Plasma Retinol Assay by Elution from Silicic Acid with Cymene (p-Isopropyl Toluene)

Abstract

Abstract We developed a microcolumn adsorption chromatographic technique for assay of plasma retinol. A petroleum ether extract of plasma was fractionated on silicic acid, to isolate retinol. The first eluent, petroleum ether, removed "unidentified fluorescent contaminant" (UFC); the second eluent, cyclohexene, removed traces of UFC and retinyl palmitate. Retinol was eluted by the third eluent, cymeme (p-isopropyl toluene). No recognized plasma vitamin A derivatives, other than retinol, were detectable in the cymene eluate. With the procedure, we could detect a minimum concentration of 7 µg of retinol per deciliter of plasma. Sixty-six plasmas from preschool children were assayed by our "cymene assay," and by the Neeld—Pearson modification of the Carr—Price procedure, the method of Thompson et al., and the column assay of Garry et al. Results by the latter three procedures were similar (P >0.025): the mean plasma retinol ranged from 29.4 to 31.2 (SD, ± 5.8 to 7.4) µg/dl. By the present assay the same samples contained 24.5 (SD, ± 6.8) µg/dl; significantly more was detected by the other procedures (P <0.001). This difference (16-22%) between the cymene assay for retinol and other assays for total vitamin A may reflect, in part, the concentration of other vitamin A derivatives in plasma.