Singh R J, Hogg N, Kalyanaraman B
Biophysics Research Institute, Medical College of Wisconsin, Milwaukee 53226-0509, USA.
Arch Biochem Biophys. 1995 Dec 20;324(2):367-73. doi: 10.1006/abbi.1995.0049.
The interaction of nitric oxide (.NO) with Rose Bengal (RB) in the presence of electron donors was investigated. Upon illumination of a mixture of RB and .NO with visible light, an enhancement in the rate of .NO consumption was observed that increased with increasing RB concentration. In the presence of electron donors (NADH, glutathione, or ascorbate), the rates of .NO depletion increased further. NADH enhanced .NO depletion to a greater extent than either glutathione or ascorbate. Photoactivated RB under anaerobic conditions reacts with NADH to form the RB anion radical (RB.-), which has a characteristic visible absorption band centered at 418 nm. Rose Bengal anion radical disporportionates to give RB and a colorless reduced form of RB, RBH-. The net result of this process is the photobleaching of RB. The presence of .NO during irradiation of RB and NADH introduced a lag time into the kinetics of RB photobleaching. The length of this lag time was proportional to the concentration of .NO. A similar lag time, which was also dependent on the .NO concentration, was observed in the kinetics of formation of RB.-. The three-line electron spin resonance (ESR) spectrum of RB.-, with an intensity ratio 1:2:1, was obtained during irradiation of RB and NADH under anaerobic conditions. .NO introduced a concentration-dependent lag time into the kinetics of the appearance of this ESR signal. We propose that .NO oxidizes RB.- to regenerate RB and thus inhibit photobleaching until .NO is consumed. This reaction predicts the formation of NO-, the one-electron reduced form of .NO. Nitrous oxide, a characteristic dimerization product of NO-, was detected by gas chromatography. This evidence indicates the occurrence of a Type I mechanism between photoactivated RB and .NO.
研究了一氧化氮（·NO）与孟加拉玫瑰红（RB）在电子供体存在下的相互作用。用可见光照射RB和·NO的混合物时，观察到·NO消耗速率增加，且随RB浓度增加而升高。在电子供体（NADH、谷胱甘肽或抗坏血酸）存在下，·NO消耗速率进一步增加。NADH比谷胱甘肽或抗坏血酸更能增强·NO的消耗。厌氧条件下光活化的RB与NADH反应形成RB阴离子自由基（RB·-），其具有以418nm为中心的特征可见吸收带。孟加拉玫瑰红阴离子自由基歧化生成RB和无色还原形式的RB，即RBH-。该过程的净结果是RB的光漂白。在RB和NADH照射期间·NO的存在给RB光漂白动力学引入了一个延迟时间。该延迟时间的长度与·NO浓度成正比。在RB·-形成动力学中也观察到类似的延迟时间，其也取决于·NO浓度。在厌氧条件下照射RB和NADH期间获得了强度比为1:2:1的RB·-的三线电子自旋共振（ESR）谱。·NO给该ESR信号出现的动力学引入了浓度依赖性的延迟时间。我们提出·NO将RB·-氧化以再生RB，从而抑制光漂白，直到·NO被消耗。该反应预测了NO-的形成，即·NO的单电子还原形式。通过气相色谱检测到一氧化二氮，它是NO-的特征二聚产物。该证据表明光活化的RB和·NO之间发生了I型机制。
