Oral Administration of Extract from Curcuma longa Lowers Blood Glucose and Attenuates Alloxan-Induced Hyperlipidemia in Diabetic Rabbits

MATERIALS AND METHODS

Plant collection, authentication and extraction: Large quantities of Curcuma longa rhizormes and leaves were purchased from Oja-Oba Market in Ibadan and were identified by the Herbarium in Botany Department of the University of Ibadan Nigeria. These were air dried at room temperature in the laboratory and were powdered using the Hammer mill in Agronomy Department of the University of Ibadan. Powdered Curcuma longa (373 g) was Soxhlet extracted for 72 h using absolute Ethanol and the concentrated extracts were stored in sample bottles. The animals in group A were given 250 mg/kg plant extract per body weight for twelve consecutive days.

In vivo antidiabetic assays
Animal: Twelve healthy male rabbits weighing between 1.10 and 1.50 kg were purchased from Sasa Market in Ibadan, Nigeria. The rabbits were fed with pellets of growers mash manufactured by Guinea Mills, Ibadan and were allowed free access to water. The animals were fed for one month and were given ascorbic acid tablets for acclimatization in the Animal House of Biochemistry Department, University of Ibadan, Nigeria. The animals were placed into three groups of four rabbits each:

Biochemical and anti-hyperglycemic assays: Diabetes was induced in the rabbits by a single inter-peritoneal injection of 100 mg/kg of Alloxan. Blood samples were collected from the ears of the rabbits into EDTA bottles containing anticoagulant and the samples were centrifuged at 3000 rpm for 10 min to obtain plasma which was stored in a refrigerator until it was used. The plasma glucose was determined by the quantitative enzymatic hexokinase determination in plasma at 520 nm wavelength using the Sigma glucose oxidase kit. Triglycerides were analyzed using enzymatic hydrolysis of triglycerides by determining colorimetrically the glycerol produced using Randox kit. Protein concentration was determined using the photometric colorimetric test (Biuret method) available in Human kit. Measurement of creatinine clearance using difference in color intensity measured at 500 nm before and after acidification was achieved using Sigma kit. Plasma cholesterol was determined using enzymatic hydrolysis and oxidation of cholesterol with quinonemine as the indicator formed from hydrogen peroxide and 4-amino antipyrine using Randox kit.

RESULTS AND DISCUSSION

Diabetes mellitus is associated with hyperglycemic effect, increased urinary output and weight loss and these were observed in diabetes induced by Alloxan (Weaver et al., 1978). Figure 1 show mean plasma glucose level (mg/dl) in diabetic animals and diabetic rabbits administered Tumeric for 12 consecutive days. There was a significant increase in blood sugar in the group A compared to the Group D, implying an increase in plasma glucose in diabetic rabbits and a significant decrease in the plasma glucose level of test rabbits administered curcumin. This result shows C. longa has a possible antihyperglycemic agent, confirming previous reports on this plant (Khan et al., 1990). The report of C. longa as a potent anti-oxidant may suggest it reaction with free radicals produced by alloxan inactivating them by activating glucokinase (WHO, 1980; Szkudelski, 2001; Weaver et al., 1978). The overall effect of the presence of reactive oxygen species leads to hyperlipidemia as there is an accumulation of triglycerides and cholesterol, due to damage to pancreatic islets and liver cells. (Lenzens-Bruning and Monster, 1978). Obtained data for mean total cholesterol and total triglycerides in mg/dl in this study are shown on Fig. 2 and 3 respectively. Figure 2 showed a significant increase in total cholesterol in the diabetic control when compared to the normal and the diabetic treated rabbits and in the Tumeric treated group did not change, the cholesterol decreased significantly especially on days 9 and 12. Triglyceride level was remarkably higher on days 6-12 compared to the normal and the test rabbits and this implies that the plant drug exhibited hypolipidemic effect. C longa has hypocholesteric effect by its use in treating liver disorders (Ammon and Wahl, 1991; Lewis, 1972 and Nippon, 1993) have reported that increased cholesterol level in diabetes mellitus is due to enhanced activity of HMG-CoA reductase and Cholesterol Acyl Transferase (ACAT) which are the major enzymes in cholesterol biosynthesis.

The effect of Curcuma longa on plasma protein concentration (g/L) at 565 nm is shown on Table 1. The result indicates a significant increase in protein concentration with regards to control (p<0.05) and the plant drug has slightly decreased effect n the plasma protein level in the diabetic treated rabbits (A).

The plasma creatinine level in μmdl at 500 nm is shown in Table 2. There was a significant increase in the creatinine concentration with respect to the control with p<0.05.

Diabetes causes a significant increase in creatinine synthesis. There was an increase in the value of test rabbits compared to the normal rabbits and a significant decrease in the values of treated rabbits, when compared with control especially from days 6-12.

Table 2 shows that Curcumin reduced the total protein concentration as it significantly reduced the creatinine level of alloxan diabetic rabbits, thus implying hypoprotein properties.

Table 1:	The effect of Curcuma longa on plasma protein concentration (mg/dl) in alloxan diabetic rabbits

Table 2:	Effect of Curcuma longa on creatinine level (μm/dl) of alloxan diabetic rabbits

Fig. 1:	Effect of curcuma longa on plasma glucose level in alloxan-diabetic rabbits

Fig. 2:	Effect of curcuma longa on plasma cholesterol of alloxan diabetic rabbits plasma cholesterol in mg/dl

Fig. 3:	Effect of curcuma longa on plasma triglyceride levels in alloxan diabetic rabbit (Tg in mg/dl)

Diabetes is associated with polyphagia (increased appetite), polydipsia (increased thirst), polyuria (increased urine) as well as general inactivity. These symptoms were markedly reduced in dietetic rabbits treated with Tumeric. Diabetes has no cure, but effective control of blood sugar levels is beneficial especially in terms of prolonging life and reducing some long term complications. We have shown in this study to C. longa reduces plasma glucose level in diabetic rabbits as well as secondary symptoms that accompany diabetes such as hypercholesterolemia and hyperlipidemia.

ACKNOWLEDGMENTS

The author for correspondence is grateful to TWAS for postdoctoral fellowship in Centres of Excellence in the South, utilized in HEJ Research Institute of Chemistry, University of Karachi, Karachi, Pakistan, where this work was presented as poster in 9^th International Symposium on Protein-Structure function relationships held in the same institute in January 2007.