Determination of Retinol and Retinoic Acid in Capillary Blood by High Performance Liquid Chromatography

Abstract

A simplified method for quantitation of retinol in capillary blood by high-performance liquid chromatography (HPLC) is described. Serum (10–20 μ1) obtained by centrifugation of capillary blood is extracted with a solvent mixture of isopropanol-dichloroethane, and the extract is directly injected into a HPLC system to quantitate retinol. The lower limit of quantitation is 0.25 ng retinol, thus permitting quantitation of serum or plasma level of 8 μg or more retinol/L. Although endogenous retinoic acid cannot be analyzed, retinoic acid, following an oral dose, can be extracted along with retinol in presence of acetic acid and quantitated. The lower limit of quantitation of retinoic acid is 0.5 ng, thus permitting quantitation of 50 ng or more of retinoic acid/ml serum or plasma. The small sample size required, their simple preparation, and rapid analysis make this method well suited for clinical studies. The method should be very useful for monitoring retinol level in blood of patients suspected of vitamin A deficiency or of retinol and retinoic acid levels in patients undergoing retinoic acid therapy for skin disorders or acute promyelocytic leukemia.