Determination Of α-cyano Pyrethroids Multi-residues By Immunoassay Methods

Immunologcal analysis technique is an important method for detection of the pesticide residue in the world. In this work, polycolonal antibodies anginst a family of theα-cyano Pyrethroids were prepared. The configuration analogyα-cyano-3-Phenoxybenzyl alcohol (CPBA) with pyrethroids was synthesized from the intermediate of 3-phenoxybenzaldehyde as the major materials. Then, theα-cyano-3-phenoxybenzyl alcohol butyric ester (CPBA-BE) was synthesized as the hapten conjugated with bovine serum albumin (BSA) as immunogen with NHS/DCC.the coating antigen conjugates were prepared by linking the CPBA-BE to big protein molecular of ovalbumin (OVA) with mixed anhydride method (MA). The coupling reaction was confirmed a success by UV-visible light spectrum scanning. The conjugation rates of immunogen and coating antigen was 17.9 and 13.2 respectively.Polyclonal antibodies PY-antibody were generated against the coupled CPBA-BE-BSA, and their antiserum gradient riched 102400, 25600, 25600, 6400 respectively. A general and broad class selective competitive enzyme-linked immunosorbent assay (ELISA) for the detection of theα-cyano Pyrethroids was developed. The assay was further optimized and tested for tolerance to co-solvent, pH and ionic strength changes. Under the optimized condition an assay curve Y was formed by the linking rate logit (B/B₀) and X was the logarithm of the Tau-Fluvalinate concentration (ng/mL) as the two influencing factors. In the range of 0.023-10ug/mL, the two factors represent a apparent linear relation, the epuation is as follows: Y = -0.2036X + 1.2051, R² = 0.9655.The cross-reaction rate to the other pyrethroids was tested, the polycolonal can recognize the metabolite PBA (IC₅₀= 4.898μg/mL) and cyhalothrin (IC₅₀=9.772ug/mL), but was no inhibition action to pyrethroids withoutα-cyano. The cross-reaction rates were 58.88% and 29.51% to PBA and cyhalothrin respectively. The limit of detection(LOD) of this assay was 0.023μg/mL for Tau-Fluvalinate.To estimate reliability, a recovery test was performed using a tap water sample. The average recoveries of spiked water were from 64.84% to 80.02%, the relative standard deviation of this method is within 2.20 %.