STABLE SILICA-BASED ETHER BONDED PHASES FOR BIOPOLYMER SEPARATIONS

摘要

Fast high resolution separations of biopolymers with retention of biological activity have been achieved by hydrophobic interaction chromatography using trialkoxy silyl ethers of the general formula (RO)3Si-(CH2)m-O-(CH2CH2O)n-(CH2)pR' chemically bonded to silica-based chromatographic supports. In the formula R is alkyl of from one to five carbons, m is an integer from two to five, n is an integer from one to five, p is an integer from zero of ten, and R' is methyl, phenyl, or substituted phenyl. Stable and reproducible bonded phases are prepared in a novel solventless procedure by a bonding process which uses a defined and controlled amount of water on the silica surface and a gaseous or volatile basic catalyst such as ammonia to produce a controlled amount of silane polymerization and cross-linking in addition to extensive bonding between silane and silica. High performance liquid chromatography on such weakly hydrophobic stationary phases using aqueous eluents and decreasing salt gradients under mild conditions permits high speed, high resolution separations of biopolymers such as proteins without destruction of their biological activity and without column degradation. Size exclusion chromatography can also be performed on these phases using low ionic strength eluents.