多肽的合成和在金属离子检测中的应用
发布时间：2018-08-27 12:50
【摘要】：近年来,全球范围内重金属污染越来越严重,重金属离子污染已经引起了世界各国政府和环保组织的高度重视,可以说,重金属污染问题已经成为一个全球性环境问题。所以,对于生命科学、环境科学以及医学来说,发展一些简便、高效的检测重金属离子的分析方法是极其重要的。多肽作为小分子量的蛋白质,既有蛋白质的一般特性,又具有自身独特的生理功能,所以多肽的功能引起人们的广泛关注。多肽的研究已成为相关学科的研究热点之一,多肽更是凭借着本身的特性和与金属离子作用的优势成为检测金属离子的传感器中的佼佼者。本文基于金属离子与多肽的相互作用,设计、合成了基于多肽的化学探针,分别研究了它们与金属离子间的相互作用,探讨了它们在金属离子检测方面的应用。主要内容介绍如下:1.特异性检测Hg~(2+)的多肽比色探针采用Fomc固相合成法合成了比色传感器多肽Cys-Pro-Gly-Cys-Pro-Gly-Cys-CONH2(P7),应用HPLC进行了分离纯化,并利用质谱法进行了表征。运用紫外可见吸收光谱和圆二色谱等技术研究了该多肽P7与金属离子的相互作用。结果表明,该多肽P7对Hg~(2+)有很强的选择性,多肽中的半胱氨酸-SH与Hg~(2+)具有特异性的结合能力。把多肽P7加入到胶体纳米金溶液中时,纳米金的颜色由红色变为蓝色。当将P7和Hg(II)离子的混合溶液加入到胶体纳米金溶液中时,纳米金的颜色保持不变。但是,当将七肽和其他金属离子的混合溶液加入到胶体纳米金溶液中时,纳米金的颜色由红色变为蓝色。2.二茂铁-多肽Fc-CPPCG的合成及电化学的研究采用Fomc固相合成法合成了二茂铁-多肽Fc-CPPCG(Fc-P5)。用循环伏安法(CV)、线性扫描伏安法(LSV)和电化学阻抗法(EIS)对二茂铁-多肽Fc-P5修饰的玻碳电极进行了电化学表征,结果表明该修饰电极具有良好的电化学性能。研究了该二茂铁-多肽Fc-P5修饰的玻碳电极对金属离子的电化学响应,即它对金属离子的选择性作用。结果发现,Hg~(2+)使二茂铁多肽的峰电流和电化学阻抗都显著减小,但其他金属离子对二茂铁多肽的峰电流和电化学阻抗几乎都没有影响。峰电流和电化学阻抗的变化均表明,Fc-P5和Hg~(2+)发生了特异性结合。峰电流降低可能是由于Hg~(2+)与Fc-P5上Cys残基发生反应,使二茂铁基团的微环境发生改变,从而降低了二茂铁基周围的电子密度。电化学阻抗减小则是由于Hg~(2+)与Fc-P5的Cys发生配位反应,减少多肽对阻抗液中[Fe(CN)6]3-/4-分子在电极表面的电子传递的阻碍作用,促进了[Fe(CN)6]3-/4-分子与电极之间的电子传递。3.多肽Dansyl-HENRYKW的合成及其对Zn~(2+)的荧光传感检测合成了三个丹磺酰基修饰的多肽Dansyl-HERYKW(D-P6(1)),Dansyl-HENRKW(D-P6(2))和Dansyl-HENRYKW(D-P7)。通过荧光光谱分别研究了这三个丹磺酰基修饰的多肽与不同金属离子的相互作用。结果表明,Dansyl-HERYKW和Dansyl-HENRKW对金属离子没有选择性特异结合作用,唯有Dansyl-HENRYKW对Zn~(2+)具有特殊的选择性作用。Zn~(2+)的加入使多肽Dansyl-HENRYKW在515 nm处的荧光强度显著增强。研究了Dansyl-HENRYKW对Zn~(2+)的荧光传感性质和影响因素,多肽与Zn~(2+)的键合比为2:1,结合常数为1.8×1011L2×mol-2,检出限为0.012μmol×L-1。4.多肽CPPC和CPPCGG的合成及其耦合树脂对汞离子的吸附研究基于多肽与金属离子的反应性,设计、合成了多肽CPPC(P4)和CPPCGG(P6)及其耦合树脂(R-P4,R-P6)。运用紫外可见吸收光谱,研究了它们与金属离子的相互作用。结果表明,P4和P6对Hg~(2+)有很强的选择性作用。研究了多肽耦合树脂R-P4和R-P6对Hg~(2+)的吸附作用。原子荧光光谱法测得多肽耦合树脂R-P4和R-P6对Hg~(2+)的吸附容量分别为96 ng/g和60 ng/g,最佳吸附pH分别为3.0和5.0。
[Abstract]:In recent years, heavy metal pollution has become more and more serious worldwide. Heavy metal ion pollution has attracted the attention of governments and environmental protection organizations all over the world. It can be said that heavy metal pollution has become a global environmental problem. Detection of heavy metal ions is very important. As a small molecular weight protein, polypeptides not only have the general characteristics of proteins, but also have their own unique physiological functions, so the function of polypeptides has attracted widespread attention. In this paper, based on the interaction between metal ions and polypeptides, polypeptide-based chemical probes were designed and synthesized. Their interactions with metal ions were studied and their applications in the detection of metal ions were discussed. The main contents are as follows: 1. Polypeptide colorimetric probe for specific detection of Hg~ (2+) was synthesized by Fomc solid-phase synthesis. The polypeptide Cys-Pro-Gly-Cys-Pro-Gly-Cys-CONH2 (P7) was purified by HPLC and characterized by mass spectrometry. The polypeptide P7 was