Journal of Lipid Research, Vol 23, 204-210, Copyright © 1982 by Lipid Research, Inc.
A method for the accurate measurement of isotope ratios of chenodeoxycholic and cholic acids in serum
BR DeMark, GT Everson, PD Klein, RB Showalter and F Kern Jr
A method for the extraction of bile acids from serum is described that
enables the stable isotopic content of chenodeoxycholic acid and cholic
acid to be determined accurately to levels as low as the natural 13C
abundance. The method uses Sep-Pak C18 reverse phase cartridges both for
extraction and purification procedures. Free bile acids, bile acid
conjugates, and 3-monosulfated bile acid conjugates are recovered in high
yield from the Sep-Pak in methanol-water 75:25 after first removing
impurities with hexane and methanol-water 40:60 washes. Other important
features of the method include the use of enzymatic rather than alkaline
hydrolysis of bile acid conjugates, the use of ammonia as the reagent gas
for chemical ionization mass spectrometric measurement of isotopic ratios,
and the exclusion of all extraneous components in the final sample from the
ion source. This method should be applicable to kinetic studies of bile
acids using bile acids labeled with stable isotopes and serum measurements,
and provides an alternative sampling point in the enterohepatic circulation
to conventional duodenal bile samples requiring intubation.
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