyl-propylamine (3 mL, 0.02 mol) was heated and stirred at 140° C. for 24 hours in an argon environment. Ice water (3 mL) was added to the material and left to stir for two hours at room temperature. The resulting solid was then collected by filtration washed with water (3×20 mL) and left to dry overnight under vacuum to produce off brown crystals. Yield, 0.41 g, 0.0008 mol, 69%.

(524) Melting point: 177.0-178.3° C.

(525) .sup.1H NMR CDCl.sub.3 (250 MHz) δ=0.61 (s, 3H, 18-CH.sub.3) 0.89 (d, 3H, 19-CH.sub.3) 1.57 (s, 3H, 21-CH.sub.3) 2.22 (s, 6H, 2×CH.sub.3) 2.36 (t, 2H, CH.sub.2, J=6.3) 3.30 (t, 2H, CH.sub.2, J=5.6) 3.59 (m, 1H, 3-CH) 6.95 (s, 1H, NH) ppm.

(526) .sup.13C NMR (62.9 MHz) δ=12.0 (CH.sub.3, C18), 18.3 (CH.sub.3, C21), 20.8 (CH.sub.2, C11), 23.3 (CH.sub.3, C19), 24.2 (CH.sub.2, C15), 24.3 (CH.sub.2, C7), 26.4 (CH.sub.2), 27.1 (CH.sub.2, C6), 28.2 (CH.sub.2, C16), 30.5 (CH.sub.2, C2), 31.7 (CH.sub.2, C22) 34.5 (C, C10), 35.3 (CH, C20), 35.5 (CH.sub.2, C1), 35.8 (CH, C8), 36.4 (CH.sub.2, C4), 40.1 (CH.sub.2, C12), 40.3 (CH, C9), 42.0 (CH, C5), 42.7 (C, C13), 43.0 (CH.sub.3) 55.3 (CH, C17), 55.9 (CH, C14), 56.4 (CH.sub.2) 71.9 (CH, C3), 174.9 (CO, C24) ppm.

(527) IR=3310-3318 (OH—NH), 2730 (alkyl), 2859 (alkyl), 2946 (alkyl), 1648 (C═O), 1047 (CH—OH) cm.sup.−1.

(528) MS (ES+APCI) m/z=Found 461.4105 (M+H)+; calculated for C.sub.29H.sub.53N.sub.2O.sub.2 461.4107; 0.4 ppm.

Synthesis of (4R)-4-[(3R,10S,12S,13R,17R)-3,12-dihydroxy-10,13-dimethyl-2,3,4,5,6,7,8,-9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]-N-[3-(dimethylamino)propyl]pentanamide (104)

(529) ##STR00076##

(530) A mixture of methyl deoxycholate (0.5 g 0.002 mol) and 3-dimethyl-1-propylamine (3.63 mL 0.03 mol) were added together and heated to 100° C. for 5 days then allowed to cool to ambient room temperature. Ice water (40 mL) was then added and left to stir for 2 hours and the precipitate collected by vacuum filtration, washed with water (3×20 mL) and dried under vacuum.

(531) Yield, 0.41 g, 0.0008 mol, 73%.

(532) Melting point: 123-127° C.

(533) .sup.1H NMR (CDCl.sub.3) (250 MHz) δ=0.64 (s, 3H, 18-CH.sub.3), 0.92 (s, 3H, 19-CH.sub.3), 2.23 (s, 6H, 2×CH.sub.3), 2.42 (t, 2H, CH.sub.2, J=5.0), 3.325 (q, 2H, CH.sub.2, J=7.5), 3.63 (m, 1H, 3-CH), 6.21 (broad singlet, 1H, NH) ppm.

(534) .sup.13C NMR (62.9 MHz) δ=12.0 (CH.sub.3, C18), 18.3 (CH.sub.3, C21), 20.8 (CH.sub.2, C11), 23.3 (CH.sub.3, C19), 24.2 (CH.sub.2, C15), 26.4 (CH.sub.2, C7), 27.2 (CH.sub.2, C6), 28.2 (CH.sub.2, C16), 30.5 (CH.sub.2, C2), 31.7 (CH.sub.2, C22), 33.5 (CH.sub.2, C23), 34.5 (C, C10), 35.3 (CH, C20), 35.5 (CH.sub.2, C1), 35.8 (CH, C8), 36.4 (CH.sub.2, C4), 36.6 (CH.sub.2), 40.2 (CH.sub.2, C12), 40.4 (CH, C9), 42.1 (CH, C5), 42.7 (C, C13), 45.1 (CH.sub.2), 56.0 (CH, C17), 56.5 (CH.sub.2), 57.9 (CH, C14), 71.8 (CH, C3), 173.7 (CO, C24) ppm.

(535) IR=3376 (NH and OH), 2938 (alkyl), 2861 (alkyl), 1643 (C═O), 1544, 1444, 1378, 1045 (R.sub.2CH—OH) cm.sup.−1.

(536) MS (+ES APCI) m/z=Found 447.3943 (M+H)+; calculated for C.sub.28H.sub.51N.sub.2O.sub.2 447.3943; 0.5 ppm.

Synthesis of 1-(4-butylpiperazin-1-yl)-4-[(3R,10S,13R,17R)-3-hydroxy-10,13-dimethyl-2,-3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]pentan-1-one (105)

(537) ##STR00077##

(538) Following a modified method (Fini et al., 1992), lithocholic acid (0.5 g 0.001 mol) was dissolved in tetrahydrofuran (20 mL) with triethylamine (0.4 mL 0.002 mol). The solution was cooled for 10 minutes in cold water. Ethyl chloroformate (0.2 mL 0.001 mol) was then added dropwise over a ten minute period. Once added, the cold water was removed and the solution was stirred for 2 hours. After 2 hours, solvent removed under reduced pressure then re-dissolved in dichloromethane and washed with water then 1-butylpiperazine (0.25 mL 0.001 mol) was added and the solution was stirred for 24 hours. Water (50 mL) was then added and the solution extracted with ethyl acetate (3×50 mL). The organic layers were combined and washed with saturated sodium hydrogen carbonate solution (3×50 mL). The organic layer was dried over magnesium sulphate. Solvent was evaporated under reduced pressure.

(539) Yield; 0.16 g, 0.0003 mol, 23.5%.

(540) Melting point: 172.7-173.1° C.

(541) .sup.1H NMR (CDCl.sub.3) (250 MHz) δ=0.61 (s, 3H, 18-CH.sub.3), 0.88 (s, 3H, 19-CH.sub.3), 1.34 (m, 4H, CH.sub.2), 2.35 (m, 6H, CH.sub.2), 3.45 (broad singlet, 2H, CH.sub.2), 3.60 (broad singlet, 2H, CH.sub.2) ppm.

(542) .sup.13C NMR (62.9 MHz) δ=12.0 (CH.sub.3, C18), 14.0 (CH.sub.3), 18.5 (CH.sub.3, C21), 20.6 (CH.sub.2, C1), 20.8 (CH.sub.2) 23.3 (CH.sub.3, C19), 24.2 (CH.sub.2, C15), 26.4 (CH.sub.2, C7), 27.2 (CH.sub.2, C6), 28.2 (CH.sub.2, C16), 28.8 (CH.sub.2), 30.2 (CH.sub.2, C2), 30.5 (CH.sub.2, C22), 31.4 (CH.sub.2, C23), 34.5 (C, C10), 35.3 (CH, C20), 35.6 (CH.sub.2, C1), 35.8 (CH, C8), 36.4 (CH.sub.2, C4), 40.1 (CH.sub.2, C12), 40.4 (CH, C9), 41.4 (CH, C5), 42.1 (C, C13), 42.8 (CH.sub.2), 45.6 (CH.sub.2), 52.9 (CH.sub.2), 53.4 (CH.sub.2), 53.4 (CH.sub.2), 56.0 (CH, C17), 56.5 (CH, C14), 58.3 (CH.sub.2), 71.7 (CH, C3), 172.1 (CO, C24) ppm.

(543) IR=3381, (OH), 2916 (alkyl), 2852 (alkyl), 1616 (C═O), 1437, 1253, 1036 (R.sub.2CH—OH) cm.sup.−1.

(544) MS (+ESI) m/z=Found 501.4409 (M+H)+; calculated for C.sub.32H.sub.57N.sub.2O.sub.2 501.4409; 1.1 ppm.

Synthesis of (4R)-1-(4-butylpiperazin-1-yl)-4-[(3R,10S,12S,13R,17R)-3,12-dihydroxy-10,-13-dimethyl-2,3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta-[a]phenanthren-17-yl]pentan-1-one (106)

(545) ##STR00078##

(546) Deoxycholic acid (0.5 g 0.001 mol) was dissolved in tetrahydrofuran (20 mL) with triethylamine (0.4 mL 0.001 mol). The solution was cooled for 10 minutes in cold water. Ethyl chloroformate (0.2 mL 0.001 mol) was then added dropwise over a ten minute period. Once added, the cold water was removed and the solution was stirred for 2 hours. After 2 hours, solvent removed under reduced pressure then re-dissolved in dichloromethane and washed with water then 1-butylpiperazine (0.25 mL 0.001 mol) was added and the solution was stirred for 24 hours. Water (50 mL) was then added and the solution extracted with ethyl acetate (3×50 mL). The organic layers were combined and washed with saturated sodium hydrogen carbonate solution (3×50 mL). The organic layer was dried over magnesium sulphate. Solvent was evaporated under reduced pressure.

(547) Yield; 0.04 g, 0.0004 mol, 6%.

(548) Melting point: 92.3-93.5° C.

(549) .sup.1H NMR (CDCl.sub.3) (250 MHz) δ=0.68 (s, 3H, 18-CH.sub.3), 0.89 (s, 3H, 19-CH.sub.3), 0.98 (d, 3H, 21-CH.sub.3 J=7.5), 2.38 (m, 6H, CH.sub.2), 3.48 (broad singlet, 2H, CH.sub.2), 3.62 (broad singlet, 2H, CH.sub.2), 3.98 (broad singlet, 1H, 12-CH) ppm.

(550) .sup.13C NMR (62.9 MHz) δ=12.7 (CH.sub.3, C18), 14.0 (CH.sub.2), 17.5 (CH.sub.3, C21), 20.6 (CH.sub.2), 23.1 (CH.sub.3, C19), 23.6 (CH.sub.2, C15), 27.5 (CH.sub.2, C7), 28.6 (CH.sub.2, C6), 28.8 (CH.sub.2, C16), 30.1 (CH.sub.2, C11), 30.5 (CH.sub.2, C2), 31.3 (CH.sub.2, C23), 33.6 (CH.sub.2, C22), 34.1 (CH, C9), 35.2 (C, C10), 35.2 (CH.sub.2, C1), 36.0 (CH, C20), 36.4 (CH.sub.2, C4), 41.4 (CH.sub.2), 42.0 (CH, C5), 45.6 (CH.sub.2) 46.5 (CH.sub.2) 47.2 (CH, C17), 48.3 (CH, C14), 52.8 (CH.sub.2) 53.4 (CH.sub.2) 71.8 (CH, C3), 73.1 (CH.sub.2, C12), 171.9 (CO, C24) ppm.

(551) IR=3300 (OH), 2930 (alkyl), 2861 (alkyl), 1594 (C═O), 1443, 1282, 1162 (C═O ester stretch), 1011 (R.sub.2CH—OH) cm.sup.1.

(552) MS (+ESI) m/z=Found 517.4359 (M+H)+; calculated for C.sub.32H.sub.57N.sub.2O.sub.3 517.4364; 0.9 ppm.

Synthesis of 4-[(3R,10S,13R,17R)-3-hydroxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,12,14,15,-16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]-N-(2-pyrrolidin-1-ylethyl)pentanamide (107)

(553) ##STR00079##

(554) Lithocholic acid (0.5 g 0.001 mol) was dissolved in tetrahydrofuran (20 mL) with triethylamine (0.20 mL, 0.001 mol). The solution was cooled for 10 minutes in cold water. Ethyl chloroformate (0.20 mL, 0.001 mol) was then added dropwise over a ten minute period. Once added, the cold water was removed and the solution was stirred for 2 hours at room temperature. After 2 hours 1-(2-aminoethyl)pyrollidine (0.25 mL, 0.002 mol) was added and the solution was stirred for 3 hours. Water (50 mL) was then added and the solution was extracted with ethyl acetate (3×50 mL). The organic layers were combined and dried over magnesium sulphate. The solvent was evaporated under reduced pressure to produce a white powder.

(555) Yield; 0.56 g, 0.001 mol, 91.8%.

(556) Melting point: 144.1-145.1° C.

(557) .sup.1H NMR (CDCl.sub.3) (250 MHz) δ=0.64 (s, 3H, 18-CH.sub.3), 0.92 (s, 3H, 19-CH.sub.3) 0.94 (d, 3H, 21-CH.sub.3, J=7.5), 1.80 (m, 4H, CH.sub.2), 2.54 (broad singlet, 4H, CH.sub.2), 2.60 (t, 2H, CH.sub.2, J=5.0) 3.63 (q, 2H, CH.sub.2, J=5.0) 3.63 (m, 1H, 3-CH), 6.13 (broad s, 1H, NH) ppm.

(558) .sup.13C NMR (62.9 MHz) δ=12.0 (CH.sub.3, C18), 18.4 (CH.sub.3, C21), 20.8 (CH.sub.2, C11), 23.4 (CH.sub.3, C19), 24.2 (CH.sub.2, C15), 26.4 (CH.sub.2, C7), 27.2 (C H.sub.2, C6), 28.2 (CH.sub.2, C16), 30.5 (CH.sub.2, C2), 31.7 (CH.sub.2, C22), 33.5 (CH.sub.2, C23), 34.5 (C, C10), 35.3 (CH, C20), 35.5 (CH.sub.2, C1), 35.8 (CH, C8), 36.4 (CH.sub.2, C4), 37.7 (CH.sub.2) 40.2 (CH.sub.2, C12), 40.4 (CH, C9), 42.1 (CH, C5), 42.7 (C, C13), 53.8 (CH.sub.2), 54.9 (CH.sub.2) 56.0 (CH, C17), 56.5 (CH, C14), 71.8 (CH, C3), 173.9 (CO, C24) ppm.

