Synthesis of novel xylofuranosyloxymethyl nucleosides

INTRODUCTION

The chemistry of nucleosides¹ was revolutionized after the discovery of modified nucleosides² as effective therapeutic agents for the treatment of AIDS and cancer.^3,4 Examples of modified nucleosides approved by US Food and Drug administration are: 3’-azidothymidine (AZT), dideoxycytidine (ddC), dideoxyinosine (ddI), 2’,3’-didehydro-2’,3’-dideoxythymidine (d₄T), 3’-azido-2’,3’-dideoxyuridine (AzddU).⁵ Another important class of sugar modified nucleosides are acyclic nucleosides, where a sugar unit is replaced by acyclic group.⁶ The first acyclic nucleoside 9-[(2-hydroxyethoxy) methyl] guanine (ACV) is a highly specific inhibitor of herpes simplex virus proliferation (Figure 1). Ganciclovir (GCV), structurally different from acyclovir by the addition of a hydroxymethyl group to the acyclic side chain is effective for cytomegalovirus infections.

 

Figure 1: Therapeutically important nucleoside analogues Click here to View Figure

 

Glycosylation reaction ⁷ is crucial for the synthesis of complex glycoconjugates, including the nucleosides. Although there has been fairly significant advancement in the field of glycoconjugate synthesis, the efficiency of glycosidic bond formation in achieving high chemical yield, stereoselectivity and regioselectivity⁸ still remains challenging.⁹ The  classical aspects of carbohydrate chemistry centers around the anomeric carbon where glycosidic bond formation takes place via glycosyl cation¹⁰ and stereochemical outcome depends on attaching the aglycones to give either α– or β–glycosides.¹¹ We were interested in developing efficient glycosylation methods for the synthesis of glycoconjugates.¹² In continuation of our efforts to develop efficient glycosylation reactions,we designed an acetoxymethoxy leaving group at the anomeric centre and demonstrated its ability as glycosyl donor.¹³ During our investigations, we found that when a glycosylation reaction was performed with O-nucleophiles, it resulted into O-glycosides, whereas N-nucleophiles afforded glycosyloxymethyl derivatives.^13,14 The formation of glycosyloxymethyl derivatives was explained based on capturing the glycosyloxymethyl cation ‘b’ by N-nucleophiles such as nucleic acid bases (Scheme 1).¹⁴ In order to confer generality to the concept developed for synthesis of novel nucleosides and to probe the reactivity of other glycosyl donors we report the activation of xylofuranosyl donor. Furthermore, given the medicinal importance of acylic nucleosides, synthesis of modified glycosyloxymethyl nucleosides may provide new therapeutic agents that can inhibit herpes simplex virus.

 

Scheme 1:  Activation of acetoxymethoxy glycosyl donor Click here to View Scheme

 

RESULTS AND DISCUSSION

The key substrate xylofuranosyl donor 4 was prepared starting from 1-O-acetyl-2,3,5-tri-O-benzoyl-α/β-D-xylofuranoside (1).14 Compound 1 on reaction with propargyl alcohol and BF3.OEt2 at room temperature gave 2-propynyl 2,3,5-tri-O-benzoyl-β-D-xylofuranoside (2) (Scheme 2).15 Compound 2 on further reaction with a catalytic amount of Hg(OCOCF3)2 in acetone-H2O (2:1) at room temperature gave the corresponding 2-(oxopropoxy)-2,3,5-tri-O-benzoyl-β-D-xylofuranoside (3). Baeyer-Villiger rearrangement of 3 with m-CPBA in CHCl3 at room temperature gave 1-[(2,3,5-tri-O-benzoyl-β-D-(xylofuranosyloxy)methyl]acetate (4) in 45% overall yield from compound 1.Compound 4 was characterized by 1H NMR spectrum from the appearance of methylene protons at δ 5.40-5.48 (m, 2H), anomeric proton (H-1) at δ 5.50 as a singlet indicating the β-anomeric configuration and by 13C NMR from the appearance of methylene carbon at δ 85.1 and anomeric carbon (C-1) at δ 104.9.

 

Scheme 2: Preparation of xylofuranosyl donor 4 Click here to View Scheme

 

To envisage the concept, we first investigated the coupling reactions of xylofuranosyl donor 4 with pyrimidine bases 5-8.¹⁶ Thus, coupling reaction of 4 with 5 in CHCl₃ and using activator SnCl₄ at room temperature gave 1-[(2,3,5-tri-O-benzoyl-β-D-xylofuranosyloxy)methyl]uracil (9) and 1-(2’,3’,5’-tri-O-benzoyl-β-D-xylofuranosyl)uracil (10) in a ratio of 1.0:1.9 (78% yield) (Scheme 3). A similar coupling reaction of 4 with bis(trimethylsilyl)thymine (6) by use of SnCl₄ gave 1-[(2,3,5-tri-O-benzoyl-β-D-xylofuranosyloxy)methyl]thymine (12) and 1-(2’,3’,5’-tri-O-benzoyl-β-D-xylofuranosyl)thymine (13) in a ratio of 1.0:2.1 (66% yield) (Scheme 3). Coupling reaction of 4 with bis(trimethylsilyl)cytosine (7) by use of SnCl₄ gave 1-[(2,3,5-tri-O-benzoyl-β-D-xylofuranosyloxy)methyl]cytosine (15) and 1-(2’,3’,5’-tri-O-benzoyl-β-D-xylofuranosyl)cytosine (16) in a ratio 1.0:2.4 (62% yield) respectively (Scheme 3).

