Indian Journal of Experimental Biology Vol. 43, December 2005, pp. 1161-1164

Beneficial effects of Zingiber officinale Roscoe on induced hyperlipidemia and hyperinsulinemia in rats

Sanjay V Kadnur & Ramesh K Goyal* Department of Pharmacology L. M. College of Pharmacy, Navarangpura, Ahmedabad 380 009, India Received 8 September 2004; revised 4 July 2005

Fructose supplementation produced cardinal features of Syndrome-X including significant elevations in serum cholesterol, triglyceride, and insulin and also in body weight. While treatment with methanolic extract of dried of Zingiber officinale produced a significant reduction in fructose induced elevation in lipid levels, body weight, and hyperinsulinemia, treatment with ethyl acetate extract of Z officinale did not produce any significant change in either of the last two parameters. However. it produced a significant reduction in elevated lipid levels and body weight. The concentration of 6-gingerol was found to be higher in methanolic extract and less in ethyl acetate extract. 1be results suggest that the methanolic extract of Z officinale produces better effects as compared to ethyl acetate extract in fructose induced hyperlipidemia associated with insulin resistance. The extent of activity appears to be dependent on the concentration of 6-gingerol present in the extracts. Keywords: Hyperlipidemia. Insulin resistance Zingiber officinale.

12 The prevalence of obesity is increasing worldwide. In stomachic , and tonic activities12. For the frrst time the last few decades there has been a shift in the types Akhani et al. have shown that possesses anti­ of nutrients included in the diet. The consumption of diabetic activity by virtue of 5-HT modulators present 13 fructose has increased, largely because of an in Z. officinale • increased consumption of soft drinks and many other Hyperlipidemia is generally associated with beverages that are high in fructose and also because of elevated levels of blood glucose and insulin levels. the consumption of foods such as breakfast cereals, Until now observations on beneficial effects of Z. baked goods, condiments and prepared desserts officinale in experimental animal models are studied sweetened with and high-fructose com syrupl. only for lipid profile in hyperlipidemic states 2 4 Fructose consumption induces insulin resistance , representing atherosclerosisll.1 • Studies on the effects impairs glucose tolerance3 and produces of Z. officinale in conditions associated with elevated 4 3 hyperinsulinemia , hypertriglyceredemia and glycemic and insulin levels representing dyslipidemia hypertension5 in animal models. This cluster of and syndrome X are scanty. Hence, the present study changes appears to be major risk factor in the has been undertaken to explore the possible beneficial pathogenesis of coronary artery diseasel. effects of Z. officinale on syndrome X and to correlate Zingiber officinale Roscoe, () with 6-gingerol concentrations in extracts for their commonly known as ginger is one of the commonly effects in fructose induced hyperlipidemia, used spices in India and around the world. It belongs hyperinsulinemia and hyperglycemia in rats. to Zingiberaceae family. Ginger is reported to possess 6 6 6 antiallergic , antinauseant , antiemetic , Materials and Methods 1 antihepatotoxic , anti-inflammatory8, antipyreticS, Plant material--Dried rhizomes of Zingiber 8 8 8 9 antiseptic , antitussive , hypoglycemic , antioxidant , officinale were collected locally, rhizomes identified lO ll antiplatelet , and hypolipidemic activities. It also and authenticated by Prof. O. P. Saxena, Head of the l2 12 has carminative , aromatic 12, stimulant , Department of Botany, Gujarat University. Ahmedabad, and the identification tests were done as per United States Pharmacopoeia 2000. *Correspondent author Preparation of extracts--Dried rhizomes of Z. Phone: 91-79-26302746 • Fax: 91-79-26304865 officinale Roscoe were powdered and used for e-mail: [email protected] extraction. Powder of dried rhizomes (500 g) was 1162 INDIAN J EXP BIOL, DECEMBER 2005 , ,.,. ( I ! 1 !' 1 i;.; · l!. J ' • ~ ••••

