| Aminoglycosides are a large class of antibiotics that are characterised by two or more amino sugars linked by glycosidic bonds to an aminocyclitol component. Aminoglycosides are classified according to the pattern of substitution of the cyclo-dolichol.The two most important subclasses are streptidine(streptomycin and dihydrostreptomycin) and deoxystreptamine(gentamicin,kanamycin,and neomycin). Aminoglycoside antibiotics can block the synthesis of bacterial protein by binding irreversibly to ribosomes,which causes damage to the cell membranes.Aminoglycoside antibiotices have been widely used for many years in the prevention and control of animal diseases,which causes some problems in foods owing to drug residues.As a result there is increasing pressure on laboratories responsible for food safety to monitor the use of these drugs and ensure the safety of food for human consumption.Therefore, it is very important to accurately detect these antibiotics of low levels of in food.At present,some methods including microbiological method,immunoassay and instrumental analysis can be used to detect aminoglycoside antibiotices in animal-derived foods,such as pork and bee products.In instrumental analysis,high performance liquid chromatography(HPLC) is widely used.However,these methods are mostly used to detect only one or more aminoglycoside antibiotices,and seldom used in other animal-derived foods except pork and bee products.Moreover,these methods play little role in routine detection owing to their troublesome operation and low efficiency.Liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) has some advantages of high sensitivity,accurate quantification,and less interference and abundant detection information,and is more effective in residual confirmation.In this thesis,simultaneous detection method for multi-residues of aminoglycoside antibiotics in aquatic products was established by using LC-MS/MS technique.Main achievements were summarized as follows.1.Obtain the optimal extraction conditions for aminoglycoside antibiotics in aquatic products:the extraction efficiencies were examined by some protein precipitants, such as perchloric acid,trichloroacetic acid,and sodium tungstate.The results showed that the extraction efficiency was the highest by 5%of trichloroacetic acid.In addition, homogenization and sonication-assisted extraction were proved efficiently to extract aminoglycoside antibiotics in aquatic products.2.Obtain the optimal purification conditions for extracts:Several solid-phase extraction(SPE) columns such as Sep-Pak C18,HR-xSPE,Oasis MCX,and Oasis SCX were used in this studied,and some purification conditions were also examined.The results showed that the purification efficiency was the highest by using HR-xSPE column when heptafluorobutyric acid was used as ion pair regent,pH of the extraction was to adjusted as 4～5,and the cartridge was washed with methanol and ammonia methanol(10:90,v/v).3.Obtain the detection conditions for aminoglycoside antibiotics:Some parameters of chromatographic and mass spectra,such as columns,mobile phases,gradient program and column temperature were examined.Thus,the detection conditions were obtained, i.e.,LC separation was performed on a Waters Acquity UPLCTM BEH Hilic(2.1×100 mm,1.7μm i.d) with gradient elution by CH3CN and 100 mmol/L of heptafluorobutyric acid at flow rate of 0.2 mL/min,the injection volume were 10μL.Multi-reaction monitoring(MRM) and external standard method were employed in quantitative analysis.4.Estabilshment of simultaneous detection method for multi-residues in aquatic products:Methodological parameters including standard curve,linearity range,limit of determination,recovery and precision were obtained.When spiked levels of 50,100 and 200 ng/g,the recoveries of ten aminoglycoside antibiotics in fish samples were in the range of 66.5%-108.0%,and relative standard deviations(RSD) were less than 18.8%, while the recoveries of aminoglycoside antibiotics in shrimp samples were in the range of 74.9%～99.1%,and RSDs were less than 12.4%.Moreover,each of aminoglycoside antibiotics was obtained by using this method,RSDs in intra-day detection were in the range of 2.6%～8.7%),while that in inter-day detection were in the range of 1.2%～8.4%for ten aminoglycoside antibiotics.In addition,the satisfactory linearity of calibration curves (50 ng/mL～1000 ng/mL) and limits of determination(20 ng/g～50 ng/g) were obtained.Compared with the currently used method,the established method in this thesis has some advantages of accuracy in qualitative and quantitative analysis,and strong anti-disturbance ability,and the sensitivities are below maximum residue limit of the international standards for simultaneous detection of 10 aminoglycoside antibiotics.This method is suitable for identification and quantification of aminoglycoside antibiotics in aquatic products.
|
PDF Full Text Request