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Antidiabetic activity of ethanolic extract of Nardostachys jatamansi on Alloxan induced diabetic rats.

Article in INTERNATIONAL JOURNAL OF ADVANCES IN PHARMACEUTICAL RESEARCH · June 2011

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Research Paper IJAPR ISSN: 2230 – 7583 Available Online through www.ijapronline.org

ANTIDIABETIC ACTIVITY OF ETHANOLIC EXTRACT OF NARDOSTACHYS JATAMANSI ON ALLOXAN -INDUCED DIABETIC RATS

S. Nelson Kumar *, K. Ravindrareddy, Rupesh.S.Kanhere, J.Yasodha Krishna, C.Raja Ram, K. Mahesh Kumar

Department of Pharmacology, P. Rami Reddy Memorial College of Pharmacy, Kadapa, Andhra Pradesh, India

Received on 20 – 04 - 2011 Revised on 21 – 05- 2011 Accepted on 01 – 06 – 2011 ABSTRACT mellitus is the most common endocrine disorder that impairs homeostasis resulting in severe diabetic complications including retinopathy, angiopathy, nephropathy, and causing neurological disorders due to perturbation in utilization of glucose. The present study was designed to investigate the hypoglycemic properties of ethanolic extract of Nardostachys jatamansi (N. jatamansi) which is widely used as a traditional treatment for diabetes mellitus. The present study was carried out to evaluate the antidiabetic activity of Nardostachys jatamansi ethanolic rhizome extract in alloxan induced diabetic rats for 7 days. The ethanolic extract at high dose (1200 mg/kg) exhibited significant antihyperglycemic activity than at low dose (400 mg/kg) in diabetic rats. The results showed that it has significant antihyperglycemic effect in experimental model of diabetes mellitus.

Keywords: Diabetes, Hypoglycemic effect, Ethanolic extract of Nardostachys jatamansi, Alloxan.

INTRODUCTION

Nature has endowed India with a unique The India Council Medical Research (ICMR) has gift of as many as 15,000 plant varieties. India been doing research work with much appreciable accounts for two third of the flowering plants of the interest on herbal antidiabetic drugs. In spite of the world of which about 2500 are reported to possess tremendous advancements made in the modern medicinal and curative qualities1. About 70-80% of system of medicine for the management of diabetes the populations depend on herbal remedies either mellitus, till date an ideal or suitable drug is not partly or entirely. The export potential of herbal available. drugs has been estimated to be Rs. 2,840 million/annum for India and Rs.16, 000 million for Diabetes is a metabolic disorder characterized by China2. symptoms like hyperglycemia, altered metabolism of lipids, carbohydrates and proteins. It also causes an For Correspondence: * increased risk of complications from vascular diseases3. National Urban Diabetes Survey (NUDS) S.Nelson Kumar reveals that out of every eight Indians (above 20 Asst.Professor, years of age) living in urban metros is diabetic and Department of pharmacology, may be one in four either diabetes or potential P.Rami Reddy Memorial College of Pharmacy, candidate for diabetes inducement. The prevalence of Kadapa, Andhra Pradesh, India, diabetes in adults in found to be 13.2%. India has the Mail I.D: [email protected] highest diabetic population with an estimated 32 Mobile No: 09866086072. million people suffering from it. It is estimated that by 2005, India will have more than one fifth of the world diabetes population. The global diabetic population is expected to double to 300 million in IJAPR / June 2011/ Vol. 2 / Issue. 6 / 263 - 268 263 S. Nelson Kumar., et al. / International Journal of Advances in Pharmaceutical Research