taken in a round bottom flask and extracted twice with The blood samples were allowed to clot for 30 methanol (1 L x 2) under reflux for 6 hr each time. minutes at roc,m temperature, and then centrifuged at The extracts were filtered and collected each time 5000 rpm for 15 min. Serum samples thus obtained were pooled and concentrated to dryness under were stored at -20°C until biochemical estimations reduced pressure. The yield obtained was 4.3 %. were carried out. Again, 500 g powder of dried rhizomes was taken in a Biochemical analysis-The serum parameters were round bottom flask and extracted with ethyl acetate analyzed spectrophotometrically by using double (1 L x 2) as above. The extracts were filtered, pooled beam UV -Visible spectrophotometer (Shimadzu UV­ and concentrated to dryness under reduced pressure. Visible spectrophotometer, model 1601). Estimation The yield obtained 4.6%. For feeding the extracts of serum glucose (GOD-POD method)16, cholesterol (250 mg!ml) were prepared by mixing them with (enzymatic method) 17, triglyceride (enzymatic polyethylene glycol 400 using a magnetic stirrer. method)1 8 and HDL-cholesterol (phosphotungstate Standardization of extract-Standardization of method)19 were carried out using respective diagnostic methanolic and ethyl acetate extracts for 6-gingerol kits from Bayer Diagnostic Ltd, India. Serum insulin content, a marker substance was carried out by HPLC was estimated by radioimmunoassay method using analysis using 8-methyl-n-vanillylnonamide as a the kit from Bhabha Atomic Research Center, reference standard. The analysis was carried out as Mumbai, India. In addition to the above parameters, 15 per He et a1 • daily body weight, food intake and water intake of the Experimental animals and induction of animals were recorded. VLDL-cholesterol and LDL­ hyperlipidemia-Male albino Wi star rats weighing cholesterol in serum were also calculated as per 2o 200-250 g, used for the study were fed on pellet diet Friedewald's equation . (Pranav Agro Pvt. Ltd, India) and water ad libitum. total serum triglyceridesl Hyperlipidemia was induced by supplementing their VIOL -c h 0 Ie stero I drinking water with 10% fructose. All the procedures 5 total serum tri glycerides-HDL-c were performed in accordance with the Institutional LDL -c = tota I serum c I10 Ie stero I ------=' -'------Animal Ethics Committee constituted as per the 5 directions of the Committee for the Purpose of Statistical analysis--Results were analyzed Control and Supervision of Experiments on Animals statistically using one way analysis of variance­ (CPCSEA), under Ministry of Animal Welfare (ANOV A) followed by Tukey's test. Values of P< Division. Government of India, New Delhi, India. 0.05 were considered as significant. Treatment--Rats (24) were divided into following 4 groups of 6 each: Results and Discussion The results are presented in Table I. Group I: normal diet and water ad libitum and received vehicle solvent. 6-gingerol content in the extracts-Standardization Group II: normal diet and water with 10% fructose of the methanolic and ethyl acetate extracts of Z. and received vehicle solvent (control) officinale for 6-gingerol content showed the presence Group III: normal diet and water with 10% fructose of 6-gingerol in methanolic extract (3.08% w/w) and and received methanolic extract ethyl acetate extract (1.64% w/w), respectively. (250 mg! kg) Hyperlipidemic control rats showed significant Group IV: normal diet and water with 10% fructose increase in body weight, serum cholesterol, and received ethyl acetate extract triglyceride, LDL and VLDL-cholesterol levels and (250 mg! kg). no significant change in serum HDL-cholesterol levels and significant increase in fasting serum Each group of animals were administered vehicle glucose and insulin levels as compared to normal or drugs daily for 3 weeks by gavage using oral control rats. feeding needle. Feeding of animals by fructose and In recent years the populations suffering from extracts were done simultaneous from the beginning. syndrome X are increasing. In the present world . At the end of the 3 weeks period, animals were kept approximately 20% of the world population are . for ; overnight fasting and the blood samples were suffering from syndrome X and this syndrome collected from the tail vein in the centrifuge tubes. involves various disorders into its fold namely KANDUR & GOYAL: ANTIHYPERLIPIDEMIC ACTIVITY OF ZINGIBER OFFICINALE 1163

