Easy-to-Use HPLC Method to Measure Intracellular Ascorbic Acid Levels in Human Peripheral Blood Mononuclear Cells and in Plasma

Antioxidants

Will Wodzig¹  Birgit Gijsbers²  Cindy Sarodnik²  Ahmed El Molla²  Catharina Van Elssen²  Gwendolyn van Gorkom²  Erik-Jan Ververs²  Gerard Bos²  Celine Riess² 

[1] Department of Clinical Chemistry, Central Diagnostic Laboratory, Maastricht University Medical Center, 6229 HXMaastricht, The Netherlands;Department of Internal Medicine, Division of Hematology, GROW School for Oncology and Developmental Biology, Maastricht University Medical Center, 6229 HXMaastricht, The Netherlands;

关键词: ascorbic acid;    vitamin C;    peripheral blood mononuclear cells;   

DOI  :  10.3390/antiox11010134

来源: DOAJ

【 摘 要 】

Given the growing interest in ascorbic acid (AA), there is a need for a reliable and reproducible method to measure AA status in the human body. Serum AA concentrations do not correlate well with tissue levels, but AA levels in leukocytes do. However, a standard method for clinical application is lacking. This present study describes a method to measure AA in the peripheral blood mononuclear cells (PBMCs) with hydrophilic interaction liquid chromatography (HILIC). The method can also be used in plasma and other leukocyte subsets. The measurements of AA in PBMCs and plasma were performed with HPLC with HILIC separation and UV detection. The sample preparation involved the isolation of PBMCs and lysis and precipitation with acetonitrile. European Medicine Agency guidelines for bioanalytic method validation were followed for the evaluation. A highly precise execution of the method was found with intra- and inter-assay variations at a maximum of 7.8%. In 40 healthy donors, a mean intracellular AA concentration of 7.9 microgram/10⁸ cells was found in PBMCs. A correlation between plasma and PBMC AA concentration was not present (r = 0.22). In conclusion, we developed a convenient, reliable, and reproducible method for the quantitative determination of AA within PBMCs and plasma from human blood.

【 授权许可】

Unknown