Quantification of Plasma and Leukocyte Vitamin C by High Performance Liquid Chromatography with Mass Spectrometric Detec
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Quantification of Plasma and Leukocyte Vitamin C by High Performance Liquid Chromatography with Mass Spectrometric Detection Attila Szőcsa, Szende Vanceab, *, István Kissa, and Gabriella Donáth-Nagyb aDepartment
of Public Health Medicine, University of Pécs, Pécs, HU-7624 Hungary Faculty of Pharmacy, University of Medicine, Pharmacy, Sciences and Technology of Targu Mures Târgu Mureş, RO-540139 Romania *e-mail: [email protected]
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Received February 28, 2018; revised June 1, 2018; accepted March 12, 2020
Abstract—In the present study a high performance liquid chromatographic–mass spectrometric (LC–MS) method was developed for the measurement of ascorbic acid in human plasma and white blood cells. Special attention was paid to the known difficulties of ascorbic acid stability, and a plasma sample preparation method was developed, accordingly. The novel objective of this work was to focus on the determination of plasma ascorbic acid concentrations considering the necessity of accurate determination for further clinical studies related to scurvy induced by various pathological conditions. The presented study describes in detail a validated LC–MS method with adequate accuracy and precision for quantitative determination of ascorbic acid in human plasma. The method was tested on human plasma and leukocyte samples. Keywords: ascorbic acid, dehydroascorbic acid, human plasma, LC–MS, stability, scurvy DOI: 10.1134/S1061934820090038
Vitamin C is a mild organic acid, structurally similar to glucose from which most animals produce their own ascorbic acid. The most important chemical property of this vitamin is the reversible oxidation to dehydroascorbic (DHA) acid, which means its underlying physiological activity. Ascorbic acid (AA) can exist as a free radical, which explains the antioxidant activity. Under physiological conditions, primarily the reduced (ascorbate) form is present [1, 2]. Vitamin C has already been studied more than any other supplement or pharmaceutical drug in the history, a fact that is not surprising given that scurvy is a fatal disease and chronic hypoascorbemia is considered today’s most widespread deficiency disorder [3, 4]. In diabetes, allergy, infectious diseases and toxic abuse, the manifestation of this avitaminosis is in most cases inevitable and intravenous administration of ascorbic acid is often a part of emergency care also in our days [5, 6]. Furthermore, the incidence of acute and subacute infantile scurvy needs a special attention. On the basis of epidemiological surveys, the number of scorbutic children is increasing [7, 8]. Based on all these facts, it is important to notice the opinion of all those physicians who declare that, especially in the mentioned pathological states, the accurate quantification of ascorbic acid is needed. Considering that the most important reducing system in humans is represented by the glutathione–ascorbic acid interaction
[9, 10], wherein the presence of both molecules is essential, and knowing also that in plasma the primarily used antioxi
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