Factors Influencing the Formation of the Carbon Dioxide Radical Anion (CO₂⁻) Spin Adduct of Pbn in the Rat Liver Metabolism of Halocarbons

Abstract

Spin trapping techniques have been used to detect free radicals generated from the in vitro metabolism by rat liver microsomes of carbon tetrachloride (CCl4) and bromotrichloromethane (BrCCl3) under conditions of varying oxygen tension and pH. Dispersions of rat liver microsomes incubated with 12CCl4, 13CCl4 or Br12CCl3, alpha-phenyl-tert-butyl nitrone (PBN) and NADPH/NADH in a phosphate buffer varying in pH from 6.6 to 8.0 under varying oxygen tensions produced various amounts of four different PBN adducts: PBN-CCl3, PBN-L, PBN-OL and PBN-CO2- where L is a carbon-centered lipid type radical and LO is an oxygen-centered lipid type radical. The relative amount of PBN-CO2- increases with the absence of oxygen. With the use of 31P-NMR in vivo spectroscopy it was possible to detect a pH change from 7.4 to 6.8 in the livers of rats treated with CCl4 or BrCCl3. These results suggest that halocarbon metabolism in biological systems may depend on both oxygen tension as well as pH.