2.6. Sulphydryl group determination
The sulphydryl group of the hydrolysates and GSH oxidation assay mixtures was determined as reported (Udenigwe et al., 2014). The sample (500 µL) was incubated in 500 µL of 0.1 M Tris-glycine buffer (pH 8.0) containing 7.5 M urea at 40 °C for 30 min. Thereafter, 25 µL of DTNB (4 mg/mL, in Tris-glycine buffer) was added to the mixture and incubated at room temperature for 10 min. Absorbance was then measured at 412 nm, and sulphydryl group content was calculated as µM SH/g = 73.53A412nm (D/C), where D is dilution factor, A is absorbance and C is concentration of the hydrolysates.