 

Scheme 3: Coupling of xylose donor 4 with pyrimidine bases 5-8 Click here to View Scheme

 

The xylofuranosyloxy methyl pyrimidine nucleosides 9, 12, 15 were separated from pyrimidine nucleo sides 10, 13, 16 by column chromatography and characterized by ¹H and ¹³C NMR. The methylene protons, characteristic of xylofuranosyloxymethyl nucleosides 9, 12, 15 appeared at δ 5.20-5.40 (2H, AB type quartet, J_gem = 9.6-11.0 Hz). Compounds 10, 13, 16 were characterized as natural nucleoside by comparison of ¹H NMR, melting point and specific rotation with that reported in the literature.^16-18 Compound 9, 12, 15 on debenzoylation gave 1-[(β-D-xylofuranosyloxy)methyl]uracil (11), 1-[(β-D-xylofuranosyloxy)methyl]thymine (14), 1-[(β-D-xylofuranosyloxy)methyl]cytosine (17) respectively in good yields. Regiochemistry (N¹ or N³ linked nucleosides) of the newly formed xylofuranosyloxymethyl nucleosides 9, 12, 15 were assigned as N-1 linked nucleosides based on the (2 nm or less) absorption difference observed in UV spectra of the compounds 11, 14, 17 in neutral vs. 0.1 N alkali.¹⁸

Once we accomplished the synthesis of xylofuranosyloxymethyl pyrimidines, we studied the coupling reactions of 4 with purine bases. Thus, coupling reaction of 4 with bis(trimethylsilyl)-N⁶-benzoyladenine¹⁹ (20) using SnCl₄ in MeCN at room temperature resulted in the isolation of N⁶-benzoyl-9-[(2,3,5-tri-O-benzoyl-β-D-xylofuranosyloxy)methyl]adenine (21) and 9-(2’,3’,5’-tetra-O-benzoyl-β-D-xylofuranosyl)adenine (22) in a ratio of 1.0:2.27 (59%) (Scheme 4). Compound 21 was characterized as xylofuranosyloxymethyl adenine by ¹H NMR and nucleoside 22 was characterized as N⁹-isomer by comparison of its ¹H NMR, melting point and specific rotation with that of reported in the literature.^18b Compound 21 on debenzoylation gave 9-[(-β-D-xylofuranosyloxy)methyl]adenine (23) and the regiochemistry was assigned based on ¹H NMR spectrum from the appearance of H-2 at δ 8.30 and H-8 at δ 8.28 (Scheme 4). A chemical shift difference of ∆δ 0.02 ppm between H-2 and H-8 observed in the ¹H NMR spectrum is characteristic of N⁹ regioisomer.²⁰

 

Scheme 4: Coupling of 4 with adenine derivative 20 Click here to View Scheme

 

Coupling of 4 with guanine derivative 24 using SnCl₄ in MeCN at room temperature resulted in the isolation of oxymethylguanine nucleosides, N²-acetyl-9-[(2,3,5-tri-O-benzoyl-β-D-xylofuran- -osyloxy)methyl]guanine (25), N²-acetyl-7-[(2,3,5-tri-O-benzoyl-β-D-xylofuranosyloxy)methyl] -guanine (26) and natural nucleosides, N²-acetyl-9-(2’,3’,5’-tri-O-benzoyl-β-D-xylofuranosyl) guanine (27) and N²-acetyl-7-(2’,3’,5’-tri-O-benzoyl-β-D-xylofuranosyl)guanine (28). The ratio of xylofuranosyloxymethyl guanine derivatives 25, 26 to natural nucleosides 27, 28 was found to be 1.0:2.2 in 57% yield (Scheme 5). Compound 25, 26 were characterized as xylofuranosyloxymethyl guanine by ¹H NMR spectra and compounds 27, 28 were characterized as natural nucleosides by comparison of their ¹H NMR spectra with that reported in the literature. Compounds 25, 26 on debenzoylation gave 9-[(-β-D-xylofuranosyloxy)methyl]guanine (29) and 7-[(-β-D-xylofuranosyloxy)methyl]guanine (30) respectively. The regiochemistry of compounds 25, 26 and 29, 30 were assigned by ¹H NMR. In case of the compound 29 derived from compound 25, the appearance of H-8 at δ 7.78 as a singlet (¹H NMR) and C-5 at δ 116.4 (¹³C NMR) was characteristic of N⁹-regioisomer.²¹ In case of compound 30 derived from compound 26, the appearance of H-8 at δ 8.30 as a singlet (¹H NMR) and C-5 at δ 106.5 (¹³C NMR) was characteristic of N⁷-regioisomer.²¹

 

Scheme 5: Coupling of 4 with guanine derivative 24 Click here to View Scheme

 

