Additive effects and potential inhibitory mechanism of some common aromatic pollutants on in vitro mitochondrial respiration

Abstract

The in vitro toxicity of multiple hydrophobic compounds was the focus of this study. A mitochondrial respiratory assay, sensitive to perturbations caused by hydrophobic chemicals, was utilized to measure the effects of individual aromatic hydrocarbon pollutants and their mixtures on mitochondrial respiratory function. Benzene, naphthalene, acenaphthene, and 1-chloronaphthalene, common industrial solvents shown to interact additively in vivo, were evaluated using this in vitro assay system. Mitochondrial respiration was inhibited 50% (EC50) by 525 ppm (6.7 mM) benzene, 15 ppm (117 microM) naphthalene, 3.9 ppm (25.5 microM) acenaphthene, or 3.8 ppm (23.4 microM) 1-chloronaphthalene. NADH:O2 oxidoreductase (NADH-->O2), NADH:ubiquinone oxidoreductase, and ubiquinol:O2 oxidoreductase activities were inhibited by all four compounds, whereas succinate:O2 oxidoreductase, cytochrome c oxidase, and duroquinol:O2 oxidoreductase activities were not inhibited. Inhibition of mitochondrial respiration occurred at the level of ubiquinone (coenzyme Q10) for all four aromatic hydrocarbons. The ultraviolet absorbance spectrum of isolated Q10 was also altered by naphthalene, acenaphthene, or 1-chloronaphthalene, suggesting a specific interaction between that component of the respiratory chain and these aromatic hydrocarbons. Inhibition by a mixture of 2, 3, or 4 of the compounds tested was additive, reflecting a summation effect of each compound present in the mixture. This additive nature is consistent with previously reported effects of these compounds in vivo and with compounds having similar modes of action. The similar mode of action in vitro is a specific interaction with coenzyme Q10, not a generalized membrane perturbation as speculated to occur in vivo, and is the likely mechanism for the observed additive toxicity.

Ability of 16 priority PAHs to be directly cytotoxic to a cell line from the rainbow trout gill

“…The work of others on the toxicity of PAHs suggested several possibilities. Using isolated mitochondria (Harmon and Sanborn, 1982;Struble and Harmon, 1983;Harmon, 1988;Beach and Harmon, 1992), as well as cells (Harmon and Sanborn, 1982), Harmon and his coworkers found that naphthalene and acenaphthylene specifically inhibited mitochondrial respiration, and did so at the level of ubiquinone (coenzyme Q 10 ). Another school of thought, which has been developed for anesthetic compounds, is that inhibitory actions are due to general changes in membrane integrity (Seeman, 1972).…”

“…Within membranes, the different PAHs could interact in either an additive or non-additive manner to elicit cytotoxicity. In other experimental systems, the toxicity of several aromatic hydrocarbons has been observed to be additive whether the compounds have been thought to be toxic through a general membrane-perturbing effect (Broderius and Kahl, 1985;Deneer et al, 1988;Muñ oz and Tarazona, 1993) or a specific action on mitochondrial function (Beach and Harmon, 1992).…”

“…As well, pronounced morphological changes were observed at naphthalene concentrations near or above the EC 50 value for oxygen consumption. The mechanism of cytotoxicity was studied with isolated mitochondria and appeared to involve the inhibition of mitochondrial respiration at the level of ubiquinone (Struble and Harmon, 1983;Beach and Harmon, 1992). Except for acenaphthene (Beach and Harmon, 1992), the ability of other PAHs to be directly and quickly cytotoxic is largely unexplored.…”

Use of fish gill cells in culture to evaluate the cytotoxicity and photocytotoxicity of intact and photomodified creosote

“…In the past, benzene, naphthalene, acenapthene, and 1-chloronaphthalene were shown to act additively in inhibiting mitochondrial respiration in vitro [27]. In the past, benzene, naphthalene, acenapthene, and 1-chloronaphthalene were shown to act additively in inhibiting mitochondrial respiration in vitro [27].…”

“…Additivity has been an adequate model in the past for explaining the toxicity of mixtures of several creosote components, but in the current study such a model only partly described the cytotoxicity of the creosote solution. In the past, benzene, naphthalene, acenapthene, and 1-chloronaphthalene were shown to act additively in inhibiting mitochondrial respiration in vitro [27]. Naphthalene, acenaphthene, phenanthrene, and anthracene were slightly less than additive at equitoxic concentrations in eliciting acute toxicity to Daphnia magna, but additivity was still thought a sufficient model [28].…”