期刊
ANALYTICAL BIOCHEMISTRY
卷 426, 期 2, 页码 109-117出版社
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.ab.2012.04.014
关键词
Ascorbic acid; Vitamin C; Red blood cells; Electrochemical detection; High-pressure liquid chromatography
资金
- National Institutes of Diabetes and Digestive and Kidney Diseases, National Institutes of Health
Although vitamin C (ascorbate) is present in whole blood, measurements in red blood cells (RBCs) are problematic because of interference, instability, limited sensitivity, and sample volume requirements. We describe a new technique using HPLC with coulometric electrochemical detection for ascorbate measurement in RBCs of humans, wild-type mice, and mice unable to synthesize ascorbate. Exogenously added ascorbate was fully recovered even when endogenous RBC ascorbate was below the detection threshold (25 nM). Twenty microliters of whole blood or 10 mu l of packed RBCs was sufficient for assay. RBC ascorbate was stable for 24 h from whole-blood samples at 4 degrees C. Processed, stored samples were stable for >1 month at -80 degrees C. Unlike other tissues, ascorbate concentrations in human and mouse RBCs were linear in relation to plasma concentrations (R = 0.8 and 0.9, respectively). In healthy humans, RBC ascorbate concentrations were 9-57 mu M, corresponding to ascorbate plasma concentrations of 15-90 mu M. Mouse data were similar. In human blood stored as if for transfusion, initial RBC ascorbate concentrations varied approximately sevenfold and decreased 50% after 6 weeks of storage under clinical conditions. With this assay, it becomes possible for the first time to characterize ascorbate function in relation to endogenous concentrations in RBCs. Published by Elsevier Inc.
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