色谱2012，Vol.30Issue(11)：1194-1202,9.
Adsorption of endotoxins on Ca2+-iminodiacetic acid by metal ion affinity chromatography
Adsorption of endotoxins on Ca2+-iminodiacetic acid by metal ion affinity chromatography
摘要
Abstract
Endotoxins (also known as lipopolysaccharides (LPS)) are undesirable by-products of recombinant proteins,purified from Escherichia coli.LPS can be considered stable under a wide range of temperature and pH,making their removal one of the most difficult tasks in downstream processes during protein purification.The inherent toxicity of LPS makes their removal an important step for the application of these proteins in several biological assays and for a safe parenteral administration.Immobilized metal affinity chromatography (IMAC) enables the affinity interactions between the metal ions (immobilized on the support through the chelating compound) and the target molecules,thus enabling high-efficiency separation of the target molecules from other components present in a mixture.Affinity chromatography is applied with Ca2+-iminodiacetic acid (IDA)to remove most of the LPS contaminants from the end product (more than 90％).In this study,the adsorption of LPS on an IDA-Ca2+ was investigated.The adsorption Freundlich isotherm of LPS-IDA-Ca2 + provides a theoretical basis for LPS removal.It was found that LPS is bound mainly by interactions between the phosphate group in LPS and Ca2+ ligands on the beads.The factors such as pH (4.0 or 5.5) and ionic strength (1.0 mol/L) are essential to obtain effective removal of LPS for contaminant levels between endotoxin' concentration values less than 100 EU/mL and 100000 EU/mL.This new protocol represents a substantial advantage in time,effort,and production costs.关键词
immobilized metal affinity chromatography (IMAC) / lipopolysaccharides (LPS) / endotoxin removal/ recombinant proteins/ isotherms/ protein purificationKey words
immobilized metal affinity chromatography (IMAC) / lipopolysaccharides (LPS) / endotoxin removal/ recombinant proteins/ isotherms/ protein purification分类
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André Moreni LOPES,Jorge Sánchez ROMEU,Rolando Páez MEIRELES,Gabriel Marquez PERERA,Rolando Perdomo MORALES,Adalberto PESSOA,Lourdes Zumalacárregui CáRDENAS..Adsorption of endotoxins on Ca2+-iminodiacetic acid by metal ion affinity chromatography[J].色谱,2012,30(11):1194-1202,9.基金项目
This research was supported by grants from the Brazilian Agency Coordination of Graduate Level Training (CAPES,project 0366/09-9) and State of S(a)o Paulo Research Support Foundation (FAPESP-Brazil,project 2005/60159-7). （CAPES,project 0366/09-9）
