Advances in Biomedicine and Pharmacy (An International Journal of Biomedicine, Natural Products and Pharmacy)

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1 ISSN: Adv. Biomed. Pharma. 2:5 (2015) Advances in Biomedicine and Pharmacy (An International Journal of Biomedicine, Natural Products and Pharmacy) Original Paper Anti-hyperlipidemic and anti-obesity activities of ethanolic extract of Trigonella foenum graecum (seeds) of Himalayan region in diet induced obese mice. Ajay Semalty 1, Rahul Kumar 1, Mona Semalty* 1 1 Department of Pharmaceutical Sciences, H.N.B. Garhwal (Central) University Srinagar Garhwal , Uttarakhand, India. *Corresponding Author: Dr. Mona Semalty. Department of Pharmaceutical Sciences, H.N.B. Garhwal (Central) University Srinagar Garhwal , Uttarakhand, India. address: monasemalty@gmail.com; Tel: Running Title: Antihyperlipidemic and antiobesity activity of Trigonella Received: 10 September, 2015; Revised: 10 October, 2015; Accepted: 22 October, 2015 Available online at Abstract Trigonella is a well-established constituent of our daily food with a variety of beneficial properties. In present study the ethanolic extract of Trigonella foenum graecum (seed) obtained from Himalayan region was prepared and evaluated for antihyperlipidemic and antiobesity activities. The extract was given orally (400 mg/kg) to the obese (high fat diet induced) mice for one month and then body weight, total lipid profile, serum creatnine and serum potassium were determined and compared with the control and standard group(atorvastatin, 400 mg/kg). At the end of treatment the standard group showed a total of % body weight gain while the extract showed % body weight gain against the obese control group (51.97 %). The extract group showed significantly lower serum, total cholesterol, triglyceride and LDL-Cholesterol and higher values of serum HDL cholesterol. The serum creatinine and serum potassium concentration were found to be in normal range at the end of the study in the standard and test group. The study suggests that the Trigonella extract might be used for antihyperlipidemic and anti-obesity activity with efficacy comparable to modern lipid lowering agent (atorvastatin). Hence, it can be used as natural and safe remedy for the prevention or delay of hyperlipidemic and cardiovascular complications of diet induced obesity. Keywords: Obesity, lipid profile, Trigonella, weight management, herbal anti-obesity. Introduction Obesity has developed as an epidemic in global population at an alarming rate. Lack of exercise, sedentary lifestyles and the consumption of energy rich diets are contributory factors towards the etiology of obesity. Currently, more than one billion adults worldwide are overweight and at least 300 million of them are clinically obese. Overweight and obesity are the fifth leading risk for global deaths. At least 2.8 million adults die each year because of being overweight or obese. In addition, 44% of the diabetes burden, 23% of the ischemic heart disease burden and 7% to 41% of certain cancer burdens are attributable to overweight and obesity [1].The obesity can cause and exaggerate many health problems both independently and in association with other diseases like diabetes, hypertension, osteoarthritis, and heart disease [2, 3]. Obesity is usually a high risk factor for cardiovascular disorders and diabetes. Therefore, the dietary supplements containing antioxidants and hypolipidemic agents may serve as a potential therapeutic approach for prevention and the management of obesity and other related complications [4]. Trigonella or Fenugreek or Greek Hayes is cultivated as a leafy vegetable, condiment and as medicinal plant. Trigonella foenum-graecum Linn. (Fabaceae). Fenugreek is reported to have anti-diabetic, anti-fertility, anticancer, anti-microbial, anti-parasitic, hypocholesterolaemic, 229

