Determination Of Sudan Ⅰ And Benzophenone By ELISA

Sudan red was used synthetic industry colorant, Benzophenone(BZP) used as photoinitiator in packaging, have verified the potential carcinogenesis. This thesis has developed a rapid enzyme-linked immunosorbent assay (ELISA) detection method to detect Sudan and Benzophenone.Two sudanI hapten derivatives were synthesized by diazotization, was characterized by Liquid Chromatography-Mass Spectrum. .One hapten was used as immunogen, while the other was applied as coating antigens, SUDAN-PAPA-CBSA immunize two New Zealand rabbits to obtain anti-serums, titres of antibodies were up to 64000. Ic- ELISA methods was established,and the coating buffer, blocking solution, sample solution, antibody solution, reaction time of antibody, reaction time of enzyme-secondary antibody were optimized Influence of ELISA. The regression equation of the final inhibition curve was y = -0.3508x + 0.6503,R²= 0.9989. The IC₅₀ of optimized indirect compete ELISA for sudanâ… were 3.0ng/mL , the linear range was 0.3～23.4ng/mL with a limit of detection (LOD) of 0.1ng/mL.The cross-reactivity towards para red was up to 140 %, sudanâ…¡,sudanâ…¢,sudan G were lower than 10 %, but towards sudanâ…£was lower than 1 %.Chili were fortified with sudanâ… by spiking two level concentrations (5 ng/g, 10 ng/g).The recoveries were 90.4%and 96.0%; the C.V were 1.8% and 4.9%.The hapten was 4-amino benzophenone conjugated with bovine serum albumin (BSA) as immunogen by diazotization, conjugated with ovalbumin (OVA) as coating antigens by glutaraldehyde. BZP-BSA immunize two New Zealand rabbits to obtain anti-serums, one anti-serum reached 56000,the other one was detected up to 128000. Indirect competitive ELISA methods for the detection of the BZP was developed.Optimization the parameters such as the coating buffer, blocking solution, sample solution, pH, ionic strength, organic solvents. The regression equation of the final inhibition curve was y = -0.3508x + 0.6503,R²= 0.9989. The sensitivity of ELISA showed that the IC50 was 23.7ng/mL,the LOD was 2ng/mL and the linear range was 3.2～177.6ng/ml. The cross-reactivity towards 4.4-bis N, N-dimethyl amino benzophenone, 2.4-Dihydroxy benzophenone were lower than 1 %. We established a rapid method was used for BZP detection in package paper samples and the recoveries were (92.8%, 94.5%) at 50 ng/g and 150 ng/g spiked level. The C.V were 2.7% and 4.5%.