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Catalytic activity of trypsin entrapped in electrospun poly(ϵ-caprolactone) nanofibers

Paper ID Volume ID Publish Year Pages File Format Full-Text
16854 42618 2015 11 PDF Available
Title
Catalytic activity of trypsin entrapped in electrospun poly(ϵ-caprolactone) nanofibers
Abstract

•Trypsin encapsulation into PCL nanofibers was achieved by emulsion electrospinning.•The optimized catalytic system showed 66% activity retention.•Immobilization enhanced storage and thermal stability compared to the free enzyme.•After 5 cycles of reuse, 59% of the activity was retained.•The proposed mechanism relies on the preferential enzyme location near the fiber surface.

Trypsin was successfully entrapped in situ into nanofibers of poly(ϵ-caprolactone) (PCL) prepared by electrospinning. The spinning dope was an emulsion consisting of an aqueous phase with the solubilized enzyme in a pH buffer plus an oil phase of the polymer solubilized in chloroform (CF)/dimethylformamide (DMF). The optimized materials were composed by random arrays of bead-free fibers with outer diameters in the range 110–180 nm without showing core–shell structure. The fiber size and morphology, membrane porosity and surface properties were shown to be influenced by the polymer concentration and the composition ratio of the solvent mixture, and also by the presence of the enzyme. The activity of the immobilized trypsin was studied toward both a low-molecular weight synthetic substrate (BAPNA) and a protein (casein). Fluorescence microscopy, the increasing hydrophilicity of the fibrous membrane and the observed catalytic activity confirmed the entrapment of the enzyme into the PCL nanofibers. The best activity retention (∼66% toward BAPNA) was achieved using 0.20 g/mL PCL in CF/DMF [75:25], with trypsin in an aqueous buffer at pH 7.1 in the presence of benzamidine and Span80. The immobilized enzyme showed satisfactory operational stability retaining ∼59% of its initial activity after five reaction cycles. Compared with the free enzyme, the storage (at 4 °C) and thermal stability of the immobilized enzyme were highly improved. The retained catalytic activity and the observed reusability can be explained by a heterogeneous distribution of the enzyme within the polymer fiber influenced by the electrostatic field during the electrospinning process, enabling a preferential location near the fiber surface but simultaneously assuring minimal leaching out during operations. Results suggest that trypsin-PCL fibrous membranes may be useful for concomitant proteolytic and separation commercial applications.

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Keywords
EFMs, electrospun fibrous membranes; tryp-PCL-EFM, PCL EFM with entrapped trypsin; fitc-tryp-PCL-EFM, labeled PCL EFM obtained by electrospinning FITC labeled trypsin (fitc-tryp); BAPNA, Nα-benzoyl-dl-arginine-p-nitroanilide; CF, chloroform; DMF, dimethyl
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Catalytic activity of trypsin entrapped in electrospun poly(ϵ-caprolactone) nanofibers
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Publisher
Database: Elsevier - ScienceDirect
Journal: Enzyme and Microbial Technology - Volumes 79–80, November 2015, Pages 8–18
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us