Spontaneous Oxidation of Methionine: Effect on the Quantification of Plasma Methionine Levels

Abstract

Plasma methionine (Met), methionine sulfoxide (MSO), and total Met concentrations were determined by reversed-phase chromatography and fluorescence detection after automated precolumn derivatization with ano-phthalic aldehyde mercaptoethanol reagent. Addition of pure, MSO-freel-Met to plasma samples resulted in the anticipated linear increase in plasma Met concentrations, but simultaneously effected a dose-dependent, linear increase in MSO levels. In contrast, the addition of purel-MSO to plasma samples rendered linear calibration curves for MSO, while the Met concentration remained constant. A strong buffering effect against the spontaneous or hydrogen peroxide induced oxidation of Met to MSO was observed in plasma samples. This protective effect could be neutralized by preincubating the plasma samples with sodium azide. The addition of relatively low concentrations of red cell lysates to plasma samples, prior to hydrogen peroxide oxidation, strongly inhibited the conversion of Met to MSO. Plasma samples from 127 healthy female volunteers were analyzed: MSO concentrations (mean, 3.6 ± 2.1 μm) exhibited a weak positive correlation (r= 0.352) with Met levels (mean, 21.3 ± 6.1 μm) but, after the exclusion of two probable outliers from the data set, no correlation was observed. Our results suggest that plasma Met concentrations should be corrected for oxidative losses incurred during storage, sample processing and because of the action of a variety ofin situoxidants, present in plasma, in order to obtain a reliable estimate of the methionine status of an individual.