Abstract
A simple and rapid method for the determination of reduced glutathione (GSH) in plants and biological samples by thermospray high-performance liquid chromatography-mass spectrometry (LC/MS) was studied. After a sample was homogenized in 0.1% trifluoroacetic acid (TFA) in a glass homogenizer with teflon pestle, the homogenate was centrifuged at 10000 rpm for 10 min. The supernatant solution was filtered through disposable syringe filter unit (cellulose acetate, 0.45 μm), and the filtrate was applied for the determination of GSH by selected ion monitoring (SIM) of a protonated molecular ion at m/z 308. Analytical conditions for LC/MS were as follows : column : Shim-pack CLC-ODS (6.0 mm φ×15 cm), mobile phase : 0.05% TFA-acetonitrile (95 : 5 v/v), flow rate : 0.8 ml/min, exit temperature from the vaporizer : 200°C, ion source block temperature : 260°C Under these conditions, GSH in plants and biological samples could be determined with recovery of 95-105%. The coefficient of variation was as low as 5%. The calibration curve was linear between 1 and 100 μg/ml (r=0.999).
