Zipfel E, Grezes J R, Seiffert W, Zimmermann H W
Histochemistry. 1982;75(4):539-55.
Analytically pure samples of the Romanowsky dyes eosin y, erythrosin b and tetrachlorofluorescein are prepared. DC of the dye samples shows no contaminations. We measured the absorption spectra of the dye dianions in alkaline aqueous solution and of the dye acids in 95% ethanol at very low dye concentrations. The molar extinction coefficients of the long wavelength absorption of the monomeric dye species are determined (Table 1). The extinction coefficients may be used for standardisation of dye samples. The absorption spectra of eosin y in aqueous solution are dependent on concentration. Using a new very sensitive method it was possible to identify two association equilibria from the concentration dependency of the spectra. Dimers are formed even in very dilute solutions, at higher concentrations tetramers. The dissociation constant of the dimers D in monomers M at 293 K, pH = 12, is K21 = 2,9 X 10(-5) M; of the tetramers Q in dimers D K42 = 2,4 X 10(-3) M. From the experimental spectra of eosin solutions at various concentrations, pH = 12, and the equilibrium constants K21, K42 the absorption spectra of the pure monomers, dimers and tetramers are calculated. M has one long wavelength absorption band, VM = 19300 cm-1, epsilon M = 1,03 X 10(5) M-1 cm-1; D also one absorption band, VD = 19300 cm-1, epsilon D = 1,74 X 10(5) M-1 cm-1; Q two absorption bands, VQ1 = 19100, VQ2 = 20200 cm-1, epsilon Q1 = 1,65 X 10(5), epsilon Q2 = 1,96 X 10(5) M-1 cm-1. The absorption spectrum of the dimers is discussed by quantum mechanics.
制备了分析纯的罗曼诺夫斯基染料伊红Y、赤藓红B和四氯荧光素样品。染料样品的薄层层析显示没有杂质。我们在极低染料浓度下测量了碱性水溶液中染料二价阴离子以及95%乙醇中染料酸的吸收光谱。测定了单体染料物种长波长吸收的摩尔消光系数（表1）。这些消光系数可用于染料样品的标准化。伊红Y在水溶液中的吸收光谱取决于浓度。使用一种新的非常灵敏的方法，从光谱的浓度依赖性中识别出两个缔合平衡。即使在非常稀的溶液中也会形成二聚体，在较高浓度下会形成四聚体。在293K、pH = 12时，二聚体D在单体M中的解离常数K21 = 2.9×10⁻⁵ M；四聚体Q在二聚体D中的解离常数K42 = 2.4×10⁻³ M。根据不同浓度、pH = 12的伊红溶液的实验光谱以及平衡常数K21、K42，计算出纯单体、二聚体和四聚体的吸收光谱。M有一个长波长吸收带，VM = 19300 cm⁻¹，εM = 1.03×10⁵ M⁻¹ cm⁻¹；D也有一个吸收带，VD = 19300 cm⁻¹，εD = 1.74×10⁵ M⁻¹ cm⁻¹；Q有两个吸收带，VQ1 = 19100，VQ2 = 20200 cm⁻¹，εQ1 = 1.65×10⁵，εQ2 = 1.96×10⁵ M⁻¹ cm⁻¹。用量子力学讨论了二聚体的吸收光谱。