(559) IR=3415 (NH), 3310 (OH), 2933 (alkyl), 2865 (alkyl), 2872 (alkyl), 1648 (C═O), 1548, 1444, 1378, 1265 (C═O ester stretch), 1064 (R.sub.2CH—OH) cm.sup.−1.

(560) MS (+ESI) m/z=Found 473.4096 (M+H)+; calculated for C.sub.30H.sub.53N.sub.2O.sub.2 473.4102; 1.2 ppm.

Synthesis of 4-[(3R,10S,12S,13R,17R)-3,12-dihydroxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,-12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]-N-(2-pyrr-olidin-1-ylethyl) pentanamide (108)

(561) ##STR00080##

(562) Deoxycholic acid (2.0 g 0.005 mol) was dissolved in dioxane (60 mL) with triethylamine (1.28 mL, 0.01 mol). The solution was cooled for 10 minutes in cold water. Ethyl chloroformate (0.37 mL, 0.003 mol) was then added dropwise over a ten minute period. Once added, the cold water was removed and the solution was stirred for 2 hours. After 2 hours 1-(2-aminoethyl)pyrollidine (0.77 mL, 0.006 mol) was added and the solution was stirred for 3 hours at room temperature. Water (50 mL) was then added and the solution was extracted with ethyl acetate (3×50 mL). The organic layers were combined and washed with saturated sodium hydrogen carbonate solution (3×50 mL). The organic layer was dried over magnesium sulphate. The solvent was evaporated under reduced pressure to produce a white powder.

(563) Yield; 1.2 g, 0.0025 mol, 49%.

(564) Melting point: 155.3-158.7° C.

(565) .sup.1H NMR (CDCl.sub.3) (250 MHz) δ=0.66 (s, 3H, 18-CH.sub.3), 0.89 (s, 3H, 19-CH.sub.3), 0.975 (d, 3H, 21-CH.sub.3, J=7.5), 1.78 (m, 4H, CH.sub.2), 2.42-2.58 (m, 6H, CH.sub.2), 3.35 (m, 2H, CH.sub.2), 3.58 (m, 1H, 3-CH), 3.95 (broad s, 1H, 12-CH), 6.49 (broad s, 1H, NH) ppm.

(566) .sup.13C NMR (62.9 MHz) δ=12.7 (CH.sub.3, C18), 17.5 (CH.sub.3, C21), 23.3 (CH.sub.3, C19), 23.7 (CH.sub.2, C15), 26.1 (CH.sub.2, C7), 27.1 (CH.sub.2, C6), 28.6 (CH.sub.2, C16), 30.4 (CH.sub.2, C11), 30.5 (CH.sub.2, C2), 33.3 (CH.sub.2, C23), 33.6 (CH.sub.2, C22), 34.1 (CH, C9), 35.2 (C, C10), 35.3 (CH.sub.2, C1), 36.0 (CH, C20), 36.5 (CH.sub.2, C4), 37.9 (CH, C8), 42.1 (CH.sub.2), 46.5 (C, C13), 46.8 (CH, C17), 48.2 (CH, C14), 53.9 (CH), 55.0 (CH.sub.2) 71.5 (CH, C3), 73.0 (CH.sub.2, C12), 173.9 (CO, C24) ppm.

(567) IR=3287 (OH), 2916 (alkyl), 2865 (alkyl), 1641 (C═O), 1539, 1441, 1040 (R.sub.2CH—OH) cm.sup.−1.

(568) MS (+ESI) m/z=Found 489.4046 (M+H)+; calculated for C.sub.30H.sub.53N.sub.2O.sub.3 489.4051; 1 ppm.

Synthesis of 4-[(3R,10S,13R,17R)-3-hydroxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,12,14,15,-16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]-N-[2-(1-piperidyl)-ethyl]pentanamide (109)

(569) ##STR00081##

(570) Methyl lithocholate 0.5 g (0.001 mci) 1,2 amino-ethyl piperidine 0.36 mL (0.002 mol) were heated at and stirred at 150° C. for 48 hours in a argon environment. More 1,2 amino-ethyl piperidine (1 mL, 0.007 mol) of was added and the reaction was continued under the same conditions overnight. The crude material was purified by flash column chromatography (1:1 EtOAc/MeOH) to give a solid.

(571) Yield; 0.01 g, 0.00001 mol, 1.6%.

(572) Melting point: 85.0-89.9° C.

(573) .sup.1H NMR CDCl.sub.3 (250 MHz) δ=0.59 (s, 3H, 18-CH.sub.3) 0.87-0.89 (s, 3H, 19-CH.sub.3 J=5.5) 1.72 (in, 4R, CH.sub.2) 2.68 (t, 2H, CH.sub.2, J=5.6) 3.42-3.44 (q, 2R, CH.sub.2, J=5.6) 3.58 (in, 1H, 3-CR) 7.18 (s/t, 1H, NR) ppm.

(574) .sup.13C NMR (62.9 MHz) δ=18.3 (CH) 23.4 (CH.sub.3) 24.2 (CH.sub.3CH) 26.4 (CH.sub.2) 42.2 (CH.sub.3) 56.5 (CH.sub.2) 71.8 (COH) 173.9 (CO) ppm.

(575) IR=3418 (NH), 3465 (OH) 2935 (alkyl), 2865 (alkyl), 1641 (C═O) cm.sup.−1.

(576) MS (ES) m/z=Found 487.4347 (M+H)+; calculated for C.sub.31H.sub.54N.sub.2O.sub.3 487.4264; 0.8 ppm.

Synthesis of (4R)-4-[(3R,10S,12S,13R,17R)-3,12-dihydroxy-10,13-dimethyl-2,3,4,5,6,7,8,-9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]-N-[2-(1-piperidyl)ethyl] pentanamide (110)

(577) ##STR00082##

(578) Methyl deoxycholate (1.0 g 0.002 mol) was dissolved in methanol (10 mL) with 1-(2-aminoethyl)piperidine (3.15 g, 0.024 mol) was added and the solution was stirred for 5 days. The solvent was evaporated under reduced pressure and the product was recrystallized from ethyl acetate to produce a white powder.

(579) Yield; 0.56 g, 0.0011 mol, 45%.

(580) Melting point: 158.5-161.1° C.

(581) .sup.1H NMR (CDCl.sub.3) (250 MHz) δ=0.66 (s, 3H, 18-CH.sub.3), 0.90 (s, 3H, 19-CH.sub.3), 0.99 (d, 3H, 21-CH.sub.3, J=5.0), 2.52 (m, 2H, CH.sub.2), 3.385 (q, 2H, CH.sub.2, J=5.0), 3.60 (m, 1H, 3-CH), 3.97 (s, 1H, 12-CH), 6.72 (broad s, 1H, NH) ppm.

(582) .sup.13C NMR (62.9 MHz) δ=12.7 (CH.sub.3, C18), 17.5 (CH.sub.3, C21), 23.1 (CH.sub.3, C19), 23.7 (CH.sub.2), 23.8 (CH.sub.2) 25.2 (CH.sub.2, C15), 26.1 (CH.sub.2) 27.1 (CH.sub.2, C7), 27.5 (CH.sub.2, C6), 28.6 (CH.sub.2, C16), 30.5 (CH.sub.2, C2), 31.6 (CH.sub.2, C23), 33.3 (CH.sub.2, C22), 33.6 (CH, C9), 34.1 (C, C10), 35.2 (CH.sub.2, C1), 35.2 (CH, C20), 35.4 (CH.sub.2, C4), 36.0 (CH, C8), 36.5 (CH, C5), 42.0 (C, C13), 46.5 (CH, C17), 47.0 (CH.sub.2) 48.3 (CH, C14), 54.2 (CH.sub.2), 57.4 (CH.sub.2) 71.6 (CH, C3), 73.0 (CH.sub.2, C12), 173.9 (CO, C24) ppm.

(583) IR=3300 (OH), 2925 (alkyl), 2861 (alkyl), 1642 (C═O), 1543, 1437, 1313, 1040 (R.sub.2CH—OH) cm.sup.−1.

(584) MS (+ESI) m/z=Found 503.4202 (M+H)+; calculated for C.sub.31H.sub.55N.sub.2O.sub.3 503.4207; 1 ppm.

Synthesis of (4R)—N-(4-benzoylphenyl)-4-[(3R,10S,13R,17R)-3-hydroxy-10,13-dimethyl-2,-3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]pentanamide (111)

(585) ##STR00083##

(586) Lithocholic acid (2.0 g 0.005 mol) was dissolved in tetrahydrofuran (60 mL) with N-methylmorpholine (1.07 mL, 0.01 mol). The solution was cooled for 10 minutes in cold water. Ethyl chloroformate (0.54 mL, 0.005 mol) was then added dropwise over a ten minute period. Once added, the cold water was removed and the solution was stirred for 2 hours. After 2 hours amino-benzophenone (1.5 g, 0.007 mol) was added and the solution was stirred for a further 48 hours. Water (50 mL) was then added and the solution was extracted with ethyl acetate (3×50 mL). The organic layers were combined and washed with saturated sodium hydrogen carbonate solution (3×50 mL) and 2 M hydrochloric acid solution (3×50 mL). The organic layer was dried over magnesium sulphate. The solvent was evaporated under reduced pressure and the product was triturated with ethyl acetate to produce a white powder.

(587) Yield 0.57 g, 0.001 mol, 19%.

(588) Melting point: 228.6-230.8° C.

(589) .sup.1H NMR (DMSO) (250 MHz) δ=0.62 (s, 3H, 18-CH.sub.3) 0.87 (s, 3H, 19-CH.sub.3) 0.92 (d, 2H, CH.sub.2 J=7.5) 4.44 (d, 1H, 3-OH, J=5.0) 7.52-7.805 (m, 9H, Ar—CH), 10.20 (s, 1H, NH) ppm.

(590) .sup.13C NMR (62.9 MHz) δ=11.8 (CH.sub.3, C18), 18.3 (CH.sub.3, C21), 20.3 (CH.sub.2, C1), 23.2 (CH.sub.3, C19), 23.8 (CH.sub.2, C15), 26.1 (CH.sub.2, C7), 26.8 (CH.sub.2, C6), 27.7 (CH.sub.2, C16), 30.3 (CH.sub.2, C2), 31.1 (CH.sub.2, C22), 33.4 (CH.sub.2, C23), 34.1 (C, C10), 34.9 (CH, C20), 35.1 (CH.sub.2, C1), 35.3 (CH, C8), 41.4 (CH, C5), 42.2 (C, C13), 55.5 (CH, C17), 56.0 (CH, C14), 69.8 (CH, C3), 118.1, 128.4, 129.3, 131.0, 131.1, 132.1, 137.5, 143.5 (Ar—CH) 172.3 (CO, C24), 194.4 (CO) ppm.

(591) IR=3488 (NH), 3249 (OH), 2925 (alkyl), 2857 (alkyl), 1675 (C═O), 1586, 1296, 1245, 1168 (C═O ester stretch), 1031 (R.sub.2CH—OH) cm.sup.−1.

(592) MS (+ESI) m/z=Found 556.3781 (M+H)+; calculated for C.sub.37H.sub.50NO.sub.3 556.3785; 0.8 ppm.

Synthesis of (4R)—N-(4-benzoylphenyl)-4-[(3R,10S,12S,13R,17R)-3,12-dihydroxy-10,13-di-methyl-2,3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]ph-enanthren-17-yl]pentanamide (112)

(593) ##STR00084##

(594) Deoxycholic acid (2.0 g 0.005 mol) was dissolved in 1, 4 dioxane (60 mL) with N-methylmorpholine (1.07 mL 0.01 mol). The solution was cooled for 10 minutes in cold water. Ethyl chloroformate (0.69 mL 0.006 mol) was then added dropwise over a ten minute period. Once added, the cold water was removed and the solution was stirred for 2 hours. After 2 hours amino-benzophenone (1.5 g 0.007 mol) was added and the solution was stirred for a further 24 hours. Water (50 mL) was then added and the solution extracted with ethyl acetate (3×50 mL). The organic layers were combined and washed with saturated sodium hydrogen carbonate solution (3×50 mL) and 2 M hydrochloric acid (3×50 mL). The organic layer was dried over magnesium sulphate. Solvent was evaporated under reduced pressure and triturated with ethyl acetate to produce a white powder.

(595) Yield; 0.24 g, 0.0004 mol, 8%.

(596) Melting point: 220-227.6° C.

(597) .sup.1H NMR (DMSO) (250 MHz) δ=0.60 (s, 3H, 18-CH.sub.3), 0.85 (s, 3H, 19-CH.sub.3), 0.97 (d, 3H, 21-CH.sub.3, J=5.0), 3.80 (s, 1H, 3-CH), 4.03 (s, 1H, 12-CH), 4.21 (d, 1H, 3-OH, J=2.5), 4.46 (d, 1H, 12-OH, J=5.0), 7.52-7.79 (multiple overlapping multiplets, 9H, Ar—CH), 10.26 (s, 1H, NH) ppm.

(598) .sup.13C NMR (62.9 MHz) δ=12.4 (CH.sub.3, C18), 17.0 (CH.sub.3, C21), 23.0 (CH.sub.3, C19), 26.9 (CH.sub.2, C7), 27.2, (CH.sub.2, C2), 32.9 (CH.sub.2, C22), 33.5 (CH, C9), 33.7 (C, C10), 35.0 (CH.sub.2, C1), 35.6 (CH, C20), 45.9 (C, C13), 46.1 (CH, C17), 47.2 (CH, C14), 70.9 (CH, C3), 118.1 (CH), 128.4 (CH), 129.3 (CH), 131.1 (CH), 132.1 (CH), 143.5 (CH), 172.4 (CO, C24), 194.5 (CO) ppm.

(599) IR=3462 (OH), 2921 (alkyl), 2861 (alkyl), 1675 (C═O), 1586 (C═O), 1441, 1279, 1168.78 (C═O ester stretch), 1036 (R.sub.2CH—OH) cm.sup.−1.

(600) MS (+ESI) m/z=Found 572.3730 (M+H)+; calculated for C.sub.37H.sub.50NO.sub.4 572.3734; 0.8 ppm.

