Development and Validation of a High-Throughput Method for the Determination of Eight Non-Steroidal Anti-Inflammatory Drugs and Chloramphenicol in Milk, using Liquid Chromatography-Tandem Mass Spectroscopy

Abstract

A high-throughput LC-MS/MS method for simultaneous identification and quantitation of eight non-steroidal anti-inflammatory drugs (NSAIDs) and chloramphenicol (CAP) in bovine and ovine milk was developed. The method is capable of detecting and quantitating: Carprofen (CPF), tolfenamic acid (TLF), 5-hydroxy flunixin (FLU-OH), diclofenac (DCL), 4-methylaminoantipyrin (4MAA), meloxicam (MLX), ibuprofen (IBU), phenylbutazone (PBZ) and chloramphenicol (CAP) at their maximum residue limits (MRLs) or target analytical levels. Addition of ascorbic acid into the milk samples before the extraction step was found to be crucial for repeatability and intensity of PBZ and 4MAA. The method consists of the single extraction step with acetonitrile and a commercially available salt mixture, followed by evaporation of the supernatant and reconstitution. Complementary LC-MS/MS and LC-MS/MS/MS methods were developed for confirmation of ibuprofen and diclofenac respectively. The method was validated according to EU Commission Decision 2002/657/EC requirements. The method accuracy was in the range of 89-108% and coefficients of variation of the interday precision assessment varied between 3% and 16%. This method has been implemented at the Israel National Residue Control Laboratory for routine monitoring of NSAIDs and CAP residues in bovine and ovine milk.