In order to probe the reactivity of xylofuranosyl donor 4, we carefully studied the coupling reactions using organic Lewis acid, TMSOTf. Thus, coupling of 4 with 5, 6 by use of TMSOTf afforded the xylofuranosyloxymethyl pyrimidine nucleosides 9 and 12 exclusively. Formation of the corresponding natural nucleosides 10 and 13 were not observed. Coupling of 4 with 7 under similar conditions using TMSOTf showed poor selectivity. It resulted into the formation of xylofuranosyloxymethyl cytosine derivative 15 and natural nucleoside 16 in a ratio of 1.0:1.8. The formation of mixture of cytosine nucleosides may be due to the basicity of cytosine. Therefore, we protected cytosine as N⁴-bz-cytosine 8 and used for coupling with 4, which resulted in the formation of N⁴-benzoyl-1-[(2,3,5-tri-O-benzoyl-β-D-xylofuranosyloxy)-methyl]cytosine (18) in a higher yield compared to natural nucleoside 1-(N⁴,2’,3’,5’-tetra-O-benzoyl-β-D-xylofuranosyl)cytosine (19) in a ratio of 2.9:1.0. Similarly, coupling of 4 with 20 using TMSOTf as an activator gave xylofuranosyloxymethyl adenine 21 and natural nucleoside 22 in a ratio of 1.0:1.27. Coupling of 4 with 24 using TMSOTf gave oxymethyl guanine nucleosides 25, 26 and natural nucleoside 27, formation of 28 was not observed (Scheme 5). The ratio of natural nucleosides to oxymethyl guanine nucleosides was 1.0:14.0. Thus, synthesis of xylofuranosyloxymethyl nucleosides was achieved by coupling of xylofuranosyl donor 4 with nucleic acid bases similar to ribofuranosyl donor.^[14] It was observed that the formation of glycosyloxymethyl nucleosides was in a higher ratio to natural nucleosides in case of ribofuranosyl donor when compared to xylofuranosyl donor during coupling reactions performed using SnCl₄. This may be due to subtle difference in the reactivity of xylose donor to the ribose donor. There is no significant difference observed between ribose and xylose donors when activated by TMSOTf.

In order to probe the reactivity of xylofuranosyl donor 4, we carefully studied the coupling reactions using organic Lewis acid, TMSOTf. Thus, coupling of 4 with 5, 6 by use of TMSOTf afforded the xylofuranosyloxymethyl pyrimidine nucleosides 9 and 12 exclusively. Formation of the corresponding natural nucleosides 10 and 13 were not observed. Coupling of 4 with 7 under similar conditions using TMSOTf showed poor selectivity. It resulted into the formation of xylofuranosyloxymethyl cytosine derivative 15 and natural nucleoside 16 in a ratio of 1.0:1.8. The formation of mixture of cytosine nucleosides may be due to the basicity of cytosine. Therefore, we protected cytosine as N⁴-bz-cytosine 8 and used for coupling with 4, which resulted in the formation of N⁴-benzoyl-1-[(2,3,5-tri-O-benzoyl-β-D-xylofuranosyloxy)-methyl]cytosine (18) in a higher yield compared to natural nucleoside 1-(N⁴,2’,3’,5’-tetra-O-benzoyl-β-D-xylofuranosyl)cytosine (19) in a ratio of 2.9:1.0. Similarly, coupling of 4 with 20 using TMSOTf as an activator gave xylofuranosyloxymethyl adenine 21 and natural nucleoside 22 in a ratio of 1.0:1.27. Coupling of 4 with 24 using TMSOTf gave oxymethyl guanine nucleosides 25, 26 and natural nucleoside 27, formation of 28 was not observed (Scheme 5). The ratio of natural nucleosides to oxymethyl guanine nucleosides was 1.0:14.0. Thus, synthesis of xylofuranosyloxymethyl nucleosides was achieved by coupling of xylofuranosyl donor 4 with nucleic acid bases similar to ribofuranosyl donor.^[14] It was observed that the formation of glycosyloxymethyl nucleosides was in a higher ratio to natural nucleosides in case of ribofuranosyl donor when compared to xylofuranosyl donor during coupling reactions performed using SnCl₄. This may be due to subtle difference in the reactivity of xylose donor to the ribose donor. There is no significant difference observed between ribose and xylose donors when activated by TMSOTf.

CONCLUSION

In summary, we have successfully demonstrated that iterative glycosylation of acetoxymethoxy xylofuranosyl donor resulted in the formation of novel xylofuranosyloxymethyl nucleosides. The coupling reactions using SnCl₄ as an activator resulted in the formation of a mixture of natural nucleosides and novel xylofuranosyloxymethyl nucleosides whereas use of TMSOTf resulted in the exclusive formation of novel xylofuranosyloxymethyl nucleosides (in case of uracil, thymine, guanine) and mixture of nucleosides (in case of cytosine and adenine). The application of this new method to other glycosyl donors exploring various Lewis acids and their biological functions will be reported in near future.

EXPERIMENTAL SECTION

General. ¹H NMR spectra were recorded using the following instruments: at 200 MHz on a Varian Gemini, at 300 MHz on a Bruker Avance; at 400 MHz on a Varian Unity; at 500 MHz on a Varian Inova, with tetramethyl silane as internal standard for solutions in CDCl₃. J values are given Hz. ¹³C NMR spectra were taken with a Varian Gemini (50 MHz), Bruker Avance (75 MHz) spectrometer with CDCl₃ as internal standard (C 77.0) for solutions in deuteriochloroform, DMSO-d6 (C 39.7) for solutions in deuteriodimethyl sulfoxide, dioxane (C 67.3) for solutions in D₂O. Optical rotations were measured with a JASCO DIP-370 instrument. Melting points were determined by using Fischer–John’s melting point apparatus and are uncorrected. IR spectra were taken with a Perkin–Elmer 1310 spectrometer. UV spectra were recorded on a Shimadzu UV 160A spectrometer. Organic solutions were dried over anhydrous Na₂SO₄.