2005 from 136 million in 1995. The increasing The plant was collected from Sri Venkateswra incidence in India during the period will be three fold Ayurvredic stores, Vijayawada, Krishna District, from 19.4 million to 57.2 million4. The ancient Andhrapradesh, India. Which was Authentified by Ayurvedic drugs have mentioned about the DR.K.Madhava Chetty, Asst.Professor of Botany, permanent cure of diabetes mellitus through S.V.University, Tirupathi. restoration of to normal functioning5. Preparation of the Extract Though this theory appears to be very sound as yet The Nardostachys jatamansi was collected, cleaned, no such drug has been clinically investigated so far dried and powdered in a grinder –mixer to obtain a which can completely bring back pancreas to normal coarse powder and then passed through 40 mesh functioning in diabetes. The therapeutic objective of sieve. About 500 gm of powdered drug was extracted the modern system of medicine is to provide successively with hexane, ethyl acetate, 80% aqueous metabolic relief by bringing down the blood glucose ethanol and water by soxhlet apparatus. The level to normal. This helps in the prevention, extraction was carried out until the drug became postponement and amelioration of the complications exhausted. The solvents were recovered from their of diabetes mellitus and also in improving the quality extract by distillation under reduced pressure .The of life. dried extract thus obtained was kept in a desiccator and was used for further experiments was well as Nardostachys jatamansi (N. jatamansi) used for identifying their chemical groups present (Valirenaceae) is indigenous to the Himalayan .The plant profile is mentioned in the table no: 1 regions of India. In Ayurveda, roots and rhizomes of N. jatamansi are used to treat hysteria, epilepsy, and Experimental Animals convulsions6. The decoction of the drug is also used in neurological disorders, insomnia and disorders of Male albino rats of (180-210 g) were used throughout cardiovascular system7. The sesquiterpenes the experiments. The animals were procured from Sri (Jatamansic acid, Jatamansone), lignans and Venkateshwara Enterprises, Bangalore. Before neolignans are reported to be present in the roots of initiation of the experiment, the rats were this plant8, 9. Rhizomes are reported to contain a acclimatized for a period of 7 days. Standard terpenoid ester, Nardostachysin I10. To date much environmental conditions such as temperature (26 + research has been undertaken to evaluate the drug to 20C), relative humidity (45-55%) and 12hrs treat various neurological and cardiovascular dark/light cycle were maintained in the quarantine. disorders in various animal models and is widely All the animals were fed with rodent pellet diet (Gold used in ayurvedic formulations. It is reported to mohr, Lipton India Ltd.) and water was allowed ad- possess antidepressant activity,11 anticonvulsant libitum under strict hygienic conditions. Ethical activity 12, antiarrhythmic activity 13 , and possess clearance for performing the experiments on animals antioxidant, and lipid peroxidation activity in was obtained from Institutional Animal Ethics doxorubicin- induced cardiac damage in rats 14. It is Committee (IAEC). also reported to improve learning and memory in Preliminary phytochemical screening mice15 and to enhance biogenic amine activity16. An Preliminary phytochemical screening were done to acetone extract of N. jatamansi has shown significant find the presence of the active chemical constituents inhibition of benzoyl peroxide-induced cutaneous such as alkaloids, proteins, tannins, terpins, lignin’s, oxidative stress, toxicity, and ear edema in mice17. It volatile oils and fats 19,20. has also been reported to possess protective activity Acute toxicity studies in 6-hydroxydopamine-induced, parkinsonism in rats Acute toxicity studies for aqueous ethanolic 18. extract of Nardostachys jatamansi was carried out in rats at different doses (1000 - 2000 mg/kg, orally), MATERIALS AND METHODS showed no gross evidence of any abnormalities in the mice up to the end of 72 hr of the observation period. Drugs and Chemicals: Ethanol (S.D. Fine chemicals This indicates the safety of extract. Further, it is Ltd, Mumbai), Sodium hydroxide (S.D. Fine reported 21 that ethanolic extract of Nardostachys chemicals Ltd, Mumbai), Sodium hydroxyl methyl jatamansi is found to be safe at a dose of 3000 cellulose (S.D. Fine chemicals Ltd, Mumbai), mg/kg. Hence, further pharmacological investigation Alloxan (sigma chemicals Co., U.S.A), was carried at dose levels of 400, 800, 1200 mg/kg. Glibenclamide (Aristo pharma, Mumbai), Glucose Acute toxicity study was done as per OECD, 2006 strips (Abbott laboratories. U.S.A). Guidelines (0ECD, 2006 Guidelines). Plant collection and Authentication Induction of diabetes

