Effect of Gymnema sylvestre R. Br. leaves extract on certain physiological parameters of diabetic rats

1 Introduction

Diabetes mellitus (DM) is a metabolic disorder characterized by hyperglycemia resulting from defective insulin secretion, resistance to insulin action or both (Jarald et al., 2008; Vasi and Austin, 2009; Khan et al., 2012). DM is a widespread disease, associates with chronic micro- and macro-vascular complications (Goycheva et al., 2006; Ahmed et al., 2011; Aralelimath and Bhise, 2012). Total recovery from diabetes has not been reported up to now and this explains the intensive studies done on it.

Untreated diabetes leads to serious complications or even premature death. The treatment of diabetes mellitus is based on insulin and/or oral hypoglycemic drugs (Daisy et al., 2009). These drugs act by various mechanisms to control the blood glucose level, but many side-effects have been reported (Patel et al., 2012). Therefore, there is considerable interest in the field of medicinal plants due to their natural origin and less side effects (Jarald et al., 2008). One of these medicinal plants is Gymnema sylvestre (Gurmar), which means sugar killer (Najafi and Deokule, 2011). It is a wild plant that grows in the open forest in India, China, Indonesia, Japan, Malaysia, Sri Lanka, Vietnam, and South Africa (Gurav et al., 2007; Spasov et al., 2008). The leaves of the plant in particular are used as antiviral, diuretic, antiallergic, hypoglycemic, hypolipidemic, antibiotic, in stomach pains and in rheumatism (Saneja et al., 2010).

Most studies on G. sylvestre leaves extract focused on its role as hypoglycemic medicinal plant, but the present study demonstrates the effect of G. sylvestre leaves extract (18 mg/kg body weight) on several etiological factors of STZ-diabetic rats including hyperglycemia, hypoinsulinemia, hyperlipidemia and oxidative stress at the same time.

2 Materials and methods

3 Results

• Effect of G. sylvestre leaves extract on plasma glucose and insulin levels.

Plasma glucose level was significantly low (p < 0.05) in the rat group supplemented with G. sylvestre leaves extract (group III) compared to those of control rat groups (groups I and II, respectively). On the other hand, plasma glucose levels were significantly high in the diabetic rat group (group IV) and the rat group treated with G. sylvestre leaves extract post diabetic induction (group V) in relation to those of control rat groups. Plasma glucose level decreased significantly in group V compared to that of group IV (Table 1).

Plasma insulin level was significantly high (p < 0.05) in group III compared to those of groups I and II. On the other hand, plasma insulin levels were significantly low in groups IV and V compared to those of groups I and II. Plasma insulin level increased significantly in group V compared to that of group IV (Table 1).

• Effect of G. sylvestre leaves extract on plasma liver function enzymes level.

Both plasma alanine aminotransaminase (ALT) and aspartate aminotransaminase (AST) levels in group III showed a non significant difference (p < 0.05) in relation to those of groups I and II. Plasma ALT and AST levels were significantly high in groups IV and V compared to those of groups I and II. Plasma ALT and AST levels were significantly low in group V compared to those of group IV (Table 2).

• Effect of G. sylvestre leaves extract on plasma lipid profile.

Plasma triglycerides, cholesterol and LDL-cholesterol levels decreased significantly (p < 0.05) in group III compared to those of groups I and II. Plasma triglycerides, cholesterol and LDL-cholesterol levels in groups IV and V were significantly high compared to those of groups I and group II. Plasma triglycerides, cholesterol and LDL-cholesterol levels decreased significantly in group V compared to those of group IV (Table 3).

Plasma HDL-cholesterol levels increased significantly (p < 0.05) in group III compared to those of groups I and II. On the other hand, plasma HDL-cholesterol levels were significantly low compared to those of groups I and II. Plasma HDL-cholesterol level was increased significantly in group V compared to that of group IV (Table 3).

• Effect of G. sylvestre leaves extract on antioxidant parameter levels.

Plasma malondialdehyde (MDA) levels in groups III and V showed a non significant difference (p < 0.05) in relation to those of groups I and II. Plasma MDA level was significantly increased in group IV compared to that of groups I and II. Plasma MDA level was significantly decreased in group V compared to that of group IV (Table 4).

Plasma catalase level in group III was significantly low (p < 0.05) compared to that of groups I and II. Plasma catalase levels in groups IV and V increased significantly compared to those of groups I and II. Plasma catalase level in group V showed non significant difference compared to that of group IV (Table 4).

Erythrocyte superoxide dismutase (SOD) levels were significantly low (p < 0.05) in groups III and IV compared to those of groups I and II. Erythrocyte SOD level in group V showed a non significant difference compared to that of groups I and II. Erythrocyte SOD level in group V showed a significant increase compared to that of group IV (Table 4).

