Development of a liquid chromatography/tandem mass spectrometry method to investigate the presence of biomarkers of DNA damage in urine related to red meat consumption and risk of colorectal cancer

Abstract

Novel, reproducible, and accurate LC/MS/MS methods were developed for the determination of the urinary content of O(6)CMG and O(6)CMdG, and of the possible formation of O(6)MeG and O(6)MedG by decarboxylation. Clinical samples from volunteers on different diets were analysed. Further studies are required to discover a link between the presence of these biomarkers in urine and red meat consumption.

Quantification of Azaserine-Induced Carboxymethylated and Methylated DNA Lesions in Cells by Nanoflow Liquid Chromatography-Nanoelectrospray Ionization Tandem Mass Spectrometry Coupled with the Stable Isotope-Dilution Method

“…Compared to the previously reported LC/MS-based quantification of O 6 -CMdG and O 6 -MedG in selected-reaction monitoring (SRM) mode on a triple quadrupole mass spectrometer, 17,26 our MS 3 detection mode in the LTQ XL linear ion trap mass spectrometer provided better specificity (see Figures S7–S12 for representative selected-ion chromatograms and tandem mass spectra acquired from MS 2 and MS 3 modes of detection). Additionally, in contrast to the solid-phase extraction method previously used for analyte enrichment, 17,26 the off-line HPLC enrichment method reported herein offered better sample cleanup by removing excess amount of unmodified nucleosides and buffer salts added during the enzymatic digestion, 30 which further improved the sensitivity of our method.…”

“…Additionally, in contrast to the solid-phase extraction method previously used for analyte enrichment, 17,26 the off-line HPLC enrichment method reported herein offered better sample cleanup by removing excess amount of unmodified nucleosides and buffer salts added during the enzymatic digestion, 30 which further improved the sensitivity of our method. Moreover, to our knowledge, our technique is the first application of the nLC-nESI-MS 3 for the simultaneous quantification of multiple carboxymethylated and methylated DNA lesions in cellular DNA, which increased the sensitivity of the analysis by improving ionization efficiency and transmission of ions from the ionization source into the mass analyzer.…”

“…24,25 In this regard, solid-phase extraction followed by LC-MS/MS analysis on a triple-quadrupole mass spectrometer was previously employed for the quantification of O 6 -CMdG and O 6 -MedG, directly or after converting them to the corresponding modified nucleobases with acid hydrolysis at elevated temperature. 17,26 Although neither approach provided adequate sensitivity for detecting these modified nucleosides in human urine samples, 26 LC-MS/MS analysis of the acid-hydrolyzed samples facilitated the successful detection of O 6 -CMdG and O 6 -MedG in DNA isolated from cultured human cells treated with diazoacetate. 17 …”

Urinary DNA adductomics – A novel approach for exposomics

“…Sample workup is simpler than for DNA, and therefore urine is a popular matrix in which to study biomarkers of exposure (Aylward et al 2017), including a wide variety of DNA adducts e.g. oxidized nucleic acid products (Broedbaek et al 2011), N -nitroso-derived DNA adducts (Da Pieve et al 2013), alkylated purines (Hu et al 2018), and aristolochic acid-derived DNA adducts (Leung and Chan 2014), illustrating the breadth of adducts in urine, which parallels that in DNA.…”

Analysis of urinary nucleosides as potential cancer markers determined using LC–MS technique

“…The most popular analytical column is C18 [6,[15][16][17][18][19][57][58][59], however HILIC columns are becoming more popular due to the polar character of compounds like nucleosides [55,60]. Besides, there is also a trend to resemble conditions like UHPLC using routine HPLC system in order to reduce eddy-diffusion and increase flow rates with controlled back pressures.…”