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 ¹⁸O for use in chemical and biochemical studies in the area of nitric oxide research. NaNO₂ (0.1 mmol) is diluted in H₂¹⁸O (45 μl) and acidified with HCl (1 μl, 5 M), and the solution is allowed to equilibrate. Subsequently, the sample is mixed by vortexing with ethyl acetate (500 μl), and the organic phase is dried over anhydrous Na₂SO(4). Ethyl acetate is evaporated to dryness, and the residue is reconstituted in phosphate-buffered saline. In human blood hemolysate, oxyhemoglobin (HbFe¹⁶O₂) was shown to oxidize N¹⁸O₂⁻ to ¹⁶ON¹⁸O₂⁻.