Study On Multi-residue Detection Of Aminoglycoside Antibiotics Based On Molecular Imprinting Technique

Aminoglycoside antibiotics are a kind of broad-spectrum antibiotics formed by the combination of amino sugars and amino cyclic alcohols through oxygen bridge.Because of its good antibacterial effect on Gram-positive and Gram-negative infections,it is widely used in animal husbandry,aquaculture and veterinary medicine.Due to the abuse and overuse of aminoglycoside antibiotics,aminoglycoside antibiotics residual in the environment and agricultural products will cause nephrotoxicity,ototoxicity and neuromuscular block.Therefore,the detection methods of aminoglycoside antibiotics have become the focus of researchers.The research and establishment of efficient pretreatment technology is of great significance for the trace detection of aminoglycoside antibiotics in complex matrix.At present,the pretreatment of this kind of antibiotics has some shortcomings,such as lack of selectivity,tedious operation steps,serious matrix interference and so on.In view of the above shortcomings,the molecularly imprinted recognition reaction with specific selection was introduced,and the molecularly imprinted polymer of novel aminoglycoside antibiotics was prepared.a method for the detection of antibiotic residues based on solid phase extraction(SPE)and magnetic solid phase extraction(MSPE)was established.The details are as follows:1.An instrumental method for liquid chromatography tandem mass spectrometry of seven aminoglycoside antibiotics was established.Under the mode of electrospray ion source,positive ion scanning and multi-reaction monitoring(MRM),the optimal parameters were determined by optimizing the mass spectrometry conditions(collision gas:5 psi;air curtain gas:10 psi;atomization gas:50 psi;auxiliary gas:50 psi;ion source temperature:500.0?;ionization voltage:5500.0 V;de-cluster voltage and collision voltage).Through the optimization of liquid chromatographic conditions,the liquid chromatographic parameters were determined(10 mM ammonium acetate/0.1%formic acid water as mobile phase An and acetonitrile as mobile phase B.the elution procedure was as follows:0 min² min,B:80%,2-3 min,B:80%⁵%,3-6 min,B:5%,6 min⁶.1min,B:5%⁸0%,6.1 min¹2 min,B:80%,HILIC column,column temperature 35?;.The flow rate was 0.3 mL/min).The seven antibiotics showed a good linear relationship in the range of 50 ng/mL¹000 ng/mL,R2 was more than 0.993,the detection limit was as low as 0.0024 ng/mL¹.24 ng/mL,the precision RSD was less than 9.5%.2.A novel virtual molecularly imprinted polymer(DMIPs)of aminoglycoside antibiotic(AAs)was prepared by precipitation polymerization using raffinose as template molecule,methacrylic acid as functional monomer and trimethylol propane triacrylate as cross-linking agent.The obtained DMIPs was characterized in detail,and its adsorption and recognition properties were evaluated.The results showed that DMIPs could specifically recognize six kinds of AAs with large adsorption capacity.The conditions of virtual molecular imprinting solid phase extraction(DMISPE)were optimized,including pH value,eluent,eluent and sample loading.Under the optimum conditions,a simple and efficient method for the determination of trace aminoglycoside antibiotics in environmental water samples based on DMISPE-HPLC-MS/MS was established.The method has a good linear relationship,and the correlation coefficients are all greater than0.9921.The recovery is 70.8%-108.3%,the relative standard deviation is 2.6%-11.4%,and the detection limit(S/N=3)of the method is 0.006-0.6 ng/mL.The results show that DMIPs has a good recovery effect,and the introduction of virtual template molecules avoids the problem of template leakage,so it has a good application prospect in the sample pretreatment of trace AAs in environmental water samples.3.The magnetic virtual template molecularly imprinted polymer was successfully prepared in this experiment.Fe₃O₄ was synthesized by coprecipitation method and modified with amino groups on its surface.Combined with surface imprinting technique,Fe3O4@SiO2-NH2@MIP was successfully prepared.The magnetic virtual molecularly imprinted polymer((DMMIPs))was characterized and analyzed by Fourier transform infrared spectroscopy(FTIR).The results show that DMMIPs is well dispersed and spherical.The adsorption effect of DMMIPs was evaluated by adsorption test and Scatchard equation,and the maximum apparent adsorption capacity was 8.7 mg/g and20.5 mg/g,respectively.Based on magnetic separation technology and magnetic solid phase extraction technology,the pH value,eluent,eluent and elution strategy were optimized,and the DMMISPE-HPLC-MS/MS method for five aminoglycoside antibiotics in milk matrix was established.The results showed that the recoveries of five antibiotics were 82.7%¹14.1%,and the relative standard deviations were 2.8%¹3.2%.The established method further reduces the time and operation steps of sample pretreatment,and realizes the selective and rapid enrichment of many kinds of aminoglycoside antibiotics in milk matrix.