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. 260^oC, split mode.

Oven temperature programmed as follows: 170^oC for 2 min, then a 35^oC/min ramp to 278^oC and hold for 6min.

 

Retention Times for Bile Acids

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.