Table l--Effects of Z officinale extracts on various parameters in fructose induced hyperlipidemia in rats. [VaJues are mean ± SE of 6 rats in each group J

Serum Normal Hyperlipidemia Methanolic Ethyl acetate parameters control control extract extract

Glucose (mg/dl) 86.5 ± 1.9 I J6.1 ± 3.4* 95.1 ± 1.7** 114.1 ± 2.5 Insulin (IlUlml) 23.6 ± 1.2 41.3 ± 0.9* 34.0 ± 1.1** 41.0 ± 0.6 Cholesterol (mg/dl) 73.0 ± 1.2 106.1 ± 1.4* 84.2 ± 1.5** 92.7 ± 1.0** Triglyceride (mg/dl) 69.7 ± 1.2 152.0 ± 3.9* 88.2 ± 2.8** 111.1 ± 2.3** HDL-Cholesterol (mg/dl) 17.5 ± 0.6 19.3 ± 0.5 17.4 ± 0.4 17.6 ± 0.4 LDL-Cholesterol (mg/dl) 41.5± 1.4 56.3 ± 1.3* 49.0 ± 1.9** 52.9 ± 0.6 VLDL-Cholesterol (mg/dl) 13.9 ± 0.2 30.3 ± 0.7* 17.6±0.5** 22.2 ± 0.4** Other related changes Body weight (g/rat) 243.3 ± 3.3 378.3 ± 3.0* 326.7 ± 2.1** 335.0 ± 2.2** Water intake (mUrat/day) 51.6 ± 1.0 52.2 ± 0.3 51.9 ± 0.5 53.3 ±0.6 Food intake (g/rat/day) 23.8 ± 0.3 25 .5 ± 0.3 24.1 ± 0.7 25.0±0.6 *significantly different from normal control **significantly different from hyperlipidemia control. (P <0.05) diabetes, coronary artery disease, atherosclerosis and Fresh juice of Z. officinale reduced elevated lipid hypertension. The hallmark of all these disorders is levels in streptozotocin induced a condition that it is associated with hyperlipidemia with or with hypoinsulinemia'3 . The present study involving a without hyperinsulinemia. Conventional drugs and condition of hyperglycemia, hyperlipidemia and allopathic drugs were evaluated for their beneficial hyperinsulinemia shows that treatment with an extract effects in such conditions. Drugs from traditional of Z. officinale is effective in reducing both elevated system of medicine are also evaluated for their lipid profile and elevated glucose levels. Methanol efficacy in such conditions. Z. officinale is one among extract of Z. offinale also prevented partially but such drugs obtained from traditional systems of significantly the elevated insulin levels in fructose fed medicine. Studies until now on Z. officinale were rats. The present results show significant reduction in evaluated for its usefulness in reducing body weight on treatment with both the extracts of Z. experimentally induced hyperlipidemia that causes officinale tested for their efficacy. atherosclerosis8.".21-22. Earlier studies showed that on treatment with Z. officinale there were significant 5-HT modulators are reported to possess significant reductions of elevated lipid profiles in conditions of beneficial effects on dyslipidemia in conditions of atherosclerosis, but there are no reports on the syndrome X26. 5-HT receptor modulators like efficacy of Z. officinale in conditions of sibutramine are reported to possess significant lipid lowering activity associated with hyperglycemia and hyperlipidemia associated with hyperglycemia and 27 2R hyperinsulinemia representing syndrome X. hyperinsulinemia - • Sibutramine l~ also reported to reduce body weight in conditions of syndrome X29. Numerous human and animal studies have Extracts of ginger are reported to possess 5-HT demonstrated myocardial alteration in conditions of modulatory effeceo. It appears the that beneficial hyperinsulinemia associated with elevated VLDL and effects obtained by ginger extracts on hyperlipidemia triglyceride levels. It is also shown that people with associated with hyperglycemia and hyperinsulit:temia hyperinsulinemia are more likely to experience 23 24 are similar to 5-HT receptor modulator sibu#-~ne. dyslipidemia - . Several other studies have also However, further experiments are required to ,~; done demonstrated that in conditions of hyperinsulinemia, in this direction. hypertriglyceredemia becomes the strongest indicator of cardiovascular disease. A strong correlation exists The results of the present study show that Z. between insulin resistance and dyslipidemia. Insulin officinale may be useful in conditions of syndrome X resistance at the adipocyte leads to release of free and associated disorders as it can reduce the elevated fatty acids and their increased flux to the liver, which hypertriglyceride, VLDL-cholesterol levels and leads to stimulation in liver for synthesis and release hyperinslinemia. This is the first study with Z. officinale, 25 of VLDL with consequent hypertriglyceredimia . which correlates its usefulness in syndrome X. 1164 INDIAN J EXP BIOL, DECEMBER 2005

