effect of infection of the filarial parasite brugia malayi - Pondicherry ...

dichromate/acetic acid mixture. The dichromate in acetic acid is reduced to chromic acetate,
whcn heated in the presence of hydrogen peroxide with the formation of perchloric acid as
an unstable intermediate. The chromic acetate thus produced is measured at 610 nm. Since
dichrornate has no absorbance in this region, the presence of this compound in the assay
mixture does not interfere with the calorimetric determination of chromic acetate.
Resgents
I. Stock dichromatdacetic acid reagent : This reagent was prepared by mixing a 5%
solution of potassium dichromate with glacial acetic acid (1 :3) by volume.
2. Working dichromatdacetic acid reagent : The stock was diluted (1:s) with water to
make the working dichromatdacetic acid solution.
3. Hydrogen peroxide (0.2 M) : 1.0 ml of 30% hydrogen peroxide was made
upto 45.0 ml with water.
4. Phosphate buffer : 0.01 M, pH 7.0
The reaction mixture contained 0.5 ml of hydrogen peroxide, 1.0 rnl of buffer, 0.4 ml
of water and 0.1 ml of diluted homogenate (1: 10). After 15, 30 and 60 seconds of incubation,
2.0 ml of dichromateJacetic acid reagent was added. To the control tube, the enzyme was
added after the addition of acid reagent The tubes were then heated and the colour
developed was read at 610 nm.The activity of catalase was amved at from the amount of
hydrogen peroxide consumed and was expressed as poles of hydrogen peroxide
consumed/rninutdmg protein.
'Ibis was assayed by the method of Beutler (1984).
GST catalyses the reaction of I-chloro, 2,4-dinitrobenzene with the -SH group of
glutahone. The conjugate formed is measured at 340 nrn.
Reagents
1. Potassium phosphate buffer : 0.5 M, pH 6.5
2. I-chlm -2,4 dinitrobemne (CDNB) : 30 mM in 95% ethandl