Synthesis of (4R)-4-[(3R,10S,13R,17R)-3-hydroxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,12,1-4,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]-1-[4-(4-pyrid-yl)piperazin-1-yl]pentan-1-one (113)

(601) ##STR00085##

(602) Lithocholic acid (0.5 g, 0.001 mol) was dissolved in tetrahydrofuran (15 mL) with triethylamine (0.20 mL, 0.001 mol). The solution was cooled for 10 minutes in cold water. Ethyl chloroformate (0.2 mL, 0.001 mol) was then added dropwise over a ten minute period. Once added, the cold water was removed and the solution was stirred for 2 hours. After 2 hours 1-(4-pyridyl)piperazine (0.21 mL, 0.001 mol) was added and the solution was stirred for 24 hours. Water (50 mL) was then added and the solution extracted with ethyl acetate (3×50 mL). The organic layers were combined and washed with saturated sodium hydrogen carbonate solution (3×50 mL). Due to impurities, re-dissolved in a methanol/water solution (20 mL 50/50) and extracted with chloroform (4×20 mL). The combined organic layers were dried over magnesium sulphate. The solvent was evaporated under reduced pressure to produce a white powder.

(603) Yield; 0.03 g, 0.00005 mol, 4%.

(604) Melting point: 150-157° C.

(605) .sup.1H NMR (CDCl.sub.3) (250 MHz) δ=0.65 (s, 3H, 18-CH.sub.3), 0.92 (s, 3H, 19-CH.sub.3), 3.35 (m, 4H, CH.sub.2), 3.64 (m, 2H, CH.sub.2), 6.68 (d, 2H, Ar—CH, J=5.0), 8.31 (s, 2H, Ar—CH) ppm.

(606) .sup.13C NMR (62.9 MHz) δ=12.0 (CH.sub.3, C18), 18.6 (CH.sub.3, C21), 20.8 (CH.sub.2, C11), 23.3 (CH.sub.3, C19), 24.3 (CH.sub.2, C15), 26.3 (CH.sub.2, C7), 27.1 (CH.sub.2, C6), 28.3 (CH.sub.2, C16), 30.2 (CH.sub.2, C2), 31.3 (CH.sub.2, C22), 34.5 (C, C10), 35.4 (CH, C20), 35.8 (CH.sub.2, C1), 36.4 (CH.sub.2, C4), 40.1 (CH.sub.2, C12), 40.4 (CH, C9), 40.7 (CH, C5), 42.1 (C, C13), 42.7 (CH.sub.2), 44.7 (CH.sub.2), 45.8 (CH.sub.2), 45.9 (CH.sub.2) 55.9 (CH, C17), 56.5 (CH, C14), 71.8 (CH, C3), 108.5 (CH), 129.7 (CH), 149.9 (CH), 150.0 (CH), 154.6 (CH), 172.7 (CO, C24) ppm.

(607) IR=3387 (OH), 2925 (alkyl), 2852 (alkyl), 1641 (C═O), 1445, 1236, 1044, 989 (R.sub.2CH—OH) cm.sup.−1.

(608) MS (ES) m/z=Found 522.4048 (M+H).+−.; calculated for C.sub.31H.sub.57N.sub.2O.sub.4 521.4313; 1.5 ppm.

Synthesis of (4R)—N-(4-aminobutyl)-4-[(3R,10S,12S,13R,17R)-3,12-dihydroxy-10,13-dimet-hyl-2,3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phena-nthren-17-yl]pentanamide (114)

(609) ##STR00086##

(610) Methyl deoxycholate (1.0 g 0.009 mol) was dissolved in methanol (10 mL). 1,4 Diaminobutane (2.47 mL 0.02 mol) was added and the solution was stirred for a further 72 hours. Water (50 mL) was then added and the resultant precipitate was collected by vacuum filtration, washed with water (3×20 mL) and dried under vacuum to produce a white powder.

(611) Yield; 0.29 g, 0.0006 mol, 25%.

(612) Melting point: 137.4-141° C.

(613) .sup.1H NMR (DMSO) (250 MHz) δ=0.58 (s, 3H, 18-CH.sub.3), 0.84 (s, 3H, 19-CH.sub.3), 0.91 (d, 3H, 21-CH.sub.3, J=5.0), 2.99 (q, 2H, CH.sub.2, J=7.5), 3.6 (m, 1H, 3-CH), 3.78 (s, 1H, 12-CH), 7.73 (broad s, 1H, NH) ppm.

(614) .sup.13C NMR (62.9 MHz) δ=12.4 (CH.sub.3, C18), 17.0 (CH.sub.3, C21), 23.0 (CH.sub.3, C19), 23.4 (CH.sub.2, C15), 26.0, (CH.sub.2), 26.6 (CH.sub.2), 26.9 (CH.sub.2) 27.1 (CH.sub.2, C7), 28.5 (CH.sub.2, C16), 30.2 (CH.sub.2, C11), 30.5 (CH.sub.2, C2), 31.7 (CH.sub.2, C23), 32.5 (CH.sub.2, C22), 32.8 (CH, C9), 33.7 (C, C10), 35.0 (CH.sub.2, C1), 35.1 (CH, C20), 35.6 (CH.sub.2, C4), 36.2 (CH, C8), 41.3 (CH, C5), 41.5 (CH) 45.9 (C, C13), 46.1 (CH, C17), 47.4 (CH, C14), 69.8 (CH, C3), 69.8 (CH.sub.2, C12), 172.2 (CO, C24)v ppm.

(615) IR=3322 (OH), 2929 (alkyl), 2857 (alkyl), 1624 (C═O), 1539, 1437, 1360, 1040 (R.sup.2CH—OH) cm.sup.−1.

(616) MS (+ESI) m/z=Found 463.3886 (M+H)+; calculated for C.sub.28H.sub.51N.sub.2O.sub.3 463.3894; 1.8 ppm.

Synthesis of (4R)—N-[2-[2-(2-aminoethoxy)ethoxy]ethyl]-4-[(3R,10S,13R,17R)-3-hydroxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]pentanamide (115)

(617) ##STR00087##

(618) Lithocholic acid (2.0 g, 0.005 mol) was dissolved in toluene (40 mL). 2, 2′-(ethylenedioxy)bis(ethylamine) (7.8 mL, 0.052 mol) was added and the solution was heated at reflux for 5 days. Water (50 mL) was then added and resultant precipitate collected by vacuum filtration, with further washes of water (3×20 mL). The product was dried under vacuum to produce an off-white powder.

(619) Yield, 1.62 g, 0.003 mol, 60.22%

(620) Melting point: 81.1-85.5° C.

(621) .sup.1H NMR (CDCl.sub.3) (250 MHz) δ=0.64 (s, 3H, 18-CH.sub.3), 0.92 (s, 3H, 19-CH.sub.3), 2.91 (broad s, 2H, CH.sub.2), 3.46-3.63 (multiple overlapping multiplets, 1OH, CH.sub.2), 6.24 (broad s, 1H, NH) ppm.

(622) .sup.13C NMR (62.9 MHz) δ=12.0 (CH.sub.3, C18), 18.4 (CH.sub.3, C21), 19.9 (CH.sub.2) 20.8 (CH.sub.2, C11), 23.3 (CH.sub.3, C19), 24.2 (CH.sub.2, C15), 26.4 (CH.sub.2, C7), 27.2 (CH.sub.2, C6), 28.2 (CH.sub.2, C16), 30.5 (CH.sub.2, C2), 31.7 (CH.sub.2, C22), 33.4 (CH.sub.2, C23), 34.5 (C, C10), 35.3 (CH, C20), 35.5 (CH.sub.2, C1), 35.8 (CH, C8), 36.4 (CH.sub.2, C4), 39.0 (CH.sub.2, C12), 39.1 (CH, C9), 40.2 (CH.sub.2), 40.4 (CH.sub.2) 42.1 (CH, C5), 42.7 (C, C13), 56.0 (CH, C17), 56.5 (CH, C14), 70.1 (CH.sub.2), 70.5 (CH, C3), 71.7 (CH.sub.2), 173.7 (CO, C24) ppm.

(623) IR=3356 (OH), 2921 (alkyl), 2857 (alkyl), 1641 (C═O), 1552, 1441, 1300, 1104 (C═O ester stretch), 1053 (R.sub.2CH—OH) cm.sup.−1.

(624) MS (+ESI) m/z=Found 507.4149 (M+H)+; calculated for C.sub.30H.sub.55N.sub.2O.sub.4 507.4156; 1.4 ppm.

Synthesis of (4R)-4-[(3R,10S,13R,17R)-3-hydroxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,12,1-4,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]-N-[2-[2-[2-(2 methylprop-2-enoylamino)ethoxy]ethoxy]ethyl]pentanamide (116)

(625) ##STR00088##

(626) Compound 115 (0.5 g 0.0009 mol) was dissolved in anhydrous tetrahydrofuran (10 mL) with triethylamine (0.20 mL, 0.001 mol). Methacrylic anhydride (0.30 mL, 0.0018) was then added. The solution was stirred for 48 hours, protected from sunlight by tin foil at ambient room temperature. Water (30 mL) was then added and the resulting precipitate was collected by vacuum filtration, washed with water (3×20 mL) and dried under vacuum.

(627) Yield; 0.07 g, 0.0001 mol, 12.5%.

(628) Melting point: 57.3-59.0° C.

(629) .sup.1H NMR (DMSO) (250 MHz) δ=0.60 (s, 3H, 18-CH.sub.3), 0.87 (s, 3H, 19-CH.sub.3), 1.84 (s, 3H, acryloyl-CH.sub.3), 3.154-3.50 (m, 13H, CH/CH.sub.2), 4.42 (d, 1H, 3-OH, J=5.0), 5.31 (s, 1H, ═CH), 5.64 (s, 1H, ═CH), 7.78 (broad s, 1H, NH), 7.91 (broad s, 1H, NH) ppm.

(630) .sup.13C NMR (62.9 MHz) δ=11.8 (CH.sub.3, C18), 18.2 (CH.sub.3, C21), 18.6 (CH), 20.3 (CH.sub.2, C11), 23.2 (CH.sub.3, C19), 23.8 (CH.sub.2, C15), 26.1 (CH.sub.2, C7), 26.8 (CH.sub.2, C6), 27.7 (CH.sub.2, C16), 30.3 (CH.sub.2, C2), 31.5 (CH.sub.2, C22), 32.2 (CH.sub.2, C23), 34.1 (C, C10), 34.9 (CH, C20), 35.1 (CH.sub.2, C1), 35.3 (CH, C8), 41.4 (CH, C5), 42.2 (C, C13), 55.2 (CH, C17), 56.0 (CH, C14), 68.8 (CH.sub.2), 69.1 (CH.sub.2), 69.5 (CH.sub.2), 69.8 (CH, C3), 118.9 (CH.sub.2), 139.8 (CH.sub.2), 167.4 (CO) 172.5 (CO, C24) ppm.

(631) IR=3415 (NH), 3292 (OH), 2933 (alkyl), 2861 (alkyl), 1658 (C═O), 1590 (C═O), 1394, 1249 (C═O ester stretch), 1036 (R.sub.2CH—OH) cm.sup.−1.

(632) MS (+ESI) m/z=Found 575.4412 (M+H)+; calculated for C.sub.34H.sub.59N.sub.2O.sub.5 575.4418; 1.1 ppm.

Synthesis of (4R)—N-(4-acetylphenyl)-4-[(3R,10S,12S,13R,17R)-3,12-dihydroxy-10,13-dim-ethyl-2,3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phe-nanthren-17-yl]pentanamide (117)

(633) ##STR00089##

(634) Deoxycholic acid (2.0 g 0.005 mol) was dissolved in 1, 4 dioxane (60 mL) with triethylamine (1.32 mL, 0.01 mol). The solution was cooled for 10 minutes in cold water. Ethyl chloroformate (0.40 mL, 0.003 mol) was then added dropwise over a ten minute period. Once added, the cold water was removed and the solution was stirred for 2 hours. After 2 hours 4′-aminoacetophenone (0.75 g, 0.005 mol) was added and the solution was stirred for a further 48 hours. Water (50 mL) was then added and the resulting precipitate was collected by vacuum filtration. The crude product was triturated with ethyl acetate to produce a white powder.

(635) Yield; 0.65 g, 0.001 mol, 25%.

(636) .sup.1H NMR (DMSO) (250 MHz) δ=0.62 (s, 3H, 18-CH.sub.3), 0.86 (s, 3H, 19-CH.sub.3), 2.52 (s, 3H, ketone-CH.sub.3), 3.81 (broad s, 1H, 12-CH), 4.21 (d, 1H, 3-OH, J=2.5), 4.46 (d, 1H, 12-OH, J=2.5), 7.73 (d, 2H, Ar—CH, J=10.0), 7.92 (d, 2H, Ar—CH, J=7.5), 10.21 (s, 1H, NH) ppm.

(637) IR=3475 (OH), 2921 (alkyl), 2861 (alkyl), 1684 (C═O), 1646 (C═O), 1594, 1539, 1407, 1044 (C═O ester stretch) cm.sup.−1.

(638) MS (+ESI) m/z=Found 510.3570 (M+H)+; calculated for C.sub.32H.sub.48NO.sub.4 510.3578; 1.3 ppm.

Synthesis of ethane; (4R)-4-[(3R,10S,13R,17R)-3-hydroxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,12,1-4,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]-1-[4-(2-hydro-xyethyl)piperazin-1-yl]pentan-1-one (118)

(639) ##STR00090##

(640) Lithocholic acid (4.0 g 0.01 mol) was dissolved in tetrahydrofuran (120 mL) with triethylamine (1.30 mL, 0.01 mol). The solution was cooled for 10 minutes in cold water. Ethyl chloroformate (1.02 mL, 0.009 mol) was then added dropwise over a ten minute period. Once added, the cold water was removed and the solution was stirred for 2 hours. After 2 hours 1-(2-hydroxyethylpiperazine) (1.5 mL, 0.01 mol) was added and the solution was stirred for a further 48 hours. Water (50 mL) was then added and the solution was extracted with ethyl acetate (3×50 mL). The organic layers were combined and washed with saturated sodium hydrogen carbonate solution (3×50 mL). The organic layer was dried over magnesium sulphate. The solvent was evaporated under reduced pressure and the product was recrystallized from ethyl acetate to produce a white powder.

