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Effect of Allium sativum leaf extracts on glucose tolerance in glucose-induced hyperglycemic mice.

INTRODUCTION

Allium sativum L. (Liliaceae) is known as garlic in English and roshun in Bengali. The plant belongs to the onion genus, Allium. It is widely cultivated throughout the world for both culinary and medicinal purposes. The cloves are the main part consumed or used medicinally, although ethnomedicinal use of the leaves is not unknown. The tribals of Chitteri Hills, Dharmapuri District, Tamil Nadu, India use leaves of the plant as anthelmintic, antiasthmatic, anticholesterolemic, antiseptic, antispasmodic, diaphoretic, diuretic, stomachic, and antidiabetic (Kadhirvel et al, 2010). The cloves of the plant are used as natural remedies against diabetes and related complications in Mauritius (Mootoosamy and Mahomoodally, 2014).

Garlic cloves have been the most scientifically studied part with relation to diabetes and diabetes-induced complications than other parts of the plant. Administration of ethanolic extract of the cloves has been shown to produce hypoglycemic and hypolipidemic effects in alloxan-induced diabetic rabbits (Sher et al., 2012). In streptozotocin-induced diabetic rats, elevated levels of glucose, cholesterol and triglycerides were found to be decreased when garlic clove juice was administered for 3 weeks prior to streptozotocin injection, thus showing that garlic has also preventive properties (Masjedi et al., 2013). The hypoglycemic effect of methanol extract of garlic cloves has also been shown in alloxan-induced diabetes in male Wistar rats. Administration of extract led to inhibition of rat intestinal alpha-glucosidase and enhancement of pancreatic and cardiac Glut-4 mRNAs expression (Moradabadi et al., 2013). S-allylcysteine, a component of garlic clove has been shown to improve streptozotocin-induced alterations of blood glucose, liver cytochrome P450 2E1, plasma antioxidant system and adipocytes hormones in diabetic rats (Saravanan and Ponmurugan, 2013). Raw garlic clove homogenate has been found to attenuate cardiac oxidative stress in fructose-fed diabetic rats via activation of PI3K/AKT/Nrf2-Keap1 pathway (Padiya et al., 2014).

We have been conducting extensive pharmacological studies on Bangladeshi medicinal plants towards identifying plants with antihyperglycemic, antinociceptive, and cytotoxic activities (Anwar et al., 2010; Jahan et al., 2010; Rahman et al., 2010; Rahmatullah et al., 2010; Shoha et al., 2010; Ali et al., 2011; Barman et al., 2011; Hossan et al., 2011; Jahan et al., 2011; Rahman et al., 2011; Sutradhar et al., 2011; Ahmed et al., 2012; Arefin et al., 2012; Haque et al., 2012; Sathi et al., 2012). The objective of this study was to evaluate the antihyperglycemic potential of methanol extract of leaves of Allium sativum.

MATERIALS AND METHODS

Leaves of Allium sativum were collected from Mullatagor, Kushtia district, Bangladesh during October 2013. The plant was taxonomically identified at the Bangladesh National Herbarium at Dhaka (Accession Number 38,573). The sliced and air-dried leaves of Allium sativum were grounded into a fine powder and 100g of the powder was extracted with 500 ml methanol for 48 hours. The extract was evaporated to dryness at 40oC. The final weight of the extract was 5.49g.

Chemicals:

Glibenclamide and glucose were obtained from Square Pharmaceuticals Ltd., Bangladesh.

Animals:

In the present study, Swiss albino mice (male), which weighed between 15-20g were used. The animals were obtained from International Centre for Diarrheal Disease Research, Bangladesh (ICDDR,B). All animals were kept under ambient temperature with 12h light followed by a 12h dark cycle. The animals were acclimatized for three days prior to actual experiments. The study was conducted following approval by the Institutional Animal Ethical Committee of the University of Development Alternative, Dhaka, Bangladesh.

Antihyperglycemic activity:

Glucose tolerance property of methanol extract of Allium sativum leaves was determined as per the procedure previously described by Joy and Kuttan (1999) with minor modifications. In brief, fasted mice were grouped into six groups of six mice each. The various groups received different treatments like Group 1 received vehicle (1% Tween 80 in water, 10 ml/kg body weight) and served as control, group 2 received standard drug (glibenclamide, 10 mg/kg body weight). Groups 3-6 received methanol extract of Allium sativum leaves (ASME) at doses of 50, 100, 200 and 400 mg per kg body weight. Each mouse was weighed and doses adjusted accordingly prior to administration of vehicle, standard drug, and test samples. All substances were orally administered. Following a period of one hour, all mice were orally administered 2 g glucose/kg of body weight. Blood samples were collected 120 minutes after the glucose administration through puncturing heart. Blood glucose levels were measured by glucose oxidase method (Venkatesh et al., 2004).

The percent lowering of blood glucose level was calculated as follows.

