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Rourea Cuspidata: Chemical Composition and Hypoglycemic Activity

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Rourea Cuspidata: Chemical Composition and Hypoglycemic Activity 712 Asian Pac J Trop Biomed 2017; 7(8): 712–718 Contents lists available at ScienceDirect Asian Pacific Journal of Tropical Biomedicine journal homepage: www.elsevier.com/locate/apjtb Original article http://dx.doi.org/10.1016/j.apjtb.2017.07.015 Rourea cuspidata: Chemical composition and hypoglycemic activity Manuela M. Laikowski1, Paulo R. dos Santos1, Debora M. Souza1, Luciane Minetto1, Natalia Girondi2, Camila Pires2, Gisiele Alano2, Mariana Roesch-Ely3, Leandro Tasso1,2, Sidnei Moura1* 1Laboratory of Natural and Synthetics Products, University of Caxias do Sul, Caxias do Sul, Brazil 2Laboratory of Pharmacology, University of Caxias do Sul, Caxias do Sul, Brazil 3Laboratory of Genomics, Proteomics and DNA Repair, University of Caxias do Sul, Caxias do Sul, Brazil ARTICLE INFO ABSTRACT Article history: Objective: To investigate the antidiabetic effect of Rourea cuspidata hydroalcoholic Received 23 Jun 2017 stem extract in normal and streptozotocin-induced diabetic rats. Received in revised form 13 Jul 2017 Methods: In order to evaluate the chemical composition, different extracts from stem in Accepted 26 Jul 2017 ascending solvent order of polarity were prepared. The extracts were analyzed by high Available online 4 Aug 2017 resolution mass spectrometry and 7 compounds were identified, including hyperin, an important and already reported active compound in the literature. Hyperin was also quantified by HPLC-UV in all the extracts. The hydroalcoholic stem extract (Ss5), which Keywords: showed the highest concentration of hyperin, was administered to STZ-induced diabetes Rourea cuspidata rats to evaluate the potential hypoglycemic activity. Total cholesterol, HDL, triglycerides, Diabetes ALT and AST were also evaluated. In the present study, the effects of oral administration Phytochemical characterization of hydroalcoholic stem extract (200 mg/kg b. wt.) for 28 days on the level of serum HRMS glucose, total cholesterol, HDL, triglycerides, aspartate amino transferase (AST) and alanine amino transferase (ALT) in normal and streptozotocin-induced diabetic rats were evaluated. Histopathological changes in diabetic rats' pancreas were also studied. Results: The extract exposition demonstrated hypoglycemic effect like the drug control glibenclamide. The extract was able to increase the HDL levels. Histopathological study on diabetic rats' pancreas after extract treatment showed morphological alterations in STZ-induced diabetes rats, which were apparently restored after extract treatment. Conclusions: This work demonstrates the potential use of R. cuspidata as hypoglycemic agent in the treatment of diabetes. 1. Introduction representative ones. Rourea is a pantropical genus with about 100 species, 48 of them are in the Neotropics [3,4]. In Brazil, Brazil is known for its biodiversity with 40–55 thousand Connaraceae family native species are found mainly in the plant species distributed across several biomes with a large Amazonic region [5]. number of plants used by the population in folk medicine [1]. Few species of Rourea genus have been screened so far for The Connaraceae family comprises twenty genera and about their biological activities. Among them, the antidiabetic and 350 species distributed in Africa, Southeast Asia and tropical antimalarial potential of Rourea minor (R. minor), antioxidant America [2]. In tropical America, this family is formed by five and hepatoprotective activities of Rourea induta and anti- genera and among them Connarus and Rourea are the most inflammatory and hepatoprotective activities of Rourea cocci- nea can be noticed [6–8]. According to phytochemical investigations on Rourea spe- *Corresponding author: Sidnei Moura, Technology Department, Biotechnology cies, it was described that two glycoside derivatives (rourinoside Institute, University of Caxias do Sul, 1130 Francisco Getúlio Vargas St., CEP 95070- 560, Caxias do Sul, Brazil. and rouremin) and derivative [1-(26-hydroxyhexacosanoyl)- Tel: +55 54 3218 2100 glycerol], actives as antimalarial, as well as nor-sesquiterpene E-mail: [email protected] (S. Moura). (dihydrovomifoliol-9-b-D glucopyranoside) were present in Peer review under responsibility of Hainan Medical University. The journal [8] implements double-blind peer review practiced by specially invited international R. minor . It was also described that the compounds quinone editorial board members. (rapanone) and cianidine (leucopelargonidine) were from 2221-1691/Copyright © 2017 Hainan Medical University. Production and hosting by Elsevier B.V. This is an open access article under the CC BY-NC-ND license (http:// creativecommons.org/licenses/by-nc-nd/4.0/). Manuela M. Laikowski et al./Asian