Synthesis of 2′‐ and 3′‐C‐Methylribonucleosides

Leonid Beigelman1, Sergey N. Mikhailov2

1 InterMune, Brisbane, California, 2 Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Moscow
Publication Name:  Current Protocols in Nucleic Acid Chemistry
Unit Number:  Unit 14.5
DOI:  10.1002/0471142700.nc1405s28
Online Posting Date:  March, 2007
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Abstract

A simple, efficient method for the synthesis of 2′‐ and 3′‐C‐methylribonucleosides starting from a common precursor is described. This synthesis achieves conversion of 1,2:5,6‐di‐O‐isopropylidene‐3‐C‐methyl‐α‐D‐allofuranose into 1,2,3‐tri‐O‐acetyl‐5‐O‐benzoyl‐3‐C‐methyl‐α,β‐D‐ribofuranose followed by condensation with nucleic acid bases, with a final ammonolysis leading to 3′‐C‐methylribonucleosides. Alternatively, the same branched allofuranose converted to l,2,3‐tri‐O‐acetyl‐5‐Op‐methylbenzoyl‐2‐C‐methyl‐β‐D‐ribofuranose, after analogous Vorbruggen condensation and ammonolysis, provides 2′‐C‐methylribonucleosides.

Keywords: branched‐chain nucleosides; anti‐HCV activity; synthesis; methylnucleosides; methylribonucleosides

     
 
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Table of Contents

  • Basic Protocol 1: Preparation of the Protected 3‐C‐Methyl‐D‐Ribofuranose
  • Basic Protocol 2: Preparation of the Protected 2‐C‐Methyl‐D‐Ribofuranose
  • Basic Protocol 3: Preparation of 1‐(3‐C‐Methyl‐β‐D‐Ribofuranosyl)Uracil
  • Alternate Protocol 1: Preparation of 3‐C‐Methyl‐β‐D‐Ribofuranosyl Derivatives of Cytosine, Adenine, and Guanine
  • Basic Protocol 4: Preparation of 1‐(2‐C‐Methyl‐β‐D‐Ribofuranosyl)Uracil
  • Alternate Protocol 2: Preparation of 2‐C‐Methyl‐β‐D‐Ribofuranosyl Derivatives of Cytosine and Adenine
  • Commentary
  • Literature Cited
  • Figures
     
 
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Materials

Basic Protocol 1: Preparation of the Protected 3‐C‐Methyl‐D‐Ribofuranose

  Materials
  • 1,2:5,6‐Di‐O‐isopropylidene‐3‐C‐methyl‐α‐D‐allofuranose (S.1; Bio et al., )
  • Acetic acid (AcOH)
  • 1‐Butanol, reagent grade
  • Ethanol (EtOH), reagent grade
  • Phosphorus pentoxide (P 2O 5)
  • Sodium periodate (NaIO 4)
  • Sodium borohydride (NaBH 4)
  • Chloroform (CHCl 3), reagent grade
  • Methanol (MeOH), analytical grade
  • Hexane, reagent grade
  • Anhydrous pyridine
  • Benzoyl chloride
  • Saturated sodium bicarbonate (NaHCO 3)
  • Anhydrous sodium sulfate (Na 2SO 4)
  • Toluene, reagent grade
  • 90% trifluoroacetic acid (F 3CCOOH)
  • Acetic anhydride, reagent grade
  • 4‐Dimethylaminopyridine (DMAP)
  • Silica gel: Kieselgel 60 (0.06 to 0.20 mm; Merck)
  • 250‐ and 500‐mL round‐bottom flasks
  • Rotary evaporator equipped with a water aspirator and an oil pump
  • Reflux condenser
  • Vacuum filtration system with glass filters (porosity 3)
  • Vacuum desiccator
  • 250‐mL and 1‐L separatory funnels
  • Dropping funnel connected to a CaCl 2 protection tube
  • Silica‐coated aluminum TLC plates with fluorescent indicator (Merck silicagel 60 F 254)
  • 254‐nm UV lamp
  • 3 × 20–cm sintered glass chromatography column, porosity 3
  • Vacuum oil pump
  • Additional reagents and equipment for thin‐layer chromatography (TLC; appendix 3D) and column chromatography ( appendix 3E)

