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38 Mahgoub Mohammed AHMED and Safaa Eid ALI water bath at 100 ◦ C for 30 min. After cooling with tap water to room temperature, 4ml n-butanol was added and shaken vigorously. After centrifugation, the color of butanol layer was measured at 535 nm. Assay for glutathione-S-transferase (GST) activity in PMS Glutathione-S-transferase activity was assayed by the method of Habig et al. (1974). The method is based on the rate of conjugate formation between GSH and 1-chloro-2,4-dinitrobenzene (CDNB). The absorbance change was recorded at 340 nm and the enzyme activity calculated as nmol CDNB conjugates formed/min/mg protein. Assay for glutathione peroxidase (GPx) activity in PMS Glutathione peroxidase activity was assayed by the method of Mohandas et al. (1984). The change in absorbance was recorded spectrophotometrically at 340 nm. GPx activity was expressed as nmol NADPH oxidized/min/mg protein. Assay for glutathione reductase (GR) activity in PMS Glutathione reductase activity was determined by the method of Carlberg and Mannervik (1975). GR was assayed by following the oxidation of NADPH at 340 nm at 37 o C. GR activity was expressed as nmol NADPH oxidized/min/mg protein. Assay for catalase (CAT) activity in PMS CAT activity measurement in PMS was measured by the method of Takahara et al. (1960). The reduction rate of H2O2 was followed at 240 nm for 30 s at room temperature. CAT activity was expressed in nmol H2O2 consumed/min/mg protein. Assay for glucose-6-phosphate dehydrogenase (GPD) activity in PMS The activity of glucose-6-phosphate dehydrogenase was determined according to the method of Zaheer et al. (1965). The changes in absorbance were recorded at 340 nm and enzyme activity was calculated as nmol NADP reduced/min/mg protein. Estimation of protein concentration The protein concentration in all samples was determined by the method of Lowry et al. (1951). Statistical analysis The results are expressed as Mean±SEM. The collected data were statistically analyzed by the least significant differences (LSD) at the level 5% of the probability procedure according to Snedecor and Cochran (1980). Results Effect of pomegranate peel extract on renal toxicity markers The effect of pre-treatment of rats with pomegranate peel extract on Fe-NTA-induced enhancement in the level of serum creatinine and urea are shown in Table (1). Fe-NTA treatment leads to about 147% and 303% enhancement in the values of creatinine and urea, respectively, as compared with saline-treated group. Prophylaxis with pomegranate peel extract at both doses resulted in 28-45% and 48-88% reduction in the values of serum creatinine and urea respectively as compared with Fe-NTA-treated group. Effect of pomegranate peel extract on glutathione metabolism Table (2) shows the effect of pretreatment of rats with pomegranate peel extracts on Fe-NTAmediated renal glutathione content and on the activities of its metabolizing enzymes, viz, glutathione-S-transferase and glutathione reductase. Treatment with Fe-NTA alone resulted in the depletion of renal glutathione and reduction in the activities of glutathione-S-transferase and glutathione reductase by 48%, 55% and 46% respectively, as compared with saline-treated group. However, pretreatment of animals with pomegranate peel extract at 100 and 200 mg/kg body weight resulted in the recovery by 79-83%, 46-73% and 40-72% respectively, as compared with Fe-NTA-treated group.
Pomegranate peel extract against renal oxidative damage 39 Table 1. Effect of pomegranate peel ethanol extract on Fe-NTA-induced enhancement of serum creatinine and urea in rats Values Mean±SEM (n=6 animals). a p
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