Preparation Of Ag@Au NPs Solid Phase Array Substrate And Its Application In SERS Detection Of Particular Antibiotics In Crayfish

The abuse of antibiotics in crayfish breeding,storage and transportation has seriously threatened food safety and human health,so it is urgent to establish a convenient,rapid,sensitive and specific detection method.Compared with traditional detection methods,such as chromatography and enzyme-linked immunoassay,SERS has attracted more and more attention because of its features,such as convenient,high sensitivity,"fingerprint"recognition and non-destructive detection.In this paper,with the specific antibiotic used in crayfish culture system as the target,SERS solid phase substrates based on Ag@Au NPs were developed to establish a series of SERS detection techniques with high sensitivity,good stability and excellent reproducibility.The specific detection was realized by combining with aptamer.The main research contents and conclusions are as follows:（1）The"core-shell"Ag@Au NPs was prepared by regulating Ag NPs by seed-mediated technique and blocking the galvanic reaction by ascorbic acid reduction.The results showed that under the conditions of p H≥10,Ag particle size 26.5 nm and Au shell thickness 6.8 nm,Ag@Au NPs had the strongest electromagnetic coupling effect,its EF was 5.7×10⁷ and storage period was 3 days.The RSD of signal intensity at different collection points was 4.91%.It has the same SERS enhancement performance and better chemical stability as Ag NPs.The detection limits of R6G were 1×10^-10 mol/L.The detection limits of ENR and AHD were 0.1ng/m L and 0.01 ng/m L,respectively.The recoveries were 90.50%-112.34%and 96.13%-104.93%,respectively.（2）In order to construct a stable SERS solid phase substrate,Ag@Au NPs was modified on glass substrate by interfacial silanization technology to obtain Ag@Au NSS.The EF was9.5×10⁶ and the storage period of 30 days.The RSD of different sites was 5.72%.The detection limit of ENR was 1 ng/m L and in the range of 5-10000 ng/m L,the intensity at 1382 cm^-1 showed a linear relationship with the logarithm of concentration.（3）Based on the solid phase substrate preparation strategy,Ag@Au NSA substrate with high EF（8.3×10⁷）and good reproducibility（3.12%RSD at different collection points）was successfully developed by means of microsphere etching and magnetron sputtering.SERS mapping imaging showed that micro-area enhancement with long-range effect was formed,resulting in a wider range of high-density"hot spots".Ag@Au NSA was applied to SERS detection of ENR,CIP,AHD,furaltadone metabolites and furazolidone metabolites in crayfish.The detection limits were 0.076 ng/m L,0.025 ng/m L,0.085 ng/m L,0.100 ng/m L and 0.050ng/m L,respectively.The recoveries in crayfish were 91.36%-112.60%,92.52-108.54%,94.00%-112.00%,92.00%-114.00%and 96.40%-110.00%,respectively.（4）In order to improve the specificity,the detection strategies of double-stranded aptamer modified Ag@Au NSA were constructed and applied to the detection of ENR and CIP.The detection limits of ENR and CIP were both 0.1 ng/m L,and the linear ranges were 7.5-100ng/m L and 5-40 ng/m L,respectively.The recoveries of ENR and CIP in crayfish were 96.00%-105.00%and 94.50%-104.00%,RSD_S were 3.81-5.19%and 4.11%-7.23%,respectively.The strategy of double-stranded aptamer modified Ag@Au NSA reduced the background interference in the detection of ENR and CIP in crayfish and could be directly applied to the detection of ENR and CIP in real samples.