Antidiabetic Effect of GII Compound Purified from Fenugreek (Trigonella foenum graecum Linn) Seeds in Diabetic Rabbits.

Aim is to study the antidiabetic effect of a compound GII purified earlier from the water extract of fenugreek (Trigonella foenum graecum) seeds by Murthy and his colleagues (patented in India and USA) in diabetic rabbits. Diabetes was induced in rabbits by injecting 80 mg/kg bw of alloxan intravenously into rabiits. Rabbits were subdivided into subdiabetic [fasting blood sugar (FBG) up to 120 mg/dl with abnormal glucose tolerance in glucose tolerance test (GTT)], moderately diabetic (FBG below 250 mg/dl) and severely diabetic (FBG above 250 mg/dl). Blood glucose and glycosylated hemoglobin (HbA1C) were estimated by procedures in the kits of Stangen Immunodiagnostics, Mumbai using, respectively, glucose oxidase method and absorbance at 415 nm. Serum insulin was estimated by the ELISA method as described in the kit of Boehringer Mannheim Immunodiagnostics, Mumbai, India. GII was found to improve blood glucose utilization in GTT and reduced FBG and HbA1C. In the present communication detailed studies were carried out with GII in the subdiabetic, moderately diabetic and severely diabetic rabbits. GII at a dose of 50 mg/kg bw per day brought down the elevated FBG levels in the untreated subdiabetic (FBG 96.6 ± 7 mg/dl), moderately diabetic (150.1 ± 14 mg/dl) and severely diabetic rabbits (427 ± 46 mg/dl) to normal in 12, 15 and 28 days of treatment. It improved serum HbA1C and insulin levels also in these rabbits. Intermittent therapy once a week for 6 weeks with GII at the same dose brought down the FBG values to normal in the subdiabetic (FBG 96.0 ± 2 mg/dl) and in the moderately diabetic rabbits to 133.0 ± 12 mg/dl. After stopping therapy of the subdiabetic and moderately diabetic rabbits whose FBG values came to normal after treatment with GII 50 mg/kg bw, the values remained normal for 1 week and showed a tendency to increase only after 15 days. If these animal studies are applicable to humans these results indicate that a diabetic person need not take GII daily when once the FBG value comes to normal or near to normal. Patients might be able to take GII only when the FBG value shows tendency to increase. So, intermittent therapy is possible with the potent product GII of the fenugreek seeds which is of a great advantage.

Mechanism of Antidiabetic Action of Compound GII Purified from Fenugreek (Trigonella foenum graecum) Seeds.

To study the mechanism of action of water soluble compound GII purified from fenugreek (Trigonella foenum graecum) seeds which was shown earlier to have antidiabetic effect in the subdiabetic, moderately and severely diabetic rabbits. In rabbits (1-1.5 kg bw) diabetes was induced by intravenous injection of 80 mg/kg bw of alloxan. They were fed with GII at a dose of 50 mg/kg bw daily once in the morning for 15 days in the subdiabetic and moderately diabetic and 30 days in the severely diabetic rabbits. Serum total cholesterol (TC), triglycerides (TG), LDL + VLDL cholesterol [(LDL + VLDL)C], HDL cholesterol [(HDL)C], total tissue lipids, glycogen and enzymes of carbohydrate metabolism (glycolysis, gluconeogenesis, polyol pathway) hexokinase, glucokinase, pyruvate kinase, malic enzyme, glucose-6-phosphatase, glucose-6-phosphate dehydrogenase, aldose reductase and sorbitol dehydrogenase and antioxidant enzymes glutathione peroxidase, glutathione reductase and superoxide dismutase were estimated. Liver and kidney function parameters were also estimated. Treatment with GII for 15 days in the subdiabetic and moderately diabetic rabbits and for 30 days in the severely diabetic rabbits (i) decreased the elevated lipids TC, TG, (LDL + VLDL)C and increased the decreased (HDL)C, (ii) decreased the elevated liver and heart total lipids, TC and TG, (iii) increased the decreased liver and muscle glycogen, (iv) increased the decreased hexokinase, glucokinase, pyruvate kinase, malic enzyme, glucose-6-phosphate dehydrogenase, superoxide dismutase, glutathione peroxidase, (v) decreased the increased glucose-6-phosphatase, sorbitol dehydrogenase, aldose reductase. Results thus show that treatment with GII compound purified from fenugreek seeds for 15 days in the subdiabetic and moderately diabetic and 30 days in the severely diabetic rabbits corrects the altered serum lipids, tissue lipids, glycogen, enzymes of glycolysis, gluconeogenesis, glycogen metabolism, polyol pathway and antioxidant enzymes. Histopathological abnormalities (fatty infiltration and other cellular changes) seen in the pancreas, liver, heart and kidneys were repaired after treatment with GII. In fact partially damaged pancreas was repaired. Liver and kidney function test results were normal in the GII treated animals indicating that GII treatment is safe and free from any side effects.

