Table of Contents

Determination of the Absolute Configuration using - acetylated butyl glycosides

Introduction

see previous page

Procedure

  1. Transfer sample (ca 0.2 mg, 1 µmol) to a 13 x 100 mm screw cap tube.
  2. Hydrolyze in 0.5 mL 2M TFA at 120 °C for 30 min.
  3. Evaporate the solution to dryness with a stream of compressed air, add 0.5 mL methanol and evaporate. Repeat once.
  4. If the polysaccharide contains aminosugars, re-N-acetylate with 25 µL acetic anhydride for 4 h at room temperature. Evaporate with a stream of dry air.
  5. Dissolve in 0.2 mL (+)-2-butanol. Add 15 µL acetyl chloride, bubble nitrogen through the solution for 30 sec., then seal the tube.
  6. Butanolyze the sample for 8-16 h at 80 °C.
  7. Evaporate the solution to dryness with a stream of air, add 0.5 mL methanol, then evaporate. Repeat once.
  8. Acetylate with 0.2 mL acetic anhydride and 0.1 mL pyridine at 120 °C for 30 min.
  9. Evaporate the solution and add 0.5 mL toluene and evaporate to dryness. Repeat once.
  10. Partition between 0.5 mL H2O and 0.5 mL EtOAc. Transfer the upper (EtOAc) phase to a new tube. Repeat the extraction twice with 0.5 mL EtOAc. Concentrate the combined EtOAc phases to dryness, dissolve in 0.2 mL EtOAc, transfer to a new sample tube and concentrate to 25-50 µL.

References