Antihyperglycemic and Antihyperlipidemic Activity of Acacia suma (Roxb.) Barks

S. Acharyya¹*, G.K. Dash², S. Pattnaik³ and R.R Chhetree⁴

¹Matushree V. B. Manvar College of Pharmacy, Dumiyani, Rajkot, Gujarat -360 440.

²Institute of Pharmacy and Technology, Salipur, Cuttack, Orissa -754 202.

³Department of Zoology, Berhampur University, Berhampur, Orissa -760 007.

⁴Regional Institute of Pharmaceutical Science and Technology, Abhoynagar, Agartala, West Tripura-799005.

ABSTRACT:

Methanolic bark extract of Acacia suma was evaluated for its antihyperglycemic and antihyperlipidemic activity in alloxan induced diabetes model in rats. The extract was given orally in two different doses (200mg/kg and 400mg/kg) for 28 days and for comparison purpose, Glibenclamide (4mg/kg), an oral hypoglycemic was used as a standard drug. Parameters under study were fasting blood glucose, serum lipids, glycosylated haemoglobin and liver glycogen. The extract showed significant antihyperglycemic activity in dose dependent manner. The orally administered extract significantly reduced elevated lipids and glycosylated haemoglobin in diabetic rats. The extract significantly improved glucose tolerance, body weight and liver glycogen of diabetic rats. It was observed that both glibenclamide and Acacia suma significantly reduced blood glucose as well as produced favorable changes in lipid profile. It was observed that methanolic bark extract of Acacia suma has potential to prevent the secondary complications of diabetes mellitus like atherosclerosis

KEYWORDS: Acacia suma, Glibenclamide, Antidiabetic, Antihyperlipidemic.

INTRODUCTION:

Diabetes mellitus is a common metabolic disorder characterized by chronic hyperglycemic due to either insufficient insulin production by pancreatic beta cells or by cellular resistance to insulin. It leads to disturbances of carbohydrate, protein and fat metabolism¹. Diabetes has significantly accelerated levels of oxidative stress which contributed massively to most neurological, cardiovascular, retinal and renal diabetic complications². Diabetes affects 5% of the worlds population and worldwide projection suggests that more than 300 million people will have diabetes by the year 2025³.

Long term complications arising due to diabetes mellitus and serious side effects shown by synthetic hypoglycemic agents^4,5has continued the search for more effective and safer antidiabetic agents. Indian Medicinal system is one of the richest system among those available around the world and it has treated diabetes with its herbals for ages^6,7. Since ancient ages people are using medicinal plants as home remedies for treatment of diabetes⁸.

Acacia suma (Roxb.) var. Acacia polyacantha (Family-Fabaceae) is a medium sized erect tree; trunk with fissured bark and knobby persistent prickles found in the greater part of India and costal districts of Orissa^9,10. The bark is reported to be used as blood purifier¹⁰ and possesses anti-cancer, insecticide and astringent properties^11-14. The seeds are reported to have hypoglycaemic effect¹¹.The leaves and roots of the plant are reported to be use as insecticide, antifungal, antivenin, aphrodisiac, antimalarial, anticrustacean, stimulant and in the treatment of sores, abcesses and asthma^14-20.Presence of proanthocyanidin¹¹, 5,4’-dihydroxy-7, 3’-dimethoxyflavone-3-0-D galactopyranoside^12,21, gallocatechin-5-7-digallate, quercetin and gallocatechin-7-gallate¹³in the barks have been reported earlier. The tribes of Ganjam district of Orissa grind the fresh bark and drink its suspension in water daily to reduce blood sugar in the patients with diabetes mellitus since time immemorial and they claim for its promising activity. Some experimental studies regarding antidiabetic activity have been carried out with this plant, but most of them have evaluated acute antidiabetic effect of the plant. The antihyperlipidemic activity of the extract is not reported. The present study of 28 days was therefore undertaken to evaluate long term antidiabetic effect of methanolic bark extracts of Acacia suma in alloxan induced diabetic rats and to evaluate its effect on lipid profile.

MATERIALS AND METHODS:

Preparation of Plant Extract:

The plant material (barks) was collected from the forests of Ganjam district of Orissa during June 2007 and identified by the taxonomists of the Botanical Survey of India, Shibpur, Howrah. A voucher specimen [Sp. No: CNH/ I-I / (17)/2009/Tech.II/28] has been kept in our research laboratory for further reference. After authentication, fresh barks were collected in bulk, washed, shade dried and pulverized in a mechanical grinder to obtain coarse powder. The powdered material was subjected to extract with methanol for 48 h in a soxhlet extractor. The liquid extract was concentrated under vacuum to yield dry extract. It was weighed and preserved for further use.

