Fluorometric kinetic studies.
The effect of AA and GPE on purified
GST (from rabbit liver, Sigma cat. no. G8261, 6 U ml21 final
concentration) as a function of GSH concentration was investigated.
Increasing concentrations of GSH (in PBS pH 5?6) in the absence
and presence of AA (0?5 or 1?0 mM) and GPE (5 or 10 mg ml21)
were added to 3 ml PBS (pH 5?6); a subsequent addition of 50 ml
GST was used to commence the reaction. Monochlorobimane
(MCB, 2 mM final concentration; lmax excitation 391 nm; lmax emission
491 nm) was used as previously described (Aon et al., 2003),
and fluorescence measured using a Cary Eclipse fluorescence
spectrophotometer (Varian).