2 Advances in Biomedicine and Pharmacy Vol. 2 (5) 2015 Semalty A et al. lactation stimulant, antioxidant and hair growth promotion effects [5-11].The seeds are used for reducing the body weight in folklore. But still the anti-hyperlipidemic and anti-obesity effects are still to investigate for the plant. As lipid profile and the body weight may be an important approach to control the obesity [12-14], the present study was designed to evaluate the ethanolic extract of Trigonella foenum graecum seed for anti-hyperlipidemic and anti-obesity activities. Materials and Methods Collection and extraction of T. foengracium seed Fenugreek seeds of Himalayan region were purchased from local market of Srinagar Garhwal. The seeds were identified by a taxonomist from Department of Botany, H.N.B. Garhwal University, Srinagar Garhwal, Uttarkhand, India. All other ingredients used were of analytical grade. The seeds were dried in shade and powdered clumsily. The powdered seeds were extracted with n-hexane for removing lipids and then air dried for 5 days in shade at room temperature; the dried residues were soaked into sufficient ethanol for two days extracted again with ethanol by cold extraction method using rotary vacuum evaporator (Buchi type, 5100, Perfit India Ltd., India). The percent yield was found to be 88 % of original weight of fenugreek seeds. In vivo studies Animals Healthy young Swiss albino mice (25.08 to g) were used in the present study. The groups were kept separately in an individual polypropylene cages and given food and water ad libitum and maintained under Good Laboratory Practices (GLP) condition at a temperature of 22±2 0 C and 12/12 hour light / dark cycle and at a relative humidity of 50±15%. In vivo anti-hyperlipidemic and anti-obesity activities Mice were divided into four groups, with 6 animals in each group and kept in the cages for one week prior to dosing, for acclimatization to the laboratory conditions. Except the group I (normal control), in animals of all other groups, hyperlipidemia was induced by high fat diet (containing 21% w/w butterfat and 0.15% cholesterol containing butter) 400 mg/kg orally for one month. After one month these hyperlipidemic mice were used for study. The mice received the drug or the standard in the following manner in different groups Group I (Normal Control) mice received only normal diet (kept on normal chow diet containing 6% by weight of fat, 400 mg/kg, orally). Group II (Obese Control) mice received only High Fat Diet i.e., HFD (400 mg/kg, orally). Group III (Standard) received the reference standard drug ATORVA (400 mg/kg, orally). Group IV Fenugreek extract (400 mg/kg, orally). At the end of one month of treatment, various biochemical parameters were obtained from the blood withdrawn from cardiac puncture method. For the determination of lipid profiles, serum was separated by immediate centrifugation of blood samples using ultra cooling centrifuge (Remi, India) at 3000 rpm for 5 min, at room temperature and was directly used for estimating Total Cholesterol (TC), Triglyceride (TG), HDL- Cholesterol (HDL-C), Serum Creatinine and Serum Potassium. LDL Cholesterol (LDL-C) and VLDL (Very Low Density Lipoprotein) Cholesterol (VLDL-C) were calculated by using the following formula: LDL-C= (Total-C)-HDL-C- (TG/5) and; VLDL C= TG/5 Statistical analysis The results were analyzed with Graph Pad Instat software and expressed as mean values ± standard deviations and the significance of the difference observed was analyzed by the Student s t-test. Results The fenugreek extract was prepared by cold extraction method. The prepared extract was given orally to test animals (diet induced obese mice) for one month. At the end of treatment the mice of standard group (received HFD + atorvastatin) showed a total of % body weight gain while that of test group (received HFD + extract) showed 43.64% body weight gain against % and % weight gain in the obese control and normal control, respectively (Fig. 1). The present study revealed that total lipid profile of the HFD group mice were significantly higher than in the normal control, and the total lipid profile of HFD+ Std. and HFD+ Test groups were normalized as that of the normal control group. At the end of treatment the levels of total cholesterol increased from 68.0 to 195 mg/dl, while the levels of triglyceride increased from 63.0 to mg/dl in hyperlipidemic mice as compared to the normal control (Table 1). High total cholesterol or LDL cholesterol concentrations are high risk factors for coronary heart diseases. High fat diets significantly 230

