Study On The Interaction Between Series Of 1,2,3-Triazoles And Blood Proteins And Chromogenic Mechanism Of Their Rhodamine B Derivatives To Hg²⁺ Ions And Application In Cell Imaging

1,2,3-triazole compounds have distinctive physical and chemical properties due to their unique nitrogen heterocyclic structure,such as good biological activity and low toxicity.They have been widely used in many fields including organic chemistry,organic metal chemistry,material chemistry and pharmaceutical chemistry.Especially the application of these compounds in medicine is particularly prominent.Some of these compounds have been used in pathogens,inflammation,cancer and so on,which showed good biological activity in the course of treatment.As for the newly synthesized 1,2,3-triazole compounds,it is necessary to study their interaction with blood proteins in view of the potential value of their development and utilization as novel drugs.It plays a vital role in vivo through binding with most drug molecules for the blood protein that is one of the important research objects of life science as a very important carrier of small drug molecules.The interaction between blood protein and drug not only affects drug distribution,but also affects drug metabolism and excretion in vivo,which is also the main factor determining whether drug has good pharmacological activity,bioavailability,low toxicity and metabolic stability.Therefore,the study of the interaction between drugs and blood proteins has been one of the important research topics in the fields of life science,chemistry and clinical medicine.Among of various blood proteins,serum albumin,smmunoglobulin and semoglobin are three kinds of representative globulin models in common use.On the other hand,1,2,3-triazole compounds can be use to synthesize many kinds of derivatives with other substituents.In particular,triazole derivatives containing ester or carboxyl groups in structure can specifically bond with some metal ions and be used as probes to detecting metal ions,which has shown its unique pharmacological activity and broad application prospects in environmental science and medicine now.As we all know,mercury is a highly toxic metal pollutant.The intake of mercury by organisms can lead to normal physiological disorders,which can cause many diseases such as kidney,digestive tract,brain or nervous system.Mercury can accumulate rapidly through aquatic biological chains after sinking into water,causing great pollution to the environment.Therefore,it is of great theoretical and practical significance for the environment and life safety to study the characteristic color recognition of mercury ions by newly synthesized derivatives of 1,2,3-triazole compounds.It is importantly helpful for to establish a method for the determination of mercury ions under physiological conditions,to determine the interaction mechanism between these triazoles and mercury ions,and to be used for detection to Hg²⁺in cell.Based on the previous research in our laboratory,eight different 1,2,3-triazole compounds and their corresponding rhodamine derivatives were studied in this paper,including ethyl 2,5-diphenyl-2H-1,2,3-triazole-4-carboxylate（S1）,ethyl 5-phenyl-2-（3-（trifluoromethyl）phenyl）-2H-1,2,3-triazole-4-carboxylate（S2）,ethyl 5-phenyl-2-（o-tolyl）-2H-1,2,3-triazole-4-carboxylate（S3）,ethyl 2-（4-methoxyphenyl）-5-phenyl-2H-1,2,3-triazole-4-carboxylate（S4）,ethyl 2-phenyl-5-（trifluoromethyl）-2H-1,2,3-triazole-4-carboxylate（S5）,ethyl 5-phenyl-2-（p-tolyl）-2H-1,2,3-triazole-4-carboxylate（S6）,5-methyl-2-phenyl-2H-1,2,3-triazole-4-carboxylic acid（S7）,5-methyl-1-phenyl-1H-1,2,3-triazole-4-carboxylic acid（S8）.Its corresponding rhodamine derivatives are abbreviated as R1,R2,R3,R4,R5,R6,R7,R8,respectively.Combining various spectroscopic techniques and computer chemistry methods,the following innovative studies have been carried out:（1）Eight 1,2,3-triazole compounds（S1-S8）were optimized by using the GaussView5.0program at HF Restricted algorithm level and Dependent Density Functional Theory（DFT）for the first time.Their absorption spectra and emission spectra were simulated by TD-DFT and compared with the experimental results.