Navpreet Kaur et al. / Journal of Pharmacy Research 2012,5(11),5202-5205 Research Article Available online through ISSN: 0974-6943 http://jprsolutions.info Phytochemical and Antidiabetic Evaluation of Phaseolus trilobus Roots

Navpreet Kaur1, Rashmi1*, Y.C. Tripathi1 and Lokesh Upadhyay2 1Chemistry Division, Forest Research Institute, P.O.-New Forest-248006, Dehradun, 2Advance Research Centre for Indian System of Medicine, PRIST University, Thanjavur-613403, India Received on:12-06-2012; Revised on: 17-07-2012; Accepted on:26-08-2012

ABSTRACT The present investigation was designed to assess the phytochemical profile of Phaseolus. trilobus Ait. syn trilobata (L) Verdc (family: ) roots and antidiabetic effect of methanol extract of the roots of the on Streptozotocin induced diabetic rats. Preliminary phytochemical screening of the methanol extract revealed the presence of different types of compounds including flavonoids, steroids, terpenoids, saponin, glycoside and tannin. Acute toxicity studies of dried extract of roots at an orally administered dose of 2000 mg/kg showed neither lethality nor any reaction and behavioral change indicating the dose to be safe. Oral administration of root extract at the dose of 400 mg/kg showed a significant increase in body weight and decrease in blood glucose level on 15th and 20th day of post induction as compared to untreated diabetic rats. The significant decrease in blood glucose level in diabetic rats is found comparable to that of standard drug Glibenclamide thereby indicating the root extract of P. trilobus to be a potent antidiabetic drug and justifying its use in the traditional system of medicine.

Key words: Phaseolus trilobus, Roots, Phytochemical profile, Antidiabetic activity. INTRODUCTION Diabetes mellitus is a carbohydrate metabolism disorder of endocrine system glabrous or with a few scattered hairs. It is extensively used by tribal people due to an absolute or relative deficiency of insulin secretion, action, or both. of Nandurbar district of Maharashtra, India in the treatment of jaundice and The disorder affects more than 100 million people worldwide and by 2030 other liver disorders[9]. are considered as sedative, coolant, it is predicted to reach 366 million[1]. The most prevalent form both in the antibilious[10-11] tonic and they are used in cataplasms for weak eyes[12]. global and Indian scenario is the non-insulin dependent diabetes mellitus Fruit is aphrodisiac, astringent, styptic, anthelmintic and good for eyes. Its (NIDDM type 2) which is associated with elevated postprandial consumption cures inflammation, fever, burning sensation, thirst, piles, hyperglycemia. Regardless of considerable progress in the management of dysentery, cough, gout and biliousness[13-14]. Roots are used for curing fever, diabetes mellitus by synthetic drugs, many traditional treatments cough, diarrhea, haemorrhoids, ophthalmology and dyspepsia[15]. A wide are still used throughout the world. The plant kingdom offers a wide field to range of active components including flavonoids, isoflavonoids, tannins, look for oral antidiabetic drugs. While plant derivatives with purported stigmasterol, vitamin K, vitamin C and proteins have been isolated from the antidiabetic properties have been used in folk medicine and traditional different parts of the plant. The plant however, has not been investigated healing systems, very few of these traditional anti-diabetic plants have for antidiabetic efficacy so far. This study was thus undertaken to evaluate received proper scientific or medical scrutiny despite recommendations by the antidiabetic efficacy of methanol extract of P. trilobus roots in World Health Organization[2]. Ayurveda and other Indian traditional Streptozotocin induced diabetic albino rats. approaches have described more than 800 plants in the Indian subcontinent, known to possess antidiabetic properties. These require to be effectively MATERIALS AND METHODS studied and in fact only few of them have been characterized for their mechanistic actions [3-6]. The lead extracts have also been subjected to Plant material phytochemical analysis to determine the probable inhibitory compounds Roots of Phaseolus trilobus were collected from the forest area of Garhwal present. Search for more safer, specific, and effective antidiabetic agents region of Uttarakhand in the month of July, 2011 and authenticated by has therefore, continued to be an important area of investigation with natural Headd, Systemic Botany Section of Botany Division, Forest Research extracts from readily available traditional medicinal plants offering great Institute (FRI), Dehradun, India. Voucher specimen of the collected plant potential for discovery of new antidiabetic drugs[7-8]. In retrospection of the material has been preserved in the Chemistry Division, FRI, Dehradun. The fact, the present study was specifically aimed to evaluate the antidiabetic collected roots were dried under shade at room temperature (25°C) for 10 effect of the roots of Phaseolus trilobus in the Streptozotocin induced days and powdered to a coarse consistency in a grinder mill. The powder diabetic albino rats and compare its efficacy with the already available oral was passed through 40# mesh particle size and stored in an air tight container hypoglycemic drug Glibenclamide. at room temperature.

