ANTI-DIABETIC ACTIVITY OF RIBES NIGRUM FRUIT EXTRACT IN ALLOXAN INDUCED DIABETIC RATS

ANTI-DIABETIC ACTIVITY OF RIBES NIGRUM FRUIT EXTRACT IN ALLOXAN INDUCED DIABETIC RATS

G. Sivalingam* and N. Sriram

Smt. Sarojini Ramulamma College of Pharmacy, Seshadrinagar, Mahabubnagar, Andhra Pradesh, India

ABSTRACT:

Different doses of Ethanolic fraction of Ribes nigrum fruit were evaluated for hypoglycemic activity in Alloxan induced diabetic rats. The oral administration of Ethanolic extract at a dosage of 100mg/kg and 500mg/kg body weight exhibited significantly reduced the serum glucose level in both acute (1, 3, 5 h) and sub-acute (1, 3, 5, 7 days) treatments. Perform the Histopathological study of pancreas.

Ribes nigrum, Antidiabetic activity, Alloxan-induced diabetic

INTRODUCTION: Diabetes mellitus is a common metabolic disorder characterized by hyperglycemia, glycosuria, polyurea and polydipsia induced by insulin deficiency ¹ and insulin resistance ².Recent estimates indicate that there were 171 million people in world with Diabetes in year 2000 and this may be projected to increase to 366 millions by 2030 ².

Diabetes mellitus is treated by using Oral hypoglycemic agents such as Sulphonyl Ureas. Biguanides, Meglitinides and Alpha glucosidase inhibitors. Traditional medicines like herbal drugs in primary form or their extractshave been used by many diabetic patients as they are assumed to be non–toxic in nature

About plant: Ribes nigrum is a common name ofBlackcurrant it belongs to Grossulariace family. It is geographically distributed in northern India, northern Asia, and central Europe It is a small shrub, growing to 1–2 m tall. The fruit is an edible berry 1 cm diameter, very dark purple in colour, almost black, with a glossy skin and a persistent calyx at the apex, and containing several seeds dense in nutrients.

An established bush can produce up to 5 kilograms of berries during summer. ³

MATERIAL AND METHODS:

Collection of plant material:The fruit of Ribes nigrumwere collected and authentication was done by botanist Dr. Madhava chetty, Assistant Professor, Department of Botany in S.V. University, Titupathi. The voucher no. 1006

Preparation of the extracts: The plant materials were dried in the shade and powdered by a mechanical grinder. The powder of  Ribes nigrum fruits was extracted with 90% Ethanol.powedered material and solvent was used at 1:4 ratio, at 65-70⁰C  by using a Soxhlet extractor for 12 hours at a temperature not exceeding the boiling point of the solvent. The extract was filtered using whattman filter paper (No 1) and then concentrated in a vacuum and dried at 45˚C for ethanol elimination. The extracts were kept in a sterile bottle under refrigeration conditions of about 2-8˚C ^{4, 5}.

The percentage yield (20%) was calculated. The extracts were used for further studies and determination of activity.

Animals: Wister albino rats (150–220 g) of male were purchased from Sainath Agencies, 1-6-197/45/D, Bapujinagar, Musheerabad, (Reg. No. 282/99/CPCSEA), Hyderabad 500048, India and Maintained under standard environmental laboratory conditions and fed with laboratory diet and water ad libitum. All the protocols were performed in accordance with the Institutional Animal Ethical committee (IAEC) as per the directions of the CPCSEA (Committee for the Purpose of Control and Supervision of Experiments on Animals) All the pharmacological studies are carried out at Smt. Sarojini Ramulamma college of Pharmacy. (Reg. No: 51/01/c/cpcsea/2012/16).

Limit test:Albino Wistar Rats are taken for the study of limit test. Three animals treated with 2000mg/kg as a single dose via oral route as per the limit test of OECD guide lines 423 and. The rats were observed continuously for 24 h for behavioral, neurological, and autonomic profiles and, after a period of 24 and 72 h, for any lethality, morbidity state or death. The animal not showed any signs of toxicity and behavioral changes after 24 hrs and 72 hrs ⁶.

Hypoglycemic activity:

Induction of diabetes: Alloxan monohydrate was first weighed individually for each animal according to its weight and then solubilized with 0.2 ml saline just prior to injection. Diabetes was induced by injecting it at a dose of 100 mg/kg b. wt. intraperitonially. After 1 hr of Alloxan administration, the animals were given feed ad labium and 5% dextrose solution was also given in feeding bottle for a day to overcome the early hypoglycemic phase.

