Dorta E, Aspée A, Pino E, González L, Lissi E, López-Alarcón C
Departamento de Química Física, Pontificia Universidad Católica de Chile, C.P. 782 0436, Santiago, Chile.
Facultad de Química y Biología, Universidad de Santiago de Chile, Casilla 40, Correo 33, Santiago, Chile.
Biomed Pharmacother. 2017 Jun;90:332-338. doi: 10.1016/j.biopha.2017.03.082. Epub 2017 Apr 2.
3-Hydroxy-anthranilic acid (3-OHAA), a tryptophan metabolite produced in the kynurenine pathway, is an efficient antioxidant towards peroxyl radicals (ROO) derived from the AAPH (2,2'-azobis(2-amidinopropane) dihydrochloride) thermolysis. However, self-reactions of ROO can give rise to alkoxyl radicals (RO), which could strongly affect the fate of scavenging reactions. In the present work, we studied the influence of RO in the scavenging activity of 3-OHAA in three different systems: i) Monitoring of the direct reaction between 3-OHAA and AAPH-derived free radicals (kinetic studies); ii) Evaluation of the protective effect of 3-OHAA on the AAPH-induced consumption of fluorescein; and, iii) Inhibition, given by 3-OHAA, of the AAPH-initiated lipid peroxidation of both, rat brain synaptosomes and homogenate preparations (assessed by chemiluminescence). For such purposes, the fraction of free radicals (f) trapped per 3-OHAA molecule was determined in each system. Kinetic results show that the oxidation of 3-OHAA follows a process dominated by ROO with a zero order kinetic limit in 3-OHAA, and a fraction (f) equal to 0.88. From the induction times, elicited by 3-OHAA in the kinetic profiles of fluorescein consumption, a fraction (f) of 0.28 was determined. 3-OHAA also generated induction times in the kinetic profiles of light emission during the AAPH-initiated lipid peroxidation of rat brain synaptosomes and homogenates. From such induction times, fractions of 0.61 and 0.63 were determined for rat brain synaptosomes (f) and homogenates (f), respectively. These results show that during the incubation of 3-OHAA and AAPH, a low fraction of ROO self-reacts to generate RO. Nevertheless, when 3-OHAA is employed to protect particular targets, such as fluorescein, rat brain synaptosomes and homogenates, reactions of ROO and/or RO should be considered.
3-羟基邻氨基苯甲酸（3-OHAA）是色氨酸在犬尿氨酸途径中产生的一种代谢产物，是一种高效的抗氧化剂，可抵御由2,2'-偶氮二异丁脒二盐酸盐（AAPH）热解产生的过氧自由基（ROO）。然而，ROO的自身反应会产生烷氧基自由基（RO），这可能会强烈影响清除反应的结果。在本研究中，我们在三种不同体系中研究了RO对3-OHAA清除活性的影响：i）监测3-OHAA与AAPH衍生自由基之间的直接反应（动力学研究）；ii）评估3-OHAA对AAPH诱导的荧光素消耗的保护作用；iii）3-OHAA对AAPH引发的大鼠脑突触体和匀浆制剂脂质过氧化的抑制作用（通过化学发光评估）。为实现这些目的，在每个体系中测定了每个3-OHAA分子捕获的自由基分数（f）。动力学结果表明，3-OHAA的氧化过程以ROO为主导，在3-OHAA中具有零级动力学极限，分数（f）等于0.88。根据3-OHAA在荧光素消耗动力学曲线中引发的诱导时间，确定分数（f）为0.28。3-OHAA在AAPH引发的大鼠脑突触体和匀浆脂质过氧化过程中的发光动力学曲线中也产生了诱导时间。根据这些诱导时间，分别测定大鼠脑突触体（f）和匀浆（f）的分数为0.61和0.63。这些结果表明，在3-OHAA与AAPH孵育期间，一小部分ROO会自身反应生成RO。然而，当使用3-OHAA保护特定靶点（如荧光素、大鼠脑突触体和匀浆）时，应考虑ROO和/或RO的反应。