P. trilobus Ait. syn Vigna trilobata (L) Verdc is commonly known as Preparation of plant extract Ranmoong or Mudgparni belongs to the family Fabaceae, native to the The dried and coarsely powdered roots of P. trilobus (100 g) were extracted tropical and warm temperate regions of the world. It is a regenerating annual with 400 ml of methanol for 48 hours three times at room temperature. The or perennial herb having numerous stems 0.6-0.9 m from a woody root- methanol extract was filtered through Whatmann filter paper (No.1) at stock. Leaves are alternate and trifoliate, 3.8-7.5 cm. long, grooved, room temperature. The combined extracts were evaporated under reduced pressure using a rotary evaporator to obtain 11.35 g of dried material yielding *Corresponding author. 11.35%. The extract was then preserved in the refrigerator for the Rashmi experimental use. Chemistry Division, Forest Research Institute, P.O.-New Forest-248006, Preliminary Phytochemical Analysis Dehradun, India Preliminary qualitative phytochemical analysis of methanolic extract of P.

Journal of Pharmacy Research Vol.5 Issue 11.November 2012 5202-5205 Navpreet Kaur et al. / Journal of Pharmacy Research 2012,5(11),5202-5205 trilobus roots was carried out by employing standard protocols[16-17] for between control and treatment groups followed by Dunnet’s t-test. P values determining the presence and/or absence of phytochemicals such as alkaloids <0.05 were considered as significant. (Dragendorff’s test), flavonoids (Shinoda test), terpenoids (Salkowski test), glycosides (Keller-Kiliani,conc.H2SO4 and Molisch tests), phenols (Ellagic RESULTS AND DISCUSSION acid and Phenol tests), saponins (Foam and Haemolysis tests), steroids (Liberman-Burchard and Salkowski tests), tannins (Gelatin test). Phytochemical Screening Phytochemical screening carried out using various chemical reactions to Experimental Animals identify the presence of different bioactive compounds in the test extract Female Swiss Albino mice (20-25g) were used for toxicity study and male and the results are shown in Table 1. P. trilobus showed the presence of albino Win star rats (175-225g) for antidiabetic activity. All the rats were flavonoids, steroids, terpenoids, saponin, glycoside and tannin. While maintained under standard laboratory conditions at 25 ± 20°C, relative negative results were obtained for alkaloids, carbohydrate and amino acid. humidity 50± 5% and normal photo period. [12h dark/12h light] were used for the experiment. Animals were fed with standard rodent diet with water Table 1. Phytochemical screening of P. trilobus roots. and adlibitum. The experimental protocol has been approved by the Phytochemicals Methanol extract institutional animal ethics committee and by the regulatory body of the Alkaloids - government. Flavonoids +++ Terpenoids ++ Steroids ++ Acute oral toxicity studies Carbohydrates - The acute oral toxicity studies of methanol extract was carried out as per Glycosides + [18] Amino acids - the OECD guidelines 423 by acute toxic class method. Swiss Albino mice Saponins + of female sex weighing 20-25g were fasted for 18 h and used. In acute Tannins + toxicity studies, a dose of 2000 mg/kg of dried extract were orally administered (-) = not detected; (+) = detected. to 12 mice, additionally 3 mice were kept as a control. Then, they were observed for any sign of toxicity, morbidity or mortality during the first 24 Antidiabetic effect h with special attention during the first 4 hours and daily thereafter for a In acute toxicity study the methanolic extract of Phaseolus trilobus did not total of 14 days [19]. No mortality was observed and the behavioral pattern produce lethality up to the dose level of 2000 mg/kg. In the antidiabetic was unaffected. activity, the effects of Phaseolus trilobus root extract on body weight was measured on 5th, 15th and 20th day of post induction and were compared Induction of Diabetes with normal and diabetic control groups as shown in Table 2. Streptozotocin All the rats were fasted overnight before the administration of Streptozotocin. induced diabetic rats showed a significant decrease (P<0.05) in body weight Diabetes was induced in rats by intra peritoneal injection of Streptozotocin compared to normal rats. Oral administration of root extract at the dose of th dissolved in 0.1M sodium citrate buffer pH 4.5 at the dose of 50mg/kg 400 mg/kg showed a significant increase (P<0.05) in body weight on 15 th body weight. After the injection they had free access to food and water. The and 20 day of post induction when compared to untreated diabetic rats animals were allowed to drink 5% glucose solution overnight to overcome (Fig. 1). hypoglycaemic shock. The development of diabetes was confirmed after Table 2. Effect of Phaseolus trilobus root extract on body weight in 48 hrs of Streptozotocin injection. The animals having fasting blood glucose Streptozotocin induced diabetic rats. level more than 200mg/dl were considered as diabetic rats and used for the [20-21] Groups Body weight in gms (Mean±SEM) experimentation . Diabetic animals were grouped for five days after Post induction days induction of diabetes to study the effect of methanolic extract of P. trilobus 5th day 15th day 20th day roots in streptozotocin induced diabetic rats. Control 167.2±3.25 173±3.54 181±3.34 Diabetic control 167.2±3.25 136.8±2.10* 125.3±2.39* Experimental Design Diabetic rats+ Control 164.3±1.98 170.3±1.764* 175.8±1.47* In the experiment rats were divided into the following groups with six Diabetic rats+ glibenclamide 165.1±2.77 170.8±82.62* 178.5±2.37 animals each Values are expressed as Mean ± S.E. n=6. P*<0.05 Experimental groups were compared with diabetic control. Group I: Normal control received 1% w/v gum acacia 1ml/kg for 15 days P*<0.05 Diabetic groups were compared with control group. orally. Group II: Diabetic control received 1% w/v gum acacia 1ml/kg for 15 days orally. Group III: Diabetic rats received methanolic extract of Phaseolus trilobus root 400mg/kg body weight once a day orally for 15 days. Group IV: Diabetic rats treated with Glibenclamide 0.5mg/kg orally once a day for 15 days.

