Quantitative determination of penicillin V and amoxicillin in feed samples by pressurised liquid extraction and liquid chromatography with ultraviolet detection

A rapid and simple method is proposed for the routine determination of amoxicillin (AMOX) and penicillin V (PENV) in swine feedingstuffs. The method is based on pressurised liquid extraction (PLE) followed by high performance liquid chromatography with ultraviolet detection (PLE–HPLC–UV) for antibiotic analysis. Parameters affecting PLE procedure, such as temperature, solvent composition, number of extraction cycles and sample cell size, were evaluated in order to achieve the highest extraction efficiency. The optimised method employed 11 mL extraction cells, acetonitrile–water mixtures (25:75, v/v) for AMOX and (50:50, v/v) for PENV, as extraction solvent, 102.07 atm of extraction pressure, 50 °C of extraction temperature, 5 min of static time and 60% flush volume of the cell size. Extracts were filtered and directly analysed by HPLC–DAD/UV without further clean-up. Mean recovery rates for feed samples fortified with 200–500 mg kg−1 of both antibiotics were 86% for AMOX (RSD ≤ 6%) and 95% for PENV (RSD ≤ 3%). The method was successfully applied to the analysis of a commercial medicated swine feedingstuff, and the results were in good agreement with those obtained using mechanical shaking or ultrasonic extraction combined with solid phase extraction (UE-SPE), previously applied in the literature for feed analysis. The extraction efficiencies were evaluated by statistical comparison (analysis of variance, ANOVA-single factor) of the results obtained using the different extraction methods. Compared to the alternative techniques, PLE offers several practical advantages: easy to perform, fast, savings in solvent volume and in time, all steps are fully automated and further clean-up is not necessary for penicillin analysis.