Synthesising And Application Of Fluorescence Moleculer Probe For Proteins Containing Thiol Groups

Thiol protein is an important part of animal tissues and fluids, and concentration detection of thiol protein has important significance for organisms. The traditional method of detecting proteins containing thiol groups are iodometry, nitroprusside, spectrophotometry, colorimetry and amperometry, but these methods exist many defects, such as complex process, complicated operation, low sensitivity, so that the research and for testing proteins containing thiol groups has been greatly restricted.Porphyrins are provided with many characteristics,such as good optical properties, high fluorescent efficiency, relatively long excitation wavelength(about 400 nm) and long emission wavelength(about 660 nm) wavelength. Moreover, near infrared fluorescence(more than the 600 nm) light area, the fluorescence intensity of biological samples is very weak, which greatly reduces the interference of background fluorescence. In this paper, using these characteristics of porphyrin four enhanced fluorescence moleculer probes have been designed and synthesized, which are composed of porphyrin as a fluorophore and maleimide as identify groups, to detect quantitatively containing thiol-containing bovine serum albumin(BSA).Via the Adler method, four new porphyrin fluorescence probes were synthesized by 4-nitrobenzaldehyde, pyrrole, 4-methylbenzaldehyde, 4-methoxylbenzaldehyde, 4-hydroxybe-nzaldehyde, 4-carboxybenzaldehyde,acetic anhydride, maleic anhydride as a raw material, which were 5-(4-Maleimidophenyl)-10,15,20-tri-(4-methyphenyl) porphrin P1, 5-(4-Maleimidophenyl)-10,15,20-tri-(4-methoxyphenyl) porphrin P2, 5-(4-Maleimidophen-yl)-10,15,20-tri-(4-hydroxyphenyl) porphrin P3, 5-(4-Maleimidophenyl)-10,15,20-tri-(4-carboxyphenyl) porphrin P4.The structure of the four kinds of porphyrin compounds(P1, P2, P3, P4) were characterized by infraredspectroscopy(IR), UV-visible spectroscopy, 1H NMR spectroscopy, and elemental analysis. The detection effect of BSA was explored with the porphyrin compounds(P1, P2, P3, P4) as enhanced fluorescence probes. The optimum conditions of BSA quantitative detection using the above four fluorescent molecular probes and the detection impact of interfering substances was explored.Results show that these compound structures were the same with the expected. The linear range of the probes P1, P2, P3 for detection of BSA is all 0 mol/L~2.0×10-8 mol/L, then the linear equation, the correlation coefficient,and the detection limit are as follows respectively: P1:F=72.404+6.95[BSA](10-8mol/L), R=0.9876, 1.06 nmol/L; P2:F=26.367+12.861[BSA](10-8 mol/L), R=0.9927, 0.42 nmol/L; P3: F=32.734+15.178[BSA](10-8mol/L), R=0.9931, 0.39 nmol/L; P4: F=66.434+35.024[BSA](10-8 mol/L), R=0.9952, 0.28 nmol/L. The results showed that the method is feasible of fluorescent moleculer probes P2 、 P3、P4 for quantitative detecting thiol protein.