Animals:

Male wistar rats (150-200 g) were acclimitised for the period of 10 days at room temperature. The animals were kept in standard polypropylene cage and maintained under standard environmental conditions: 12:12 h light: dark cycle and with free access to food and water ad libitum. All the procedures were performed in accordance to Institutional Animal Ethics Committee.

Experimental Design:

a) Induction of Diabetes:

The method of Perfumi et al., 1996 was followed ²². The acclimatized animals were kept fasting for 24 hours with water ad libitum and injected intraperitoneally a dose of 150 mg/kg of alloxan monohydrate in normal saline²³. After one hour, the animals were provided standard laboratory diet ad libitum. The fasting blood glucose was estimated after 48 h of alloxan administration to confirm the diabetic state. Rats showing fasting blood glucose more than 225 mg/dl²⁴ were considered diabetic and used for the study.

b) Experimental Groups:

Rats were divided in following groups of 6 each as follows:

Group 1 : Control group

Group 2 : Diabetic control group

Group 3 : Diabetic group treated with 200 mg/kg bark extract

Group 4 : Diabetic group treated with 400 mg/kg bark extract

Group 5 : Diabetic group treated with 4 mg/kg of glibenclamide

Group 6 : Normoglycemig rats treated with 400 mg/kg extract.

c) Experimental Procedure:

The extracts/ drug treatment was given orally for 28 days after which rats were kept for overnight fasting with free access of water. During the experiment, evaluation of following parameters was done-

i) Blood glucose

ii) Serum lipid profile

iii) Liver glycogen

iv) Glycosylated haemoglobin

v) Body weight and organ weight (liver, kidney and pancreas).

Blood glucose levels were estimated after every week by collecting the blood from the tip of the tail of the rats and using glucostix (Senso card blood glucose meter supplied by M/s Avecon health care Pvt. Ltd., Himachal Pradesh).

Lipid profile was done after 28 days of study and estimation of serum parameters like total cholesterol, triglycerides, and high density lipoproteins was done using commercial kits (Span Diagnostics, Mumbai).

For serum glycogen content, the liver was homogenized in 5%w/v trichloroacetic acid and its glycogen content was determined by the method of Caroll et al ²⁵.

Two days before the termination of the experiment, the oral glucose tolerance test was performed to assess the glucose tolerance. For this purpose, overnight fasted rats were fed orally 2g/kg body weight glucose. Blood was collected at 0, 30, 60, 90, and 120 min interval for glucose estimation ²⁶.

During the experiments, body weight was measured after every week and weight of organs i.e. liver, kidney and pancreas was measured after the termination of the experiment.

Statistical analysis:

All the data is expressed as mean ± SEM. The differences between groups were evaluated by one way- Analysis of Variance (ANOVA) followed by Dennett’s Multiple Comparison test. P<0.05 was considered significant.

RESULTS:

i) Effect of bark extract of Acacia suma on fasting blood glucose in experimental rats:

As shown in Table 1, the extract showed significant decrease in blood glucose level gradually. In the diabetic rats blood glucose level was reduced from 300.12mg% to 198 and 187.8mg% with 200 and 400mg/kg doses respectively on the 28^th day. Glibenclamide (4 mg/kg) reduced blood glucose level to 117.47mg%. The effect of bark extract (400 mg/kg) was comparable to that of glibenclamide while that of 200mg/kg was lower (Graph1).

Graph 1: Effect of bark extract of A. suma on fasting blood glucose in experimental rats.

n=6; Data expressed as mean ± SEM. Evaluation by one way-Analysis of Variance

(ANOVA) followed by Dunnett’s Multiple Comparison test.

ii) Effect of bark extract of Acacia suma on oral glucose tolerance test (OGTT) in experimental rats:

As shown in Table 2, chronic administration of drug extract significantly inhibited the rise in blood glucose level in glucose loaded rats. The fasting blood glucose level of diabetic rats was higher even after 2 h of glucose load. However rats treated with bark extract and glibenclamide showed improvement in oral glucose tolerance in which the significant fall in blood glucose was observed after 60 min of glucose load.

iii) Effect of bark extract of Acacia suma on lipid profile, glycogen content and glycosylated haemoglobin in experimental rats:

As shown in Table 3, the diabetic rats showed elevated levels of serum cholesterol, triglycerides and LDL-CH. Oral treatment with 400mg/kg methanolic bark extract of Acacia suma significantly reduced the above stated lipid parameters and the effect was comparable to that of glibenclamide (Graph 2). The alloxan induced diabetic rats showed significant reduction in liver glycogen and HDL-cholesterol. As shown in Table 4, oral treatment with 400mg/kg methanolic bark extract of Acacia suma significantly improved the glycogen content from 0.87g/100g to 2.84g/100g, the effect was comparable to that of glibenclamide which restored glycogen content to 3.11g/100g. Glycosylated haemoglobin levels were reduced from 9.99% in diabetic rats to 8.36% and 7.22% in rats treated with 200 and 400mg/kg methanolic bark extract of Acacia suma respectively. The glycosylated haemoglobin levels were found to be 6.72% in glibenclamide treated diabetic rats (Table 4).