2-Propynyl 2,3,5-tri-O-benzoyl-β-D-xylofuranoside (2)

To a solution of 1-O-acetyl-2,3,5-tri-O-benzoyl-α/β-D-xylofuranoside (1) (30.0 g, 59.5 mmol) in dry CHCl₃ (300 mL) was added propargyl alcohol (4.2 mL, 71.4 mmol) and BF₃.OEt₂ (9 mL, 71.4 mmol) at 0 ^oC. The reaction mixture was stirred at room temperature for 8 h. After completion of the reaction, anhydrous K₂CO₃ (9.0 g) was added, stirred for a further 30 min., filtered, and the residue was washed with CHCl₃ (75 mL). The filtrate was transferred to a separating funnel, washed with water (2×150 mL) and brine (150 mL). The organic phase was separated, dried (Na₂SO₄) and concentrated to obtain a residue. The residue was chromatographed [SiO₂; 60-120 mesh; hexane-ethyl acetate (9:1)] to isolate the title compound 2 as syrup (19.2 g, 66%). [α]_D +10.0^o (c 1.0, CHCl₃); ν_max (KBr)/cm^-1 1723; ¹H NMR (200 MHz, CDCl₃) δ 2.42 (1H, t, CºCH), 4.40 (2H, m, CH₂-CºC), 4.50 (1H, dd, J_5,5’ =13.0, J_4,5 =6.0, H-5), 4.65 (1H, dd, J_4,5’ = 4.7, H-5’), 4.96 (1H, 2d, J_3,4 = 4.8, H-4), 5.42 (1H, s, H-1), 5.60 (1H, s, H-2), 5.82 (1H d, H-3), 7.20-8.20 (15H, m, Ar-H); ¹³C NMR (50 MHz, CDCl₃) δ 54.6 (HCºC-), 63.4 (C-5), 75.0 (C-4), 75.2 (C-3), 78.6 (O-CH₂-C), 79.2 (HCºC-), 81.1 (C-2), 104.1 (C-1), 125-135 (aromatic), 164.9, 165.2 and 166.1 (C=O, ester); Mass (FAB-MS): m/z 523 [M⁺+Na]; Anal. Calcd for C₂₉H₂₄O₈: C, 69.59; H, 4.83. Found: C, 69.38; H, 4.80.

2-(Oxopropoxy) 2,3,5-tri-O-benzoyl-β-D-xylofuranoside (3)

To a solution of 2 (19.0 g, 38.0 mmol) in acetone:H₂O (300 mL, 2:1) was added Hg(OCOCF₃)₂ (3.16 g, 7.60 mmol). The reaction mixture was stirred at room temperature for 6 h. After completion of the reaction, acetone was evaporated; the resulting residue was dissolved in ethyl acetate (300 mL), washed with 10% aq. KI solution (2×150 mL), 20% aq. hypo solution (2×150 mL) and brine (150 mL). The organic layer was separated, dried over Na₂SO₄ and concentrated to obtain the title compound 3 as syrup (18.2 g, 91%). [α]_D +18.0^o (c 1.0, CHCl₃); ν_max (KBr)/cm^-1 1715 and 1600; ¹H NMR (200 MHz, CDCl₃) δ 2.25 (3H, s, CH₃), 4.20, 4.40 (2H, AB type quartet, J_gem = 15.0, OCH₂C), 4.60 (1H, dd, J_5,5’ = 13.0, J_4,5 = 5.0, H-5), 4.70 (1H, dd, J_4,5’ = 6.0, H-5’), 5.00 (1H, 2d, J_3,4 = 5.0, H-4), 5.30 (1H, s, H-1), 5.65 (1H, s, H-2), 5.90 (1H, d, H-3), 7.20-8.20 (15H, m, Ar-H); ¹³C NMR (50 MHz, CDCl₃) δ 26.5 (CH₃), 63.2 (C-5), 72.5 (C-4), 75.0 (C-3), 79.3 (C-2), 80.9 (-OCH₂-), 105.7 (C-1), 125-133 (aromatic), 164.9, 165.2 and 165.9, 205.2 (C=O, ester); Mass (FAB-MS): m/z 541 [M⁺+Na]; Anal. Calcd for C₂₉H₂₆O₉: C, 67.17; H, 5.05. Found: C, 66.97; H, 5.16.

1-[(2,3,5-Tri-O-benzoyl-β-D-xylofuranosyloxy)methyl]acetate (4)

To a solution of 3 (17.5g, 33.7 mmol) in dry CHCl₃ (175 mL) was added m-CPBA (14.0 g, 81.1 mmol). The reaction mixture was stirred at room temperature for 14 h. After completion of the reaction, the reaction mixture was diluted with CHCl₃ (150 mL), washed with saturated aq. NaHCO₃ solution (150 mLx3, water (150 mL), brine (150 mL) and dried (Na₂SO₄). The solvent was concentrated to give a residue which was purified by silica gel chromatography [SiO₂; 60-120 mesh; hexane:ethyl acetate (9:1)] to obtain the title compound 4 as a solid (13.4 g, 75%). mp 115-117 ^oC; [α]_D +14.0^o (c 1.0, CHCl₃); ν_max (KBr)/cm^-1 1723; ¹H NMR (200 MHz, CDCl₃) δ 2.10 (3H, s, CH₃), 4.58 (1H, dd, J_5,5’=13.0, J_4,5 = 4.65, H-5), 4.68 (1H, dd, J_4,5’ = 6.0, H-5’), 4.98 (1H, 2d, J_3,4  = 5.0, H-4), 5.40 – 5.48 (2H, m, OCH₂O), 5.50 (1H, s, H-1), 5.55 (1H, s, H-2), 5.88 (1H, d, H-3), 7.20-8.20 (15H, m, Ar-H); ¹³C NMR (50 MHz, CDCl₃) δ 20.9 (CH₃) 63.2 (C-5), 74.9 (C-4), 79.5 (C-3), 81.0 (C-2), 85.1 (OCH₂O), 104.9 (C-1), 128-135 (aromatic), 164.8, 165.1, 166.0 and 170.2 (C=O, ester); Mass (FAB-MS): m/z 535 [M⁺+H]; Anal. Calcd for C₂₉H₂₆O₁₀: C, 65.16; H, 4.90. Found: C, 65.17; H, 5.14.

