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【答案】应助回帖
铁（Fe）是人体中含量丰富的金属元素，在生物学和病理学方面发挥着重要作用。铁元素在生物体中主要以亚铁（Fe2+）和铁（Fe3+）离子的形式存在，由于生物体内的Fe3+容易被还原成Fe2+及在偏碱性的生理条件下溶解性不好；另一方面，生物体内的Fe2+会催化芬顿反应（Fenton reaction）产生自由基，可能引发一系列疾病。因此，设计一个快速、准确检测生物体内亚铁离子的探针具有十分重要的意义。
过氧亚硝基阴离子（Peroxynitrite anion, ONOO-）作为生物体内活性氮（Reactive nitrogen species, RNS）分子之一，它是由超氧自由基（O2•-）和一氧化氮（NO）反应生成。ONOO-作为很强的生物氧化剂，具有不稳定性和较高的反应活性，是一些生物体内循环途径的信号传导分子。但这种传导分子也可能对一些生物靶细胞造成不可逆转的损害，比如引发癌症、炎症和心血管疾病等多种疾病。因此，设计一个合适的探针对ONOO-检测显得至关重要。
（一）设计合成探针K1检测Fe2+。在苯并吡喃腈衍生物中引入氮氧化物（N-Oxide）合成荧光探针K1，探究探针K1与Fe2+发生反应可能的机理，并对其结构进行表征。本文通过光物理性质研究发现，该探针在PBS/DMF=1:9(0.01 M, pH 7.4)缓冲溶液中几乎没有荧光，加入Fe2+后，N-Oxide被还原，荧光恢复，在波长685 nm处有最大发射。该探针具有选择性好和灵敏度高（检测极限为2.96×10-6 mM）等特点，被应用于秀丽隐杆线虫和活体细胞中的Fe2+荧光成像。
（二）设计合成探针K2检测ONOO-。以苯并吡喃腈和香豆素合成荧光探针K2，探究探针K2与ONOO-发生反应可能的机理，并对其结构进行表征。在HEPES/DMF=1:9 (0.01 M, pH 7.4)缓冲体系下进行光物理性质测试，当加入饱和当量ONOO-时，该探针荧光光谱发生明显蓝移，并且能够实现对ONOO-的专一性识别，检测极限低至6×10-7 mM。更重要的是，该探针被成功应用于活体细胞中的ONOO- 荧光成像。
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关键词：亚铁离子；过氧亚硝基阴离子；苯并吡喃腈；荧光探针；荧光成像
Iron (Fe) is rich in human body, existing mainly as ferrous iron (Fe2+) and iron (Fe3+) ions , and plays an important role in biology and pathology.
It is very important to design a probe to detect (quickly and accurately) ferrous ions in living organisms, as Fe3+ soluble poorly under alkaline physiological conditions and tending to reduced to Fe2+,
and Fe2+( in the body) would catalyze the Fenton reaction generating free radicals which may cause a series of diseases,
It is critical to design a suitable probe for ONOO-detection, because Peroxynitrite anion (ONOO-), being one of the reactive nitrogen species (RNS) molecules and a strong biological oxidant (with high instability and reactivity) in the body, may damage (some) biological target cells and cause cancer, inflammation and cardiovascular diseases.
(1) Designing a synthetic probe K1 to detect Fe2+.
The fluorescent probe K1 was synthesized by introducing nitrogen oxide (N-Oxide) into the benzopyranonitrile derivative, its structure was characterized and the possible mechanism of the reaction of probe K1 and Fe2+ was investigated.
The probe had almost no fluorescence in the buffer solution of PBS/DMF=1:9 (0.01 M, pH 7.4), but N-Oxide was reduced and the fluorescence recovered nm by adding Fe2+, with a maximum emission at 685.
The probe was applied to Fe2+ fluorescence imaging in C. elegans and living cells and showed high selectivity and good sensitivity (detection limit 2.96×10-6 mM).
(B) Designing synthetic probe K2 to detect ONOO-.
The fluorescent probe K2 was synthesized with benzopyranonitrile and coumarin, its structure was characterized , and the possible mechanism of the reaction of probe K2 and ONOO- was investigated.
The fluorescence spectrum of the probe was significantly blue-shifted when saturated equivalent ONOO- was added, in HEPES/DMF=1:9 (0.01 M, pH 7.4) buffer, and the ONOO-specialization was realized, with a detection limit as low as 6 × 10-7 mM.
More importantly, the probe was successfully applied to ONOO-fluorescence imaging in living cells.
Key words: ferrous ion; peroxynitrite; benzopyranonitrile; fluorescent probe; fluorescence imaging |
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