Syntheses And Characterization Of Aminofluorescein And Its Derivatives

Aminofluorescein is one of the most important fluorescent probes. The fluorescent intensity of aminofluorescein derivatives is enhanced after the amine group is bonded to carboxyl group of the biomolecules. So aminofluorescein has been widely applied in the protein analysis and immunology.In this paper, aminofluorescein was obtained from 4-nitrophthalic acid, resoreinol in the presence of zinc chloride through condensation and reduction, and 5-aminofluorescein and 6-aminofluorescein were successfully isolated by recrystallization of the obtained mixture of 5- and 6-aminofluorescein in 6% (w) hydrochloric acid. Compared with the traditional method, this synthetic route is more convenient and the total yield of 5-aminofluorescein was increased from 18% to 40%.The ability of aminofluorescein linking with carboxyl group was reduced due to the conjugation of amine group with aromatic ring. Aminofluorescein was water-soluble fluorescent probe, which restricted its application in biological analysis. In search of new better fluorescent probes, a series of derivatives of aminofluorescent were designed and synthesized such as fluorescein isothiocyanate, 2',7'-dichloro- fluorescein-5(6)-isothiocyanate, 5-(4,6-dichlorotriazin-2-yl)aminofluorescein, fluore- scein-5-chloroacetamide, 5-succinimidylfluorescein. The structures of the derivatives were confirmed by IR, MS and NMR, and their fluorescent properties were determined. The fluorescent intensity, maximum excitation wavenumber and maximum emission wavenumber of the derivatives were changed compared with aminofluorescein, which indicated better fluorescent properties.Besides, the compounds of 2,4,6-triarylpyrylium salts were sensitive fluorescent probes in the dyeing of inorganic ions and fluorescent analysis. Pyrylium perchlorates were prepared. The dyes with double fluorophore 1-(fluorescein-5-yl) pyridinium salts were yielded by the treatment of pyrylium salts with 5-aminofluorescein. 1-(Fluorescein-5-yl)-2,4,6-trimethylpyridinium perchlorate showed good fluorescent properties.