0. The homogenate was centrifuged in cold at 12,000 g for 12 min. The supernatant, thus obtained, was then collected and incubated with 0.01 ml of absolute ethanol at 4 °C for 30 minutes, after which 10% Triton X-100 was added so as to have a final concentration of 1%. The sample, thus obtained, was used to determine catalase activity by measuring the breakdown of H2O2 spectrophotometrically at 240 nm. The enzyme activity was expressed as μmoles of H2O2 consumed/min/mg tissue protein. The activity of GR was determined according to the following method [24]. The assay mixture in a final volume of 3 ml contained

50 mM phosphate buffer, 200 mM KCl, 1 mM EDTA and water. The blank was set with this mixture. Then, 0.1 mM NADPH was added with suitable amount of homogenate (enzyme) into the cuvette. cAMP inhibitor The reaction was initiated with 1 mM oxidized glutathione (GSSG). The decrease in NADPH absorption was monitored spectrophotometrically at 340 nm. The specific activity of the enzyme was calculated as units/min/mg tissue protein. The GPx activity was measured according to the method of [32] with some modifications [13]. A weighed amount of gastric tissue was homogenized (10%) in ice cold 50 mM phosphate buffer containing 2 mMEDTA, pH 7.0. The assay system in a final volume of selleck chemical 1 ml contained 0.05 M phosphate buffer with

2 mM EDTA, pH 7.0, 0.025 mM sodium azide, 0.15 mM glutathione, and 0.25 mM NADPH. The reaction was started by the addition of 0.36 mM H2O2. The linear decrease of absorbance at 340 nm was recorded using a UV/VIS spectrophotometer. The specific activity of the enzyme was expressed as nmol of NADPH produced/min/mg tissue protein. The GST activity of the rat gastric tissue was measured spectrophotometrically according to the method as described by [20]. The

enzymatic reaction was measured by observing the conjugation of 1-chloro, 2,4-dinitrobenzene (CDNB) with reduced glutathione (GSH). One unit of enzyme conjugates 10.0 nmol of CDNB with reduced glutathione per minute at 25 °C. The rate where the reaction was linear was noted at 340 nm. The molar extinction of CDNB is 0.0096 μM −1/cm. The enzyme activity was expressed as units/min/mg of tissue protein. The °OH generated in the stomach were measured using DMSO as °OH scavenger [4]. DMSO Erastin ic50 forms a stable product [methanesulfonic acid (MSA)] on reaction with fast blue BB salt. Four groups of rats containing six animals per group were used for each experiment. The first group served as control and the animals were injected (i.p.) with 0.4 mL of 25% DMSO in saline per 100 g body weight. The second group served as Cu LE administered group and the animals were injected DMSO in the earlier mentioned dose 30 mins before oral administration of Cu LE at a dose of 200 mg/kg body weight. The third group was injected DMSO in the above mentioned dose exactly 30 mins before feeding piroxicam only at 30 mg/kg body weight.