Simultaneous Determination of Homocysteine, Cysteine, Glycine, Glutamic Acid, Methionine, Serine and Taurine in Human Plasma by Using Reverse Phase-High Performance Liquid Chromatography

Alterations in the plasma aminothiol levels can be considered as biomarkers for the diagnosis and screening of human disorders namely cardiovascular diseases. Among the common method, ELISA is available for homocysteine estimation, the analytical method like HPLC is considering as a gold standard. In the proposed study, we have developed a RP-HPLC method for the simultaneous determination of homocysteine, cysteine, glycine, glutamic acid, methionine, serine and taurine in human plasma using fluorescence detector. Separation was achieved on C18 column with gradient elution using 80% methanol, 0.05 M acetate buffer (pH-7.0) and the column was maintained at 26°C and the mobile phase was pumped at flow rate of 1.0 mL min-1. The detection of eluent was carried out at fluorimetrically (excitation 340 nm, emission 450 nm) with a total run time 30 min. The calibration curve ranged from 25 to 100 ng/mL for the analytes. Inter and intraday precision and accuracy, stability studies and sample reanalysis were investigated for all the analytes, and the results met the acceptance criteria. This method will be a useful tool in the investigation of the roles of these important thiol-containing compounds and their corresponding disulfides in physiological and pathological processes