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Genetic Algorithm Based Optimisation of HPLC Separation of Veterinary Pharmaceuticals

Veterinary drugs are continually being released in the environment mainly as a result of manufacturing processes, disposal of unused or expired products, and excreta. Because of their physical and chemical properties, many of these substances or their bioactive metabolites end up in soils, sediments and water, where they can accumulate and induce adverse effects in terrestrial or aquatic organisms. High performance liquid chromatography (HPLC) has been developed for simultaneous analysis of three different classes of antimicrobials: fluoroquinolones (enrofloxacin), tetracyclines (enrofloxacin) and diaminopyrimidines (trimethoprim), commonly used for veterinary purposes and agricultural feed additives and suspected to leach into ground and surface water. HPLC separation was examined on Varian ProStar HPLC/DAD with C-18 column. Detection and quantification was performed at 254 nm. Optimum mobile phase composition was determinated using genetic algorithm approach. The best separations were achieved with mobile phase 0.5 % formic and 1% trifluoroacetic acid in 0.05 M ammonium acetate (pH=3)-methanol, 70+30 v/v. Limit of detection (LOD) and limit of quantification (LOQ) for determination at 254 nm were calculated: enrofloxacin (2 ug/mL, 3 ug/mL), oxytetracycline (1 ug/mL, 2 ug/mL), and trimethoprim (0.5 ug/mL, 1 ug/mL), respectively. The method was examined on spiked water samples. The pharmaceuticals were extracted from water using solid-phase extraction (SPE) procedure on C18 Empore extraction disks (Varian, USA).

enrofloxacin; oxytetracycline; trimethoprim; antibiotics; Solid-phase Extraction (SPE); High-performance liquid chromatography (HPLC/DAD)

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