Measurement of nitric oxide synthase activity in vivo and in vitro by gas chromatography-mass spectrometry.

摘要: Measurement of the nitric oxide (NO) metabolites nitrite and nitrate in biological matrices is a reliable method to assess NO synthase (NOS) activity. Unlike "L-citrulline assay," "nitrite/nitrate assay" generally applicable permits acquisition maximum information about what NOS did actually produce. From various analytical approaches methods available so far for quantitation nitrate, mass-spectrometry-based methods, notably gas chromatography-mass spectrometry (GC-MS), offer highest reliability terms accuracy, precision, specificity, minimum interferences, sensitivity, no limitation regarding matrix. Among GC-MS that one utilizing derivatization reagent pentafluorobenzyl bromide single assay permitting measurement nitrate. This possesses unique opportunity measure simultaneously stable-isotope-labeled (i.e. [15N]nitrite [15N]nitrate), which are exclusively produced by NOS-catalyzed oxidation L-[guanidino-15N2]-arginine, L-arginine analog. *NOS may concomitantly produce *NO O2*-, thus forming peroxynitrite (ONOO-), also degrades Thus, nitrite/nitrate provides biologically relevant activity chapter describes protocols vitro (e.g., isolated preparations cells) vivo (in animals humans).