EVALUATION OF ANTIOBESITY EFFECT OF GYMNEMA SYLVESTRE AND ACORUS CALAMUS ON DIET INDUCED MODEL OF OBESITY IN RATS

EVALUATION OF ANTIOBESITY EFFECT OF GYMNEMA SYLVESTRE AND ACORUS CALAMUS ON DIET INDUCED MODEL OF OBESITY IN RATS

Dheeraj Kumar Singh^*1, Anjula Sachan ², Narendra Kumar ¹, Preet Lakhani ¹, Sachin Tutu ¹, Pratap Shankar ¹, R. Nath ¹, Amod Kumar ¹ and R. K. Dixit ¹

Department of Pharmacology ¹, King George’s Medical University, Lucknow - 226003, Uttar Pradesh, India.

Department of Pharmacology ², Hind Institute of Medical Sciences, Mau Ataria, Sitapur Road, Lucknow - 261303, Uttar Pradesh, India.

ABSTRACT: Introduction: Obesity is a morbid condition characterized by abnormal or excess deposition of body fat to an extent that it adversely affects health. The basic etiology behind obesity is the disparity between the energy intake and energy expenditure. Lifestyle modifications and drug treatment is the mainstay of treatment. Unfortunately the benefits are for short term that might be because of cessation of drugs use or side effects which ultimately leads to rebound weight gain. Methods: Experiment was conducted on 42 adult healthy female wistar rats weighing around 150-200g, divided into 7 groups with 6 rats in each group. Group 1 given normal chow while others were given High Fat Diet (HFD). From 5^th week till the end of 8^th week group 3 to 7 were given respective drugs (extracts of Gymnema sylvestre and Acorus calamus). Obesity parameters (body weight, BMI, locomotor activity) were recorded on day 0, at the end of 8^th week and at the end of 8^th week. Internal organs (kidney, liver, spleen and heart) and fat pads (perirenal, mesenteric and gonadal) were also removed and weighed. Results: Feeding rats on HFD significantly changed the obesity parameters and lead to development of obesity. Combination of GS and AC significantly reduced body weight, BMI and improved locomotor activity. It also significantly reduced the weight of internal organs and fat pads. Both GS and AC also improved the obesity parameters in a dose dependent manner. Conclusions: Extracts of Gymnema sylvestre and Acorus calamus has a potent antiobesity effect. Combination of both is superior to the individual drugs.

Body mass index, Extract, Fat

INTRODUCTION: Obesity is a morbid condition characterized by abnormal or excess deposition of body fat to an extent that it adversely affects health¹.

It is a multifactorial disease, with its causative agents ranging from genetic, metabolic, social, behavioural, and cultural factors ². In the last few decades the prevalence has increased to such an extent that can be attributed to psychological and behavioural factors instead of biological factors only. Overall in 2014, about 13% of the world’s adult population was obese and 39% adults were overweight. Obesity affects the health adversely and can lead to variety of other co-morbid conditions like insulin resistance, type II diabetes mellitus, hypertension, dyslipidaemia, metabolic syndrome, coronary heart disease and even some cancers ³. The basic etiology behind obesity is the disparity between the energy intake and energy expenditure ⁴. The type of food that we eat plays a major role in disturbing the energy balance. Increase in energy density of foods, better palatability of food, portion size, and easy availability as well as low cost promote the development of overweight and obesity ⁵. Decreased physical activity and sedentary lifestyle contributes equally in the occurrence of obesity ⁶.

Lifestyle modification is considered as the initial modality to control weight which includes increased physical activity, dietary and behavioural modifications ⁷. However to continue these modification for long is itself a challenge. Pharmacotherapy is the other way to control the obesity but these are indicated in some particular conditions. Currently available drugs have their own side effects. Unfortunately the benefits are for short term that might be because of cessation of drugs use or side effects which ultimately leads to rebound weight gain ⁸. Gymnema sylvestre is an indigenous herb widely distributed in India, tropical Africa, Australia, Japan, Indonesia, Vietnam, and some regions of China. The plant is known to be a good source of number of bioactive molecules ⁹. Medicinal uses of this plant are well known since ages. Traditionally it has been found useful in constipation, dyspepsia, jaundice, renal calculi, haemorrhoids, asthma, bronchitis, conjunctivitis and number of other conditions ¹⁰.

