Scheideler L, Ninnemann H
Anal Biochem. 1986 Apr;154(1):29-33. doi: 10.1016/0003-2697(86)90491-4.
Nitrate reductase activity is usually measured by colorimetric determination of the nitrite formed. Since reduced pyridine nucleotides interfere with color formation, the use of NADPH or NADH in the assay requires a specific postassay treatment to remove excess substrate. A "stop mix" containing 1.5 mM phenazine methosulfate and 4.0 mM ferricyanide (final concentrations 0.136 and 0.36 mM, respectively) can remove excess NAD(P)H and terminate the enzymatic reaction quickly in a single, time-saving step. For activity tests containing dithionite we recommend the use of a 1:1 mixture of the two color reagents to avoid incomplete color formation. This may occur during longer time intervals between addition of the color reagents due to destruction of the diazonium salt formed with the first reagent by oxidation product(s) of dithionite.
硝酸还原酶活性通常通过比色法测定所形成的亚硝酸盐来衡量。由于还原型吡啶核苷酸会干扰显色反应，因此在测定中使用NADPH或NADH时，需要进行特定的测定后处理以去除过量的底物。一种含有1.5 mM吩嗪硫酸甲酯和4.0 mM铁氰化物（最终浓度分别为0.136和0.36 mM）的“终止混合液”可以去除过量的NAD(P)H，并在一个省时的步骤中快速终止酶促反应。对于含有连二亚硫酸盐的活性测试，我们建议使用两种显色试剂1:1的混合物，以避免显色不完全。在加入显色试剂之间较长的时间间隔内，由于连二亚硫酸盐的氧化产物破坏了与第一种试剂形成的重氮盐，可能会发生这种情况。
