Is the aquatic macrophyte Crassula helmsii a genuine copper hyperaccumulator?

Aims: The Australian native hemi-aquatic herb C. helmsii (Crassulaceae) is a copper (Cu) accumulator, tolerant to a wide range of climatic conditions and able to concentrate >9000 μg Cu g⁻¹ in its living tissues. These characteristics suggest practical potential for decontamination of Cu-polluted mine waste waters using a phyto-extraction approach. The aim of this study was to investigate Cu uptake in C. helmsii at different Cu²⁺ concentrations in solution and to test for the effect of pH on the concentration of free Cu²⁺ and Cu accumulation in this species. Methods: Different solutions were tested in acid (pH 4.0) and mildly acidic (pH 5.8) conditions at five different Cu²⁺ concentrations (ranging from 0 to 5 mg Cu L⁻¹). The concentration of free Cu²⁺ was evaluated using Geochem-EZ software. The distribution of Cu in C. helmsii tissues was examined with micro-X-ray Fluorescence (μXRF) elemental mapping and with Scanning Electron Microscopy with Energy Dispersive X-ray Spectroscopy (SEM-EDS). Results: The highest shoot Cu concentration in C. helmsii was 5870 μg Cu g⁻¹ in the 5 mg Cu L⁻¹ treatment. Copper bioaccumulation was positively correlated with the Cu²⁺ treatment in the different solutions. The μXRF and SEM-EDS analysis revealed that roots accumulated higher Cu concentrations than the shoots. The concentration of free Cu²⁺ was shown to be dependent upon the type of Fe-chelator (DTPA, EDDHA, HBED) used in the solution due to the displacement of Fe³⁺ and complexation of Cu²⁺. Conclusions: Crassula helmsii is highly tolerant to Cu²⁺ in solution at acidic and mildly acidic pH, and able to accumulate high Cu concentrations in its tissue. This confirms its potential for applications in Cu phyto-extraction of acidic mine effluent waters high in Cu²⁺.