上海交通大学学报(医学版)

• 技术与方法 • 上一篇    

高效液相色谱-串联质谱法检测糖尿病患者血浆甲基乙二醛和乙二醛浓度

孔 祥1,2,黄 凯3,张洪梅1,杨 震1,李晓永1,苏 青1   

  1. 1.上海交通大学 医学院附属新华医院内分泌科, 上海 200092; 2.皖南医学院 药理学教研室, 芜湖 241002; 3.无锡市人民医院Ⅰ期临床研究室, 无锡 214023
  • 出版日期:2013-08-28 发布日期:2013-09-16
  • 通讯作者: 苏 青, 电子信箱: suqingxinhua@163.com。
  • 作者简介:孔 祥(1982—), 男, 讲师, 博士生; 电子信箱: wnmcyaolikx@sina.com。
  • 基金资助:

    国家自然科学基金(30872727,81000332)

Determination of plasma methylglyoxal and glyoxal concentrations in patients with diabetes mellitus by high-performance liquid chromatographic/tandem mass spectrometry

KONG Xiang1,2, HUANG Kai3, ZHANG Hong-mei1, YANG Zhen1, LI Xiao-yong1, SU Qing1   

  1. 1.Department of Endocrinology, Xinhua Hospital, Shanghai Jiaotong University School of Medicine, Shanghai 200092, China; 2.Department of Pharmacology, Wannan Medical College, Wuhu 241002, China; 3.Phase I Clinical Laboratory, Wuxi People's Hospital, Wuxi 214023, China
  • Online:2013-08-28 Published:2013-09-16
  • Supported by:

    National Natural Science Foundation of China, 30872727,81000332

摘要:

目的 建立一种测定糖尿病患者血浆甲基乙二醛和乙二醛浓度的高效液相色谱-串联质谱法(HPLC-MS/MS)。方法 采用HPLC-MS/MS技术测定1型(n=15)和2型糖尿病(n=17)患者以及正常受试者(n=10)血浆甲基乙二醛和乙二醛的浓度。待测血浆加入2,3-二氨基萘(2,3-DAN)和内标乙二酮,充分反应及用乙腈沉淀蛋白后,以乙腈-0.1%甲酸水溶液(57∶43,v/v)为流动相,通过Hypersil GOLD色谱柱(50 mm×2.1 mm,5 μm)分离;采用电喷雾电离源,选择正离子检测,甲基乙二醛-DAN结合物、乙二醛-DAN结合物和乙二酮-DAN结合物用于定量分析的选择性离子分别为m/z 195.18→m/z 126.25、m/z 181.14→m/z 126.10和m/z 237.34→m/z 209.34。结果 甲基乙二醛-DAN结合物和乙二醛-DAN结合物线性范围分别为2~1 000 ng/mL和50~1 000 ng/mL,平均回收率均>70%。1型和2型糖尿病患者血浆甲基乙二醛和乙二醛浓度显著高于正常受试者。结论 成功建立测定糖尿病患者血浆中甲基乙二醛和乙二醛浓度的HPLC-MS/MS方法,灵敏度、准确度和回收率均较高,专属性强,适用于大批临床样本的分析。

关键词: 甲基乙二醛, 乙二醛, 高效液相色谱-串联质谱法, 糖尿病

Abstract:

Objective To develop a high-performance liquid chromatographic/tandem mass spectrometry (HPLC-MS/MS) method for the determination of plasma methylglyoxal and glyoxal concentrations in patients with diabetes mellitus. Methods HPLC-MS/MS method was employed to measure the plasma methylglyoxal and glyoxal concentrations in patients with type 1 diabetes mellitus (n=15) and type 2 diabetes mellitus (n=17) and normal subjects (n=10). 2,3-diaminonaphthalene (2,3-DAN) and hexanedione (using as internal standard) were added to plasma samples. After sufficient reaction and precipitation with acetonitrile from the plasma, methylglyoxal-DAN, glyoxal-DAN and hexanedione-DAN were isolated chromatographically on a Hypersil GOLD column (50 mm×2.1 mm, 5 μm). The mobile phase consisted of acetonitrile-0.1% formic acid (57∶43, v/v). Electrospray ionization source was applied and operated in the positive ion mode. The transitions of m/z 195.18→m/z 126.25, m/z 181.14→m/z 126.10 and m/z 237.34→m/z 209.34 were used to quantify methylglyoxal-DAN, glyoxal-DAN and hexanedione-DAN respectively. Results The linear calibration curves were obtained in the concentration range of 2-1 000 ng/mL and 50-1 000 ng/mL for methylglyoxal-DAN and glyoxal-DAN respectively. The average recovery rate was more than 70%. The plasma methylglyoxal and glyoxal concentrations in patients with type 1 and type 2 diabetes mellitus were significantly higher than those in normal subjects. Conclusion The established HPLC-MS/MS method is proved to be suitable for determination of plasma methylglyoxal and glyoxal concentrations in patients with diabetes mellitus, which offers advantages of high sensitivity and selectivity.

Key words: methylglyoxal, glyoxal, high-performance liquid chromatographic/tandem mass spectrometry, diabetes mellitus