Anti-hyperglycemic compound (GII) from fenugreek (Trigonella foenum-graecum Linn.) seeds, its purification and effect in diabetes mellitus.

An anti-hyperglycemic compound named GII was purified from the water extract of the seeds of fenugreek (T. foenum-graecum) and shown to be different from trigonelline and nicotinic acid isolated earlier from the same plant. GII (50 mg/kg body weight, po) reduced blood glucose in glucose tolerance test (GTT) in the sub-diabetic and moderately diabetic rabbits and significantly reduced the area under the curve (AUC) of GTT. Treatment for 7 days of the sub-diabetic rabbits with GII (50 mg/kg body weight, po) improved glucose tolerance without reducing fasting blood glucose (FBG) which was nearly normal. The results suggest that there is no risk of hypoglycemia in near normal animals (may be humans also) with abnormal GTT. Treatment of the moderately diabetic rabbits with GII (100 mg/kg body weight for 3 weeks) reduced FBG to nearly normal value and improved GTT. GII was more effective than the standard drug tolbutamide. Intermittent therapy given on days 1-5, 11-15, 26-30 and 56-60 to moderately diabetic rabbits leaving in between days without treatment brought down FBG to normal and AUC during GTT was normal. After 15 days treatment with GII (100 mg/kg body weight for 3 weeks) glycosylated hemoglobin came down and insulin increased to normal values in the sub-diabetic, moderately diabetic and severely diabetic rabbits. GII treatment (100 mg/kg body weight for 15 days) brought down all the altered serum lipids (TC, HDLC, TAG, PLs and FFAs) to normal levels. The results suggest that intermittent therapy, instead of daily therapy is possible and GII has good potential as an oral anti-diabetic drug with intermittent therapy.

Mechanism of anti-diabetic action, efficacy and safety profile of GII purified from fenugreek (Trigonella foenum-graceum Linn.) seeds in diabetic animals.

Mechanism of action of GII (100 mg/kg body weight, po for 15 days) purified from fenugreek (T. foenum-graecum) seeds was studied in the sub-diabetic and moderately diabetic rabbits. In the sub-diabetic rabbits it did not change much the content of total lipids, glycogen and proteins in the liver, muscle and heart (glycogen was not studied in the heart). However, in the moderately diabetic rabbits same treatment decreased total lipids more in the liver (21%) than those in the heart and muscle. Total protein content increased (14%) in the liver but negligible change (5-7%) was observed in heart and muscle. Glycogen increased (17%) in the liver but not in the muscle of the moderately diabetic rabbits (glycogen was not estimated in the heart). Among the enzymes of glycolysis, activity of glucokinase was not affected in the liver of both the sub-diabetic and moderately diabetic rabbits. Phosphofructokinase and pyruvate kinase activity in both sub-diabetic and moderately diabetic rabbits increased (13-50%) indicating stimulation of glycolysis. The activity of gluconeogenic enzymes glucose-6-phosphatase and fructose-1,6-diphosphatase of the sub-diabetic rabbits decreased in the liver (15-20%) but not in the kidneys. In the moderately diabetic rabbits after treatment with GII, glucokinase in the liver was not affected much (-9%) but increased well in the muscle (40%). Phosphofructokinase and pyruvate kinase were moderately increased both in the liver and the muscle (18-23%). The gluconeogenic enzyme glucose-6-phosphatase decreased reasonably well in the liver and kidneys (22, 32%). Fructose-1,6-diphosphatase decreased only slightly (10, 9%) in the moderately diabetic rabbits. Thus GII seems to decrease lipid content of liver and stimulate the enzymes of glycolysis (except glucokinase) and inhibit enzymes of gluconeogenesis in the liver of the diabetic especially moderately diabetic rabbits.

Coronary risk variables in young asymptomatic smokers.