Standardization of the methanolic extract and ethyl mice, J NuJr, 130 (2000) 1124. acetate extract of Z officinale for 6-gingerol content 12 Perry L M, Medicinal plants of east and southeast Asia (The MIT Press, Boston. Massachusetts), 1980, p 443. showed the presence of 3.08 % w/w and 1.64 % w/w of 13 Akhani S P, Vishwakarma S L & Goyal R K, Anti-diabetic 6-gingerol in each of them respectively. This shows activity of Zingiber officinale in streptozotocin-induced type I that higher concentration of 6-gingeral present in the diabetic rats, J Pharm Pharmacol, 56 (2004) 10 1. methanol extract as compared to ethyl acetate extract 14 Bhandari U, Sharma J N & Zafar R, The protective action of may be responsible for the better efficacy of methyl ethanolic Z officinale (Zingiber officinale) extract in extract as compared to ethyl extract. cholesterol fed rabbits, J Ethnopharmacol, 61 (1998) 161. In conclusion, the present data suggest that treatment 15 He X, Bemart M, Lian L & Lin L, High-performance liquid chromatography-electrospray mass spectrometric analysis of with methanolic extract of Zingiber officinale is better pungent constituents of ginger, J. Chromatography A. 196 in controlling the elevated glycemic levels and elevated (1998) 321. lipid profile in experimentally induced hyperlipidemia 16 Trinder P, Detennination of blood glucose using an oxidase­ representing syndrome X in animals as compared to its peroxidase system with a non-carcinogenic chromogen, J Clill ethyl acetate extract:, which correlates with the Pathoi, 22 (1%9) 158. 11 Allain C C, Poon L S, Chan C S, Richmond W & Fu P C, concentration of 6-gingerol a marker compound in Enzymatic determination of total serum cholesterol. Clin Z officinale. Chem, 20 (1914) 410. 18 Bucolo G & David M. Quantitative determination of serum Acknowledgement triglycerides by the use of , Clin Chem, 19 (1913) 416. Thanks are due to the University Grants 19 Lopes-Virella M F, Stone P, Ellis S & Colwell JA, Cholesterol Commission, New Delhi for financial assistance. determination in high density lipoproteins separated by three different methods, Clin Chem, 23 (1911) 882. References 20 Friedewald W T, Levy R I & Fredrickson D S, Estimation of Elliot S S, Keirn N L, Stem J S, Teff K & Havel P J, Fructose, low-density lipoprotein cholesterol in plasma, without use of weight gain, and the insulin resistance syndrome, Am J Clin the preparative centrifuge. Clin Chem, 18 (1972) 499. NuJr, 16 (2002) 91 J. 21 Ahmed R S & Sharma S B. Biochemical studies on combined 2 Harati M & Ani M, Vanadyl sulfate ameliorates insulin effects of (Allium sativum Linn) and ginger (Zingiber resistance and restores plasma -sulfate officinale Roscoe) in albino rats, Indian J Exp Bioi, 35 (1991) levels in fructose-fed, insulin-resistant rats, Clin Biochem, 31 841. (2004)694. 