Abstract

DEVELOPMENT AND VALIDATION OF A HPLC WITH SOLIDPHASE CLEAN-UP AND PRE-COLUMN DERIVATIZATION METHOD FOR THE SIMULTANEOUS DETERMINATION OF THREE ?- LACTAM ANTIBIOTICS RESIDUES IN EGGS

Suying Ma, Yongli Shi, Haixia Lv, Xiaojun Shang*

Abstract

An accurate and sensitive HPLC method with UV detector was developed for the simultaneous determination of three β- lactam antibiotics residues in eggs. The residues in the eggs samples were extracted with acetonitrile, cleaned up with C18 solid phase cartridge and derivated with 1, 2, 4 – triazole. The analytes were separated on an ODS-BP C18 column. The mobile phase was a mixture of acetonitrile – 0.1mol/L phosphate buffer pH 6.5 (25:75; V/V) containing 0.015 mol/L tetrabutyl ammonium bromide and sodium thiosulfate. The flow rate was 1.0 mL/min and the wavelength was set at 325nm. The method was validated according to the Directive 2002/657/EC and subsequently applied to twenty commercial eggs samples. The method showed good linear relationship over the concentration range of 2.5 to 100.0μg/kg for amoxicillin, ampicillin and 30.0 to 300.0 μg/kg for oxacillin with the coefficient of variation was over 0.99. The average recoveries of amoxicillin and ampicillin residues in eggs spiked from 2.5 μg/kg to 25.0 μg/kg ranged from 69.3% to 81.8 % with the relative standard deviations of less than 9.2 %, while the average recoveries of oxacillin residues in eggs spiked from 30.0 μg/kg to 300.0 μg/kg ranged from 72.4% to 82.7 % with the relative standard deviations of less than 9.6%. The limit of detection and quantification were 2.5 μg/kg, 2.5μg/kg and 30.0μg/kg for amoxicillin, ampicillin and oxacillin, respectively. Only one egg sample was found to contain amoxicillin at concentration of 7.2±3.6 μg/kg. The proposed HPLC method could be applied to the determination of amoxicillin, ampicillin and oxacillin residues in eggs.

Keywords: HPLC, UV, Amoxicillin, Ampicillin, Oxacillin, Residues, Eggs.