St John R T, Hollocher T C
J Biol Chem. 1977 Jan 10;252(1):212-8.
The pathway of anaerobic reduction of nitrite to nitrogen gas (N2) by cell suspensions of the denitrifier, Pseudomonas aeruginosa, was studied using the techniques of gas chromatography and mass spectrometry. While release of nitrous oxide (N2O) is not normally detected during the reduction of nitrite to N2 by this organism, 15N from [15N]nitrite nevertheless can be trapped quantitatively as 15N2O in a pool of added N2O. In such experiments the abundance of 15N in N2O always exceeds that in product N2, consistent with the absence of a major reductive route from nitrite to N2 which by-passes N2O. During the reduction of a mixture of [15N]nitrite and nitric oxide (NO), 15NO produced at most only in trace amounts. The final products are chiefly 15N2 and 14N2 with only a small fraction of the scrambled product, 14N15N. Much of the 14N15N can be accounted for as an artifact caused by traces of molecular oxygen, which promote the conversion of NO to nitrite by autooxidation and thereby degrade slightly the isotopic purity of [15N]nitrite. Nitrous oxide shows all the properties of a free obligatory intermediate during the denitrification of nitrite to N2 by P. aeruginosa, whereas NO does not. The inability to trap 15NO in a pool of NO indicates that NO is not a free obligatory intermediate in the reduction of nitrite. The small mole fractions of 14N15N produced from a mixture of [15N]nitrite and NO require that the main reductive pathways for these nitrogen oxides cannot share any freely diffusible mono-nitrogen intermediate in common. The simplest interpretation is that nitrite and NO are denitrified by separate pathways, at least prior to the formation of the first bi-nitrogen compound.
利用气相色谱法和质谱法技术，对反硝化菌铜绿假单胞菌细胞悬液将亚硝酸盐厌氧还原为氮气（N₂）的途径进行了研究。虽然在该生物体将亚硝酸盐还原为N₂的过程中通常检测不到一氧化二氮（N₂O）的释放，但来自[¹⁵N]亚硝酸盐的¹⁵N仍可作为¹⁵N₂O定量捕获在添加的N₂O池中。在这类实验中，N₂O中¹⁵N的丰度总是超过产物N₂中的丰度，这与不存在从亚硝酸盐到N₂的绕过N₂O的主要还原途径一致。在[¹⁵N]亚硝酸盐和一氧化氮（NO）混合物的还原过程中，产生的¹⁵NO最多仅为痕量。最终产物主要是¹⁵N₂和¹⁴N₂，只有一小部分是混和产物¹⁴N¹⁵N。大部分的¹⁴N¹⁵N可解释为由痕量分子氧引起的假象，分子氧通过自氧化促进NO转化为亚硝酸盐，从而略微降低了[¹⁵N]亚硝酸盐的同位素纯度。在铜绿假单胞菌将亚硝酸盐反硝化生成N₂的过程中，一氧化二氮表现出游离必需中间体的所有特性，而NO则不然。无法在NO池中捕获¹⁵NO表明NO不是亚硝酸盐还原过程中的游离必需中间体。由[¹⁵N]亚硝酸盐和NO混合物产生的¹⁴N¹⁵N的小摩尔分数表明，这些氮氧化物的主要还原途径不能共享任何自由扩散的单氮中间体。最简单的解释是，亚硝酸盐和NO通过不同途径进行反硝化，至少在形成第一个双氮化合物之前是这样。
