Characterization of oxidized and glucuronidated metabolites of retinol in monkey plasma by thermospray liquid chromatography/mass spectrometry

Abstract

Eight metabolites of retinol were isolated by high-performance liquid chromatography (HPLC) from the plasma of the non-human primate Macaca fascicularis after acute exposure to 150,000 IU of vitamin A per kilogram body weight. After enrichment and further chromatographic purification, the metabolites were reinjected individually into a second HPLC system which was connected on-line by a thermospray interface to a mass spectrometer operated in the positive ionization mode. Six retinoids were identified by (i) a comparison of their retention times with those of appropriate reference compounds in the two chromatographic systems and (ii) by comparison of their mass spectra with those of reference compounds. These retinoids were: 13-cis-4-oxoretinoic acid, all-trans-4-oxoretinoic acid, 13-cis-retinoic acid, all-trans-retinoic acid, all-trans-retinoyl beta-glucuronide and all-trans-retinyl beta-glucuronide. One further metabolite could be identified for the first time as all-trans-4-oxoretinoyl beta-glucuronide by its mass spectrum and, after treatment of the unknown metabolite with beta-glucuronidase, by its hydrolysis product all-trans-4-oxoretinoic acid. The molecular structure of one metabolite could not be elucidated. A major metabolic pathway of high-dose vitamin A in the non-human primate is apparently the oxidation of the primary alcohol group of retinol resulting in the formation of all-trans-retinoic acid. Subsequently, a broad spectrum of various metabolites of all-trans-retinoic acid, including beta-glucuronides and retinoids with a 13-cis configuration, appear in the plasma.

4-Methylpyrazole partially ameliorated the teratogenicity of retinol and reduced the metabolic formation of all-trans-retinoic acid in the mouse

“…Previous studies have suggested that the embryotoxic sequelae result from the biotransformation of retinol to all-trans RA Eckhoff et al 1989), although it is difficult to attribute the teratogenicity of retinol to a single metabolic product because there are a number of retin01 metabolites in addition to the parent compound (e.g., Eckhoff et al 1990). There is evidence that all-trans-RA can be synthesized from retinol by several different enzymatic systems.…”

Methods for detecting and identifying retinoids in tissue

“…Reproduced with permission from Garwin and Saari (2000). in a series or Superspher 100 RP-18e column in combination with acetonitrile-water-ammonium acetateacetic acid mixtures and gradient elution for separation of various polar retinoids (Wyss and Bucheli, 1988a,c, 1990, 1997Wyss, 1990;Wyss et al, 1998). Schmidt et al (2003) used a binary gradient formed from 60 mM ammonium acetate (pH 5.7)/ methanol (1:1) and 100% methanol in combination with a Spherisorb ODS2 (2.1 Â 150 mm, 3 m) reverse phase column for polar retinoids, and a YMC, J'sphere ODS-H80 (4.6 Â 150 mm, 4 m) eluted with methanol, acetonitrile, and chloroform in a gradient for the apolar retinoids ROH, RAL, and RE.…”

“…Historically only a handful of articles have described the use of LC-MS for the determination of retinoids in biological samples (Eckhoff et al, 1990;Fayer et al, 1991;Ranalder et al, 1993;Bempong et al, 1995). During the last decade however, modern soft atmospheric pressure ionization techniques like electrospray (ESI) and atmospheric pressure chemical ionization (APCI) have been explored for retinoid analysis.…”

Plasma Retinoids after Topical Use of Retinaldehyde on Human Skin

“…Polar retinoids (all-trans-RA, 13-cis-RA, 13-cis-4-oxo-RA) were determined by the method of Ekhoff and Nau [10] without internal standardization [11]; the limit of quantification was 0.5 ng/ml. ROL, RAL and RE concentrations were obtained by the method of Collins et al [12].…”