Scholarly article on topic 'Methanolic extract of Marrubium vulgare ameliorates hyperglycemia and dyslipidemia in streptozotocin-induced diabetic rats'

Methanolic extract of Marrubium vulgare ameliorates hyperglycemia and dyslipidemia in streptozotocin-induced diabetic rats Academic research paper on "Clinical medicine"

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Abstract of research paper on Clinical medicine, author of scientific article — Ahmed A. Elberry, Fathalla M. Harraz, Salah A. Ghareib, Salah A. Gabr, Ayman A. Nagy, et al.

Abstract Aim and background Marrubium vulgare is used in traditional medicine in some countries such as Mexico in the treatment of diabetes. On the other hand, some studies reported the antioxidant effect of the extract due to its flavonoid content. The current study was designed to investigate the antidiabetic and antidyslipidemic effects of the methanolic extract of the aerial part of M. vulgare in streptozotocin-induced diabetic rats. Materials and methods The antidiabetic activity of a daily single oral dose of 500mg/kg/day of M. vulgare for 28days was evaluated by measuring the fasting blood glucose and the peak of blood glucose level within 120min of oral glucose tolerance test (OGTT) in diabetic rats. In addition, the effect of the extract on blood plasma insulin was measured as well as its effect on tissue glycogen contents in muscles and liver. Moreover, its effect on the oxidant status was evaluated. Results M. vulgare significantly reduced the blood glucose level starting on the second week. Furthermore, the extract of M. vulgare showed significant increase in plasma insulin and tissue glycogen contents. The antidyslipidemic effect was demonstrated by a significant reduction in plasma total cholesterol (TC), triglycerides (TG), and low density lipoprotein-cholesterol (LDL-C), while the cardio-protective lipid, high density lipoprotein-cholesterol (HDL-C), was increased. Conclusion The present data suggest that M. vulgare’s methanolic extract has antihyperglycemic with antidyslipidemic effect. The protective effect of the extract may be in part due to its antioxidant activity.

Academic research paper on topic "Methanolic extract of Marrubium vulgare ameliorates hyperglycemia and dyslipidemia in streptozotocin-induced diabetic rats"

International Journal of Diabetes Mellitus (2011) xxx, xxx-xxx

Diabetes Science International International Journal of Diabetes Mellitus

diabetes

www.elsevier.com/locate/ijdm www.sciencedirect.com

ORIGINAL ARTICLE

Methanolic extract of Marrubium vulgare ameliorates hyperglycemia and dyslipidemia in streptozotocin-induced diabetic rats

Ahmed A. Elberry a *, Fathalla M. Harraz b, Salah A. Ghareib c, Salah A. Gabr d Ayman A. Nagy e, Essam Abdel-Sattar f

a Department of Clinical Pharmacy, Faculty of Pharmacy, King Abdulaziz University, Jeddah 21589, Saudi Arabia b Department of Pharmacognosy, Faculty of Pharmacy, Alexandria University, Alexandria, Egypt

c Department of Pharmacology and Toxicology, Faculty of Pharmacy, King Abdulaziz University, Jeddah, Saudi Arabia d Department of Isotopes Applications, Nuclear Research Center, Atomic Energy Authority, Egypt e Department of Forensic Medicine and Clinical Toxicology, Faculty of Medicine, Tanta University, Egypt f Department of Natural Products, Faculty of Pharmacy, King Abdulaziz University, Jeddah, Saudi Arabia

KEYWORDS

Marrubium vulgare;

Antihyperglycemic;

Antioxidant;

Dyslipedimia;

Diabetes;

Abstract Aim and background: Marrubium vulgare is used in traditional medicine in some countries such as Mexico in the treatment of diabetes. On the other hand, some studies reported the anti-oxidant effect of the extract due to its flavonoid content. The current study was designed to investigate the antidiabetic and antidyslipidemic effects of the methanolic extract of the aerial part of M. vulgare in streptozotocin-induced diabetic rats.

