There has been a renewed interest in the enzyme arginase for its role in various physiological and pathological processes that go beyond the urea cycle. One such role ascribed to arginase has been that of regulating nitric oxide (NO) production by a substrate (l-arginine) competition between arginase and nitric oxide synthase (NOS). Several arginase inhibitors have been developed to investigate the biological roles of arginase, of which Nω-hydroxy-l-norarginine (nor-NOHA) is commercially available and is used widely from cell culture models to clinical investigations in humans. Despite the prevalence of nor-NOHA to investigate the substrate competition between arginase and NOS, little is known regarding interferences that nor-NOHA could have on common methods to assess NO production. Therefore, we investigated if nor-NOHA has unintended consequences on common NO assessment methods. We show that nor-NOHA spontaneously releases biologically active NO-like molecule in cell culture media by reacting with riboflavin. This NO-like molecule is indistinguishable from an NO donor (NOR-3) using common methods to assess NO. Besides riboflavin, nor-NOHA spontaneously reacts with H2O2 to diminish H2O2 content and produce NO-like molecule in the process. Our investigation provides detailed evidence on unintended artefacts related to nor-NOHA that can limit its use in cell culture, as well as some ex vivo and in vivo models. Future studies on arginase should take into consideration the limitations presented here when using nor-NOHA as a research tool, not only in investigations related to arginase and NOS competition, but also for investigating other biological roles of arginase.

中文翻译：

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精氨酸酶抑制剂Nω-羟基-L-精氨酸自发释放具有生物活性的类NO分子：研究应用的局限性。

由于精氨酸酶在尿素循环以外的各种生理和病理过程中的作用，引起了人们的新兴趣。归因于精氨酸酶的这种作用之一是通过精氨酸酶和一氧化氮合酶（NOS）之间的底物（1-精氨酸）竞争来调节一氧化氮（NO）产生。已经开发了几种精氨酸酶抑制剂来研究精氨酸酶的生物学作用，其中Nω-羟基-1-正精氨酸（nor-NOHA）是可商购的，并且广泛用于从细胞培养模型到人类临床研究中。尽管nor-NOHA广泛用于研究精氨酸酶和NOS之间的底物竞争，但对于nor-NOHA对评估NO产生的常见方法可能产生的干扰知之甚少。因此，我们调查了nor-NOHA是否会对常见的NO评估方法产生意想不到的后果。我们表明，nor-NOHA通过与核黄素反应自发地在细胞培养基中释放具有生物活性的NO样分子。使用常规方法评估NO，这种NO样分子与NO供体（NOR-3）是无法区分的。除了核黄素外，nor-NOHA还会自发地与H2O2反应，以减少H2O2的含量并在此过程中产生类似NO的分子。我们的研究提供了与nor-NOHA相关的意外制品的详细证据，这些制品可能会限制它在细胞培养以及某些离体和体内模型中的使用。在将nor-NOHA用作研究工具时，以后对精氨酸酶的研究应考虑到此处提出的局限性，不仅限于与精氨酸酶和NOS竞争有关的研究，