High-performance liquid chromatography of bile pigments: separation and characterization of the urobilinoids.

The detailed analysis of faecal bile pigments by high-performance liquid chromatography is described. Non-aqueous reversed-phase systems with acetonitrile-dimethyl sulphoxide or acetonitrile-dimethyl sulphoxide-methanol as the mobile phase on C1, C8 or C18-bonded silica are used for the group separation of verdinoids, violinoids and urobilinoids. A silica column, with acetonitrile-water-tetraethylene-pentamine as mobile phase, separates the laevorotatory stercobilin (C33H46N4O6) and half-stercobilin (C33H44N4O6) from the optically inactive urobilin (C33H42N4O6). The diastereoisomers are resolved by converting the urobilinoids into their dimethyl esters before chromatography on a silica column with n-heptane-methyl acetate-methanol containing 1% of diethylamine as the solvent system.

High-performance liquid chromatography of bile pigments: application to the determination of unconjugated bilirubin in serum.

A rapid, sensitive and specific method for the determination of unconjugated bilirubin in serum is described. Reversed-phase chromatography on short-chain alkylsilica (C2) or octadecylsilica (C18) with acetonitrile--dimethylsulphoxide--water as eluent is used for the separation of bilirubin. The method is applied to the analysis of bilirubin in patients with disorders associated with jaundice.