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Hypoglycemic Effects of Apigenin from Morus Indica in Streptozotocin Induced Diabetic Rats

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Hypoglycemic Effects of Apigenin from Morus Indica in Streptozotocin Induced Diabetic Rats International Journal of Current Research and Review Original Study DOI: http://dx.doi.org/10.31782/IJCRR.2021.13213 Hypoglycemic Effects of Apigenin from Morus Indica in Streptozotocin Induced Diabetic Rats IJCRR 1,2 1 Section: Healthcare Satish Anandan , Asna Urooj ISI Impact Factor (2019-20): 1.628 1 IC Value (2019): 90.81 Department of Studies in Food Science and Nutrition, University of Mysore, Manasagangothri, Mysuru -570 006, Karnataka, India; SJIF (2020) = 7.893 2Department of Clinical Nutrition & Dietetics, Sri Devaraj Urs Academy of Higher Education and Research, Kolar, Karnataka, India. Copyright@IJCRR ABSTRACT Introduction: Morus Indica L occupies an important position in the holistic system of Indian medicine ‘Ayurveda’ which has its roots in antiquity and has been followed for centuries. Objective: Methods: The hypoglycemic potential of apigenin isolated from MI leaves in streptozotocin (STZ) induced diabetic rats was studied. The rats were divided into four groups (n=6 animals in each group) viz. Group I- Normal healthy control rats (NC); Group II- STZ-induced diabetic control (DC) rats without treatment; Group III- STZ-induced diabetic rats treated with Ami- noguanidine (AG) (30 mg kg-1 body weight by intraperitoneal); Group IV- STZ-induced diabetic rats treated with Apigenin (API) (50 mg kg-1 body weight was given by orally). The protective effect of API was evaluated by determining the biochemical param- eters (lipid profile, liver, and kidney) and by studying the histopathological alterations in liver and kidney. Results: Diabetic control group had altered biochemical values (lipid profile, liver and kidney) when compared with the NC group. However, treatment with API showed significant improvement in the biochemical parameters and values are comparable to the NC group. Histopathological data revealed destruction of the kidney and liver architecture in DC, which was reverted in the group treated with API. Conclusions: The present findings suggest that API might be useful in the management of diabetes mellitus. Key Words: Hyperglycemic, Mulberry, Flavonoids, Streptozotocin, Oxidative stress INTRODUCTION proach supported by scientific documentation can serve as a novel, affordable medicine with minimum side effects.7,8 Diabetes mellitus (DM) is a metabolic disorder characterized by insulin resistance and pancreatic β-cell dysfunction as a Morus Indica L. is used as traditional medicine for its hypo- 8 consequence of unsettled hyperglycemia.1 Prolonged hyper- glycemic and diuretic properties. In our laboratory, Morus glycaemia may lead to a variety of secondary complications Indica (MI) has been screened for various biological prop- such as retinopathy, nephropathy, neuropathy and cardiovas- erties such as antioxidant, toxicological studies, anti-hyper- cular disease.2 Type 2 diabetes mellitus (T2DM) is one of cholesterolemic, and anti-diabetic effect in in-vitro, ex-vivo 9-12 the most common forms of diabetes in worldwide. In India, and in-vivo models. In our previous study, Morus Indica- around 69.2 million people are prone to T2DM and it has the G4 (MI-G4) variety exhibited the highest AGEs inhibition in second-highest number of people living with DM worldwide BSA-glucose model which could be due to the presence of 13 following China.3 polyphenols and phenolic compounds. Further, we isolated the bioactive compound of apigenin (API) and showed poten- Even though powerful anti-diabetic drugs are offered for the tial AGEs inhibition in all stages of protein glycation.14 Be- management of diabetes, there has been no drug developed sides, API is also proved to inhibit Aldose reductase (ALR) 4 which has no side effect(s) with low cost-effective. There- activity, one of the major complications of diabetes (cataract) fore, natural products have stimulated a new wave of research in the lens.15 Literature reports research studies carried out in 5, to look for more efficacious agents with lesser side effects. the crude extract of MI and very limited studies have investi- 6 Traditional knowledge with its holistic and systematic ap- gated the role of active ingredients from MI of G4 variety for Corresponding Author: Dr. Asna Urooj, Professor, DOS in Food Science and Nutrition, University of Mysore, Manasagangotri, Mysuru-570006, Karnataka, India; Contact: +91-9448489334; Telefax: +91 821 241 9632; Email: [email protected] ISSN: 2231-2196 (Print) ISSN: 0975-5241 (Online) Received: 23.07.2020 Revised: 27.09.2020 Accepted: 04.11.2020 Published: 16.01.2021 Int J Cur Res Rev | Vol 13 • Issue 02 • January 2021 100 Anandan et al.: Hypoglycemic effects of apigenin from morus indica in