A typical procedure for coupling of xylofuranoside donor 4 with nucleic acid bases 5–8, 20, and 24. Method A: SnCl₄; Method B: Trimethylsilyltrifluoromethanesulfonate (TMSOTf)

To a solution (20 mL) of xylofuranoside donor 4 (1.0 mmol) and silylated nucleic acid bases 5–8, 20, 24 (1.2 mmol) at 0 ^oC was added the catalyst (1.2–2.0 mmol, 0.5N in CHCl₃ or MeCN or 1,2-dichloroethane) under nitrogen atmosphere. The reaction mixture was stirred at room temperature for 4–18 h or reflux temperature for 1–6 h, progress of the reaction was monitored by TLC. After completion of the reaction it was neutralized with saturated aqueous NaHCO₃ solution and filtered through a celite pad. The filtrate was extracted into CHCl₃ (30 ml), organic phase was separated, washed with brine, dried over anhydrous Na₂SO₄ and the solvent was removed under reduced pressure to obtain a residue. The residue was separated by column chromatography [SiO₂, 60–120 mesh, ethyl acetate:chloroform] to isolate the title compounds.

 1-[(2,3,5-Tri-O-benzoyl-β-D-xylofuranosyloxy)methyl]uracil (9)

Compound 9: Mp 83-85 ^oC; [α]_D +17.5^o (c 1.0, CHCl₃); λ_max/nm 259 (MeCN); ν_max (KBr)/cm^-1 1730; ¹H NMR (200 MHz, CDCl₃) δ 4.50 (1H, dd, J_5’,5”  =13.0, J_4’,5’ 5.0, H-5’), 4.64 (1H, dd, J_4’,5” = 6.0, H-5”), 4.92 (1H, m, H-4’), 5.26, 5.40 (2H, AB type quartet, J_gem = 11.0, OCH₂N), 5.44 (1H, s, H-1’), 5.52 (1H, s, H-2’), 5.64 (1H, d, J _3’,4’ = 5.1, H-3’), 5.80 (1H, d, J_5,6 = 5.0, H-5), 7.24-8.10 (16H, m, H-6, Ar-H); Mass (FAB-MS): m/z 609 [M⁺+Na]; Anal Calcd for C₃₁H₂₆O₁₀N₂: C, 63.48; H, 4.47; N, 4.78 Found: C, 63.26; H, 4.38; N, 4.59. Compound 10: Mp. 108-111 ^oC [lit.,¹³ Mp 112-118 ^oC]; [α]_D +73.3^o (c 1.0, CHCl₃) [lit.,¹³ [α]_D +75.0^o (c 0.8, CHCl₃)].

1-[(β-D-Xylofuranosyloxy)methyl]uracil (11)

To a solution of 9 (0.8 g, 1.36 mmol) in methanol (15 mL) was added a catalytic amount of NaHCO₃ (0.1 g) and refluxed for 3 h. After completion of the reaction the reaction mixture was neutralized with IR 120 H⁺ resin, filtered, washed with methanol (15 mL) and filtrate was evaporated to a residue. The residue was dissolved in water (30 mL) washed with chloroform (2×25 mL); the water phase was concentrated to isolate the title compound 11 as a thick syrup (0.32 g, 84%). [α]_D –55.0^o (c 1.0, H₂O); λ_max/nm 260 (H₂O), 261 (0.1N, NaOH); ν_max (KBr)/cm^-11685; ¹H NMR (400 MHz, D₂O) δ 3.60 (1H, dd, J_5’,5” = 10.0, J_4’,5’ = 5.5, H-5’), 3.75 (1H, dd, J_4’,5” = 4.7, H-5”), 4.10 (1H, s, H-2’), 4.18 (1H, m, H-4’), 4.30 (1H, m, H-3’), 5.12 (1H, s, H-1’), 5.20, 5.30 (2H, AB type quartet, J_gem = 10.0, OCH₂N), 5.80 (1H, d, J_5,6 = 5.6, H-5), 7.65 (1H, d, H-6); ¹³C NMR (50 MHz, D₂O) δ 66.0 (C-5’), 81.0 (C-4’), 81.2 (C-3’), 86.3 (C-2’), 89.2 (OCH₂N), 108.0 (C-1’), 113.0 (C-5), 152.0 (C-6), 158.0 (C=O,C-2), 172.0 (C=O,C-4); Mass (FAB-MS): m/z 275 [M⁺+H]; Anal. Calcd for C₁₀H₁₄O₇N₇: C, 43.80; H, 5.15; N, 10.22. Found: C, 43.56; H, 5.22; N, 10.28.

