Study On The Determination Of Melamine,dicyclanil And Cyromazine And In Animal Tissues

A high performance liquid chromatography-tandem mass spectrometric detection（HPLC-MS/MS） method for melamine,dicyclanil,and cyromazine in milk was established.The protein in the milk sample was precipitated and the target analyte was extracted with a 1%trichloroacetic acid-acetonitrile solution.The target analyte was enriched and purified through a strong cation solid phase extraction column,and finally Waters UPLC-The tandem mass spectrometer was used for the determination of the residual contents of melamine,chlorhexazine and cyromazine.The chromatographic column was Acquity HSS T3（2.1 mm×50 mm,1.7μm）.Gradient washing was performed using 0.1%formic acid aqueous solution-acetonitrile as the mobile phase.The quantification was performed by external standard method.The pretreatment method and the liquid chromatography and mass spectrometry conditions for melamine,strychnil,and cyromazine were optimized.The results showed that the qualitative limits of melamine,dicyclanil,and cyromazine were 0.02,0.01 and 0.01mg/kg,respectively.The limits of quantification were 0.06,0.03 and 0.03 mg/kg,respectively.The recovery was between94%and 104%,which met the requirement.The method is sensitive and stable,and can qualitatively and quantitatively analyze melamine,chlorhexine and cyromazine in milk.In addition,an indirect enzyme-linked immunosorbent assay（ELISA）of dicyclanil in goat tissues and milk was established.The immunogen and the coating antigen were synthesized by cross-linking of the dicyclanil with the carrier protein using glutaraldehyde,and the polyclonal antibody was obtained after multiple immunizations of the rabbit.The antibodies were evaluated by indirect enzyme-linked immunosorbent assay and competitive enzyme-linked immunosorbent assay.The specificity of the antibody against dicyclanil was tested by cross-reaction with other compounds,and the conditions for sample extraction were optimized.The results indicate that an indirect competitive enzyme-linked immunosorbent assay for dicyclanil in sheep tissues and goat milk was established.The IC₅₀ for dicyclanil was 9 ng/mL.The recoveries were between 52.5%and 72.4%for sheep tissues and milk,the coefficient of variation was between 4.7%and 11.9%.The method is sensitive,specific,and easy operation.It can be used to simultaneously screen the residues sheep tissue and milk.The suspected positive samples can be further confirmed by high performance liquid chromatography tandem mass spectrometry.