Anti-Diabetic, Antidyslipoproteinemic and Antioxidant Activities of Tinospora Cordifolia

ANTI-DIABETIC, ANTIDYSLIPOPROTEINEMIC AND ANTIOXIDANT ACTIVITIES OF TINOSPORA CORDIFOLIA IN DIABETIC TYPE 2 PATIENTS

Vishnu Kumar1, Farzana Mahdi1, Dharam Raj Singh2 , Ramesh Chander1, Ashok Kumar Khanna4, Jitendra Kumar Saxena4 , Ranjana Singh3, Abbas Ali Mahdi3, and Raj Kumar Singh5.

1Department of Biochemistry Era’s Lucknow Medical College & Hospital Lucknow-226 001 2Kaya Chikitsha Vibhag, Rajkiya Ayurvedic Chikitsha Mahavidyalaya, Touriya Ganj, Lucknow - 226 003. 3Department of Biochemistry, CSM Medical University Lucknow-226 003 4Division of Biochemistry, Central Drug Research Institute, Lucknow-226 003 5Shri Guru Ram Rai Institute of Medical & Health Sciences, Patel Nagar, Dehradun-248 001. Inrtoduction Tinospora cordifolia (Menispermaceae) is widely used in Ayurvedic medicine as a remedy for metabolic disorders however, anti-diabetic and anti-dyslipoproteinemic activities are not well studied in diabetics this study is carried out to explore these effects in diabetic type 2 patients. Aims and Objectives To explore anti-diabetic and anti-dyslipoproteinemic activities of T. cordifolia in Diabetic type 2 patients as well as healthy human volunteers. For This purpose fasting blood glucose, serum lipid and lipoprotein profile were analyzed in serum.

Methodology Selection Diabetic Subjects: The study was carried out on 60 subjects attending outpatient department of, Kaya Chikitsha Vibhag, Rajkiya Ayurvedic Chikitsha Mahavidyalaya, Touriya Ganj, Lucknow 60 diabetic subjects (M = 30, F = 30 as well as normal healthy control (M = 15, F = 15) who attended for their periodic health checkups 30 ((M = 15, F = 15) from families of these patients as per guide line of ethics in supervision of Kaya Chikitsha Vibhag. All biochemical assays were done by standard kit methods. Exclusion Criteria: Patients with evidence of acute or chronic inflammatory or infectious disease, cancer, persons on insulin, or other medications that could affect glucose metabolism and pregnant or lactating women were excluded from the study.

Inclusion Criteria: All individuals were subjected to a complete medical evaluation by a physician including a full medical history and physical examination. Both males and females between 40 - 60 years of age, weight 60 – 70 kg were included in the study. Material and Methods Collection of blood sample: Blood sample were collected from cubital vein followed by overnight fasting, for biochemical estimations in fluoride (sodium fluoride and potassium oxalate, 5.4 mg NaF and 3.0 mg K-oxalate in each vial) and plain vials respectively. Preparation of plant powder: Healthy ( free from diseases insecticidal & pesticidal effects) T. cordifolia stems were collected from local area of Lucknow, check and identified taxonomically with the help of Taxonomist and plant pathologist by the Department of Pharmacology, Era’s Lucknow Medical College & Hospital, Lucknow and a voucher specimen was also submitted (TC-001/06). Stem was dried under shade and made into fine powder using laboratory mill. Biochemical analysis of blood and plasma lipoproteins: The blood was centrifuged and plasma was separated. The glycosylated hemoglobin (HbA1C) in RBC and plasma total lipid were assayed by standard spectrophotometric methods (12, 13). Plasma was also used for the assay of lecithin cholesterol acyl transferase activity (LCAT) (14), glucose (10), and lipid peroxide (17) (LPO), SOD, Catalase and GSH. A portion of plasma was fractionated into very low density lipoprotein (VLDL), low density lipoprotein (LDL) and high density lipoprotein (HDL) by polyanionic precipitation methods (18). Plasma as well as lipoproteins were measured for their total cholesterol (19) (TC), phospholipids (20) (PL), triglyceride (21) (TG) and protein (22) by standard spectrophotometric methods. Results:

Effect of T. cordifolia stems powder on blood glucose, Glycated Hemoglobin, Lecithin Cholesterol Acyl Transferase and serum lipid profile in diabetic type-2 patients.

