DEVELOPMENT OF A WATER-SOLUBLE, SULFATED(1-]3)-BETA-D-GLUCAN BIOLOGICAL RESPONSE MODIFIER DERIVED FROM SACCHAROMYCES-CEREVISIAE

This report describes a method for the solubilization of micro-particulate (1 --> 3)-beta-D-glucan. Insoluble glucan is dissolved in methyl sulfoxide and urea (8 M) and partially sulfated at 100-degrees. The resulting water-soluble product is called glucan sulfate. The conversion rate is 98%, and the preparation is endotoxin free as determined by the Limulus lysate procedure. Glucan sulfate is composed of 34.06% C, 6.15% H, 50.30% O, 5.69% S and 3.23% N, and has a repeating unit empirical formula of (C6H10O5)8.3 SO3NH4+.4 H2O, suggesting that, on the average, a sulfate group is substituted on every third glucose subunit along the polymer. Molecular weight averages, polydispersity, and intrinsic viscosity were determined by aqueous high-performance size-exclusion chromatography (HPSEC). Two polymer peaks were resolved. Peak 1 (M(w) = 1.25 x 10(6) g/mol) represents < 1% of the total polymer mass. Peak 2 (M(w) = 1.45 x 10(4) g/mol) comprises > 99% of polymers. C-13 NMR spectroscopy con firmed the beta-(1 --> 3) interchain linkage. In solution, glucan sulfate polymers self-associate in a triple helix. Glucan sulfate stimulates murine bone marrow proliferation following intravenous administration. The ability to prepare a immunologically active, water-soluble (1 --> 3)-beta-D-glucan preparation will greatly enhance the clinical utility of this class of compounds.