Plasma reduced glutathione (GSH) level was significantly decreased (p < 0.05) in group II compared to that of group I. On the other hand, plasma GSH levels in groups III and V showed a non significant difference compared to that of group I. Plasma GSH level in group IV increased significantly compared to that of group I. Plasma GSH level in group V showed a non significant difference compared to that of group IV (Table 4).

4 Discussion

G. sylvestre is an important medicinal plant used in different systems of medicine as a remedy for the treatment of diabetes (Jarald et al., 2008).

The present study showed a significant decrease in plasma glucose level (20.20%) in diabetic rats treated with G. sylvestre leaves extract (18 mg/kg body weight) compared to that of untreated diabetic rats. Significant increase in diabetic rats’ insulin level post treated with G. sylvestre leaves extract confirmed our result. Moreover, glucose level might be decreased in treated diabetic rats as a result of decreasing gluconeogenesis that was indicated by low levels of ALT and AST in treated diabetic rats compared to untreated diabetic rats.

This result was in agreement with Shanmugasundaram et al. (1983) who reported that administration of dried leaf powder of G. sylvestre decreased glucose levels as it controlled gluconeogenic enzymes (ALT and AST) and increased glycogen levels in liver, kidney and muscle. Treated 27 type 1 diabetic patients with water extract of G. sylvestre leaves (400 mg/day) for 12 months reduced blood glucose level (up to 35%) as a direct effect of increasing exogenous insulin level (up to 50%) (Shanmugasundaram et al., 1990). Chattopadhyay (1998) reported that administration of alcoholic extract of G. sylvestre leaves to Swiss albino rats fed by glucose increases the insulin level. Bolkent et al. (2000) explained that the decrease in plasma glucose levels might be due to the increase in insulin levels after administration of gymnemic acid to diabetic rats. Gholap and Kar (2005) discovered that gymnemic acid was capable of reducing corticosteroid hormones in dexamethasone-induced hyperglycemia in mice. Daisy et al. (2009) reported that plasma glucose level reduced more than 50% in diabetic rats treated by dihydroxy gymnemic triacetate (20 mg/kg b.w.) for 40 days compared to untreated diabetic rats and they explain this reduction as a result of increasing plasma insulin (50%), muscle glycogen (77.1%) and liver glycogen (59.09%) content. In vivo leaf and in vitro callus methanol extract of G. sylvestre decreased blood glucose level (72.4%) in diabetic rats (Ahmed et al., 2008) and that might be due to increased liver glycogen content (Ahmed et al., 2010). The activities of phosphorylase enzymes and sorbitol dehydrogenase were increased in diabetic rabbits after treated with G. sylvestre extract (Venkatesham et al., 2010).

This result was supported also by Yoshioka (1986) who demonstrated that Gymnemic acid inhibited Na⁺ dependent active glucose transport in the small intestine. Fushiki et al. (1992) suggested that the G. sylvestre leaves extract inhibited gastric inhibitory peptide (GIP). Sahu et al. (1996) reported that the gymnemic acid molecules fill the receptor location in the absorptive external layers of the intestine thereby preventing the glucose absorption by the rat intestine. It was suggested that gymnemic acid extract inhibited glucose uptake in the intestine of guinea pig which occurred due to the effect of gymnemic acid extract on the suppression of high K⁺ induced contraction in guinea pig ileac longitudinal muscles (Shimizu et al., 1997). The treatment with high concentrations of G. sylvestre leaves extract reduced the levels of glucose in albino Wistar rats and that might be due to the effect of gymnemic acid on the inhibition of glucose uptake in the intestine (sujin et al., 2008). Hypoglycemic potential of G. sylvestre, Tinospora cordifolia, Eugenia jambolana and Aegle marmelos might be due to their effect on inhibition of the activity of α-amylase resulting in the delayed digestion of the dietary carbohydrates, so the liberated glucose from the intestine lumen to the circulation decreased (Ahmed et al., 2011).

Plasma insulin levels significantly increased (83.40%) in diabetic rats treated with G. sylvestre leaves extract (18 mg/kg body weight) compared to that of untreated diabetic rats. Our result was in accordance with Persaud et al. (1999) who suggested that alcoholic extract of G. sylvestre leaves increased insulin release in vitro by two mechanisms: (1) the major mode of action was through β-cell plasma membrane permeability, (2) the pores formed by plasma membrane disruption. The number of pancreatic β-cells in the diabetic rats doubled after administration of alcoholic extract of G. sylvestre leaves (Shane-McWhorter, 2001). Daisy et al. (2009) reported that the insulin levels in diabetic rats increased (50%) after treatment with dihydroxy gymnemic triacetate and that might be due to the stimulation of insulin secretion from regenerated or residual β-cells by dihydroxy gymnemic triacetate. Liu et al. (2009) suggested that the increase in insulin secretion from mouse β-cells and isolated human islets after the administration of low concentration of G. sylvestre leaves extract might be partially dependent on the increase in Ca⁺ influx. Degenerative changes in pancreatic β-cells were minimized and normal morphology maintained in alloxan induced diabetic rats after administration of gymnemic acid (Ahmed et al., 2010). Microscopy studies of pancreas of animals receiving G. sylvestre extract showed that nuclei of endocrinocytes were significantly enlarged in all sections of the organ with the same volume fraction and area of pancreatic islets. The result agreed completely with investigations proposed that use of G. sylvestre increased the endogenous levels of insulin, possibly due to regeneration of pancreas cells (Venkatesham et al., 2010).

