Development of a whole liquid egg certified reference material for accurate measurement of enrofloxacin residue

•A certified reference material of enrofloxacin in whole liquid egg was developed.•Egg raw materials were obtained from laying hens following oral administration.•The certified reference material retained the resemblance to real samples.•Characterization, homogeneity, stability and uncertainty were fully studied. To ensure accurate and reliable measurement results, a whole liquid egg certified reference material (CRM) retaining the resemblance to real samples was first developed for improving measurement quality of enrofloxacin residue in eggs. The CRM raw material naturally containing enrofloxacin was obtained by sampling eggs from laying hens following oral administration of enrofloxacin. The homogenizing and bottling procedure was evaluated and applied on the bulk raw material to prepare the CRM candidate. For characterization of the CRM, liquid chromatography tandem isotope dilution mass spectrometry as a reliable reference method was studied and used for value assignment by all participating laboratories. The certified value was assigned to be 30.6 μg/kg with expanded uncertainty of 3.1 μg/kg (coverage factor k = 2, approximate 95% confidence interval) for enrofloxacin. In addition, homogeneity, long-term stability at −70 °C for one year and short-term stability at −18 °C, 4 °C and room temperature for nine days were assessed.