Sayo H, Saito M, Lee E, Kariya K
Chem Pharm Bull (Tokyo). 1989 Dec;37(12):3347-50. doi: 10.1248/cpb.37.3347.
Although in the absence of halide ion chloroperoxidase did not catalyze the ethylhydroperoxide (EHP)-supported oxidation of aminopyrine, in the presence of Br- or Cl-, chloroperoxidase did catalyze the oxidation of aminopyrine, generating the aminopyrine cation radical (AP+). The initial rate of AP+ formation was determined by monitoring the absorbance at 565 nm. The pH optimum of the reaction was centered around 5.0. The rate of AP+ formation showed typical Michaelis-Menten saturation kinetics with respect to EHP, aminopyrine and Br-. The rate of formation of bromine in the chloroperoxidase-EHP-Br- system was also determined by measuring the change in absorbance at 267 nm. In the system containing 1 mM EHP and 0.2 M KBr at pH 5.0, the rate was 1.8 nmol of bromine/s/micrograms of chloroperoxidase, which was slower than that of AP+ formation under the same conditions. The present results suggest that the formation of AP+ is initiated by the halogenation of the N,N-dimethylamino group followed by the homolysis of the haloammonium cation, and that the most likely halogenating reagent is an enzyme-bound halogenating intermediate.
虽然在没有卤离子的情况下，氯过氧化物酶不催化乙基过氧化氢（EHP）支持的氨基比林氧化反应，但在存在Br-或Cl-的情况下，氯过氧化物酶确实催化了氨基比林的氧化反应，生成了氨基比林阳离子自由基（AP+）。通过监测565 nm处的吸光度来测定AP+形成的初始速率。该反应的最适pH值集中在5.0左右。AP+形成的速率相对于EHP、氨基比林和Br-呈现出典型的米氏饱和动力学。氯过氧化物酶-EHP-Br-体系中溴的形成速率也通过测量267 nm处吸光度的变化来确定。在pH 5.0时含有1 mM EHP和0.2 M KBr的体系中，该速率为1.8 nmol溴/秒/微克氯过氧化物酶，这比相同条件下AP+形成的速率要慢。目前的结果表明，AP+的形成是由N，N-二甲基氨基的卤化作用引发，随后卤化铵阳离子发生均裂，并且最可能的卤化试剂是一种酶结合的卤化中间体。
