Lysozyme Adsorption onto Different Supports: A Comparative Study
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Lysozyme Adsorption onto Different Supports: A Comparative Study ´ KATIA CEC´ILIA DE SOUZA FIGUEIREDO, VERA MARIA MARTINS SALIM, TITO L´IVIO MOITINHO ALVES AND JOSE´ CARLOS PINTO∗ Programa de Engenharia Qu´ımica/COPPE, Universidade Federal do Rio de Janeiro, Cidade Universit´aria, CP 68502, Rio de Janeiro, 21945-970 RJ, Brasil [email protected]
Received July 21, 2004; Revised June 2, 2005; Accepted August 2, 2005
Abstract. The interaction between proteins and solid surfaces has been investigated. The aim of this work is to compare three different materials (hydroxyapatite, polystyrene with core-shell structure (PE-CS) and a functionalized styrene divinylbenzene copolymer) to be used as adsorbents for lysozyme, known as a “hard” protein. Tests were performed according to an experimental design in order to compare the effects of pH, lysozyme and phosphate buffer concentration onto the adsorbed amount of protein. A 23 factorial design and a cross design, which was performed in triplicate, were used to distinguish the most important variables and to infer about the interaction between them. Hydroxyapatite showed the best performance—higher adsorbed amount of lysozyme and smaller dispersion (72.2 ± 0.9 mg/g). However, PE-CS can be regarded as a promising support as high amounts of lysozyme are adsorbed onto this material with relatively small dispersion. Keywords: adsorption, lysozyme, hydroxyapatite, polystyrene, amberlite
1.
Introduction
Biomolecules separation has been intensively studied over the last decades (Barroug et al., 1989; Kandori et al., 1997; Kawasaki et al., 2003). This is partially due to the high importance of biomolecules for medical, pharmaceutical, chemical, dentistry and other industrial purposes. Liquid chromatography is one of the classical methods used to separate proteins. So, the interaction between proteins and solid surfaces is the basis of this process. Proteins are long chains of amino acids. Depending on the nature of the amino acids, protein charge in solutions can be positive or negative due to the amino or carboxyl groups, respectively. Lysozyme is a globular protein and is known for its antibiotic activity. It can be found in the body fluids and hen egg-white. Lysozyme is a basic protein since its isoelectric point ∗ To
whom all correspondence should be addressed.
is 11.1. It also displays a relatively large hydrophobic patch on its surface and despite its recognized hardness on hydrophilic surfaces, it remains unclear whether lysozyme compromises its structural integrity upon adsorption onto hydrophobic surfaces (Weaver and Carta, 1996). Many materials have been investigated to be used as support in chromatographic columns as ceramics (Kandori et al., 1997), polymers (Skidmore et al., 1990) and composites (Weaver and Carta, 1996). Hydroxyapatite (Hap) is one of the most important adsorbents designed for proteins separation. Besides, it is the major mineral component of bones and teeth and this similarity with hard tissues can elucidate the mechanism of adsorption of some p
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