METHOD FOR COMBINED ANALYSIS OF PROFILES OF CONJUGATED PROGESTERONE METABOLITES AND BILE-ACIDS IN SERUM AND URINE OF PREGNANT-WOMEN

A method for analysis of profiles of conjugated progesterone metabolit es and bile acids in 10 mi of urine and 1-4 mi of serum from pregnant women is described. Total bile acids and neutral steroids from serum a nd urine were extracted with octadecylsilane-bonded silica. Groups of conjugates were separated on the lipophilic ion-exchanger triethylamin ohydroxypropyl Sephadex LH-20 (TEAP-LH-20). Fractions were divided for steroid or bile acid analyses. Sequences of hydrolysis/solvolysis and separations on TEAP-LH-20 permitted separate analyses of steroid gluc uronides, monosulfates and disulfates and bile acid aminoacyl amidates , sulfates, glucuronides and sulfate-glucuronides. Radiolabelled compo unds were added at different steps to monitor recoveries and completen ess of separation, and hydrolysis/solvolysis of conjugates was monitor ed by fast-atom bombardment mass spectrometry. The extraction and solv olysis of steroid disulfates in urine were studied in detail, and extr action recoveries were found to be pH-dependent. Following methylation of bile acids, all compounds were analysed by capillary gas chromatog raphy and gas chromatography-mass spectrometry of their trimethylsilyl ether derivatives. Semiquantification of individual compounds in each profile by gas-liquid chromatography had a coefficient of variation o f less than 30%. The total analysis required 3 days for serum and 4 da ys for urine.