Development of new chromatographic support based on gellan gum for pharmaceutical biomolecules isolation

• Autor(a) principal: Gonçalves, Armanda Isabel Carvalho
• Data de Publicação: 2013
• Tipo de documento: Dissertação
• Idioma: eng
• Título da fonte: Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
• Texto Completo: http://hdl.handle.net/10400.6/3241
• topic: Cromatografia; Polímero de gelan

Higher separation efficiency and resolution have been of great interest in chromatography and have become increasingly important in recent years mainly driven by the challenges of either more complex samples or high sample quantity. Therefore, for the development of new chromatographic matrices is increasingly important to improve the purification efficiency and to decrease the use of resources. Gellan gum is a polysaccharide polymer with natural anionic nature and ability to form thermoreversible gels, seeming to have potential to be used as a chromatographic matrix. In the presence of cations, the gellan polymer suffers conformational transition accompanied by the formation of a three dimensional network, forming a gel. In this work, it was intended to prepare of a stable gellan gum gel to be used as a chromatographic matrix. In order to increase the stability of the gellan gels, different experimental conditions were tested. Experimental design was used to obtain optimal conditions for the gel stability. Due to negative charge of these gels, it was possible to study the interactions established with three model proteins (bovine seric albumin (BSA), α-chymotrypsin and lysozyme). Gellan gum was able to interact with two of these proteins, being able to elute them with an increase in the ionic strength. In this assays a MES buffer with pH 6.2 was utilized. This pH conferred negative charge to BSA and positive charge to α-chymotrypsin and lysozyme, due to their isoelectric points. Assays of dynamic binding capacity were performed to find more characteristics of this new matrix and comparing with commercial resins. The values of dynamic binding capacity of the gellan gum to 10% and 50% breakthrough were 3,9 mg/ml and 17,4 mg/ml, respectively. These values were similar to commercial resins. These results showed that gellan gum might be an innovative and promising chromatographic matrix due to its versatility to interact with different biomolecules and its gelling ability. Thus, the gellan gum gel could have potential to be applied in different scientific domains (purification of complexes cellular extracts or nucleic acids).