Biliary metabolites of all-trans-retinoic acid in the rat: isolation and identification of a novel polar metabolite.

The biliary metabolites from normal rats dosed with either pharmacological or physiological doses of all-trans-[11,12-3H2]retinoic acid were investigated. Biliary metabolites excreted during the first 24 h account for approximately 60-65% of the radiolabeled dose. A major polar metabolite was purified to homogeneity by using Sephadex LH-20 chromatography and several high-performance liquid chromatographic procedures. This metabolite was negatively charged as revealed by high-performance liquid chromatography on ion-exchange columns and accounts for 10% of the total biliary radioactivity (6% of the dose). The polar compound was positively identified by using Fourier transform proton nuclear magnetic resonance spectroscopy, high- and low-resolution mass spectrometry, fast atom bombardment mass spectrometry, ultraviolet absorption spectrophotometry, Fourier transform infrared spectroscopy, amino acid analysis, and chemical derivatization as 2-[8-[6-(hydroxymethyl)-2,6-dimethyl-3-oxo-1-cyclohexen-1-yl]-2,6- dimethyl-5,7-octadienamido]ethanesulfonic acid. The metabolic transformations required for the generation of this metabolite from all-trans-retinoic acid are the following: (1) allylic oxidation at carbon 4 of the cyclohexene ring to produce a 4-keto group, (2) hydroxylation of one of the methyl groups at carbon 1 of the cyclohexene ring, (3) saturation of the two terminal double bonds in the side chain, (4) loss of the terminal carboxyl group of the side chain via decarboxylation, and (5) conjugation of the resulting retinoid with taurine. To our knowledge, this metabolite represents the first taurine conjugate of a fat-soluble vitamin to be identified.