characterized by UV-Vis absorption spectroscopy and circular dichroism. The results showed that the polypeptide P7 had strong selectivity to Hg~ (2+) and the cysteine-SH in the polypeptide had specific binding ability to Hg~ (2+). When the polypeptide P7 was added to the colloidal nanogold solution, the colour of gold nanoparticles changed from red to blue. When the mixture of P7 and Hg (II) ions was added to the colloidal nanoparticles, the colour of gold nanoparticles changed from red to blue. However, when heptapeptide and other metal ions were mixed into colloidal gold nanoparticles, the color of gold nanoparticles changed from red to blue. 2. Synthesis and electrochemical study of ferrocene-polypeptide Fc-CPPCG Cyclic voltammetry (CV), linear sweep voltammetry (LSV) and electrochemical impedance spectroscopy (EIS) were used to characterize the ferrocene-polypeptide Fc-P5 modified glassy carbon electrode. The results showed that the modified electrode had good electrochemical performance. The electrochemical response of the ferrocene-polypeptide Fc-P5 modified glassy carbon electrode to metal ions was studied. It was found that Hg~ (2+) significantly reduced the peak current and electrochemical impedance of ferrocene polypeptide, but other metal ions had little effect on peak current and electrochemical impedance of ferrocene polypeptide. The decrease of peak current may be due to the reaction of Hg~ (2+) with Cys residue on Fc-P5, which changes the microenvironment of ferrocene group and reduces the electron density around ferrocene group. The hindrance of electron transfer promotes electron transfer between [Fe(CN)6]3-/4-molecule and electrode. 3. Synthesis of polypeptide Dansyl-HENRYKW and its fluorescence detection of Zn~ (2+) have synthesized three polypeptides, Dansyl-HERYKW (D-P6(1)), Dansyl-HENRKW (D-P6(2)) and Dansyl-HENRYKW (D-P7), respectively. The interaction of these three polypeptides with different metal ions was investigated. The results showed that Dansyl-HERYKW and Dansyl-HENRKW had no selective binding to metal ions, only Dansyl-HENRYKW had special selective effect on Zn~ (2+). The fluorescence intensity of Dansyl-HENRYKW at 515 nm was enhanced by the addition of Zn~ (2+). The fluorescence sensing properties and influencing factors of Dansyl-HENRYKW to Zn~ (2+) were studied. The binding ratio of polypeptide to Zn~ (2+) was 2:1, the binding constant was 1.8 Polypeptides CPPC (P4) and CPPCGG (P6) and their coupling resins (R-P4, R-P6) were synthesized. Their interaction with metal ions was studied by UV-Vis absorption spectroscopy. The results showed that P4 and P6 had strong selectivity for Hg ~ (2+). The adsorption of polypeptide coupling resins R-P4 and R-P6 on Hg ~ (2+) was studied. The adsorption capacities of peptide-coupled resins R-P4 and R-P6 for Hg~ (2+) were 96 ng/g and 60 ng/g, respectively. The optimum adsorption pH was 3.0 and 5.0, respectively.