Serum analysis of diabetic type-II patients showed marked increase in blood glucose (1.87 Fold), HbA1c, LCAT, TC (1.34 Fold), TG (1.59 Fold), LDL-TC (1.58 Fold), VLDL-TC (1.59 Fold), lipid peroxide (3.52 Fold) following decrease in the level of serum HDL-TC (39%) (Table 1)

Effect of T. cordifolia stems powder on lipoprotein profile in diabetic type 2 patients

Diabetes type 2 causes increase in the lipids and apoprotein levels of serum β lipoproteins these are very low density lipoprotein (VLDL) and low density lipoprotein (LDL) following decrease in lipid and protein constituents of α lipoprotein this is high density lipoprotein (HDL) were observed in Diabetic type 2 patients with respect to healthy human volunteer. However, treatment with T. cordifolia powdered stem for 15 days significantly decrease the fasting blood glucose level (15.0%), serum TC (12%), serum TG (18%) serum LDL-TC (11%), serum VLDL-TC (18%), serum lipid peroxide (42%) following recovery of HDLTC (4.0%) in diabetic type 2 patients. The decrease of lipids and apoprotein levels of β lipoproteins were followed by stimulation of plasma lecithin cholesterol acyltransferase. Lipid and apoprotein level of α lipoprotein (HDL) were also recovered partially on treatment with powder T. cordifolia (Table 2). On the other hand these effect of T. cordifolia on healthy human volunteers were non significant.

Effect of T. cordifolia stems powder on GSH, serum lipid peroxide SOD and Catalase in diabetic type 2 patients Diabetes type 2 causes increase in LPO, as well as decrease in GSH, SOD and Catalase. However

Statistical analysis: One-way-analysis of variance (ANOVA- Newman’s student test) was performed by comparison of values for alloxan-treated group with control, alloxan and drug-treated with alloxan only. All hypothesis testing were two-tailed. P <0.05 was considered statistically significant and the results were expressed as mean ± SD. The Graph pad INSTAT 3.0 software was used to carried out the statistical analysis.

Discussion:

It is well documented that hyperglycemia induced pathologic changes also suppresses the synthesis of glucosominaglycons in capillary endothelium surface that lead to defect in LPL binding and consequent poor clearance of VLDL in diabetics (33). This may be true; the diminution of capillary endothelial and hepatic lipases had been involved to produce hyper  -lipoproteinemia; and their reactivation by the treatment with T. cordifolia had played a significant role in regulation of lipoprotein metabolism in diabetic rats. Further more, structural modifications in lipoproteins would have made them a defective substrate for their catabolism through LPL and hepatic LDL receptors. The treatment with T. cordifolia and glibenclamide might also improved antioxidatve status in animals following inhibition of oxidative changes in body biomolicules including to that of LDL, facilitating its catabolism through hepatic receptors. Induction of diabetes adversely affects on HDL, as it caused decrease in levels of lipids and apoprotein components of this lipoprotein by 24-27% (Table 5). Hyperglycemia also interfere with function of CETP and LCAT, which are reported to be responsible for abnormalities with HDL metabolism and appearance of high atherogenic sd-LDL particles in diabetics (28 ). In the present study, we found that treatment with T. cordifolia but not glibenclamide could stimulated LCAT enzyme and partially recovered the level of HDL in diabetic rats. The early studies have shown that feeding with T. cordifolia root alcoholic-extract caused lowering in blood sugar levels, serum and tissue lipids in alloxan induced diabetic rats (34). However the alcoholic extract of leaves of this plant, though showed hypoglycemic activity but failed to exert lipid lowering action in above model (35). The chemical constituents reported from stem of this shrub belong to different classes such as alkaloids, diterpenoid lactones, glycosides, steroids, sesquiterpenoid, phenolics, aliphatic compounds and polysaccharides, root contains alkaloids, and however the leaf contains only steroids (36). Furthermore, stem as well as root both contain alkaloids; Berberine. Palmatine, Tembetarine, Magnoflorine, Choline, Tinosporin Isocolubin, Palmatine, Tetrahydropalmatine, Magnoflorine.Therefore ,to that of root (34) antidiabetic and antidyslipidemic activities of T. cordifolia stem may be by part due to presence of these typical alkaloids. In conclusion, T. cordifolia might suppress hyperglycemia induced alterations in biochemical path ways that are responsible to cause abnormities with lipid metabolism in diabetic rats. Besides its antidiabetic and antilipoperoxidative effects, T. cordifolia may have regulated functioning of various enzymes and metabolites to afford a normal lipid metabolism in dyslipidemic animals. This may be due to reactivation of PHLA, LCAT and tissue LPL enzymes. Treatment caused beneficial effect on HDL synthesis that may also be contributed to regulate lipid metabolism. T. cordifolia enhanced excretion of bile acids through feces and thus contributed to regress the hepatic cholestesteosis in diabetic rats. The study reveals that T. cordifolia is a better drug as a natural product to regress diabetic-dyslipidemia and oxidative stress in diabetes. Further work to assess the antidyslipidemic activity of different fractions of T. cordifolia stem in alloxan induced diabetic rats is under progress to substantiate the present findings.

Conclusion:

The results of the present study leads to research and development of a potent anti diabetic, antidyslipoproteinemic antioxidant drug, from T. cordifolia stem powder.