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Alloxan (2, 4, 5, 6-tetraoxypyrimidine; 2, 4, 5, 6- ANOVA and post hoc Dunnet’s t test. Differences tetrone) is an oxygenated pyrimidine between groups were considered significant at P < derivative 22 and was originally isolated in 1818 by 0.05 levels. Brugnatelli and got its name in 1838 by Friedrich Wohler and 23. Alloxan is a toxic RESULTS glucose analogue, which selectively destroys - The qualitative phytochemical analysis of aqueous producing cells in the pancreas when administered to ethanolic extract of Nardostachys jatamansi reveals rodents and many other animal species. This causes that the extract is rich in Alkaloids, proteins, tannins, an insulin-dependent diabetes mellitus (called terpins, lignin’s, volatile oils and fats (Table No: 2) "Alloxan Diabetes") in these animals, with Effect of Nardostachys jatamansi rhizome extract characteristics similar to type 1 diabetes in humans. on blood glucose level in diabetic rats (Table 3) Alloxan monohydrate was obtained from S.D. Fine, Ethanolic extract of Nardostachys jatamansi rhizome Mumbai and all the other chemicals used were of was subjected to anti-diabetic activity in rats where analytical grade and were acquired from commercial alloxan monohydrate (150 mg/kg b.w., p.o) was used sources. as the diabetogenic agent. A marked rise in fasting Anti-diabetic activity blood glucose level observed in diabetic control Fasting blood glucose was determined after depriving compared to normal control rats. Ethanolic extract of food for 16 hrs with free access of drinking water. Nardostachys jatamansi (at 400, 800, 1200 mg/kg) Hyperglycemia was induced by a single i.p. injection exhibited a dose dependent significant anti- of 150 mg/kg of alloxan monohydrate (s.d. fine- hyperglycemic activity on 1st, 2 nd, 3rd, 4th, 5th, 6th, and chem. Ltd., Mumbai, India) in sterile saline. After 3 7th day post treatment. The extract dose of 400 days of alloxan injection, the hyperglycemic rats mg/kg also caused less reduction in blood glucose (glucose level > 250 mg/dl) were separated and level but the results were found statistically divided into different groups comprising of 6 rats insignificant. The antihyperglycemic effect t of each for the anti-diabetic study. The treatment (p.o.) ethanol extract was found less effective than the was started from the same day except normal control reference standard, Glibenclamide. Glibenclamide and diabetic control groups for a period of 7 days. produced a significant reduction in blood glucose During this period, animals in all groups had free compare to diabetic control. The results are shown in access to standard diet and water. Body weight and the Table No. 3. blood glucose levels were estimated on 1st, 2 nd,3rd, Effect of Nardostachys jatamansi rhizome extract 4th,5th, 6th, and 7th day of the treatment. On the 10th on body weight in diabetic rats day, blood samples were collected from overnight Normal control animals were found to be stable in fasted rats by cardiac puncture under mild ether their body weight but diabetic rats showed significant anesthesia for biochemical estimations. reduction in body weight during 7 days. Alloxan The various groups used in experiment mediated body weight reduction was significantly Group 1 - Served as normal control and received 1% reversed by the ethanolic extract in dose dependant SCMC 2ml/kg per orally. fashion (at 400, 800, and 1200 mg/kg). Results are Group 2 – Alloxan 150 mg/kg and received 1% shown in Table No. 4 SCMC 2ml/kg per orally Group 3 - Alloxan 150 mg/kg + Glibenclamide (10 DISCUSSION mg/kg, p.o.). Pancreas is the primary organ involved in sensing the Group 4 - Alloxan 150 mg/kg + AEENJ (400 mg/kg, organism’s dietary and energetic states via glucose p.o.) concentration in the blood and in response to elevated Group 5 - Alloxan 150 mg/kg + AEENJ (800 mg/kg, blood glucose, insulin is secreted 24. Alloxan is one of p.o.) the usual substances used for the induction of Group 6 - Alloxan 150 mg/kg + AEENJ (1200 diabetes mellitus apart from . Alloxan mg/kg, p.o.) has a destructive effect on the beta cells of the pancreas 25, 26. Alloxan causes a massive reduction in AEENJ—Aqueous ethanolic extract of Nardostachys insulin release by the destruction of β-cells of the jatamansi islets of langerhans, thereby inducing SCMC--- Sodium carboxy methyl cellulose hyperglycaemia 27. Insulin deficiency leads to various metabolic alterations in the animals viz increased Statistical Analysis blood glucose, increased cholesterol, increased levels All the values of body weight, fasting blood sugar, of alkaline phosphate and transaminases 28, 29. and biochemical estimations were expressed as mean ± standard error of mean (S.E.M.) and analyzed for

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The results of the present study indicate that diabetic activity. The improvement of function Nardostachys jatamansi rhizome extract was found and subsequent increase in uptake of blood glucose to reduce the glucose level in animals made diabetic and its utilization may be another mechanism of with alloxan. Alloxan has been shown to induce free action of the extract. production and cause tissue injury. The pancreas is especially susceptible to the action of ACKNOWLEDGEMENT alloxan induced free radical damage. In the present We are thankful to Management of P. Rami Reddy investigation ethanolic extract of Nardostachys memorial College of Pharmacy, Kadapa, A.P for the jatamansi rhizome demonstrated the significant anti- providing the all facilities for carriing out this work.

