Synergistic effects of a photooxidized polycyclic aromatic hydrocarbon andcopper on photosynthesis and plant growth: Evidence that in vivo formationof reactive oxygen species is a mechanism of copper toxicity

Heavy metals and polycyclic aromatic hydrocarbons (PAHs) are often cocontam inants in industrialized environments, yet little is known about either the extent or mechanisms of their cotoxicity. To address this shortfall, the c ombined effects of an oxygenated PAH, 1,2-diliydroxyanthraquinone (1,2-dhAT Q), and a heavy metal, Cu2+, on photosynthesis and growth of the duckweed ( Lemna gibba) were evaluated. Using assays of chlorophyll a fluorescence and photosystem I activity, 1,2-dhATQ inhibited electron transport at the cyto chrome b(6)/f complex Conversely, Cu2+ alone (at low concentrations) had li ttle effect on photosynthesis. When Cu2+ was combined with 1,2-dhATQ, an in crease in transient and steady-state chlorophyll a fluorescence quenching o ccurred relative to 1,2-dhATQ alone. Treatment of isolated thylakoid membra nes with 1,2-dhATQ inhibited whole-chain linear electron transport, measure d as O-2 consumption using methyl viologen as the electron acceptor. Howeve r, Cu2+ plus 1,2-dhATQ resulted in active O-2. consumption with or without methyl viologen as an electron acceptor. From these data, we conclude that 1,2-dhATQ renders the plastoquinone pool to a highly reduced state by inhib iting at cytochrome b(6)/f. Then, Cu2+ is able to mediate the transfer of e lectrons from reduced plastoquinone to O-2, forming reactive oxygen species . At the whole-organism level, when Cu2+ and 1,2-dhATQ were mixed at concen trations that resulted in the above-mentioned impacts on photosynthesis, sy nergistic inhibition of plant growth was observed. This suggests a catalyti c mechanism of toxicity for redox active metals, a process that could be in strumental in explaining their impacts at low concentrations.