A thermospray liquid chromatography/mass spectrometry method for analysis of human urine for the major malondialdehyde-guanine adduct

Abstract

A method is described for detection and quantitation of the major malondialdehyde-guanine adduct (M1G) based on thermospray liquid chromatography/mass spectrometry. A stable isotope analog of M1G ([2H2]M1G) was used as an internal standard. Thermospray mass spectra of M1G and [2H2]M1G showed intense protonated molecular (MH+) ions that were suitable for use in quantitation of M1G. M1G was purified from human urine and reduced with NaBH4 to a dihydro derivative that was cleanly separated from the contaminants in the urine. The detection limit of reduced M1G by thermospray liquid chromatography/mass spectrometry in the selected ion monitoring mode was 250 fmol on column. Six human urine samples were analyzed, and the concentrations of M1G were below the limit of detection of the assay (500 fmol/mL).

Development of Monoclonal Antibodies to the Malondialdehyde−Deoxyguanosine Adduct, Pyrimidopurinone1

“…Furthermore, Seto detected M 1 G-R in urine and confirmed its structure by mass spectrometry (). There is a report of the detection of the base, M 1 G, in human urine by HPLC with fluorescence detection, but we were unable to confirm this finding by mass spectrometry ( , ).…”

Measurement of the malondialdehyde–2′‐deoxyguanosine adduct in human urine by immuno‐extraction and liquid chromatography/atmospheric pressure chemical ionization tandem mass spectrometry

“…The low excretion level obtained with the immuno‐extraction LC/APCI‐MS/MS method, combined with the complexity of urine, have clearly demonstrated that detection and quantification of M 1 G‐dR in urine represent a major analytical challenge. This might explain the scarcity of data previously published on lipid peroxidation DNA adducts in humans, particularly urinary excretion 18, 20, 22. The measurement of M 1 G‐dR excretion rates in exposed individuals, e.g.…”

“…Protocols for antibody preparation and immuno‐extraction are given in detail elsewhere 10, 12, 17. The purified sample was chemically reduced with 10 µl of 10 mg ml −1 NaBH 4 18. The reagent was removed on a 50 mg C 18 solid‐phase extraction column, vacuum centrifuged to dryness and dissolved in a final volume of 100 µl of 5 m M ammonium acetate buffer (pH 5.0) before LC/MS/MS analysis.…”

Monitoring of 8-oxo-7,8-dihydro-2′-deoxyguanosine in urine by high-performance liquid chromatography after pre-column derivatization with glyoxal reagents

“…Of these markers, 8-OHdG has been widely used for the diagnosis of oxidative stress caused by smoking (Cooke et al, 2002(Cooke et al, , 2005Harman et al, 2003) and has also been reported as the DNA damage marker of cancer (Kasai, 2003;Cooke et al, 2005), Parkinson's disease (Sato et al, 2005) and Alzheimer's disease (de la Monte et al, 2000). Urinary 8-OH-dG has been monitored by gas chromatographymass spectrometry (GC-MS; Teixeira et al, 1995), high-performance liquid chromatography with electrochemical detection (HPLC-ECD; Cooke et al, 2002;Kasai, 2003), HPLC with mass spectrometry (HPLC-MS; Harman et al, 2003;Jajoo et al, 1992) and enzymelinked immunoassay (ELISA; Chiou et al, 2003). HPLC-ECD required sample purification to avoid interferences from charged species in urine samples and required frequent maintenance of the cell electrode (Cooke et al, 2002, Kasai, 2003.…”

“…HPLC-ECD required sample purification to avoid interferences from charged species in urine samples and required frequent maintenance of the cell electrode (Cooke et al, 2002, Kasai, 2003. HPLC-MS required the addition of expensive stable isotope-labeled internal standard (Harman et al, 2003;Jajoo et al, 1992). The urinary data by ELISA were higher than those from the HPLC-ECD method (Kasai et al, 2003), possibly because of the quality of the commercial monoclonal antibody.…”