3 Antihyperlipidemic and antiobesity activity of Trigonella Advances in Biomedicine and Pharmacy Vol. 2 (5) 2015 increase the total cholesterol levels in the serum and liver as compared with the normal control diet. The herbal extract improved the lipid profile by lowering serum total cholesterol, triglyceride and LDL cholesterol concentration as compared with the HFD groups, after treating with the ethanolic fenugreek seed extract and Standard drug (Atorvastatin). The total cholesterol concentration was significantly lower in HFD+Std. (74.32±1.20mg/dl) and HFD+Test (86.33 ± 1.50mg/dl) groups as compared with that of the obese control / HFD (195±7.93mg/dl) groups at the end of treatment. The triglyceride concentration was also significantly lower in HFD+Std. (69.30±1.40mg/dl) and HFD+TEST (73.84±1.40 mg/dl) groups as compared with that of the HFD (165.33±2.08mg/dl) groups at the end of treatment. The HDL cholesterol concentration was significantly higher in HFD+Std. (24.06±1.02 mg/dl) and HFD+Test (23.94±1.10 mg/dl) groups as compared with that of the HFD (11.66±1.15mg/dl) groups. The LDL cholesterol concentration was also significantly lower in HFD+Std. (36.40±0.10mg/dl) and HFD+Test (47.62 ± 0.12mg/dl) groups as compared with that of the HFD (150.27±4.03 mg/dl) groups. normal group (0.84±0.12) as compared to that of obese control/ HFD group. Therefore, it was concluded that the prepared herbal extract improved the HDL/ LDL ratio and was comparable to that of standard drug in maintaining the same in the normal range. Serum creatinine concentration was normal in HFD+Std. (0.69±0.05mg/dl) and HFD+Test (0.70±0.20mg/dl) groups (because that falls in the normal range i.e., mg/dl). But the HFD group showed increased serum creatinine level (3.2±0.1mg/dl). Serum potassium concentration was normal in HFD+Std. (5.26±0.15mg/dl) and HFD+TEST (5.83±0.28 mg/dl) groups (that falls in normal range i.e., mg/dl). But HFD group showed significantly higher serum potassium level (8.2±0.20mg/dl). The higher serum creatinine and serum potassium are closely associated with a group of complex pathological condition comprised of hypertension (or any cardio vascular diseases), obesity and disturbance in lipid profile. The significant lowering of this serum creatinine and serum potassium values in test groups showed that the prepared herbal formulation might be of excellent potential as anti-obesity and anti-hyperlipidemic agent. The HDL /LDL ratio was significantly higher in HFD+Std. (0.66±0.09) and HFD+Test (0.50±0.06) groups as compared with that of the HFD (0.07±0.01) groups (Fig. 2). The HDL / LDL ratio of HFD+Std., and HFD+Test were quite closer to the normal values of Table 1: Effect of fenugreek seed extract on lipid, creatinine and potassium concentrations in serum. Serum Lipid, Creatinin and Potassium levels in different groups (mg/dl) a Parameter Normal HFD HFD+Std HFD+Test Total Cholesterol 68± ± ± ±1.52 Triglyceride 63± ±2.08 b 69.30±1.40 b 73.84±1.40 b HDL-Cholesterol 25.33± ±1.15 b 24.06±1.02 b 23.94±1.10 b LDL-Cholesterol 30.07± ±0.03 b 36.40±0.10 b 47.62±0.12 b VLDL- Cholesterol 12.6± ±1.5 b 13.86±0.3 b 14.77±0.3 b HDL / LDL Ratio 0.84± ±0.01 b 0.66±0.09 b 0.50±0.06 b Serum Creatinin 0.56± ±0.15 b 0.69± ±0.20 b Serum Potassium 5.4± ± ±0.15 b 5.83±0.28 a n=6; b p<

4 Advances in Biomedicine and Pharmacy Vol. 2 (5) 2015 Semalty A et al. Figure 1: Effect of ethanolic fenugreek seed extract on body weights in high-fat diet induced obese mice against standard and control Figure 2: Effect of ethanolic fenugreek seed extract (HFD + Test) on HDL/LDL ratio in high-fat diet induced obese mice against standard (HFD+Std) and control (obese control: HFD and normal control: Normal) Discussion Several natural products like green tea, tea catechins etc. have been investigated for the anti-hyperlipidemic and anti-obesity activities [15 17]. This study investigated the anti-obesity and anti-hyperlipidemic activity of the Trigonella seeds in the high fat diet (HFD) induced obesity and hyperlipidemic mice. HFD induced obesity is the most common obesity model which mimics the obesity of humans in a very high similarity. In humans HFD induce central abdominal adiposity which is the most commonly found form of obesity and high risk factor for cardiovascular and diabetic complications. In present study the high fat diet induced hyperlipidemia and increased the 232

