title: Induction of oxidative DNA damages and enhancement of cell proliferation in human lymphocytes in vitro by butylated hydroxyanisole.
abstract: The food additive butylated hydroxyanisole (BHA) has been shown to induce gastrointestinal hyperplasia in rodents by an unknown mechanism. The relevance of this observation for human risk assessment is not clear. We therefore analysed the effect of BHA and its primary metabolites tert-butylhydroquinone (TBHQ) and tert-butylquinone (TBQ) on 8-oxo-deoxyguanosine formation and labelling induces in human lymphocytes in vitro. Analysis of culture medium and cell lysate fractions after administration of BHA or metabolites of BHA revealed that BHA and TBHQ undergo biotransformation in whole blood cultures. Moreover, TBQ can be reduced to TBHQ. While in cultures treated with BHA 50-60% of the dose administered was recovered, a much lower dose recovery was found in cultures treated with either TBHQ or TBQ. This indicates a considerable binding of these compounds to macromolecules. BHA and TBHQ, as well as TBQ, induced a dose-dependent increase in cell proliferation of phytohaemagglutinin-stimulated lymphocytes, 50 microM being the optimal dose. Since BHA is metabolized to TBHQ, it is not clear which compound is responsible for the proliferation enhancing effects observed in culture. Inhibition of IBHQ metabolism to its semiquinone radical by acetylsalicylic acid (ASA) reduced the increase in labelling indices induced by TBHQ. This indicates that this metabolic pathway is involved in the enhancement of cell proliferation induced by the hydroquinone. HPLC-ECD analysis of oxidative DNA damage in lymphocytes exposed to 10, 50 and 100 microM BHA, TBHQ or TBQ respectively showed that BHA was not capable of inducing oxidative DNA damage to a significant degree. TBQ and, in particular, TBHQ at a dose of 50 microM (the optimal dose for induction of cell proliferation), however, increased lymphocyte 7-hydroxy-8-oxo-2'-deoxyguanosine formation by 320 and 680% respectively. Inhibition of prostaglandin H synthase by ASA in cultures treated with TBHQ decreased the oxidation ratio significantly, confirming the significance of this enzyme system in the mechanism of toxicity of BHA.
authors.affiliation: Department of Health Risk Analysis and Toxicology, University of Limburg, Maastricht, The Netherlands.
authors.forename: P A
authors.initials: PA
authors.lastname: Schilderman
authors.forename: E
authors.initials: E
authors.lastname: Rhijnsburger
authors.forename: I
authors.initials: I
authors.lastname: Zwingmann
authors.forename: J C
authors.initials: JC
authors.lastname: Kleinjans
mesh_terms: D000818:Animals; D016227:Benzoquinones / Q000633:toxicity; D002083:Butylated Hydroxyanisole / Q000378:metabolism / Q000633:toxicity*; D002455:Cell Division; D002478:Cells, Cultured; D004249:DNA Damage; D004305:Dose-Response Relationship, Drug; D006801:Humans; D006873:Hydroquinones / Q000633:toxicity; D008214:Lymphocytes / Q000166:cytology / Q000187:drug effects*; D010084:Oxidation-Reduction; D012377:Rodentia
chemical_list: D016227:Benzoquinones; D006873:Hydroquinones; D002083:Butylated Hydroxyanisole; C018855:2-tert-butylhydroquinone; C051290:2-tert-butyl-4-quinone