Expanded bed adsorption on supermacroporous cross-linked cellulose matrix.
Rigid spherical macroporous adsorbent beads (CELBEADS) prepared by cross-linking of cellulose were characterized and found eminently suitable for use as expanded bed affinity chromatography matrix. Chromatographic runs were performed on a 10 mm diameter column with three solutes tyrosine, papain and bovine serum albumin under non-retaining conditions on CELBEADS and Streamline DEAE, a commercial agarose based expanded bed matrix. Performance of the runs was measured in terms of height equivalent to theoretical plate, HETP. Variation in HETP with velocity on Streamline DEAE gave flat profiles in packed bed and increasing trend in expanded bed. On CELBEADS, the HETP curves in both packed and expanded bed modes followed profiles typical of macroporous adsorbents i.e. increasing and levelling with velocity. HETP values obtained for papain and bovine serum albumin on CELBEADS were lower than those obtained on Streamline DEAE at all velocities. Lactate dehydrogenase was purified from porcine muscle homogenate using Cibacron blue conjugated to CELBEADS using a protocol reported for supports with surface hydroxyl groups. Elution of the enzyme was investigated both in packed mode as well as in expanded mode at a flow rate of 1 ml min-1. The purification procedure took about 60 minutes and a purification fold of about 14 was achieved in both cases. The adsorbent could be cleaned in place with 5 M urea and used repeatedly without loss of performance.
Pai
A
A
Department of Chemical Technology, University of Mumbai, Matunga, India.
Gondkar
S
S
Sundaram
S
S
Lali
A
A
D000327:Adsorption; D000818:Animals; D002482:Cellulose / Q000737:chemistry*; D003432:Cross-Linking Reagents; D004983:Ethanolamines; D007770:L-Lactate Dehydrogenase / Q000302:isolation & purification; D013552:Swine
D003432:Cross-Linking Reagents; D004983:Ethanolamines; D002482:Cellulose; D007770:L-Lactate Dehydrogenase; C007369:2-diethylaminoethanol