 1-[(2,3,5-Tri-O-benzoyl-β-D-xylofuranosyloxy)methyl]thymine (12)

Compound 12: Mp 137-139^oC; [α]_D +14.2^o (c 1.0, CHCl₃); λ_max/nm 263 (MeOH); ν_max (KBr)/cm^-1 1723; ¹H NMR (200 MHz, CDCl₃) δ 1.80 (3H, s, CH₃), 4.40-4.70 (2H, m, H-5’,5”), 4.85-5.10 (1H, m, H-4’), 5.22,5.34 (1H, AB type quartet, J_gem = 11.0, OCH₂N), 5.40 (1H, s, H-1’), 5.52 (1H, s, H-2’), 5.80 (1H, d, J_3’,4’ = 5.4, H-3’), 7.10 (1H, s, H-6), 720-8.10 (15H, m, Ar-H), 9.05 (1H, br s, NH); ¹³C NMR (50 MHz, CDCl₃) δ 12.2 (CH₃), 63.5 (C-5’), 74.5 (C-3’), 75.0 (C-2’), 79.5 (C-4’), 81.0 (OCH₂N), 105.0 (C-1’), 111.5 (C-5), 125-135 (aromatic), 139.1 (C-6), 150.5 (C=O,C-2) 164.0 (C=O,C-4) 164.5, 165.0, 166.0 (C=O, ester); Mass (FAB-MS): m/z 601 [M⁺+H]; Anal. Calcd for C₃₂H₂₈O₁₈N₂: C, 63.99; H, 4.70; N, 4.66. Found: C, 63.28; H, 4.62; N, 4.73.

Compound 13: Mp 196-199 ^oC [lit.¹⁴ mp. 196-198 ^oC]; [α]_D +49.3^o (c 0.85, CHCl₃) [lit.¹⁴ [α]_D +50.6^o (c 0.85, CHCl₃)].

 1-[(β-D-Xylofuranosyloxy)methyl]thymine (14)

The benzoylated derivative 12 (0.8g, 1.33 mmol) in methanol (16 mL) was deprotected as described for 11 to isolate the title compound 14 as a white solid (0.35 g, 91.0%). mp 175-177 ^oC; [α]_D –31.0^o (c 1.0, H₂O); λ_max/nm 265 (H₂O), 266 (0.1N NaOH); ν_max (KBr)/cm^-1 1690; ¹H NMR (200 MHz; D₂O) δ 1.88 (3H, s, CH₃), 3.70 (1H, dd, J_5’,5” = 14.0, J_4’,5’ =5.2, H-5’), 3.85 (1H, dd, J_4’,5” = 4.0, H-5”), 4.15 (1H, d, J_2’,3’ = 4.65, H-3’), 4.26 (1H, d, H-2’), 4.30-4.45 (1H, m, H-4’), 5.18 (1H, s, H-1’), 5.20, 5.35 (2H, AB type quartet, J_gem = 14.0, OCH₂N), 7.45 (1H, s, H-6), 8.00 (1H, br s, NH); ¹³C NMR (50 MHz, D₂O) δ 12.5 (CH₃), 60.3 (C-5’), 73.9 (C-4’), 75.5 (C-3’), 80.5 (C-2’), 83.2 (OCH₂N), 106.5 (C-1’), 109.0 (C-5), 140.0 (C-6), 154.0 (C=O,C-2), 168.0 (C=O,C-4); Mass (FAB-MS): m/z 311 [M⁺+Na]; Anal. Calcd for C₁₁H₁₆O₇N₂: C, 45.83, H, 5.60, N, 9.2. Found: C, 45.67; H, 5.48; N, 8.91.

1-[(2,3,5-Tri-O-benzoyl-β-D-xylofuranosyloxy)methyl]cytosine (15)

Compound 15: Mp 122-126^oC; [α]_D +44.6^o (c 1.0, CHCl₃); λ_max/nm 274 (MeOH); ν_max (KBr)/cm^-1 1723, 1661 and 1615; ¹H NMR (200 MHz, CDCl₃) δ 4.48 (1H, dd, J_5’,5” = 13.0, J_4’,5’ = 5.0, H-5’), 4.62 (1H, dd, J_4’,5” =6.0, H-5”), 4.90 (1H, dd, J_3’,4’ = J_4’,5’ 5.20, H-4’), 5.25, 5.35 (2H, AB type quartet, J_gem = 9.6, OCH₂N), 5.48 (1H, s, H-1’), 5.50 (1H, s, H-2’), 5.76 (1H, d, H-3’), 5.82 (1H, d, J_5,6 = 7.6, H-5), 7.20-7.60 (10H, m, H-6, Ar-H), 7.85-8.20 (6H, m, Ar-H); ¹³C NMR (50 MHz, CDCl₃) δ 63.3 (C-5’), 75.1 (C-4’), 76.3 (C-3’), 79.4 (C-2’), 81.0 (OCH₂N), 95.6 (C-5), 104.8 (C-1’), 128.0-135.0 (aromatic), 144.7 (C-6), 155.9 (C-4), 165.0 (C=2, C=O), (C=O,C-2) 165.6, 166.0, 166.2 (C=O, ester); Mass (FAB-MS): m/z 586 [M⁺+H]; Anal. Cacld for C₃₁H₂₇O₉N₃: C, 63.58; H, 4.65; N, 7.18. Found: C, 63.32; H, 4.63; N, 7.11.