Rats were fasted overnight and the blood samples were taken from the orbital sinus of the eye on the 5th, 15th and 20th day of post induction to determine blood glucose by GOD-POD kit method. The change in body weight was observed throughout treatment period in experimental animals.

Statistical Analysis All values were expressed as Mean ± Standard error mean (SEM). The differences were compared using one way analysis of variance (ANOVA) Fig. 1: Effect of Phaseolus trilobus root extract on body weight in Streptozotocin induced diabetic rats. Journal of Pharmacy Research Vol.5 Issue 11.November 2012 5202-5205 Navpreet Kaur et al. / Journal of Pharmacy Research 2012,5(11),5202-5205 The effect Phaseolus trilobus root extract on fasting blood glucose level is may also be due to presence of similar phytoconstitutes as evident by th th th measured on 5 , 15 and 20 day of post induction and compared with phytochemical screening. normal and diabetic control groups as shown in Table 3. Streptozotocin induced rats showed a significant increase (P<0.05) in fasting blood glucose CONCLUSION level compared to normal rats. Oral administration of root extract at the Regardless of important progress in the management of diabetes using syn- dose of 400mg/kg body weight showed a significant decrease (P<0.05) in thetic drugs, many traditional plants treatments are still used throughout blood glucose level in 10 and 15 days of treatment. The fasting blood the world. However, few traditional antidiabetic plants have received proper glucose level on 15th day of post induction (10 days of treatment) was scientific validation [27]. In the present study, antidiabetic activity of roots 135.4 b ± 13.99 mg/dl compared to fasting blood glucose of diabetic control of Phaseolus trilobus was evaluated in Streptozotocin induced diabetic animal 260.2 ± 1.34mg/dl. The group treated with Glibenclamide 0.5 mg/kg rats. The study concluded with the inference that methanolic extract of P. showed fasting blood glucose level of 127.06 ± 8.07 mg/dl. On 20th day of trilobus roots have beneficial effects on blood glucose levels which adds to post induction, the root extract treated group showed a fasting blood glucose the existing knowledge on pharmacological actions of P. trilobus useful for level of 70.61 ± 2.24 mg/dl, compared to untreated diabetic animal which development of antidiabetic formulations. However, further pharmacologi- showed a fasting blood glucose level of 269.8±11.88 mg/dl. The group cal and biochemical investigations will clearly elucidate the mechanism of treated with Glibenclamide 0.5 mg/kg orally showed fasting blood glucose action and will be helpful in projecting this plant as a therapeutic target in level of 69.06 ± 1.28 mg/dl (Fig. 2). diabetes management. Table 3. Effect of Phaseolus trilobus root methanolic extract on blood sugar level in streptozotocin induced diabetic rats. ACKNOWLEDGEMENT The authors sincerely thank the Director, Forest Research Institute, Groups Blood glucose level in mg/dl (Mean±SEM) Dehradun for providing necessary facilities and support at each and every Post induction days step for completing this research work successfully. 5th day 15th day 20th day Control 62.2±1.22 61.05±1.11 60.47±1.16 REFERENCES Diabetic control 273.46±14.7 260.2±1.34* 269.8±11.88* Diabetic rats+ Control 273.10±17.04 135.4±13.99* 70.61±2.24* 1. Alberti, K.G. and Zimmet, P.Z., Definition, diagnosis and Diabetic rats+ glibenclamide 263.20±3.59 127.06±8.07* 69.06±1.28* classification of diabetes mellitus and its complications. Part 1: diagnosis and classification of diabetes mellitus. Report of a WHO Values are expressed as Mean ± S.E. n=6. Consultation, Diabetic Medicine, 15(7), 1998, 539-553. P*<0.05 Experimental groups were compared with diabetic control. P*<0.05 Diabetic groups were compared with control group. 2. 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Journal of Pharmacy Research Vol.5 Issue 11.November 2012 5202-5205