Basic Protocol 2: Preparation of the Protected 2‐C‐Methyl‐D‐Ribofuranose

  Materials
  • 1,2:5,6‐Di‐O‐isopropylidene‐3‐C‐methyl‐α‐D‐allofuranose (S.1; Bio et al., )
  • Anhydrous dimethyl sulfoxide (DMSO)
  • 80% (w/w) sodium hydride (NaH) in mineral oil
  • Benzyl chloride, reagent grade
  • Chloroform (CHCl 3), reagent grade
  • Anhydrous sodium sulfate (Na 2SO 4)
  • Silica gel: Kieselgel 60 (0.06 to 0.20 mm; Merck)
  • 75% (v/v) acetic acid (AcOH)
  • 1‐Butanol, reagent grade
  • Ethanol (EtOH), reagent grade
  • Phosphorus pentoxide (P 2O 5)
  • Dibutylstannoxane
  • Dry methanol (MeOH)
  • Triethylamine, reagent grade
  • p‐Methylbenzoyl chloride (TolCl)
  • Saturated sodium bicarbonate (NaHCO 3)
  • 90% (v/v) trifluoroacetic acid (F 3CCOOH)
  • Toluene, reagent grade
  • 1,4‐Dioxane
  • 1 M sodium periodate (NaIO 4)
  • Ethyl acetate (EtOAc), reagent grade
  • 1 M sodium methoxide (NaOMe) in methanol
  • Dowex 50 (H+) resin
  • Anhydrous pyridine
  • Acetic anhydride, reagent grade
  • Cyclohexene
  • 20% palladium hydroxide on carbon (Pd(OH) 2/C)
  • Hyflo Super Cel
  • 4‐Dimethylaminopyridine (DMAP)
  • 250‐mL round‐bottom flasks
  • Reflux condensers equipped with a CaCl 2 protection tube
  • Oil bath with temperature control
  • 250‐ and 500‐mL separatory funnels
  • Rotary evaporator equipped with a water aspirator and an oil pump
  • 3 × 20– and 5 × 30–cm sintered glass chromatography columns, porosity 3
  • Silica‐coated aluminum TLC plates with fluorescent indicator (Merck silicagel 60 F 254)
  • 254‐nm UV lamp
  • Vacuum oil pump
  • Vacuum filtration system with glass filters (porosity 3)
  • Vacuum desiccator
  • 100‐mL funnel with sintered disc (porosity 3)
  • Additional reagents and equipment for thin‐layer chromatography (TLC; appendix 3D) and column chromatography ( appendix 3E)

Basic Protocol 3: Preparation of 1‐(3‐C‐Methyl‐β‐D‐Ribofuranosyl)Uracil

  Materials
  • Uracil
  • Anhydrous pyridine
  • 1,1,1,3,3,3‐Hexamethyldisilazane (HMDS), reagent grade
  • Anhydrous toluene, reagent grade
  • Anhydrous 1,2‐dichloroethane
  • 1,2,3‐Tri‐O‐acetyl‐5‐O‐benzoyl‐3‐C‐methyl‐α,β‐D‐ribofuranose (S.6; see protocol 1)
  • 2 M trimethylsilyl trifluoromethanesulfonate (TMSOTf)
  • Chloroform (CHCl 3), reagent grade
  • Ethanol (EtOH), reagent grade
  • Saturated sodium bicarbonate (NaHCO 3)
  • Hyflo Super Cel
  • Anhydrous sodium sulfate (Na 2SO 4)
  • Silica gel: Kieselgel 60 (0.06 to 0.20 mm; Merck)
  • 5 M ammonia in methanol (half saturated at 0°C)
  • Methylene chloride
  • Acetone
  • Diethyl ether, reagent grade
  • Phosphorus pentoxide (P 2O 5)
  • 100‐ and 250‐mL round‐bottom flasks
  • Reflux condenser equipped with a CaCl 2 protection tube
  • Oil bath with temperature control
  • Rotary evaporator equipped with a water aspirator
  • Silica‐coated aluminum TLC plates with fluorescent indicator (Merck silica gel 60 F 254)
  • 254‐nm UV lamp
  • 100‐mL funnel with a sintered disc (porosity 3)
  • Vacuum pump for filtration
  • 100‐ and 250‐mL separatory funnels
  • 3 × 20–cm sintered glass chromatography column, porosity 3
  • Vacuum oil pump
  • Glass filters (porosity 3)
  • Vacuum desiccator
  • Additional reagents and equipment for thin‐layer chromatography (TLC; appendix 3D) and column chromatography ( appendix 3E)

Alternate Protocol 1: Preparation of 3‐C‐Methyl‐β‐D‐Ribofuranosyl Derivatives of Cytosine, Adenine, and Guanine

  Materials
  • Uracil
  • Anhydrous pyridine
  • 1,1,1,3,3,3‐Hexamethyldisilazane (HMDS), reagent grade
  • Anhydrous toluene, reagent grade
  • Anhydrous 1,2‐dichloroethane
  • 1,2,3‐Tri‐O‐acetyl‐5‐Op‐methylbenzoyl‐2‐C‐methyl‐β‐D‐ribofuranose (S.13; see protocol 2)
  • 2 M Trimethylsilyl trifluoromethanesulfonate (TMSOTf)
  • Chloroform (CHCl 3), reagent grade
  • Ethanol (EtOH), reagent grade
  • Saturated sodium bicarbonate (NaHCO 3)
  • Hyflo Super Cel
  • Anhydrous sodium sulfate (Na 2SO 4)
  • Silica gel: Kieselgel 60 (0.06 to 0.20 mm; Merck)
  • 5 M ammonia in methanol (half saturated at 0°C)
  • Acetone
  • Diethyl ether, reagent grade
  • Phosphorus pentoxide (P 2O 5)
  • 100‐ and 250‐mL round‐bottom flasks
  • Reflux condenser equipped with a CaCl 2 protection tube
  • Oil bath with temperature control
  • Rotary evaporator equipped with a water aspirator
  • Silica‐coated aluminum TLC plates with fluorescent indicator (Merck silicagel 60 F 254)
  • 254‐nm UV lamp
  • 100‐mL funnel with a sintered disc (porosity 3)
  • Vacuum pump for filtration
  • 100‐ and 250‐mL separatory funnels
  • 3 × 20–cm sintered glass chromatography column, porosity 3
  • Vacuum oil pump
  • Glass filters (porosity 3)
  • Vacuum desiccator
  • Additional reagents and equipment for thin‐layer chromatography (TLC; appendix 3D) and column chromatography ( appendix 3E)
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Literature Cited

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