Pac J Trop Biomed 2017; 7(8): 712–718 713 Rourea santaloides [9]. From Rourea microphylla, flavonoids respectively. The samples were separated by liquid chromatog- (quercetin, quercetin 3-O-b-L rhamnopyranoside, astilbin, raphy (UFLC system), consisted of a LC-20ADXR pump, a SIL- hyperin, rutin, and kaempferol), anthraquinones (physcion and 30AC autosampler (Shimadzu®). Chromatographic separations erythroglaucin), triterpenes (23-hydroxybetulinic acid, ursolic were performed on a Shim-pack XR-ODS (30 mm × 2.0 mm, acid and hederagenin), coumarin (daphnetin), phytosteroids (b- 2.2 mm) column. A hybrid high-resolution and high accuracy ® sitosterol, b-sitosteryl-b-D-glucopyranoside), besides fatty acids, microTof (Q-TOF) (Bruker Scientific) was used for detection, alkane, alcohol, and glycery derivative were found [10,11]. From with electrospray ionization (ESI) source (MicrOTOF-QII the leaves of Rourea induta, flavonoids were found (such as Bruker® Scientific) in positive and negative mode. The range of quercetin, hyperin, quercetin 3-O-a-L-arabinofuranoside, and mass was 50–1 200 m/z with two scans per second, providing the quercetin 3-O-b-D-xylopyranoside) [3]. resolution of 50 000 (FWHM). The drying temperature was 200 C Rourea cuspidata (R. cuspidate) Benth ex. Baker popularly and nitrogen was used for drying gas, in a 10 L/min flow. The known in Brazil as ‘miraruíra’, ‘cipo´ miraruíra’, and ‘muirar- ionization energy was 3.0 eV and the capillary voltage was uíra’, is a shrub of the Connaraceae family, common in Ama- 4 500 eV. The software was used to read the spectrum, Compass zonic region [12], which is used for diabetes treatment in folk DataAnalysis version 4.3 was used along with the following tools: medicine. The same activity was reported in R. minor [7]. Smart formula, Smart formula 3D and Send formula to Compound Although these studies involved Rourea species, there are few Crawler (Compound Crawler version 3.0). MetFrag 2010 was also scientific papers focusing on R. cuspidata. Therefore, the used to predict fragments with mass spectra in order to compare present work aimed to evaluate the chemical composition of withthepractice results,corroborating withtheidentificationofthe R. cuspidata stem extracts as well as the hypoglycemic compound. activity of the stem hydroalcoholic extract of the species. 2.5. Quantitative analysis by HPLC-UV 2. Material and methods The study was conducted according to Kalegari et al. [6] with 2.1. Chemical reagents slight modifications. Analytical HPLC experiments were performed with Shimadzu LC-20AD fitted with an analytical Streptozotocin and hyperin were purchased from Sigma– column (Agilent LiChrosfer 100 RP C18, 5 mm, 250 × 4.6 mm) Aldrich (Saint-Louis, Missouri, USA). Accu-check active and UV–vis detector (l 356 nm). The mobile phase consisted of monitor and test stripes were purchased from Roche Diagnostics water buffer acid solution (H2SO4 0.01 mol/L: H3PO4 0.02 mol/ (Mannheim, Germany). Acetonitrile, sulphuric acid, phosphoric L)-solvent A, and acetonitrile-solvent B, with 1 mL/min flow acid, formic acid, ammonia hydroxide, hexane, chloroform, rate following linear gradient over a total run time of 25 min, ethyl acetate, ethanol, haematoxylin, eosin, formaldehyde and initially 98%: 2% for A: B and at 25 min, 74%: 26% for A: B. paraffin were supplied by Merck (São Paulo, SP, Brazil). All chemicals were of analytic grade. 2.6. Animals 2.2. Plant material The study was conducted according to Abeeleh et al. [13] with slight modifications, and it was previously approved by R. cuspidata plant material was collected in Uaicurapa river, in Ethics Committee on Animal Use (CEUA)-University of Santo Expedito community, Parintins (02 370 4200 S, 56 440 0900 Caxias do Sul (Project number: 002/2013). Healthy, young W), Amazonas, Brazil, under authorization from Ibama number male adult Wistar rats, weighing 250–350 g, purchased from 02001.004236/2013-63, and identified by Dr. Juan David Revilla Technology and Science Foundation (Santa Maria, Brazil), Cardenas, from Herbarium of INPA (Instituto Nacional de Pes- were used in the study. The animals were housed under quisas da Amazonia),ˆ where a voucher specimen was deposited. standard conditions, kept on a 12 h light: 12 h dark cycle, and fed with a commercial rodent diet (Nuvital®) and water ad 2.3. Extraction libitum. The plant samples were dried in air oven at 45 C. The stem 2.7. In vivo study was separated from the bark and it was powdered. Stem was extracted under reflux (10 g of plant material with 200 mL of For the experiment, male Wistar rats were randomly solvent for 2 h) in ascending polarity order. The extracts were distributed into 4 groups (n = 5). Diabetes was induced through nominated according to the following condition: S = under i.p. administration of 55 mg/kg streptozotocin (STZ). After 24 h, reflux extract,
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