(641) Yield; 2.27 g, 0.004 mol, 43%.

(642) Melting point: 153.0-154.8° C.

(643) .sup.1H NMR (CDCl.sub.3) (250 MHz) δ=0.65 (s, 3H, 18 CH.sub.3), 0.92 (s, 3H, 19-CH.sub.3), 2.50 (m, 4H, CH.sub.2), 2.58 (t, 2H, CH.sub.2, J=5.0), 3.49 (t, 2H, CH.sub.2, J=2.5), 3.65 (t, 4H, CH.sub.2, J=5.0) ppm.

(644) .sup.13C NMR (62.9 MHz) δ=12.0 (CH.sub.3, C18), 18.5 (CH.sub.3, C21), 20.8 (CH.sub.2, C11), 23.3 (CH.sub.3, C19), 24.2 (CH.sub.2, C15), 26.4 (CH.sub.2, C7), 27.2 (CH.sub.2, C6), 28.2 (CH.sub.2, C16), 30.2 (CH.sub.2, C2), 30.5 (CH.sub.2, C22), 31.4 (CH.sub.2, C23), 34.5 (C, C10), 35.3 (CH, C20), 35.6 (CH.sub.2, C1), 35.8 (CH, C8), 36.4 (CH.sub.2, C4), 40.2 (CH.sub.2, C12), 40.4 (CH, C9), 41.4 (CH, C5), 42.1 (C, C13), 42.7 (CH.sub.2), 45.6 (CH.sub.2), 52.6 (CH.sub.2), 53.1 (CH.sub.2) 56.0 (CH, C17), 56.5 (CH, C14), 57.7 (CH.sub.2), 59.3 (CH.sub.2) 71.8 (CH, C3), 172.1 (CO, C24) ppm.

(645) IR=3381 (OH), 2916 (alkyl), 2840 (alkyl), 1620 (C═O), 1445, 1258, 1044 (R.sup.2CH—OH) cm.sup.−1.

(646) MS (ES) m/z=Found 489.4045 (M+H)+; calculated for C.sub.30H.sub.53N.sub.2O.sub.3 489.4051; 1.2 ppm.

Synthesis of (4R)-4-[(3R,10S,12S,13R,17R)-3,12-dihydroxy-10,13-dimethyl-2,3,4,5,6,7,8,-9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]-1-[4-(2-hydroxyethyl) piperazin-1-yl]pentan-1-one (119)

(647) ##STR00091##

(648) Deoxycholic acid (4.0 g 0.01 mol) was dissolved in 1, 4 dioxane (120 mL) with triethylamine (2.70 mL 0.02 mol). The solution was cooled for 10 minutes in cold water. Ethyl chloroformate (0.8 mL 0.005 mol) was then added dropwise over a ten minute period. Once added, the cold water was removed and the solution was stirred for 2 hours. After 2 hours 1-(2-hydroxyethylpiperazine) (1.32 mL 0.01 mol) was added and the solution was stirred for 48 hours. Water (50 mL) was then added and the solution extracted with ethyl acetate (3×50 mL). The organic layers were combined and washed with saturated sodium hydrogen carbonate solution (3×50 mL). The organic layer was dried over magnesium sulphate. Solvent was evaporated under reduced pressure and recrystallized from methanol to produce a white powder.

(649) Yield; 2.41 g, 0.004 mol, 46.8%.

(650) Melting point: 241.0-243.9° C.

(651) .sup.1H NMR (CDCl.sub.3/MeOH) (250 MHz) δ=0.70 (s, 3H, 18-CH.sub.3), 0.93 (s, 3H, 19-CH.sub.3), 2.58 (multiple overlapping multiplets, 6H, CH.sub.2), 3.53 (broad t, 3H, 3-CH/CH.sub.2), 3.65-3.72 (multiple overlapping multiplets, 4H, CH.sub.2), 3.96 (broad s, 1H, 12-CH) ppm.

(652) IR=3407 (OH), 2929 (alkyl), 1620 (C═O), 1454, 1215, 1044 (R.sub.2CH—OH) cm.sup.−1.

(653) MS (+ESI) m/z=Found 505.3994 (M+H)+; calculated for C.sub.31H.sub.57N.sub.2O.sub.4 505.4000; 1.2 ppm.

Synthesis of (4R)—N-[3-(dibutylamino)propyl]-4-[(3R,10S,13R,17R)-3-hydroxy-10,13-dime-thyl-2,3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phen-anthren-17-yl]pentanamide (120)

(654) ##STR00092##

(655) Lithocholic acid (0.5 g 0.001 mol) was dissolved in tetrahydrofuran (15 mL) with triethylamine (0.20 mL 0.001 mol). The solution was cooled for 10 minutes in cold water. Ethyl chloroformate (0.2 mL 0.001 mol) was then added dropwise over a ten minute period. Once added, the cold water was removed and the solution was stirred for 2 hours. After 2 hours 3-(dibutylamino)-1-propylamine (0.25 mL, 0.001 mol) was added and the solution was stirred for 24 hours. Water (50 mL) was then added and the solution extracted with ethyl acetate (3×50 mL). The organic layers were combined and washed with saturated sodium hydrogen carbonate solution (3×50 mL). This crude material was further purified using column chromatography (8:2 ethyl acetate/methanol). The compound containing fractions were dried over magnesium sulphate. Solvent was evaporated under reduced pressure to produce a white powder.

(656) Yield; 0.24 g, 0.0004 mol, 33%.

(657) Melting point: 62.9-69.3° C.

(658) .sup.1H NMR (CDCl.sub.3) (250 MHz) δ=0.64 (s, 3H, 18-CH.sub.3), 0.93 (s, 3H, 19-CH.sub.3), 0.94 (m, 6H, CH.sub.3), 1.35 (m, 8H, CH.sub.2), 2.45 (t, 4H, CH.sub.2, J=5.0), 3.32 (q, 2H, CH.sub.2, J=5.0), 7.51 (broad s, 1H, NH) ppm.

(659) .sup.13C NMR (62.9 MHz) δ=12.0 (CH.sub.3, C18), 14.0 (CH), 18.3 (CH.sub.3, C21), 20.7 (CH.sub.2, C11), 23.3 (CH.sub.3, C19), 24.2 (CH.sub.2, C15), 25.1 (CH.sub.2), 26.4 (CH.sub.2, C7), 27.2 (CH.sub.2, C6), 28.2 (CH.sub.2, C16), 28.4 (CH.sub.2), 28.5 (CH.sub.2) 30.4 (CH.sub.2, C2), 30.5 (CH.sub.2, C22), 33.8 (CH.sub.2, C23), 34.5 (C, C10), 35.3 (CH, C20), 36.4 (CH.sub.2, C1), 39.4 (CH.sub.2, C4), 40.1 (CH.sub.2, C12), 40.4 (CH, C9), 42.0 (CH, C5), 42.1 (C, C13), 42.7 (CH.sub.2) 56.1 (CH, C17), 56.4 (CH, C14), 71.6 (CH, C3), 173.6 (CO, C24) ppm.

(660) IR=3296 (OH), 2916 (alkyl), 2861 (alkyl), 1650 (C═O), 1548, 1441, 1377, 1070, 1066, 1036 (R.sub.2CH—OH) cm.sup.−1.

(661) MS (+ESI) m/z=Found 545.5032 (M+H)+; calculated for C.sub.35H.sub.65N.sub.2O.sub.2 545.5041; 1.6 ppm.

Synthesis of (4R)-4-[(3R,10S,13R,17R)-3-hydroxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,12,1-4,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]-N-octadecyl-p-entanamide (121)

(662) ##STR00093##

(663) Lithocholic acid (2.0 g 0.005 mol) was dissolved in tetrahydrofuran (60 mL) with 4-methylmorpholine (2.14 mL, 0.02 mol). The solution was cooled for 10 minutes in cold water. Ethyl chloroformate (0.51 mL, 0.005 mol) was then added dropwise over a ten minute period. Once added, the cold water was removed and the solution was stirred for 2 hours. After 2 octadecylamine (1.43 mL, 0.005 mol) was added and the solution was stirred for a further 21 hours. Water (50 mL) was then added and the solution was extracted with ethyl acetate (3×50 mL). The crude product was purified by column chromatography (100% ethyl acetate). The product containing fractions were dried over magnesium sulphate. The solvent was evaporated under reduced pressure to produce a white powder.

(664) Yield; 0.8 g, 0.001 mol, 23.9%.

(665) Melting point: 94.8-95.4° C.

(666) .sup.1H NMR (250 MHz) CDCl.sub.3 δ=0.64 (s, 3H, 18-CH.sub.3), 0.92 (s, 3H, 19-CH.sub.3), 1.25 (s, 30H, aliphatic CH.sub.2) 3.23 (q, 3H, chain terminal CH.sub.3, J=5.0), 3.64 (m, 1H, 3-CH), 5.36 (broad s, 1H, NH) ppm.

(667) .sup.13C NMR (62.9 MHz) δ=12.0 (CH.sub.3, C18), 18.4 (CH.sub.3, C21), 20.8 (CH.sub.2, C11), 22.7 (CH.sub.3, C19), 23.3 (CH.sub.2, C15), 24.2 (CH.sub.2) 26.4 (CH.sub.2, C7), 26.9 (CH.sub.2, C6), 27.2 (CH.sub.2, C16), 28.2 (CH.sub.2), 29.3 (CH.sub.2), 29.5 (CH.sub.2), 29.7 (CH.sub.2), 30.5 (CH.sub.2, C2), 31.8 (CH.sub.2, C22), 31.9 (CH.sub.2, C23), 33.7 (C, C10), 35.3 (CH, C20), 35.4 (CH.sub.2, C1), 35.8 (CH, C8), 39.5 (CH.sub.2, C4), 40.2 (CH.sub.2, C12), 40.4 (CH, C9), 42.1 (C, C5), 56.0 (CH, C17), 56.5 (CH, C14), 71.9 (CH, C3), 185.3 (CO, C24) ppm.

(668) IR=3411 (NH), 3325 (OH), 2912 (alkyl), 2849 (alkyl), 1653 (C═O), 1544, 1464, 1367, 1308, 1040 (R.sub.ZCH—OH) cm.sup.−1.

(669) MS (+ESI) m/z=Found 628.6205 (M+H)+; calculated for C.sub.42H.sub.78NO.sub.2 628.6207; 0.6 ppm.

Synthesis of (4R)-4-[(3R,10S,12S,13R,17R)-3,12-dihydroxy-10,13-dimethyl-2,3,4,5,6,7,8,-9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]-N-oc-tadecyl-pentanamide (122)

(670) ##STR00094##

(671) Deoxycholic acid (2.0 g 0.005 mol) was dissolved in 1, 4 dioxane (60 mL) with triethylamine (1.32 mL, 0.01 mol). The solution was cooled for 10 minutes in cold water. Ethyl chloroformate (0.40 mL, 0.003 mol) was then added dropwise over a ten minute period. Once added, the cold water was removed and the solution was stirred for 2 hours. After 2 hours octadecylamine (1.37 g, 0.005 mol) was added and the solution was stirred for a further 48 hours. Water (50 mL) was then added and the solution was extracted with ethyl acetate (3×50 mL). The organic layers were combined and washed with saturated sodium hydrogen carbonate solution (3×50 mL). The organic layer was dried over magnesium sulphate. The solvent was evaporated under reduced pressure and the product was recrystallized from ethyl acetate to produce a transparent glass-like solid.

(672) Yield; 2.32 g, 0.003 mol, 70%.

(673) Melting point: 54.2-57.3° C.

(674) .sup.1H NMR (CDCl.sub.3) (250 MHz) δ=0.68 (s, 3H, 18-CH.sub.3), 0.91 (s, 3H, 19-CH.sub.3), 1.25 (s, 32H, aliphatic CH.sub.2) 3.22 (q, 2H, CH.sub.2, J=7.5), 3.61 (m, 1H, 3-CH), 3.98 (1H, 12-CH), 5.53 (broad s, 1H, NH) ppm.

(675) .sup.13C NMR (62.9 MHz) δ=12.7 (CH.sub.3, C18), 14.1 (CH.sub.2), 17.4 (CH.sub.3, C21), 22.6 (CH.sub.2), 23.1 (CH.sub.3, C19), 23.6 (CH.sub.2, C15), 26.1 (CH), 26.9 (CH.sub.2, C7), 28.6 (CH.sub.2, C16), 29.3 (CH.sub.2, C11), 29.5 (CH.sub.2, C2), 29.7 (CH.sub.2), 30.5 (CH) 31.7 (CH.sub.2, C23), 31.9 (CH.sub.2, C22), 33.5 (CH, C9), 33.6 (CH), 34.1 (CH), 35.2 (C, C10), 35.2 (CH) 36.0 (CH.sub.2, C1), 36.4 (CH, C20), 39.5 (CH.sub.2, C4), 42.1 (CH, C5), 46.5 (C, C13), 47.2 (CH, C17), 48.2 (CH, C14), 71.7 (CH, C3), 73.1 (CH.sub.2, C12), 173.4 (CO, C24) ppm.

(676) IR=3292 (OH), 2912, 2852 (alkyl), 1637 (C═O), 1548, 1189.78 (C═O ester stretch), 1036 (RCHOH) cm.sup.−1.

(677) MS (+ESI) m/z=Found 644.5971 (M+H)+; calculated for C.sub.42H.sub.78NO.sub.3 644.5976; 0.8 ppm.

Synthesis of 2-[[(4R)-4-[(3R,10S,13R,17R)-3-hydroxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,-12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]pentanoyl]-amino]ethyl-trimethyl-ammonium iodide (123)

(678) ##STR00095##

(679) Compound 101 (0.2 g 0.0004 mol) was dissolved in chloroform (10 mL) with Iodomethane (0.31 mL 0.002 mol). The solution was left overnight at which point a precipitate had formed which was collected by vacuum filtration, washed with water (3×20 mL) and dried under vacuum to produce a white powder.

