Lohse J, Nielsen P E, Harrit N, Dahl O
Department of Chemistry, H. C. Orsted Institute, University of Copenhagen, Denmark.
Bioconjug Chem. 1997 Jul-Aug;8(4):503-9. doi: 10.1021/bc9700704.
Fluorescein ethyl ester, I, was used to prepare the fluorescent mixed ester/ether 6-O-(carboxymethyl)-fluorescein ethyl ester, III. Conjugation of III to the epsilon-amino group of alpha-N-Boc-L-lysine, via the N-hydroxysuccinimde ester, IV, gave the Boc-protected fluorescein-conjugated lysine monomer V. Removal of the Boc group, followed by reaction with Fmoc chloride, gave the Fmoc-protected monomer, VI (Figure 1). These Boc- and Fmoc-protected fluorescein-conjugated lysines were readily incorporated into peptides and PNA oligomers during solid phase synthesis to give fluorescent products. Mass spectroscopy and UV studies showed that the fluorophore remains unchanged during solid phase synthesis. In contrast to fluorescein, the photophysical properties of these derivatives are pH independent from pH 3 to 8, with a molar absorption coefficient, epsilon max 456, of 2.9 x 10(4) M-1 cm-1 and fluorescence quantum yield, phi f, of 0.18.
荧光素乙酯（I）用于制备荧光混合酯/醚6-O-（羧甲基）-荧光素乙酯（III）。III通过N-羟基琥珀酰亚胺酯（IV）与α-N-叔丁氧羰基-L-赖氨酸的ε-氨基偶联，得到叔丁氧羰基保护的荧光素偶联赖氨酸单体V。除去叔丁氧羰基后，再与芴甲氧羰基氯反应，得到芴甲氧羰基保护的单体VI（图1）。这些叔丁氧羰基和芴甲氧羰基保护的荧光素偶联赖氨酸在固相合成过程中很容易掺入肽和肽核酸低聚物中，得到荧光产物。质谱和紫外研究表明，荧光团在固相合成过程中保持不变。与荧光素不同，这些衍生物的光物理性质在pH 3至8范围内与pH无关，摩尔吸收系数εmax 456为2.9×10⁴ M⁻¹ cm⁻¹，荧光量子产率φf为0.18。
