Study On Magnetic Prussian Blue Catalyzed Signal Amplification Mediated Dual-readout Immunochromatography For Clenbuterol Hydrochloride And Ractopamine Detection

As a typicalβ-adrenoceptor agonist,Clenbuterol hydrochloride（CLE）remains in animal-derived foods.After being ingested by the human body through the food chain,it will cause adverse reactions and even induce malignant tumors.Strict limits have been established for CLE worldwide,but then Ractopamine（RAC）has gradually become a substitute for CLE due to its fast metabolism.However,in view of the potential hazards of RAC to human health,many countries,including China,have specified limit standards for RAC.However,due to the differences limit standards of RAC in different regions,trade disputes are likely to occur.Therefore,on the one hand,strengthening the detection ofβ-agonist residues in animal-derived foods,especially the development of rapid detection technology using immunochromatographic test strips as the detection platform is extremely important for food safety;on the other hand,it is also important to develop methods that can meet the needs of testing the same target under different limit standards for reducing trade disputes.This research is oriented to the hazards ofβ-receptor agonists and the reality that CLE and RAC have different limit standards in different countries and regions,a magnetic Prussian blue nanozyme（MPBN）catalyzed signal amplification mediated dual-readout immunochromatographic test strip forβ-adrenoceptor agonist detection was constructed and used for the immediate and rapid detection of CLE and RAC residues in animal-derived foods.The main research contents and results are as follows:1.Preparation and characterization proof of new signal label MPBN:The new signal tag MPBN with dual signal output capability was constructed by the post-synthesis strategy,Prussian blue nanoparticles（PBNPs）were compounded on the magnetic substrate Fe₃O₄ through the shell growth strategy.Scanning electron microscopy（SEM）and transmission electron microscopy（TEM）proved that MPBN has an approximately spherical core-shell structure with 250 nm.Ultraviolet-visible absorption spectroscopy（UV-vis）,Fourier transform infrared spectroscopy（FT-IR）,X-ray photoelectron spectroscopy（XPS）and hysteresis loop proved the successful preparation of the new signal label MPBN.At the same time,the peroxidase-mimicking catalytic activity of MPBN was verified by ultraviolet absorption peak at 652 nm and the color change of the corresponding solution,and it was confirmed that MPBN has an output colorimetric signal（derived from the color of MPBN as a signal label）and a catalytic signal（derived from MPBN’s analogue peroxidase catalytic activity）,to provide dual-signal capability,established a foundation for the construction of nanozyme catalyzed amplification mediated dual-readout immunochromatographic test strip.2.Construction and performance identification of MPBN catalyzed signal amplification mediated dual-readout immunochromatographic test strip:Under the strategy of parallel detection lines,a dual-readout immunochromatographic test strip was constructed by introducing MPBN as dual-function signal tag.Double signal and broadened detection range simultaneous detection of RAC and CLE were realized,the on-demand output mode of colorimetric signal and catalytic signal fulfill the detection requirements for different limits of the same target in various regions.Through the optimal selection for the key parameters that affect the detection performance,the visual detection limit（v LOD）based on the colorimetric signal for the detection of RAC and CLE were both 1.0 ng m L^-1,the cut-off values were 3.5 ng m L^-1and 6.0 ng m L^-1,respectively.The v LOD based on catalytic signals were both 2.0 ng m L^-1,and the cut-off values were 6.0 ng m L^-1 and 12 ng m L^-1,respectively,and the calculated limits of detection（c LOD）of 0.12 ng m L^-1 and 0.20 ng m L^-1 were obtained,respectively.Indicating that the proposed method possesses relatively sensitive performances,dual-signal output capability,low cost and wide detection range.3.The detection performance in actual sample detection based on the proposed method:Although the fat and other substances in actual samples interfered with the actual detection to a certain extent,the detection results similar to the theoretical values in the RAC and CLE spiked detection.At the same time,the good recovery ranged from84.01%to 119.94%and the coefficient of variation（CV）of 2.10%to 9.69%indicate the proposed method has good repeatability and reliability for the simultaneous detection of RAC and CLE in practical applications,and had a great application prospect in the monitoring of food hazard factors.