22 Tanabe M, Chen Y D. Saito K & Kano Y. Cholesterol & 3 Suzuki T Hara H, Ingestion of guar gum hydrolysate, a biosynthesis inhibitory component from Zingiber officinale, soluble and fermentable non-digestible saccharide, improves Chem Pharm Bull (Tokyo), 41 (1993) 110. glucose intolerance and prevents hypertriglyceridemia in rat'> 23 Garg A & Grundy S M, Management of dyslipidemia in fed fructose, J NuJr, 134 (2004) 1942. NIDDM, Diabetes. 13 (1990) 153. 4 Wu L Y, Juan C C, Hwang L S, Hsu Y P, Ho P H & Ho L T, & 0, Green tea supplementation ameliorates insulin resistance and 24 Howard B V, Cowman L D Go Adverse effects of diabetes on multiple cardiovascular disease risk factors in increases glucose tran!.porter IV content in a fructose-fed ral women. The Strong Heart Study, Diabetes Care. 21 (1998) model, Eur J Nwr, 43 (2004) 116. 5 Sudesh V, Linda L & Vicki G, Prevention of fructose-induced 1258. hypertension by dietary vitamins, Clin Biochem, 31 (2004) 1. 25 Ginsberg H N. Insulin resistance and cardiovascular disease, J 6 Lumb A B, Mechanism of antiemetic effect of Z officinale, Clin Invest. 103 (2000) 453. Anesthesia, 48 (1993) 1118. 26 Cheetham S C, Jackson H C, Vickers S P, Dickinson K. Jones 1 Hikino H, lGso Y, Kato N, Hamada Y, Shioiri T, Aiyama R, R B & Heal D J, Novel targets for the treatment of obesity: A ltokawa H, lGuchi F & Sankawa U, Antihepatotoxic actions of review of progress, Drug Discovery Today, 1 (2004) 221. gingerols and , J Ethnopharmacol 14 (1985) 21 James W P T, Astrup A & Finer N, Effect of sibutramine on 31. weight maintenance after weight loss: A randomized trial. 8 Mascolo N, Jain R, Jain S C & Capasso F, Umcet, 356 (2000) 2119. Ethnopharmacological investigation of Z officinale (Zil/giber 28 Finer N, James W P T & Kopelman P G. One year treatment officina/e), J £thnopharmacol, 21 (1989) 129. of obesity: A randomized, double blind, placebo controlled, 9 Reddy A C & Lokesh B R, Studies on spice principles as multicentre study of orlistal, a gastrointestinal lipase inhibitor. antioxidants in the inhibition of lipid peroxidation of rat liver Int JObes, 24 (2000) 306. microsomes, Mol Cell Biochem, III (1992) 111. 29 McLaughlin T, Abbasi F & Lamendola C, Metabolic changes 10 Srivastava K C, Aqueous extracts of , garlic and Z following sibutramine-assisted weight loss in obese officinale inhibit platelet aggregation and alter individuals: Role of plasma free fatty acids in the insulin metabolism. Biomed Biochem Acta, 43 (1984) S335. resistance of obesity, Metabolism, 50 (200 1) 819. II Fuhrman B, Rosenblat M, Hayek T, Coleman R & Aviram M. 30 Huang Q, Iwamoto M. Aoki S, Tanaka N, Tajima K & Z officinale extract consumption reduce plasma cholesterol. Yamahara J, Anti-5-hydroxytryptamine effect of inhibits LDL oxidation and attenuates development of galanolactone. diterpenoid isolated from Z officinale, Chem atherosclerosis in atherosclerotic. apolipoprotein E-deficient Pharm Bull (Tokyo). 39 (1991) 391.