The mixture was cooled
in an ice bath for 15 min and centrifuged at 10,621g for 5 min at
room temperature. Aliquot of 0.6 ml of the supernatant was added
to 0.8 ml of the complexing reagent (0.5 g of 2,20-bipiridine and
65 ll of thioglycolic acid dissolved in 50 ml of 0.2 M HCl) and
absorbance was read at 540 nm using a microplate reader (BioTek
Instruments, Winooski, VT, USA) controlled by the Gene 5TM software
version 1.1. (BioTek Instruments).