Journal of Chromatography A, 1216 (2009) 4290–4294 Contents lists available at ScienceDirect Journal of Chromatography A journal homepage: www.elsevier.com/locate/chroma High-speed counter-current chromatographic isolation of ricinine, an insecticide from Ricinus communis Cristiane de Melo Cazal, Jaqueline Raquel Batalhão, Vanessa de Cássia Domingues, Odair Corrêa Bueno, Edson Rodrigues Filho, Moacir R. Forim, Maria Fátima G. Fernandes da Silva, Paulo Cezar Vieira, João Batista Fernandes ∗ Universidade Federal de São Carlos, Laboratório de Produtos Naturais, Departamento de Química, São Carlos, SP 13560-970, Brazil article info Article history: Available online 10 February 2009 Keywords: High-speed counter-current chromatography Ricinine Leaf-cutting ant Ricinus communis abstract The alkaloid ricinine, an insecticide for leaf-cutting ant (Atta sexdens rubropilosa), was obtained from Ricinus communis. A two-phase solvent system composed of CH 2 Cl 2 /EtOH/H 2 O (93:35:72, v/v/v) was used for high-speed counter-current chromatographic (HSCCC) isolation of ricinine in high yield and with over 96% purity, as determined by liquid and gas chromatography–mass spectrometry (LC–MS and GC–MS). Identification of ricinine was performed by comparison of 1 H NMR, 13 C NMR and LC–MS/MS data. © 2009 Elsevier B.V. All rights reserved. 1. Introduction Ricinus communis Jussieu (Euphorbiaceae) is distributed over many tropical areas and has other common names, such as Cas- tor bean tree and African oil tree. It is an exotic plant found in all regions of Brazil, known by the names of “mamona” or “car- rapateira” or “mamoneira” or “palma-cristi” [1]. R. communis was described as a plant toxic to the leaf-cutting ants, Atta sexdens rubropilosa [2]. Seeds of this oleaginous, R. communis, have been used in Brazil to produce castor oil, one of starting materials for biodiesel produc- tion, in the government program for bioenergy [3]. The extraction residue of the castor oil, as well as the leaves and flowers of R. communis contain the alkaloid ricinine (1, Fig. 1), at 0.77, 0.38 and 1.07% of their dried weights, respectively [4]. Ricinine is responsible by R. communis insecticide effect to leaf-cutting ants (Atta sexdens rubropilosa) [5]. Ricinine showed low toxicity to mice (LD 50 , intraperitoneal, subcutaneous, intravenous and orally: 19–20 mg kg -1 ) when compared with ricin, a protein toxin from R. communis, that is listed in the USA as a potential threat for the population and not available commercially in USA. Ricinine may be considered as a promising cognition-enhancing drug that may be used for the treatment of human amnesias [6]. ∗ Corresponding author at: Universidade Federal de São Carlos, Departamento de Química, Rodovia Washington Luiz Km 235, 13565-905 São Carlos, SP, Brazil. Tel.: +55 16 33518085; fax: +55 16 33518350. E-mail address: djbf@power.ufscar.br (J.B. Fernandes). The leaf-cutting ants of the genera Atta and Acromyrmex use as part of their food, sap from mostly fresh plant fragments and to grow their symbiotic fungi that metabolize polysaccharide to mono- and disaccharide, the main food of ants and causing considerable eco- nomic damage, due to defoliation of plants [7–9]. Control of this pest is still a challenge. Insecticides usually present only temporary effects and are sometimes, harmful to the environment, to man and other animals. Consequently, an extensive search for alterna- tive methods to control these insects has been made in an attempt to substitute traditional agrochemicals by highly specific insecticides that would be less aggressive to the environment. The counter-current chromatography benefits from several advantages when compared with the more traditional liquid–solid separation methods such as no irreversible adsorption, total recov- ery of injected sample, low solvent consumption and low time of analyses [10–14]. In this context, the subject of this work was the isolation of the alkaloid ricinine in enough amounts to perform biological assays against leaf-cutting ants and to make its nano-encapsulation, using high-speed counter-current chromatography (HSCCC). 2. Experimental 2.1. Apparatus The HSCCC instrument employed was from P.C. Inc. (Potomac, MD, USA). It was equipped with a quadruple multilayer coil of 1.68mm I.D. polytetrafluoroethylene (PTFE) tubing and had a total capacity of 443 mL. The  value varied from 0.50 at the internal 0021-9673/$ – see front matter © 2009 Elsevier B.V. All rights reserved. doi:10.1016/j.chroma.2009.02.008