Percent lowering of blood glucose level = (1 ? [W.sub.e]/[W.sub.c]) X 100, where [W.sub.e] and [W.sub.c] represents the blood glucose concentration in glibenclamide or ASME administered mice (Groups 2-6), and control mice (Group 1), respectively.

Statistical analysis:

Experimental values are expressed as mean [+ or -] SEM. Independent Sample t-test was carried out for statistical comparison. Statistical significance was considered to be indicated by a p value < 0.05 in all cases.

RESULTS AND DISCUSSION

Preliminary phytochemical analysis revealed the presence of alkaloids, tannins, and saponins in the extract.

In oral glucose tolerance tests conducted with glucose-loaded Swiss albino mice, methanolic extract of leaves significantly and dose-dependently reduced blood glucose concentrations. At extract doses of 50, 100, 200 and 400 mg per kg body weight mice, the percent lowering of blood sugar by the extract was, respectively, 16.7, 37.0, 42.6, and 52.8. A standard antihyperglycemic drug, glibenclamide, when administered to glucose-loaded mice at a dose of 10 mg per kg body weight, reduced blood sugar levels by 47.3%. Thus at the highest dose of the extract (400 mg), the percent lowering of blood sugar was greater than that of glibenclamide. The results are shown in Table 1 and demonstrate that the methanolic extract possesses significant antihyperglycemic potential, which can prove useful in lowering of blood glucose in diabetic patients.

Leaves of the plant are known to contain among other constituents cholorogenic acid, phloroglucinol, quercetin-3-O-beta-D-glucoside, rutin, and stigmasterol (Duke, 1992). Chlorogenic acid has been shown to inhibit both alpha-amylase and alpha-glucosidase enzymes in a dose-dependent manner (Oboh et al., 2014) and as such, can play a role in the observed antihyperglycemic activity of the extract. Notably, postprandial hyperglycemia in diabetes could be ameliorated by inhibiting intestinal a-glucosidases, and ethanol extract of banana pseudostems (containing stigmasterol) has been shown to inhibit mammalian intestinal alpha-glucosidases activity (Ramu et al., 2014). A phloroglucinol derivative from the brown alga Eisenia bicyclis has been found to be effective in inhibiting glycation and alpha-amylase (Okada et al., 2004). Quercetin-3-O-beta D-glucoside has been isolated from a Chinese medicine, Tang-Zhi-Qing, used for the treatment of Type 2 diabetes in China (Tao et al., 2013). Rutin has been implicated as one of the active constituent in the blood glucose lowering effect of Tectona grandis flowers in Type 2 diabetic rats (Ramachandran and Rajasekaran, 2014). Any or a combination of the above compounds may be responsible for the observed glucose-lowering effects as seen in the present study with ASME. As such, the extract merits further studies towards isolation and identification of the active component(s).

ARTICLE INFO

Article history:

Received 2 April 2014

Received in revised form 13 May 2014

Accepted 28 May 2014

Available online 27 June 2014

REFERENCES

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Diponcor Ghosh, Indrani Mandal, Jannatul Ferdous Rumi, Ummay Kawchur Trisha, Humayra Jannat, Mousumi Ahmed, Mohammed Rahmatullah

Faculty of Life Sciences, University of Development Alternative, Dhanmondi, Dhaka-1209, Bangladesh.

Corresponding Author: Mohammed Rahmatullah, Faculty of Life Sciences, University of Development Alternative, Dhanmondi, Dhaka-1209, Bangladesh. Tel: 88-01715032621; Fax: 88-02-815739; E-mail: rahamatm@hotmail.com
Table 1: Effect of methanol extract of Allium sativum leaves on blood
glucose level in hyperglycemic mice following 120 minutes of glucose
loading.

Treatment                 Dose (mg/kg      Blood glucose
                          body weight)     level (mmol/l)

Control (Group 1)            10 ml       5.98 [+ or -] 0.27
Glibenclamide (Group 2)      10 mg       3.15 [+ or -] 0.29
ASME (Group 3)               50 mg       4.98 [+ or -] 0.21
ASME (Group 4)               100 mg      3.77 [+ or -] 0.23
ASME (Group 5)               200 mg      3.43 [+ or -] 0.19
ASME (Group 6)               400 mg      2.82 [+ or -] 0.10

Treatment                 % lowering of blood
                             glucose level

Control (Group 1)                 --
Glibenclamide (Group 2)         47.3 *
ASME (Group 3)                  16.7 *
ASME (Group 4)                  37.0 *
ASME (Group 5)                  42.6 *
ASME (Group 6)                  52.8 *

All administrations were made orally. Values represented as mean [+ or
-] SEM, (n = 6); * P < 0.05; significant compared to hyperglycemic
control animals.
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Author:Ghosh, Diponcor; Mandal, Indrani; Rumi, Jannatul Ferdous; Trisha, Ummay Kawchur; Jannat, Humayra; Ah
Publication:Advances in Natural and Applied Sciences
Article Type:Report
Date:Jul 1, 2014
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