(680) Yield; 0.212 g, 0.0003 mol, 81.5%.

(681) Melting point: 212.8-216.1° C.

(682) .sup.1H NMR (MeOD) (250 MHz) δ=0.69, (s, 3H, 18-CH.sub.3), 0.94 (s, 3H, 19-CH.sub.3), 3.19, (s, 9H, 3×CH.sub.3), 3.45 (q, 2H, CH.sub.2, J=7.5), 3.55 (m, 1H, 3-CH), 3.64 (t, 2H, CH.sub.2, J=7.5) ppm.

(683) .sup.13C NMR (62.9 MHz) δ=11.0 (CH.sub.3), 12.5 (CH.sub.3), 17.3 (CH.sub.3), 18.8 (CH.sub.2), 21.9 (CH) 23.9 (CH.sub.2), 27.6 (CH), 28.3 (CH), 31.2 (CH), 33.0 (CH), 33.9 (CH), 36.9 (CH.sub.2), 37.2 (CH.sub.2), 41.9 (CH), 43.5 (C, C13), 57.4 (CH, C17), 57.9 (CH, C14), 65.8 (CH.sub.2) 72.4 (CH), 181.2 (CO) ppm.

(684) IR=3368 (NH), 3245 (OH), 2938 (alkyl), 2852 (alkyl) 1639 (C═O), 1560, 1441, 1258 (C═O ester stretch), 1040 (R.sup.2CH—OH) cm.sup.−1.

(685) MS (+ESI) m/z=Found 461.4105 M+; calculated for C.sub.29H.sub.53N.sub.2O.sub.2 461.4102; 0.7 ppm.

Synthesis of 2-[[(4R)-4-[(3R,10S,12S,13R,17R)-3,12-dihydroxy-10,13-dimethyl-2,3, 4,5,6,-7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]p-entanoyl]amino]ethyl-trimethyl-ammonium iodide (124)

(686) ##STR00096##

(687) Compound 102 (0.15 g 0.0003 mol) was dissolved in chloroform (5 mL). Iodomethane (0.22 mL, 0.001 mol) was added and the solution was stirred overnight at ambient temperature. The resultant precipitate was collected via vacuum filtration and dried in a desiccator overnight to give the product as an off-white solid.

(688) Yield 0.09 g, 0.0001 mol, 78.9%.

(689) Melting point: 160.1-169.1° C.

(690) .sup.1H NMR (MeOD) (250 MHz) δ=0.70 (s, 3H, 18-CH.sub.3), 0.93 (s, 3H, 19-CH.sub.3), 1.02 (d, 3H, 21-CH.sub.3, J=5.0), 3.20 (s, 9H, 3×CH.sub.3), 3.47 (t, 2H, CH.sub.2, J=7.5), 3.64 (t, 2H, CH.sub.2, J=5.0), 3.95 (s, 1H, 12-CH) ppm.

(691) .sup.13C NMR (62.9 MHz) δ=13.2 (CH.sub.3, C18), 17.6 (CH.sub.3, C21), 23.7 (CH.sub.3, C19), 24.8 (CH.sub.2, C15), 27.4 (CH.sub.2, C7), 28.4 (CH.sub.2, C6), 28.7 (CH.sub.2, C16), 29.9 (CH.sub.2, C11), 31.0 (CH.sub.2, C2), 33.0 (CH.sub.2, C23), 33.8 (CH.sub.2), 34.6 (CH.sub.2), 36.4 (CH.sub.2, C1), 36.8 (CH, C20), 37.2 (CH.sub.2, C4), 37.4 (CH, C8), 43.6 (CH, C5), 54.0 (CH), 54.0 (CH), 54.1 (CH), 72.5 (CH, C3), 74.0 (CH.sub.2, C12), 177.3 (CO, C24) ppm.

(692) IR=3377 (NH), 3249 (OH), 2921 (alkyl), 2861 (alkyl), 1641 (C═O), 1573, 1454, 1373, 1249, 1031 (R.sub.2CH—OH) cm.sup.1.

(693) MS (+ESI) m/z=Found 477.4060 M.sup.+; calculated for C.sub.29H.sub.53N.sub.2O.sub.3 477.4051; 1.9 ppm.

Synthesis of ethyl-[3-[[(4R)-4-[(3R,10S,13R,17R)-3-hydroxy-10,13-dimethyl-2,3,4,5,6,7,-8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]pen-tanoyl]amino]propyl]-dimethyl-ammonium iodide (125)

(694) ##STR00097##

(695) Compound 101 (0.2 g 0.0004 mol) was dissolved in chloroform (10 mL). Ethyl Iodide (0.34 mL, 0.001 mol) was added and the solution was stirred overnight at ambient temperature. The resulting precipitate was collected by vacuum filtration and was dried overnight in a desiccator to give the product as an off-white solid.

(696) Yield; 0.2 g, 0.0003 mol, 76%.

(697) Melting point: 120.0-123.0° C.

(698) .sup.1H NMR (MeOD) (250 MHz) δ=0.69 (s, 3H, 18-CH.sub.3), 0.95 (s, 3H, 19-CH.sub.3), 3.13 (s, 6H, 2×CH.sub.3), 3.43 (m, 4H, 3-CH/CH.sub.2), 3.60 (q, 2H, CH.sub.2, J=7.5) ppm.

(699) .sup.13C NMR (62.9 MHz) δ=8.4 (CH) 12.5 (CH.sub.3, C18), 18.8 (CH.sub.3, C21), 21.4 (CH.sub.2, C11), 21.9 (CH.sub.3, C19), 23.9 (CH.sub.2, C15), 25.2 (CH.sub.2, C7), 28.3 (CH.sub.2, C16), 29.3 (CH.sub.2, C2), 31.2 (CH.sub.2, C23), 33.0 (CH.sub.2), 33.9 (CH.sub.2), 34.2 (C, C10), 35.6 (CH, C20), 36.4 (CH.sub.2, C1), 37.1 (CH, C8), 37.2 (CH.sub.2, C4), 41.5 (CH, C9), 41.9 (CH, C5), 43.5 (C, C13), 43.9 (CH.sub.2), 51.1 (CH.sub.2) 57.3 (CH, C17), 57.9 (CH, C14), 62.4 (CH) (CH, C3), 177.3 (CO, C24) ppm.

(700) IR=3373 (OH), 2942 (alkyl), 2844 (alkyl), 1646 (C═O), 1441, 1270, 1036 (R.sup.2CH—OH) cm.sup.−1.

(701) MS (+ESI) m/z=Found 475.4247 M.sup.+; calculated for C.sub.30H.sub.55N.sub.2O.sub.2 475.4258; 2.3 ppm.

Synthesis of 3-[[(4R)-4-[(3R,10S,12S,13R,17R)-3,12-dihydroxy-10,13-dimethyl-2,3,4,5,6,-7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]p-entanoyl]amino]propyl-ethyl-dimethyl-ammonium iodide (126)

(702) ##STR00098##

(703) Compound 102 (0.2 g 0.0004 mol) was dissolved in chloroform (5 mL). Ethyl Iodide (0.62 mL, 0.003 mol) was added and the solution was stirred overnight at ambient temperature. The resulting precipitate was collected by vacuum filtration and was dried overnight in a desiccator. The product was an off-white solid.

(704) Yield; 0.13 g, 0.0002 mol, 52%.

(705) Melting point: 115.3-120.3° C.

(706) .sup.1H NMR (CDCl.sub.3) (250 MHz) δ=0.70 (s, 3H, 18-CH.sub.3), 0.93 (s, 3H, 19-CH.sub.3), 1.02 (d, 2H, CH.sub.2), 2.21 (m, 6H, 3-CH/CH.sub.2), 3.4 (broad s, 2H, CH.sub.2), 3.45-3.61 (multiple overlapping multiplets, 8H, CH.sub.2) ppm.

(707) .sup.13C NMR (62.9 MHz) δ=8.4 (CH.sub.2) 12.4 (CH.sub.3, C18), 16.9 (CH.sub.3, C21), 21.9 (CH.sub.2) 22.9 (CH.sub.3, C19), 24.0 (CH.sub.2, C15), 26.6 (CH.sub.2, C7), 27.9 (CH.sub.2, C16), 29.1 (CH.sub.2, C11), 30.3 (CH.sub.2, C2), 32.2 (CH.sub.2, C23), 33.1 (CH.sub.2, C22), 34.0 (CH, C9), 34.5 (C, C10), 35.6 (CH.sub.2, C1), 36.0 (CH, C20), 36.4 (CH.sub.2, C4), 36.6 (CH, C8), 42.8 (CH, C5), 55.6 (CH), 57.1 (CH), 63.2 (CH) 71.7 (CH, C3), 73.2 (CH.sub.2, C12), 176.6 (CO, C24) ppm.

(708) IR=3368 (NH), 3253 (OH), 2921 (alkyl), 2852 (alkyl), 1650 (C═O), 1522, 1445, 1364, 1253, 1036 (R.sub.2CH—OH) cm.sup.−1.

(709) MS (+ESI) m/z=Found 517.4354 M.sup.+; calculated for C.sub.32H.sub.57N.sub.2O.sub.3 517.4364; 1.9 ppm.

Synthesis of allyl-[2-[[(4R)-4-[(3R,10S,13R,17R)-3-hydroxy-10,13-dimethyl-2,3,4,5,6,7,-8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]pen-tanoyl]amino]ethyl]-dimethyl-ammonium bromide (127)

(710) ##STR00099##

(711) Compound 101 (0.2 g, 0.0004 mol) was dissolved in a solution of chloroform (10 mL). Allyl bromide (0.27 mL, 0.002 mol) was added and the solution was stirred overnight at which point a precipitate was formed. The precipitate was collected by vacuum filtration and was dried overnight in a desiccator. The product was an off-white solid.

(712) Yield; 0.13 g, 0.0002 mol, 54%.

(713) Melting point: 198.7-203.8° C.

(714) .sup.1H NMR (MeOD) (250 MHz) δ=0.68 (s, 3H, 18-CH.sub.3), 0.94 (s, 3H, 19-CH.sub.3), 3.12 (s, 6H, 2×CH.sub.3), 3.39 (t, 2H, CH.sub.2), 3.65 (t, 2H, CH.sub.2), 5.70 (m, 2H, CH.sub.2), 6.10 (m, 1H, ═CH—) ppm.

(715) .sup.13C NMR (62.9 MHz) δ=12.5 (CH.sub.3, C18), 18.8 (CH.sub.3, C21), 21.9 (CH.sub.2, C11), 23.9 (CH.sub.3, C19), 25.2 (CH.sub.2, C15), 27.6 (CH.sub.2, C7), 28.3 (CH.sub.2), 28.0 (CH.sub.2, C16), 33.0 (CH.sub.2) 34.2 (C, C10), 37.1 (CH.sub.2), 41.5 (CH.sub.2), 41.9 (CH.sub.2), 43.8 (CH), 57.3 (CH.sub.2), 57.9 (CH.sub.2), 63.0 (CH.sub.2), 72.4 (CH, C3), 126.0 (CH), 129.8 (CH), 177.3 (CO, C24) ppm.

(716) IR=3266 (OH), 2929 (alkyl), 2848 (alkyl), 1646 (C═O), 1569, 1420, 1066, 1036 (R.sub.2CH—OH) cm.sup.1.

(717) MS (+ESI) m/z=Found 487.4252 M.sup.+; calculated for C.sub.31H.sub.55N.sub.2O.sub.2 487.4258; 1.2 ppm.

Synthesis of allyl-[2-[[(4R)-4-[(3R,10S,12S,13R,17R)-3,12-dihydroxy-10,13-dimethyl-2,3-,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]pentanoyl]amino]ethyl]-dimethyl-ammonium iodide (128)

(718) ##STR00100##

(719) Compound 102 (0.15 g, 0.0003 mol) was dissolved in chloroform (5 mL). Allyl bromide (0.20 mL, 0.0016 mol) was added and the solution was stirred overnight at ambient temperature. The resulting precipitate was collected by vacuum filtration and was dried overnight in a desiccator. The product was an off-white solid.

(720) Yield; 0.19 g, 0.0001 mol, 53%.

(721) Melting point: 184.2-186.7° C.

(722) .sup.1H NMR (MeOD) (250 MHz) δ=0.70 (s, 3H, CH.sub.3), 0.93 (s, 3H, CH.sub.3), 3.13 (s, 6H, CH.sub.3), 3.40 (t, 2H, CH.sub.2, J=7.5), 3.65 (t, 2H, CH.sub.2, J=5.0), 3.95 (s, 1H, 12-CH), 4.05 (d, 2H, CH.sub.2, J=5.0), 5.75 (m, 2H, CH.sub.2), 6.10 (m, 1H, ═CH—) ppm.

(723) .sup.13C NMR (62.9 MHz) δ=13.2 (CH.sub.3, C18), 17.6 (CH.sub.3, C21), 23.7 (CH.sub.3, C19), 24.8 (CH.sub.2, C15), 27.4 (CH.sub.2, C7), 28.4 (CH.sub.2, C6), 28.7 (CH.sub.2, C16), 29.9 (CH.sub.2, C11), 31.1 (CH.sub.2, C2), 33.0 (CH.sub.2, C23), 33.8 (CH.sub.2, C22), 34.8 (CH, C9), 36.9 (CH), 37.2 (CH), 37.4 (CH), 43.6 (CH, C5), 47.5 (CH, C17), 48.0 (CH, C14), 63.0 (CH.sub.2), 63.1 (CH.sub.2), 67.9 (CH.sub.2), 68.0 (CH.sub.2), 68.0 (CH.sub.2) 72.5 (CH, C3), 74.0 (CH, C12), 126.1 (CH.sub.3), 129.8 (CH.sub.2), 177.4 (CO, C24) ppm.

(724) IR=3415 (NH), 3245 (OH), 2925 (alkyl), 2852 (alkyl), 1646 (C═O), 1441, 1364, 1292 cm.sup.−1.

(725) MS (+ESI) m/z=Found 503.4202 M.sup.+; calculated for C.sub.31H.sub.55N.sub.2O.sub.3 503.4207; 1.0 ppm.

