基于BODIPY的反应型荧光探针对生物硫醇和苯硫酚的选择性检测

发布时间：2019-03-09 09:44

【摘要】：含巯基化合物包括硫醇和苯硫酚,其在生命活动、工业生产以及自然环境中都扮演着十分重要的角色。生物硫醇主要有半胱氨酸Cys,同型半胱氨酸Hcy和谷胱甘肽GSH,在多个生理过程中起着重要作用,其浓度以及分布的异常与许多疾病有着紧密联系;而苯硫酚及其衍生物为有毒化合物,但是是重要的化工原料,工业生产中含苯硫酚的废水排入自然水体,势必会对水体和土壤造成污染,从而危害人体健康。因此,发展选择性检测生物硫醇和苯硫酚的荧光探针有着重要意义。本论文以光物理性能优异的BODIPY作为荧光团,结合巯基化合物的结构以及反应活性的差别,设计、合成了选择性检测GSH,Cys/Hcy以及苯硫酚的荧光探针,研究了其对含巯基化合物的识别检测性能,取得了如下研究成果:1.在BODIPY的3位引入了香豆素基团,合成了探针BC,用于半胱氨酸Cys和谷胱甘肽GSH的同时检测。生物硫醇的巯基通过亲核取代反应释放出蓝色荧光的香豆素作为内标分子,此外,GSH/Hcy生成巯基取代的黄色和红色BODIPY,而Cys的氨基可进一步经过五元环或六元环过渡态通过分子内重排生成氨基取代的蓝色BODIPY,从而体系呈现不同的发光颜色,实现了对GSH/Hcy和Cys的同时检测。特别是,在谷胱甘肽GSH浓度为0.4 mM时,溶液显示出白色的荧光。该探针成功用于活细胞内谷胱甘肽和半胱氨酸的同时成像检测。2.通过点击化学反应,在BODIPY的3位引入了具有线粒体定位功能的三苯基膦基团,合成了荧光探针BODIPY-PPh3,其能够准确定位于细胞线粒体,并且其与Cys/Hcy反应生成氨基取代产物,而与GSH反应生成巯基取代产物,不同产物的发光波长不同,从而实现了线粒体内GSH的选择性成像检测。3.通过调节BODIPY的3位取代基的推拉电子能力,合成并筛选了选择性检测苯硫酚的荧光探针Amide-BODIPY。利用苯硫酚和脂肪硫醇的反应活性不同,在生理条件下Amide-BODIPY能够与苯硫酚发生亲核取代反应,发射光谱红移,而脂肪硫醇则不发生反应,实现了对苯硫酚的选择性检测。该探针成功用于实际水样以及活细胞中苯硫酚的选择性检测。4.合成了一种新型的"O代F"成环BODIPY结构,通过用核磁、质谱和单晶衍射技术对目标产物进行了结构表征,测试数据表明该结构有着较长的吸收、发射波长,较高的量子产率和理想的荧光寿命,是一种潜在的有着良好光学性能荧光染料。
[Abstract]:Mercapto compounds, including mercaptan and phenol, play an important role in life activities, industrial production and natural environment. Biomercaptan mainly includes cysteine Cys, homocysteine Hcy and glutathione GSH, which play an important role in many physiological processes. The abnormal concentration and distribution of biomercaptan are closely related to many diseases. However, phenol and its derivatives are toxic compounds. However, the waste water containing phenol in industrial production is discharged into natural water, which is bound to cause pollution to water body and soil, thus endangering human health. Therefore, it is of great significance to develop fluorescent probes for selective detection of biomercaptan and phenylene mercaptan. In this paper, BODIPY, which has excellent photophysical properties, is used as fluorescent group. Combining with the difference of structure and reaction activity of sulfhydryl compounds, a fluorescent probe for selective detection of GSH,Cys/Hcy and phenol has been synthesized. The recognition and detection of sulfhydryl compounds were studied, and the following results were obtained: 1. The coumarin group was introduced into BODIPY and the probe BC, was synthesized for simultaneous detection of cysteine Cys and glutathione GSH. The sulfhydryl group of biomercaptan releases blue fluorescent coumarin as internal standard by nucleophilic substitution reaction. In addition, GSH/Hcy produces yellow and red sulfhydryl substituted BODIPY,. On the other hand, the amino groups of Cys can pass through five-or six-membered ring transition states to produce amino-substituted blue BODIPY, through intramolecular rearrangement, which results in different luminescence colors, and realizes simultaneous detection of GSH/Hcy and Cys. In particular, when the concentration of glutathione GSH was 0.4 mM, the solution showed white fluorescence. The probe was successfully used for simultaneous imaging detection of glutathione and cysteine in living cells. 2. By clicking chemical reaction, triphenylphosphine group with mitochondrial localization function was introduced in the third position of BODIPY, and fluorescence probe BODIPY-PPh3, was synthesized, which can accurately locate the cell mitochondria and react with Cys/Hcy to produce amino substituted products. The sulfhydryl substituted products were produced by the reaction with GSH, and the luminescence wavelengths of different products were different, thus the selective imaging detection of GSH in mitochondria was realized. 3. By adjusting the push-pull electron capacity of the three substituents of BODIPY, a fluorescent probe Amide-BODIPY. for selective detection of phenol was synthesized and screened. According to the different reactivity of thiophenol and fatty mercaptan, the nucleophilic substitution reaction between Amide-BODIPY and phenylene mercaptan can take place under physiological conditions, and the emission spectrum is red-shifted, but the fatty mercaptan does not react, so the selective detection of p-phenylene mercaptan is realized. The probe was successfully applied to the selective detection of phenol in real water samples and living cells. 4. A novel "O-substituted F" ring-forming BODIPY structure was synthesized and characterized by NMR, MS and single crystal diffraction. The results show that the structure has a long absorption and emission wavelength. High quantum yield and ideal fluorescence lifetime are potential fluorescent dyes with good optical properties.
【学位授予单位】：北京科技大学
【学位级别】：博士
【学位授予年份】：2017
【分类号】：O657.3

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