Determination of clavulanic acid residue in milk by high performance liquid chromatography-tandem mass spectrometry

Abstract

An analytical method was developed for the determination of clavulanic acid (CLAV/ in milk by high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS). A 2 g milk sample was deproteinized by ethanol. The supernatant was transferred into a pear-shaped bottle to be evaporated to about 0.5 mL, and the residue was dissolved with ammonium acetate solution. The sample was determined by HPLC-MS/MS after the purification. The chromatographic separation was achieved on a Luna 5u C8 column using 0.1% formic acid in water and acetonitrile as mobile phases with gradient elution. The identification of CLAV was carried out by MS/MS equipped with electrospray realization in negative scanning and multiple reaction monitoring (MRM) modes. Matrix-matched calibration standard was used for the quantification. The calibration curve showed perfect linear in the range of 10-400 μg/kg with the correlation coefficient of 0.999. The limit of detection (LOD, S/N ≥ 3) was 10 μg/kg in milk, and the limit of quantification (LOQ, S/N ≥ 10) was 20 μg/kg. The mean recoveries varied from 80.00% to 91.25% at me four spiked levels of LOQ, 1/2MRL (the maximum residue limit), MRL, and 2MRL with the relative standard deviations of 5.60%-8.77%. In conclusion, the established mthod can be applied for the determination of CLAV residues in milk. © 2010 Editorial Office of Chinese Journal of Chromatography, Dalian Institute of Chemical Physics, CAS.