Preparation Of Copper Nanoclusters With Aggregation Induced Emission And Their Application In Hydrolase Activity Detection

With respect to organic dyes and semiconductor quantum dots,copper nanoclusters(CuNCs)have gradually become an active research area owing to their inherent merits,including desirable luminescence performance,much lower cost,good water solubility,good biocompatibility,low toxicity and wide availability,which enables them attractive candidates of promising nanomaterial suited for catalysis,biological imaging,chemical sensors and biomedical applications.Thiolcontaining small molecules are more suited for synthesis of CuNCs since they can function as both redutant and stabilizing agent and the synthesis process of CuNCs is usually quite simple without the requirement of expensive equipment and complicated operations compared to DNA oligonulceotides,proteins and peptides.It has been found that these CuNCs synthesized from small thiol-containing molecules possess a unique aggregation induced emission(AIE)property.CuNCs with this AIE property have been utilized for detection some chemical species of interest.However,the synthesis of CuNCs are still at an early stage in the application and analysis.In this dissertation,we proposed a facile one-step synthesis to prepare brightly luminescent CuNCs stabilized by glutathione and penicillamine.The synthesized CuNCs with inherent AIE property were employed as a luminescent sensing platform for the detection of ultralow level of water in organic solvents,real-time monitoring of acid phosphatase activity and β-galactosidase activity.The main contents are as follows:(1)Brightly luminescent CuNCs were prepared via a facile one-step synthesis in organic phase,and a novel luminescent nanoswitch on the basis of CuNCs through alternation of the physical states between aggregation and dispersion in response to specific external stimuli was designed.Two states including aggregation state and disaggregation state corresponding to luminescence on and off signaling can be readily switched in a reversible way based on the aggregation-induced emission and disaggregation induced quenching mechanism,respectively.This reversible nanoswitch can be controlled by the external stimulus water or N,N’-dicyclohexylcarbodiimide(DCC).The bright luminescence due to aggregation of CuNCs in organic solvents can be effectively quenched by the introduction of a small amount of water,where a disaggregation induced quenching takes place.This specific sensing behavior is capable to quantify an ultralow level(ppm)of water in aprotic solvents.The excellent reversibility of the nanoswitch enables one to monitor water content in a continuous and recyclable way.(2)A reversible luminescence nanoswitch through competitive hydrophobic interaction among CuNCs,p-nitrophenol and a-cyclodextrin is established,and a reliable real-time luminescent assay for acid phosphatase(ACP)activity is developed on the basis of this luminescence nanoswitch.Stable and intensely luminescent CuNCs were synthesized via a green one-pot approach.The hydrophobic nature of CuNCs aggregate surface is identified,and further used to drive the adsorption of p-nitrophenol on the surface of CuNCs aggregate due to their hydrophobic interaction.This close contact switches off the luminescence of CuNCs aggregate through static quenching mechanism.However,the introduction of a-cyclodextrin switches on the luminescence since stronger host-guest interaction between a-cyclodextrin and p-nitrophenol causes the removal of p-nitrophenol from the surface of CuNCs.This nanoswitch in response to external stimulus p-nitrophenol or a-cyclodextrin can be run in a reversible way.Luminescence quenching by p-nitrophenol is further utilized to develop ACP assay using p-nitrophenyl phosphate ester as the substrate.Quantitative measurement of ACP level with a low detection limit of 1.3 U/L was achieved based on this specific detection strategy.This work reports a luminescence nanoswitch mediated by hydrophobic interaction,and provides a sensitive detection method for ACP level which is capable for practical detection in human serum and seminal plasma.(3)A facile one-step synthesis of CuNCs AIE dots based on aluminum ions aggregation controlled by p-nitrophenol for the monitoring of Beta-galactosidase(β-Gal)activity was designed.The stable CuNCs AIE dots was prepared by introducing aluminum ions into the dispersed CuNCs in alkaline solution accompanying the luminescence switching on.p-Nitrophenol is utilized to quench the luminescent CuNCs AIE dots via the cooperative function by the static quenching and dynamic quenching mechanism through the formation of a non-luminescent complex between p-nitrophenol and aluminum ions of CuNCs aggregate with the signal readout of luminescence off.Luminescence quenching by p-nitrophenol is further utilized to develop P-Gal assay using p-nitrophenyl-β-D-galactopy-ranoside as the substrate.The correlation between quenched luminescence and P-Gal level is established to achieve sensitive measurement of β-Gal level.The detection limit can be lowered to be 0.7 U/L depending on the amount of the substrate.In summary,we successfully achieved a sensitive analysis for ultralow level of water in organic solvents by exploiting the AIE property of CuNCs in this research.Subsequently,a reversible luminescence nanoswitch through competitive hydrophobic interaction based on CuNCs is established as a reliable real-time luminescent assay for ACP activity which is the first time by utilizing thiolcontaining small molecules stabilized CuNCs for the detection of hydrolase.Ultimately,we developed a innovative assay based on CuNCs AIE dots by aluminum ions aggregation controlled by p-nitrophenol for the monitoring of β-Gal activity.This work demonstrates an example of AIE underlying detection strategy,broadens the application of copper nanoclusters in bioanalysis,and provides a reliable and sensitive assay for the detection of hydrolase.