Methods of preparing nicotinamide riboside and derivatives thereof

Abstract

Methods of preparing nicotinamide riboside and derivatives thereof. In an aspect, the invention relates to a method of preparing a compound of formula (I) ##STR00001##
wherein
n is 0 or 1;
m is 0 or 1;
Y is O or S; R.sub.1 is selected from H, substituted or unsubstituted alkyl, substituted or unsubstituted alkenyl, substituted or unsubstituted alkynyl, substituted or unsubstituted aryl, substituted or unsubstituted primary or secondary amino, and substituted or unsubstituted azido; R.sub.2-R.sub.5, which may be the same or different, are each independently selected from H, substituted or unsubstituted alkyl, substituted or unsubstituted alkenyl, substituted or unsubstituted alkynyl, and substituted or unsubstituted aryl; and X.sup.− is an anion, selected from an anion of a substituted or unsubstituted carboxylic acid, a halide, a substituted or unsubstituted sulfonate, a substituted or unsubstituted phosphate, a substituted or unsubstituted sulfate, a substituted or unsubstituted carbonate, and a substituted or unsubstituted carbamate.

Claims

1. A compound of formula (IV) ##STR00011## wherein n is 0 or 1; m is 0 or 1; Y is S; R.sub.1 is selected from H, substituted or unsubstituted alkyl, substituted or unsubstituted alkenyl, substituted or unsubstituted alkynyl, substituted or unsubstituted aryl, substituted or unsubstituted primary or secondary amino, and unsubstituted azido; R.sub.2-R.sub.5, which may be the same or different, are each independently selected from H, substituted or unsubstituted alkyl, substituted or unsubstituted alkenyl, substituted or unsubstituted alkynyl, and substituted or unsubstituted aryl; and R.sub.6, R.sub.7 and R.sub.8, which may be the same or different, are each independently a hydroxyl protecting group; and X.sup.−is an anion selected from an anion of a substituted or unsubstituted carboxylic acid, a halide, a substituted or unsubstituted sulfonate, a substituted or unsubstituted phosphate, a substituted or unsubstituted sulfate, a substituted or unsubstituted carbonate, and a substituted or unsubstituted carbamate, provided that when n is 0, m is 1, and R.sub.2-R.sub.5 are each H, then R.sub.1 is not NH.sub.2.

2. The compound of claim 1, wherein R.sub.2-R.sub.5 are each H.

3. The compound of claim 1 of formula (IVA) ##STR00012## wherein n is 0 or 1; m is 0 or 1; Y is S; R.sub.1 is selected from H, substituted or unsubstituted alkyl, substituted or unsubstituted alkenyl, substituted or unsubstituted alkynyl, substituted or unsubstituted aryl, substituted or unsubstituted primary or secondary amino, and unsubstituted azido; and R.sub.2-R.sub.5, which may be the same or different, are each independently selected from H, substituted or unsubstituted alkyl, substituted or unsubstituted alkenyl, substituted or unsubstituted alkynyl, and substituted or unsubstituted aryl; and R.sub.6, R.sub.7 and R.sub.8, which may be the same or different, are each independently a hydroxyl-protecting group X.sup.−is an anion, selected from an anion of a substituted or unsubstituted carboxylic acid, a halide, a substituted or unsubstituted sulfonate, a substituted or unsubstituted phosphate, a substituted or unsubstituted sulfate, a substituted or unsubstituted carbonate, and a substituted or unsubstituted carbamate, provided that when n is 0, m is 1, and R.sub.2-R.sub.5 are each H, then R.sub.1 is not NH.sub.2.

4. A substantially isomerically pure compound of formula (III) having a beta-D-ribofuranosyl configuration: ##STR00013## wherein, n is 0 or 1; m is 0 or 1; Y is O or S; R.sub.1 is selected from H, substituted or unsubstituted alkyl, substituted or unsubstituted alkenyl, substituted or unsubstituted alkynyl, substituted or unsubstituted aryl, substituted or unsubstituted primary or secondary amino, and substituted or unsubstituted azido; and R.sub.2-R.sub.5, which may be the same or different, are each independently selected from H, substituted or unsubstituted alkyl, substituted or unsubstituted alkenyl, substituted or unsubstituted alkynyl, and substituted or unsubstituted aryl; and R.sub.6, R.sub.7 and R.sub.8, which may be the same or different, are each independently a hydroxyl-protecting group; or R.sub.6, R.sub.7 and R.sub.8 are each H; and wherein the compound has greater than 90% chemical purity, and comprises at least 98% diastereomeric purity of the beta-D-ribofuranosyl configuration.

