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Journal of Lipid Research, Vol 28, 589-595, Copyright © 1987 by Lipid Research, Inc.
SS Rossi, JL Converse and AF Hofmann
A reversed phase high pressure liquid chromatography (HPLC) system capable
of simultaneously separating four lithocholyl species (sulfated and
unsulfated forms of lithocholylglycine and lithocholyltaurine) as well as
the eight other major conjugated bile acids present in human bile is
described. The system uses a C18 octadecylsilane column and isocratic
elution with methanol phosphate buffer, pH 5.35. Relative bile acid
concentration is determined by absorbance at 200 nm. Retention times
relative to chenodeoxycholylglycine are reported for the four lithocholic
acid forms, the glycine and taurine amidate of the four major bile acids
present in human bile (cholic, chenodeoxycholic, ursodeoxycholic, and
deoxycholic), and for their corresponding unconjugated forms. Retention
times are also reported for the glycine and taurine amidates as well as the
unconjugated form of the C23 norderivatives of these bile acids. Maximal
absorbance of bile acid amidates is at 200 nm and is very similar for the
(unsulfated) glycine and taurine amidates. Sulfated lithocholyl amidates
exhibit molar absorptivities at 200 nm which are 1.4 times greater than
that of non- sulfated lithocholyl amidates. Unconjugated bile acid
absorbance at 200 nm or 210 nm is 20 to 30 times less than that of
corresponding peptide conjugates. The method has been applied to samples of
gallbladder bile obtained from 14 healthy subjects to define the pattern of
conjugated bile acids present in human bile.
ARTICLES
High pressure liquid chromatographic analysis of conjugated bile acids in human bile: simultaneous resolution of sulfated and unsulfated lithocholyl amidates and the common conjugated bile acids
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