Synthesis of 2-[[(4R)-4-[(3R,10S,13R,17R)-3-hydroxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,-12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]pentanoyl]-amino]ethyl-dimethyl-[(4-vinylphenyl)methyl]ammonium chloride (129)

(726) ##STR00101##

(727) Compound 101 (0.2 g 0.0004 mol) was dissolved in chloroform (10 mL). Vinyl benzyl chloride (0.64 mL, 0.004 mol) was added and the solution was stirred overnight at ambient temperature. The resulting precipitate was collected by vacuum filtration and was dried overnight in a desiccator. The product was an off-white solid.

(728) Yield; 0.07 g, 0.0016 mol, 26.9%.

(729) Melting point: 143.8-148.9° C.

(730) .sup.1H NMR (MeOD) (250 MHz) δ=0.66 (s, 3H, 18-CH.sub.3), 0.94 (s, 3H, 19-CH.sub.3), 3.09 (s, 6H, 2×CH.sub.3), 3.41 (t, 2H, CH.sub.2, J=7.5), 3.73 (t, 2H, CH.sub.2, J=5.0), 4.56 (s, 2H, CH.sub.2), 5.37 (d, 1H, ═CH—, J=12.5), 5.91 (d, 1H, ═CH, J=20.0) 6.80 (dd, 1H, ═CH, J=10.0), 7.57 (dd, 4H, Ar—CH, J=5.0) ppm.

(731) .sup.13C NMR (62.9 MHz) δ=11.7 (CH.sub.3, C18), 18.0 (CH.sub.3, C21), 21.1 (CH.sub.2, C11), 23.1 (CH.sub.3, C19), 24.4 (CH.sub.2, C15), 26.8 (CH.sub.2, C7), 27.5 (CH.sub.2, C6), 28.4 (CH.sub.2, C16), 30.3 (CH.sub.2, C2), 33.0 (CH.sub.2), 33.9 (CH.sub.2), 35.7 (CH.sub.2), 36.8 (CH), 37.2 (CH), 41.9 (CH), 43.5 (CH), 57.4 (CH, C17), 57.9 (CH, C14), 63.5 (CH.sub.2) 71.6 (CH, C3), 115.8 (CH), 126.9 (CH), 127.2 (CH), 133.6 (CH), 136.2 (CH), 140.8 (CH), 176.5 (CO, C24) ppm.

(732) IR=3339 (NH), 3215 (OH), 2921 (alkyl), 2857 (alkyl), 1667 (C═O), 1646, 1445, 1356, 1070 (R.sup.2CH—OH) cm.sup.−1.

(733) MS (+ESI) m/z=Found 563.4558 M.sup.+; calculated for C.sub.37H.sub.59N.sub.2O.sub.2 563.4571; 2.3 ppm.

Synthesis 3-[[(4R)-4-[(3R,10S,13R,17R)-3-hydroxy-10,13-dimethyl-2,3,4,5,6,-7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]p-entanoyl]amino]propyl-trimethyl-ammonium iodide (130)

(734) ##STR00102##

(735) Compound 103 (0.5 g 0.002 mol) was dissolved in a mixture of chloroform (15 mL) and methanol (4 mL). Iodomethane (3.63 mL, 0.02 mol) was added and the solution was stirred for four days at ambient temperature. To induce precipitation the solution was placed in a acetone/dry ice mixture (−78° C.). The resulting precipitate was collected by vacuum filtration and was dried overnight in a desiccator. The product was an off-white solid.

(736) Yield; 0.42 g, 0.06 mol, 64.6%.

(737) Melting point: 253.8-255.4° C.

(738) .sup.1H NMR (CDCl.sub.3) (250 MHz) δ=0.69 (s, 3H, 18-CH.sub.3), 0.94 (s, 3H, 19-CH.sub.3), 3.13 (s, 9H, 3×CH.sub.3), 3.54 (m, 1H, 3-CH) ppm.

(739) .sup.13C NMR (62.9 MHz) δ=10.9 (CH.sub.3, C18), 17.3 (CH.sub.3, C21), 20.4 (CH.sub.2, C11), 22.4 (CH.sub.2), 23.0 (CH.sub.3, C19), 23.7 (CH.sub.2, C15), 26.1 (CH.sub.2, C7), 26.8 (CH.sub.2, C6), 28.0 (CH.sub.2, C16), 29.6 (CH.sub.2, C2), 31.6 (CH.sub.2, C22), 32.5 (CH.sub.2, C23), 34.1 (C, C10), 34.9 (CH, C20), 35.4 (CH.sub.2, C1), 35.6 (CH, C8), 35.7 (CH.sub.2, C4), 40.0 (CH.sub.2, C12), 40.4 (CH, C9), 42.0 (CH, C5), 42.4 (C, C13), 52.2 (CH, C17), 55.8 (CH, C14), 56.8 (CH), 64.2 (CH.sub.2) 70.8 (CH, C3), 175.7 (CO, C24) ppm.

(740) IR=3386 (OH), 2921 (alkyl), 2852 (alkyl), 1641 (C═O), 1552, 1441, 1368, 1253 (C═O ester stretch), 1031 (R.sup.2CH—OH) cm.sup.1.

(741) MS (+ESI) m/z=Found 475.4256 M+; calculated for C.sub.30H.sub.55N.sub.2O.sub.2 475.4258; 0.4 ppm.

Synthesis of 3-[[(4R)-4-[(3R,10S,12S,13R,17R)-3,12-dihydroxy-10,13-dimethyl-2,3,4,5,6,-7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]p-entanoyl]amino]propyl-trimethyl-ammonium iodide (131)

(742) ##STR00103##

(743) Compound 104 (0.1 g 0.0002 mol) was dissolved in chloroform (5 mL). Iodomethane (0.29 mL, 0.002 mol) was added and the solution was stirred overnight at ambient temperature. The resulting precipitate was collected by vacuum filtration and was dried overnight in a desiccator. The product was an off-white solid.

(744) Yield; 0.09 g, 0.0014 mol, 75%.

(745) Melting point: 150.1-153.3° C.

(746) .sup.1H NMR (MeOD) (250 MHz) δ=0.71 (s, 3H, 18-CH.sub.3), 0.93 (s, 3H, 19-CH.sub.3), 1.03 (d, 3H, 21-CH.sub.3, J=7.5), 3.15 (s, 6H, 2×CH.sub.3), 3.53 (m, 1H, 3-CH), 3.95 (s, 1H, 12-CH) ppm.

(747) .sup.13C NMR (62.9 MHz) δ=13.2 (CH.sub.3, C18), 17.7 (CH.sub.3, C21), 23.7 (CH.sub.3, C19), 27.4 (CH.sub.2, C7), 28.4 (CH.sub.2, C6), 28.7 (CH.sub.2, C16), 29.9 (CH.sub.2, C11), 31.1 (CH.sub.2, C2), 33.1 (CH.sub.2, C23), 35.3 (C, C10), 37.2 (CH.sub.2, C4), 43.6 (CH, C5), 72.5 (CH, C3), 74.0 (CH.sub.2, C12), 177.3 (CO, C24) ppm.

(748) IR=3364 (OH), 2933 (alkyl), 2852 (alkyl), 1654 (C═O), 1548, 1437 cm.sup.−1.

(749) MS (ES) m/z=Found 491.4195 M.sup.+; calculated for C.sub.30H.sub.55N.sub.2O.sub.3 491.4207; 2.5 ppm.

Synthesis of allyl-[3-[[(4R)-4-[(3R,10S,13R,17R)-3-hydroxy-10,13-dimethyl-2,3,4,5,6,7,-8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]pen-tanoyl]amino]propyl]-dimethyl-ammonium bromide (132)

(750) ##STR00104##

(751) Compound 103 (0.5 g 0.001 mol) was dissolved in dichloromethane (20 mL). Allyl bromide (1.88 mL, 0.01 mol) was added and the solution was stirred for a further 5 days. The reaction mixture was chilled to −78° C. A precipitate formed. This was collected by vacuum filtration.

(752) Yield; 0.45 g, 0.0007 mol, 71%.

(753) Melting point: 191.1-209.9° C.

(754) .sup.1H NMR (CDCl.sub.3) (250 MHz) δ=0.68 (s, 3H, 18-CH.sub.3), 0.94 (s, 3H, 19-CH.sub.3), 3.10 (s, 6H, 2×CH.sub.3), 3.27 (q, 2H, CH.sub.2), 3.54 (m, 1H, 3-CH), 4.01 (d, 2H, CH.sub.2, J=7.5), 5.74 (t, 2H, CH.sub.2, J=10.0), 6.08 (m, 1H, NH) ppm.

(755) .sup.13C NMR (62.9 MHz) δ=11.1 (CH.sub.3, C18), 17.5 (CH.sub.3, C21), 20.5 (CH.sub.2, C11), 22.5 (CH.sub.3, C19), 23.8 (CH.sub.2, C15), 26.2 (CH.sub.2, C7), 26.9 (CH.sub.2, C6), 27.0 (CH.sub.2, C16), 29.7 (CH.sub.2, C2), 31.7 (CH.sub.2, C22), 32.5 (CH.sub.2, C23), 34.2 (C, C10), 35.0 (CH, C20), 35.4 (CH.sub.2, C1), 35.7 (CH, C8), 35.9 (CH.sub.2, C4), 40.0 (CH.sub.2, C12), 40.4 (CH, C9), 42.0 (CH, C5), 42.4 (C, C13), 55.9 (CH, C17), 56.4 (CH, C14), 61.7 (CH.sub.2), 66.1 (CH.sub.2) 70.9 (CH, C3), 124.8 (CH), 128.0 (CH.sub.2), 175.8 (CO, C24) ppm.

(756) IR=3411 (NH), 3270 (OH), 2925 (alkyl), 2857 (alkyl), 1646 (C═O), 1543, 1437, 1373, 1036 (R.sub.2CH—OH) cm.sup.−1.

(757) MS (+ESI) m/z=Found 501.4405 M.sup.+; calculated for C31H.sub.55N.sub.2O.sub.2.

Synthesis of cyclopentylmethyl-[3-[[(4R)-4-[(3R,10S,13R,17R)-3-hydroxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthr-en-17-yl]pentanoyl]amino]propyl]-dimethyl-ammonium bromide (133)

(758) ##STR00105##

(759) Compound 103 (0.5 g 0.001 mol) was dissolved in chloroform (20 mL) with benzyl bromide (1.37 mL, 0.008 mol) and the solution was stirred for 48 hours. The solvent was evaporated under reduced pressure, then re-dissolved in methanol and washed with petroleum ether 60/80 (3×20 mL). The solvent was removed under reduced pressure to produce a white powder.

(760) Yield; 0.24 g, 0.0003 mol, 35%.

(761) Melting point: 145-147.9° C.

(762) .sup.1H NMR (MeOD) (250 MHz) δ=0.68 (s, 3H, 18-CH.sub.3), 0.94 (s, 3H, 19-CH.sub.3), 3.05 (s, 6H, 2×CH.sub.3), 3.54 (m, 1H, 3-CH), 4.56 (s, 2H, CH.sub.2), 7.55 (broad s, 5H, Ar—CH) ppm.

(763) .sup.13C NMR (62.9 MHz) δ=10.9 (CH.sub.3, C18), 17.3 (CH.sub.3, C21), 20.4 (CH.sub.2, C11), 22.4 (CH.sub.3, C19), 22.6 (CH.sub.2, C15), 23.7 (CH) 26.1 (CH.sub.2, C7), 26.8 (CH.sub.2, C6), 27.7 (CH.sub.2, C16), 29.6 (CH.sub.2, C2), 31.6 (CH.sub.2, C22), 32.5 (CH.sub.2, C23), 34.1 (C, C10), 34.9 (CH, C20), 35.3 (CH.sub.2, C1), 35.6 (CH, C8), 35.6 (CH.sub.2, C4), 39.9 (CH.sub.2, C12), 40.3 (CH, C9), 41.9 (CH, C5), 42.3 (C, C13), 55.8 (CH, C17), 56.3 (CH, C14), 61.7 (CH), 67.4 (CH), 70.8 (CH, C3), 127.3 (CH), 128.8 (CH), 130.3 (CH), 132.6 (CH), 175.6 (CO, C24) ppm.

(764) IR=3253 (OH), 2929 (alkyl), 2865 (alkyl), 1637 (C═O), 1565, 1441 cm.sup.−1.

(765) MS (+ESI) m/z=Found 551.4570 M.sup.+; calculated for C36H.sub.59N.sub.2O.sub.2 551.4571; 0.2 ppm.

Synthesis of 3-[[(4R)-4-[(3R,10S,13R,17R)-3-hydroxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,-12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]pentanoyl]-amino]propyl-dimethyl-[(4-vinylphenyl)methyl]ammonium chloride (134)

(766) ##STR00106##

(767) Compound 103 (1.0 g 0.001 mol) was dissolved in dichloromethane (20 mL). Vinyl benzyl chloride (0.89 mL, 0.005) was added and the solution was stirred for 48 hours at ambient temperature. The resulting precipitate was collected by vacuum filtration and was washed with petroleum ether 60/80 (3×20 mL). The resultant crude product was dissolved in methanol and again washed with petroleum ether 60/80 (3×20 mL).

(768) The solvent was removed under reduced pressure to produce off-white solid.

(769) Yield; 1 g, 0.001 mol, 75%.

(770) Melting point: >350° C.

(771) .sup.1H NMR (MeOD) (250 MHz) δ=0.67 (s, 3H, 18-CH.sub.3), 0.94 (s, 3H, 19-CH.sub.3), 3.05 (s, 6H, 2×CH.sub.3), 4.54 (s, 2H, CH.sub.2), 5.37 (d, 1H, ═CH—, J=12.5), 5.91 (d, 2H, ═CH, J=15.0), 6.80 (dd, 1H, ═CH, J=12.5), 7.56 (q, 5H, Ar—CH, J=7.5) ppm.

