Turk J Biol 36 (2012) 574-579 © TÜBİTAK doi:10.3906/biy-1112-11 Antihyperglycemic, antioxidant, and cytotoxic activities of Alocasia macrorrhizos (L.) rhizome extract Md. Masudur RAHMAN1, Md. Aslam HOSSAIN2, Saiful Alam SIDDIQUE1, Kaishar Parvej BIPLAB1, Md. Helal UDDIN1 1Department of Pharmacy, International Islamic University Chittagong, Chittagong 4203 - BANGLADESH 2Department of Pharmaceutical Chemistry, Faculty of Pharmacy, University of Dhaka, Dhaka 1000 - BANGLADESH Received: 06.12.2011 ● Accepted: 10.05.2012 Abstract: Alocasia macrorrhizos (L.) (Araceae), a coarse and erect plant that is widely cultivated to be eaten as a vegetable throughout Bangladesh, has been reported to possess a number of medicinal properties. Th e antihyperglycemic, antioxidant, and cytotoxic eff ects of the methanol extract of A. macrorrhizos rhizomes were investigated. Th e extract was given at a single dose (250 and 500 mg/kg) in alloxan-induced hyperglycemic mice. Th e extract at 500 mg/kg produced a signifi cant (P < 0.05) decrease in the blood glucose level (55.49%) at 8 h of treatment when compared with the control and was comparable with the reference drug, metformin. Th e extract was also subjected to antioxidant potentiality and brine shrimp lethality bioassays. Th e IC50 value of the extract was 693.0 μg/mL and the LC50 was 188.14 μg/mL. Key words: Antihyperglycemic, alloxan, Alocasia macrorrhizos, antioxidant activity, cytotoxicity, rhizome Introduction Alocasia macrorrhizos (L.) G.Don (synonym: Diabetes mellitus is the seventh leading cause of death Alocasia indica) is a very robust evergreen herb, among high-income countries; it is characterized by stem caulescent, leaves petiolate, green, stout, leaf blade erect, broadly ovate, large, margin entire or high levels of blood glucose resulting from defects subundulate, growing up to 1.2-1.8 m in height in insulin production (1). Statistical projection under favorable conditions. It is commonly known as suggests that India, China, and the United States Mankachu in Bengali and Giant Taro or Elephant Ear will have the highest number of people with diabetes Taro in English, and it belongs to the family Araceae, (2) and Bangladesh will have the seventh highest distributed throughout Bangladesh and other parts number of diabetics in the world by 2030 (3,4). of Asia. Th is plant contains fl avonoids, cynogenetic Th e increasing worldwide incidence of diabetes glycosides, ascorbic acid, gallic acid, malic acid, mellitus in adults constitutes a signifi cant impact oxalic acid, alocasin, amino acids, succinic acid, on the health, quality of life, and life expectancy of and β-lectins (6). Th e plant has been used as folk patients, as well as on the global public health care medicine in diff erent districts of Bangladesh. Th e system. Th e long-term manifestation of this disease various parts are cooked and eaten as a vegetable due can result in the development of vascular disorders to being an enriched source of protein. It has been such as retinopathy, nephropathy, neuropathy, and traditionally used (all parts) for the treatment of angiopathy (5). diabetes (7). Stem juice is applied to prevent edema, 574 M. RAHMAN, A. HOSSAIN, S. A. SIDDIQUE, K. P. BIPLAB, H. UDDIN pain, and bleeding from cuts and wounds (8). Th e Drugs leaves of A. macrorrhizos are also used to prevent Alloxan monohydrate (Sisco Research Laboratories iron defi ciency and to enhance eye sight, while the Pvt. Ltd., India), metformin hydrochloride (Square whole plant is utilized for pus in the ears, jaundice, or Pharmaceuticals Ltd., Bangladesh), 1,1-diphenyl-2- constipation (9-11). It is experimentally reported that picrylhydrazyl (DPPH; Sigma Chemical Co., USA), the leaf extract possesses antimicrobial, antifungal, and ascorbic acid (Fine Chem. Ltd., India) were used. antioxidant, hepatoprotective, antidiarrheal, and All of the other chemicals and reagents used were of antiprotozoal activities (12-16). Alocasin, an analytical grade. antifungal protein, and a new ceramide from A. macrorrhiza have been isolated (17,18). Th e literature Induction of diabetes and treatments survey revealed that there are no scientifi c studies Th e alloxan was fi rst weighed individually for each carried out regarding the antidiabetic, antioxidant, animal according to its weight and then solubilized and cytotoxic activities of A. macrorrhizos rhizomes with 0.2 mL of saline (154 mM NaCl) just prior to substantiate its therapeutic claim. Hence, in the to injection. Diabetes was induced by injecting present study, the methanolic extract of the rhizomes the alloxan at a dose of 150 mg/kg body weight was examined for its antihyperglycemic, antioxidant, intraperitoneally aft er overnight fasting. Aft er 48 h, and cytotoxic properties. animals with fasting blood glucose levels from 16 to 19 mmol/L were separated and included in the study. Materials and methods Twenty-fi ve male