In this tutorial review, an overview of the why, what and how of enzyme immobilisation for use in biocatalysis is presented. The importance of biocatalysis in the context of green and sustainable chemicals manufacture is discussed and the necessity for immobilisation of enzymes as a key enabling technology for practical and commercial viability is emphasised. The underlying reasons for immobilisation are the need to improve the stability and recyclability of the biocatalyst compared to the free enzyme. The lower risk of product contamination with enzyme residues and low or no allergenicity are further advantages of immobilised enzymes. Methods for immobilisation are divided into three categories: adsorption on a carrier (support), encapsulation in a carrier, and cross-linking (carrier-free). General considerations regarding immobilisation, regardless of the method used, are immobilisation yield, immobilisation efficiency, activity recovery, enzyme loading (wt% in the biocatalyst) and the physical properties, e.g. particle size and density, hydrophobicity and mechanical robustness of the immobilisate, i.e. the immobilised enzyme as a whole (enzyme + support). The choice of immobilisate is also strongly dependent on the reactor configuration used, e.g. stirred tank, fixed bed, fluidised bed, and the mode of downstream processing. Emphasis is placed on relatively recent developments, such as the use of novel supports such as mesoporous silicas, hydrogels, and smart polymers, and cross-linked enzyme aggregates (CLEAs).

/pdf/enzyme-immobilisation-in-biocatalysis-why-what-and-how-2xy4zx4fuj.pdf

Enzyme immobilisation in biocatalysis : Why, what and how

Immobilization is often the key to optimizing the operational performance of an enzyme in industrial processes, particularly for use in non-aqueous media. Different methods for the immobilization of enzymes are critically reviewed. The methods are divided into three main categories, viz. (i) binding to a prefabricated support (carrier), (ii) entrapment in organic or inorganic polymer matrices, and (iii) cross-linking of enzyme molecules. Emphasis is placed on relatively recent developments, such as the use of novel supports, e.g., mesoporous silicas, hydrogels, and smart polymers, novel entrapment methods and cross-linked enzyme aggregates (CLEAs).

http://largens.com/biochemlab/peroxidase2.pdf

Enzyme immobilization: The quest for optimum performance

Roger Sheldon (1942) received a PhD in organic chemistry from the University of Leicester (UK) in 1967. This was followed by post-doctoral studies with Prof. Jay Kochi in the U.S. From 1969 to 1980 he was with Shell Research in Amsterdam and from 1980 to 1990 he was R&D Director of DSM Andeno. In 1991 he moved to his present position as Professor of organic chemistry and catalysis at the Delft University of Technology (The Netherlands). His primary research interests are in the application of catalytic methodologies—homogeneous, heterogeneous and enzymatic—in organic synthesis, particularly in relation to fine chemicals production. He developed the concepts of E factors and atom utilization for assessing the environmental impact of chemical processes.

Biocatalysis in ionic liquids

Outlook for cellulase improvement: screening and selection strategies.

Enzymes are versatile catalysts in the laboratory and on an industrial scale. To broaden their applicability in the laboratory and to ensure their (re)use in manufacturing the stability of enzymes can often require improvement. Immobilisation can address the issue of enzymatic instability. Immobilisation can also help to enable the employment of enzymes in different solvents, at extremes of pH and temperature and exceptionally high substrate concentrations. At the same time substrate-specificity, enantioselectivity and reactivity can be modified. However, most often the molecular and physical–chemical bases of these phenomena have not been elucidated yet. This tutorial review focuses on the understanding of enzyme immobilisation.

Understanding enzyme immobilisation

Industrial enzyme applications.

Lipases from Candida antarctica: Unique Biocatalysts from a Unique Origin

The invention relates to substrates which develops colour on oxidation of general formula (I). The invention also relates to the use of said substrates for dyeing, e.g. keratinous fibers, in particular hair, dyeing textiles, a dyeing composition and a method for dyeing keratinous fibers.

Diaminobenzoic acid derivatives as dye precursors

Industrial lipase immobilization

The activity of three different lipases, a glycosidase and a protease in ionic liquids has been studied. Ambient temperature ionic liquids are a new class of solvents that are nonvolatile and nonflammable and thus an interesting alternative to classical organic solvents. Monitoring the synthesis of a simple ester, all lipases were found to exhibit both excellent activity and stability in the non-polar ionic liquid 1-butyl-3-methylimidazohum hexaflurophosphate ([bmin][PF6], 1). Furthermore, β-galactosidase from E. coli and the Subtilisin protease SavinaseTM were both found to exhibit a hydrolytic activity in a 50% aqueous solution of the water-miscible ionic liquid 1-butyl-3-methyhmidazoUum tetra-fluoroborate ([bmin][BF4], 2) comparable to the activity observed in 50% aqueous solutions of ethanol and acetonitrile.

Ole Kirk

Papers

Industrial enzyme applications.

Lipases from Candida antarctica: Unique Biocatalysts from a Unique Origin

Diaminobenzoic acid derivatives as dye precursors

Industrial lipase immobilization

Enzyme Catalysed Synthesis in Ambient Temperature Ionic Liquids