高位阻卟啉类荧光探针的设计合成及其荧光性能的研究

发布时间：2019-05-19 13:22

【摘要】：卟啉是一种性能优良的近红外荧光发色团,以其为母体设计合成的荧光探针,在近红外光区进行检测时可以有效减小复杂生物样品中背景荧光的干扰,提高检测的准确性和分辨率。目前基于卟啉结构设计合成的荧光探针以阳离子探针居多,而中性分子及阴离子探针鲜少报道。中性分子和阴离子在探针识别过程中的相互作用较弱,且大多数的中性分子及阴离子探针是通过与分析物之间发生化学反应导致体系荧光强度变化来达到识别目的。因此,设计合成卟啉类中性分子和阴离子探针较阳离子探针更具有挑战性。同时,研究表明,简单地改变荧光团中取代基的位置可以很好地提高荧光探针分子的选择性。选择性的改变探针会更好地避免生物体及环境中干扰离子的影响,使探针对目标化合物的检测更为精准。本文以卟啉为母体,根据PET机理设计合成了高位阻可识别硫离子、苯硫酚及巯基氨基酸的荧光探针。因为这些探针受光激发后发生PET过程,并且拥有高的位阻,所以探针本身的空白荧光很弱。而当加入目标化合物后,分子内PET过程被抑制,体系荧光得到明显的提高。主要内容如下:首先,我们以不同取代位置的单羟基苯基卟啉为母体,2,4-二硝基苯磺酰氯为识别部分合成得到了两种硫离子荧光探针M1、M2,利用核磁、质谱等分析手段进行了结构表征。优化探针M1、M2的测试条件后,得知两个探针对硫离子均有较好的响应。但将两个探针的选择性比较之后发现:探针M1较M2更具有选择的专一性。经分析,探针M1的位阻较高,在一定程度上抑制了巯基化合物的进攻,而促使体积较小的硫离子进入发生亲核反应。因此,增大探针的空间位阻有利于探针选择性的提高。其中,探针M1的线性范围为3×10-6 mol/L~9×10-6 mol/L,检出限为40 nM,探针M2的检出限为62nM,线性范围为1×10-6 mol/L~8×10-6 mol/L。其次,基于上述结论,将2-单羟基卟啉和NBD-Cl为原料合成了一种高位阻的苯硫酚荧光探针M3。该探针在加入苯硫酚前后荧光发生变化,并且在1×10-6 mol/L~7×10-6mol/L的浓度范围内呈现良好的线性关系,检出限为54 nM。最后,我们在2-单氨基卟啉接入马来酰亚胺基团,得到了一种荧光增强型的可识别巯基氨基酸的探针M4。当加入巯基氨基酸(GSH、Cys、Hcy)后,与探针之间发生环合反应,抑制了PET过程,体系的荧光强度得到增强。同时,还检测了6种非巯基氨基酸,结果发现体系的荧光光谱并没有发生变化,表明探针M4可以用来区分巯基氨基酸和非巯基氨基酸。
[Abstract]:Porphyrin is a kind of near infrared fluorescence chromophore with excellent properties. The fluorescence probe designed and synthesized with it as the parent can effectively reduce the interference of background fluorescence in complex biological samples when it is detected in the near infrared region. Improve the accuracy and resolution of detection. At present, cationic probes are the most common fluorescent probes based on porphyrin structure design, while neutral molecules and anion probes are rarely reported. The interaction between neutral molecules and anions in the process of probe recognition is weak, and most neutral molecules and anion probes achieve the recognition purpose by chemical reaction with analytes to cause the change of fluorescence intensity of the system. Therefore, it is more challenging to design and synthesize porphyrin neutral molecules and anion probes than cationic probes. At the same time, it is shown that simply changing the position of the substituted group in the fluorescent cluster can improve the selectivity of the fluorescent probe molecule. Selective change of probe will better avoid the influence of interference ions in organism and environment, and make the detection of target compounds more accurate. In this paper, a fluorescence probe with high resistance to recognize sulfur ions, phlorophenol and sulfhydryl amino acids was designed and synthesized according to the mechanism of PET. Because the PET process occurs after these probes are excited by light and has high steric resistance, the blank fluorescence of the probes itself is very weak. When the target compound was added, the intramolecular PET process was suppressed and the fluorescence of the system was improved obviously. The main contents are as follows: first, we synthesized two kinds of sulfur ion fluorescence probes M1, M2by using monohydroxyphenyl porphyrins with different substitution positions as parent and 2, 4-dinitrobenzenesulfonyl chloride as recognition part. The structure was characterized by mass spectrometry and other analytical methods. After optimizing the test conditions of probe M _ 1 and M _ 2, it was found that the two probes had good responses to sulfur ions. However, after comparing the selectivity of the two probes, it is found that probe M1 is more selective and specific than M2. The results show that the steric hindrance of probe M1 is high, which suppresses the attack of sulfhydryl compounds to a certain extent, and causes the smaller sulfur ions to enter the nucleophilic reaction. Therefore, increasing the steric hindrance of the probe is beneficial to the improvement of the selectivity of the probe. The linear range of probe M1 is 3 脳 10 鈮，

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