（2）The interactions of 1,2,3-triazole compounds（S2,S4,S5,S7）with three blood model globular proteins（human serum albumin（HSA）,human immunoglobulin（HIgG）and bovine hemoglobin（BHb））were studied by UV-visible spectroscopy,uluorescence spectroscopy and molecular docking techniques.（3）For the above rhodamine derivatives（R1-R8）based on 1,2,3-triazoles,their complexation structures with Hg²⁺were calculated by B3LYP method of DFT,and their chromogenic mechanism for Hg²⁺was determined by electrospray ionization mass spectrometry.（4）The measurements on cell imaging indicated that eight rhodamine derivatives（R1-R8）could be used to label human cervical cancer cell line HeLa under physiological conditions so as to discover the properties and uses of these new compounds.The main contents of this thesis are divided into two parts and nine chapters.The first part:Theoretical calculation of series 1,2,3-triazole compounds and their interactions with three blood globular proteins.In the first chapter,the 1,2,3-triazole compounds and their research status were reviewed.In the second chapter,the structures of eight 1,2,3-triazole compounds（S1-S8）were studied by computational chemistry.The results showed that the simulation results of HF and DFT methods are similar.But the compounds calculated by B3LYP method are closer to stable configurations.Meanwhile,the simulated spectra are basically consistent with the experimental spectra.In the third chapter to sixth chapter,the interactions of four 1,2,3-triazole compounds（S2,S4,S5,S7）with three blood model globular proteins（HSA,HIgG and BHb）were studied by using UV-vis,various fluorescencespectroscopiesandmoleculardockingtechnique.Thefluorescence spectroscopies showed that the four compounds enhanced the fluorescence of HSA/HIgG/BHb.The results of UV-vis,synchronous and three-dimensional（3D）fluorescence spectra revealed that the presentence of different 1,2,3-triazole compounds caused the changes of the conformation and microenvironment of proteins.The results of thermodynamic parameters and molecular docking demonstrated that the interaction forces of S5 and S7 with the three proteins were hydrophobic and hydrogen bonding forces.While it is different for the binding forces of S2 or S4 binding to the three proteins respectively,which there are also electrostatic and van der Waals forces.The second part:The chromogenic mechanism of their rhodamine derivatives（R1-R8）based on 1,2,3-triazoles to Hg²⁺and application in cell imaging.In seventh chapter,the studies on chromogenic rhodamine derivatives to Hg²⁺and cell imaging were reviewed.In the eighth chapter,the complex structures of eight rhodamine derivatives（R1-R8）were calculated by B3LYP method.The mass spectra of different complexing systems were determined by electrospray ionization mass spectrometry（ESI-MS）,and their chromogenic mechanism for Hg²⁺were further determined.The results of B3LYP calculation showed that the low-energy configurations of R1,R2,R3,R7 and R8 complexed with two Hg²⁺respectively were more stable.They exhibited more stable when the low energy configurations of complexes were performed by R4 and R6 complexed with one Hg²⁺,respectively.Their complexation ratios of R1-R8 and with Hg²⁺were 1:2 or 1:1 that were also confirmed by the corresponding electrospray ionization mass spectrometry.Simulated configuration of R5 with Hg²⁺was not obtained.While mass spectra results revealed that R5could also stably complex with two Hg²⁺.The chromogenic mechanism is predicted that Hg²⁺induces the ring opening of rhodamine spirals in rhodamine derivatives based on 1,2,3-triazoles.Finally,the results of cell imaging showed that eight rhodamine derivatives（R1-R8）based on 1,2,3-triazoles could be used to label human cervical cancer cell line HeLa under physiological conditions.In ninth chapter,there are the summary and outlook.It is summarized about the interaction of a series of 1,2,3-triazole compounds with blood proteins and the chromogenic mechanism and cell imaging studies of rhodamine derivatives on Hg²⁺.In view of the above results,the existing problems and possible further research ideas were put forward.Meanwhile,it was prospected on the further development and utilization of these series of 1,2,3-triazole compounds.