| Quinolones(QNs)is widely used in aquiculture due to the good efficacy and low price.However,the drug resistance of pathogenic bacteria and the potential carcinogenicity of some QNs have attracted wide attention.Since the residue of QNs in animal-derived foods are usually very low,high-sensitivity detection methods are required to determine the residue.This study used Mylopharyngodon piceus,Misgurnus anguillicaudatus and Penaeus vannamei as experimental animals to establish the UPLC-MS/MS for testing residue of seven QNs drugs of these samples.The drugs are Enrofloxacin(ENR)?Ciprofloxacin(CIP)?Ofloxacin(OFL)?Norfloxacin(NOR)?Lomefloxacin(LOM)?Sarafloxacin(SAR)and Pefloxacin(PEF).This method provide a reference for the municipal(county)agricultural product quality and safety testing center to carry out the determination of QNs drug residue in aquatic food.Through the established method,sampling and detecting the status of seven QNs drug residues in aquatic products of a city,and analyzing the residual and elimination rules of enrofloxacin and ciprofloxacin in the Misgurnus anguillicaudatus.The main research contents are as follows.1.We established a method to test the residue of seven QNs in the muscle tissues of M.piceus,M.anguillicaudatus and P.vannamei,by the comparison of extraction solvents,optimization of purification conditions,chromatographic conditions and mass spectrometry conditions.The muscle tissue of M.piceus,M.anguillicaudatus,and P.vannamei was ultrasonically extracted with 1%formic acid-acetonitrile,purified by Oasis(?)Prime HLB cartridge,and determined by UPLC-MS/MS,The seven QNs were all quantified by internal standard method.0.01%formic acid water and pure methanol were used as mobile phases,the flow rate was 0.3 mL/min,and electrospray-multi-reaction monitoring positive ion mode monitoring.The result showed that the seven QNs had a good linear relationship in the concentration range of 1.0 ?g/kg to 100 ?g/kg in M.piceus,M.anguillicaudatus and P.vannamei,and the correlation coefficient(R2)was higher than 0.998.When enrofloxacin,ciprofloxacin,ofloxacin,norfloxacin,lomefloxacin,sarafloxacin,and pefloxacin are added to the blank sample at 0.5 MRL,1.0 MRL and 2.0 MRL,the recovery rates of the seven QNs drugs in M.piceus were 95.96%?104.72%?95.57%?105.08%?92.71%?99.56%?99.56%?105.31%?97.29%?100.14%?94.64%?103.05%?96.39%?102.77%.the relative standard deviations(RSD)are 4.91?6.28?4.13?5.29?1.76?5.82?2.41?3.92?2.33?4.57?1.02?2.16?3.17?5.67,Intra-day RSD were 2.19?6.54?2.05?4.64?0.95?4.44?3.35?6.04?0.68?3.81?2.85?5.23?3.24?6.06,Inter-day RSD were 4.28?7.54?2.05?4.64?0.95?4.44?3.35?6.04?0.68?3.81?2.85?5.23?3.24?6.06,Limit of detection(LOD)were 0.34?0.32?0.03?0.11?0.01?0.02?0.06,Limit of quantification(LOQ)were 0.87?0.56?0.05?0.50?0.04?0.05?0.10;The recovery rates of the seven QNs drugs in M.anguillicaudatus were 92.35%?105.03%?92.51%?109.38%?92.31%?104.36%?96.61%?104.36%?99.86%?103.78%?98.25%?103.37%?94.36%?106.04%,the relative standard deviations(RSD)were 3.14?3.43?3.21?4.63?2.14?5.21?1.54?3.65?0.91?3.76?2.06?4.34?1.18?6.17,Intra-day RSD were 4.09?6.37?4.44?6.75?3.18?5.24?1.84?4.49?2.68?6.94?3.70?5.46?1.85?4.46,Inter-day RSD were 6.83?7.88?6.29?7.30?4.62?7.27?3.29?6.19?3.47?8.27?5.21?6.98?2.48?6.59,Limit of detection(LOD)were 0.39?0.30?0.03?0.15?0.02?0.01?0.05,Limit of quantification(LOQ)were 0.94?0.50?0.06?0.53?0.05?0.05?0.10;The recovery rates of the seven QNs drugs in P.vannamei were 98.02%-103.07%?96.11%?104.39%?92.31%?99.78%?97.35%?106.76%?99.32%?105.03%?96.05%?102.09%?98.35%?106.72%,the relative standard deviations(RSD)were 1.46?3.91?2.37?4.31?1.95?4.66?2.95?5.00?1.79?3.89?1.48?3.42?2.90?5.09,Intra-day RSD were 3.04?5.08?3.27?4.56?1.96?7.05?4.61?7.73?3.91?6.00?0.29?5.29?3.74?8.29,Inter-day RSD were 