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

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Abstract
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 1st 24 h account for .apprx. 60-65% of the radiolabeled dose. A major polar metabolite was purified to homogeneity by 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 Fourier transform .H NMR spectroscopy, high- and low-resolution mass spectrometry, fast atom bombardment mass spectrometry, UV absorption spectrophotometry, Fourier transform IR 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: allylic oxidation at C4 of the cyclohexene ring to produce a 4-keto group, hydroxylation of one of the methyl groups at Ca1 of the cyclohexene ring, saturation of the 2 terminal double bonds in the side chain, loss of the terminal carboxyl group of the side chain via decarboxylation and conjugation of the resulting retinoid with taurine. This metabolite apparently represents the first taurine conjugate of a fat-soluble vitamin to be identified.