PORE-SIZE CONTROLLED AND POLY(epsilon-LYSINE)-IMMOBILIZED CELLULOSE SPHERICAL PARTICLES FOR REMOVAL OF LIPOPOLYSACCHARIDES

摘要

Poly(e-lysine) was covalently immobilized onto cellulose spherical particles and used for selective adsorption of pyrogenic lipopolysaccharides (LPS) from protein solutions. The resulting poly(epsilon-lysine)-immobilized cellulose particles (PL-cellulose), which had diameters of 44 to 105 mum and matrix's pore-sizes of 2 x 10~3, 1 x 10~4, and >2 x 10~6 as molecular mass exclusions (M_(lim)), were used as adsorbents. The adsorption of LPS and protein to the adsorbent were determined using a batchwise method. The larger the pore size (M_(lim)) of the adsorbent, the larger is the LPS-adsorbing activity of the adsorbent.The apparent dissociation constant between the LPS (E. coli O111:B4) and the adsorbent decreased from 3.8 x 10~(-10) to l.lxl0~(-11)M with an increase in the M_(lim) from 2 x 103 to >2 x 10~6 at an ionic strength of mu=0.05 and a pH of 7.0. On the other hand, the adsorbing activity of bovine serum albumin also increased with the increasing M_(lim) of the adsorbent, but sharply decreased with increasing ionic strength of the buffer.When PL-cellulose-10~3, having a small M_(lim) of 2 x 10~3, was used as the adsorbent at a wide ionic strength of mu=0.05 to 0.4 and a pH of 7.0, it selectively reduced LPS in various protein solutions, including lmgmL~(-1) of protein and 1500 to 32,000pgmL~(-1) of natural LPS. When PL-cellulose-10~6, having a large M_(lim) of over 2 x 10~6, was used as the adsorbent, it only selectively removed LPS at a high ionic strength of mu=0.4 and a pH of 7.0. The LPS-removing activity of PL-cellulose-10~6 was always stronger than that of PL-cellulose-10~3. PL-cellulose-10~6 decreased the concentration of LPS in each protein solution to less than l0pgrnL~(-1).On the other hand, the recovery rate (99%) of protein for PL-cellulose-103 was higher than that for PL-cellulose-10~6 (96 to 97%) in all cases.