Acorus calamus is a well known drug in traditional system of medicine and rhizomes of the plants are being used since ages ¹¹. In various studies it has shown antidiabetic activity, antiinflammatory and many other properties ^{12, 13}. Present study was designed to explore and compare the antiobesity effect of Gymnema sylvestre and Acorus calamus individually as well as in combination.

MATERIAL AND METHODS: Present work was started after getting ethical approval from Institutional Animal Ethics Committee of King George’s Medical University.

Experimental Procedure: Experiment was conducted on 42 adult healthy female wistar rats weighing around 150-200g that were procured from Indian Institute of Toxicology Research, Lucknow. Animals were kept under standard condition of temperature (25 ± 2 °C), humidity (55 ± 5%) and twelve hour light- dark cycle at the institutional animal house of King George’s Medical University. All the guidelines of CPCSEA were followed while conducting experiment. After an acclimatization period of 7 days animals were divided into 7 groups with 6 rats in each group. During this period all the animals were given normal chow and water ad libitum.

On day 0 all the parameters of obesity including body weight, Body mass index (BMI) and locomotor activity (ambulatory activity and rearing) was measured. During phase I of the study which lasted till 4 weeks obesity was induced in group 2 to 6 by feeding them on high fat diet (HFD) while group 1 was given normal chow. HFD was purchased from Bharat Science Solution Company, Lok Nagar, Unnao, Uttar Pradesh. Nutritional composition of HFD included carbohydrate 44%, crude fat 25%, crude protein 18%, fiber 13%, moisture 8 %, Vitamins, minerals and other ingredients in required quantity. All the parameters were measured again at the end of 4^th week.

From 5^th week phase II of the study commenced and lasted till the end of 8^th week. During this phase group 1 was continued on normal chow while group 2 was continued with HFD. Group 3 to 6 were kept on HFD along with the respective drugs. Group 3 was given Gymnema sylvestre 100mg/kg and group 4 was given Gymnema sylvestre 200mg/kg. Group 5 was treated with Acorus calamus 100mg/kg while group 6 was given Acorus calamus 200mg/kg. All the drugs were given by oral route as a suspension using distilled water. At the end of 8^th week all the parameters were measured. After that all the rats were sacrificed with high dose of anaesthesia. Their internal organs (kidney, liver, spleen and heart) and fat pad (perirenal, mesenteric and gonadal) were identified, removed and weighed.

Test Drugs:

Gymnema sylvestre: Dried powdered extract of leaves of Gymnema sylvestre was obtained from Ekgaon Company, New Delhi.

The drug was administered in a dose of 100 and 200 mg/kg bw ¹⁴.

Acorus calamus: Dried powdered extract of rhizomes of Acorus calamus was obtained from Sierra India Organics, Indore. It was administered orally in a dose of 100 and 200 mg/kg bw ¹⁵.

Measurement of Body Weight and Body Mass Index (BMI): Body weight was measured by digital weighing machine. BMI was calculated by using the following formula:

BMI = Body weight (kg) / (Naso-anal length in meter) ²

Measurement of Locomotor Activity (Ambulatory Activity and Rearing): Locomotor activity (ambulatory activity and the frequency of rearing) was calculated by using open field behaviour test apparatus (68×68×45 cm). Rats were kept in the apparatus for a period of 5 minutes and their ambulatory activity (AA) and the frequency of rearing was recorded.

Removal of Different Organs and Fat Pads: Rats were sacrificed by using high dose of anaesthesia (sodium pentobarbitone 150mg/kg i.p.). Different organs (kidney, liver, spleen and heart) and fat pads (perirenal, mesenteric and gonadal) were identified, removed and weighed.

Statistical Analysis: Statistical analysis was done using SPSS Statistics 20 software (Armonk, NY: IBM Corp.). Data were expressed as mean ± standard error of mean (S.E.M.). Different groups were compared with one way analysis of variance (ANOVA) followed by Dunnett’s t-test while to compare similar group at different time intervals, paired t test was used. A p-value < 0.05 was considered statistically significant.

RESULTS: At day 0 all the groups were compared by analysis of variance (ANOVA). All the groups were found comparable to each other for obesity parameters (body weight, BMI and Locomotor activity) as no significant difference was found (Table 1).

