METHOD VALIDATION FOR THE DETERMINATION OF RACTOPAMINE CHLORIDE IN RAW MATERIAL BY HPLC/UV

TF16

Ellen Figueiredo Freire (ellen.freire@ourofino.com) (1), Keyller Bastos Borges (2), Hélio Tanimoto (3), Cristiane Masetto de Gaitani (1)

(1) Departament of Pharmaceutical Sciences, Faculty of Pharmaceutical Sciences of Ribeirão Preto, University of São Paulo, Ribeirão Preto, Brazil.(2) Departament of Physics and Chemistry, Faculty of Pharmaceutical Sciences of Ribeirão Preto, University of São Paulo, Ribeirão Preto, Brazil.(3) Departament of Research and Analytical Development, Ouro Fino Saúde Animal, Cravinhos, Brazil.

The quantity of drugs used in animal production has grown exponentially, mainly due to new forms of intensive livestock. In special, the use of b-adrenergic agonists as growth promoters have grown considerably in recent years. Among the b-adrenergic agonists, ractopamine (RAC) is the most used as a nutrient repartitioning agent by diverting nutrients from fat deposition in animals to the production of muscle tissues. It promotes reduction of fat, increased muscle mass, and improved feed utilization efficiency in swine, cattle, and turkeys. The goal of this research was to optimize and validate a method for determination of the RAC chloride in raw material by HPLC/UV for using in quality control in veterinary industries. The best optimized HPLC conditions obtained were: Gemini C18 column Phenomenex^® (250 mm x 4.6 mm I.D., 5.0 mm particle size) at room temperature under isocratic conditions, acetonitrile/sodium acetate buffer (25:75, v/v) as mobile phase at a flow rate of 1.0 mL min^-1, UV detector at 275nm. Before starting the study for method validation, it is necessary to realize System Suitability through a series of tests to ensure that equipment used is able to generate results of precision and accuracy acceptable. The method shows theoretical plates (N) > 6290, retention time (Tr = 5.2 min.) with CV = 0.03%, area with CV = 0.18% and tailing factor (Tf) 10% < 1.16. After verifying that the system was in compliance, the method validation was performed. Some validations parameters obtained were: linearity 160-240 mg mL^-1 (r ≥ 0.99), limit of detection of 2.2 mg mL^-1, limit of quantification of 7.5 mg mL^-1, coefficients of variation and relative errors in precision and accuracy studies (within-day and between-day) were below 2%. In addition, the stability study showed that the sample were stable (CV < 0.2%), the robustness (flow rate, column and temperature) presented CVs < 2.0%. A simple and rapid high-performance liquid chromatography method using UV detection was developed and validated for the determination of RAC in raw material. (Acknowledgements: Ouro Fino Saúde Animal, FAPESP, CNPq and CAPES).