|Chronic hyperglycemia is a major cause of oxidative stress in human. High oxidative stress leads to macrovascular and microvascular dysfunction, which are the important cause of diabetic cardiovascular and renal complications. The present study aims to investigate the antioxidative effect of Bauhinia penicillioba (BP) leaf extracts on the protection of vascular and kidney function in diabetic rats including primary toxicity study. The 95% ethanolic extract of BP leaves were prepared using percolation techniques and consequencely partial purified by column chromatography. The most antioxidative fractions were investigated. The protective effects of vascular and kidney functions were evaluated in streptozotocin (70 mg/kg, iv.) -induced diabetic rats. Vascular function was evaluated by comparing the percent of maximal aortic contraction responded to phenylephrine between groups. The 2-week interval blood urea nitrogen (BUN), serum creatinine (SCr), and the last week creatinine clearance (ClCr) were used to evaluate the kidney function. Both vascular and kidney function were compared between normal rats, diabetic control rats, and diabetic rats orally received BP leaf extract (100 or 200 mg/kg), glyburide (10 mg/kg), or vitamin E (300 mg/kg) consecutively for 6 weeks. Acute toxicity study was performed by administrating a single various doses of BP leaf extract intragastrically or intraperitoneally to both sexes of mice and the LD50 were then determined. Sub-chronic toxicity was conducted by orally giving the BP leaf extract to both sexes of mice for 8 weeks. The body weight, organ weight, AST, ALT, ALP, Cr, and BUN serum levels were determined and compared between treated and control group.
The results showed that the ethanolic extract of BP leaf was accounted for 23.45 % of dry weight. The partial purified fraction containing the highest antioxidative activity determined by DPPH assay (IC50=5.55 g/ml) consisted of flavonoids as a major constituent, and it was accounted for 1.84 % of crude extract. Evaluation of kidney function of diabetic rats demonstrated that BUN of diabetic rats was significantly higher than those of normal rats in all weeks. The week-2 BUN of diabetic rats received glyburide, vitamin E, glyburide + vitamin E, glyburide + BP 100 mg/kg, and vitamin E+BP 100 mg/kg significantly reduced when compared to diabetic rats. In week 4, only glyburide + vitamin E significantly reduced the BUN level, however it was not observed in week 6. Serum creatinine level was not significant difference between groups. ClCr of diabetic control rats was highest (7.29±2.57 ml/min) and higher than those of other groups. The aortic contraction responses to phenylephrine showed that the percent of maximal aortic contraction of diabetic control rats (379.29±41.42%) were higher than those of normal rats (276.42±37.96%). The percent of maximal aortic contraction of glyburide-treated diabetic rats were reduced but it was not significant. Vitamin E and BP leaf extract (200 mg/kg) significantly reduced the percent of maximal aortic contraction (249.79±75.24 and 277.13±99.69%, respectively) from diabetic control rats. In acute toxicity study, oral administration of the maximal dose of 6,000 mg/kg of the extract to both male and female mice showed neither clinical adverse effect nor animal death (LD50 > 6,000 mg/kg). Peritoneal administration of the maximal dose of 4,000 mg/kg of the extract to male mice did not cause 50% death (LD50 > 4,000 mg/kg). However, the dose of extract that causes 50% death in female mice was 2,765.5 mg/kg (LD50 = 2,765.5 mg/kg). The results of sub-chronic toxicity showed no difference in apparent adverse event, behavioral change, overall health and growth rate between the treatment and the control group. The animal body weight, major organ weight, ALT, AST, ALP, BUN and Cr serum levels were also not different between both groups.
The ethanolic extracts of BP leaf exhibited potent antioxidative effect in vitro but moderate effect for protection of vascular and kidney functions. However, the BP extracts were safe and practically non toxic in mice.