Compound 16: Mp 137-139 ^oC [lit.,¹⁴ mp 140-141 ^oC]; [α]_D +61.5^o (c 0.6, CHCl₃) [lit.,¹⁴ [α]_D +63.4^o (c 0.6, CHCl₃)].

 1-[(β-D-Xylofuranosyloxy)methyl]cytosine (17)

The benzoylated derivative 15 (0.6 g, 0.9 mmol) was debenzoylated as described for 14 to isolate the title compound 17 as a syrup (0.19 g, 81.5%). [α]_D +17.8^o (c 1, H₂O); λ_max/nm 268 (H₂O), 270 nm (0.1N NaOH); ν_max (KBr)/cm^-1 1653; ¹H NMR (200 MHz, D₂O) δ 3.70 (1H, dd, J_5’,5” = 12.8, J_4’,5’ = 5.0, H-5’), 3.84 (1H, dd, J_4’,5” = 6.0, H-5”), 4.20 (1H, s, H-2’), 4.22-4.40 (2H, m, H-3’, H-4’), 5.20 (1H, s, H-1’), 5.24, 5.40 (2H, AB type quartet, J_gem = 10.0, OCH₂N), 6.20 (1H,d, J_5,6 = 7.8, H-5), 7.70 (1H, d, H-6); ¹³C NMR (50 MHz, D₂O) δ 59.3 (C-5’), 73.5 (C-4’), 74.9 (C-3’), 78.7 (C-2’), 81.6 (OCH₂N), 104.9 (C-1’), 94.6 (C-5), 144.8 (C-6), 156.0 (C=O, C-2), 165.2 (C=O,C-4); Mass (FAB-MS) m/z 274 [M⁺+H]; Anal. Calcd for C₁₀H₁₅O₆N₂: C, 43.95; H, 5.53; N, 15.38. Found: C, 44.11; H, 5.38; N, 15.21.

N⁶-Benzoyl-9-[(2,3,5-tri-O-benzoyl-β-D-xylofuranosyloxy)methyl]adenine (21)

Compound 21: Mp 77-79 ^oC; [α]_D +32.6^o (c 1, CHCl₃); λ_max/nm 277 (MeCN); ν_max (KBr)/cm^-1 1723 and 1600. ¹H NMR (200 MHz, CDCl₃) δ 4.50 (2H, m, H-5’,5”), 4.98 (1H, m, H-4’), 5.50 (1H, s, H-1’), 5.56 (1H, s, H-2’), 5.80 (1H, d, J_3’,4’ = 3.5, H-3’), 5.84, 6.02 (2H, AB type quartet, J_gem = 9.8, OCH₂N), 7.20-7.60 (12H, m, Ar-H), 7.80-8.16 (8H, m, Ar-H), 8.22 (1H, s, H-8), 8.78 (1H, s, H-2), 9.24 (1H, br s, NH); Mass (FAB-MS): m/z 714 [M⁺+H]; Anal. Cacld for C₃₉H₃₁O₉N₅: C, 65.68; H, 4.37; N, 9.82. Found: C, 63.23; H, 4.48; N, 9.38. Compound 22: Mp 104-107 ^oC [lit.,^14,17 mp 105-110 ^oC]; [α]_D +10.73^o (c 1.0, CHCl₃) [lit.,^14,17 [α]_D +5.2^o (c 0.8, CHCl₃)];

9-[(β-D-Xylofuranosyloxy)methyl]adenine (23)

The benzoylated derivative 21 (0.6 g, 0.84 mmol) was debenzoylated as described for 11 to isolate the title compound 23 as a solid (0.19 g, 76.0%). mp 220 ^oC (decom.); [α]_D –38.5^o (c 1, DMSO); λ_max/nm 259 (H₂O); ν_max (KBr)/cm^-1 1678 and 1610; ¹H NMR (300 MHz, DMSO-d₆) δ 3.20-4.30 (6H, m, H-2’,H-3’,H-4’,H-5’,H-5”,OH), 4.82 (1H, br s, OH), 4.98 (1H, s, H-1’), 5.15 (1H, br s, OH), 5.64, 5.85 (2H, AB type quartet, J_gem = 10.0, OCH₂N)), 7.30 (2H, br s, NH₂), 8.28 (1H, s, H-8), 8.30 (1H,s, H-2); ¹³C NMR (50 MHz, DMSO-d₆) δ 63.1 (C-5’), 68.8 (C-4’), 71.8 (C-3’), 74.4 (C-2’), 86.1 (OCH₂N), 105.5 (C-1’), 119.3 (C-5), 143.1 (C-8), 149.8 (C-4), 153.6 (C-2), 156.0 (C-6); Mass (FAB-MS): m/z 298 [M⁺+H]; Anal. Calcd for C₁₁H₁₅O₅N₅: C, 44.44; H, 5.09; N, 23.56. Found: C, 43.91; H, 5.14; N, 23.42.

