Tissue distribution and macromolecular binding of extremely low doses of [14C]-benzene in B6C3F1 mice

Abstract

The tissue distribution and macromolecular binding of benzene was studied over a dose range spanning nine-orders of magnitude to determine the nature of the dose-response and to establish benzene's internal dosimetry at doses encompassing human environmental exposures. [14C]-Benzene was administered to B6C3F1 male mice at doses ranging between 700 pg/kg and 500 mg/kg body wt. Tissues, DNA and protein were analyzed for [14C]-benzene content between 0 and 48 h post-exposure (625 Ng/kg and 5 microg/kg dose) by accelerator mass spectrometry (AMS). [14C]-Benzene levels were highest in the liver and peaked within 0.5 h of exposure. Liver DNA adduct levels peaked at 0.5 h, in contrast to bone marrow DNA adduct levels, which peaked at 12-24 h. Dose-response assessments at 1 h showed that adducts and tissue available doses increased linearly with administered dose up to doses of 16 mg/kg body wt. Tissue available doses and liver protein adducts plateau above the 16 mg/kg dose. Furthermore, a larger percentage of the available dose in bone marrow bound to DNA relative to liver. Protein adduct levels were 9- to 43-fold greater than DNA adduct levels. These data show that benzene is bioavailable at human-relevant doses and that DNA and protein adduct formation is linear with dose over a dose range spanning eight orders of magnitude. Finally, these data show that the dose of bioactive metabolites is greater to the bone marrow than the liver and suggests that protein adducts may contribute to benzene's hematoxicity.

Ion-optics calculations of the LLNL AMS system for biochemical 14C measurements

“…Experiments have proven that accelerator mass spectrometry can be successfully applied to areas of biochemical interest, including studies of nutrition, toxicology, pharmacokinetics and human carcinogenesis [e.g., [1][2][3][4]. Such experiments were conducted at large multi-purpose accelerator facilities, mainly dedicated to the more the "traditional" fields of AMS research: geochemistry, geochronology, archeology and the environmental sciences.…”

Attomole level protein sequencing by Edman degradation coupled with accelerator mass spectrometry

“…In these equations, the mass m carrier ϩ buffer was expressed in mg of C, and the isotope ratios R carrier , R fraction were expressed in mol of 14 C͞mg of C. Fig. 1 shows the 14 C level of HPLC fractions for ten cycles.…”

“…Automated protein sequencing was performed on a 477A sequencer equipped with a 120A HPLC system (PE Applied Biosystems). All reagents and solvents used for the sequencer were obtained from PE Applied Biosystems and checked for 14 C content by AMS before use. The solution of [ 14 C]GST was diluted to concentrations of 10-1,000 amol͞ l with 0.1% TFA in water containing ␤-lactoglobulin (2.5 pmol͞ l).…”

“…AMS can accurately measure Ͻ10 6 atoms [1 attomole (amol)] of 14 C in a sample containing Ϸ1 mg of total carbon and can analyze hundreds of samples per day. Unlike liquid scintillation counting (LSC), which counts decay events of a radioisotope, AMS is a mass spectrometric technique that detects individual 14 C nuclei. Over the past decade, AMS has been applied to life science studies and has increased precision and sensitivity of detection for 3 H and 14 C by several magnitudes (12)(13)(14)(15)(16)(17)(18).…”

Frequent p53 and H-ras Mutations in Benzene- and Ethylene Oxide-Induced Mammary Gland Carcinomas from B6C3F1 Mice

“…Several studies indicate that metabolism of benzene to its reactive metabolites is a prerequisite for its cytotoxic, genotoxic, and carcinogenic properties (Gut et al, 1996;Snyder and Hedli, 1996;Valentine et al, 1996;Abernethy et al, 2004). It has long been recognized that several metabolites of benzene can bind covalently to proteins and nucleic acids (Creek et al, 1997), and certain metabolites can be oxidized by peroxidases resulting in the production of reactive oxygen species (ROS) and DNA reactive quinones (Hiraku and Kawanishi, 1996;Farris et al, 1997). Several studies suggest that p53 dysfunction is a potential factor in benzene-induced carcinogenesis (Donehower, 1996;Boley et al, 2002;Yoon et al, 2003;Hirabayashi et al, 2004).…”