Construction Of Novel SERS Nanoprobes Based On Gold Substrate For The Detection Of Hg²⁺ And Hydrogen Peroxide In Living Cells

Surface-enhanced Raman scattering(SERS)has become a powerful and sensitive analysis techniques in analytical,materials,and biomedicine due to its advantages of low autofluorescence,resistance to photobleaching,low phototoxicity,and narrow emission peaks.At present,use the SERS technology to construct new SERS nanoprobes with high-sensitivity and high-selectivity to detect the active substances in vitro,cells,microorganisms or in vivo became one of the main research hotspots in Raman analysis.In this paper,two substrates with high SERS activity gold nanospheres(AuNPs)and gold nanoflower substrates with core-molecule-shell structure were synthesized based on noble metal gold,as substrates for the construction of SERS nanoprobes,and their surfaces were decorated with the functionalized molecules CPBA and TP-Tyr as analyte recognition units and Raman reporter units,respectively.Two novel SERS probes,AuNPs@CPBA and Au@4-MBN@AuNFs@TP-Tyr,were constructed for the detection of Mercury ions(Hg2+)and hydrogen peroxide(H2O2)in vitro and in cells,respectively.The specific research contents are as follows:1.Ratiometric SERS nanoprobes based on gold nanospheres are used for the detection of intracellular Hg2+.In this chapter,60 nm gold nanoparticles were synthesized by seed growth method as SERS substrate,and functionalized CPBA molecules were synthesized by amidation reaction as Hg2+ recognition group and Raman reporter group.Then,the CPBA molecules were modified on the surface of gold spheres through Au-S bonds,and a ratiometric SERS probe AuNPs@CPBA that could specifically recognize Hg2+ was successfully constructed.When Hg2+ exists,Hg2+ will replace the boronic acid site of the CPBA molecule to form a new C-HgCl bond,causing the signal of B-O-H at 1002 cm-1 was weaken and the signal of C-HgCl at 1080 cm-1 was increased,thereby realized the ratiometric of Hg2+.According to the intensity ratio of I1080/I1303,this study calculated that the detection limit of this probe for Hg2+ was 0.12 μM.In addition,using this SERS probe,not only realized the detection of Hg2+ in four water samples,but also realized ratiometric Raman imaging of Hg2+ in living cells.The probe has strong specificity,high stability and good biocompatibility,which is of great significance for the detection of Hg2+in environmental water samples and biological samples.2.Interference-free SERS nanoprobes based on core-molecule-shell structures gold nanoflower for the detection of intracellular H2O2.In this chapter,core-moleculeshell structures gold nanoflowers embedded with 4-mercaptobenzonitrile(4-MBN)molecules were synthesized by seed growth method as highly active SERS substrates,and the functionalized TP-Tyr molecules were prepared by amidation reaction as H2O2 recognition group and Raman reporter group.A SERS probe Au@4MBN@AuNFs@TP-Tyr that can specifically recognize H2O2 was successfully constructed by modifying TP-Tyr on the surface of gold nanoflowers.In the coexistence of horseradish peroxidase(HRP)and H2O2,HRP can catalyze the H2O2,which induces the cross-linking reaction of the phenol site in the TP-Tyr molecule,promoting Au@4-MBN@AuNFs@TP-Tyr nanoprobe reunion.The nitrile signal(2223cm-1)of the 4-MBN molecule located in the cell silencing region is enhanced to achieve high selectivity and high sensitivity detection of H2O2.Based on the linear relationship between the signal intensity of nitrile group and the concentration of H2O2,the detection limit of this probe for H2O2 was calculated to be 0.3 μM.The SERS nanoprobe has the advantages of accuracy and reliability,no interference,high stability and strong specificity,and successfully achieved Raman imaging of H2O2 in A549 cells.