Materials and methods: The antidiabetic activity of a daily single oral dose of 500 mg/kg/day of M. vulgare for 28 days was evaluated by measuring the fasting blood glucose and the peak of blood glucose level within 120 min of oral glucose tolerance test (OGTT) in diabetic rats. In addition, the effect of the extract on blood plasma insulin was measured as well as its effect on tissue glycogen contents in muscles and liver. Moreover, its effect on the oxidant status was evaluated. Results: M. vulgare significantly reduced the blood glucose level starting on the second week. Furthermore, the extract of M. vulgare showed significant increase in plasma insulin and tissue glycogen

* Corresponding author. Tel.: +966 543430919; fax: +966 2 6951696. E-mail address: berry_ahmed@yahoo.com (A.A. Elberry).

1877-5934 © 2011 International Journal of Diabetes Mellitus. Published by Elsevier Ltd. All rights reserved. doi:10.1016/j.ijdm.2011.01.004

contents. The antidyslipidemic effect was demonstrated by a significant reduction in plasma total cholesterol (TC), triglycerides (TG), and low density lipoprotein-cholesterol (LDL-C), while the cardio-protective lipid, high density lipoprotein-cholesterol (HDL-C), was increased. Conclusion: The present data suggest that M. vulgare's methanolic extract has antihyperglycemic with antidyslipidemic effect. The protective effect of the extract may be in part due to its antioxidant activity.

© 2011 International Journal of Diabetes Mellitus. Published by Elsevier Ltd. All rights reserved.

1. Introduction

Diabetes mellitus is a metabolic disease, characterized by hyperglycemia together with impaired metabolism of glucose and other energy-yielding fuels, such as lipids and proteins

[I]. This metabolic disorder is the result of a deficiency in insulin secretion or a resistance to insulin action, or both [2]. More than 220 million people worldwide have diabetes and this number is likely to more than double by the year of 2030 [3]. Diabetic patients also exhibit oxidative stress, which leads to lipid peroxidation and tissue damage including retinopathy, nephropathy, and coronary heart disease [4,5]. Dyslipidemia or hyperlipidemia is also involved in the development of cardiovascular complications, which are a major cause of morbidity and mortality [6].

Renewed attention to alternative medicines and natural therapies has raised researchers' interest in traditional herbal medicine. Because of their perceived effectiveness, with minimal side effects in clinical experience and relatively low costs, herbal drugs are prescribed widely, even when their contents of the biologically active constituents are unknown [7]. Marrubium vulgare L. (Lamiaceae), commonly known as "Horehound", is a widespread Mediterranean plant used in folk medicine to cure a variety of diseases, and is widely distributed in Saudi Arabia. M. vulgare has been widely used in Mexican traditional medicine for the treatment of DM [8], and its hypoglycemic effect was demonstrated during the study of the hypoglycemic effect of some Mexican and Brazilian plants [9,10]. The plant is reported to possess vasorelaxant

[II], antihypertensive [12], analgesic [13], anti-inflammatory [14], and antioxidant properties [15]. The main objective of our study is to elucidate the effect of M. vulgare on hypergly-cemia and dyslipidemia in streptozotocin (STZ)-induced diabetes in rats.

2. Materials and methods

2.1. Chemicals

Streptozotocin (STZ) and a-D-glucose were purchased from Sigma-Aldrich (St. Louis, MO, USA). Carboxymethylcellulose sodium (CMC-Na) was purchased from Acros Organics (NJ, USA), while heparin sodium was purchased from Merck (Dramstadt, Germany). An insulin kit (Coat-A-Count Insulin) was purchased from Siemens, Medical Solutions Diagnostics (Los Angeles, USA), while lipid profile kits were purchased from Randox Laboratories Ltd. (Antrim, UK). Marrubiin, the major diterpene in M. vulgare, was isolated according to the method reported by Knoss et al. [16], and was identified by NMR analysis. All other biodiagnostic kits were purchased from Diagnostic and Research Reagents (Giza, Egypt).

