title: Quantitative determination of ɛ-N-carboxymethyl-L-lysine in human plasma by liquid chromatography-tandem mass spectrometry.
abstract: ɛ-N-carboxymethyl-L-lysine (CML) is a stable chemical modification of protein lysine residues resulting from glycation and oxidation reactions and a potential biomarker of oxidative stress caused by sugar and lipid oxidation. In this study, a rapid, simple and sensitive method based on liquid chromatography-tandem spectrometry (LC-MS/MS) for the determination of CML in human plasma has been developed and validated. Sample preparation involved protein precipitation using trichloroacetic acid after addition of deuterated CML as internal standard. Chromatography was performed on an amino column by gradient-elution with a mobile phase containing acetonitrile:ultrapure water (80:20, v/v). CML and CML-d2 were detected by multiple reaction monitoring mode with ion pairs 205.0/130.1 and 207.2/84.1 respectively. The assay was linear in the range 10-1000 ng/mL with a lower limit of quantitation (LLOQ) of 10 ng/mL and recovery >90%. Assay validation showed that inter- and intra-day precision and accuracy were satisfactory. The method was applied to compare plasma CML levels in healthy Chinese subjects and patients with diabetes and uremia. In healthy subjects CML concentration (mean±SD) was 16.6±7.8 ng/mL. CML level in diabetic patients was not significantly different from healthy subjects whereas the level in patients with uremia was significantly higher than both healthy subjects and diabetic patients (P<0.001). The method will be useful to assess the value of CML as a biomarker of diabetic vascular complications resulting from elevated oxidative stress.
authors: Center of Drug Metabolism and Pharmacokinetics, China Pharmaceutical University, Nanjing 210009, China.
authors: Liqing
authors: L
authors: Kuang
authors: Department of Clinical Pharmacy, China Pharmaceutical University, Nanjing 210009, China.
authors: Zhiqiang
authors: Z
authors: Jing
authors: Center of Drug Metabolism and Pharmacokinetics, China Pharmaceutical University, Nanjing 210009, China.
authors: Jing
authors: J
authors: Wang
authors: Center of Drug Metabolism and Pharmacokinetics, China Pharmaceutical University, Nanjing 210009, China.
authors: Liyuan
authors: L
authors: Ma
authors: Center of Drug Metabolism and Pharmacokinetics, China Pharmaceutical University, Nanjing 210009, China.
authors: Xiaoqiang
authors: X
authors: Liu
authors: Center of Drug Metabolism and Pharmacokinetics, China Pharmaceutical University, Nanjing 210009, China. Electronic address: yjcpu@yahoo.com.
authors: Jin
authors: J
authors: Yang
mesh_terms: D015415:Biomarkers / Q000097:blood; D016022:Case-Control Studies; D002853:Chromatography, Liquid / Q000379:methods*; D003920:Diabetes Mellitus / Q000097:blood; D003925:Diabetic Angiopathies / Q000097:blood; D006801:Humans; D057230:Limit of Detection; D008239:Lysine / Q000031:analogs & derivatives* / Q000097:blood; D018384:Oxidative Stress; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D053719:Tandem Mass Spectrometry / Q000379:methods*; D014511:Uremia / Q000097:blood
chemical_list: D015415:Biomarkers; C048496:N(6)-carboxymethyllysine; D008239:Lysine
keywords: Biomaker; Diabetic vascular complications; Human plasma; LC–MS/MS; ɛ-N-carboxymethyl-l-lysine