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J Chromatogr A ; 1338: 77-84, 2014 Apr 18.
Artigo em Inglês | MEDLINE | ID: mdl-24630982

RESUMO

The aim of this work was to optimize the preparation of a capillary human purine nucleoside phosphorylase (HsPNP) immobilized enzyme reactor (IMER) for characterization and affinity screening studies of new inhibitors by frontal affinity chromatography coupled to mass spectrometry (FAC-MS). For this purpose two monolithic supports, a Chromolith Speed Rod (0.1mm I.D.×5cm) and a methacrylate-based monolithic epoxy polymeric capillary column (0.25mm I.D.×5cm) with epoxy reactive groups were considered and compared to an IMER previously developed using an open fused silica capillary. Each HsPNP-IMER was characterized in terms of catalytic activity using Inosine as standard substrate. Furthermore, they were also explored for affinity ranking experiments. Kd determination was carried out with the based fused silica HsPNP-IMER and the results are herein discussed.


Assuntos
Cromatografia de Afinidade/métodos , Purina-Núcleosídeo Fosforilase/química , Enzimas Imobilizadas/química , Humanos , Cinética , Espectrometria de Massas , Microscopia Eletrônica de Varredura
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