2.2. Plant materials and extraction

The aerial part of M. vulgare was collected in December 2009 from Wadi Kama, Al-Taif governorate, Saudi Arabia. It was dried in the shade and the specimen was deposited in the herbarium of the College of Pharmacy, King Abdulaziz University, Jeddah, Saudi Arabia (M. vulgare #MV1105). The dried aerial part (500 g) was homogenized with methanol for 15 min, three times each with 1000 mL using Ultra-turrax T25 homogenizer (Janke and Kunkel, IKA Labortechnik, Stauten, Germany) followed by distillation of the solvent under reduced pressure. The percentage yield was calculated as 12%, and the dried methanolic extract was kept at 4 0C.

2.3. Phytochemical analysis

A preliminary phytochemical test to identify the chemical constituents of M. vulgare was carried out according to the methods of Treas and Evans [17].

2.4. TLC profile of extract

The methanolic extract was analyzed using thin layer chroma-tography as per fingerprinting using a chromatographic system as reported by Wagner and Bladt [18]. TLC on Si gel 60 F254 plates, using CHCL3-MeOH (95:5) as a solvent system, was used for the identification of terpenoid class compounds, after spraying with Komarowsky reagent. The extract was also chromatographed on TLC (Si gel 60 F254 plates) using AcOEt-HCOOH-CH3COOH-H2O (100:11: 11:26) as the mobile phase. TLC was observed under UV 254 and 366 nm, before and after spraying with natural products reagent NP/PEG for the detection of flavonoid class compounds.

2.5. Determination of total flavonoids content

Total flavonoid content was measured by means of an aluminum chloride colorimetric assay [19] with slight modifications. An aliquot (1 ml) of the extract (14.5 mg/10 ml methanol) or standards solution of quercetin (0.60-1.8 mg/ml) was added to a 10 ml volumetric flask, containing 4 ml of 50% solution of methanol. To the flask, 0.3 ml 5% NaNO2 was added. After 5 min, 0.3 ml 10% AlCl3 was added. At the sixth minute, 2 ml 1 M NaOH solution was added and the total volume was made up to 10 ml with dd H2O. The solution was well mixed, and absorbance was measured against the prepared reagent, blank at 510 nm. The total flavonoid content (mg/g) was determined from the calibration curve, and expressed as mg quercetin equivalents. All the determinations were carried out in triplicate, and the mean values were calculated.

2.6. Animals and experimental protocol

Male Wistar rats, weighing 200-250 g, were used in this study, in accordance with the guidelines of the Biochemical and Research Ethical Committee at King Abdulaziz University, Jeddah, Saudi Arabia. Animals were purchased from the animal house of King Fahd Medical Research Center, King Abdulaziz University and were housed for one week under standard conditions (well ventilated, temperature 22 ± 2 0C, relative humidity 50-60% and 12 h day and night cycle). Food consisted of normal rat chow, and water was provided ad libitum. Care was taken to avoid stressful conditions, and all procedures were performed between 8 and 10 a.m.

Diabetes was induced through a fresh solution of STZ (in 0.1 M sodium citrate buffer, pH 4.5), injected intraperitoneally (i.p.) at a dose of 55 mg/kg, as a single dose to overnight fastening rats [20]. Ten days after STZ administration, the rats with blood glucose concentration of between 20 and 30 mmol/L (360-540 mg/dl) were selected, and divided into three groups (groups II-IV).

Animals were randomly divided into 4 groups, with 8 animals in each group. Group I (normal control), received an equivalent volume of the 0.1 M sodium citrate buffer. Group II (diabetic control), diabetic rats received a single daily dose of 1% CMC-Na, as a vehicle for the tested extract, starting on the 11th day. This group served as a —ve control group. In Group III (the glibenclamide group), diabetic rats received a single daily dose of glibenclamide in a dose of 5 mg/kg, starting on the 11th day. This group served as a +ve control group. In Group IV (M. vulgare group), diabetic rats received a plant extract of M. vulgare in a dose of 500 mg/kg as a single daily dose, starting on the 11th day. The 500 mg/kg extract dose used in the treatment was chosen from a preliminary short-term study in our laboratory. Vehicle, glibenclamide and plant extract were given orally by gavage as single daily treatments, for 28 days. Blood samples were collected from tail vein, and fasting blood glucose of the overnight fasted animals was measured before and at days 7, 14, 21, and 28 from treatment.

