Toxicon 51 (2008) 181–190 Purification and renal effects of phospholipase A 2 isolated from Bothrops insularis venom $ Marcus Davis Machado Braga a , Alice Maria Costa Martins b , Claudeˆnio Dio´genes Alves b , Dalgimar Beserra de Menezes a , Rene´ Duarte Martins c , Paulo Se´rgio Ferreira Barbosa c , Isadora Maria de Sousa Oliveira c , Marcos Hikari Toyama d , Daniela Oliveira Toyama e , Eduardo Brito dos Santos Diz Filho d , Fabio Henrique Ramos Fagundes d , Manasse´s Claudino Fonteles e , Helena Serra Azul Monteiro c,Ã a Department of Pathology, Federal University of Ceara, Fortaleza, Ceara´, Brazil b Department of Clinical and Toxicological Analyses, Federal University of Ceara, Fortaleza, Ceara´, Brazil c Department of Physiology and Pharmacology, Faculty of Medicine, Federal University of Ceara´, Fortaleza, Ceara´, Brazil d Sa˜o Vicente Unity, Campus of Litoral Paulista, Paulista State University (UNESP), Sa˜o Paulo, Brazil e Biological Science, Exact and Experimental Faculty, Presbiterian Mackenzie University, Sa˜o Paulo, Brazil Received 30 May 2007; received in revised form 21 August 2007; accepted 31 August 2007 Available online 11 September 2007 Abstract Bothrops insularis venom contains a variety of substances presumably responsible for several pharmacological effects. We investigated the biochemical and biological effects of phospholipase A 2 protein isolated from B. insularis venom and the chromatographic profile showed 7 main fractions and the main phospholipase A 2 (PLA 2 ) enzymatic activity was detected in fractions IV and V. Fraction IV was submitted to a new chromatographic procedure on ion exchange chromatography, which allowed the elution of 5 main fractions designated as IV-1 to IV-5, from which IV-4 constituted the main fraction. The molecular homogeneity of this fraction was characterized by high-performance liquid chromatography (HPLC) and demonstrated by mass spectrometry (MS), which showed a molecular mass of 13984.20 Da; its N-terminal sequence presented a high amino acid identity (up to 95%) with the PLA 2 of Bothrops jararaca and Bothrops asper. Phospholipase A 2 isolated from B. insularis (Bi PLA 2 ) venom (10 mg/mL) was also studied as to its effect on the renal function of isolated perfused kidneys of Wistar rats (n ¼ 6). Bi PLA 2 increased perfusion pressure (PP), renal vascular resistance (RVR), urinary flow (UF) and glomerular filtration rate (GFR). Sodium (%TNa + ) and ARTICLE IN PRESS www.elsevier.com/locate/toxicon 0041-0101/$ - see front matter r 2007 Elsevier Ltd. All rights reserved. doi:10.1016/j.toxicon.2007.08.017 $ Ethical statement: The experiments follow the methodology recommended by the international ethical standards of the scientific committee of our university (Comitteˆ de E ` tica e Pesquisa com Animais). Ã Corresponding author. Unidade de Pesquisas Clı´nicas/UFC, Departamento de Fisiologia e Farmacologia, Faculdade de Medicina, Universidade Federal do Ceara´ , CP 3229 Fortaleza, Ce 60420-970, Brazil. Tel.: +55 85 3223 6982; fax: +55 85 32815212. E-mail addresses: serrazul@baydenet.com.br, martinsalice@gmail.com (H.S. Azul Monteiro).