BACKGROUND AND OBJECTIVES: Smoking plays a dominant role in premature atherosclerosis particularly among males in South Asian countries. It initiates and promotes atherosclerosis by altering cardiac haemodynamics, causing dyslipidaemia and producing oxidative damage. Not much information is available from our country. We therefore undertook this study to see the effect of smoking on electrocardiogram (ECG), blood pressure, lipids, apolipoprotein B level and free radical activity in young asymptomatic male smokers. METHODS: The study included 100 consecutive male subjects (50 smokers and 50 non smokers) aged 30-40 yr. Smoking profile, detailed cardiovascular assessment including ECG and lipid profile were evaluated in each subject. RESULTS: Of the 50 smokers, 22 (44%) had grade I hypertension as against 5 of 50 non smokers. Sinus tachycardia (10%) and P-pulmonale (8%) were the only notable ECG abnormalities. Dyslipidaemia was detected in 92 per cent smokers and 48 per cent non smokers (P<0.001). Total serum cholesterol, low density lipoprotein (LDL)-cholesterol, triglycerides and apolipoprotein B levels were significantly higher (P<0.001) in smokers compared to non smokers. LDL-cholesterol was > or =135 mg/dl in 94 per cent dyslipidaemic smokers. However, no significant difference was found in high density lipoprotein (HDL)-cholesterol. Smokers had significantly higher serum malondialdehyde levels (P<0.001) and low superoxide dismutase (P<0.001) compared to non smokers. INTERPRETATION AND CONCLUSION: Our data indicate that young asymptomatic male smokers tend to have hypertension, dyslipidaemia and increased production of free oxygen radicals, perhaps by attenuation of oxidative stress by cigarette smoking. This makes them prone for premature coronary artery disease. However, the findings need to be confirmed on a larger sample.

Antioxidant effect of aqueous extract of the bark of Ficus bengalensis in hypercholesterolaemic rabbits.

The antioxidant effect of aqueous extract of the bark of Ficus bengalensis has been evaluated in hypercholesterolaemic rabbits. Rabbits were divided into three groups, Group I served as healthy control; groups II and III were made hypercholesterolaemic by feeding cholesterol suspended in groundnut oil (100 mg/kg body weight per day) for 6 weeks. Rabbits of Group III received water extract of the bark of Ficus bengalensis at a dose of 50 mg/kg body weight per day in addition to cholesterol suspended in oil. Feeding cholesterol increased serum cholesterol, triacylglycerol and LDL + VLDL-cholesterol significantly in Group II as compared to Group I (P = 0.001). Treatment with water extract decreased the serum cholesterol level by 59%, triacylglycerol by 54% and LDL + VLDL-cholesterol by 60% in Group III as compared to Group II. In addition, treatment with this extract led to a decrease in lipid peroxidation as evidenced by fall in thiobarbituric acid reactive substances with a corresponding increase in blood glutathione content (P = 0.001). Further, there was significant increase in the activities of antioxidant enzymes; superoxide dismutase (P < 0.001), catalase (P < 0.03), glutathione peroxidase (P = 0.03) and glutathione reductase (P < 0.01); which were depressed in Group II rabbits after cholesterol feeding. Thus, our results show that the water extract of the bark of Ficus bengalensis has significant antioxidant effect, in addition to hypolipidaemic effect.

Hypoglycaemic and hypolipidemic effect of ethanolic extract of seeds of Eugenia jambolana in alloxan-induced diabetic rabbits.

The hypoglycaemic and hypolipidemic effect of ethanolic extract obtained from seeds of E. jambolana was investigated in alloxan-induced diabetic rabbits. Hypoglycaemic activity was assessed by reduction in fasting blood glucose (FBG) at 90min and also fall in peak blood glucose during glucose tolerance test (GTT) in sub-diabetic and mild diabetic (MD) rabbits, but in severe diabetic (SD) rabbits by reduction in FBG at 90min. Ethanolic extract (100mg/kg body weight) when given orally to sub-diabetic (AR) for 1 day, MD for 7 days and SD for 15 days showed significant fall in FBG at 90min (12% AR, 18.9% MD and 29% SD) and also produced 16.9% fall in peak blood glucose in AR and 21% in MD rabbits during GTT. When administered daily for 15 days to MD and SD rabbits, significant fall in FBG (41.3% MD, 31.6% SD) and glycosylated haemoglobin (GHb) levels (23.3% MD, 26.6% SD) were observed, while serum insulin level showed significant increase (32.8% MD, 26.9% SD). Liver and muscle glycogen content also increased. The ethanolic extract of seeds also exhibited significant hypolipidemic effect as evident from fall in total serum cholesterol (TC)/high density lipoprotein cholesterol (HDL-c) ratio, serum low density lipoprotein cholesterol (LDL-c) levels and decreased activity of HMG-CoA reductase. The histopathological studies of liver, pancreas and aorta in alcoholic extract treated diabetic groups revealed almost normal appearance.

