Affinity surfactants as reversibly bound ligands for high-performance affinity chromatography.
Pyridine was coupled covalently to a nonionic ethoxylated alcohol: octaethylene glycol n-hexadecyl ether. This modified surfactant was found to be a reversible, competitive inhibitor of horse serum cholinesterase. The surfactant bound irreversibly, in aqueous media, to octadecyl-bounded reverse phase silica particles commonly used for high-performance liquid chromatography. The amount of ligand bound was found to be 550 mumol/ml of packing, a concentration that is over 100 times higher than what can be normally bound to agarose affinity chromatography supports. With this packing, a 280-fold purification of cholinesterase from horse serum and a 79-fold purification of human serum cholinesterase were accomplished, with yields greater than 80%, using a 2-cm-long column and a 7-min elution time. The affinity surfactant could be eluted from the column using a 6:4 (v/v) mixture of methanol and isopropanol. This technique should be generally applicable in the development of biospecific supports for high-performance affinity chromatography.
Torres
J L
JL
Department of Chemical Engineering, North Carolina State University, Raleigh 27695.
Guzman
R
R
Carbonell
R G
RG
Klipatrick
P K
PK
D000818:Animals; D001665:Binding Sites; D002800:Cholinesterase Inhibitors / Q000097:blood; D002846:Chromatography, Affinity / Q000379:methods*; D002851:Chromatography, High Pressure Liquid; D006736:Horses; D006801:Humans; D011726:Pyridinium Compounds; D013501:Surface-Active Agents / Q000494:pharmacology*
D002800:Cholinesterase Inhibitors; D011726:Pyridinium Compounds; D013501:Surface-Active Agents