【作者】 马立军；

【导师】 吴玉清；

【作者基本信息】 吉林大学 ， 物理化学， 2008， 博士

【摘要】 重金属离子是一类重要的环境污染物,它们对人类的健康存在着巨大的威胁,因此对它们在生物、环境中的分析和检测成为生命科学、环境化学以及化学等多学科的重要课题。荧光离子探针方法不仅使用便捷,而且在灵敏度、选择性、时间分辨以及实时、原位检测等方面均具有突出优点。本论文利用常见的氨基酸作为金属离子的识别基团,设计、合成了几种重金属离子荧光探针,并在水溶液中测试了它们的识别特性。主要内容包括:（1）设计、合成了芘丁酰色氨酸,它以芘基团激基缔合物的特征发射峰为荧光响应信号,对Pb²⁺表现出很高的选择性和响应灵敏度（0.15μM）;（2）合成了含有双羧基氨基酸的荧光离子探针——芘丁酰谷氨酸和芘丁酰天冬氨酸,它们以芘基团基态二聚体（GSD）的特征发射峰和激发峰作为荧光响应信号,对铅离子表现出极高的选择性和较低的检测限（分别为1.5和1.6μM）;（3）合成了天冬氨酸修饰的荧光探针——丹酰天冬氨酸,该分子单独存在时对重金属离子无特异性识别。但当它与含有疏水空腔的牛血清白蛋白分子复合后,却对Hg²⁺表现出极高的选择性和响应灵敏度（检测限为0.5μM）。同时利用核磁共振波谱、拉曼和红外光谱以及理论计算等方法对这些探针分子和重金属离子的识别模式和作用机理进行了详细分析。这些研究对于水相中重金属离子的检测具有很好的应用前景,同时也为进一步理解金属离子-蛋白质分子之间的相互作用奠定了基础。更多还原

【Abstract】 Heavy metal ions （HMTs） are significant pollution for the environment due to their tremendous harms to human health. Since the analysis and detection of the heavy metal ions are important subjects for biology, environmental chemistry and chemistry etc. The designed Fluorometric sensors（Fls） based on the theory of fluorescent probe are not only easy to use but also can realize real space, real time, high sensitive and selective. And therefore, in recent years, the design and synthesis of Fls for HMTs are attracting more and more attentions. Because of the best water-solubility and the various possible interactions with HMTs, amino acid and peptide are general efficient recognizing groups for HMTs.Firstly, we designed and synthesized N-[4（1-pyrene）-butyroyl]-L-tryptophan （PLT） for Pb²⁺ by combining a pyrene-containing fluorophore, 4-（1-pyrenyl）-butyric acid, with a tryptophan （recognizing group）. PLT exhibited high sensitivity and selectivity for Pb²⁺ in solution with the responded signal of specific excimer of pyrene and very low detected limit （0.15μM）. NMR, Raman spectra, Infrared spectra and DFT calculation were performed to explore the coordination mechanism of PLT and Pb²⁺. The results show that the chelating bidentate coordination of PLT and Pb²⁺, an interaction between Pb²⁺ and the indole ring, and a unique hydrogen bonding between amide groups are crucial in the specific recognition of Pb²⁺. The process of recognition can be summarized as follows: the electrostatic interaction between one Pb²⁺ and two carboxylates in a chelating bidentate model induced a coordination between indole ring and Pb²⁺, the hydrogen bonding between amide groups as well; subsequently, a pyrene dimer is generated; thereby the specific excimer of PLT/Pb²⁺ is appeared. During the process the chelating bidentate model of Pb²⁺ and carboxylate is essential for Pb²⁺ recognition. It could induce the formation of hydrogen bonding between amide groups and coordination between indole ring and Pb²⁺, which are absolutely necessary for the formation of excimer.Based on the investigation of PLT, we have designed and synthesized another two new Fls, N-[4（1-pyrene）-butyroyl]-L-glutamic acid （PLE） and N-[4（1-pyrene）-butyroyl]-L-aspartic acid （PLD）, with double carboxyl groups which act as the Pb²⁺ recognition site and a pyrene moiety that serves as the photosignal transducer. PLE and PLD show high sensitivity （detection limit up to 1.5 and 1.6μM） and dual illustration of specific selectivity for Pb²⁺ over fourteen familiar metal ions in aqueous solution. The responded signal for Pb²⁺ comes from the specific emission and excitation of ground-state dimer （GSD） of pyrene. NMR, DFT and time-resolved emission decay experiments were performed to explore the coordination mechanism of PLE and Pb²⁺. These results show that during the interaction of Pb²⁺ and one carboxyl （near chiral carbon） of PLE, three species （monomer, excimer, and quenched species） coexist in aqueous solution. A certain degree of quenching of HMTs appeared because of the flexibility of PLE, which will be a guide for the construction of a more rigid moiety in future designs of Fls. In addition, literatures report when environment sensitive probe is coupled to protein, the conformational transforms of protein can induce the changes of fluorescent properties. If the probe is non-covalent located in the cavity of protein, the addition of Hg²⁺ should bring the structural changes of protein and/or participate in the interaction of probe with protein. So, the fluorescent properties of probe maybe change. Based on the concept and works above, we have designed and synthesized the third Fls, dansyl-L-aspartic acid （DLD）, with aspartic acid acting as the HMTs recognition site and a dansyl group serving as the photosignal transducer. Dansyl group is ubiquitous photophysical probes, which shows very high sensitivity in the interaction with cavity of protein. We select bovine serum albumin （BSA） by the way of its cavitation to DLD. The complex of DLD and BSA shows high sensitivity （detection limit up to 0.5μM） and specific selectivity for Hg²⁺ over fourteen familiar metal ions in aqueous solution with intensity enhancement and blue-shift （ΔλMAX = 35 nm） of fluorescence peak. NMR and else experiments show that the specific recognition of DLD/BSA for Hg²⁺ due to the dimethylamino group in DLD and the heteroatoms in cavites of BSA.In conclusion, we have successfully obtained three Fls of Pb²⁺ and one of Hg²⁺ by using amino acid as the recognition site of HMTs, which exhibit excellent performance of Fls in aqueous solution. Such results possess promising potential for the further development of Fls and better understanding of biological system.更多还原