Acute and chronic toxicity studies on partially purified hypoglycemic preparation from water extract of bark ofFicus bengalensis.

Acute and chronic toxicity studies were conducted to assess toxicity of a partially purified preparation from the water extract of the bark ofFicus bengalensis, which was demonstrated in our earlier studies to have significant hypoglycemic and hypocholesteroiemic effect on alloxan induced, mild and severe diabetes in rabbits. LD(50) of this preparation was found to be ∼1 gm/kg in rats when given orally. For chronic toxicity studies 3 doses of aqueous preparation were given to 3 groups of rats. First group received 5 times ED(50) (50 mg/kg), second group 10 times ED(50) (100 mg/kg) and the third group 15 times ED(50) (150 mg/kg) for 3 months. Fourth group which served as control was given water. After three months, blood was collected for studying biochemical and hematological parameters. Blood glucose, serum cholesterol, liver and kidney function tests, haemoglobin, total and differential leukocyte count were determined. Animals were sacrificed and histopathological examination of liver, heart and kidneys was carried out. Results of the study showed that partially purified preparation fromFicus bengalensis is not toxic by all the above mentioned parameters.

Mechanism of action of a hypoglycemic principle isolated from fenugreek seeds.

Mechanism of action of an orally active hypoglycemic principle isolated from water extract of seeds of Trigonella foenum graecum (fenugreek) was investigated in alloxan induced subdiabetic and overtly diabetic rabbits of different severities. The active principle was orally administered to the subdiabetic and mild diabetic rabbits (five in each group) at a dose of 50 mg/kg body weight for 15 days. The treatment produced significant attenuation of the glucose tolerance curve and improvement in the glucose induced insulin response, suggesting that the hypoglycemic effect may be mediated through stimulating insulin synthesis and/or secretion from the beta pancreatic cells of Langerhans. Prolonged administration of the same dose of the active principle for 30 days to the severely diabetic rabbits (n = 5) lowered fasting blood glucose significantly, but could elevate the fasting serum insulin level to a much lower extent, which suggests an extra-pancreatic mode of action for the active principle. The effect may also be by increasing the sensitivity of tissues to available insulin. The hypoglycemic effect was observed to be slow but sustained, without any risk of developing severe hypoglycemia.

Lipoprotein(a) as an independent risk factor for coronary artery disease in patients below 40 years of age.

Coronary artery disease has assumed alarming proportions in Indians and often affects people at younger age. Traditional risk factors fail to explain the high incidence of disease. Although lipoprotein(a) has been shown to be a powerful risk factor for atherosclerosis, there is very limited data with regard to its significance in premature coronary artery disease. The present study was therefore undertaken to assess lipoprotein(a) levels and its role as a marker of coronary artery disease in patients below the age of 40 years. Lipid profile and lipoprotein(a) levels were estimated in 50 patients of angiographically proven coronary artery disease and an equal number of age-matched healthy controls. There was no significant difference in the family history of coronary artery disease, body mass index and waist-hip ratio between the two groups. Total plasma cholesterol, triglyceride and LDL-cholesterol levels were significantly higher and HDL-cholesterol significantly lower in patients as compared to controls. In patients of coronary artery disease, mean lipoprotein(a) levels, measured by ELISA method, were 35.0 +/- 32.4 mg/dL and the median was 26.7 mg/dL. These values were significantly higher than the mean of 20.3 +/- 17.0 mg/dL (p < 0.002) and the median of 13.8 mg/dL (p < 0.015) in controls. Multiple regression analysis, to assess the influence of various risk factors, showed that low HDL-cholesterol (odds ratio 4.62, 95% CI 1.84-11.60; p < 0.015) and elevated lipoprotein(a) levels (odds ratio 3.06, 95% CI 1.24-7.55; p < 0.001) were independent risk factors, whereas high total cholesterol and triglyceride levels did not have any independent influence on premature coronary artery disease. Our data thus suggest that lipoprotein (a) levels are elevated and constitute an independent risk factor in patients with premature coronary artery disease below 40 years of age.