In the present study, rats treated with G. sylvestre leaves extract post STZ-diabetic induction showed a significant decrease in triglyceride, cholesterol and LDL-cholesterol and showed a significant increase in HDL-cholesterol as compared to that of untreated diabetic rats. Decreasing levels of triglyceride, cholesterol and LDL-cholesterol and increasing level of HDL-cholesterol might be due to an increase in insulin which caused an increased activity of lipoprotein lipase (Facilitated chylomicron transport through cell membranes) and a decreased activity of hormone-sensitive lipase (converted neutral fats into free fatty acids). This result was in agreement with Daisy et al. (2009) and Aralelimath and Bhise (2012) who reported that increasing insulin secretion after administration of G. sylvestre extract led to a decrease of cholesterogenesis and fatty acid synthesis.

This result was supported also by Mall et al. (2009) who reported that G. sylvestre decreases total cholesterol, LDL-cholesterol, VLDL-cholesterol and triglyceride levels in diabetic rats and that could be due to the presence of hypolipidemic agent such as sitosterol in the aqueous leaf extract. G. sylvestre inhibited the intestinal absorption of oleic acid in rats, which suggested the possibility of G. sylvestre to inhibit lipid absorption (Wang et al., 1998). Shigematsu et al. (2001) reported that administration of G. sylvestre leaves extract decreased the total cholesterol, LDL-cholesterol, VLDL-cholesterol and triglyceride levels in the serum of rats fed with a high fat diet or normal fat diet and that might be due to the effect of G. sylvestre in increasing neutral sterols and acid steroids excretion into feces. Luo et al. (2007) discovered the ability of G. sylvestre water extract to prevent the genetic obesity by improving the cholesterol metabolism and inhibiting polyphagia. Ishijima et al. (2008) suggested that gymnemic acid might have some pharmacological activities including antidiabetic activity and lipid lowering effects via the inhibition of glycerol-3-phosphate dehydrogenase activity in vitro. The intra peritoneal fat and fat drop vacuoles on the epithelium of renal tubules were scattered by administration of the G. sylvestre extract (Mahajan et al., 2011). Lipid lowering activity of flavonoids and saponins extracted from G. sylvestre might be due to the inhibition of pancreatic lipase activity (Manish et al., 2011).

It was apparent from our results that plasma lipid peroxidation level decreased and erythrocyte superoxide dismutase level increased in rats treated with G. sylvestre leaves extract post STZ-diabetic induction compared to untreated diabetic rats.

Ohmori et al. (2005) discovered the antioxidant ability of G. sylvestre, when they studied the antioxidant activity of six teas against free radicals and LDL oxidation in 10 healthy volunteers ranging from 20 to 22 years of age. In vitro G. sylvestre alcoholic leaf extract showed antioxidant ability by inhibiting 1,1-diphenyl-2-picrylhdrazyl (DPPH) and scavenging superoxide and hydrogen peroxide (Rachh et al., 2009). Administration of G. sylvestre extract to diabetic rats increased superoxide dismutase activity and decreased lipid peroxide by either directly scavenging the reactive oxygen species, due to the presence of various antioxidant compounds, or by increasing the synthesis of antioxidant molecules (albumin and uric acid) (Vasi and Austin, 2009). Fazal et al. (2011) found that the maximum scavenging activity of ethanolic extract of G. sylvestre in vitro was 54.4% at concentration 250 μg/ml. Some poly herbal ayurvedic formulations like Hyponidd and Dihar containing G. sylvestre extract had shown antioxidant activity by increasing superoxide dismutase, glutathione and catalase levels in rats (Mahajan et al., 2011). Kang et al. (2012) reported that feeding G. sylvestre extract to diabetic rats decreased lipid peroxidation levels by 31.7% in serum, 9.9% in liver and 9.1% in kidney. It also decreased the activity of glutathione peroxidase in cytosolic liver and glutamate pyruvate transaminase in serum to normal levels.

5 Conclusion

It can be concluded that, G. sylvestre leaves extract exhibit antidiabetic activity, so it is recommended to use it as a complementary medicine in diabetes mellitus. Hence, clinical studies on G. sylvestre leaves extract as a complementary medicine are needed.