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An alginate lyase from Alteromonas sp. H-4 was used for the preparation of protoplast from young sporophytes of Laminaria japonica. Optimum conditions for the isolation of protoplasts were as follows: a) fine pieces of the sporophytes chopped by a razor were incubated in an enzyme mixture for 3 to 5h at 15°C with shaking, b) the composition of the enzyme mixture was purified alginate lyase (30U/ml) with cellulase Onozuka R-10 (1.5%) in a hypertonic solution (0.7M mannitol, 25mM MgCl2, and 5mM HEPES in 50% seawater, pH7.8). Yields of protoplast were 106-107 cells/g (sporophytes) and protoplast viability was more than 75%. Protoplasts were brown-greenish and spherical with a diameter of 10-25μm. The purity of alginate lyase from the strain H-4 scarcely influenced the yield of protoplast, but purified enzyme produced the highest viability of cells

Application of an Alginate Lyase from Alteromonas sp. for Isolation of Protoplasts from a Brown Algae Laminaria japonica.