Measurement of Vitamin C in Blood Components by High‐Performance Liquid Chromatography

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Abstract
The fact that platelets, PMN leukocytes, and MN leukocytes concentrate ascorbic acid suggests that vitamin C has an important role in their physiological functions. The question still remains as to which one of the cells best reflects vitamin C status. The ascorbic acid content of PMNs and platelets correlates positively with plasma concentration and supplementation with vitamin C, as shown in Evans et al. They also found that MN leukocytes, in contrast, do not show any such relationship; however, MN leukocytes maintain the highest levels of ascorbic acid and play a very important function in immunocompetence. We have found that with a limited number of subjects, ascorbic acid content of MN and PMN leukocytes correlates positively with plasma ascorbic acid, but there was no correlation between platelets and plasma ascorbic acid (unpublished results). Therefore, further work is necessary to evaluate these three blood components for the best cellular marker of vitamin C status. We have developed a reversed-phase HPLC method for ascorbic acid that can be used in conjunction with our cellular differential centrifugation technique for the determination of ascorbic acid in relatively pure blood cell fractions. The chromatographic method is simple, sensitive, and automated. It clearly resolves ascorbic acid, which is the major form of the vitamin found in vivo and is not prone to interference by sugars, carbohydrates, or nucleotides.