厌氧发酵有机酸体系中NAD+和NADH测定方法的建立  

Establishment of the determination of NAD+ and NADH in the anaerobic fermentation of organic acids

作　　者：李建[1] 陈可泉[1] 黄秀梅[1] 杨卓娜[1] 姜岷[1] 韦萍[1] 

机构地区：[1]南京工业大学制药与生命科学学院材料化学工程国家重点实验室,南京210009

出　　处：《食品科技》2008年第12期254-257,共4页Food Science and Technology

基　　金：国家高技术研究发展计划"863"(2006AA02Z235);国家自然科学基金项目(20606017);江苏省高技术研究计划项目(BG2007001)

摘　　要：建立一种酶法高效测定厌氧发酵有机酸体系中NAD+和NADH的方法。利用酶循环机理,以1.0 mol/L Bicine buffer(pH8.0)、纯乙醇、40 mmol/L EDTA(pH8.0)、4.2 mmol/L MTT、16.6mmol/L PES和50μL的乙醇脱氢酶作为混合反应体系,采用紫外可见分光光度计在570 nm处测定吸光值变化情况,能将发酵体系中的NAD+和NADH准确定量,检出限分别为1.3×10-6 mol/L和5.9×10-6 mol/L,回收率分别在94%～98%和96%～101%范围内。能够快速、精确测定厌氧发酵体系中的NAD+和NADH含量,对于指导厌氧发酵体系辅酶代谢调控分析具有重要意义。An enzyme method for determining NAD^＋ and NADH highly efficient in the anaerobic fermentation broth was developed. Using the recycling mechanism, 1.0 mol/L Bicine buffer（pH8.0）, pure ethanol, 40 mmol/L EDTA（pH8.0）, 4.2 mmol/L MTT, 16.6 mM PES and 50 μL alcohol dehydrogenase as mixed reaction system, and determining the change of absorbance at the wavelength of 570 nm in UV-Vis spectrophotometer could quantitate NAD^＋ and NADH of fermentation system exactly. The detection limit was 5.9×10^-7 mol/L and 1.3×10^-7 mol/L, and the recovery were 94%-98% and 96%-101% respectively. The contents of NAD^＋ and NADH in the anaerobic fermentation broth could be rapidly and exactly determined by this method. It is very useful for the anaerobic metabolic regulation analysis.

关 键 词：NAD^+ NADH 厌氧发酵 酶法 

分 类 号：Q525[生物学—生物化学]