Comparison of ion-pairing and ion-suppressing liquid chromatographic methods for the determination of pyrimethamine and ormetoprim in chicken feed

Abstract

A high-performance liquid chromatographic (HPLC) method is developed to simultaneously determine pyrimethamine (PYR) and ormetoprim (OMT) in chicken feed. In the ion-pairing HPLC determination of PYR and OMT, the relation between the retention factor (k1) and the concentration of the organic phase (acetonitrile) shows a characteristic curve. The k1 value first decreases and then increases slowly with increasing concentrations of acetonitrile, but then increases rapidly when the acetonitrile concentration increases to 90%. Resolutions (Rs) of PYR and OMT decrease gradually when the concentration of organic phase increases. Increasing the concentration of the pairing ion sodium 1 -octanesulfonate (PIC B-8) can decrease the k1 and Rs values. Optimum values of k1 and Rs are obtained using 82% acetonitrile in 0.005M PIC B-8. In ion-suppressing HPLC, varying the concentration of Na2HPO4 has little effect on either the k1 or Rs values of PYR or OMT at pH 7.5. However, at pH 4.0, k1 and Rs decline when the concentration of Na2HPO4 increases. In general, ion-pairing HPLC generates more satisfactory results than ion-suppressing HPLC. Using 82% acetonitrile in water containing 0.001M PIC B-8 as the mobile phase, linear calibration curves are obtained in the range from 1 to 5 mg/L of PYR and OMT. Sulfamonomethoxine, sulfadimethoxine, sulfaquinoxaline, trimethoprim, amprolium, clopidol, and nicarbazin do not interfere with the detection of PYR or OMT. The recoveries of PYR from spiked feed at 1 and 5 mg/Kg are 73.0% and 72.0%, respectively, and those of OMT from spiked feed at 3 and 7 mg/Kg are 50.3% and 53.6%, respectively.