Development and validation of screening and confirmatory methods for the detection of chloramphenicol and chloramphenicol glucuronide using SPR biosensor and liquid chromatography-tandem mass spectrometry

Biacore Q biosensor and liquid chromatography-tandem mass spectrometry (LC-MS/MS) based methods for the screening and confirmation of trace levels of chloramphenicol (CAP) and the mammalian metabolite chloramphenicol glucuronide (CAP-Glu) is reported. Both methods employ solvent extraction and clean-up by solid phase extraction (SPE) prior to analysis. The biosensor screening method utilises surface plasmon resonance (SPR) to determine chloramphenicol concentration in a range of matrices including honey and prawns. As the antibody used in the biosensor has a high cross-reactivity with CAP-Glu, direct detection of this metabolite is possible in matrices such as porcine kidney. LC-MS/MS is used in negative ion electrospray mode for the confirmatory procedure. Parent CAP is determined via the use of an internal standard. In the case of porcine kidney parent CAP is released from CAP-Glu following a short digestion with a beta-glucuronidase. All methods have been validated to the latest EU requirements (Commission Decision 2002/657/EC). The calculated decision limits (CCalpha) and detection capabilities (CCbeta) are less than 0.1 and 0.2 mug kg(-1) respectively for the screening and confirmatory techniques. Incurred tissues were used to study and confirm the long-term reproducibility and agreement between methods. In addition, the stability of CAP and CAP-Glu has been investigated. The analytes are stable under nearly all storage conditions for at least 20 weeks. Crown Copyright (C) 2004 Published by Elsevier B.V. All rights reserved.