Determination of glyoxal and methylglyoxal in serum by UHPLC coupled with fluorescence detection
Determination of glyoxal and methylglyoxal in serum by UHPLC coupled with fluorescence detection
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DOI：
10.1016/j.ab.2019.02.014
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发表时间：
2019-05-15
影响因子：
2.9
通讯作者：
Muncha, Gerald
中科院分区：
文献类型：
--
作者：
Dhananjayan, Karthik;Irrgang, Felix;Muncha, Gerald
乙二醛（GO）和甲基乙二醛（MGO）是糖尿病的两种重要生物标志物。血清样品中氧化石墨烯和MGO的分析方法为紫外-可见高效液相色谱法（灵敏度低）或质谱联用法（成本高）。这些缺点阻碍了将这些生物标志物作为糖尿病诊断的常规分析物引入临床实验室。在本研究中，我们引入了一种UHPLC荧光检测方法，通过在中性pH下，用5,6 -二氨基-2,4-二羟基嘧啶硫酸酯（DDP）柱前衍生生成lumazines，来测定血清样品中的GO和MGO。该方法按照FDA指南进行了验证。利用该方法，我们测定了多种动物血清样品中的GO和MGO，例如测定了成年牛血清中GO和MGO的浓度分别为852 +/- 27和192 +/- 10 nmol/L。在人类血清中，非糖尿病受试者（n = 14）的GO和MGO水平分别为154 +/- 88和98 +/- 27 nmol/L，糖尿病受试者（n = 14）的血清样品中GO和MGO水平分别为244 +/- 137和190 +/- 68 nmol/L。此外，干扰研究表明，生理血清成分不会导致氧化石墨烯和MGO水平的人为增加。
Glyoxal (GO) and methylglyoxal (MGO) are two important biomarkers in diabetes. Analytical methods for determination of GO and MGO in serum samples are either HPLC with UV-Vis (low sensitivity) or MS/MS (expensive) detection. These disadvantages have hampered the introduction of these biomarkers as a routine analyte for diabetes diagnostics into the clinical laboratory. In this study, we introduce a UHPLC method with fluorescence detection for the measurement of GO and MGO in serum samples by pre-column derivatization at neutral pH with 5, 6-diamino-2,4-dihydroxypyrimidine sulfate (DDP) to form lumazines. The method was validated as per FDA guidelines. Using this method, we have determined GO and MGO in a variety of animal serum samples, and for example, determined the GO and MGO concentration in adult bovine serum to be 852 +/- 27 and 192 +/- 10 nmol/L, respectively. In human serum, GO and MGO levels in non-diabetic subjects (n = 14) were determined to be 154 +/- 88 and 98 +/- 27 nmol/L, and in serum samples from subjects with diabetes (n = 14) 244 +/- 137 and 190 +/- 68 nmol/L, respectively. In addition, interference studies showed that physiological serum components did not lead to an artificial increase in the levels of GO and MGO.