Robak J, Marcinkiewicz E, Michalska Z, Gryglewski R J
Department of Pharmacology, Medical College of Jagiellonian University, Kraków, Poland.
Pol J Pharmacol. 1997 Aug;49(4):255-62.
Nitric oxide (NO) or nitrite (NO2-) were assayed using the Werringloer's method or the Griess' method, respectively, in the presence or absence of various thiols, amino acids, or albumin. This has been done because both methods are used to determine the generation of endogenous NO from L-arginine or exogenous NO from drugs in vivo, paying little attention to biological constituents which may affect results of these assays. Albumin, reduced glutathione (GSH), cysteine and N-acetylcysteine, but not other amino acids lowered the amount of NO2- as detected by Griess' method no matter whether sodium nitrite or 3-morpholinosydnonimine (SIN-1) were used as a source of NO2-. This happened probably because at low pH of the reaction mixture the corresponding nitrosothiols were formed and thus NO2- was not accessible for detection. However, this phenomenon was not seen when instead of SIN-1 another NO donor--S-nitroso-N-acetylpenicillamine (SNAP) was used. SNAP is a nitrosothiol itself and physiological low molecular thiols (e.g. GSH or cysteine) displaced NO from SNAP. An increase in the amount of released NO was detectable by both Werringloer's and Griess' methods. Only the presence of 700 microns of albumin steadily suppressed the detection of NO or NO2- no matter what was the source of these species. It is concluded that low molecular thiols and albumin may differently influence the detection of both NO and NO2- which derive from various NO donors or sodium nitrite.
分别采用韦林勒尔法或格里斯法，在存在或不存在各种硫醇、氨基酸或白蛋白的情况下，对一氧化氮（NO）或亚硝酸盐（NO₂⁻）进行测定。之所以这样做，是因为这两种方法都用于测定体内由L-精氨酸产生的内源性NO或药物产生的外源性NO，而很少关注可能影响这些测定结果的生物成分。白蛋白、还原型谷胱甘肽（GSH）、半胱氨酸和N-乙酰半胱氨酸，但其他氨基酸则不然，无论使用亚硝酸钠还是3-吗啉代 sydnonimine（SIN-1）作为NO₂⁻的来源，通过格里斯法检测到的NO₂⁻量都会降低。这可能是因为在反应混合物的低pH值下形成了相应的亚硝基硫醇，因此无法检测到NO₂⁻。然而，当使用另一种NO供体——S-亚硝基-N-乙酰青霉胺（SNAP）代替SIN-1时，这种现象并未出现。SNAP本身就是一种亚硝基硫醇，生理性低分子硫醇（如GSH或半胱氨酸）会将NO从SNAP中置换出来。通过韦林勒尔法和格里斯法都可检测到释放的NO量增加。无论这些物质的来源是什么，只有存在700微米的白蛋白会持续抑制NO或NO₂⁻的检测。结论是，低分子硫醇和白蛋白可能会对源自各种NO供体或亚硝酸钠的NO和NO₂⁻的检测产生不同影响。