streptozotocin induced diabetic rats their beneficial effects on DM. Therefore, the present study At the end of the study, the rats were made to fast for 12 h was undertaken to assess the role of isolated API from MI for before blood collection. For ease of handling, before blood anti-diabetic efficacy in STZ-induced diabetic rats. sampling, the rats were anaesthetized with diethyl ether. The blood samples were collected by cardiac puncture using 25 G, 1” needle. Approximately 5 ml of blood was taken and MATERIALS AND METHODS dispensed into labelled plain tubes. The blood samples were Chemicals and reagents then centrifuged at 3000 rpm at 4 °C for 15 min to separate Clinical diagnostics kits were purchased from M/s. Agappe the serum. The serum was stored at -40°C until biochemical Diagnostics Ltd, Kerala, India. All the chemical reagents assays were carried out. used in this experiment were of analytical grade. Estimation of body weight Collection of Plant material During the experimental period, the body weights of the ex- Morus Indica G4 leaves ((ISGR Reg. No.: 050564) were perimental rats were recorded on alternate weeks, i.e on day collected from Centre for Sericulture Research and Techni- 0, 14, 28 and 45 by using a digital balance. These weights cal Institute (CSRTI), Mysore district of Karnataka, India. were determined at the fixed time in the morning session Apigenin was isolated from 80% methanol extract of leaves throughout the experimental period. by preparative HPLC and characterized through Ultra per- formance liquid chromatography-mass spectra (UP-LCMS), Estimation of blood glucose Nuclear magnetic resonance (NMR), Fourier transform in- Blood glucose levels of experimental rats were measured us- frared spectroscopy (FTIR) and Scanning electron micro- ing a glucometer (Roche, Germany) by taking 0.5 µL blood scope (SEM).14 from lateral tail vain onto the test strip on days of bodyweight determination. Experimental rats In this experimental study, twenty-four healthy adult male Biochemical studies Wistar Strain rats (140-190 g), were procured from the ani- Blood serum was used for the evaluation of lipid profile [total mal house facility of the University of Mysore (UoM), Mys- cholesterol (TC), triglyceride (TG) and high-density lipoprotein uru. The obtained animals were kept in polyacrylic cages. (HDL)], renal function test [Creatinine (CR), bilirubin (BIL), The animals were maintained under standard conditions, blood urea nitrogen (BUN)] and liver function test [alanine with a temperature of 22 ± 2˚C, a regular 12/12 hour light- aminotransferase (ALT), aspartate aminotransferase (AST), al- dark cycle. The animals were fed with pellet diet and water kaline phosphatase (ALP) including, total protein (TP), albumin ad libitum. The animal experimental protocol was approved (ALB)], using commercial kits from (Agappe Diagnostics Ltd., by the Ethics Committee of the UoM, Manasagangotri, Mys- India), according to the manufacturer’s protocol. uru. (Animal Sanction Order No: UOM/IAEC/05/2017). Histopathological studies Induction of diabetes and treatment Small portions of the kidney and liver were fixed in 10% Diabetes was induced by a single intraperitoneal injection of formaldehyde and were dehydrated with graded ethanol se- streptozotocin (STZ) to animals fasted overnight at a dose ries (50-100%) and were embedded in paraffin. The paraffin of 45 mg kg-1 body weight citrate buffer (pH 4.5) and the blocks were subsequently cut into (4-5 m) and stained with normal control rats received freshly prepared citrate buffer haematoxylin and eosin (HE) dyes. The slides were exam- (pH 4.5) alone.16 Diabetic condition in rats was confirmed ined under a microscope for histopathological휇 changes. by measuring the altered fasting blood glucose level (by Glucometer) after 72 h of STZ injection. The rats with fast- Statistical analysis ing blood glucose above 250 mg dL-1 were considered to be The values are expressed as mean ± SD. The data were sub- hyperglycemic and used for the experiment. Based on their jected to one-way ANOVA followed by Tukey’s multiple weights using randomized block design, the rats were divid- comparisons test for significant difference (≤0.05) using ed into four groups (consisting of 6 rats in each group) viz. SPSS16.0 software. Group I- Normal healthy control rat (NC), Group II- STZ- induced hyperglycemic rats without treatment (DC), Group III- STZ-induced hyperglycemic rats treated with Amino- RESULTS -1 guanidine (30 mg kg body weight by intraperitoneal) as Changes in blood glucose level and body a positive control, Group IV- STZ-induced hyperglycemic weight of experimental rats -1 rats treated with Apigenin (API) (50 mg kg body weight The blood glucose level and body weight of all the groups was given by orally) and treated accordingly for 45 days. of experimental rats are shown in Table 1. There was a 101 Int
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