5. The compound of formula (III) of claim 4, containing <3000 ppm methanol, under 0.1% (w/w) other solvents and under 0.2% (w/w) water.

6. A compound of claim 4, wherein the compound is 1-(beta-D-ribofuranosyl)-1,4-dihydronicotinamide: ##STR00014##

7. A compound according to claim 4, wherein R.sub.6, R.sub.7 and R.sub.8 are each independently an ester-type protecting group selected from acetyl, propionyl, isopropionyl, benzoyl, and trihaloacetyl.

8. A compound according to claim 7, wherein the ester-type protecting group is a protecting group selected from trifluoroacetyl and trichloroacetyl.

9. A compound according to claim 4, wherein the R.sub.6, R.sub.7 and R.sub.8 moieties are selected from substituted and unsubstituted acetyl, and substituted and unsubstituted benzoyl.

10. A compound according to claim 4, wherein at least two of R.sub.6, R.sub.7 and R.sub.8 are selected from unsubstituted acetyl or unsubstituted benzoyl.

11. A compound according to claim 10, wherein the compound is triacetyl-1,4-dihydronicotinamide riboside: ##STR00015##

12. A compound according to claim 10, wherein the compound is tribenzoyl-1,4-dihydronicotinamide riboside: ##STR00016##

13. A compound according to claim 4, wherein the compound is 1-(beta-D-ribofuranosyl)-1,4-dihydronicotinic acid: ##STR00017##

14. The compound of claim 1, wherein n is 1; m is 0 or 1; and Y is S.

15. The compound of claim 1, wherein X.sup.−is an anion selected from an anion of a substituted or unsubstituted carboxylic acid, a halide, a substituted or unsubstituted phosphate, a substituted or unsubstituted sulfate, a substituted or unsubstituted carbonate, and a substituted or unsubstituted carbamate.

16. The compound of claim 1, wherein n is 0 or 1; m is 0 or 1; Y is S; and R.sub.1 is selected from substituted or unsubstituted alkyl, substituted or unsubstituted alkenyl, substituted or unsubstituted alkynyl, and substituted or unsubstituted aryl.

17. The compound of claim 1, wherein n is 0 or 1; m is 0 or 1; Y is S; R.sub.1 is selected from substituted or unsubstituted alkyl, substituted or unsubstituted alkenyl, substituted or unsubstituted alkynyl, and substituted or unsubstituted aryl and X.sup.−is an anion selected from an anion of a substituted or unsubstituted carboxylic acid, a halide, a substituted or unsubstituted phosphate, a substituted or unsubstituted sulfate, a substituted or unsubstituted carbonate, and a substituted or unsubstituted carbamate.

18. The compound of claim 4, wherein n is 0 or 1; m is 0 or 1; and Y is S.

19. The compound of claim 4, wherein n is 1 and Y is S.

Description

THE EXAMPLES

(1) Embodiments of the present invention will now be described, with reference to the accompanying, non-limiting examples and drawings, in which:

(2) FIG. 1 shows the β-anomer forms of nicotinamide riboside, nicotinate riboside, nicotinamide mononucleotide and nicotinate mononucleotide, without counter ions;

(3) FIG. 2 depicts Scheme A, which is a scheme illustrating, in general terms, that compounds of formula (IV) may be used to prepare compounds of formula (III), as described in Example 1; that compounds of formula (III) may be used to prepare compounds of formula (II), as described in Example 2; and that compounds of formula (II) may be used to prepare compounds of formula (I), as described in Example 3; wherein n, m, Y, R.sub.1-R.sub.8 and X.sup.− are as defined above;

(4) FIG. 3 depicts Scheme B, which is a scheme illustrating, in general terms, that triacetyl nicotinamide riboside, triflate salt may be used to prepare triacetyl-1,4-dihydronicotinamide riboside, as described in Example 1(A), and that triacetyl-1,4-dihydronicotinamide riboside may be used to prepare 1-(beta-D-ribofuranosyl)-1,4-dihydronicotinamide, as described in Example 2, and that 1-(beta-D-ribofuranosyl)-1,4-dihydronicotinamide may be used to prepare nicotinamide riboside, chloride salt, as described in Examples 3(A), 3(E) and 3(F). The β-anomers of all of the mentioned compounds are shown. It will be appreciated that Scheme B is merely exemplary and is not to be construed as limiting the invention thereto;

(5) FIG. 4 shows the β-anomer forms of triacetyl-1,4-dihydronicotinamide riboside (Example 1(A)), triacetyl O-ethyl-1, 4-dihydronicotinate riboside, (Example 1(B)), tribenzoyl-1, 4-dihydronicotinamide riboside (Example 1(C)), and 1-(beta-D-ribofuranosyl)-1,4-dihydronicotinamide (Example 2); and

(6) FIG. 5 shows the β-anomer forms of nicotinamide riboside, chloride salt (Examples 3(A), 3(E) and 3(F)), nicotinamide riboside, acetate salt (Example 3(B)), nicotinamide riboside, formate salt (Example 3(C)), and nicotinamide riboside, trifluoroacetate salt (Example 3(D)).