Graph 2: Effect of bark extract of A. suma on lipid profile in experimental rats.

T-CH: Total cholesterol; TG: Triglycerides; HDL-CH: HDL cholesterol;

LDL-CH: LDL cholesterol; VLDL-CH: VLDL cholesterol

n=6; Data expressed as mean ± SEM. Evaluation by one way-Analysis of Variance

(ANOVA) followed by Dunnett’s Multiple Comparison test.

Table 1: Effect of bark extract of A. suma on fasting blood glucose in experimental rats.

Experimental groups	Fasting Blood Glucose (mg/dl)
Experimental groups	0 Day	7 Day	14Day	21Day	28Day
Control	80.02±9.9	78.07±10.8	80.07±12.3	81.47±12.4	81.35±10.8
Diabetic control	292.27±17.8	293.3±19.1	295.95±20	297.5±20.8	300.12±23.1
Diabetic + 200mg extract	293.07±18.8	281.8±20.2	260.32±21.4	242.1±23.7	198±15.9**
Diabetic + 400mg extract	290.67±16.5	266.2±19.6	239.8±21.7	211.7±22.5*	187.8±25.4**
Diabetic + Glibenclamide	288.57±12.7	251.7±16.80	213.9±19.1*	152.9±13.7**	117.47±22.3**
Control + 400mg extract	79.32±4.5	76.75±5.3	78.67±5.5	75.5±5.9	70.45±6.3

Data expressed as mean ± SEM. Evaluation by one way-Analysis of Variance (ANOVA) followed by Dunnett’s Multiple

Comparison test. *P<0.05, **P<0.01 as compared to diabetic control.

Table 2: Effect of bark extract of A. suma on oral glucose tolerance test (OGTT) in experimental rats.

Experimental groups	Time (min)
Experimental groups	0min	30 min	60 min	90min	120min
Control	75.97±2.7	177.05±6.6	141.77±7	127.35±5.8	93.6±4.1
Diabetic control	298.95±17.8	368.5±18.5	358.4±19.8	340.42±19.1	321.22±18.7
Diabetic + 200mg extract	243.1±16.5	312.08±19.5	295.17±21.4	285.11±19.7	267.62±18.7
Diabetic + 400mg extract	212.14±18.2	294.67±21.5	278.47±16.4*	252.35±16.4**	227.21±13.4**
Diabetic + Glibenclamide	155.5±19.2	241.6±26.3	229.06±16.2**	198.14±21.5**	179.16±19.8**
Control + 400mg extract	77.47±9.3	165.05±11.8	148±7.02	129.72±12.04	85.72±5.6

Data expressed as mean ± SEM. Evaluation by one way-Analysis of Variance (ANOVA) followed by Dunnett’s Multiple

Comparison test. *P<0.05, **P<0.01 as compared to diabetic control.

Table 3: Effect of bark extract of A. suma on lipid profile in experimental rats.

Experimental groups	T-CH	TG	HDL-CH	LDL-CH	VLDL-CH
Control	75.02±7.5	44.92±4.8	24.2±2.7	4102±2.4	8.49±0.41
Diabetic control	197.02±4.5	155.8±5.2	16.4±2.2	151.96±4.1	31.22±1.8
Diabetic + 200mg extract	160.02±3.9**	123±3.5**	21.3±2.1	121.0±12.4*	22.67±2.8*
Diabetic + 400mg extract	113.82±5.7**	83.27±4.3**	25.12±1.7*	75.45±5.9**	17.15±2.11**
Diabetic + Glibenclamide	100.8±4.9**	77.25±4.8**	27.65±2.2**	59.52±5**	16.57±1.8**
Control + 400mg extract	77.07±8.11	45.57±2.7	26.53±3.3	43.25±5.4	9.55±1.5

T-CH: Total cholesterol; TG: Triglycerides; HDL-CH: HDL cholesterol; LDL-CH: LDL cholesterol; VLDL-CH: VLDL cholesterol

n=6; Data expressed as mean ± SEM. Evaluation by one way-Analysis of Variance (ANOVA) Followed by Dunnett’s Multiple Comparison test. *P<0.05, **P<0.01 as compared to diabetic control.

Table 4: Effect of bark extracts of A. suma on liver glycogen and glycosylated haemoglobin in experimental rats.

