Pharmacognostical, Phytochemical Studies and Evaluation of Antidiabetic Potential of Biophytum sensitivum DC. and Biophytum intermedium Wt

Though there are wide developments in the treatment of diabetes mellitus with synthetic drugs, side effects such as hypoglycemia at higher dose administration, low oral bioavailability due to degradation in stomach makes it necessary to find other alternatives. Therefore, there is an owing interest in discovering natural treatments without negative side effects that can reduce these risk factors in diabetic patients. The macroscopical, microscopical studies of leaflet, stem and root to ascertain the arrangement of tissues of both selected medicinal plants Biophytum sensitivum (L.) DC. var. sensitivum and Biophytum intermedium Wight. were evaluated. The T.S. of leaflet through midrib, crystal distribution around the vascular bundle of the midrib, leaf lamina, leaf margin, stomata, T.S. of stem, T.S. of root and the powder analysis of both plants were evaluated. Quantitative microscopy, physico-chemical evaluation, extractive values and fluorescence analysis for both plants were determined. The shade dried coarsely powdered material (500 gm) extracted separately in Soxhlet apparatus using various solvents according to their polarity. As per literatures, the ethanolic extracts were selected for both plants for further study. The ethanolic extracts of both plants showed the presence of plant constituents like flavonoids, alkaloids, carbohydrates, glycosides, phytosterol, saponins, tannins and phenolic compounds, proteins and free amino acids, lignins, volatile oils by qualitative chemical tests. The various extracts of whole plant powders of Biophytum sensitivum DC. and Biophytum intermedium Wt. were subjected to Thin Layer Chromatography using mobile phase, Toluene : Methanol : Acetic acid 20:2:1, (v/v) was found to be effective, showed six spots with Rf values were calculated as 0.25, 0.36, 0.40, 0.51, 0.65 and 0.77 in EEBS and five spots with Rf values were calculated as 0.29, 0.38, 0.64, 0.71 and 0.76 in EEBI detected in UV 365nm. The EEBS and EEBI were subjected to HPTLC for the separation of phytoconstituents using Toluene : Methanol : Glacial acetic acid, 20:2:1, (v/v) as mobile phase with the standard flavonoid compound at 254nm and observed that the extracts showed the presence of flavonoid and confirmed from the chromatogram after derivatization. The Rf value of the different compounds of flavonoid present in the EEBS was found to be 0.02, 0.20, 0.26, 0.35, 0.42, 0.50, 0.63, 0.69 and 0.76 of peak 1, 2, 3, 4, 5, 6, 7, 8 and 9 respectively. The Rf value of the different compounds of flavonoid present in the EEBI was found to be 0.19, 0.27, 0.36, 0.63, 0.72 and 0.74 of peak 1, 2, 3, 4, 5 and 6 respectively. Among them, peak 5 (Rf value 0.42) was identified as flavonoid glycoside in EEBS and peak 5 (Rf value 0.72) was identified as flavonoid glycoside in EEBI as per literatures. The EEBS and EEBI were subjected to column chromatography for the separation of phytoconstituents. All the fractions were subjected to TLC studies by using Toluene : Methanol : Glacial acetic acid, 20:2:1, (v/v) as mobile phase. Fractions showing similar Rf value, melting point and identification test, were pooled together and solvents evaporated to get residues. The residues were named as BSC 1 and BIC 2. BSC 1 was collected from the fractions of ethyl acetate : ethanol (20:80, 40:60, 50:50). The fractions of ethyl acetate : ethanol (40:60, 50:50, 60:40) yielded BIC 2. From the FT-IR, 1H NMR, 13C NMR and LC-MS spectral studies, the isolated compounds BSC 1 and BIC 2 were identified as (2S,3S,4S,5R,6S)-6-[(5,6-dihydroxy-4-oxo-2-phenyl-4H-chromen-7-yl)oxy]-3,4,5-trihydroxytetrahydro-2H-pyran-2-carbxylic acid (baicalin, Mol. formula C21H18O11) ands 2-(3,4-dihydroxyphenyl)-5,7-dihydroxy-3-{[2S,3R,4R,5S)-3,4,5-trihydroxytetrahydro-2Hpyran-2-yl]oxy}-4H-chromen-4-one (reynoutrin, Mol. formula C20H18O11) respectively. The acute toxicity study of ethanolic extracts of whole plants of Biophytum sensitivum DC. and Biophytum intermedium Wt. revealed the nontoxic nature of the plants. Administration of ethanolic extracts of whole plants of Biophytum sensitivum DC. and Biophytum intermedium Wt. (EEBS and EEBI) to diabetic rats significantly reduced the levels of blood glucose, recovered body weight, serum profiles such as total cholesterol, HDL cholesterol, LDL cholesterol, VLDL cholesterol, triglycerides, total protein, serum creatinine, serum urea, SGOT, SGPT, antioxidant enzyme levels such as SOD liver tissue, CAT liver tissue, LPO, GPx liver tissue, organ weight such as kidney, liver, mesenteric fat pad, perirenal fat, liver cholesterol and triglyceride level compared to untreated diabetic rats in type 2 diabetes models. It was found that oral administration of extract shows nearly equal effectiveness in controlling diabetes when compared with diabetic rats treated with standard drug (Glibenclamide). Histopathological studies of kidney, pancreas and liver showed better improvement after EEBS and EEBI treated rats compared to diabetic rats. It was observed that, treatment of diabetic rats with ethanolic extracts of whole plants of Biophytum sensitivum DC. and Biophytum intermedium Wt. had not only shown a significant antidiabetic activity but also possess an effective antioxidant activity which was evident from the observations. All the diabetic rats treated with ethanolic extracts elevated the levels of antioxidant enzymes. The extracts are thus expected to possess antioxidant activity. In conclusion, ethanolic extracts of whole plants of Biophytum sensitivum DC. and Biophytum intermedium Wt. showed significant antidiabetic and antioxidant effect in diabetic rats after oral administration. It could be speculated that the observed antihyperglycemic and antioxidant activity of Biophytum sensitivum DC. and Biophytum intermedium Wt. might be due to the presence of flavonoid glycosides as active constituents