At the end of the experiment, blood samples were withdrawn, and animals were sacrificed through cervical dislocation, under light ether anesthesia, to separate the liver. Livers were dissected out and retained in liquid nitrogen to determine the antioxidant status. A 5% tissue homogenate in ice-cold 0.9% saline was made, and the supernatant was obtained from tissue homogenate by centrifugation (1000g, 4 0C, 10 min) for determination of the oxidant status.

2.7. Toxicity study

The rats were given M. vulgare at increasing doses of 100, 250, 500, and 1000 mg/kg daily, for a period of 3 weeks (six rats were taken for each dose). The animals were observed continuously during the first hour, and then every hour for 6 h, then after 12 and 24 h, and finally after every 24 h, up to 3 weeks, for any physical signs of toxicity such as writhing, gasping, palpitation and respiratory rate, or mortality.

2.8. Evaluation of the effect of M. vulgare extract on plasma insulin level

At the end of the 28th day, 3 h after the last dose of the solvent, glibenclamide or the extract, blood samples were with-

drawn from the orbital sinus of rats, under light ether anesthesia, into heparinized tubes. Samples were centrifuged at 3500 rpm for 15min to separate the plasma. The plasma samples were separated and kept at —20 0C for analysis when required. Insulin concentrations were determined through a radioimmunoassay procedure, using insulin kits according to the manufacturer's instructions.

2.9. Evaluation of the effect of M. vulgare extract on plasma glucose and tissue glycogen

Fasting plasma glucose was estimated using the glucose oxidase peroxidase method [21]. Tissue glycogen was estimated through the method of Morales et al. [22].

2.10. Evaluation of the effect of M. vulgare extract on plasma lipid profile

Total cholesterol (TC), triglycerides (TG) and HDL-choles-terol (HDL-C) were determined spectrophotometrically, using commercial kits. Low density lipoprotein (LDL) was calculated by using Friedewald formula [23].

2.11. Evaluation of the effect of M. vulgare extract on liver antioxidant status

Hepatic superoxide dismutase (SOD) activity was determined according to the method described by Sun and Zigman [24], while, the activities of hepatic glutathione peroxidase (GPx), glutathione reductase (GR) and glutathione-S-transferase (GST) activity were determined according to Mohandas et al. [25]. Reduced glutathione (GSH) level was measured according to the chemical method described by Moron et al. [26], while lipid peroxidation products were determined by measuring malondialdehyde (MDA) content in tissue homog-enates, according to the method of Uchiyama and Mihara [27].

2.12. Evaluation of the effect of M. vulgare extract on oral glucose tolerance test (OGTT)

Twenty four male Wistar rats were divided into other four groups (six rats in each group) as the first protocol. At the end of the 28th day, 3 h after the last dose of the solvent, gli-benclamide or the extract, blood samples were withdrawn from tail vein of overnight fasting rats and blood glucose was determined, indicating zero time of the test. Glucose solution (50%) in a dose of 2.5 g/kg [28] was given orally. Blood samples were withdrawn at 15, 30, 60, 90, and 120 min after glucose loading, and blood glucose level was determined at these time intervals using glucometer (Accu-Check Active, Roche). Curves of blood glucose levels (mg/dl) versus time intervals (min) were constructed, and the area under the curve (AUC) was calculated according to the trapezoidal method. The AUCs of the curves of each group were compared and tested for significance to the control diabetic group, to represent glucose utilization by the tissues.

2.13. Determination of protein content

The protein content of cardiac tissue homogenates was determined by the Lowry protein assay using bovine serum albumin as the standard [29].

2.14. Statistical analysis

Data are expressed as mean ± standard error (SE) of mean. Statistical analyses were performed using one-way analysis of variance (ANOVA). If the overall F-value was found to be statistically significant (P < 0.05), further comparisons among groups were made according to post hoc Tuckey's test. All statistical analyses were performed using GraphPad InStat 3 (GraphPad Software, Inc. La Jolla, CA, USA) software.

