Isolation, purification and characterization of glycosaminoglycans in the fluids of the mollusc Anodonta cygnea.

The isolation, purification and characterization of Anodonta cygnea haemolymph, and extrapallial fluid glycosaminoglycans (Gags), which have high calcium affinity, were carried out in order to better understand the process of nacreous shell biomineralization. Our results show the existence of two different Gags with similar contents in the two fluids, throughout the year, but with significant seasonal variations for both. In the winter months, we identified by electrophoresis, only one kind of Gag chain (hyaluronic acid) while two different Gags (hyaluronic acid and heparan sulphate-like) from both fluids of A. cygnea were found in summer months. Quantification showed the total Gag fractions of both fluids in average, at their highest concentration (79.8 mg/L) in the highest calcification summer period. In contrast, the period of the year with the lowest concentration of total Gags (54.6 mg/L) occurred during the winter when calcification is reduced. This significant decrease between seasons is correlated mainly with the sulphated fraction, being 37.1 mg/L in the summer while only 9.2 mg/L in the winter haemolymph. The present data suggest that a heparan sulphate-like Gag has a relevant role in the biomineralization mechanisms acting as the calcium carbonate nucleator in the shell.