Carotenoids, retinoids and alpha-tocopherol in human serum: Identification and determination by reversed-phase HPLC

A rapid, simple and specific high performance liquid chromatographic procedure for assaying alpha- and beta-carotene is described. The method also enables the simultaneous determination of retinol and dl-alpha-tocopherol in human serum. The same chromatographic procedure can be used to assay the major carotenoids in human serum, provided analyses are replicated and the effluent is monitored at 450 nm. The conditions described also enable determination of licopene, cryptoxanthine and lutein with zeaxanthine. An aliquot of 0.5 ml serum is deproteinized with ethanol (0.5 ml) and extracted with petroleum ether (0.75 ml). The petroleum ether extract is evaporated until dry and then redissolved immediately with 0.5 ml of an eluent mixture consisting of methanol-hexane (85:15, v/v). Aliquots of 50 microl are then injected onto a 250 x 4.6 mm column packed with Spherisorb ODS-2. Owing to its good reproducibility, the procedure can be used for assays with external standards. Clinical applications are described for cases of hypercarotinemia associated with endocrine dysfunctions such as hypothyroidism and diabetes.