5 Antihyperlipidemic and antiobesity activity of Trigonella Advances in Biomedicine and Pharmacy Vol. 2 (5) 2015 body weight. The levels of total cholesterol and triglyceride were also increased significantly in hyperlipidemic mice (HFD group). The herbal extract improved lipid profile by lowering serum total cholesterol, triglyceride and LDL cholesterol concentration as compared with the HFD groups, after the treatment with Fenugreek extract and Standard drug (Atorva). Fenugreek seeds contain 45.4% dietary fiber (32% insoluble and 13.3% soluble), and the gum is composed of galactose and mannose. The latter compounds are associated with reduced glycemia and cholesterolemia [17]. At the end of treatment the extract showed only 43.64% body weight gain against that of standard group which showed a % body weight gain which indicates significant weight management effect of the extract. The significant hypolipdiemic effect of Trigonella seeds reported in this study is in agreement of the various previous studies [18-21]. But in contrast of the previous studies, the weight management effect of Trigonella of Himalayan region is more pronounced. Trigonella seeds have been shown to modulate feeding behavior in animals. The repeated administration of a fenugreek seed extract slightly but significantly decreased dietary fat consumption in healthy overweight subjects in a short-term study [22]. It has been reported that Fenugreek dose-dependently reduced the hepatic triglyceride and total cholesterol levels. Trigonella foenum-graecum (fenugreek) can ameliorate dyslipidemia, but the detailed mechanism is unclear. Recently, a study concluded that Fenugreek also dose-dependently increased the excretion of cholesterol and total bile acids into the faeces. It was suggested that fenugreek inhibited lipid accumulation in the liver by increasing the lipid excretion in the faeces [23].In another study it was shown that fenugreek ameliorates dyslipidemia by decreasing the hepatic lipid content in obese diabetic mice and that its effect is mediated by diosgenin, the main aglycon of fenugreek [24]. Conclusion Ethanolic extract of Trigonella seeds (Himalayan region) was almost equally efficacious as that of standard in terms of lowering the levels of total cholesterol, triglyceride, LDL cholesterol and improving the HDL/LDL ratio in short term treatment of diet induced obese mice. The present study depicts that Fenugreek seed extract could be used as a potential anti-hyperlipidemic and anti-obesity therapeutic agent in diet induced hyperlipidemia and obesity. Conflict of interest The authors declare that there is no conflict of interest to reveal. References [1] WHO, Obesity and overweight, Available at Accessed on February 06, [2] Yun JW., Possible anti-obesity therapeutics from nature A review, Phytochemistry, 71: , [3] Athyros VG, Tziomalos KA, Karagiannis and Mikhailidis DP., Dyslipidaemia of obesity, metabolic syndrome and type 2 diabetes mellitus: the case for residual risk reduction after statin treatment, Open Cardio. Med. J., 5:34-39, [4] Khairunnuur FA, Zulkhairi A, Hairuszah I, et al., Hypolipemic and weight reducing properties from Tamarindusindica L. pulp extract in diet-induced obese rats, Int. J. Pharmacol., 6: , [5] Al-Habori M and Rama A., Pharmacological properties, In: Petropoulos GA (eds). Fenugreek - the Genus Trigonella. Taylor and Francis: London and New York, pp , [6] Broca C, Breil V, Cruciani-Guglielmacci C, et al., The insulin trophic agent 1D1101(4-hydroxyisoleucine) activates insulin signalling in rat, Am. J. Physiol. Endocrinol. Metab., 287: E463-E471, [7] Acharya S, Srichamroen A, Basu S, Ooraikul B and Basu T., Improvement in the nutraceutical properties of fenugreek, Songklanakarin J. Sci. Technol., 28: 1-9, [8] Dixit P, Ghaskadbi S, Mohan H and Devasagayam TP., Antioxidant properties of germinated fenugreek seeds, Phytother. Res., 19: , [9] Semalty M, Semalty A, Joshi GP and Rawat MSM., Comparison of in vitro antioxidant activity of Trigonellafoenumgraecum and T. corniculata seeds, Res. J. Phytochem., 3: 63-67,

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