Studies on oxidative stress, serum iron and iron binding capacity in subjects prone to the risk of coronary artery disease.

Lipid peroxidation in vitro and in vivo has been postulated to be involved in the development of atherosclerosis. It is also known that free iron catalyses the lipid peroxidation. Therefore, we assessed the status of oxidative stress in smokers, hypertensives and non-insulin dependent subjects, who were prone to coronary artery disease. In addition, superoxide dismutase levels and iron binding capacity were also measured to know their antioxidant defences. One hundred seventy-five consecutive subjects below 60 years of age were examined; they were then divided into three groups: one with coronary artery disease, another without coronary artery disease and a healthy control group. The patients having either of the one risk factors for coronary artery disease i.e. smoking, hypertension and/or diabetes were studied. Serum lipid peroxides, superoxide dismutase, serum iron and iron binding capacity were estimated. Oxidative stress was highest in smokers with coronary artery disease (3.11+/-0.79 mmol/ml) as compared to hypertensives (2.69+/-0.20 mmol/nl) and non-insulin dependent diabetics (2.78+/-0.19 mmol/ml). Superoxide dismutase activity was also significantly decreased (p<0.001) in smokers with coronary artery disease as compared to hypertensives and non-insulin dependent diabetes mellitus. Final step of stepwise logistic regression based on malondialdehyde and superoxide dismutase correctly predicted coronary artery disease status in 90 percent smokers. Serum iron and total iron binding capacity were not significantly different in risk prone subjects. However, among all risk prone subjects, smokers with coronary artery disease showed highest serum iron levels and decreased iron binding capacity.

Medicinal plants for treatment of diabetes mellitus.

Many plants have been used for the treatment of diabetes mellitus in Indian system of medicine and in other ancient systems of the world. Out of these only a few have been evaluated as per modern system of medicine. From many such plants only extracts have been prepared and their usefulness evaluated in experimental diabetes in animals. In some plants likeAllium cepa, Allium sativum, Ficus bengalensis, Gymnema sylvestre, Pterocarpus marsupium etc. active hypoglycemic principles have been isolated and their mechanism of action studied. Most of them seem to act directly on pancreas (pancreatic effect) and stimulate insulin level in blood. Some have extra pancreatic effect also by acting directly on tissues like liver, muscle etc. and alter favourably the activities of the regulatory enzymes of glycolysis, gluconeogenesis and other pathways. Since the plant products have less side effects, they have the potential as good hypoglycemic drugs. They may also provide clues for the development of new and better oral drugs for diabetes.

Preliminary studies on serum lipids, apolipoprotein-B and oxidative stress in xanthelasma.

Serum lipid profile, apolipoprotein-B (apo-B), malondialdehyde levels(MDA) and superoxide dismutase (SOD) activity were assessed in 12 cases of xanthelasma with and without coronary artery disease (CAD)/hypertension (HTN) and results are compared with healthy controls. Dyslipidemia was found in 65% cases of xanthelasma as compared to 20% healthy controls. Xanthelasma patients had significantly high malondialdehyde (MDA) levels (p<0.01) and significantly decreased (p<0.05) SOD activity as compared to controls. Among xanthelasma patients, xanthelasma with CAD/HTN showed higher total cholesterol (236±32.7 vs 188±24.7 mg/dl), low density lipoprotein cholesterol (157±35.5 vs 113±16 mg/dl) and Apo-B (120.5±9.4 vs 114±19.2 mg/dl) levels as compared to xanthelasma without CAD/HTN. Results of our study indicate that xanthelasma patients with increased apo-B, MDA and decreased SOD need cardiovascular monitoring.

Effect of captopril on serum angiotensin converting enzyme and blood pressure in hypertensive patients.

The effect of chronic captopril therapy on serum angiotensin converting enzyme (ACE) was studied in 30 patients with essential hypertension. Patients were assessed for serum ACE levels serially every week for 4 weeks. Thirty healthy individuals served as controls. The basal serum ACE level among hypertensives (57.4 +/- 37.2 u/l) was found to be significantly higher (p < 0.001) than the controls (33.3 +/- 8.8 u/l). One week after starting captopril therapy, serum ACE levels fell to almost half the basal values (p < 0.001). However, thereafter, it rose to levels higher than the basal level even though the blood pressure remained well controlled. Our study suggests that besides its action on ACE, captopril may lower the blood pressure by other mechanisms as well.