Example 1

(7) Compounds of formula (III) were prepared in accordance with the invention as follows. The pH of the reaction media described in the following examples was in the region of about 6-8.

Example 1(A): Preparation of Reduced Triacetyl Nicotinamide Riboside, Namely Triacetyl-1,4-Dihydronicotinamide Riboside, a Compound of Formula (III) (the β-Anomer Form of Which is Shown in FIG. 4)

(8) Reduction: All solvents were degassed prior to use by sonication and argon bubbling. Sodium dithionite (0.656 g, 3.76 mmol, 2 eq) and sodium hydrogencarbonate (0.79 g, 9.40 mmol, 5 eq) were added to a clean, dry round bottom flask with a magnetic stirrer and placed under inert atmosphere. A compound of formula (IV), namely triacetyl nicotinamide riboside, triflate (CF.sub.3SO.sub.3.sup.−, also known as −OTf) salt (1 g, 1.88 mmol, 1 eq) was then dissolved in a minimum amount of water (<10 ml) and slowly added to the reaction vessel. Once the reaction settled, further water was added to the reaction until all of the reactants had dissolved (<10 ml) and was left to stir for 20 minutes. The aqueous solution was then extracted with three half portions of dichloromethane (DCM). The DCM fractions were collected and concentrated under reduced pressure, affording the triacetyl-1, 4-dihydronicotinamide riboside derivative (triacetyl-NRH) with residual amounts of starting material (<5%). The aqueous layer was subjected to the above conditions a second time to increase yields which averaged 65%. Ethyl acetate was also an excellent alternative extraction solvent in place of DCM, yielding a 75% yield.

(9) .sup.1H-NMR (MeOD, 400 MHz)-δ7.15 (s, 1H, H-5), 5.95 (d, 1H, J=7.21 Hz, H-6), 5.25 (d, 1H, J=2.84 Hz) & 5.17 (d, 1H, J=1.80 Hz) (H-8 & H-7), 4.96 (d, 1H, J=7.09 Hz, H-4), 4.87 (ABX, 1H, Jaa=8.18 Hz, Jab=3.60 Hz, H-9), 4.26 (d, 2H, J=3.20 Hz, H-10 & H-10′) 4.19 (m, 1H, J=3.00 Hz, H-3), 3.13 (m, 2H, J=1.18 Hz, H-2), 2.13 (s, 3H), 2.11 (s, 3H), 2.10 (s, 3H) (H-13, H-15, H-17). 13C-NMR (MeOD, 125 MHz)-δ172.80 (C-11), 170.40 (C-12, C-14, C-16), 137.90 (C-5), 125.20 (C-4), 105.12 (C-6), 95.24 (C-3), 83.49 (C-9), 71.18 (C-8), 70.26 (C-7), 61.55 (C-10), 22.16 (C-2), 21.52 (C-13, C-15, C-17). HMRS m/z: 383.1445; Calc. Mass: 383.1454.

(10) The compound of formula (IV), namely triacetyl nicotinamide riboside, triflate (-OTf) salt was prepared as follows. Nicotinamide (10 g, 81.89 mmol, 1 eq) was silylated using TMSCI (15.6 ml, 122.85 mmol, 1.5 eq) in HMDS (100 ml) at 130° C. in quantitative yield, in order to force the β-selectivity via the following Vorbrüggen reaction. Ribose tetraacetate (also known as tetraacetate riboside) was reacted with the resultant silylated nicotinamide in the presence of 5 equivalents of TMSOTf. The reactants were shaken in a 1.5 ml steel vessel with a 5 mm diameter steel ball bearing in a Retsch MM400 mixer mill at 25 Hz for 0.5 h. At this point the formed triacetylated nicotinamide riboside (compound of formula (IV)) could be isolated. It will be appreciated that the triacetyl nicotinamide riboside is not limited to being produced by this exact method, and could, for example, be produced using a conventional Vorbrüggen reaction as described, for example, in International PCT patent publication no. WO 2007/061798 or in T. Yang, N. Y. K. Chan and A. A. Sauve, Journal of Medicinal Chemistry, 2007, 50, 6458-6461.