Swiss albino mice were used, divided randomly into 5 groups containing 5 mice Plant material each. Group 1 served as a control, which received a A. macrorrhizos rhizomes were collected from a vehicle alone. In groups 2-5, each animal received a local vegetable market in Chittagong, Bangladesh, single dose of alloxan (150 mg/kg intraperitoneally) in the month of August, and were authenticated by aft er overnight fasting. Group 1 received only Dr Shaikh Bokhtear Uddin, Associate Professor, dimethyl sulfoxide (DMSO) as the normal control Department of Botany, University of Chittagong, group and group 2 was the diabetic control group, Chittagong, Bangladesh. which did not receive either metformin or rhizome Preparation of extract extract. Metformin (150 mg/kg body weight) was injected intraperitoneally into group 3, and extract Th e rhizomes were sun-dried and ground. Th e at a dose of 250 mg/kg body weight and 500 mg/ ground rhizomes (250 g) were soaked in a suffi cient kg body weight was injected intraperitoneally into amount of methanol for 1 week at room temperature, group 4 and group 5, respectively. Metformin and with occasional shaking and stirring, and were then the extract were both dissolved in 0.2% DMSO fi ltered through a cotton plug followed by Whitman vehicle. Blood samples were then analyzed for blood fi lter paper No. 1. Th e solvent was evaporated under glucose content at 0, 2, 4, 8, and 16 h, respectively, vacuum at room temperature to yield a semisolid. using a glucometer kit (Accu-Check Active, Roche Th e extract was then preserved in a refrigerator until Diagnostic GmbH, Germany). further use. Antioxidant activity Experimental animals Male Swiss albino mice, about 25-30 g in weight and DPPH free radical scavenging activity 4-6 weeks of age, were collected from the International DPPH scavenging activity was carried out using the Center for Diarrheal Diseases Research, Bangladesh method of Braca et al. (19). Diff erent concentrations (ICDDRB), and housed in polypropylene cages under (500, 300, 100, 50, and 10 μg/mL) of Alocasia controlled conditions. Th e animals were exposed to macrorrhizos extract were dissolved in methanol and an alternating cycle of 12 h of light and 12 h of dark. placed in diff erent test tubes, and 3 mL of a 0.004% Th e animals were allowed free access to drinking w/v methanol solution of DPPH was added to each water and pellet diet, obtained from the ICDDRB. test tube. Absorbance at 517 nm was determined aft er Th e mice were acclimatized for 7 days. 30 min against a blank, and the percent inhibition 575 Antihyperglycemic, antioxidant, and cytotoxic activities of Alocasia macrorrhizos (L.) rhizome extract activity was calculated from [(A0 – A1) / A0] × 100, inspected using a magnifying glass and the number where A0 is the absorbance of the control and A1 is of surviving nauplii in each vial was counted. From the absorbance of the sample. Ascorbic acid was used these data, the percent (%) mortality of the brine as a reference standard and dissolved in methanol to shrimp nauplii was calculated for each concentration make the stock solution with the same concentration. using the following formula: Th e control sample was prepared containing the % mortality = (N / N ) × 100, same volume without any extract or reference drug. t 0 where N = the number of killed nauplii aft er 24 h Methanol served as a blank. Th e inhibition curves t of incubation and N = the number of total nauplii were prepared and the half maximal inhibitory 0 concentration (IC ) values were calculated using transferred, i.e. 10. Th e median lethal concentration 50 (LC ) was then determined. linear regression analysis. 50 Reducing power capacity Statistical analysis Th e reducing power of the extract was evaluated using Th e experimental data are presented as the means ± the method of Oyaizu (20). Diff erent concentrations standard error of the mean (SEM). Th e results were of rhizome extract of A. macrorrhizos (125, 250, 500, analyzed for statistical signifi cance using one-way and 1000 μg/mL) in 1 mL of distilled water were ANOVA followed by Dunnett’s test using GraphPad mixed with phosphate buff er (2.5 mL, 0.2 M, pH 6.6) Prism Version 5.0 for Windows (GraphPad Soft ware, USA). P < 0.05 was considered signifi cant. and potassium ferricyanide [K3Fe(CN)6] (2.5 mL, 1% w/v). Th e mixture was incubated at 50 °C for 20 min. Aft er incubation, 2.5 mL of 10% trichloroacetic acid Results and discussion solution was added to each tube and the mixture was centrifuged at 3000 rpm for 10 min. Subsequently, 5 Th e methanol extract of A. macrorrhizos rhizomes mL of the upper layer solution was mixed with 5 mL (250 and 500 mg/kg) exhibited a dose-dependent of distilled water and 1 mL of ferric chloride solution antihyperglycemic activity. Although the reduction (0.1% w/v), and the absorbance was measured at 700 of blood glucose level (41.70%) at a dose of 250 mg/ nm.