Yokozawa Yuka, Watanabe Haruki, Matsubara Aya, Horinouchi Rina, Iida Sayaka, Yamamoto Yorihiro, Fujisawa Akio
School of Bioscience and Biotechnology, Tokyo University of Technology, 1404-1 Katakura-cho, Hachioji, Tokyo 192-0982, Japan.
J Clin Biochem Nutr. 2025 Jan;76(1):8-15. doi: 10.3164/jcbn.24-152. Epub 2024 Nov 9.
Uric acid, a water-soluble antioxidant ubiquitous in human bodily fluids at relatively high concentrations, reacts with a variety of reactive oxygen and nitrogen species. Ours and other previous studies identified allantoin, oxaluric acid, triuret, and 5--carboxyimino-6-aminopyrimidine-2,4(3)-dione as specific metabolites reactive against free radicals, singlet oxygen, peroxynitrite, and hypochlorous anion, respectively. We analyzed human plasma spiked with these products using high-performance liquid chromatography-tandem mass spectrometry. We observed recoveries of 40-110% and coefficients of variance within 7%. Samples remained stable at -80°C for at least 4 weeks, indicating the analytical method is sound. Detection of these metabolites in biological samples enables the identification of each species generated . We observed changes in the products in human blood during pseudo-inflammation induced by treatment with lipopolysaccharide. Levels of allantoin, oxaluric acid, triuret, and 5--carboxyimino-6-aminopyrimidine-2,4(3)-dione increased after addition of lipopolysaccharide. The formation of singlet oxygen was confirmed by increased formation of Trp-derived -hydroxide ([2,3a,8a]-3a-hydroxy-1,2,3,3a,8,8a-hexahydropyrrolo[2,3-b]indole-2-carboxylic acid) (-WOH). We believe our method will aid development of strategies to treat oxidative stress-associated diseases.
尿酸是一种在人体体液中普遍存在且浓度相对较高的水溶性抗氧化剂，它能与多种活性氧和氮物种发生反应。我们的研究以及其他先前的研究分别鉴定出尿囊素、草尿酸、三聚氰酸和5-羧基亚氨基-6-氨基嘧啶-2,4(3)-二酮是分别对自由基、单线态氧、过氧亚硝酸盐和次氯酸根阴离子具有反应活性的特定代谢产物。我们使用高效液相色谱-串联质谱法分析了添加这些产物的人血浆。我们观察到回收率为40%-110%，变异系数在7%以内。样品在-80°C下至少4周保持稳定，表明该分析方法可靠。在生物样品中检测这些代谢产物能够鉴定所产生的每种物质。我们观察到在用脂多糖处理诱导的假炎症过程中人体血液中这些产物的变化。添加脂多糖后，尿囊素、草尿酸、三聚氰酸和5-羧基亚氨基-6-氨基嘧啶-2,4(3)-二酮的水平升高。通过色氨酸衍生的-羟基化物（[2,3a,8a]-3a-羟基-1,2,3,3a,8,8a-六氢吡咯并[2,3-b]吲哚-2-羧酸）（-WOH）生成量的增加证实了单线态氧的形成。我们相信我们的方法将有助于开发治疗氧化应激相关疾病的策略。
