Angelica Kresnamurti, et al.
REDUCTION OF BLOOD GLUCOSE LEVELS OF ETHANOLIC
Reduction of Blood Glucose Levels of Ethanolic………
rise to be 5th on the world. At this time, it was reported that in big cities like Jakarta and Surabaya, was nearly 10% of the population suffer from DM (Tandra, 2008).
Indonesia's natural wealth of source material has been used by Indonesian people through generations. The culture to back to nature, or better known as Back to Nature is now a trend worldwide, including Indonesia. The result of research conducted by Andayani in 2000 showed that the crude extract of chickpea can lower blood glucose levels up to 30% in rabbits alloxan-induced diabetes. Bitter leaf extract at a dose of 0.5 g / kg bw, 1 g / kg bw, and 1.5 g / kg bw was able to inhibit the increase in blood glucose levels in normal rats (Suryadhana, 2000).
Several research reports indicated the potential of Bungur leaf extract in lowering blood glucose levels in DM patients (Liu et al., 2001, Hayashi et al., 2002, Saha et al., 2009). Based on the studies of water extract of leaves Bungur (EADB) it was shown that hypolipidemic activity was found at all doses of treatment, 0.1 g/200gBw, 0.2 g/200gBW and 0.5 g/200gBW. The dose of 0.5 g/200gBW Bungur leaf water extract treatment showed a higher hypoglycemic activity compared with glibenclamide treatment (Hernawan et al., 2004).
In this research the effect of decreasing blood glucose levels of the ethanol Bungur leaf extract at a dose 250mg/kgBW, 500mg/kgBW, and 1000mg/kgBW given orally to male Wistar albino rats that have been induced alloxan were determined. Alloxan inhibit insulin secretion from pancreatic β cells and stimulated a type 2 diabetites mellitus model, so that we could predicted the influence of ethanol extract of Bungur leaves to decrease blood glucose levels in hyperglycemic conditions, as well as the use of metformin at the dose of 63 mg / kgBW as a comparison.
METHODS Plants
Bungur leaves were collected and determined from Purwodadi Botanical Garden, Pasuruan, East Java. Bungur leaves were dried and then extracted with 70% ethanol by maceration. The extract obtained was concentrated with an evaporator.
Animals and diet
25 male Wistar Rats (Rattus norvegicus albinus), 2 months old, weight between 100-150 gram were used for the present study. They were housed in room temperature at 26±2 °C with a 12-h light and 12-h dark cycle. The animals were kept in the experimental animal room for 7 days with free access to food and water. Blood samples were withdrawn from the vein tail for glucose determinations.
Estimation of blood sugar
Blood sugar estimation was estimated by using standard glucose kit essentially followed by glucose oxidase-peroxidase (GOD-POD) methods.
Experimental protocol
25 male albino rats were induced with alloxan monohidrate i.m. at the dose of 150 mg / KgBW to create conditions of type 2 diabetes mellitus (Kresnamurti and Sugiarso, 2003). Blood glucose levels were measured every day after induction. Seven days after the occurrence of hyperglycemia, all rats were randomly divided into five groups where each group consisted of five rats, the first group was given 0.5% CMC Na, the 2nd , 3rd , 4th groups were given the suspension of the ethanolic extract at the dose of 250, 500, and 1000 mg / kgBW and the 5th group was given metformin as comparison group at the dose of 65 mg / KgBW. The Blood glucose level were measured every day until 8 days administration of extracts.
Data analysis
All data are expressed as mean ± S.E.M. ANOVA and Tukey test were used for statistical analysis. Values were considered to be significantly different when the p value was less than 0.05.
Angelica Kresnamurti
RESULTS AND DISCUSSION.
Table 1. Average of Rat Blood Glucose Levels All Groups on the 8th day after 7th Days of Treatments
No Treatment Groups ∆ Blood Glucose Level (mg/dl)
After 7th days of treatment
1 Na CMC 0.5% 184.40 ± 70.12
2 Bungur Ethanol extract 250 mg/KgBW -340.60 ± 147.06 3 Bungur Ethanol extract 500 mg/KgBW -340.80 ± 192.12 4 Bungur Ethanol extract 1000 mg/KgBW -376.40 ± 151.47
5 Metformin 63 mg / KgBW -297.40 ± 48.19
Fig 1. Percentage of blood glucose level vs time (days)
Table 2. Percentage of Reduction of Blood Glucose Level All Groups at the 8th day After 7th days of Treatments
No Treatment groups Percentage of reduction of Blood Glucose Level
1 Ethanolic extract 250 mg/KgBW 78.23 %
2 Ethanolic extract 500 mg/KgBW 80.49 %
3 Ethanolic extract 1000 mg/KgBW 83.53 %
4 Metformin 63 mg / KgBW 71.35 %
Based on the results of the study all doses of ethanol extract of leaves Bungur given orally for 7 days gave high decreasing effect of the blood glucose level of diabetic rats which more than 75%
higher than metformin which was 71.35%. Increasing doses of ethanol extract of leaves Bungur enhanced the effect of blood glucose levels, but not statistically significant by Tukey test at α > 0.05.
Blood glucose lowering effect of leaves Bungur was thought to originate from the content of the ellagitannin or ellagic acid from Bungur leaves extract that can be dissolved in 70% ethanol.
Ellagitannin is hydrolyzed tannin compound which is exactly a heksaoxidiphenate acid esters.
Ellagitanin consists of lagerstroemin, flosin B, and reginin A which has properties similar to the hormone insulin (insulin-like compound). In vitro, all three compounds are able to increase the activity
Reduction of Blood Glucose Levels of Ethanolic………
of glucose transport in adipose cells. The Ability of lagerstroemin and flosin B is almost half time the ability of insulin to increase glucose transport rate. In fact, reginin A has a capability similar to insulin (Hayashi et al., 2002). From the description it was known that the hypoglycemic activity of leaves Bungur occur through the increament of glucose transport rate. The involvement of transport speed occurs via the same pathway with the hormone insulin pathway. It is based on in vitro studies that show the work of ellagitannin in the Bungur leves which can be inhibited by wortmannin compound, i.e a compound that is able to close the path furanosteroid the hormone insulin through PI3-kinase activation barriers (phosphatidylinositol 3-kinase) in insulin signaling pathway.
It is estimated that the mechanism of action of Bungur leaves active compounds i.e. ellagitannin are as follows the active compound of Bungur leaves binds to the insulin receptor (IR) protein, causing activation of tyrosine kinase-autophosphorylation and phosphorylation of Tyr residues followed. These reactions cause the activation of insulin receptor substance that generate docking sites of SH2- containg protein molecule (a protein subunit of p85/p110 on PI 3-kinase). Then the molecule PI 3- kinase becomes active and produces PIP3 (phosphatidylinositol 3,4,5-phosphate) and then binds to the PH-domain of PDK-1 (PIP3-dependent kinase) and Akt (protein Ser / Thr kinase B). This reaction lead to Akt and PDK-1 becomes active. Active Akt molecule can cause the translocation of GLUT4 proteins that mediate the glucose transport mechanism.
From these findings, it is very likely that the hypoglycemic activity of bungur ethanolic extract is derived, at least in part, from the decrease in insulin resistance, presumably because of the increase in GLUT4 translocation in total muscle membrane. These results suggest that the clinical use of bungur ethanolic extract in the treatment of diabetes mellitus, especially type 2 diabetes, may be appropriate.
ACKNOWLEDGEMENT
Researchers would like to acknowledge the advice and input provided by Dr. Junaidi Khotib, Dr.
Bambang Prayoga, and Dr. Ratna Megawati Widharna on this project.
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