Simultaneous determination of three aminothiols in human plasma by high performance liquid chromatography with gold nanoparticle enrichment

Abstract

A high performance liquid chromatography (HPLC) method with ultraviolet detection (UVD) for the simultaneous determination of cysteine (Cys), homocysteine (Hcys) and glutathione (GSH) in human plasma was established. Tween 20-capped gold nanoparticles (Tween 20-AuNPs) were used as selective probes for the extraction and enrichment of amino-thiols. The aminothiols removed from the Tween 20-AuNPs by dithiothreitol were separated on a Spursil C18 column (250 mm x 4.6 mm, 5 microm) with a mobile phase of 60 mmol/L phosphate buffer solution (pH 2.0). The column eluent was monitored using UVD at the wavelength of 200 nm. The linear ranges were found between peak area and concentration in the ranges of 0.025-350 micromol/L for Cys, 0.02-60 micromol/L for Hcys and 0.01-50 micromol/L for GSH, with linear regression coefficients all above 0.99. The limits of detection (LOD, signal to noise ratio of 3) for the aminothiols were between 2.5 nmol/L and 6.0 nmol/L and the recoveries were between 92.8% and 106.0%. Due to the significant interference reduction of endogenous substances in the plasma and the subsequent improvement of the selectivity and sensitivity of HPLC-UVD, the present method was applied for the simultaneous analysis of the forementioned aminothiols in the plasma samples of cardiovascular disease patients. The results showed that the levels of Hcys and GSH had significant differences between the patients and the controls, while that of Cys had no significant difference.