Liquid purification or separation – Processes – Liquid/liquid solvent or colloidal extraction or diffusing...
Patent
1996-01-16
1997-06-24
Therkorn, Ernest G.
Liquid purification or separation
Processes
Liquid/liquid solvent or colloidal extraction or diffusing...
210656, 2101982, 2105021, B01D 1508
Patent
active
056414030
DESCRIPTION:
BRIEF SUMMARY
The invention relates to separating materials for hydrophobic chromatography.
BACKGROUND OF THE INVENTION
Two groups of separating materials are customary for liquid chromatography which is based on hydrophobic interactions: (RP-chromatography), which have a high ligand density, are mainly used for separating low molecular weight substances in non-aqueous eluents. which have lower ligand densities, and which in general have ligands of lower hydrophobicity (e.g. C.sub.4 -alkyl or C.sub.7 -aryl, instead of c.sub.8 -alkyl or C.sub.18 -alkyl as in RP chromatography), are mainly employed for separating proteins using aqueous eluents. A chaotropic salt, for example 2M ammonium sulfate, is added to the starting eluent; the salt concentration is decreased during the course of the gradient.
Separating materials for hydrophobic interaction chromatography are known in the state of the art. Polysaccharides, which can also be cross-linked, and also cross-linked poly(methyl)acrylates, are customarily used as basal supports. In each case, the basal supports are substituted by lower alkyl or aryl radicals, for example by butyl, phenyl or benzyl. A known process for preparing these materials is based on reacting a basal support which contains oxirane groups with an alcohol in the presence of boron trifluoride etherate.
An inadequate separating effect is frequently observed when using separating materials which are known from the state of the art. Additional, frequently observed, disadvantages of these materials are diminution in the biological activity and/or losses due to non-specific irreversible binding of the analyte.
SUMMARY OF THE INVENTION
The object of the present invention is to provide separating materials having improved properties, in particular for hydrophobic interaction chromatography.
Application DE 43 10 964 discloses oxirane-containing activated support materials in which monomers of the formula I are grafted onto a hydroxyl group-containing basal support, ##STR2## in which R.sup.1, R.sup.2 and R.sup.3 are, independently of each other, H or CH.sub.3,
It has been found that these activated support materials can be converted, in a manner known per se, into separating materials for hydrophobic interaction chromatography. The resulting separating materials exhibit improved properties.
The invention consequently relates to separating materials for hydrophobic chromatography based on hydroxyl group-containing basal supports onto whose surfaces polymers are covalently bonded, characterized in that a terminal monomer unit, ##STR3## in which R.sup.1, R.sup.2 and R.sup.3 are, independently of each other, H or CH.sub.3, -C.sub.25 -alkylaryl or C.sub.7 -C.sub.25 -arylalkyl which can also be derivatized, once or more than once, with nitrile or C.sub.1 -C.sub.5 -alkoxy, where one or more non-adjacent CH.sub.2 groups can also be replaced by NH or O or else one or more CH groups can be replaced by N, or
The invention relates to the use of the novel separating materials for hydrophobic interaction chromatography in association with the separation of mixtures of at least two substances, in particular for separating proteins.
The invention relates to processes for preparing separating materials for hydrophobic interaction chromatography, characterized in that oxirane-containing, activated support materials, which are disclosed in DE 43 10 964, are reacted with alcohols or phenols of the formula III alcohols is preferably carried out in the presence of boron trifluoride etherate.
The invention relates to processes for the separation of mixtures of at least two substances, in particular for separating proteins, by means of hydrophobic interaction chromatography using the novel separating materials.
BRIEF DESCRIPTION OF THE DRAWINGS
FIG. 1a depicts separation of the proteins cytochrome C, myoglobin, lysozyme and chymotrypsinogen on a novel separating material; for comparison, FIG. 1b shows separation of the same mixture on a material which is known from the state of the art.
DESCRIPTION OF THE PREFERRED EMBODIMENTS
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Derwent Abstract of Japan Patent 62 267 663 vol. 12 No. 147 (P-698) May 1988.
Merck Patent Gesellschaft mit beschrankter Haftung
Therkorn Ernest G.
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