(11) .sup.1H-NMR (MeOD, 400 MHz)-δ 9.61 (s, 1H, aromatic), 9.30 (dt, 1H, J=6.3, 1.4 Hz, aromatic), 9.10 (dt, 1H, J=8.2, 1.4 Hz, aromatic), 8.37 (dd, 1H, J=8.2, 6.3 Hz, aromatic), 6.60 (d, 1H, J=3.9 Hz, H-1 (anomeric)), 5.60 (dd, 1H, J=5.6, 3.9 Hz, H-2), 5.46 (t, 1H, J=5.6 Hz, H-3), 4.81-4.84 (m, 1H, H-4), 4.61 (ABX, 1H, J.sub.a,a′=13.1 Hz, J.sub.a,b=3.5 Hz, H-5), 4.51 (ABX, 1H, J.sub.a,a′=13.0 Hz, J.sub.a,b=2.8 Hz, H-5), 2.20 (s, 3H, OAc), 2.17 (s, 3H, OAc), 2.16 (s, 3H, OAc).
.sup.13C-NMR (MeOD, 125 MHz)-δ 172.1, 171.6, 171.2 (3x C═OCH.sub.3), 164.9 (C═ONH.sub.2) 147.0, 144.3, 142.3, 136.2, 129.6, (aromatic), 121.6 (q, J=320.2 Hz, CF.sub.3), 99.4 (C-1 (anomeric)), 84.4 (C-4), 77.6 (C-2), 70.7 (C-3), 63.5 (C-5), 20.7 (OAc), 20.3 (OAc), 20.2 (OAc).
.sup.19F-NMR (MeOD, 376 MHz)-δ −79.9 (triflate counterion)

Example 1(B): Preparation of Reduced Triacetyl Nicotinate Ester Riboside, Namely Triacetyl O-ethyl-1, 4-Dihydronicotinate Riboside, a Compound of Formula (III) (the β-Anomer Form of Which is Shown in FIG. 4)

(12) Reduction: A compound of formula (IV), namely triacetyl O-ethyl nicotinate riboside, triflate (-OTf) salt (2.30 g, 4.2 mmol, 1 eq) was dissolved in 20 mL H.sub.2O and a solution of a solution of NaHCO.sub.3 (1.77 g, 21.0 mmol, 5 eq) and sodium dithionite (1.47 g, 8.22 mmol, 2 eq) in 30 mL H.sub.2O was added and stirred for 2 hrs. The yellow solution obtained was then washed with 2× ethyl acetate (EtOAc, 40 mL), the organic layer dried over MgSO.sub.4, filtered and concentrated to provide 900 mg (39% yield) of 2,3,5-triacetyl O-ethyl-1, 4-dihydronicotinate riboside (a yellow oil) without further purification. 80% purity based on .sup.1H-NMR.

(13) .sup.1H-NMR-δ 7.27 (1H, s, H-6), 6.05 (1H, dd, J=8.2, 1.5 Hz, H-7), 5.26 (1H, dd, J=5.8, 2.8 Hz, H-3), 5.23 (1H, dd, J=6.9, 5.8 Hz, H-2), 5.08 (1H, d, J=6.9 Hz, H-1), 4.91 (1H, dt, J=8.3, 3.5 Hz, H-8), 4.24-4.30 (3H, m, H-4, H-5, H-5′), 4.11 (2H, q, J=7.2 Hz, H-11), 3.04-3.06 (2H, m, H-9), 2.16 (3H, s, OAc), 2.12 (3H, s, OAc), 2.09 (3H, s, OAc), 1.25 (3H, t, J=7.2 Hz, H-12).
.sup.13C-NMR-δ 172.2, 171.5, 171.3, 169.8, (3x C═O—CH.sub.3 and C═O—OEt), 139.9 (C-6), 126.3 (C-7), 106.4 (C-8), 101.5 (C-10), 94.2 (C-1), 80.4 (C-4), 72.3 (C-2), 72.1 (C-3), 64.8 (C-5), 61.0 (C-11), 23.4 (C-9), 20.7, 20.5, 20.3 (3x C═O—CH.sub.3), 14.8 (C-12).