3. Results

3.1. Phytochemical analysis

Phytochemical screening of the extracts indicated the presence of flavonoids, tannins, and sterols and/or terpenoids, and the absence of alkaloids and saponins.

3.2. Determination of total flavonoids content

Assay of total flavonoid content of dried aerial parts of M. vulgare was determined to be 15.53 mg quercetin equivalent/g of dry plant material.

3.3. Toxicity study

A toxicity study revealed the non-toxic nature of the extract. The rats treated with different doses of M. vulgare did not show any drug-induced physical signs of toxicity during the whole experimental period, and no deaths were reported.

3.4. Effect of M. vulgare extract on plasma glucose level

Diabetic rats showed a significant increase in the levels of plasma glucose when compared to normal rats. Oral administra-

Table 1 Effect of pretreatment with methanolic extract of M. vulgare (500 mg/kg/day for 28 days) on blood glucose levels of diabetic rats.

Groups Blood glucose level (mg/dl)

Before treatment After treatment

Day 0 7th day 14th day 21st day 28th day

Normal control Diabetic control Diabetic + glibenclamide Diabetic + M. vulgare 113 ± 3.9 112 ± 2.8 404 ± 27.2 416 ± 22.8 421 ± 18.8 466 ± 19.5 435 ± 29.6 495 ± 3.1 110 ± 3.7 443 ± 17.6 453 ± 22.8 405 ± 19.2a'b 100 ± 5.8 480 ± 31.7 386 ± 21.6a'b 355 ± 30.5a'b 121 ± 7.4 498 ± 31.3 345 ± 25.7ab 337 ± 23.6ab

The values are expressed as the mean ± SE of the mean of eight rats in each group. a Significantly reduced from values in day 0 in each corresponding group at P < 0.05. b Significantly reduced from control values of diabetic rats in the corresponding day at P < 0.05.

Table 2 Effects of oral pretreatment with methanolic extract of M. vulgare (500 mg/kg/day for 28 days) on plasma insulin level, muscle glycogen, and liver glycogen in diabetic rats.

Groups Insulin (iIU/ml) Muscle glycogen (mg/g tissue) Liver glycogen (mg/g tissue)

Normal control 8.48 ± 1.32 11.12 ± 0.9 54.43 ± 5.4

Diabetic control 3.22 ± 0.38a 4.13 ± 0.2a 22.12 ± 2.3a

Diabetic + glibinclamide 8.47 ± 2.39b 8.41 ± 0.6b 46.13 ± 4.2b

Diabetic + M. vulgare 6.16 ± 1.26b 9.83 ± 0.8b 48.13 ± 5.2b

The values are expressed as the mean ± SE of the mean of eight rats in each group. a Significantly different from the values of the normal rats at P < 0.05. b Significantly different from the control values of diabetic rats at P < 0.05.

Table 3 Effects of oral pretreatment with methanolic extract of M. vulgare (500 mg/kg/day for 28 days) on triglycerides (TG), high density lipoprotein (HDL-C) and low density lipoprotein (LDL-C) of diabetic rats. total cholesterol (TC),

Groups TC (mg/dL) TG (mg/dL) HDL-C (mg/dL) LDL-C (mg/dL)

Normal control Diabetic control Diabetic + glibinclamide Diabetic + M. vulgare 125.71 ± 4.04 239.99 ± 21.53a 178.28 ± 15.05ab 181.28 ± 17.20a'b 87 ± 2.52 113.33 ± 11.15a 121.6 ± 14.62a 108.80 ± 15.09a 29.57 ± 5.47 20.56 ± 3.09a 41.84 ± 7.15a'b 42.14 ± 4.81ab 78.70 ± 4.58 168.68 ± 8.75a 112.10 ± 8.02a,b 123.85 ± 16.28a,b

The values are expressed as the mean ± SE of the mean of eight rats. a Significantly different from the values of the normal control rats at P < 0.05. b Significantly different from the control values of diabetic rats at P < 0.05.

