Abstract
A new strategy to increase the enzyme-loading capacity of porous supports was investigated. Lipase from Pseudomonas fluorescens (PFL) was immobilized on octyl-agarose (OA) beads and treated with polyethyleneimine (PEI). Then, PFL was immobilized on the previous PFL layer. Next, the biocatalyst was coated with PEI and a third layer of PFL was added. Sodium dodecyl sulfate polyacrylamide electrophoresis showed that the amount of PFL proportionally increased with each enzyme layer; however, the effects on biocatalyst activity were not as clear. Hydrolyzing 50 mM of triacetin at 25 °C, the activity of the three-layer biocatalyst was even lower than that of the bi-layer one; on the contrary its activity was higher when the activity was measured at 4 °C in the presence of 30% acetonitrile (that reduced the activity and thus the relevance of the substrate diffusion limitations). That is, the advantage of the multilayer formation depends on the specific activity of the enzyme and on the diffusion limitations of the substrate. When octyl agarose (OA)-PFL-PEI-PFL preparation was treated with glutaraldehyde, the activity was reduced, although the enzyme stability increased and the immobilization of the last PFL layer offered results similar to the one obtained using the three-layer preparation without glutaraldehyde modification (90%).
Highlights
Enzyme immobilization is a key step in the enzyme biocatalyst design [1]
Immobilization of a second layer of Pseudomonas fluorescens (PFL) increased the biocatalyst activity versus triacetin, and in this case, the coating of this second layer of PFL with PEI greatly increased the biocatalyst activity. This difference with the effect of the first PEI coating may be based on the different immobilization strategy in the two enzyme layers: the first one was immobilized via interfacial activation, with the active center oriented towards the rigid support surface, while the second one was immobilized via ion exchange on a polymeric mobile and flexible bed
Addition of the third layer of PFL produced a small increase of biocatalyst activity, lower that the values expected from the second PFL layer, but enough to increase the total volumetric activity of the biocatalyst
Summary
Enzyme immobilization is a key step in the enzyme biocatalyst design [1]. Initially, its purpose was just to facilitate the reuse of these expensive biocatalysts. Price is considered relevant in some instances, but that depends on the degree of enzyme improvement that can be achieved after immobilization and on other features of the support [11]. Their mechanical properties should be compatible with the reactor system. Porous rigid supports like silicates are useful for fixed bed reactors, but they can be broken in stirred systems, while flexible supports are adequate for stirred systems but produce problems in fixed bed reactors [1]. The particle size should be adequate for the filters and catalysts retention equipment
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