(14) The compound of formula (IV), namely triacetyl O-ethyl nicotinate riboside, triflate (-OTf) salt was prepared as follows. Ribose tetraacetate (also known as tetraacetate riboside) was reacted with ethyl nicotinate (Sigma Aldrich) using the general ball milling Vorbrüggen procedure described in Example 1(A) above. The reactants, namely 1 eq tetraacetate riboside, 1 eq TMSOTf, 1 eq ethyl nicotinate, were reacted for 30 mins in a 1.5 ml steel vessel with a 1.5 cm diameter steel ball bearing in a Retsch MM400 mixer mill at 25 Hz. The crude reaction mixture (containing some unreacted ethyl nicotinate and starting sugar, <10%) was used for the reduction step (described above) without further purification. It will be appreciated that the triacetyl O-ethyl nicotinate riboside, triflate (-OTf) salt is not limited to being produced by this exact method, and could, for example, be produced using a conventional Vorbrüggen reaction as described, for example, in International PCT patent publication no. WO 2007/061798 or in T. Yang, N. Y. K. Chan and A. A. Sauve, Journal of Medicinal Chemistry, 2007, 50, 6458-6461.

(15) .sup.1H-NMR (D.sub.2O, 400 MHz)-δ 9.45 (s, 1H, aromatic), 9.14 (d, 1H, J=6.1 Hz, aromatic), 9.02 (d, 1H, J=7.8 Hz, aromatic), 8.18 (t, 1H, J=6.7 Hz, aromatic), 6.51 (d, 1H, J=4.1 Hz, H-1 (anomeric)), 5.47 (t, 1H, J=4.4 Hz, H-2), 5.36 (t, 1H, J=4.7 Hz, H-3), 4.81-4.84 (m, 1H, H-4), 4.45-4.48 (m, 2H, H-5), 4.36 (q, 2H, J=7.0 Hz, C═OCH.sub.2CH.sub.3), 2.04 (s, 3H, OAc), 2.02 (s, 3H, OAc), 1.98 (s, 3H, OAc), 1.25 (t, 3H, J=7.0 Hz, C═OCH.sub.2CH.sub.3).
.sup.19F-NMR (D.sub.2O, 376 MHz)-δ −79.0 (triflate counterion)

Example 1(C): Preparation of Reduced Tribenzoyl Nicotinamide Riboside, Namely Tribenzoyl-1, 4-Dihydronicotinamide Riboside, a Compound of Formula (III) (the β-Anomer Form of Which is Shown in FIG. 4)

(16) Reduction (unoptimised): A compound of formula (IV), namely tribenzoyl nicotinamide riboside, triflate (-OTf) salt was dissolved in minimal methanol and transferred to a round bottomed flask, 10 mL of H.sub.2O was added to the solution and most of the methanol removed via rotary evaporation. The starting material crashed out of solution and 20 mL of diethyl ether (Et.sub.2O) was added until the solids solubilized into a biphasic system. A solution of NaHCO.sub.3 (420 mg, 5 mmol, 5 eq) and sodium dithionite (348 mg, 2 mmol, 2 eq) in 10 mL H.sub.2O was added and stirred for 2 hrs. The layers were separated and the ether layer was dried over MgSO.sub.4 and concentrated to provide 428 mg (76% yield) of tribenzoyl-1, 4-dihydronicotinamide riboside (yellow solid) without further purification. 80% purity based on .sup.1H-NMR. Pure material is obtained by Biotage purification.

(17) .sup.1H-NMR-δ 8.01-8.04 (2H, m, OBz), 7.81-7.86 (4H, m, OBz), 7.25-7.55 (9H, m, OBz), 7.13 (1H, s, H-6), 6.01 (1H, dd, J=8.2, 1.5 Hz, H-7), 5.68 (1H, dd, J=6.2, 3.5 Hz, H-3), 5.57 (1H, dd, J=6.7, 6.2 Hz, H-2), 5.29 (1H, d, J=6.7 Hz, H-1), 4.61-4.68 (2H, m, H-8, H-5), 4.50-4.55 (2H, m, H-4, H-5′), 3.93-3.94 (2H, m, H-9).
.sup.13C-NMR-δ 172.7, 167.6, 166.7, 166.6 (3x C═O—C.sub.6H.sub.5, C═ONH.sub.2), 138.1 (C-6), 134.9, 134.8, 134.6, 130.9, 130.8, 130.7, 130.3, 130.0, 129.8, 129.7 (3x OBz), 125.7 (C-7), 105.9 (C-8), 94.9 (C-1), 80.3 (C-4), 72.9 (C-2), 72.7 (C-3), 65.4 (C-5), 23.6 (C-9).