tion of M. vulgare showed a highly significant reduction in the plasma glucose level starting at the 14th day of treatment, compared to before treatment (day 0) as well as compared to the control values of diabetic rats in the corresponding days. This reduction was marked on the 28th day as reaching a 23% reduction, compared to day 0, and a 42% reduction com-

hand, glibenclamide treatment of diabetic rats showed a significant reduction in plasma glucose level on the 21st day of treatment, compared to before treatment and compared also to the control values of diabetic rats in the corresponding days. The plasma glucose level was reduced by 18% compared to day 0 and 31% compared to control values of the diabetic group

pared to the control values of the diabetic group. On the other on the 28th day (Table 1).

125n C 100-

<u о a. u> E 3

<z сэ

° 4-| .

Figure 1 Effects of pretreatment with methanolic extract of M. vulgare (500 mg/kg/day for 28 days) on superoxide dismutase (SOD), glutathione peroxidase (GPx), glutathione reductase (GR), and glutathione-S-transferase (GST, reduced glutathione (GSH) and Malondialdehyde (MDA) in diabetic rats.

3.5. Effect of M. vulgare extract on plasma insulin level, liver glycogen and muscle glycogen

The plasma insulin level was significantly reduced in the diabetic group. Pretreatment of diabetic rats with M. vulgare extract or glibenclamide for 4 weeks significantly increased the insulin blood level compared to diabetic control rats (48%) (Table 2). The muscle and liver glycogen contents decreased significantly in diabetic rats when compared to normal rats. M. vulgare extract and glibenclamide treatment significantly increased both muscle and liver glycogen, compared to the diabetic control rats (58% and 54%, respectively) (Table 2).

Table 4 Total area under the curve induced after glucose loading in diabetic rats pretreated with the methanolic extract of M. vulgare (500 mg/kg/day) for four weeks.

Groups Total AUC (mg/dl min) % Reduction

Normal control 14,479 ± 480.42 -

Diabetic control 70,108 ± 2077.06 0.00

Diabetic + glibenclamide 59,692 ± 1785.26a 15%

Diabetic + M. vulgare 61,350 ± 1512.88a 12%

a Significantly different from the control values of diabetic rats at P < 0.05.

3.6. Effect of M. vulgare extract on plasma lipid profile

Diabetic rats showed a significant increase in the blood levels of TC, TG, and LDL-C (Table 3). On the other hand, HDL-C was significantly reduced when compared to the values of normal control rats. Oral administration of glibenclamide significantly decreased the blood levels of TC and LDL-C. However, the level of HDL-C significantly increased. Glibinclamide did not significantly change the blood level of TG, when compared to diabetic control rats. The pretreatment with M. vulgare extract significantly decreased the blood levels of TC (24%) and LDL-C (27%); however, the blood level of HDL-C was significantly increased when compared to the values of the diabetic control rats (27%) (Table 3).

3.7. Effect of M. vulgare extract on hepatic oxidative stress

Streptozotocin administration resulted in a significant decrease in antioxidant enzymatic activity, including GPx, GR, and GST. Moreover, it caused a significant increase in MDA, while GSH was reduced significantly (Fig. 1). M. vulgare extract exhibited improvements in antioxidant enzymatic activity, compared to the diabetic control group, and nearly normalized the level of both GSH and MDA. The activity of SOD was not significantly changed (Fig. 1).

3.8. Effect of M. vulgare extract on OGTT and AUC

The blood levels of glucose in normal control, diabetic control, and diabetic treated with gibenclamide or M. vulgare extract

—■—Normal control -a- Diabetic control -▼-Diabetic + Glibenclamide Diabetc + M. vulgare

25 50 75 100 Time (Min)

Figure 2 Effect of methanolic extract of M. vulgare (500 mg/kg/ d) on blood glucose level. The values are expressed as the mean ± SE of the mean of six rats in each group.

groups demonstrated a significant change in blood glucose level after oral loading, with 50% glucose solution (Fig. 2 and Table 4). The rats in the diabetic group had a significant elevation in blood glucose level throughout the total measurement period (120 min) with respect to normal control (Fig. 2); moreover, it did not come back to the initial value (0 min level) even at the end of the period tested. Pretreatment of diabetic rats with gibenclamide or M. vulgare extract induced a significant reduction in AUC relative to the diabetic control group by 15% and 12%, respectively (Fig. 2 and Table 4).

