Characterization and estimation of sulfaquinoxaline metabolites in animal tissues using liquid chromatography coupled to tandem mass spectrometry

Abstract

Sulfaquinoxaline (SQX) is a sulfonamide that is widely used in veterinary medicine, with a maximum residue limit (MRL) established for several food matrices. In Brazil, the MRL for liver and muscle is 100 μg kg⁻¹ for equine, bovine, poultry and swine. This value includes not only free drug but also the sum of all metabolites. Several reports showed limitations for SQX residue analysis, especially when mass spectrometry methods were used. These limitations include poor recoveries and unacceptable accuracy responses. In this work a metabolite of SQX, present in liver and kidney samples, was identified. The structure proposed was a hydroxylated form of SQX, called SQX-OH, with an m/z of 317. SQX-OH is also produced in vitro in equine, swine and bovine liver samples. The influence of time, temperature, solvent and dehydration was evaluated in the formation of SQX-OH. Different degrees of hydroxylation were observed in matrices. The N₄-acetyl derivates for both SQX and SQX-OH were also detected. In equines, the metabolism of SQX is complete. The mass spectrometry analysis of SQX-OH was determined in vitro using equine liver microsomal fraction. The characterization of this compound was performed using liquid chromatography coupled to mass spectrometry in tandem mode. The fragmentation profile of SQX-OH was seen to be similar to that of the sulfonamides group, producing two high abundant daughter ions: 317 > 156 and 317 > 108, common to most sulfonamides. The main conclusion of this work is that the residue analysis of SQX needs to consider the presence of the SQX-OH in order to give more realistic results, especially when using MRM transitions.