(18) The compound of formula (IV), namely tribenzoyl nicotinamide riboside, triflate (-OTf) salt was prepared as follows. Ribose tetraacetate (also known as tetraacetate riboside) was reacted with TMS-nicotinamide (trimethylsilyl N-trimethylsilylpyridine-3-carboximidate, available from Sigma-Aldrich) using the general ball milling Vorbrüggen procedure described in Example 1(A) above. The reactants, namely 1 eq 1-acetate-tribenzoate riboside, 1 eq TMSOTf and 1 eq TMS-nicotinamide, were reacted for 30 mins in a 1.5 ml steel vessel with a 1.5 cm diameter steel ball bearing in a Retsch MM400 mixer mill at 25 Hz. 1 eq of DCE (dichloroethylene) was required and the crude reaction mixture (containing some unreacted nicotinamide and starting benzoate sugar, <10%) was used for the reduction step (described above) without further purification. It will be appreciated that the tribenzoyl nicotinamide riboside, triflate (-OTf) salt is not limited to being produced by this exact method, and could, for example, be produced using a conventional Vorbrüggen reaction as described, for example, in International PCT patent publication no. WO 2007/061798 or in T. Yang, N. Y. K. Chan and A. A. Sauve, Journal of Medicinal Chemistry, 2007, 50, 6458-6461.

(19) .sup.1H-NMR (MeOD, 400 MHz)-δ 9.59 (s, 1H, aromatic), 9.31 (d, 1H, J=6.4 Hz, aromatic), 8.94 (d, 1H, J=8.1 Hz, aromatic), 8.15 (dd, 1H, J=8.1, 6.4 Hz, aromatic), 7.90-7.94 (m, 6H, OBz), 7.50-7.54 (m, 3H, OBz), 7.31-7.38 (m, 6H, OBz), 6.79 (d, 1H, J=3.9 Hz, H-1 (anomeric)), 5.97 (dd, 1H, J=5.6, 3.9 Hz, H-2), 5.87 (t, 1H, J=5.6 Hz, H-3), 5.13-5.16 (m, 1H, H-4), 4.83-4.91 (m, 2H, H-5).
.sup.19F-NMR (MeOD, 376 MHz)-δ −79.1 (triflate counterion)

Example 2

(20) A compound of formula (II), namely NRH (reduced nicotinamide riboside, also known as 1-(beta-D-ribofuranosyl)-1,4-dihydronicotinamide (the β-anomer form of which is shown in FIG. 4) was prepared as follows. The pH of the reaction medium described in the following example was in the region of about 6-8.

(21) Reduced triacetyl nicotinamide riboside, namely triacetyl-1, 4-dihydronicotinamide riboside, a compound of formula (III), prepared in Example 1(A) above, was deprotected using mechanochemical (MeOH, NaOH) processes to remove the acetyl moiety afforded NRH quantitatively. 100 mgs of (III) was dissolved in 0.5 mL of MeOH containing 0.05 g of NaOH. The compounds were reacted for 30 mins in a 1.5 ml steel vessel with a 1.5 cm diameter steel ball bearing in a Retsch MM400 mixer mill at 25 Hz.

(22) 1H-NMR (MeOD, 400 MHz)-δ7.18 (s, 1H, H-5), 6.14 (d, 1H, J=8.28 Hz, H-6), 4.85 (m, 1H, H-3), 4.76 (d, 1H, J=5.77 Hz, H-4), 4.04 (m, 2H, H-7&H-8), 3.93 (m, 1H, J=2.76, H-9), 3.72 (ABX, 1H, Jaa=12.55 Hz, Jab=3.51 Hz, H-10), 3.65 (ABX, 1H, Jaa=12.55 Hz, Jab=4.02 Hz, H-10′), 3.10 (q, 2H, J=1.51 Hz H-2). 13C-NMR (MeOD, 125 MHz)-δ172.88 (C-11), 137.83 (C-5), 125.29 (C-4), 105.19 (C-6), 95.00 (C-3), 83.54 (C-9), 71.10 (C-8), 70.20 (C-7), 61.61 (C-10), 22.09 (C-2); HRMS m/z: 257.1130; Calc. Mass: 257.1137.

(23) It will be appreciated that the deprotection step as described above may be used to deprotect any other compound of formula (III), including, but not limited to, reduced triacetyl nicotinate ester riboside, namely 2,3,5-triacetyl O-ethyl-1, 4-dihydronicotinate riboside, prepared in Example 1(B), and reduced tribenzoyl nicotinamide riboside, namely tribenzoyl-1, 4-dihydronicotinamide riboside, prepared in Example 1(C). The deprotection step may also be modified to suit particular requirements.

Example 3

(24) Compounds of formula (I) were prepared in accordance with the invention as follows. The pH of the reaction media described in the following examples was in the region of about 6-8.

