Purification of IgY using aqueous biphasic systems composed of good’s buffers ionic liquids

Detalhes bibliográficos
Autor(a) principal: Gomes, Joana Margarida Mota
Data de Publicação: 2015
Tipo de documento: Dissertação
Idioma: eng
Título da fonte: Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
Texto Completo: http://hdl.handle.net/10773/15391
Resumo: The increased inefficiency of antibiotics observed in the past few years, derived from the growing number of drug-resistant organisms and the appearance of individuals with impaired immune system, led to a significant research on antibodies for use in passive immunotherapy. Antibodies obtained from the egg yolk of immunized hens, immunoglobulin Y (IgY), are a promising alternative to mammalian antibodies. They can be obtained in higher titres and by less invasive techniques, thus opening the door for a new kind of more economic biopharmaceuticals. However, the available techniques for the IgY extraction and purification are time-consuming, labor intensive, low yielding, provide low-purification levels, are of high cost and cannot easily be scaled-up for industrial applications. In this work, ABS composed of C6H5K3O7 and ILs synthetized by the combination of anions with buffer capacity (Good’s buffers), with the tetrabutylphosphonium ([P4444]+) and tetrabutylammonium ([N4444]+) cations, were investigated. The use of different ILs allowed the study of the cation and anion nature on the phase diagrams behaviour and thus on their ability to form ABS. Further, the ability of these systems as an alternative purification platform for IgY, by a selective extraction, was studied. The complete extraction and purification of IgY was not achieved in a single with the investigated ABS. However, the study of the proteins profile of the coexisting phases allowed to conclude that all proteins migrate for the top phase during the partitioning process, and that a complex between the ILs and the different proteins present in the water-soluble fraction of proteins from egg yolk was formed. Still, the results obtained reveal that the system composed of 20.1 wt % [N4444][CHES] + 16.9 wt % C6H5K3O7 is the most promising ABS for the IgY extraction.
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spelling Purification of IgY using aqueous biphasic systems composed of good’s buffers ionic liquidsBioquímicaImunoglobulinas - Purificação químicaSistemas aquosos bifásicosSoluções iónicasSoluções tampãoExtracção líquido-líquidoImunoterapiaThe increased inefficiency of antibiotics observed in the past few years, derived from the growing number of drug-resistant organisms and the appearance of individuals with impaired immune system, led to a significant research on antibodies for use in passive immunotherapy. Antibodies obtained from the egg yolk of immunized hens, immunoglobulin Y (IgY), are a promising alternative to mammalian antibodies. They can be obtained in higher titres and by less invasive techniques, thus opening the door for a new kind of more economic biopharmaceuticals. However, the available techniques for the IgY extraction and purification are time-consuming, labor intensive, low yielding, provide low-purification levels, are of high cost and cannot easily be scaled-up for industrial applications. In this work, ABS composed of C6H5K3O7 and ILs synthetized by the combination of anions with buffer capacity (Good’s buffers), with the tetrabutylphosphonium ([P4444]+) and tetrabutylammonium ([N4444]+) cations, were investigated. The use of different ILs allowed the study of the cation and anion nature on the phase diagrams behaviour and thus on their ability to form ABS. Further, the ability of these systems as an alternative purification platform for IgY, by a selective extraction, was studied. The complete extraction and purification of IgY was not achieved in a single with the investigated ABS. However, the study of the proteins profile of the coexisting phases allowed to conclude that all proteins migrate for the top phase during the partitioning process, and that a complex between the ILs and the different proteins present in the water-soluble fraction of proteins from egg yolk was formed. Still, the results obtained reveal that the system composed of 20.1 wt % [N4444][CHES] + 16.9 wt % C6H5K3O7 is the most promising ABS for the IgY extraction.Recentemente, a utilização de anticorpos para imunoterapia passiva tornou-se uma área de investigação atrativa. Tal facto deve-se à diminuição da eficiência dos antibióticos pelo aumento do número de organismos resistentes. Os anticorpos obtidos a partir da gema do ovo de galinhas imunizadas, imunoglobulinas Y (IgY), são uma alternativa viável e promissora aos anticorpos de mamíferos. Estes apresentam inúmeras vantagens, dado que podem ser obtidos em maior quantidade e serem recolhidos por técnicas menos invasivas. Apesar das vantagens apresentadas, as técnicas disponíveis para a extração e purificação de IgY são demoradas, trabalhosas, apresentam baixo rendimento, levam a baixos fatores de pureza, apresentam custos elevados e não são facilmente aplicáveis a nível industrial. Assim, o principal objetivo deste trabalho consistiu no desenvolvimento de uma plataforma de purificação alternativa para a extração e purificação de IgY, utilizando sistemas aquosos bifásicos (SAB) constituídos por líquidos iónicos com capacidade tampão. Neste trabalho foram estudados SAB constituídos por C6H5K3O7 e líquidos iónicos, sendo os últimos sintetizados pela combinação de aniões