【学位授予单位】：聊城大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：O629.72;O657.3
本文编号：2207337
[Abstract]:In recent years, heavy metal pollution has become more and more serious worldwide. Heavy metal ion pollution has attracted the attention of governments and environmental protection organizations all over the world. It can be said that heavy metal pollution has become a global environmental problem. Detection of heavy metal ions is very important. As a small molecular weight protein, polypeptides not only have the general characteristics of proteins, but also have their own unique physiological functions, so the function of polypeptides has attracted widespread attention. In this paper, based on the interaction between metal ions and polypeptides, polypeptide-based chemical probes were designed and synthesized. Their interactions with metal ions were studied and their applications in the detection of metal ions were discussed. The main contents are as follows: 1. Polypeptide colorimetric probe for specific detection of Hg~ (2+) was synthesized by Fomc solid-phase synthesis. The polypeptide Cys-Pro-Gly-Cys-Pro-Gly-Cys-CONH2 (P7) was purified by HPLC and characterized by mass spectrometry. The polypeptide P7 was characterized by UV-Vis absorption spectroscopy and circular dichroism. The results showed that the polypeptide P7 had strong selectivity to Hg~ (2+) and the cysteine-SH in the polypeptide had specific binding ability to Hg~ (2+). When the polypeptide P7 was added to the colloidal nanogold solution, the colour of gold nanoparticles changed from red to blue. When the mixture of P7 and Hg (II) ions was added to the colloidal nanoparticles, the colour of gold nanoparticles changed from red to blue. However, when heptapeptide and other metal ions were mixed into colloidal gold nanoparticles, the color of gold nanoparticles changed from red to blue. 2. Synthesis and electrochemical study of ferrocene-polypeptide Fc-CPPCG Cyclic voltammetry (CV), linear sweep voltammetry (LSV) and electrochemical impedance spectroscopy (EIS) were used to characterize the ferrocene-polypeptide Fc-P5 modified glassy carbon electrode. The results showed that the modified electrode had good electrochemical performance. The electrochemical response of the ferrocene-polypeptide Fc-P5 modified glassy carbon electrode to metal ions was studied. It was found that Hg~ (2+) significantly reduced the peak current and electrochemical impedance of ferrocene polypeptide, but other metal ions had little effect on peak current and electrochemical impedance of ferrocene polypeptide. The decrease of peak current may be due to the reaction of Hg~ (2+) with Cys residue on Fc-P5, which changes the microenvironment of ferrocene group and reduces the electron density around ferrocene group. The hindrance of electron transfer promotes electron transfer between [Fe(CN)6]3-/4-molecule and electrode. 3. Synthesis of polypeptide Dansyl-HENRYKW and its fluorescence detection of Zn~ (2+) have synthesized three polypeptides, Dansyl-HERYKW (D-P6(1)), Dansyl-HENRKW (D-P6(2)) and Dansyl-HENRYKW (D-P7), respectively. The interaction of these three polypeptides with different metal ions was investigated. The results showed that Dansyl-HERYKW and Dansyl-HENRKW had no selective binding to metal ions, only Dansyl-HENRYKW had special selective effect on Zn~ (2+). The fluorescence intensity of Dansyl-HENRYKW at 515 nm was enhanced by the addition of Zn~ (2+). The fluorescence sensing properties and influencing factors of Dansyl-HENRYKW to Zn~ (2+) were studied. The binding ratio of polypeptide to Zn~ (2+) was 2:1, the binding constant was 1.8 Polypeptides CPPC (P4) and CPPCGG (P6) and their coupling resins (R-P4, R-P6) were synthesized. Their interaction with metal ions was studied by UV-Vis absorption spectroscopy. The results showed that P4 and P6 had strong selectivity for Hg ~ (2+). The adsorption of polypeptide coupling resins R-P4 and R-P6 on Hg ~ (2+) was studied. The adsorption capacities of peptide-coupled resins R-P4 and R-P6 for Hg~ (2+) were 96 ng/g and 60 ng/g, respectively. The optimum adsorption pH was 3.0 and 5.0, respectively.
【学位授予单位】：聊城大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：O629.72;O657.3
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