Table No.1, The Nardostachys jatamansi plant scientific profile Kingdom Plantae

Division Magnoliophyta

Class Magnoliopsida Order Dipsacales Family Valerianaceae Genus Nardostachys

Species Jatamansi.

Botanical name Nardostachys Jatamansi.

Table No.2, Qualitative chemical tests for phytoconstituents in Nardostachys Jatamansi Linn

Tests Aqueous ethanolic extract of Nardostachys Jatamansi Alkaloids + Carbohydrates + Flavonoids -- Triterpenoids + Protiens + Resins -- Sapoinins -- Steroids + Tannins + volatile oils + lignin’s +

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Table No: 03, Effect of Nardostachys jatamansi rhizome extract on blood glucose level in diabetic rats

GROUPS BLOOD GLUCOSE LEVEL 1st Day 2NDDay 3rdDay 4thDay 5thDay 6thDay 7thDay Group I (Control) 79.16±0.43 81.41±0.18 80.60±0.25 80.60±0.24 81.50±0.17 81.08±0.11 81.30±0.19 Group I (Alloxan) 203.83±0.3 209.83±0.4 219.60±0.7 225.16±0.5 234.50±0.7 240.00±0.5 253.00±0.80 7 1 9 7 2 2 Group III (Alloxan 206.5±0.37 184.5±0.49 166.0±0.50 144.83±0.6 122.50±1.0 99.50±1.11 85.60±0.66 +Glibenclamide) * * 5 2 3 Group IV (Alloxan+AEENJ4 208.00±1.4 197.3±0.54 187.3±1.04 178.5±0.65 168.83±0.8 157.30±0.7 148.5±1.02* 00 mg/kg) 3 * * * * * Group V (Alloxan+AEENJ8 208.30±0.5 196.0±0.89 175.0±0.39 154.00±0.4 136.83±0.7 119.6±0.53 102.0 00 mg/kg) 4 * * * * * ±1.12* Group IV (Alloxan+AEENJ1 208.16±0.4 189.0±0.48 168.0±0.75 148.0±0.73 128.6±0.72 115.0±0.63 96.3±1.08* 200 mg/kg) 8 * * * * * Values are Mean S.E.M; n=6; Statistical significant test for comparison was done by ANOVA, followed by Dunnets test- “t” Test*P < 0.05 vs Diabetic Control

Table No: 4, Effect of Nardostachys jatamansi rhizome extract on body weight in diabetic rats

GROUPS Body weight of the animal (g) 1st Day 2NDDay 3rdDay 4thDay 5thDay 6thDay 7thDay Group I (Control) 201.3±2.34 202.2±2.62 204.4±2.73 206.2±2.84 208.2±2.86 209.1±2.88 211.4±2.95 Group I (Alloxan) 203.83±2.3 195.4±2.23 189.60±2.1 183.57±2.1 175.50±2.0 170.47±2.0 161.0±2.98 6 9 4 9 5 Group III (Alloxan 205.5±2.37 203.5±2.29 199.4±2.25 195.2±2.21 191.1±2.18 187.1±2.09 184.0±1.94 +Glibenclamide) * * * * Group IV (Alloxan+AEENJ4 206.00±1.4 202.3±2.23 197.4±1.97 193.3±1.94 187.2±1.89 180.3±1.85 173.5±1.79* 00 mg/kg) 3 * * * * *

Group V (Alloxan+AEENJ8 205.30±1.9 201.2±1.86 198.6±1.83 195.5±1.80 191.3±1.77 187.4±1.75 181.4±1.69* 00 mg/kg) 4 * * * * *

Group IV (Alloxan+AEENJ1 206.16±.18 202.4±1.87 198.5±1.79 192.3±1.74 189.1±1.70 186.3±1.67 184.3±1.66* 200 mg/kg) 8 * * * * * Values are Mean S.E.M; n=6 *P < 0.05 vs Diabetic Control

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