 N²-Acetyl-9-[(2,3,5-tri-O-benzoyl-β-D-xylofuranosyloxy)methyl]guanine (25) and N²-acetyl-7-[(2,3,5-tri-O-benzoyl-β-D-xylofuranosyloxy)methyl]-guanine (26)

Compound 25: Mp 149-151 ^oC; [α]_D –4.89^o (c 1.0, CHCl₃); λ_max/nm 276 (MeCN); ν_max (KBr)/cm^-1 1715, 1615 and 1570; ¹H NMR (200 MHz, CDCl₃) δ 2.20 (3H, s, OCOCH₃), 4.60 (1H, dd, J_5’,5” =12.8, J_4’,5’ = 6.0, H-5’), 4.90-5.20 (2H, m, H-4’, H-5), 5.30, 5.78 (2H, AB type quartet, J_gem = 10.0, OCH₂N), 5.50 (1H, s, H-1’), 5.54 (1H, s, H-2’), 5.84 (1H, d, J_3’,4’ = 6.5, H-3’), 7.08-7.70 (10H, H-8, Ar-H), 8.00 (6H, d, Ar-H), 10.49 (1H, br s, NH), 11.90 (1H, br s, NH); Mass (FAB-MS): m/z 668 [M⁺+H]; Anal. Calcd for C₃₄H₂₉O₁₀N₅: C, 61.16; H, 4.38; N, 10.49. Found: C, 61.08; H, 4.12; N, 9.98.

Compound 26: Mp 115-120 ^oC; [α]_D +2.7^o (c 1.0, CHCl₃); λ_max/nm 261 (MeCN); ν_max (KBr)/cm^-1 1715, 1615 and 1570; ¹H NMR (200 MHz, CDCl₃) δ 2.40 (3H, s, OCOCH₃), 4.38-4.52 (2H, m, H-4’, H-5”), 4.94 (1H, m, H-4’), 5.58 (1H, s, H-1’), 5.62 (1H, s, H-2’), 5.80 (d, J_3’,4’ 6.3, H-3’), 5.98, 6.05 (2H, AB type quartet, J_gem = 10.0, OCH₂N), 7.30-7.64 (9H, m, Ar-H), 7.92-8.10 (7H, m, H-8, Ar-H), 11.10 (1H, br s, NH), 12.30 (1H, br s, NH); Mass (FAB-MS): m/z 668 [M⁺+H]; Anal. Cacld for C₃₄H₂₉O₁₀N₅: C, 66.16; H, 4.38; N, 10.49. Found: C, 66.21; H, 4.23; N, 10.23.

9-[(β-D-Xylofuranosyloxy)methyl]guanine (29)

The benzoyl derivative 25 (0.4g, 0.59 mmol) in methanol (10 mL) was deprotected as described for 11 to isolate the title compound 29 as a solid (0.14 g, 78%). mp 201-203 ^oC; [α]_D –43.4^o (c 1.0, DMSO); λ_max/nm 251 (H₂O); ν_max (KBr)/cm^-1 3400, 1723, 1685 and 1623; ¹H NMR (200 MHz, DMSO-d₆) δ 3.40-3.70 (2H, m, H-5’, H-5”), 3.80 (1H, m, H-2’), 3.90 (1H, m, H-3’), 4.10 (1H, dd, J_3’,4’ =3.5, J_4’,5’ = 6.0, H-4’), 4.50 (1H, s, OH), 4.80 (2H, br s, 2xOH), 5.30 (1H, s, H-1’), 5.36, 5.46 (2H, AB type quartet, J_gem = 9.8, OCH₂N), 6.58 (2H, br s, NH₂), 7.78 (1H, s, H-8), 10.68 (1H, br s, NH); ¹³C NMR (50 MHz, DMSO-d₆) δ 60.5 (C-5’), 68.3 (C-4’), 75.5 (C-3’), 80.9 (C-2’), 83.3 (OCH₂N), 105.9 (C-1’), 116.4 (C-5), 137.7 (C-8), 151.3 (C-4), 153.8 (C-2), 156.7 (C-6); Mass (FAB-MS): m/z 314 [M⁺+H]; Anal. Calcd for C₁₁H₁₅O₆N₅: C, 42.17; H, 4.83; N, 22.36. Found: C, 42.31; H, 4.67; N, 23.43.

7-[(β-D-Xylofuranosyloxy)methyl]guanine (30)

The benzoylated derivative 26 (0.2 g, 0.29 mmol) in methanol (6 mL) was deprotected as described for 11 to isolate the title compound 30 as a white solid (0.07 g, 75.0%). Mp 235 ^oC (decom.); [α]_D –47.3 (c 0.5, DMSO); λ_max/nm 281 (H₂O); ν_max (KBr)/cm^-1 1677 and 1485; ¹H; NMR (200 MHz, DMSO-d₆) δ 3.70-4.20 (5H, m, H-2’, 3’, 4’, 5’, 5”), 4.60 (1H, s, OH), 4.80 (1H, s, OH), 4.95 (1H, s, H-1’), 5.00 (1H, s, OH), 5.62, 5.70 (2H, AB type quartet, J_gem = 8.8, OCH₂N), 6.70 (2H, br s, NH₂), 8.30 (1H, s, H-8), 10.90 (1H, br s, NH); ¹³C NMR (75 MHz, DMSO-d₆) δ 60.5 (C-5’), 72.6 (C-4’), 75.4 (C-3’), 80.9 (C-2’), 83.6 (OCH₂N), 106.2 (C-1’), 106.7 (C-5), 143.2 (C-8), 154.1 (C-2), 155.2 (C-6), 160.8 (C-4); Mass (FAB-MS): m/z 314 [M⁺+H]; Anal. Calcd for C₁₁H₁₅O₆N₅: C, 42.17; H, 4.83; N, 22.36. Found: C, 42.25; H, 4.41; N, 22.62.

ACKNOWLEDGEMENTS

The authors are grateful to the Indian Institute of Chemical Technology, Hyderabad and thankful to Council of Scientific and Industrial Research, New Delhi for the financial support.

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