Bile acids methyl esters, relative retention

TABLE XII. Relative Retention Times of Different Bile Acid Methyl Ester Derivatives on Methyl (SE-30 and OV-1) and Methyl-Phenyl (SE-52, PhSi-20, and OV-17 5, 20, and 50% Phenyl Groups, Respectively) Substituted Silicones... 

TABLE XIII. Relative Retention Times" of Different Bile Acid Methyl Ester Derivatives on QF-1... 

With a purified enzyme preparation from Clostridium perfringens, Roovers et al. (36) cleave conjugated bile acids directly in plasma. Proteins are then precipitated with Ba(OH)2-saturated ethanol (123) and the supernatant is taken to near dryness. The residue is dissolved in a toluene-isopropanol-methanol-30 % aqueous NaOH (10 20 20 6, v/v) mixture, water is added, and neutral lipids removed by light petroleum extraction. Bile acids are then obtained by acidification and diethyl ether extraction. The bile acids are then methylated and analyzed as above except that the bile acid methyl esters are acetylated before chromatography on 1 % XE-60 columns at 250°C. With this column the following retention times relative to that of the diacetoxy derivative of methyl deoxycholate were found for the following acetate methyl ester derivatives lithocholic, 0.60 23-nor-deoxycholic acid (internal standard), 0.77 chenodeoxycholic acid, 1.24 and cholic acid, 1.88. The deoxycholic acid derivative was eluted after 9.0 min and methyl 5 3-cholanoate after 0.32 min. 

Slightly hindered hydroxyl groups of bile acids can be silylated according to the procedure of Makita and Wells [319] methyl esters of bile acids, prepared by treatment with diazomethane, are dissolved in anhydrous pyridine (ca. 1 ml per 10 mg) and 0.1 ml of HMDS and 0.03 ml of TMCS are added to this solution. After standing for 10 min at room temperature an aliquot of the reaction mixture is injected directly. The method was applied to the determination of faecal bile acids in the rat. Other workers [320] investigated the retention behaviour of 52 persilylated methyl esters of urine acids on QF-1, OV-1 and OV-17 stationary phases. They correlated the data expressed as relative retention times with the structures of the compounds. 

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