Multi-class/residue method for determination of veterinary drug residues, mycotoxins and pesticide in urine using LC-MS/MS technique

Abstract

Background Veterinary drugs are widely used in animals to prevent diseases and are a complex set of drugs with very different chemical properties. Multiclass and multi-residue methods for simultaneous detection of residues from veterinary drugs and contaminants in urine are very rare or non-existent. Therefore, the aim of this study was to develop and validate a sensitive and reliable quantitative LC-MS/MS method for simultaneous determination of a wide range of veterinary drug and pesticide residues and mycotoxins in bovine urine. This involved 42 veterinary drug residues (4 thyreostats, 6 anabolic hormones, 2 lactones, 10 beta agonists, 15 antibiotics, 5 sulphonamides), 28 pesticides and 2 mycotoxins. Stable isotopically labelled internal standards were used to facilitate effective quantification of the analytes. Analysis was performed in both positive and negative ionization modes with multiple reaction monitoring transitions over a period of 12 min. Results The parameters validated included linearity, limit of detection (LOD), limit of quantification (LOQ), detection capability (CCβ), decision limit (CCα), stability, accuracy and precision. The process followed guidelines of the regulation 2021/808/EC. The calibration curves were linear with coefficient of correlation (R2) from 0.991 to 0.999. The LODs were from 0.01 to 2.71 µg/L, while the LOQs were from 0.05 to 7.52 µg/L. The CCα and CCβ were in range 0.05–12.11 µg/L and 0.08–15.16 µg/L. In addition, the average recoveries of the spiked urine samples were from 71.0 to 117.0% and coefficient of variation (CV) < 21.38% (intraday and interday). Conclusion A new isotopic LC-MS/MS method has been developed, validated and applied for identification and quantification of 72 residues of veterinary drugs and pesticides and other contaminants such as mycotoxins in bovine urine. The most appropriated sample preparation procedures involved sodium acetate buffer, enzymatic hydrolysis using β-glucuronidase and cleanup solid phase extraction with OASIS SPE cartridges. The parameters were satisfactorily validated fulfilling requirements under Regulation 2021/808/EC. Consequently, the method could be used in routine analysis of bovine urine samples for simultaneous detection of veterinary drug and pesticide residues as well as contaminants such as mycotoxins.

Sample Preparation Techniques for Growth-Promoting Agents in Various Mammalian Specimen Preceding MS-Analytics

“…In the reviewed period, SPE was observed to be either combined with other extractions [ 27 , 28 ] or utilized as an independent method [ 10 , 28 , 29 , 30 , 31 , 32 ]. Sample preparation applying solely to SPE was focused primarily on BA only [ 10 , 29 , 30 , 31 ].…”

“…Overall, cation exchanger-based mixed-mode phases were the most prevalent sorbent type ( Figure 4 ). However, Hajrulai-Musliu et al [ 28 ] opted to use hydrophilic lipophilic balanced (HLB)-based SPE cartridges for the extraction of the residues of several BAs, divers other veterinary drugs, mycotoxins, and pesticides. In their manuscript, a mixed-mode strong cation exchanger-based SPE was compared to an HLB-based SPE as part of the method development.…”

“…In their manuscript, a mixed-mode strong cation exchanger-based SPE was compared to an HLB-based SPE as part of the method development. The authors reported a better recovery for the HLB cartridge in combination with previous enzymatic digestion by protease and deglucuronidation by β-glucuronidase (β-glc) [ 28 ]. Yikilmaz et al [ 27 ] omitted the deglucuronidation and applied C18-based SPE cartridges prior to LLE.…”

“…The LOD was determined to be as low as 0.009 ng/g (Table 2). Hajrulai-Musliu et al [28] tested an LLE protocol for BA, steroids, and a variety of other compounds and compared it to the previously described SPE protocol. However, the determined recoveries were not satisfactory compared to SPE [28].…”