On-line low-volume transesterification-based assay for immobilized lipases

A method has been developed for fast evaluation of transesterification activity of immobilized lipases using microlitre and submicrolitre volumes of substrate solutions. The model reaction (acylation of isopropanol with vinyl acetate) is catalyzed by microbial lipases immobilized on ceramic particle...

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Bibliographic Details
Published in:Journal of biotechnology 2006-12, Vol.126 (4), p.508-518
Main Authors: Urban, Pawel L., Goodall, David M., Bergström, Edmund T., Bruce, Neil C.
Format: Article
Language:English
Subjects:
Acetonitriles - chemistry
Bacillariophyceae
Biological and medical sciences
Bioreactors
Biotechnology
Burkholderia cepacia - enzymology
Candida - enzymology
Candida - genetics
CMOS
Enzymatic microreactor
Enzyme assay
Enzymes, Immobilized - metabolism
Esterification
Fundamental and applied biological sciences. Psychology
Imaging
Immobilized lipase
Lipase - chemistry
Lipase - metabolism
Online Systems
Solubility
Solutions - chemistry
Solvents - chemistry
Substrate Specificity
Transesterification
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Summary:A method has been developed for fast evaluation of transesterification activity of immobilized lipases using microlitre and submicrolitre volumes of substrate solutions. The model reaction (acylation of isopropanol with vinyl acetate) is catalyzed by microbial lipases immobilized on ceramic particles, diatoms or acrylic resin, packed into a Teflon tube connected to fused-silica capillary tubing. The substrate solution is pumped through the microreactor and the product of transesterification, acetaldehyde, quantified on capillary by UV absorbance at 280 nm. Using this system in the continuous-flow mode, comparisons were made of transesterification catalysed by PS-C II lipase in two different solvent mixtures. Acetonitrile was found to be a compatible solvent that can be used as a solubilizer without suppressing enzymatic activity. The method has been used to compare conversions at a fixed flow rate using a single pass of substrate through packed beds containing ∼0.2 mg of supported enzyme. Spatial distributions of the product have been visualized using a complementary metal oxide semiconductor (CMOS) imaging detector in conjunction with the microreactor system operated in the stopped-flow mode.
ISSN:0168-1656
1873-4863
DOI:10.1016/j.jbiotec.2006.05.004