BILE-ACID ANALYSIS IN BIOLOGICAL-FLUIDS - A NOVEL-APPROACH

In contrast to current methods of bile acid analysis that require the separation of bile acids into different groups prior to their analysis , the HPLC method using a reverse phase column and gradient elution th at we developed permits the separation and detection of nonconjugated, glycine-conjugated, and esterified bile acids as their fluorescent di methoxycoumarin esters. The mild conditions for ester formation make p ossible the identification of allylic bile acids characteristic of met abolic errors in bile acid synthesis. Quantification is obtained using 7 alpha,12 alpha-dihydroxy-5 beta-cholanoic acid as an internal stand ard. In addition to identification based on retention time, peak-shift strategy is used by treatment of aliquots with cholyglycine hydrolase and/or solvolysis. sis. Loss of the parent peak and appearance of the derivative provide further assurance of the identity of each bile aci d in biologic fluids that contain other organic acids.