2.90?7.49?3.60?5.14?2.96?8.14?4.85?7.81?4.85?5.29?1.45?5.34?3.42?8.06,Limit of detection(LOD)were 0.45?0.34?0.03?0.16?0.02?0.02?0.07,Limit of quantification(LOQ)were 1.02?0.67?0.08?0.62?0.05?0.06?0.12.The detection method has good correlation and high recovery rate,and meet the detection of 7 QNs in M.piceus,M.anguillicaudatus and P.vannamei.It provides a new detection method for the simultaneous detection of enrofloxacin,ciprofloxacin,ofloxacin,norfloxacin,lomefloxacin,sarafloxacin,and pefloxacin residues in animal-derived foods.2.Sampling and testing of 7 QNs drug residues in aquaculture aquatic products in a city 218 samples of farmed aquatic products were randomly selected,The detection method established in Chapter 1 were used to analyze enrofloxacin,ciprofloxacin,ofloxacin,norfloxacin,pefloxacin,lomefloxacin,and sarafloxacin in 218 batches of samples..The detection rate in 218 samples of seven kinds of QNs was 0.0%?68.2%,and the range of residual amount was 0.0?350.0 ?g/kg.According to the category of aquatic products,the detection rate of sea fishes(16 samples)was 25.0%,the detection rate of freshwater fish(118 samples)was 26.3%,freshwater prawns(26 samples)were not detected,and sea prawns(57 samples))The detection rate was 3.5%.Freshwater crabs(1 sample)were not detected.Freshwater fishes was the aquatic product category with a higher detection rate.According to the detected drugs,the detection rate of ENR was 17.4%,the detection range was 2.06?350.0?g/kg,the detection rate of CIP was 4.1%,and the detection content range was 2.65?12.0?g/kg.Ofloxacin,Pefloxacin,Lomefloxacin,Sarafloxacin and Norfloxacin were not detected.The safety evaluation of the consumption of aquatic products in this area was conducted with children,adults and elderly people.The result showed that the ratio of children to ADI was 21.9%,and the ratio of adults to ADI was 10.7%.The ratio of elderly people to ADI is 9.73%,which is still at a relatively safe intake level.3.We analyzed the residue and elimination rules of Enrofloxacin and Ciprofloxacin in M.anguillicaudatus,and provided medication guidance and withdrawal time for the cultured species of M.anguillicaudatus.At(20 ± 2)? water temperature,with Enrofloxacin and Ciprofloxacin as the target compounds,the method of drug administration,analysis of the residual amount and elimination of Enrofloxacin and CIP in the M.anguillicaudatus muscle tissue law.The result showed that the recovery rate of Enrofloxacin was 86.58%?92.36%,the RSD was 2.48?3.53,the recovery rate of Ciprofloxacin was 108.92%?114.45%,and the RSD was 2.17?4.55.The method has high recovery rate and repeatability.Enrofloxacin and Ciprofloxacin are eliminated in the M.anguillicaudatus according to the first-order kinetic process,and the half-lives(t1/2 ?)are 3.51 d and 5.49 d,respectively.According to the metabolic elimination model,the residual amount of Enrofloxacin in the M.anguillicaudatus was 102.14 ?g/kg on the 16th day,83.76?g/kg on the 17th day,11.52?g/kg on the 27th day,and 9.44 ?g/kg on the 28th day.The residual amount of Ciprofloxacin in the M.anguillicaudatus was 108.13?g/kg on the 14th day,94.47?g/kg on the 15th day,10.87?g/kg on the 31th day,9.50?g/kg on the 32th day.According to the MRL of Enrofloxacin and Ciprofloxacin in the Ministry of Agriculture Announcement No.235,under the conditions,the withdrawal period of Enrofloxacin should be more than 17 days,and the withdrawal period of Ciprofloxacin shall be more than 15 days.South Korea is a major importer of M.anguillicaudatus in the area,and its MRL requirement for seven QNs drugs does not exceed 10?g/kg.Therefore,it is recommended that the reasonable withdrawal period for exported M.anguillicaudatus should be more than 28 days. |
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