Prasad Ankush, Kumar Aditya, Matsuoka Ryo, Takahashi Akemi, Fujii Ryo, Sugiura Yamato, Kikuchi Hiroyuki, Aoyagi Shigeo, Aikawa Tatsuo, Kondo Takeshi, Yuasa Makoto, Pospíšil Pavel, Kasai Shigenobu
Department of Biophysics, Centre of the Region Haná for Biotechnological and Agricultural Research, Faculty of Science, Palacký University, Olomouc, Czech Republic.
Biomedical Engineering Research Center, Tohoku Institute of Technology, Sendai, Japan.
PeerJ. 2017 Jul 13;5:e3050. doi: 10.7717/peerj.3050. eCollection 2017.
The growth and development of plants is deleteriously affected by various biotic and abiotic stress factors. Wounding in plants is caused by exposure to environmental stress, mechanical stress, and via herbivory. Typically, oxidative burst in response to wounding is associated with the formation of reactive oxygen species, such as the superoxide anion radical (O), hydrogen peroxide (HO) and singlet oxygen; however, few experimental studies have provided direct evidence of their detection in plants. Detection of O formation in plant tissues have been performed using various techniques including electron paramagnetic resonance spin-trap spectroscopy, epinephrine-adrenochrome acceptor methods, staining with dyes such as tetrazolium dye and nitro blue tetrazolium (NBT); however, kinetic measurements have not been performed. In the current study, we provide evidence of O generation and its kinetics in the leaves of spinach () subjected to wounding.
Real-time monitoring of O generation was performed using catalytic amperometry. Changes in oxidation current for O was monitored using polymeric iron-porphyrin-based modified carbon electrodes ( = 1 mm) as working electrode with Ag/AgCl as the reference electrode.
The results obtained show continuous generation of O for minutes after wounding, followed by a decline. The exogenous addition of superoxide dismutase, which is known to dismutate O to HO, significantly suppressed the oxidation current.
Catalytic amperometric measurements were performed using polymeric iron-porphyrin based modified carbon electrode. We claim it to be a useful tool and a direct method for real-time monitoring and precise detection of O in biological samples, with the potential for wide application in plant research for specific and sensitive detection of O.
植物的生长和发育受到各种生物和非生物胁迫因素的有害影响。植物受到的创伤是由环境胁迫、机械胁迫以及食草动物造成的。通常，植物受伤后的氧化爆发与活性氧的形成有关，如超氧阴离子自由基（O）、过氧化氢（HO）和单线态氧；然而，很少有实验研究能提供在植物中检测到这些物质的直接证据。已经使用多种技术检测植物组织中O的形成，包括电子顺磁共振自旋捕获光谱法、肾上腺素 - 肾上腺色素受体法、用诸如四唑染料和硝基蓝四唑（NBT）等染料染色；然而，尚未进行动力学测量。在本研究中，我们提供了菠菜（）叶片受伤后O生成及其动力学的证据。
使用催化电流分析法对O的生成进行实时监测。以基于聚合铁卟啉的修饰碳电极（ = 1毫米）作为工作电极，Ag/AgCl作为参比电极，监测O氧化电流的变化。
所得结果表明，受伤后数分钟内O持续生成，随后下降。已知能将O歧化为HO的超氧化物歧化酶的外源添加显著抑制了氧化电流。
使用基于聚合铁卟啉的修饰碳电极进行催化电流测量。我们认为它是一种有用的工具和直接方法，可用于实时监测和精确检测生物样品中的O，在植物研究中具有广泛应用潜力，可用于特异性和灵敏地检测O。
