选择性检测谷胱甘肽的荧光探针

doi:10.7517/j.issn.1674-0475.2017.04.015

摘要/Abstract

摘要：

本文设计合成了一种基于BODIPY衍生物选择性检测谷胱甘肽的比率式荧光探针1。荧光探针1中BODIPY的3位连有苯乙炔基团，5位连有咪唑盐离去基团，利用其与谷胱甘肽和半胱氨酸反应机理的不同实现了对谷胱甘肽的选择性检测。紫外可见吸收光谱和荧光光谱实验结果表明探针分子1与谷胱甘肽反应后的光谱发生明显红移，可以实现对谷胱甘肽的比率式检测。探针分子1对谷胱甘肽有极高的选择性，不受其它氨基酸尤其是半胱氨酸的干扰。荧光滴定实验表明探针分子1可实现对谷胱甘肽的定量检测，检测限为3.3×10^-8 mol/L。探针分子成功地应用于活体细胞中检测谷胱甘肽。

关键词: BODIPY, 荧光探针, 谷胱甘肽

Abstract:

We developed a ratiometric fluorescent probe based on BODIPY which can detect GSH selectively. We did UV-Vis and fluorescence experiments and found that the UV-Vis and fluorescence spectra of probe 1 with GSH exhibited obvious red shifts. Probe 1 exhibited high selectivity and the detection inhibited the interference of Cys and was not interfered by other amino acids. The fluorescence titration experiments showed that probe 1 realized the quantitative detection of GSH with detection limit of 3.3×10^-8 mol/L. The probe was successfully applied to the detection of GSH in living cells.

Key words: BODIPY, fluorescent probe, GSH

王媛, 陈潇潇, 刘学良, 陈玉哲, 牛丽亚, 吴骊珠, 杨清正. 选择性检测谷胱甘肽的荧光探针[J]. 影像科学与光化学, 2017, 35(4): 536-545.

WANG Yuan, CHEN Xiaoxiao, LIU Xueliang, CHEN Yuzhe, NIU Liya, WU Lizhu, YANG Qingzheng. A Fluorescent Probe for Selective Detection of GSH[J]. Imaging Science and Photochemistry, 2017, 35(4): 536-545.

