Miller R W, Macdowall F D
Biochim Biophys Acta. 1975 Apr 14;387(1):176-87. doi: 10.1016/0005-2728(75)90062-6.
Wheat chloroplasts photochemically reduced molecular oxygen, as a Hill oxidant in the Mehler reaction, to superoxide anion which then oxidized added 1,2-dihydroxybenzene-3,5-disulfonate to its semiquinone, a comparatively stable free radical at pH 7. The last mentioned reaction was rapid in aqueous solution, but the rate of formation of 1,2-dihydroxybenzene-3,5-disulfonate semiquinone by the chloroplast system was calculated as T1 of 0.6 s. The Mehler reaction, or more specifically the univalent reduction of oxygen by Photosystem I, was rate-limiting so that the 1,2-dihydroxybenzene-3,5-disulfonate seniquinone was a useful spin probe for superoxide anion production at room temperature. The ESR signal of 1,2-dihydroxybenzene-3,5-disulfonate semiquinone was proportional to its steady state concentration and decayed in the dark with a T1/2 of 5-6 s. This oxygen-dependent signal was enhanced by mediation of chloroplastic oxygen reduction through methyl viologen. The superoxide anion scavengers ascorbate and L-epinephrine competitively obscured 1,2-dihydroxybenzene-3,5-disulfonate semiquinone formation, butadded superoxide dismutase was not as effective in this role. Partial inhibition by superoxide dismutase was achieved only by preincubation of Photosystem I enriched particles with ten times the endogenous concentration of superoxide dismutase. This and the persistence of a small amount of a 1,2-dihydroxybenzene-3,5-disulfonate (Tiron) oxidizing species in the dark supports the concept of Tiron accessibility but not the superoxide dismutase accessibility of superoxide anion bound in its formative enzyme complex. Benzoquinone and naphthoquinone disulfonate also reacted with superoxide anion, and supported both the Hill reaction and the Mehler reaction as final oxidants of both water and superoxide anion.
小麦叶绿体通过光化学作用将分子氧作为梅勒反应中的希尔氧化剂还原为超氧阴离子，超氧阴离子随后将添加的1,2 - 二羟基苯 - 3,5 - 二磺酸盐氧化为其半醌，在pH 7时这是一种相对稳定的自由基。上述最后一个反应在水溶液中很快，但叶绿体系统形成1,2 - 二羟基苯 - 3,5 - 二磺酸盐半醌的速率经计算为T1为0.6秒。梅勒反应，或者更具体地说是光系统I对氧的单价还原，是限速反应，因此1,2 - 二羟基苯 - 3,5 - 二磺酸盐半醌是室温下超氧阴离子产生的一种有用的自旋探针。1,2 - 二羟基苯 - 3,5 - 二磺酸盐半醌的电子自旋共振（ESR）信号与其稳态浓度成正比，在黑暗中以5 - 6秒的半衰期衰减。这种依赖氧的信号通过甲基紫精介导叶绿体氧还原而增强。超氧阴离子清除剂抗坏血酸和L - 肾上腺素竞争性地抑制1,2 - 二羟基苯 - 3,5 - 二磺酸盐半醌的形成，但添加的超氧化物歧化酶在这方面效果不佳。只有通过用比内源性超氧化物歧化酶浓度高十倍的超氧化物歧化酶预孵育富含光系统I的颗粒，才能实现超氧化物歧化酶的部分抑制。这以及在黑暗中少量的1,2 - 二羟基苯 - 3,5 - 二磺酸盐（钛铁试剂）氧化物种的持续存在支持了钛铁试剂可及性的概念，但不支持超氧化物歧化酶对结合在其形成酶复合物中的超氧阴离子的可及性概念。苯醌和萘醌二磺酸盐也与超氧阴离子反应，并支持希尔反应和梅勒反应作为水和超氧阴离子的最终氧化剂。
