Research On The Synthesis And Application Of A Long-wavelength Chemiluminescence Probe

Compared with fluorescence,chemiluminescence does not require an external excitation source,and has the advantages of no autofluorescence and light scattering background,no light stress on the detected object.Therefore,the chemiluminescence analysis method has high signal-to-noise ratio,high sensitivity,and is particularly suitable for life object applications.Although chemiluminescence has been applied in medicine,environment,food analysis,clinical diagnosis,and therapeutic effect monitoring,it is still less used than fluorescence analysis.There are several reasons for the above contrast.First,the type and quantity of chemiluminescent probes are seriously insufficient compared with fluorescence.Secondly,the existing chemiluminescent probes are inferior to fluorescence in object selectivity.Therefore,the development of new chemiluminescent probes with high selectivity and high quantum yield is the inevitable course to expand the application range of chemiluminescence analysis.Studies by Shabat et al.have shown that the nature of chemiluminescent probes depends on the structure of the illuminant,the chemiluminescence wavelength and quantum yield of the probe can be regulated by adjusting the structure of the illuminant in the probe,and the difference between the luminescent group and the substituent may affect the selectivity of the probe for the analytical object.However conventional luminophores often emit strongly as isolated molecules but meet with varying degrees of Aggregation-caused quenching(ACQ)effect when they are aggregated or clustered.From the viewpoint of real-world applications,it does harm than good in most cases.Thus,academician Tang Benzhong discovered and proposed the concept of Aggregation-induced emission(AIE).Fluorescence molecules with AIE characteristics are greatly enhanced when they are concentrated in aqueous solution.This excellent fluorescence property makes the AIE molecule a new choice for the design and synthesis of a new type of chemiluminescent probe.The long-term research goal based on the laboratory is to synthesize chemiluminescent probes suitable for 1O2 detection and in vivo imaging in life analysis,taking into account that longer wavelength probes have better penetrability on tissue,the background autofluorescence and light damage to tissues are relatively small,and has a higher signal-to-noise ratio in imaging applications.Combining the design idea of AIE material,we choose a red light-emitting aggregate structure connected with a chemiluminescent group,aiming to synthesize a long wavelength aggregation induced chemiluminescence probes,and study the aggregation-induced emission properties and biological analysis applications.The content of the paper consists of two parts:review and research report.The review section briefly describes the history of chemiluminescence and the luminescence behavior of common chemiluminescence systems,focusing on the current development of chemiluminescence probes.The research report in this paper is divided into the following two parts:1.Synthesis and Chemiluminescence Properties of TPE-N(Ph)-DBT-PH ProbesCombined with the synthesis of hydrazide chemiluminescent reagents in the laboratory and the synthetic route of aggregates in the literature,a red chemiluminescent probe(TPE-N(Ph)-DBT-PH)with aggregation-induced emission properties was designed and synthesized.The structure was characterized by nuclear magnetic resonance,mass spectrometry and infrared spectroscopy.The chemiluminescence properties of TPE-N(Ph)-DBT-PH were studied and the following results were obtained:(1)The emission wavelength of TPE-N(Ph)-DBT-PH in aqueous solution was 612 nm,and the absolute fluorescence quantum yield was 27.33%;(2)TPE-N(Ph)-DBT-PH has typical aggregation-induced emission(AIE)properties,and its fluorescence and chemiluminescence signals are greatly enhanced in the state of aggregation in aqueous solution;(3)According to the change of particle size and chemiluminescence intensity of aggregate nanoparticles with different F127 content,we get the optimum content of F127 is 2.5%;(4)TPE-N(Ph)-DBT-PH shows the strongest chemiluminescence signal at pH 7.4,it is most suitable for application under physiological conditions;(5)Under physiological conditions,TPE-N(Ph)-DBT-PH has good selectivity for singlet oxygen and is expected to be applied to analysis of 1O2 related biological processes;(6)The prepared TPE-N(Ph)-DBT-PH solution has good chemiluminescence stability,it does not need to be prepared while using,and is more convenient for experimental use.2.The application of TPE-N(Ph)-DBT-PH probe in bilogical analysis(1)Based on the better selectivity of probe 1O2,we studied the linearity of detection with 1O2 by TPE-N(Ph)-DBT-PH probe,and the detection limit was 23.5 nmol·L-1,indicating that the probe has high sensitivity to 1O2;(2)The cytotoxicity of TPE-N(Ph)-DBT-PH in Hela cells was evaluated by MTT assay,and the results showed that it was less toxic to living cells;(3)Fluorescence imaging of RAW 264.8 macrophages with probe confirmed that the probe has good cell permeability and can be used for intradlular 1O2 detection.(4)Detection of 1O2 in LPS and PMA-stimulated RAW 264.8 macrophages to evaluate cell stress levels under different conditions,and provides a basis for subsequent chemiluminescence imaging experiments.