Multi-residue Detection Technology And Application Of Antibacterial Veterinary Drugs In Dairy

In recent years,the extensive use of antimicrobial veterinary drugs in animal husbandry has led to veterinary drug residues in milk,which becomes an important issue affecting the quality and safety of milk and dairy.The simultaneous detection and rapid screening of veterinary drug residues such asβ-lactam and their inhibitors is an important link which can control the quality and safety of milk and dairy products.Time of flight mass spectrometry has a high resolution,which can screen and identify the target compounds and unknown compounds conveniently and rapidly,thus it can reduce the probability of“false positive”,and analyze hundreds of compounds in a detection process,then realize the accurate quantify and screening of veterinary drugs in milk.In this study,raw milk and UHT milk were used as the research objects,and the simultaneous detection conditions and stability of 8 kinds of target substances such as lactam antibiotics and inhibitors in milk were studied by liquid mass coupled method and time-of-flight mass spectrometry,and rapid screening of 27 veterinary drugs was performed in 200 milk samples.The main research contents and results are summarized as follows:1.A method for determination of residues ofβ-lactam antibiotics and their enzyme inhibitors in milk by liquid mass spectrometry was established.Taking milk as the object of study,β-lactam antibiotics(penicillin V,penicillin G,benzocillin,cloxacillin,ampicillin)and inhibitors(potassium clavulanate,tazobactam,sulbactam)were targeted.The pretreatment conditions,chromatographic conditions and mass spectrometry conditions were optimized.The samples were extracted with acetonitrile,and then defated with n-hexane,0.1%formic acid-acetonitrile solution was used as mobile phase for gradient elution,electrospray(ESI)multi-reaction monitoring(MRM)scanning was used,and matrix calibration was performed by external standard method.Within the range of 0.1~200μg/L,the correlation coefficient was r≥0.997;the detection limit(S/N=3)was 0.1~5.0μg/kg,and the quantitative limit(S/N=10)was 0.5~10.0μg/kg.At low,medium and high addition levels,the recovery of blank samples was 75.2%~109.5%,and the relative standard deviation RSD was 0.20%~4.5%;the recovery rate of in UHT milk was 75.7%~108.1%,and the RSD was 0.2%~1.9%.This method is accurate,rapid,simple,practical and feasible for the determination ofβ-lactam antibiotics and their inhibitors in milk.2.The stability ofβ-lactam antibiotics andβ-lactase inhibitors.At 20℃at different times(0 h,2 h,6 h,10 h,24 h),UHT milk as research object.100μL 10 ng/mL standard solution was added into the volume of 20mL.2 mL solution was filled into the centrifuge tube,deposited the protein with acetonitrile,blown with nitrogen gas,then redissolved,the detection concentration was 50 ng/mL.Continuous samples:the results showed that all drugs were basically stable within 10h,except for clavuric acid,which could decompose by 4%~6%,when laying them to 24 h at room temperature 20℃.The decomposition of clavulanic acid was about 10%,and penicillin G,penicillin V,benzeneazole also appeared different degree of10~20%of the loss contrast to the peak area of 1h.It is recommended to complete the sample determination and analysis within 10 hours.3.Time-of-Flight mass spectrometry was used to determine 27 veterinary drug residues in milk.The pretreatment conditions,the selection of purification column,the chromatographic conditions and the mass spectrometry conditions were optimized.The samples were extracted with 80%acetonitrile water,purified by Prime HLB column,blown with nitrogen,and redissolved.Mobile phase:A was 0.1%formic acid and B was acetonitrile in the positive ion mode,gradient elution,sample collection in MS~E mode.Through the collection of MS and MS/MS data,the veterinary drug database of Q-Tof was established.It can be used for the preliminary screening of suspected veterinary drug compounds.The screening parameters were optimized,and the mass deviation of all compounds was lower than 10 ppm,which could be used to screen dairy in batch.