(772) .sup.13C NMR (62.9 MHz) δ=11.9 (CH.sub.3, C18), 18.2 (CH.sub.3, C21), 20.6 (CH.sub.2, C11), 23.3 (CH.sub.3, C19), 24.0 (CH.sub.2, C15), 26.3 (CH.sub.2, C7), 27.1 (CH.sub.2, C6), 28.0 (CH.sub.2, C16), 30.3 (CH.sub.2, C2), 30.9 (CH.sub.2, C22), 30.9 (CH.sub.2, C23), 34.2 (C, C10), 35.1 (CH, C20), 35.3 (CH.sub.2, C1), 35.6 (CH, C8), 36.3 (CH.sub.2, C4), 40.0 (CH.sub.2, C12), 40.2 (CH, C9), 41.9 (CH, C5), 42.4 (C, C13), 55.8 (CH, C17), 56.3 (CH, C14), 70.5 (CH, C3), 178.1 (CO, C24) ppm.

(773) IR=3356 (OH), 2929 (alkyl), 2852 (alkyl), 1633 (C═O), 1548, 1441, 1031 (R.sub.2CH—OH) cm.sup.−1.

(774) MS (+ESI) m/z=Found 577.4723 M.sup.+; calculated for C.sub.38H.sub.61N.sub.2O.sub.2 577.4728; 0.8 ppm.

Synthesis of (4R)-4-[(3R,10S,12S,13R,17R)-3,12-dihydroxy-10,13-dimethyl-2,3,4,5,6,7,8,-9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]-N-[2-(1-methylpyrrolidin-1-ium-1-yl)ethyl]pentanamide iodide (135)

(775) ##STR00107##

(776) Compound 108 (0.1 g 0.0002 mol) was dissolved in chloroform (10 mL). Methyl iodide (0.09 mL 0.0006 mol) was added and the solution was stirred for 1 week. The solvent was evaporated under reduced pressure. The residue was re-dissolved in methanol and washed with petroleum ether 60/80 (3×20 mL). The methanol phase was evaporated under reduced pressure to give the product as a green-brown oil was produced.

(777) Yield; 0.08 g, 0.0001 mol, 66%.

(778) .sup.1H NMR (CDCl.sub.3) (250 MHz) δ=0.70 (s, 3H, 18-CH.sub.3), 0.93 (s, 3H, 19-CH.sub.3), 1.00 (d, 3H, 21-CH.sub.3 J=10.0), 3.15 (s, 3H, CH.sub.3), 3.50-3.70 (multiple overlapping multiplets, 7H, CH/CH.sub.2), 3.95 (s, 1H, 12-CH) ppm.

(779) .sup.13C NMR (250 MHz) δ=13.2 (CH.sub.3, C18), 17.7 (CH.sub.3, C21), 22.56 (CH.sub.2) 23.7 (CH.sub.3, C19), 24.9 (CH.sub.2, C15), 27.5 (CH.sub.2, C7), 28.44 (CH.sub.2, C6), 28.7 (CH.sub.2, C16), 29.9 (CH.sub.2, C11), 31.1 (CH.sub.2, C2), 33.0 (CH.sub.2, C23), 33.9 (CH.sub.2, C22), 34.8 (CH, C9), 35.3 (C, C10), 36.4 (CH.sub.2, C1), 36.9 (CH, C20), 37.2 (CH.sub.2, C4), 37.4 (CH, C8), 43.6 (CH, C5), 63.5 (CH.sub.2), 66.1 (CH.sub.2), 72.5 (CH, C3), 74.0 (CH.sub.2, C12), 176.5 (CO, C24) ppm.

(780) IR=3394 (OH), 2916 (alkyl), 2857 (alkyl), 1646 (C═O), 1530, 1445, 1368, 1253, 1036 (RCH—OH) cm.sup.−1.

(781) MS (+ESI) m/z=Found 503.4201 M.sup.+; calculated for C.sub.31H.sub.55N.sub.2O.sub.3 503.4207; 1.2 ppm.

Synthesis of (4R)—N-[2-(1-allylpyrrolidin-1-ium-1-yl)ethyl]-4-[(3R,10S,13R,17R)-3-hyd-roxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]pentanamide bromide (136)

(782) ##STR00108##

(783) Compound 107 (0.5 g 0.001 mol) was dissolved in dichloromethane (15 mL). Ally bromide (2.79 mL, 0.02) was added and the solution was stirred for 48 hours at 50° C. The resulting precipitate was collected by vacuum filtration, triturated chloroform (3×20 mL) and dried under vacuum.

(784) Yield; 0.168 g, 0.0002 mol, 27%.

(785) Melting point: 211.6-214.8° C.

(786) .sup.1H NMR (MeOD) (250 MHz) δ=0.68 (s, 3H, 18-CH.sub.3), 0.94 (s, 3H, 19-CH.sub.3), 3.62 (overlapping mutliplets, 4H, CH.sub.2), 4.02 (d, 2H, CH.sub.2, J=7.5), 5.74 (multiple overlapping multiplets, 2H, ═CH.sub.2), 6.11 (m, 1H, ═CH—) ppm.

(787) .sup.13C NMR (62.9 MHz) δ=11.0 (CH.sub.3, C18), 17.3 (CH.sub.3, C21), 20.4 (CH.sub.2, C11), 22.3 (CH.sub.3, C19), 22.4 (CH) 23.7 (CH.sub.2, C15), 26.1 (CH.sub.2, C7), 26.8 (CH.sub.2, C6), 27.7 (CH.sub.2, C16), 29.6 (CH.sub.2, C2), 31.4 (CH.sub.2, C22), 32.3 (CH.sub.2, C23), 33.2 (C, C10), 34.1 (CH, C20), 34.9 (CH.sub.2, C1), 35.3 (CH, C8), 35.6 (CH.sub.2, C4), 40.0 (CH.sub.2, C12), 40.3 (CH, C9), 42.0 (CH, C5), 42.4 (C, C13), 55.6 (CH, C17), 56.4 (CH, C14), 57.8 (CH.sub.2), 61.3 (CH.sub.2), 62.1 (CH.sub.2) 70.8 (CH, C3), 125.2 (CH), 127.3 (CH.sub.2) 175.8 (CO, C24) ppm.

(788) IR=3411 (NH), 3198 (OH), 2925 (alkyl), 2852 (alkyl), 1637 (C═O), 1560, 1441, 1368, 1066 (R.sub.2CH—OH) cm.sup.−1.

(789) MS (+ESI) m/z=Found 513.4405 M+; calculated for C.sub.33H.sub.57N.sub.2O.sub.2 513.4415; 1.9 ppm.

Synthesis of (4R)—N-[2-(1-allylpyrrolidin-1-ium-1-yl)ethyl]-4-[(3R,10S,12S,13R,17R)-3-,12-dihydroxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahy-dro-1H-cyclopenta[a]phenanthren-17-yl]pentanamide bromide (137)

(790) ##STR00109##

(791) Compound 108 (0.1 g 0.0002 mol) was dissolved in chloroform (10 mL) with allyl bromide (0.08 mL, 0.0006 mol). The solution was stirred for 1 week. The solvent was evaporated under reduced pressure and the product was re-dissolved in methanol and washed with petroleum ether (3×10 mL). The solvent was then removed under reduced pressure to produce a white solid.

(792) Yield; 0.1 g, 0.0001 mol, 83%.

(793) Melting point: 176.2-179.8° C.

(794) .sup.1H NMR (MeOD) (250 MHz) δ=0.70 (s, 3H, 18-CH.sub.3), 0.93 (s, 3H, 19-CH.sub.3), 1.02 (d, 3H, 21-CH.sub.3, J=5.0), 2.22 (broad s, 4H, CH.sub.2), 3.515 (q, 2H, CH.sub.2), 3.65 (m, 3H, CH.sub.2/3-CH), 4.01 (broad s, 1H, 12-CH), 4.025 (d, 2H, CH.sub.2 J=7.5), 5.74 (m, 2H, CH.sub.2), 6.11 (m, 1H, ═CH—) ppm.

(795) .sup.13C NMR (62.9 MHz) δ=12.4 (CH.sub.3, C18), 16.8 (CH.sub.3, C21), 21.8 (CH.sub.2), 22.9 (CH.sub.3, C19), 24.0 (CH.sub.2, C15), 26.6 (CH.sub.2), 27.6 (CH.sub.2, C7), 27.9 (CH.sub.2, C6), 28.1 (CH.sub.2, C16), 30.9 (CH.sub.2, C2), 32.2 (CH.sub.2, C23), 33.0 (CH.sub.2, C22), 34.0 (CH, C9), 36.0 (CH.sub.2), 36.6 (CH, C20), 42.7 (CH, C5), 44.8 (C, C13), 47.1 (CH, C17), 58.4 (CH.sub.2), 61.9 (CH.sub.2), 62.8 (CH.sub.2) 71.7 (CH, C3), 73.1 (CH.sub.2, C12), 126.7 (CH.sub.2), 128.9 (CH.sub.2) 177.4 (CO, C24) ppm.

(796) IR=3305 (OH), 2921 (alkyl), 2857 (alkyl), 1641 (C═O), 1539, 1449, 1040 (R.sub.2CH—OH) cm.sup.−1.

(797) MS (+ESI) m/z=Found 529.4349 M.sup.+; calculated for C.sub.33H.sub.57N.sub.2O.sub.3, 529.4364; 2.8 ppm.

Synthesis of (4R)—N-[2-(1-benzylpyrrolidin-1-ium-1-yl)ethyl]-4-[(3R,10S,13R,17R)-3-hy-droxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]pentanamide bromide (138)

(798) ##STR00110##

(799) Compound 107 (0.5 g 0.001 mol) was dissolved in dichloromethane (20 mL). Benzyl bromide (1.42 mL 0.008 mol) was added and the solution was stirred for 24 hours. The resultant precipitate was collected by vacuum filtration to produce a white powder.

(800) Yield; 0.01 g, 0.0001 mol, 1.5%.

(801) Melting point: 216.2-220.8° C.

(802) .sup.1H NMR (CDCl.sub.3) (250 MHz) δ=0.66 (s, 3H, 18-CH.sub.3), 0.94 (s, 3H, 19-CH.sub.3), 3.54-3.76 (multiple overlapping multiplets, 7H, 3-CH/CH.sub.2), 4.57 (s, 2H, CH.sub.2), 7.56 (m, 5H, Ar—CH) ppm.

(803) .sup.13C NMR (62.9 MHz) δ=10.9 (CH.sub.3, C18), 17.3 (CH.sub.3, C21), 20.4 (CH.sub.2), 20.7 (CH.sub.2, C11), 22.4 (CH.sub.3, C19), 23.7 (CH.sub.2, C15), 27.7 (CH.sub.2, C16), 29.6 (CH.sub.2, C2), 31.5 (CH.sub.2, C22), 32.3 (CH.sub.2, C23), 34.1 (C, C10), 34.9 (CH, C20), 35.3 (CH.sub.2, C1), 35.6 (CH, C8), 35.7 (CH.sub.2, C4), 40.0 (CH.sub.2, C12), 40.3 (CH, C9), 41.9 (CH, C5), 42.4 (C, C13), 55.8 (CH, C17), 56.4 (CH, C14), 57.0 (CH), 61.2 (CH), 70.8 (CH, C3), 127.7 (CH), 129.0 (CH), 130.4 (CH), 132.2 (CH) 175.8 (CO, C24) ppm.

(804) IR=3343 (NH), 3241 (OH), 2929 (alkyl), 2844 (alkyl), 1667 (C═O), 1535, 1445, 1360, 1070 (R.sub.2CH—OH) cm.sup.−1.

(805) MS (+ESI) m/z=Found 563.4563 M.sup.+; calculated for C.sub.37H.sub.59N.sub.2O.sub.2 563.4571; 1.4 ppm.

Synthesis of (4R)-4-[(3R,10S,13R,17R)-3-hydroxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,12,1-4,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]-N-[2-[1-[(4-vinylphenyl)methyl]pyrrolidin-1-ium-1-yl]ethyl]pentanamide chloride

(806) ##STR00111##

(807) Compound 107 (0.5 g 0.001 mol) was dissolved in dichloromethane (20 mL). Vinyl benzyl chloride (1.74 mL 0.01 mol) was added and the solution was stirred for 48 hours at ambient temperature followed by heating at 50° C. for 15 h. The solvent was evaporated under reduced pressure. The material re-dissolved in methanol and washed with petroleum ether 60-80 (3×20 mL). A white solid was produced. This was collected by vacuum filtration and dried under vacuum.

(808) Yield: 0.08 g, 0.0001 mol, 12%.

(809) Melting point: >350° C.

(810) .sup.1H NMR (CDCl.sub.3) (250 MHz) δ=0.65 (s, 3H, 18-CH.sub.3), 0.94 (s, 3H, 19-CH.sub.3), 3.56 (multiple overlapping multiplets, 4H, CH.sub.2), 4.57 (s, 2H, CH.sub.2), 5.37 (d, 1H, ═CH—, J=12.5), 5.90 (d, 1H, ═CH, J=17.5), 6.80 (dd, 1H, ═CH, J=12.5), 7.60 (s, 4H, Ar—CH) ppm.

(811) .sup.13C NMR (62.9 MHz) δ=10.9 (CH.sub.3, C18), 17.3 (CH.sub.3, C21), 20.4 (CH.sub.2, C11), 20.7 (CH.sub.2), 22.4 (CH.sub.3, C19), 23.7 (CH.sub.2, C15), 26.1 (CH.sub.2, C7), 27.7 (CH.sub.2, C16), 29.6 (CH.sub.2, C2), 31.5 (CH.sub.2, C22), 32.3 (CH.sub.2, C23), 33.2 (C, C10), 34.1 (CH, C20), 34.9 (CH.sub.2, C1), 35.3 (CH, C8), 35.7 (CH.sub.2, C4), 40.0 (CH.sub.2, C12), 40.3 (CH, C9), 41.9 (CH, C5), 42.4 (C, C13), 55.8 (CH, C17), 56.4 (CH, C14), 57.0 (CH.sub.2), 61.2 (CH), 61.8 (CH.sub.2) 70.8 (CH, C3), 115.0 (CH), 126.6 (CH), 132.5 (CH), 135.4 (CH), 139.9 (CH), 175.8 (CO, C24) ppm.

