Copper toxicity to five Parmelia lichens in vitro

Treatment of Parmelia caperata, P. perlata, P. subrudecta, P. sulcata and P. tiliacea with CuSO4 resulted in a time- and copper-concentration-dependent decrease in the total and intracellular potassium concentrations of the thallus, indicating that copper damaged the cytoplasmic membrane. Treatment with copper also resulted in a time-dependent increase in the total copper concentration of the thallus. After 4 h of exposure to copper, the process of potassium efflux was essentially completed but the absorption of copper was still increasing; moreover, the amount of copper bound to the thallus exceeded twice the amount of potassium released from the thallus, suggesting that cupric ions reached intracellular sites in the thallus, and K+/Cu2+ exchange was not electroneutral. After 5 h of exposure to copper, the extent of decrease in the total and intracellular potassium concentrations of the thallus was positively correlated with copper absorption levels, but only at 0.05<P<0.10, suggesting that membrane damage was proportional to the amount of bound copper, but other factors could have been operative, namely binding of copper to the cell wall. Acetone extracts of untreated thalli contained low concentrations of amino acids, polyols, and sugars, but considerable amounts of lichen substances: atranorin, caperatic, constictic, lecanoric, menegazziaic, protocetraric, salazinic, stictic, and usnic acids. Titration of the extracts with copper and assay of the free Cu2+ concentration revealed the presence of copper-binding ligands, and several successive absorption cycles, most probably corresponding to the binding of Cu2+ to each of the lichen substances detected in the extracts. However, no significant correlation (P>0.10) was found between the Cu2+-complexing capacity of acetone extracts and copper-induced membrane damage. It was concluded that in the studied Parmelia species, and in the experimental conditions used in this work, copper toxicity was not a simple function of the Cu2+-binding properties of the lichen substances present in the thallus. Several hypotheses were formulated to interpret the results.