Study And Application Of The Analysis Method For Detecting Trace Mycotoxins In Food And Drug

Part 1:Determination of aflatoxins in potassium clavulanate by high performance liquid chromatography-laser-induced fluorescence detector and fluorescence detectorObjective:To establish a high performance liquid chromatography-laser-induced fluorescence determination method?HPLC-LIF?for detection aflatoxins in potassium clavulanate and compare with the traditional high-performance liquid chromatography-fluorescence deter-mination method?HPLC-FD?,providing a more sensitive method for the detection of residual aflatoxins in drug produced by fermentation and ensuring drug safety.Method:precisely 5 g sample was added 1 g sodium chloride and 25 mL methanol-water?50:50,v/v?,shook vigorously for 10 min,filtered and collected the filtrate.Precisely took 20 mL of filtrate in a 50 mL volumetric flask,diluted with water to volume.25 mL of diluted extract was cleaned-up through immunoaffinity column,20?L eluted sample was taken for analysis.Results:The detection limits of four aflatoxins?AFB₁,AFB₂,AFG₁ and AFG₂?by HPLC-FD method were 0.5,0.15,0.5 and 0.15 ng/mL,respectively.The detection limits of the four aflatoxins by HPLC-LIF method were 0.08,0.024,0.08 and 0.024 ng/mL,respectively.Conclusions:Without changing the sample pretreatment,the HPLC-LIF method improves the detection sensitivity of aflatoxins by one order of magnitude compared to the HPLC-FD method,providing a more sensitive and reliable means for ensuring drug safety.Part 2:Determination of aflatoxins in soy milk by graphene oxide coated stir bar sorptive extraction methodObjective:To establish a graphene oxide coated stir bar adsorption and extraction pretreatment method combined with high performance liquid chromatography-laser-induced fluorescence detector?HPLC-LIF?for the detection of aflatoxins in soy milk,providing a simple and sensitive method for the determination of aflatoxin in foods.Method:Firstly,the graphene oxide coated stir bar was placed in the soy milk sample solution,stirred and extracted for 40 min,then the graphene oxide coated stir bar was taken out and desorbed in 1.5 mL methanol solution for 10 min.The desorption solution was dried with nitrogen and reconstituted in 100?L mobile phase.20?L was injected into the HPLC-LIF system for analysis.Results:The detection limits of four aflatoxins?AFB₁,AFB₂,AFG₁ and AFG₂?were 2.4-8.0 pg/mL and the limits of quantification were 7.5-25 pg/mL.The recoveries were 80.5%-102.3%.Conclusions:The pre-treatment process requires only two steps,that is,the graphene oxide coated stir bar was placed in the sample solution to extract the target analytes,and then the stir bar was taken out and desorbed in methanol.The method is simple,sensitive and it is suitable for detection of trace aflatoxins in soy milk.PART 3:Determination of ochratoxin A in chicken liver by graphene oxide dispersed solid phase microextraction methodObjective:To develop a sample pretreatment technique based on graphene oxide dispersive solid-phase microextraction for purifying and enriching ochratoxin A in chicken liver samples,providing a simple,low-cost and sensitive pretreatmen method for the detection of ochratoxin A in liver samples.Method:The chicken liver samples were pretreated with appropriate samples preparation and vortexed with graphene oxide solution for 5 min.The graphene oxide was centrifuged and ochratoxin A was desorbed twice with 1mL of methanol.The desorption solution was dried with nitrogen and reconstituted in 100?L of mobile phase.20?L was injected into HPLC-FD analysis.Results:A good linearity was obtained for ochratoxin A in a concentration range of 0.05-2 ng/mL with an R² of 0.9962.The limit of detection was 0.04?g/kg,the recovery was 84.1%-103.9%,RSD was less than3.0%.Five batches of chicken liver samples were analyzed with this method.Two batches of ochratoxin A were found at levels of 1?g/kg and 4.3?g/kg,respectively.Conclusions:The method can effectively remove the interference of other impurities in the chicken liver sample,and has the advantages of simple operation,high recovery and good repeatability.It provides a method for the analysis of ochratoxin A in chicken liver samples and other biological substrates.