(812) IR=3360 (OH), 2925 (alkyl), 2852 (alkyl), 1641 (C═O), 1539, 1437, 1368, 1036 (R.sub.2CH—OH) cm.sup.−1.

(813) MS (+ESI) m/z=Found 589.4719 M.sup.+; calculated for C.sub.39H.sub.61N.sub.2O.sub.2 589.4728; 1.5 ppm.

Synthesis of (4R)-4-[(3R,10S,13R,17R)-3-hydroxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,12,1-4,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]-1-[4-(2-hydro-xyethyl)-4-methyl-piperazin-4-ium-1-yl]pentan-1-one iodide (140)

(814) ##STR00112##

(815) Compound 118 (0.2 g 0.0004 mol) was dissolved in chloroform (5 mL). Iodomethane (0.29 mL, 0.002 mol) was added and the solution was stirred overnight at ambient temperature. The resulting precipitate was collected by vacuum filtration and was dried overnight in a desiccator. The product was an off-white solid.

(816) Yield; 0.01 g, 0.00001 mol, 4%.

(817) Melting point: 245.3-251.3° C.

(818) .sup.1H NMR (MeOD) (250 MHz) δ=0.69 (s, 3H, 18-CH.sub.3), 0.94 (s, 3H, 19-CH.sub.3), 0.98 (d, 3H, 21-CH.sub.3, J=5.0), 3.54-3.65 (multiple overlapping multiplets, 6H, CH.sub.2), 4.01 (multiple overlapping multiplets, 6H, CH.sub.2) ppm.

(819) .sup.13C NMR (62.9 MHz) δ=11.5 (CH.sub.3, C18), 16.4 (CH.sub.3, C21), 22.9 (CH.sub.2, C11), 25.9 (CH.sub.2), 27.8 (CH.sub.2), 32.2 (CH), 33.2 (CH), 34.6 (C, C10), 35.1 (CH, C20), 41.1 (CH, C5), 42.4 (C, C13), 44.5 (CH.sub.2), 45.8 (CH), 47.4 (CH), 60.4 (CH.sub.2), 61.3 (CH), 61.8 (CH), 67.1 (CH.sub.2), 70.1 (CH, C3), 71.2 (CH.sub.2), 75.7 (CH.sub.2), 76.3 (CH.sub.2), 80.6 (CH.sub.2) 174.8 (CO, C24) ppm.

(820) IR=3313 (OH), 2929 (alkyl), 2852 (alkyl), 1607 (C═O), 1466, 1249, 1044 (R.sub.2CH—OH) cm.sup.−1.

(821) MS (+ESI) m/z=Found 503.4195 M+; calculated for C.sub.31H.sub.55N.sub.2O.sub.3 503.4207; 2.4 ppm.

Synthesis of (4R)-1-[4-allyl-4-(2-hydroxyethyl)piperazin-4-ium-1-yl]-4-[(3R,10S, 13R, 17-R)-3-hydroxy-10, 13-dimethyl-2,3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahyd-ro-1H-cyclopenta[a]phenanthren-17-yl]pentan-1-one bromide (141)

(822) ##STR00113##

(823) Compound 118 (0.2 g 0.0004 mol) was dissolved in chloroform (5 mL). Allyl bromide (0.44 mL, 0.003 mol) was added and the solution was stirred overnight at ambient temperature. The resulting precipitate was collected by vacuum filtration and was dried overnight in a desiccator. The product was an off-white solid.

(824) Yield; 0.02 g, 0.00003 mol, 8%.

(825) Melting point: 194.9-197.8° C.

(826) .sup.1H NMR (MeOD) (250 MHz) δ=0.70 (s, 3H, 18-CH.sub.3), 0.94 (s, 3H, 19-CH.sub.3), 0.98 (d, 3H, 21-CH.sub.3, J=7.5), 3.54-3.65 (multiple overlapping multiplets, 6H, CH.sub.2), 4.01 (multiple overlapping multiplets, 6H, CH.sub.2), 4.27 (d, 2H, CH.sub.2, J=7.5), 5.76 (dd, 2H, ═CH.sub.2), 6.10 (m, 1H, ═CH—) ppm.

(827) .sup.13C NMR (62.9 MHz) δ=12.5 (CH.sub.3, C18), 18.9 (CH.sub.3, C21), 21.9 (CH.sub.2, C11), 23.9 (CH.sub.3, C19), 25.3 (CH.sub.2, C15), 27.6 (CH.sub.2, C6), 28.3 (CH.sub.2, C16), 29.3 (CH.sub.2, C2), 30.6 (CH.sub.2, C22), 31.1 (CH.sub.2, C23), 32.2 (C, C10), 35.6 (CH, C20), 36.4 (CH.sub.2, C1), 36.9 (CH, C8), 37.2 (CH.sub.2, C4), 41.5 (CH.sub.2, C12), 41.9 (CH, C9), 56.4 (CH, C17), 57.4 (CH, C14), 57.9 (CH.sub.2), 59.4 (CH.sub.2), 61.6 (CH.sub.2), 63.7 (CH.sub.2) 72.4 (CH, C3), 125.6 (CH), 129.9 (CH), 174.8 (CO, C24) ppm.

(828) IR=3350 (NH), 3241 (OH), 2938 (alkyl), 2850 (alkyl), 1633 (C═O), 1439, 1244, 1189.78 (C═O ester stretch), 1244 (R.sub.2CH—OH) cm.sup.−1.

(829) MS (ES) m/z=Found 529.4349 M.sup.+; calculated for C.sub.33H.sub.57N.sub.2O.sub.3 529.4364; 2.8 ppm.

Synthesis of (4R)-4-[(3R,10S,13R,17R)-3-hydroxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,12,1-4,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]-1-[4-(2-hydro-xyethyl)-4-pentyl-piperazin-4-ium-1-yl]pentan-1-one bromide (142)

(830) ##STR00114##

(831) Compound 103 (0.5 g 0.001 mol) was dissolved in dichloromethane (20 mL). Iodopentane (1.57 mL, 0.007 mol) was added and the solution was stirred overnight at 40° C. The resulting precipitate was collected by vacuum filtration and was dried overnight in a desiccator. The product was an off-white solid.

(832) Yield; 0.36 g, 0.0005 mol, 50.7%.

(833) Melting point: 104.8-108.6° C.

(834) .sup.1H NMR (MeOD) (250 MHz) δ=0.66 (s, 3H, 18-CH.sub.3), 0.92 (s, 3H, 19-CH.sub.3), 0.94 (multiple overlapping multiplets consisting of both CH.sub.3 and CH.sub.2), 3.08 (s, 5H, CH.sub.3/CH.sub.2). 3.51 (m, 1H, 3-CH) ppm.

(835) .sup.13C NMR (62.9 MHz) δ=11.0 (CH.sub.3, C18), 12.7, 17.3 (CH.sub.3, C21), 20.4 (CH.sub.2, C11), 21.8 (CH.sub.3, C19), 26.1 (CH.sub.2, C7), 29.6 (CH.sub.2, C2), 34.1 (C, C10), 35.4 (CH, C20), 35.7 (CH.sub.2, C1), 40.0 (CH.sub.2, C12), 40.4 (CH, C9), 42.0 (CH, C5), 42.4 (C, C13), 49.8, 55.9 (CH, C17), 56.4 (CH, C14), 70.8 (CH, C3), 175.7 (CO, C24) ppm.

(836) IR=3449 (OH), 2921 (alkyl), 2857 (alkyl), 1641 (C═O), 1548, 1445, 1364, 1036 (R.sup.2CH—OH) cm.sup.−1.

(837) MS (+ESI) m/z=Found 531.4880 M+; calculated for C.sub.34H.sub.63N.sub.2O.sub.2 531.4884; 0.8 ppm.

Synthesis of (4R)-4-[(3R,10S,13R,17R)-3-hydroxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,12,1-4,15,16,17-tetradecahydro-1H-cyclopenta[a]phenanthren-17-yl]-1-[4-(2-hydro-xyethyl)-4-[(4-vinylphenyl)methyl]piperazin-4-ium-1-yl]pentan-1-one chloride (143)

(838) ##STR00115##

(839) Compound 118 (0.2 g 0.0004 mol) was dissolved in chloroform (5 mL). Vinyl benzyl chloride (0.31 mL, 0.002 mol) was added and the solution was stirred overnight at ambient temperature. The resulting precipitate was collected by vacuum filtration and was dried overnight in a desiccator. The product was an off-white solid.

(840) Yield; 0.1 g, 0.0001 mol, 38%.

(841) Melting point: 143.6-146.0° C.

(842) .sup.1H NMR (MeOD) (250 MHz) δ=0.680 (s, 3H, 18-CH.sub.3), 0.94 (s, 3H, 19-CH.sub.3), 3.61 (multiple overlapping multiplets, 6H, CH.sub.2), 5.37 (d, 1H, ═CH—, J=12.5), 5.90 (d, 1H, ═CH, J=17.5), 6.80 (dd, 1H, ═CH, J=12.5), 7.59 (broad s, 4H, Ar—CH) ppm.

(843) .sup.13C NMR (62.9 MHz) δ=12.5 (CH.sub.3, C18), 18.9 (CH.sub.3, C21), 21.9 (CH.sub.2, C11), 23.9 (CH.sub.3, C19), 25.3 (CH.sub.2, C15), 27.6 (CH.sub.2, C7), 28.3 (CH.sub.2, C6), 29.3 (CH.sub.2, C16), 30.6 (CH.sub.2, C2), 31.2 (CH.sub.2, C22), 32.2 (CH.sub.2, C23), 35.6 (C, C10), 36.4 (CH, C20), 36.8 (CH.sub.2, C1), 37.2 (CH, C8), 40.6 (CH.sub.2, C12), 41.5 (CH, C9), 41.9 (CH, C5), 43.5 (C, C13), 56.5 (CH, C17), 57.4 (CH, C14), 57.9 (CH), 58.1 (CH.sub.2), 58.8 (CH.sub.2) 67.1 (CH) 72.4 (CH, C3), 116.7 (CH), 127.2 (CH), 128.0 (CH), 134.9 (CH), 137.0 (CH), 141.6 (CH), 174.8 (CO, C24) ppm.

(844) IR=3283 (OH), 2925 (alkyl), 2857 (alkyl), 1616 (C═O), 1445, 1044 (R.sub.2CH—OH) cm.sup.−1.

(845) MS (ES) m/z=Found 605.4669 M+; calculated for C.sub.39H.sub.61N.sub.2O.sub.3 605.4677; 1.3 ppm.

Germination Tests

(846) Many of the synthesised compounds were tested for their germinating and antimicrobial abilities against C. Difficile. Due to the insolubility of some of the compounds in water, DMSO, ethanol and methanol were used to dissolve the compounds.

Methods

(847) The C. Difficile reference strain, NCTC 11204 and C. Difficile ribotype 027 (R20291) (Anaerobic Reference Laboratory, Cardiff, UK) were used during testing.

Preparation of Spore Suspensions

(848) Spore suspensions of C. Difficile were prepared following the method proposed by Shetty et al. (1999). Briefly, Columbia base agar plates were inoculated with the relevant strain of C. Difficile and incubated for 72 hours anaerobically at 37° C. (MiniMACS anaerobic cabinet, Don Whitley Scientific, Shipley, UK). Then, the plates were removed and left or 24 hours in aerobic conditions at room temperature. Colonies were then harvested into 20 mL of 50% (w/v) ethanol and 50% saline, and vortex thoroughly. These were stored at 4° C. until needed.

(849) All experiments were performed in triplicate using spore suspensions containing 1×10.sup.7 CFU mL.sup.−1 spores of C. difficile NCTC 11204 and ribotype 027.

(850) Before use, 1 mL of spores were centrifuged at 13000 rpm for 10 minutes (Spectrafuge 24D; Labnet, Woodbridge, USA). The supernatant was discarded, and the pellet resuspended in 1 mL sterile distilled water and vortex mixed thoroughly.

(851) Germination solutions were prepared using 2% (w/v) of the compound in diluent (DMSO, ethanol, methanol, water) plus double strength thioglycollate medium (Oxoid, UK).

Heat Shock Method

(852) For the heat shock method, 100 μl spores were exposed to 100 μl of the germination solution and incubated at room temperature in air for 1 hour. The entire 200 μl sample was then added to 800 μl sterile distilled water to dilute out the germinant to ineffective concentrations. Samples were placed on heat at 70° C. for 20 minutes to eliminate any germinated, metabolically active spores. Control samples were kept on ice. Solutions were then diluted accordingly using sterile distilled water, and cultured onto fastidious anaerobic agar (Lab M, Bury, UK), supplemented with 0.1% (w/v) sodium taurocholate (ST) and 5% (w/v) defibrinated horse blood using the Miles and Misra method (Miles et al., 1938). These were then incubated anaerobically for 48 hours at 37° C. (MiniMACS anaerobic cabinet, Don Whitley Scientific, Shipley, UK) and the CFU mL.sup.−1 counted.

Germination Results for Strains 11204 and 027

(853) TABLE-US-00003 >1 log >1 log Com- reduc- reduc- pound Compound tion tion number name Structure on heat on ice 34 3- cholanamidopro pyl-allyl- dimethyl- ammonium bromide yes X 50 N-[2-(1- benzylpyrrolidin- 1-ium-1-yl)ethyl] cholanamide bromide yes yes 29 3-cholan amidopropyl- hexyl-dimethyl- ammonium iodide yes X 45 N-[2-(1- propylpyrrolidin- 1-ium-1-yl)ethyl] cholanamide iodide yes yes 16 N-octadecyl cholanamide 0 Yes (11204 and 027) X 7 N-[3- (dibutylamino) propyl] cholanamide Yes (11204) Yes (11204) 2 cholic acid benzyl amide X Yes (11204) 2/150 X Yes (11204) 2/148 Yes (027) X 10 N-(2-pyrrolidin-1- ylethyl) cholanamide Yes (11204) X

(854) The results from the table above show the compounds that gave a 1 log reduction or more between the initial spore count and the spore count after the heat and ice treatment. A reduction in the heat count suggests the compound is a germinant, whereas a reduction in the ice suggests a sporicide or a germinant/antimicrobial compound. Due to the nature of this test, it is difficult to distinguish between the two.