Example 3(A): Preparation of Nicotinamide Riboside, Chloride Salt (the β-Anomer Form of Which is Shown in FIG. 5)

(25) A compound of formula (II), namely NRH (reduced nicotinamide riboside, shown in FIG. 2; 50 mg, 0.20 mmol, 1 eq), was dissolved in 5 mL H.sub.2O and then 1 eq (i.e. 0.20 mmol) of ammonium chloride was added in one portion. Activated charcoal (˜10 mg, i.e. 0.80 mmol) was then added and the mixture stirred at RT for ˜1 hr and then filtered and freeze-dried to give the chloride salt of nicotinamide riboside, quantitatively, i.e. 100% conversion and pure product.

(26) .sup.1H-NMR (D.sub.2O, 400 MHz)-δ 9.46 (s, 1H, aromatic), 9.12 (dt, 1H, J=6.3, 1.4 Hz, aromatic), 8.83 (dt, 1H, J=8.2, 1.4 Hz, aromatic), 8.13 (dd, 1H, J=8.2, 6.3 Hz, aromatic), 6.13 (d, 1H, J=4.3 Hz, H-1 (anomeric)), 4.37 (t, 1H, J=4.7 Hz, H-2), 4.31-4.34 (m, 1H, H-4), 4.21 (t, 1H, J=4.7 Hz, H-3), 3.90 (ABX, 1H, J.sub.a,a′=13.0 Hz, J.sub.a,b=3.5 Hz, H-5), 3.75 (ABX, 1H, J.sub.a,a′=13.0 Hz, J.sub.a′,b=2.8 Hz, H-5).

(27) It will be appreciated that the NRH may be that obtained in Example 2, or may be obtained commercially from e.g. Diverchim, 100, rue Louis Blanc, 60 765 Montataire Cedex, France—(CAS Registry Number:19132-12-8) either as a pure product or as a mixture of anomers.

Example 3(B): Preparation of Nicotinamide Riboside, Acetate Salt (the β-Anomer Form of Which is Shown in FIG. 5)

(28) The method described in Example 3(A) was carried out, except that 1 eq (i.e. 0.20 mmol) of ammonium acetate was added. The acetate salt of nicotinamide riboside was obtained, quantitatively.

(29) .sup.1H-NMR (D.sub.2O, 400 MHz)-δ 9.46 (s, 1H, aromatic), 9.12 (d, 1H, J=6.3 Hz, aromatic), 8.83 (d, 1H, J=8.2 Hz, aromatic), 8.12 (m, 1H, aromatic), 6.09 (d, 1H, J=4.4 Hz, H-1 (anomeric)), 4.36 (t, 1H, J=4.7 Hz, H-2), 4.32-4.35 (m, 1H, H-4), 4.21 (t, 1H, J=4.7 Hz, H-3), 3.91 (ABX, 1H, J.sub.a,a′=13.1 Hz, J.sub.a,b=2.8 Hz, H-5), 3.75 (ABX, 1H, J.sub.a,a′=13.0 Hz, J.sub.a′,b=3.5 Hz, H-5), 1.79 (s, 3H, OAC).

Example 3(C): Preparation of Nicotinamide Riboside, Formate Salt (the β-Anomer Form of Which is Shown in FIG. 5)

(30) The method described in Example 3(A) was carried out, except that 1 eq (i.e. 0.20 mmol) of ammonium formate (methanoate) was added. The formate salt of nicotinamide riboside was obtained, quantitatively.

(31) .sup.1H-NMR (D.sub.2O, 400 MHz)-δ 9.46 (s, 1H, aromatic), 9.12 (d, 1H, J=6.3 Hz, aromatic), 8.83 (d, 1H, J=8.2 Hz, aromatic), 8.29 (s, 1H, formate), 8.12 (m, 1H, aromatic), 6.09 (d, 1H, J=4.4 Hz, H-1 (anomeric)), 4.36 (t, 1H, J=4.7 Hz, H-2), 4.31-4.34 (m, 1H, H-4), 4.21 (t, 1H, J=4.7 Hz, H-3), 3.91 (ABX, 1H, J.sub.a,a′=13.1 Hz, J.sub.a,b=3.5 Hz, H-5), 3.79 (ABX, 1H, J.sub.a,a′=13.0 Hz, J.sub.a′,b=2.8 Hz, H-5).

Example 3(D): Preparation of Nicotinamide Riboside, Trifluoroacetate Salt (the β-Anomer Form of Which is Shown in FIG. 5)

(32) The method described in Example 3(A) was carried out, except that 1 eq (i.e. 0.20 mmol) of ammonium trifluoroacetate was added. The trifluoroacetate salt of nicontinamide riboside was obtained, quantitatively.

