Bile Acids
Bile acid derivative: Methyl acetate.
Reagents:
1. 0.5N HCl in methanol.
2. Pyridine
3. Acetic anhydride.
Procedure:
1. Incubate unlabeled bile acid standard and samples
with 0.5ml of methanolic HCI at 70¡ãC overnight.
2. Dry the sample under nitrogen.
3. Add
100ml of pyridine and 100ml of acetic anhydride and heat at 100¡ãC for 90 minutes. Dry under nitrogen.
4. Add hexane as solvent
for GC/MS analysis.
GC/MS
Instrument: HP6890 GC system connected to the HP5973 mass selective detector.
GC column: HPUltra1, length-12m,
250mM, programmed at a constant He flow of 1ml/min.
Inlet temp. 260oC, split mode.
Oven temperature programmed as follows: 170oC for
2 min, then a 35oC/min ramp to 278oC and hold for 6min.
Bile Acid Retention Times
Cholic Acid 7.70 min
Lithocholic Acid 6.38 min
Chenodoxycholic Acid 7.24 min
Deoxycholic Acid 6.97 min
Reference: Fukada, Y., Kimura, K., Ayaki, Y. Effect of Chitosan on Intestinal Bile Acid
Metabolism in Rats. Lipids. Vol.
26. pp 395-399.