TABLE 1: COMPARISON OF ALL THE GROUPS AT DAY 0

Effect on Body Weight and BMI: HFD deleteriously affected the obesity parameters. Body weight and BMI of the groups on HFD increased significantly at week 4 from their previous values. After starting drug treatment, body weight and BMI reduced significantly in all the drug groups. AC at 100mg/kg caused 24.0% weight reduction while at 200mg/kg 32.8% reduction as compared to disease control group. GS 100mg/kg decreased body weight by 32.9% while at 200mg/kg it reduced weight by 38.1%. Combined GS 200mg/kg and AC 200mg/kg caused maximum reduction in body weight by 42.0%. Regarding BMI, AC 100mg/kg and 200mg/kg caused 32.6% and 37.9% reduction while GS 100mg/kg and 200mg/kg caused 34.7% and 42.1% decrease in BMI respectively. Combined dose of GS 200mg/kg and AC 200mg/kg caused maximum reduction by 49.5% (Table 2 and 3).

TABLE 2: PRE AND POST TREATMENT BODY WEIGHT (g) OF ALL THE GROUPS (MEAN ± SEM, n = 6)

^*Significant as compared to day 0; ^#Significant as compared to week 4; ^$Significant as compared to day 0.

At the end of 8^th week ANOVA revealed significant difference (p-value < 0.01) among the groups. Dunnett’s post hoc test showed that all the drug groups differed significantly from the disease control group 2 for both body weight and BMI (Table 4).

TABLE 3: PRE AND POST TREATMENT BODY BMI OF ALL THE GROUPS (MEAN ± SEM, n = 6)

^*Significant as compared to day 0; ^#Significant as compared to week 4; ^$Significant as compared to day 0.

TABLE 4: ANOVA FOLLOWED BY DUNNETT’S POST HOC TEST (COMPARISON GROUPS- 2) AT THE END OF 8^th WEEK FOR BODY WEIGHT AND BMI

*Significant.

Effect on Locomotor Activity: Feeding rats with HFD lead to significant decrease in ambulatory activity and increase in frequency of rearing suggesting development of obesity by 4^th week. Later on after giving drugs both the parameters improved i.e. AA increased and rearing decrease (Table 5 and 6). AC 100mg/kg increased AA by 22.7% and decreased rearing by 23.8%.

Similarly AC 200mg/kg increased AA by 34.8% and decreased rearing by 19.4%. GS 100mg/kg increased AA by 38.3% and decreased rearing by 22.6%. At 200mg/kg GS improved AA by 46.5% and reduced rearing by 26.2%. Combined dose of GS and AC was found more effective and increased AA by 51.5% and decreased rearing by 36.5%.

TABLE 5: PRE AND POST TREATMENT AMBULATORY ACTIVITY OF ALL THE GROUPS (MEAN ± SEM, n=6)

^*Significant as compared to day 0; ^#Significant as compared to week 4; ^$Significant as compared to day 0.

TABLE 6: PRE AND POST TREATMENT VALUE OF REARING OF ALL THE GROUPS (MEAN ± SEM, n = 6)

^*Significant as compared to day 0; ^#Significant as compared to week 4; ^$Significant as compared to day 0.

All the groups were compared by ANOVA at the end of 8^th week. Significant difference was found among the groups (p-value < 0.01). Post hoc test, considering group 2 as comparison group, revealed that both the drugs significantly improved obesity parameter by increasing AA and decreasing rearing (Table 7).

TABLE 7: ANOVA FOLLOWED BY DUNNETT’S POST HOC TEST (COMPARISON GROUPS- 2) AT THE END OF 8^th WEEK FOR AMBULATORY ACTIVITY AND REARING

*Significant.

At the end of 8^th week all the parameters were recorded again and then rats were sacrificed by high dose of anaesthesia. Their internal organs (kidney, liver, spleen and heart) and fats pads (perirenal, gonadal and mesenteric) were removed and then weighed for comparison. ANOVA revealed that there was significant difference between the groups regarding these parameters (Table 8).

TABLE 8: ANOVA AT THE END OF 8^th WEEK FOR INTERNAL ORGANS

TABLE 9: ANOVA FOLLOWED BY DUNNETT’S POST HOC TEST (COMPARISON GROUPS-2) AT THE END OF 8^th WEEK FOR INTERNAL ORGANS

*Significant.

TABLE 10: ANOVA FOLLOWED BY DUNNETT’S POST HOC TEST (COMPARISON GROUPS-2) AT THE END OF 8^th WEEK FOR INTERNAL FAT PADS

*Significant.

Intergroup comparison was made by ANOVA followed by dunnett’s post hoc test with group 2 as comparison group. As shown in Table 9 weight of internal organs reduced significantly (p-value <0.01) by both the drugs as compared to the disease control group.

