Copper isotope fractionation during excretion from a phototrophic biofilm

The metal excretion and metal sorption are two important microbial processes in the regulation of intracellular metal concentration. However, the contribution of freshwater phototrophic biofilm to metal efflux in the environment and its related isotopic fractionation remain poorly known. In this study, Cu efflux from a mature phototrophic biofilm and related Cu isotopic fractionation between the biofilm and aqueous solution were studied over 4 days in closed batch and open drip-flow reactors (BR and DFR, respectively). The impact of a 48-h drying event on Cu efflux from the biofilm and its isotopic signature was quantified in the BR. Cu excretion from the biofilm depended on Cu concentration in the biomass and did not show a latent phase. For the wet biofilm, in DFR, the Cu efflux rate decreased over time whereas in BR, Cu efflux exhibited fluctuations with a net re-sorption phase. During short-term early excretion over the first 4 h (BR) and 24 h (DFR), the enrichment of solution in heavy isotopes with a Delta Cu-65((sol-biofilm)) of +0.7 +/- 0.2% could be explained by a combination of i) desorption via competition between H+ and Cu2+ for surface sites and ii) release of isotopically heavy Cu(II)-organic complexes. In addition, there was a retention of isotopically light Cu (I) within the cells which is consistent with the biofilm ability to reduce Cu. With further exposure of the wet biofilm to the solution, a progressive decrease of Delta Cu-65((sol-biofilm)), down to 0 and -0.36% respectively in BR and DFR after 96 h, could be explained by an active efflux of Cu(I) and by a passive diffusion of Cu2+, both of these processes favoring light isotopes. The re-hydrated and wet biofilms showed distinctly different behavior during excretion, in terms of magnitude of the flux and direction of the fractionation for long term exposure. Indeed, the efflux from the re-hydrated biofilm was ten times higher than the one from the wet biofilm and the re-hydrated biofilm did not exhibit a decrease of Delta Cu-65((sol-biofilm)) after 24 h of reaction. This could be related to partial devitalization of the biofilm during drying, producing a release of isotopically heavy Cu and decreasing the intensity of Cu(I) active efflux of light isotopes. Taken together, the Cu isotopic signature in natural aquatic environments containing phototrophic biofilms varies up to 1% on a daily scale.