Biological control over the formation and storage of amorphous calcium carbonate by earthworms

The earthworm calciferous gland produces a concentrated suspension of calcium carbonate and in certain species precipitates as concretions of CaCO3, which then enter the soil. Here we investigated the initial stages of CaCO3 formation in the earthworm Lumbricus friendi by means of Fourier transform infrared and electron microscopy techniques (field-emission scanning electron microscopy, transmission electron microscopy, high resolution electron microscopy and selected area electron diffraction). In addition, comparisons between the IR spectra of the water-dissolved carbonic anhydrase (CA) and the glandular secretion ('milky fluid') were performed in order to investigate the mechanisms involved in CaCO3 precipitation. Our results strongly suggest that carbonation starts with the dissolved CO2, which is transformed via deprotonation to HCO3-, then to CO32- and finally to amorphous calcium carbonate (ACC). While ACC stabilization takes place under the biological control, further transformation stages leading to calcite concretions seem to be inorganically driven by an Ostwald ripening process.