Isolation and Characterization of a Cat 2+ ‐Binding Polysaccharide Associated with Coccoliths of Emiliania huxleyi (Lohmann) Kamptner

Coccolithophoridae, a group of mostly unicellular algae, possess a cell wall containing calcified plates, called coccoliths. The coccoliths from the species Emiliania huxleyi (Lohmann) Kamptner contain a water‐soluble acid polysaccharide. In this paper we describe the isolation and some characteristic properties of the polysaccharide, in particular its Ca 2+ ‐binding capacity. A large‐scale cultivation of the Coccolithophoridae was worked out and a new procedure for isolating coccoliths was developed. The polysaccharide obtained from the coccoliths contained two types of monobasic acid groups in a total amount of 1.8 μmol/mg polysaccharide. One type consisted of weakly acid groups which were identified as uronic acids. The nature of the stronger acid groups remains to be established. The ratio between the respective groups was 1:0.8. Studies with; 45 Ca 2+ demonstrated that the isolated polysaccharide is capable of binding Ca 2+ . Equilibrium dialysis revealed that the maximum amount of Ca 2+ which can be bound is 0.92 ± 0.05ltmol/mg polysaccharide. Flow‐rate dialysis experiments strongly suggested the presence of two classes of Ca 2+ ‐binding sites differing in affinity for Ca 2+ . High‐affinity sites (dissociation constant Kd for Ca 2+ : 2.2 + 1.0 × 10 −5 M) were found to be present in amounts (0.38 ± 0.04 μmol/ mg polysaccharide) approximately equivalent to the strongly acid monovalent groups mentioned above (0.8 μmol/mg polysaccharide). Low‐affinity sites (K d for Ca 2+ : 111 ± 39 × 10 −s M) were estimated at 0.74 ± 0.11 μmol/ mg polysaccharide. Although this figure could be determined less accurately, it is suggested that the uronic acids (1.0 μmol/mg polysaccharide) are identical to the low‐affinity sites. Preferential binding of Ca 2+ occurred in a 100‐fold excess of Na + and Mg 2+ as was shown by gel filtration. A 100‐fold excess ofSr 2+ inhibited Ca 2+ binding to a great extent while no Ca 2+ was bound in the presence of an equimolar amount of La 3+ . The dissociation constants of the high‐affinity sites for Na + , Mg 2+ , Sr 2+ and La 3+ (in the presence of Ca 2+ ) were determined with the flow‐rate dialysis technique. They confirm the order of binding preference found with gel filtration. A polysaccharide with similar properties could be isolated from subfossil coccoliths of E. huxieyi (about 1000 years old). The possible role of the polysaccharide as a heterogeneous matrix in coccolith formation is discussed.