Measurement of 8-oxo-2'-deoxyguanosine and 8-oxo-2'-deoxyadenosine in DNA and human urine by high performance liquid chromatography-electrospray tandem mass spectrometry

Free Radic Biol Med. 2001 Apr 1;30(7):757-64. doi: 10.1016/s0891-5849(01)00462-2.


A method for the determination of 8-oxo-2'-deoxyguanosine and 8-oxo-2'-deoxyadenosine in DNA and urine by High Performance Liquid Chromatography (HPLC)-Tandem Mass Spectrometry is described. For the urine samples there is no sample preparation except for addition of buffer and internal standards followed by redissolvation of precipitate containing 8-oxo-2'-deoxyguanosine and a centrifugation step before the samples are injected onto the HPLC column. The detection limit for 8-oxo-2'-deoxyguanosine and 8-oxo-2'-deoxyadenosine is approximately 0.3 nM corresponding to 7.5 fmol injected. Long runs, that is, > 50 samples, can be analyzed with only minimal loss of sensitivity. The concentrations excreted into urine samples from humans are between 1 and 100 nM for 8-oxo-2'-deoxyguanosine and below 0.3 nM for 8-oxo-2'-deoxyadenosine. In calf thymus DNA levels down to about 1 oxidized guanosine and adenosine per 10(6) unmodified bases can be detected. High levels of 8-oxo-2'-deoxyguanosine were found, 30 per 10(6) 2'-deoxyguanosine, levels of 8-oxo-2'-deoxyadenosine are at or below the detection limit. These findings indicate that High Performance Liquid Chromatography-Tandem Mass Spectrometry is a highly sensitive and specific method for analysis of oxidative DNA modifications in tissue as well as for analysis of excretion of oxidized nucleotides into urine that ensures a minimum artifact formation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • 8-Hydroxy-2'-Deoxyguanosine
  • Animals
  • Buffers
  • Cattle
  • Chromatography, High Pressure Liquid*
  • DNA / analysis*
  • Deoxyadenosines / analysis*
  • Deoxyadenosines / urine
  • Deoxyguanosine / analogs & derivatives*
  • Deoxyguanosine / analysis*
  • Deoxyguanosine / urine
  • Humans
  • Hydrolysis
  • Mass Spectrometry*
  • Sensitivity and Specificity
  • Thymus Gland / chemistry


  • Buffers
  • Deoxyadenosines
  • 2'-deoxy-7,8-dihydro-8-oxoadenosine
  • 8-Hydroxy-2'-Deoxyguanosine
  • DNA
  • Deoxyguanosine