The animals were kept under observation and after 48 hr, blood glucose was measured. One group served as a control which received vehicle alone. The diabetic rats (glucose level > 150 mg/dl) were separated and divided into four different groups for experimental study ⁷.

Experimental design:

Acute treatment and sub acute treatment: Normal rats are kept into group, diabetic induced rat are grouped into 2, 3, 4, 5. Each group contains six rats.

1. Group I: Normal control (saline),
2. Group II: Diabetic control (saline),
3. Group III: Standard (Glibenclamide 10mg/kg)
4. Group IV: Test-dose (100mg/kg of Ribes nigrum fruit Extract)
5. Group V: Test-dose (500 mg/kg of Ribes nigrum fruit Extract)

Drugs are administered via oral route, Treatment continued for seven days ⁷.

Acute study (single day study): Blood samples were collected from rat tale vain and serum glucose levels were estimated at 0, 1, 3 and 5 h after the extract administration.

Sub acute stud (seven day study): Blood samples were collected from rat tale vain and serum glucose levels were estimated at 1, 3, 5, and 7 days Blood glucose levels were determined by god-pod method.

Histopathological study of Pancreas: Rats are sacrifice by using euthanasia. Pancreases are collected in 10% formalin solution. The section cuttings of 5μ thickness were is stained with haematoxylin and eosin. The section was observed in the Photomicrograph ⁸.

Statistical analysis: All parameters were analyzed statistically by using Graph pad Prism version 5.0.Rats was analyzed by one way ANOVA followed by "Dennett's Test”. All the values of the experimental results were expressed as mean±sem. Differences were considered significant at p < 0.05 ⁹.

RESULTS:

TABLE 1: EFFECT OF RIBES NIGRUM FRUIT EXTRACT ON SERUM GLUCOSE LEVEL FOR ACUTE STUDY (SINGLE DAY STUDY)

Values are mean±SEM; N=6. *P value < 0.01, **p < 0.001, ***p < 0.0001vs. Diabetic control.

TABLE 2: EFFECT OF RIBES NIGRUM FRUIT EXTRACT ON BLOOD GLUCOSE LEVEL FOR SUB ACUTE STUDY (MULTI DAY STUDY)

Values are mean±SEM; N=6. *P value < 0.01, **p < 0.001, ***p < 0.0001vs. Diabetic control.

Graphical representation of serum glucose levels in the acute study:

(A) Diabetic control                         (B) Normal control

(C) Standard                                   (D) Extract 100mg/kg

(E) Extract 500mg/kg

FIG. 7.1. GRAPHICAL REPRESENTATION OF BLOOD GLUCOSE LEVELS IN THE ACUTE STUDY (A) DIABETIC CONTROL, (B) NORMAL CONTROL, (C) STANDARD, (D) EXTRACT 100mg/kg AND (E) EXTRACT 500mg/kg

Graphical representation of serum glucose levels in the sub acute study:

(A) Diabetic control                                 (B) Normal control

(C) Standard                                   (D) Extract 100mg/kg

(E) Extract 500mg/kg

FIG. 7.2: (A) DIABETIC CONTROL, (B) NORMAL CONTROL, (C) STANDARD (D) EXTRACT 100mg/kg (E) EXTRACT 500mg/kg

Histopathological study: The whole pancreas from each animal was removed after sacrificing the animal and was collected in 10% formaline solution. Sections of 5μ thickness were cut and stained by haematoxylin and eosin (H & E) for histological examination.

NORMAL CONTROL           DIABETIC CONTROL

STANDARD    TEST 100MG/KG    TEST 500MG/KG

DISCUSSION:

• In acute study, the Ribes nigrum fruit extract at the dose of 500 mg/kg shows very significant Antidiabetic activity at 3^rd  hour, the dose of 100 mg/kg did not show any significant antidiabetic activity.
• In sub acute study the Ribes nigrum fruit extract at the dose of 500 mg/kg shows very significant activity from the first day to seventh day, the dose of 100mg/kg shows significant from 5^th day to 7^th day.
• Histopathological study shows that 500 mg/kg extract have shows high level regeneration of beta cells as compared to 100 mg

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    How to cite this article:

    Sivalingam G and Sriram N: Anti-diabetic activity of Ribes nigrum fruit extract in Alloxan induced diabetic rats, 2011. Int J Pharm Sci Res 2013; 4(3); 1196-1201.

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