1. Detection of 8-hydroxydeoxyguanosine (8-OHdG) as a biomarker of oxidative damage in peripheral leukocyte DNA by UHPLC-MS/MS.
- Author
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Wu D, Liu B, Yin J, Xu T, Zhao S, Xu Q, Chen X, and Wang H
- Subjects
- 8-Hydroxy-2'-Deoxyguanosine, Adolescent, Adult, Aged, Biomarkers chemistry, Biomarkers urine, DNA blood, DNA chemistry, Deoxyguanosine blood, Deoxyguanosine chemistry, Deoxyguanosine urine, Female, Humans, Limit of Detection, Linear Models, Male, Middle Aged, Oxidative Stress physiology, Reproducibility of Results, Young Adult, Biomarkers blood, Chromatography, High Pressure Liquid methods, DNA Damage physiology, Deoxyguanosine analogs & derivatives, Leukocytes, Mononuclear chemistry, Tandem Mass Spectrometry methods
- Abstract
8-Hydroxydeoxyguanosine (8-OHdG) is a widely-used biomarker of oxidative DNA damages. 8-OHdG in peripheral blood leukocyte is associated with mutation and cancer risk. The level of 8-OHdG in peripheral blood leukocytes can indicate a long-term response to oxidative stress rather than that in urine. Accurate identification and quantification of leukocyte 8-OHdG are essential for understanding its mechanism of formation, repair, and biological consequences. In this study, a fast and accurate ultra-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method was developed and validated to detect 8-OHdG in human peripheral leukocyte. DNA in blood samples were extracted and digested, then subjected onto UHPLC-MS/MS using an isocratic elution on a Zorbax Eclipse Plus C18 column (2.1×100mm, 1.8μm). Multiple reaction monitoring (MRM) mode was adopted by using [
15 N5 ]-8-OHdG as an internal standard. The assay was linear over the concentration range of 1.0-100nM with R2 =0.999. The accuracy for spiked samples was 90.9%-94.8%, and the intra-day precision was within 3.7%. The limit of detection (LOD) is 0.30nM and limit of quantification (LOQ) is 1.0nM with 5μl of sample injection. By the analysis of human leukocyte 8-OHdG (n=121) using the developed UHPLC-MS/MS method, it demonstrates that the level of leukocyte 8-OHdG of the cancer patients group (n=46) is significantly higher than that of the health control (n=75)., (Copyright © 2017 Elsevier B.V. All rights reserved.)- Published
- 2017
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