Capillary Blood GSH Level Monitoring, Using an Electrochemical Method Adapted for Micro Volumes

Abstract

Glutathione (γ-glutamyl-cysteinyl-glycine; also known as GSH) is an endogenous antioxidant that plays a crucial role in cell defense mechanisms against oxidative stress. It is thus not surprising that this molecule can serve as a biomarker for oxidative stress monitoring. As capillary blood is a highly accessible target for biomarking, it is a valuable bodily fluid for diagnosing human GSH levels. This study focused on the optimization of GSH measurements from micro volumes of capillary blood prior to using electrochemical detection. The optimization of experimental parameters, including the sample volume and its stability, was performed and evaluated. Moreover, we tested the optimized method as part of a short-term study. The study consisted of examining 10 subjects within 96 h of their consumption of high amounts of antioxidants, attained from a daily dose of 2 g/150 mL of green tea. The subjects' capillary blood (5 μL) was taken at 0 h, 48 h, and 96 h for subsequent analysis. The short-term supplementation of diet with green tea showed an increase of GSH pool by approximately 38% (between 0 and 48 h) within all subjects.

Keywords: antioxidant molecules; blood drop analysis; electrochemical analysis; nutritional study; sample pretreatment.

Figure 1

The effect of sample preparation on glutathione (GSH) signal (A,C) and on the ratio of GSH to oxidized, dimeric glutathione (GSSG) (B,D) over a certain amount of time. Values are means of three replicates (n = 3). Vertical bars indicate standard error.

Figure 2

Scheme of glutathione (GSH) and oxidized, dimeric glutathione (GSSH) blood analysis procedure.

Figure 3

The effect of the tea supplementation on glutathione (GSH) levels (A) and the level of total SH-moieties in capillary blood (B) (analyzed subjects n = 10). Data are presented as median with error line (the lowest and highest value in the file).