com capacidade tampão (Good’s buffers), com os catiões tetrabutilamónio ([N4444]+) e tetrabutilfosfónio ([P4444]+). Os líquidos iónicos utilizados permitiram o estudo da influência do anião e do catião nos diagramas de fase, ou seja sob a capacidade dos diferentes líquidos iónicos para formar SAB. De seguida, foi testada a capacidade destes sistemas como uma plataforma alternativa para a extração seletiva, e posterior purificação, de IgY. A purificação de IgY não foi conseguida num único passo de extração com os sistemas estudados. No entanto, o estudo do perfil proteico de ambas as fases dos sistemas permitiu concluir que, durante a partição, todas as proteínas migraram para a fase superior e que houve formação de um complexo entre o IL e algumas das proteínas constituintes da fração aquosa proteica da gema do ovo. Os resultados obtidos revelaram que, entre os vários SAB estudados, o mais promissor para a extração de IgY, é aquele constituído por 20.1 (m/m) % [N4444][CHES] + 16.9 (m/m) % C6H5K3O7.Universidade de Aveiro2015-07-302015-07-30T00:00:00Z2020-07-30T14:00:00Zinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/masterThesisapplication/pdfhttp://hdl.handle.net/10773/15391TID:201578905engGomes, Joana Margarida Motainfo:eu-repo/semantics/openAccessreponame:Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)instname:Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informaçãoinstacron:RCAAP2024-02-22T11:28:27Zoai:ria.ua.pt:10773/15391Portal AgregadorONGhttps://www.rcaap.pt/oai/openaireopendoar:71602024-03-20T02:50:45.219708Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) - Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informaçãofalse
dc.title.none.fl_str_mv Purification of IgY using aqueous biphasic systems composed of good’s buffers ionic liquids
title Purification of IgY using aqueous biphasic systems composed of good’s buffers ionic liquids
spellingShingle Purification of IgY using aqueous biphasic systems composed of good’s buffers ionic liquids
Gomes, Joana Margarida Mota
Bioquímica
Imunoglobulinas - Purificação química
Sistemas aquosos bifásicos
Soluções iónicas
Soluções tampão
Extracção líquido-líquido
Imunoterapia
title_short Purification of IgY using aqueous biphasic systems composed of good’s buffers ionic liquids
title_full Purification of IgY using aqueous biphasic systems composed of good’s buffers ionic liquids
title_fullStr Purification of IgY using aqueous biphasic systems composed of good’s buffers ionic liquids
title_full_unstemmed Purification of IgY using aqueous biphasic systems composed of good’s buffers ionic liquids
title_sort Purification of IgY using aqueous biphasic systems composed of good’s buffers ionic liquids
author Gomes, Joana Margarida Mota
author_facet Gomes, Joana Margarida Mota
author_role author
dc.contributor.author.fl_str_mv Gomes, Joana Margarida Mota
dc.subject.por.fl_str_mv Bioquímica
Imunoglobulinas - Purificação química
Sistemas aquosos bifásicos
Soluções iónicas
Soluções tampão
Extracção líquido-líquido
Imunoterapia
topic Bioquímica
Imunoglobulinas - Purificação química
Sistemas aquosos bifásicos
Soluções iónicas
Soluções tampão
Extracção líquido-líquido
Imunoterapia
description The increased inefficiency of antibiotics observed in the past few years, derived from the growing number of drug-resistant organisms and the appearance of individuals with impaired immune system, led to a significant research on antibodies for use in passive immunotherapy. Antibodies obtained from the egg yolk of immunized hens, immunoglobulin Y (IgY), are a promising alternative to mammalian antibodies. They can be obtained in higher titres and by less invasive techniques, thus opening the door for a new kind of more economic biopharmaceuticals. However, the available techniques for the IgY extraction and purification are time-consuming, labor intensive, low yielding, provide low-purification levels, are of high cost and cannot easily be scaled-up for industrial applications. In this work, ABS composed of C6H5K3O7 and ILs synthetized by the combination of anions with buffer capacity (Good’s buffers), with the tetrabutylphosphonium ([P4444]+) and tetrabutylammonium ([N4444]+) cations, were investigated. The use of different ILs allowed the study of the cation and anion nature on the phase diagrams behaviour and thus on their ability to form ABS. Further, the ability of these systems as an alternative purification platform for IgY, by a selective extraction, was studied. The complete extraction and purification of IgY was not achieved in a single with the investigated ABS. However, the study of the proteins profile of the coexisting phases allowed to conclude that all proteins migrate for the top phase during the partitioning process, and that a complex between the ILs and the different proteins present in the water-soluble fraction of proteins from egg yolk was formed. Still, the results obtained reveal that the system composed of 20.1 wt % [N4444][CHES] + 16.9 wt % C6H5K3O7 is the most promising ABS for the IgY extraction.
publishDate 2015
dc.date.none.fl_str_mv 2015-07-30
2015-07-30T00:00:00Z
2020-07-30T14:00:00Z
dc.type.status.fl_str_mv info:eu-repo/semantics/publishedVersion
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dc.identifier.uri.fl_str_mv http://hdl.handle.net/10773/15391
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dc.publisher.none.fl_str_mv Universidade de Aveiro
publisher.none.fl_str_mv Universidade de Aveiro
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