Experimental groups	Liver glycogen (g/100g)	Glycosylated haemoglobin (%)
Control	3.77±0.12	5.9±0.07
Diabetic control	0.87±0.03	9.99±0.17
Diabetic + 200mg extract	2.15±0.09**	8.36±0.15**
Diabetic + 400mg extract	2.84±0.1**	7.22±0.1**
Diabetic + Glibenclamide	3.11±0.09**	6.72±0.14**
Control + 400mg extract	3.92±0.08	5.38±0.21

n=6; Data expressed as mean ± SEM. Evaluation by one way-Analysis of Variance (ANOVA) followed by Dunnett’s Multiple Comparison test. *P<0.05, **P<0.01 as compared to diabetic control.

Table 5: Effect of bark extract of A. suma on body weight and organ weight in experimental rats.

Experimental groups	Body weight (g)	Organ weight (g)
Experimental groups	Body weight (g)	Liver	Kidney	Pancreas
Control	261.02±4.5	7.23±0.12	1.13±0.1	0.56±0.04
Diabetic control	172.75±6.9	4.27±0.11	0.75±0.04	0.39±0.01
Diabetic + 200mg extract	181.35±4.7	5.95±0.23**	0.85±0.05	0.4±0.02
Diabetic + 400mg extract	192.02±5.9	6.16±0.17**	0.97±0.04*	0.42±0.01
Diabetic + Glibenclamide	204.72±5.8*	6.98±0.16**	0.99±0.04*	0.46±0.02
Control + 400mg extract	260.95±6.1	7.28±0.22	1.14±0.06	0.54±0.03

n=6; Data expressed as mean ± SEM. Evaluation by one way-Analysis of Variance (ANOVA) followed by Dunnett’s Multiple Comparison test. *P<0.05, **P<0.01 as compared to diabetic control.

iv) Effect of bark extract of Acacia suma on body weight and organ weight in experimental rats:

Alloxan diabetic rats show significant reduction in body weight from 261.02 to 172.75 g as compared to control group (Table 5). Oral administration of methanolic bark extract and glibenclamide significantly improved the body weight after 28 days. Also there is no further reduction in organ weight (liver, kidney, pancreas) of extract treated group as compared to diabetic control group.

v) Phytochemical analysis of bark extract of Acacia suma

The extract was tested for phytochemical constituents^27,28and showed presence of alkaloids, terpenoids, flavonoids and saponins.

DISCUSSION:

In present study, antidiabetic activity of methanolic extract of Acacia suma was evaluated in Wister rats using alloxan induced diabetes model. Rats treated with alloxan developed a diabetic state which exhibited symptoms of insulin dependent diabetes mellitus like hyperglycemia, glycosuria and loss of body weight. However the animals survived without insulin treatment indicating incomplete destruction of pancreatic islet cells by alloxan treatment. Diabetes mellitus is known to cause hyperlipidemia through various metabolic derangements amongst which insulin deficiency has been known to stimulate lipolysis in the adipose tissues and give rise to hyperlipidemia and fatty liver. Thus in diabetes hyperlipidemia and hypertriglyceridaemia often occur²⁹. The subsequent hyperlipidemia shown by diabetic rats can be used as an index for hyperglycemia.

The present study showed significant reduction in i) blood glucose on different days ii) glycosylated haemoglobin levels iii) serum cholesterol and triglycerid levels in Acacia suma bark extract and glibenclamide treated diadetic rats. Rats treated with glibenclamide and Acacia suma bark extract showed improvement in liver glycogen, HDL cholesterol and body weight as compared to diabetic control group. Glycosylated haemoglobin present time averaged values for blood glucose over the last or next few months thus providing tool for assessing treatment efficacy and patient compliance. Further the methanolic bark extract of Acacia suma has shown ability to enhance the glycogenesis process in the liver of diabetic rats. Ability of extract to improve body weight in diabetic rats shows its possible ability in restoration of protein metabolism.

Methanolic bark extract of Acacia suma when administered orally to diabetic rats significantly reduced the levels of LDL cholesterol and increased that of HDL cholesterol. The facilitation of atherogenesis by LDL cholesterol is due to its role in depositing cholesterol in the vascular bed. HDL cholesterol however carries out the reverse transport of excess cholesterol from cells of tissues to the liver. Thus along with antidiabetic activity, the methanolic bark extract of Acacia suma has potential to prevent the formation of atherosclerosis and coronary heart disease which are the secondary complications of diabetes mellitus.

ACKNOWLEDGEMENTS:

The authors are thankful to the management of Matushree V. B. Manvar College of Pharmacy, Dumiyani, Rajkot district, Gujarat for providing necessary facilities to carry out the present research works.

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Received on 27.12.2010

Accepted on 17.01.2011

Research J. Pharmacology and Pharmacodynamics. 3(2): March –April, 2011, 67-71