参考文献

[1] Shahrokhian S. Lead phthalocyanine as a selective carrier for preparation of a cysteine-selective electrode[J]. Analytical Chemistry, 2001, 73: 5972-5978.
[2] Townsend D M, Tew K D, Tapiero H. The importance of glutathione in human disease[J]. Biomedicine & Pharmacotherapy, 2003, 57: 145-155.
[3] Lu S C. Regulation of glutathione synthesis[J]. Molecular Aspects of Medicine, 2009, 30: 42-59.
[4] Jung H, Chen X, Kim J, Yoon J. Recent progress in luminescent and colorimetric chemosensors for detection of thiols[J]. Chemical Society Reviews, 2013, 42: 6019-6031.
[5] Niu L Y, Chen Y Z, Zheng H R, Wu L Z, Tung C H, Yang Q Z. Design strategies of fluorescent probes for selective detection among biothiols[J]. Chemical Society Reviews, 2015, 44: 6143-6160.
[6] Hewage H S, Anslyn E V. Pattern-based recognition of thiols and metals using a single squaraine indicator[J]. Journal of the American Chemical Society, 2009, 131: 13099-13106.
[7] Yi L, Li H, Sun L, Liu L, Zhang C, Xi Z. A highly sensitive fluorescence probe for fast thiol-quantification assay of glutathione reductase[J]. Angewandte Chemie International Edition, 2009, 48: 4034-4037.
[8] Umezawa K, Yoshida M, Kamiya M, Yamasoba T, Urano Y. Rational design of reversible fluorescentprobes for live-cell imaging and quantification of fast glutathione dynamics[J]. Nature Chemistry, 2017, 9: 279-286.
[9] Pires M M, Chmielewski J. Fluorescence imaging of cellularglutathione using a latent rhodamine[J]. Organic Letters, 2008, 10: 837-840.
[10] Cao X, Lin W, Yu Q. A ratiometric fluorescent probe for thiols based on a tetrakis(4-hydroxyphenyl)porphyrin-coumarin scaffold[J]. The Journal of Organic Chemistry, 2011, 76: 7423-7430.
[11] Zheng Z, Chen P, Xie M, Wu C, Luo Y, Wang W, Jiang J, Liang G. Cell environment-differentiated self-assembly of nanofibers[J]. Journal of the American Chemical Society, 2016, 138: 11128-11131.
[12] Zhang J, Shibata A, Ito M, Shuto S, Ito Y, Mannervik B, Abe H, Morgenstern R. Synthesis and characterization of a series of highly fluorogenic substrates for glutathione transferases, a general strategy[J]. Journal of the American Chemical Society, 2011, 133: 14109-14119.
[13] Shao J, Sun H, Guo H, Ji S, Zhao J,Wu, W, Yuan X, Zhang C, James T D. A highly selective red-emitting FRET fluorescent molecular probe derived from BODIPY for the detection of cysteine and homocysteine: an experimental and theoretical study[J]. Chemical Science, 2012, 3: 1049-1061.
[14] Isik M, Guliyev R, Kolemen S, Altay Y, Senturk B, Tekinay T, Akkaya E U. Designing an intracellular fluorescent probe for glutathione: two modulation sites for selective signal transduction[J]. Organic Letters, 2014, 16: 3260-3263.
[15] He L, Xu Q, Liu Y, Wei H, Tang Y, Lin W. Coumarin-based turn-on fluorescence probe for specific detection of glutathione over cysteine and homocysteine[J]. ACS Applied Materials & Interfaces, 2015, 7: 12809-12813.
[16] Liu Y, Lv X, Liu J, Sun Y Q, Guo W. Construction of a selective fluorescent probe for GSH based on a chloro-functionalized coumarin-enone dye platform[J]. Chemistry A European Journal, 2015, 21: 4747-4754.
[17] Yang X, Guo Y, Strongin R M. Conjugate addition/cyclization sequence enables selective and simultaneous fluorescence detection of cysteine and homocysteine[J]. Angewandte Chemie International Edition, 2011, 50: 10690-10693.
[18] Gao X, Li X, Li L, Zhou J, Ma H. A simple fluorescent off-on probe for the discrimination of cysteine fromglutathione[J]. Chemical Communications, 2015, 51: 9388-9390.
[19] Miao Q, Li Q, Yuan Q, Li L, Hai Z, Liu S, Liang G. Discriminative fluorescence sensing of biothiols in vitro and in living cells[J]. Analytical Chemistry, 2015, 87: 3460-3466.
[20] Zhang J, Wang J, Liu J, Ning L, Zhu X, Yu B, Liu X, Yao X, Zhang H. Near-infrared and naked-eye fluorescence probe for direct and highly selective detection of cysteine and its application in living cells[J]. Analytical Chemistry, 2015, 87: 4856-4863.
[21] Niu W, Guo L, Li Y, Shuang S, Dong C, Wong M. Highly selective two-photon fluorescent probe for ratiometric sensing and imaging cysteine in mitochondria[J]. Analytical Chemistry, 2016, 88: 1908-1914.
[22] Yue Y, Huo F, Ning P, Zhang Y, Chao J, Meng X, Yin C. Dual-site fluorescent probe for visualizing the metabolism of Cys in living cells[J]. Journal of the American Chemical Society, 2017, 139: 3181-3185.
