硝酸盐还原酶一步法测定血清中硝酸盐浓度

Determination of Serum Nitrate Concentration by Nitrate Reductase One Step Method

为建立简便快速的一氧化氮 (NO)间接测定方法 ,根据硝酸盐还原酶还原硝酸盐 (NO3- )至亚硝酸盐 (NO2 - )的过程中 ,所消耗的还原型辅酶Ⅱ (NADPH)的量与NO3- 浓度呈正比的原理 ,用 340nm处NADPH吸光度的下降值 ,推算出NO3- 的量。所建体系的反应时间为 90min ,NADPH浓度为 6mmol/L ,硝酸盐还原酶浓度为 5 0 0U/L。NaNO3 在 0u mol/L～ 2 0 0umol/L范围内 ,与NADPH吸光度的变化量具有良好的线性关系 ,r =0 993。批内变异系数 (CV) =4 4% ,批间CV =10 5 % ;回收率为 92 36 %。 2 0例 2 5～ 5 5岁的正常人血清中NO3- 浓度为 5 2 42± 19 76umol/L。用此法测定血清中NO3- 浓度简便快速、特异性高、干扰因素少 ,不需昂贵仪器 。

In order to set up a simple and swift method for indirect determination of nitric oxide,in accordance with the principle that the amount of nicotinamide adenine dinucleotide phosphate (NADPH) and NO 3 - are in direct proportion,consumed in the process from nitrate ( NO 3 -) to nitrite ( NO 2 -) by nitrate reductase, NO 3 - could be calculated by the descending value of NADPH of absorbandce at 340nm.The reaction time of the established system was 90 minutes,NADPH concentration was 6 mmol/L and the nitrate reductase concentration was 500U/L.In the scope of 0umol/L～200umol/L,there was a better liner relationship between NaNO 3 and the changing amount of NADPH absorbance,with r=0 993,coefficient of variation (CV)=4 4% in group processing,CV=10 5% between group processing,and the recovery rate in 92 36%,and serum NO 3 - concentration (52 42±19 76)umol/L in 20 normal adults aging from 25 to 55.This method to determine serum NO 3 - concentration is simple,swift and higher in specificity,with less interfering factors and without expensive instruments,and is worthy to be extensivey applied for routine determination in the laboratory.