Treatment by both the drugs significantly (p-value < 0.01) reduced the weight of internal fat pads (perirenal, gonadal and mesenteric) as compared to disease control group suggesting the antiobesity and weight reducing properties of GS and AC (Table 10).

DISCUSSION AND CONCLUSION: Obesity is a condition characterized by abnormal or excess deposition of body fat to an extent that it adversely affects health. The basic cause behind obesity is the disparity between the energy intake and energy expenditure. The type of food that we eat plays a major role in maintaining the energy balance. The most widely employed method to assess overweight and obesity is by the calculation of body mass index (BMI).

In the present study obesity was induced by feeding rats with high fat diet. The parameters which were used to assess obesity were body weight, body mass index, ambulatory activity, frequency of rearing, weight of internal organs and internal fat pads.  HFD for 4 weeks led to significant increase (p-value < 0.01) in the body weight, BMI and ambulatory activity along with significant reduction in frequency of rearing. This study supports the fact that although there are various models of obesity but HFD induced obesity can act as a rapid method that even lasts for long ^{16, 17}.

After 4 weeks drug treatment was started along with HFD in groups 3-7 while group 1 was continued on normal chow and group 2 on HFD only. Both the test drugs significantly (p-value < 0.01) reduced the body weight. Highest reduction (42%) occurred by the combination of GS and AC when compared to the disease control group while GS alone at 200mg/kg caused 38.1% reduction. All the test drugs also significantly reduced the BMI with maximum improvement in the combination group (49.5%) as compared to disease control.

Locomotor activity has been used by few studies as a marker of obesity ¹⁸. To assess the locomotor activity, ambulatory activity and frequency of rearing was observed. Both the drugs significantly increased the ambulatory activity and decreased the rearing. Combination was found superior than the individual drugs with 51.4% increase in the AA and 36.5% reduction in the rearing.

At the end of 8^th week changes in the internal organs (kidney, liver, spleen and heart) and fat pads (perirenal, gonadal and mesenteric) were also measured and compared to the normal control group and the disease control group. Due to high fat diet the weight of internal organs and fad pads increased significantly in disease control group as compared to the normal control and other groups. Both Gymnema and Acorus significantly reduced the weight of internal organs and fat pads. Highest reduction occurred in the group taking combination of both drugs followed by GS 200mg/kg. Weight of liver, kidney, spleen and heart found comparable to the normal control group. Weight of internal fat pads also decreased significantly when compared with the disease control group with maximum reduction by the combination of drugs.

Both the plants, Gymnema sylvestre and Acorus calamus, are rich in phytochemicals and are well known in traditional system of medicine. Leaves extract of Gymnema is rich in gymnemic acids that help in the inhibition of fat and oil hydrolyzates absorption from the digestive tract ¹⁹. Gymnemic acids not only reduce the craving for food but also increase the fecal excretion of bile acids and neutral steroids ^{20, 21}. Asarones are the phytochemicals that are thought to be responsible for the weight reducing property of Acorus calamus. Asarone inhibits the differentiation of adipocytes ²². From the present study it can be concluded that leaves extract of both Gymnema sylvestre and Acorus calamus has a potent dose-dependent antiobesity activity as shown by reduction in all the parameters of obesity taken into account. Combination of both the drugs was found superior to the individuals drugs. Based on these findings it is suggested to extend the domain of this study onto higher animals and human beings to further substantiate the results.

ACKNOWLEDGEMENT: Nil.

CONFLICT OF INTEREST: Nil.

REFERENCES:

