Methods: 2,2-diphenyl-1-picrylhydrazyl (DPPH) and Ferric reducing antioxidant power assay (FRAP) were applied to evaluate the antioxidant activity of carob. In vitro cytotoxicity of carob was conducted on human hepatocytes (WRL68) and rat pancreatic β-cells (RIN-5F). Acute oral toxicity of carob was conducted on a total of 18 male and 18 female Sprague-Dawley (SD) rats, which were subdivided into three groups (n = 6), namely: high and low dose carob-treated (CS5000 and CS2000, respectively) as well as the normal control (NC) receiving a single oral dose of 5,000 mg kg-1 carob, 2,000 mg kg-1 carob and 5 mL kg-1 distilled water for 14 days, respectively. Alkaline phosphatase, aspartate aminotransferase, alanine aminotransferase, total bilirubin, creatinine and urea were assessed. Livers and kidneys were harvested for histopathology. In vitro inhibitory effect against α-amylase and α-glucosidase was evaluated. In vivo glycemic activity was conducted on 24 male SD rats which were previously intraperitoneally injected with 55 mg kg-1 streptozotocin (STZ) followed by 210 mg kg-1nicotinamide to induce type 2 diabetes mellitus. An extra non-injected group (n = 6) was added as a normal control (NC). The injected-rats were divided into four groups (n = 6), namely: diabetic control (D0), 5 mg kg-1glibenclamide-treated diabetic (GD), 500 mg kg-1 carob-treated diabetic (CS500) and 1,000 mg kg-1 carob-treated diabetic (CS1000). All groups received a single oral daily dose of their treatment for 4 weeks. Body weight, fasting blood glucose (FBG), oral glucose tolerance test, biochemistry, insulin and hemostatic model assessment were assessed. Pancreases was harvested for histopathology.
Results: Carob demonstrated a FRAP value of 3191.67 ± 54.34 µmoL Fe++ and IC50 of DPPH of 11.23 ± 0.47 µg mL-1. In vitro, carob was non-toxic on hepatocytes and pancreatic β-cells. In acute oral toxicity, liver and kidney functions and their histological sections showed no abnormalities. Carob exerted an in vitro inhibitory effect against α-amylase and α-glucosidase with IC50 of 92.99 ± 0.22 and 97.13 ± 4.11 µg mL-1, respectively. In diabetic induced rats, FBG of CS1000 was significantly less than diabetic control. Histological pancreatic sections of CS1000 showed less destruction of β-cells than CS500 and diabetic control.
Conclusion: Carob pod did not cause acute systemic toxicity and showed in vitro antioxidant effects. On the other hand, inhibiting α-amylase and α-glucosidase was evident. Interestingly, a high dose of carob exhibits an in vivo antihyperglycemic activity and warrants further in-depth study to identify the potential carob extract composition.
METHODS: Wistar rats employed for this study consisted of normoglycaemic and diabetic rats in nine experimental groups. The normoglycaemic and diabetic rats were either treated with metformin (500 mg/kg b.w.), quercetin (10 mg/kg b.w.), or ethanol extract of H. verticillata leaf (250 mg/kg b.w. and 500 mg/kg b.w.) administered orally for 28 days.
KEY FINDINGS: Results revealed that H. verticillata significantly lowered blood glucose level, attenuated dyslipidaemia, decreased atherogenic coefficient, atherogenic and coronary risk indices, and increased cardioprotective index in diabetic rats. Also, H. verticillata significantly decreased serum urea, creatinine, aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase and unconjugated bilirubin levels, relative to untreated diabetic rats. Further, H. verticillata increased serum superoxide dismutase, catalase and glutathione peroxidase activities and glutathione level, and decreased malondialdehyde level in diabetic rats in a manner similar to metformin and quercetin. Histopathological investigation of the liver and kidney revealed restored hepatocytes and amelioration of congested interstitial blood vessel of the Bowman's space of the kidneys upon intervention with H. verticillata.
SIGNIFICANCE: H. verticillata in addition to its anti-hyperglycaemic activity ameliorates oxidative stress, dyslipidaemia, atherogenicity and hepatorenal lesions in DM.