[23] Hakuna L, Escobedo J, Lowry M, Barve A, McCallum N, Strongin R M. A photochemical method for determining plasma homocysteine with limited sample processing[J]. Chemical Communications, 2014, 50: 3071-3073.
[24] Lee H, Choi Y, Kim S, Yoon T, Guo Z, Lee S, Swamy K M K, Kim G, Lee J, Shin I, Yoon J. Selective homocysteine turn-on fluorescent probes and their bioimaging applications[J]. Chemical Communications, 2014, 50: 6967-6969.
[25] Loudet A, Burgess K. BODIPY dyes and their derivatives: syntheses and spectroscopic properties[J]. Chemical Reviews, 2007, 107: 4891-4932.
[26] Boens N, Leen V, Dehaen W. Fluorescent indicators based on BODIPY[J]. Chemical Society Reviews, 2012, 41: 1130-1172.
[27] Kowada T, Maeda H, Kikuchi K. BODIPY-based probes for the fluorescence imaging of biomolecules in living cells[J]. Chemical Society Reviews, 2015, 44: 4953-4972.
[28] Niu L Y, Guan Y S, Chen Y Z, Wu L Z, Tung C H, Yang Q Z, BODIPY-based ratiometroc fluorescent sensor for highly selective detection of glutathione over cystein and homocystein[J]. Journal of the American Chemical Society, 2012, 134: 18928-18931.
[29] Niu L Y, Guan Y S, Chen Y Z, Wu L Z, Tung C H, Yang Q Z. A turn-on fluorescent sensor for the discriminationof cystein from homocystein and glutathione[J]. Chemical Communications, 2013, 49: 1294-1296.
[30] Yin J, Kwon Y,Kim D, Lee D, Kim G, Hu Y, Ryu G H, Yoon J. Cyanine-based fluorescent probe for highly selective detection of glutathione in cell cultures and live mouse tissues[J]. Journal of the American Chemical Society, 2014, 136: 5351-5358.
[31] Liu J, Sun Y Q, Huo Y, Zhang H, Wang L, Zhang P, Song D, Shi Y, Guo W. Simultaneous fluorescence sensing of Cys and GSH from different emission channels[J]. Journal of the American Chemical Society, 2014, 136: 574-577.
[32] Kim S Y, Hong K H, Kim D, Kwon H, Kim H J. Tunable heptamethine-azo dye conjugate as an NIR fluorescent probe for the selective detection of mitochondrial glutathione over cysteine and homocysteine[J]. Journal of the American Chemical Society, 2014, 136: 7018-7025.
[33] Wang X, Lv J, Yao X, Li Y, Huang F, Li M, Yang J, Ruan X, Tang B. Screening and investigation of a cyanine fluorescent probe for simultaneous sensing of glutathione and cysteine under single excitation[J]. Chemical Communications, 2014, 50: 15439-15442.
[34] Liu J, Sun Y Q, Zhang H, Huo Y, Shi Y, Guo W. Simultaneous fluorescent imaging of Cys/Hcy and GSH from different emission channels[J]. Chemical Science, 2014, 5: 3183-3188.
[35] Guan Y S, Niu L Y, Chen Y Z, Wu L Z, Tung C H, Yang Q Z. A near-infrared fluorescent sensor for selective detection of cysteine and its application in live cell imaging[J]. RSC Advances, 2014, 4: 8360-8364.
[36] Niu L Y, Zheng H R, Chen Y Z, Wu L Z, Tung C H, Yang Q Z. Fluorescent sensors for selective detection of thiols: expanding the intramolecular displacement based mechanism to new chromophores[J]. Analyst, 2014, 139: 1389-1395.
[37] Jia M Y, Niu L Y, Zhang Y, Yang Q Z, Tung C H, Guan Y F, Feng L. BODIPY-based fluorometric sensor for the simultaneous determination of Cys, Hcy and GSH in human serum[J]. ACS Applied Materials & Interfaces, 2015, 7: 5907-5914.
[38] Niu L Y, Yang Q Q, Zheng H R, Chen Y Z, Wu L Z, Tung C H, Yang Q Z. BODIPY-based fluorescent probe for thesimultaneous detection of glutathione and cysteine/homocysteine at different excitation wavelengths[J]. RSC Advances, 2015, 5: 3959-3964.
[39] Niu L Y, Jia M Y, Chen P Z, Chen Y Z, Zhang Y, Wu L Z, Duan X F, Tung C H, Guan Y F, Feng L, Yang Q Z. Colorimetric sensors with different reactivity for the quantitative determination of cysteine, homocysteine and glutathione in a mixture[J]. RSC Advances, 2015, 5: 13042-13045.
[40] Liu X L, Niu L Y, Chen Y Z, Zheng M L, Yang Y, Yang Q Z. A mitochondria-targeting fluorescent probe for the selective detection of glutathione in living cells[J]. Organic & Biomolecular Chemistry, 2017, 15: 1072-1075.
[41] Liu X L, Niu L Y, Chen Y Z, Yang Y, Yang Q Z. A multi-emissive fluorescent probe for the discrimination of glutathione and cysteine[J]. Biosensors and Bioelectronics, 2017, 90: 403-409.
[42] Rohand T, Qin W, Boens N, Dehaen W. Palladium-catalyzed coupling reactions for the functionalization of BODIPY dyes with fluorescence spanning the visible spectrum[J]. European Journal of Organic Chemistry, 2006, 20: 4658-4663.