(33) .sup.1H-NMR (D.sub.2O, 400 MHz)-δ 9.46 (s, 1H, aromatic), 9.12 (d, 1H, J=6.3 Hz, aromatic), 8.83 (d, 1H, J=8.2, aromatic), 8.13 (dd, 1H, J=8.2, 6.3 Hz, aromatic), 6.13 (d, 1H, J=4.3 Hz, H-1 (anomeric)), 4.35 (t, 1H, J=4.7 Hz, H-2), 4.31-4.34 (m, 1H, H-4), 4.20 (t, 1H, J=4.7 Hz, H-3), 3.89 (ABX, 1H, J.sub.a,a′=13.0 Hz, J.sub.a,b=3.6 Hz, H-5), 3.74 (ABX, 1H, J.sub.a,a′=13.0 Hz, J.sub.a′,b=2.9 Hz, H-5). .sup.19F-NMR (D.sub.2O, 376 MHz)-δ −75.7 (CF.sub.3COO.sup.−).

Example 3(E): Preparation of Nicotinamide Riboside, Chloride Salt (the β-Anomer Form of Which is Shown in FIG. 5)

(34) An alternative method to that described in Example 3(A) was carried out as follows. NRH (reduced nicotinamide riboside, shown in FIG. 4; 50 mg, 0.20 mmol, 1 eq) was dissolved in 5 mL H.sub.2O:EtOAc (1:1) and then 1 eq. (i.e. 0.20 mmol) of ammonium chloride was added in one portion. Upon work-up after 1 hr, no oxidation had taken place and the starting materials were fully recovered. The recovered NRH and ammonium chloride were re-suspended in the same solvent system with addition of activated charcoal (˜10 mg, i.e. 0.8 mmol) and stirred at RT for 1 hr. Subsequent filtration and freeze-drying afforded the chloride salt of nicotinamide riboside in quantitative yield. Thus it was concluded that a carbon-containing catalyst, e.g. activated charcoal, was essential to the method.

(35) .sup.1H-NMR (D.sub.2O, 400 MHz)-δ 9.46 (s, 1H, aromatic), 9.12 (dt, 1H, J=6.3, 1.4 Hz, aromatic), 8.83 (dt, 1H, J=8.2, 1.4 Hz, aromatic), 8.13 (dd, 1H, J=8.2, 6.3 Hz, aromatic), 6.13 (d, 1H, J=4.3 Hz, H-1 (anomeric)), 4.37 (t, 1H, J=4.7 Hz, H-2), 4.31-4.34 (m, 1H, H-4), 4.21 (t, 1H, J=4.7 Hz, H-3), 3.90 (ABX, 1H, J.sub.a,a′=13.0 Hz, J.sub.a,b=3.5 Hz, H-5), 3.75 (ABX, 1H, J.sub.a,a′=13.0 Hz, J.sub.a′,b=2.8 Hz, H-5).

Example 3(F): Preparation of Nicotinamide Riboside, Chloride Salt (the β-Anomer Form of Which is Shown in FIG. 5)

(36) The method described in Example 3(E) was carried out, except that NRH (reduced nicotinamide riboside, shown in FIG. 4; 50 mg, 0.20 mmol, 1 eq) was dissolved in 5 mL H.sub.2O:THF (1:1), instead of H.sub.2O:EtOAc (1:1), and then 1 eq (i.e. 0.20 mmol) of ammonium chloride was added in one portion. Upon work-up after 1 hr, no oxidation had taken place and the starting materials were fully recovered. The recovered NRH and ammonium chloride were re-suspended in the same solvent system with addition of activated charcoal (˜10 mg, i.e. 0.8 mmol) and stirred at RT for 1 hr. Subsequent filtration and freeze-drying afforded the chloride salt of nicotinamide riboside in quantitative yield. Thus it was concluded that a carbon-containing catalyst, e.g. activated charcoal, was essential to the method.

(37) .sup.1H-NMR (D.sub.2O, 400 MHz)-δ 9.46 (s, 1H, aromatic), 9.12 (dt, 1H, J=6.3, 1.4 Hz, aromatic), 8.83 (dt, 1H, J=8.2, 1.4 Hz, aromatic), 8.13 (dd, 1H, J=8.2, 6.3 Hz, aromatic), 6.13 (d, 1H, J=4.3 Hz, H-1 (anomeric)), 4.37 (t, 1H, J=4.7 Hz, H-2), 4.31-4.34 (m, 1H, H-4), 4.21 (t, 1H, J=4.7 Hz, H-3), 3.90 (ABX, 1H, J.sub.a,a′=13.0 Hz, J.sub.a,b=3.5 Hz, H-5), 3.75 (ABX, 1H, J.sub.a,a′=13.0 Hz, J.sub.a′,b=2.8 Hz, H-5).