Mårtensson J
Anal Biochem. 1986 Apr;154(1):43-9. doi: 10.1016/0003-2697(86)90493-8.
A method for determination of 4-methylthio-2-hydroxybutyrate and 4-methylthio-2-oxobutyrate in human urine has been devised, based on metoxime formation of the keto acid and a clean-up procedure using a strong anion-exchange resin AG 2 X 8 and ethyl acetate extraction. After alkylation, the compounds were quantified by GC, using a flame photometric sulfur-selective detector. A normal urinary excretion of 0.14 to 0.25 mmol/mol creatinine and 0.07 to 0.22 mmol/mol creatinine of the alpha-hydroxy and alpha-keto acid, respectively, was found, whereas a markedly elevated excretion of the hydroxy acid was noted in subjects with hypermethioninemia. The enzymatic reduction of 4-methylthio-2-oxobutyric acid by lactate dehydrogenase: NAD+ oxidoreductase (EC 1.1.1.17) was also studied. The Km and Kequil values for 4-methylthio-2-oxobutyrate were 1.41 mM and 0.92 X 10(8) M-1. The Vmax value of the enzyme at infinite concentrations of the two substrates was 7.2 mumol/s/mumol enzyme, which indicates low affinity and reduced catalytic activity compared to other known substrates of lactate dehydrogenase. The reaction product 4-methylthio-2-hydroxybutyrate was not inhibitory on the reaction. The M4 isoenzyme of lactate dehydrogenase (rabbit and pig muscle) possessed approximately 20% of the activity of the H4 isoenzyme (pig heart) for the substrate.
已设计出一种测定人尿中4-甲硫基-2-羟基丁酸酯和4-甲硫基-2-氧代丁酸酯的方法，该方法基于酮酸的甲氧肟形成以及使用强阴离子交换树脂AG 2 X 8和乙酸乙酯萃取的净化程序。烷基化后，使用火焰光度硫选择性检测器通过气相色谱对化合物进行定量。发现α-羟基酸和α-酮酸的正常尿排泄量分别为0.14至0.25 mmol/mol肌酐和0.07至0.22 mmol/mol肌酐，而在高甲硫氨酸血症患者中观察到羟基酸排泄量明显升高。还研究了乳酸脱氢酶：NAD +氧化还原酶（EC 1.1.1.17）对4-甲硫基-2-氧代丁酸的酶促还原作用。4-甲硫基-2-氧代丁酸酯的Km和Kequil值分别为1.41 mM和0.92×10（8）M-1。在两种底物无限浓度下该酶的Vmax值为7.2 μmol/s/μmol酶，这表明与乳酸脱氢酶的其他已知底物相比，其亲和力较低且催化活性降低。反应产物4-甲硫基-2-羟基丁酸酯对该反应无抑制作用。乳酸脱氢酶的M4同工酶（兔和猪肌肉）对该底物的活性约为H4同工酶（猪心脏）的20%。
