Pou S, Keaton L, Surichamorn W, Frigillana P, Rosen G M
Department of Pharmaceutical Sciences, University of Maryland School of Pharmacy, Baltimore 21201.
Biochim Biophys Acta. 1994 Sep 28;1201(1):118-24. doi: 10.1016/0304-4165(94)90160-0.
Increasing interest in the study of nitric oxide (NO.) in many facets of biological research necessitates a search for accurate techniques to directly identify the free radical. One recently employed strategy for NO. detection is the method of electron spin resonance (ESR) used in combination with nitrone and nitroso spin traps. Applying this technique to our studies with nitric oxide synthase (NOS), we found that NO. generated directly from the enzyme system could not be detected. Further investigation revealed that 3,5-dibromo-4-nitrosobenzenesulfonic acid (DBNBS) inhibited NO. generation by NOS at concentrations used for spin trapping. Reexamining the ability of various nitrones and DBNBS to spin trap authentic NO. dissolved in buffer, we obtained ESR spectra similar to those previously reported for the spin trap DBNBS. However, continuing our studies with 15NO. and N-hydroxylamine, we found these spectra to be artifactual. Our results emphasize the need to synthesize new spin traps, since currently available compounds are not capable of spin trapping NO. generated by NOS.
在生物研究的诸多方面，对一氧化氮（NO.）研究的兴趣与日俱增，这就需要寻找精确的技术来直接鉴定这种自由基。一种最近采用的检测NO.的策略是将电子自旋共振（ESR）方法与硝酮和亚硝基自旋捕获剂结合使用。将该技术应用于我们对一氧化氮合酶（NOS）的研究时，我们发现无法检测到直接由酶系统产生的NO.。进一步的研究表明，3,5-二溴-4-亚硝基苯磺酸（DBNBS）在用于自旋捕获的浓度下会抑制NOS产生NO.。重新审视各种硝酮和DBNBS自旋捕获溶解在缓冲液中的真实NO.的能力时，我们获得了与先前报道的自旋捕获剂DBNBS相似的ESR光谱。然而，在用15NO.和N-羟胺继续我们的研究时，我们发现这些光谱是人为的。我们的结果强调需要合成新的自旋捕获剂，因为目前可用的化合物无法自旋捕获由NOS产生的NO.。
