See also previous procedure.
- Transfer sample (ca 0.2 mg, 1 µmol) to a 13 x 100 mm screw cap tube.
- Hydrolyze in 0.5 mL 2M TFA at 120 °C for 30 min.
- Evaporate the solution to dryness with a stream of compressed air, add 0.5 mL methanol and evaporate.
- 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.
- Dissolve in 0.2 mL (+)-2-butanol. Add 15 µL acetyl chloride, bubble nitrogen through the
solution for 30 sec., then seal the tube.
- Butanolyze the sample for 8-16 h at 80 °C.
- Evaporate the solution to dryness with a stream of air, add 0.5 mL methanol, then evaporate.
- Acetylate with 0.2 mL acetic anhydride and 0.1 mL pyridine at 120 °C for 30 min.
- Evaporate the solution and add 0.5 mL toluene and evaporate to dryness. Repeat once.
- 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.