Validation of an analytical method for the determination of spiramycin, virginiamycin and tylosin in feeding-stuffs by thin-layer chromatography and bio-autography

Abstract

An inter-laboratory validation was carried out to determine the performance characteristics of an analytical method based on thin-layer chromatography (TLC) coupled to microbiological detection (bio-autography) for screening feed samples for the presence of spiramycin, tylosin and virginiamycin. Twenty-four samples including blank samples and samples with concentrations of the target analytes ranging between 1 and 5 mg kg⁻¹ (expressed in microbiological activity) were analysed by seven laboratories participating in the study. The required detection limit was 1 mg kg⁻¹ (expressed in microbiological activity). For spiramycin, acceptable values for the sensitivity (at least 95%) indicating the rate of correct positive results were obtained for samples containing this substance at or above 2 mg kg⁻¹, whereas at 1 mg kg⁻¹, the sensitivity rate dropped to about 70%. Therefore, it was concluded that the detection limit was 2 mg kg⁻¹. For tylosin and virginiamycin, acceptable values of the sensitivity were obtained for all concentrations including 1 mg kg⁻¹. Therefore, the method fulfils the criterion regarding the required sensitivity at the target detection limit for tylosin and virginiamycin.

Keywords:

• Tylosin
• virginiamycin
• spiramycin
• antibiotics
• inter-laboratory comparison
• feeding-stuffs
• thin-layer chromatography (TLC)

Acknowledgements

The authors would like to thank the different institutes participating in this ring trial, more specifically: DGCCRF-L35 (Rennes, France); Bayerisches Landesamt für Gesundheid und Lebensmittelsicherheit (Oberschleissheim, Germany); LUFA-ITL (Kiel, Germany); LUFA Dep. Microbiology (Speyer, Germany); Istit. Zoo. Sper. Lobardia E. Romagna. Bromat. Feeds Lab. (Brescia, Italy); AGES (Vienna, Austria); and EZN INIAP (Vale de Santarem, Portugal). The SIMBAG-FEED project was funded by the European Commission, Direction General RTD, 5th Framework Programme, Competitive and Sustainable Growth (GROWTH) Programme, Measurements and Testing Generic Activity, Contract No. G6RD-CT-2000-00413. The authors are also very grateful to Gerald Kramer (IRMM) and Hakan Emteborg for the test samples prepared within this project and utilized in the present study.