1. WHO Obesity and overweight. Fact sheet N° 311. Geneva: World Health Organization, 2015.
2. Cefalu WT, Bray GA, Home PD et al.: Advances in the science, treatment, and prevention of the disease of obesity: reflections from a Diabetes Care editors' expert forum. Diabetes Care 2015; 38(8): 1567-1582.
3. Hisalkar PJ, Patne AB and Fawade MM: Assessment of plasma antioxidant levels in type 2 diabetes patients. Int J Biol Med Res 2012; 3(2): 1796-1800.
4. Grant-Guimaraes J, Ronald F, Erica L and Jennifer K: Childhood Overweight and Obesity. Gastroenterology Clinics of North America 2016; 45(4): 715-728.
5. Forde CG, Van Kuijk N, Thaler T, de Graaf C and Martin N: Oral processing characteristics of solid savoury meal components and relationship with food composition, sensory attribute and expected satiation. Appetite 2013; 60: 208–19.
6. Hill JO and Wyatt HR: Role of physical activity in preventing and treating obesity. J Appl Physiol 2005; 99: 765-70.
7. Östman J, Britton M and Jonsson E: Treating and Preventing Obesity. An Evidence Based Review. Wiley-VCH Verlag GmbH and KgaA: Weinheim 2004; 355.
8. Hasani-Ranjbar S, Larijani B and Abdollahi M: A systematic review of the potential herbal sources of future drugs effective in oxidant-related diseases. Inflamm Allergy Drug Targets 2009; 8: 2–10.
9. Manohar SH, Naik PM, Praveen N and Murthy HN: Distribution of gymnemic acid in various organs of Gymnema sylvestre. Journal of Forestry Research 2009; 20(3): 268–270.
10. Samar R, Agrawal MK, Varma A and Jain M: Ethno-botanical documentation of some vegetable plants in the villages of Guna district, Madhya Pradesh, India. Indian J. Life Sci. 2012; 1(2): 75 78.
11. Balakumbahan R, Rajamani K and Kumanan K: Acorus calamus: An overview. J Med Plant Res 2010; 4: 2740‑
12. Prisilla DH, Balamurugan R and Shah HR: Antidiabetic activity of methanol extract of Acorus calamus in STZ induced diabetic rats. Asian Pac J Trop Biomed 2012; 2(2): 941‑
13. Prasad L, Khan TH, Jahangir T and Sultana S: Acorus calamus extracts and nickel chloride: prevention of oxidative damage and hyperproliferation response in rat kidney. Biol Trace Elem Res2006; 113(1): 77-92.
14. Rachh PR, Rachh MR, Ghadiya NR et al.: Antihyperlipidemic activity of Gymenma sylvestreBr. leaf extract on rats fed with high cholesterol diet. International Journal of Pharmacology 2010; 6(2): 138–141.
15. Patel P, Vaghasiya J, Thakor A and Jariwala J: Antihypertensive effect of rhizome part of Acorus calamus on renal artery occlusion induced hypertension in rats. Asian Pac J Trop Dis 2012; 2(1): 6-10.
16. Benoit SC, Kemp CJ, Elias CF, Abplanalp W, Herman JP, Migrenne S, Lefevre AL, Cruciani-Guglielmacci C, Magnan C, Yu F, Niswender K, Irani BG, Holland WL and Clegg DJ: Palmitic acid mediates hypothalamic insulin resistance by altering PKC-theta subcellular localization in rodents. J Clin Invest 2009; 119: 2577–2589.
17. Hariri N and Thibault L: High-fat diet-induced obesity in animal models. Nutr Res Rev 2010; 23: 270–299.
18. Mikael B, Anna-Karin G, Christopher J, Egecioglu LE, Elmgren A, Tornell J, Oscarsson J and Mohammad BY: Acutely reduced locomotor activity is a major contributor to Western diet-induced obesity in mice. Am J Physiol Endocrinol Metab 2008; 294: 251–260.
19. Ahmed ABA, Rao AS and Rao MV: Role of in vivo leaf and in vitro callus of Gymnema sylvestre in maintaining normal levels of blood glucose and lipid profile in diabetic Wistar rats.; Biomedicine 2008; 2: 134-38.
20. Shirouchi B, Kawamura S, Matsuoka R, Baba S, Nagata K, Shiratake S, Tomoyori H, Imaizumi K and Sato M: 2011; 46(8): 789-93.
21. Santas J, Espadaler J, Cuñé J and Rafecas M: Partially hydrolyzed guar gums reduce dietary fatty acid and sterol absorption in guinea pigs independent of viscosity; Lipids 2012; 47(7): 697-705.
22. Lee MH, Chen YY, et al.: Inhibitory effect of β-asarone, a component of Acorus calamus essential oil, on inhibition of adipogenesis in 3T3-L1 cells. Food Chemistry 2011; 126: 1–7.
    

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    How to cite this article:

    Singh DK, Sachan A, Kumar N, Lakhani P, Tutu S, Shankar P, Nath R, Kumar A and Dixit RK: Evaluation of antiobesity effect of Gymnema sylvestre and Acorus calamus on diet induced model of obesity in rats. Int J Pharm Sci Res 2017; 8(10): 4341-47.doi: 10.13040/IJPSR.0975-8232.8(10).4341-47.

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