High-Performance Liquid Chromatography Electrospray Ionization Tandem Mass Spectrometry for the Detection and Quantitation of Benzo[a]pyrene-DNA Adducts

Details

• Personal Author:
  Beland FA ; Bowman ED ; Chen SJ ; Churchwell MI ; Culp SJ ; Doerge DR ; Fu PP ; Gyorffy E ; Minarovits J ; Poirier MC ; Schoket B ; Von Tungeln LS ; Weston A
• Description:
  A method, using HPLC combined with electrospray tandem mass spectrometry (ES-MS/MS), was developed and validated to detect and quantify the major DNA adduct resulting from exposure to the ultimate tumorigenic benzo[a]pyrene (BP) metabolite, trans-7,8-dihydroxy-anti-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene (BPDE). Calf thymus DNA was reacted with BPDE, digested enzymatically to nucleosides, and the major DNA adduct, 10-(deoxyguanosin-N(2)-yl)-7,8,9-trihydroxy-7,8,9,10-tetrahydrobenzo[a]pyrene (dG-BPDE), was purified by HPLC. Similar procedures were applied to prepare dG-BPDE-d(8) from [1,2,3,4,5,6,11,12-(2)H(8)]BPDE for use as an internal standard. The HPLC-ES-MS/MS method was validated using a mixture of hydrolyzed salmon testis DNA (82 mug) and 10 pg dG-BPDE (analogous to 6.9 adducts/10(8) nucleotides). The results indicated an inter- and intraday accuracy of 99-100% and precision of 1.6-1.7% (relative standard deviation). When applied to a calf thymus DNA sample modified in vitro with [1,3-(3)H]BPDE, the method gave a value very similar to those obtained by radiolabeling, (32)P-postlabeling, and immunoassay. HPLC-ES-MS/MS analysis of hepatic DNA from mice treated intraperitoneally with 0.5 and 1.0 mg of [7,8-(3)H]BP gave values comparable to those determined by (32)P-postlabeling and immunoassay. Lung DNA from mice fed a 0.3% coal tar diet (containing approximately 2 mg BP/g coal tar) for one month had 0.6 +/- 0.04 dG-BPDE adducts/10(8) nucleotides. This value is much lower than the 102 +/- 14 total DNA adducts/10(8) nucleotides determined by (32)P-postlabeling, which suggests that dG-BPDE makes only a minor contribution to the DNA adducts formed in lung tissue of mice administered coal tar. The HPLC-ES-MS/MS method was used to assess human lung DNA samples for the presence of dG-BPDE. Based upon a limit of detection of 0.3 dG-BPDE adducts/10(8) nucleotides, when using 100 mug of DNA, dG-BPDE was detected in only 1 out of 26 samples. These observations indicate that HPLC-ES-MS/MS is suitable to assess the contribution of BP to DNA damage caused by exposures to polycyclic aromatic hydrocarbon (PAH) mixtures. The results further suggest that dG-BPDE may contribute only a small fraction of the total DNA adducts detected by other DNA adduct methodologies in individuals exposed to PAHs. [Description provided by NIOSH]
• Subjects:
  Animals DNA Environmental Monitoring Hydrocarbons Laboratories Lung Mass Spectrometry Occupational Health Respiratory System Safety
• Keywords:
  Analytical-methods Animal-studies DNA-adducts DNA-damage Exposure-assessment Exposure-levels In-vitro-studies Ionization Laboratory-animals Lung-tissue Mass-spectrometry Polynuclear-aromatic-hydrocarbons Sampling

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• ISSN:
  0893-228X
• Document Type:
  Text
• Genre:
  Journal Article
• Place as Subject:
  Maryland ; OSHA Region 3 ; West Virginia
• CIO:
  National Institute for Occupational Safety and Health (NIOSH)
• Division:
  HELD - Health Effects Laboratory Division
• Topic:
  Workplace Safety & Health
• Location:
  North America
• Volume:
  18
• Issue:
  8
• NIOSHTIC Number:
  nn:20028224
• Citation:
  Chem Res Toxicol 2005 Aug; 18(8):1306-1315
• CAS Registry Number:
  Benzo[a]pyrene (CAS RN 50-32-8)
• Federal Fiscal Year:
  2005
• Peer Reviewed:
  True
• Source Full Name:
  Chemical Research in Toxicology
• Collection(s):
  National Institute for Occupational Safety and Health
• Main Document Checksum:
  urn:sha-512:71661a3df053dde52ce6395cf1d1eb1d5bbc014659cf88915dd3865ea474c1486411b8cd7d6d85e843bc1d46ce63688d79273ced5141b4afa700eaa1fed31a30
• Download URL:
  https://stacks.cdc.gov/view/cdc/189822/cdc_189822_DS1.pdf
• File Type:
  [PDF - 285.91 KB ]