4. Discussion

In the present study, diabetes mellitus was induced in rats through a STZ injection that causes the destruction of b-cells of islets of Langerhans, as proposed by many authors [30,31]. This effect was represented in the current study through the elevation of blood glucose and a decrease of insulin levels in diabetic control rats. The elevated plasma glucose levels in diabetic rats were lowered through the administration of M. vulgare, which showed an elevated plasma insulin level compared to diabetic control rats. The action of M. vulgare seems to be similar to that of glibenclamide [32].

Earlier phytochemical investigation of M. vulgare led to the characterization of several flavonoids [33] that possess hypo-glycemic, as well as antioxidant properties. Some flavonoids have hypoglycemic properties because they improve altered glucose and oxidative metabolisms of the diabetic states [34]. They also exert a stimulatory effect on insulin secretion by changing Ca+ + concentration [35].

Glycogen synthesis in the rat liver and skeletal muscles was impaired during diabetes [36]; hence glycogen content of skeletal muscle and liver was markedly decreased in diabetic rats [37]. The results of the present study showed an increase in skeletal muscle and liver glycogen content in diabetic rats after the oral administration of M. vulgare which may be due to the stimulation of insulin release from beta cells [38]. Furthermore, the diabetic control rats showed a significant increase in the AUC of the glucose concentration after oral glucose loading. This effect may be due to the reduction of glucose tissue utilization and an increased hepatic glucose production, as a result of decreased insulin production [39]. The administration of M. vulgare extract produced a significant reduction in the AUC of diabetic control rats. These results revealed that the M. vulgare extract induced an increase in glucose utilization and glucose tolerance through the body tissues of diabetic rats.

It is well known that dyslipidemia is associated with uncontrolled diabetes mellitus. The plasma levels of TC, LDL-C, and

TG increases, while the HDL levels decline, contributing to secondary complications of diabetes [40,41]. Acute insulin deficiency initially causes an increase in free fatty acid mobilization from adipose tissue. This results in an increased production of LDL-C particle [42]. In the present study, diabetic rats exhibited a significant elevation of TC, TG, and LDL-C, while HDL-C was decreased. M. vulgare administration resulted in lowering the plasma levels of TC, TG, and LDL-C with elevation of HDL-C level. It is known that the administration of insulin to diabetic subjects not only elevates lipoprotein lipase activity, but also lowers the plasma TG concentrations [43]. The presently observed decline in plasma lipid profiles in M. vulgare administered diabetic rats suggests that the extract's potential is possibly due to the elevation of insulin level.

The possible mechanism of the extract may in part be attributed to its antioxidant activities. Complementing our findings, earlier studies have reported that the extract may have antioxidant activity. M. vulgare leaves have been reported to be rich in phenolic compounds [44] and these compounds were previously served as free radical scavengers [45]. Hyper-glycemia generates reactive oxygen species (ROS), which in turn cause lipid peroxidation and membrane damage [46]. The current study showed a significant reduction in antioxi-dant enzymes including GPx, GR, and GST in the diabetic rats with decreased GSH and increased MDA levels indicating oxi-dative stress. A significant improvement in these indictors of oxidative stress in the liver of M. vulgare treated diabetic rats is indicative of its ability to reduce body glucose concentration, and its subsequent oxidation. These effects of M. vulgare on antioxidants were found to be better than those of glibencla-mide treated diabetic rats.

In conclusion the M. vulgare has both a hypoglycemic effect and an antidyslipidemic activity. The possible mechanism of the antidiabetic action may be through a stimulation of insulin release from the remnant pancreatic b-cells. Both antidiabetic and antidyslipedimic effects may in part be due to its antioxi-dant activity.

Conflict of interest statement

The authors declare that there are no conflicts of interest. References

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