18O-Labeled nitrous acid and nitrite: Synthesis, characterization, and oxyhemoglobin-catalyzed oxidation to 18O-labeled nitrate

We describe a simple laboratory method for specific labeling of nitrite with O-18 for use in chemical and biochemical studies in the area of nitric oxide research. NaNO2 (0.1 mmol) is diluted in (H2O)-O-18 (45 mu l) and acidified with HCl (1 mu l, 5 M), and the solution is allowed to equilibrate. Subsequently, the sample is mixed by vortexing with ethyl acetate (500 mu l), and the organic phase is dried over anhydrous Na2SO4. Ethyl acetate is evaporated to dryness, and the residue is reconstituted in phosphate-buffered saline. In human blood hemolysate, oxyhemoglobin (HbFe(16)O(2)) was shown to oxidize (NO2-)-O-18 to (ONO2-)-O-16-O-18. (C) 2011 Elsevier Inc. All rights reserved.