Construction Of Ag₂S-based Nanozyme With Synergistic Photothermal/Imaging Process For Biosensing And Tumor Therapy

Enzymatic reaction has been widely applied in the fields of biosensing,imaging and therapy.However,the high price,complex preparation procedure,low stability and low cycle utilization of natural enzyme limit its wide application.Nanozyme,namely nanomaterial with biocatalytic activity that can mimic natural enzyme,has attracted increasing attention owing to advantages of simple preparation,high stability,easy purification,low cost and high recycling ability.In the complex biological tissue microenvironment,the catalytic efficiency of nanozyme is still insufficient and needs to be further improved.Particularly,facing the urgent demand of modern medicine for“all-in-one”diagnostic and therapeutical nanoplatform,the design and development of new multifunctional nanozyme also have important application and research value.In addition to their catalytic activity,the near-infrared(NIR)fluorescence property of Ag₂S quantum dots(QDs)has been widely studied in the field of biological imaging.It is feasible and meaningful to design multifunctional nanozyme with high catalytic efficiency based on the catalytic activity and optical properties of Ag₂S QDs.In this thesis,based on the enzyme-like activity of Ag₂S QDs,combined with their NIR fluorescence,photoacoustics and photothermal effects,a series of nanozyme samples with high catalytic efficiency and multifunction were designed.Moreover,the designed Ag₂S-based nanozyme was applied in the fields of visualized biosensing and tumor therapy.The main research contents were summarized as follows:(1)Ag₂S QDs with peroxidase-like activity and NIR fluorescence effect were synthesized in one step with glutathione(GSH)as surface ligands.At physiological p H and temperature,Ag₂S QDs catalyzed H₂O₂ to generate hydroxyl radical(·OH),and then the generated·OH oxidized colorless 3,3',5,5'-tetramethylbenzidine(TMB)to produce blue product(ox TMB).The catalytic activity of Ag₂S QDs remained above 80%after six cycles of testing.The peroxidase-like activity of Ag₂S QDs was increased to?2 times by laser irradiation.The Cu-GSH complex formed by the coordination of Cu(II)and GSH enhanced the catalytic activity of Ag₂S QDs,which made the above colorimetric reaction more obvious.The enhancement effect of Cu(II)on colorimetric reaction was further improved by laser irradiation.Additionally,the NIR fluorescence intensity of Ag₂S QDs significantly decreased with the increase of Cu(II)concentration.On the basis of above results,a colorimetric and NIR fluorescence dual-modal sensing method of Cu(II)was established,and the detection limit was 1.14?g/d L.Naked eye detection of Cu(II)in human serum was achieved by the established method.This study promoted the further application of nanozyme in the field of biosensing.(2)To improve the catalytic efficiency of Ag₂S nanozyme,Cu doping was used to regulate the enzyme-like activity.Ag_2-xCu_xS QDs with 1.0%Cu doping exhibited the best catalytic effect.At physiological p H,Ag_1.8Cu_0.2S QDs possessed tandem enzyme-like activity(i.e.peroxidase-like activity and glucose oxidase-like activity).Ag_1.8Cu_0.2S QDs oxidized glucose to produce H₂O₂,and then catalyzed the H₂O₂to generate·OH,which oxidized colorless TMB to blue ox TMB.Under the irradiation of 635 nm laser,the photothermal conversion efficiency of Ag_1.8Cu_0.2S QDs was 26%.The photothermal effect increased the catalytic activity of Ag_1.8Cu_0.2S QDs to?10 times,and made the colorimetric reaction more obvious.Based on the photothermal-enhanced tandem enzyme-like activity of Ag_1.8Cu_0.2S QDs,a one-step colorimetric glucose detection method with detection limit as low as 0.37 m M was constructed.Glucose in unprocessed human urine was accurately detected by the constructed method.These contributions would boost the development of one-step“nonenzyme”colorimetric glucose detection.(3)Nanozyme has a great potential in reactive oxygen species-mediated cancer therapy,however,high complexity of tumor microenvironment leads to decrease in catalytic efficiency of nanozyme,which makes therapeutic effect weaken.To overcome the obstacle,Cu-doped amorphous Ag₂S(a-Ag_2-xCu_xS)QDs were designed.Cu doping in amorphous Ag₂S QDs caused red-shift of NIRF to the lowest tissue absorption wavelength in so called“first biowindow”(?820 nm)and improved photothermal conversion efficiency from 11.9%to 44.0%simultaneously.Density functional theory simulation results revealed these fluorescence red-shift and enhanced photothermal/PA properties could be attributed to the generation of intragap states introduced by Cu doping.Cellular toxicity assay and serum chemistry analysis confirmed a-Ag_2-xCu_xS QDs possessed long-term high biocompatibility.Cell fluorescence results using 2',7'-dichlorodihydrofluorescein diacetate(DCFH-DA)as probe showed that a-Ag_2-xCu_xS QDs catalyzed endogenic H₂O₂ to produce highly toxic·OH under the photothermal effect,thus killing tumor cells.In addition,synergistic photothermal therapy could further improve the therapeutic effect.On the basis,a-Ag_2-xCu_xS QDs as“all-in-one”nanomedicine achieved efficient catalytic therapy and photothermal therapy guided by NIR fluorescence and photoacoustics dual-modal imaging.The tumor inhibition rate was>90%.This work provided an extensive paradigm to explore the treatment of cancer.