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Luuk M. van Langen

Other affiliations: Moscow State University
Bio: Luuk M. van Langen is an academic researcher from Delft University of Technology. The author has contributed to research in topics: Penicillin amidase & Acylation. The author has an hindex of 19, co-authored 24 publications receiving 1577 citations. Previous affiliations of Luuk M. van Langen include Moscow State University.

Papers
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TL;DR: The pros and cons of carrier-free versus carrier-bound immobilised enzymes and of each type of carriers-free enzyme are discussed.

547 citations

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TL;DR: It is proposed that macromolecular cross‐linkers are too large to penetrate the protein active site and react with catalytically essential amino acid residues.
Abstract: Cross-linked enzyme aggregates (CLEAs) were prepared from several enzymes (penicillin G acylase, hydroxynitrile lyase, alcohol dehydrogenase, and two different nitrilases) by precipitation and subsequent cross-linking using dextran polyaldehyde. In most cases, higher immobilization yields were obtained using the latter cross-linker as compared with the commonly used glutaraldehyde. Active site titration of penicillin acylase CLEAs showed that the higher activity originated from a significantly lower loss in active sites using dextran polyaldehyde as a cross-linking agent. It is proposed that macromolecular cross-linkers are too large to penetrate the protein active site and react with catalytically essential amino acid residues.

273 citations

Journal ArticleDOI
TL;DR: The (R)-oxynitrilase from almonds was immobilized as a cross-linked enzyme aggregate (CLEA) via precipitation with 1,2-dimethoxyethane and subsequent cross-linking using glutaraldehyde and the beneficial effect of these latter conditions on the hydrocyanation of slow-reacting aldehydes is demonstrated.

85 citations

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TL;DR: Penicillin G acylase from Escherichia coli was immobilized on Eupergit C with different enzyme loading and demonstrated that the immobilized enzyme molecules on average had turnover rates much lower than that of the dissolved enzyme.
Abstract: Penicillin G acylase from Escherichia coli was immobilized on Eupergit® C with different enzyme loading. The activity of the immobilized preparations was assayed in the hydrolysis of penicillin G and was found to be much lower than would be expected on the basis of the residual enzyme activity in the immobilization supernatant. Active-site titration demonstrated that the immobilized enzyme molecules on average had turnover rates much lower than that of the dissolved enzyme. This was attributed to diffusion limitations of substrate and product inhibition. Indeed, when the immobilized preparations were crushed, the activity increased from 587 U g -1 to up to 974 U g -1 . The immobilized preparations exhibited up to 15% lower turnover rates than the dissolved enzyme in cephalexin synthesis from 7-ADCA and D-(-)-phenylglycine amide. The synthesis over hydrolysis ratios of the immobilized preparations were also much lower than that of the dissolved enzyme. This was partly due to diffusion limitations but also to an intrinsic property of the immobilized enzyme because the synthesis over hydrolysis ratio of the crushed preparations was much lower than that of the dissolved enzyme.

70 citations

Journal ArticleDOI
TL;DR: The asymmetric synthesis of (R)-o-chloromandelic acid, a key intermediate for the anti-thrombotic agent clopidogrel, via the almond meal catalyzed hydrocyanation of 2-chlorobenzaldehyde and subsequent acidic hydrolysis was developed into an industrially viable procedure as mentioned in this paper.

62 citations


Cited by
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Journal ArticleDOI
TL;DR: An overview of the why, what and how of enzyme immobilisation for use in biocatalysis is presented and 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).
Abstract: 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).

2,013 citations

Journal ArticleDOI
TL;DR: Different methods for the immobilization of enzymes are critically reviewed, with emphasis 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).
Abstract: 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).

1,857 citations

Journal ArticleDOI
TL;DR: A review of the state of the art in the use of alternative reaction media for green, sustainable organic synthesis is presented in this article, where a novel and effective method for the immobilisation of enzymes as cross-linked enzyme aggregates (CLEAs) is discussed and a combi CLEA, containing two enzymes, for the one-pot conversion of benzaldehyde to S-mandelic acid is reported.

1,392 citations

Journal ArticleDOI
TL;DR: The advantages and disadvantages of the different existing immobilization strategies to solve the different aforementioned enzyme limitations are given and some advice to select the optimal strategy for each particular enzyme and process is given.
Abstract: Enzyme biocatalysis plays a very relevant role in the development of many chemical industries, e.g., energy, food or fine chemistry. To achieve this goal, enzyme immobilization is a usual pre-requisite as a solution to get reusable biocatalysts and thus decrease the price of this relatively expensive compound. However, a proper immobilization technique may permit far more than to get a reusable enzyme; it may be used to improve enzyme performance by improving some enzyme limitations: enzyme purity, stability (including the possibility of enzyme reactivation), activity, specificity, selectivity, or inhibitions. Among the diverse immobilization techniques, the use of pre-existing supports to immobilize enzymes (via covalent or physical coupling) and the immobilization without supports [enzyme crosslinked aggregates (CLEAs) or crystals (CLECs)] are the most used or promising ones. This paper intends to give the advantages and disadvantages of the different existing immobilization strategies to solve the different aforementioned enzyme limitations. Moreover, the use of nanoparticles as immobilization supports is achieving an increasing importance, as the nanoparticles versatility increases and becomes more accessible to the researchers. We will also discuss here some of the advantages and drawbacks of these non porous supports compared to conventional porous supports. Although there are no universal optimal solutions for all cases, we will try to give some advice to select the optimal strategy for each particular enzyme and process, considering the enzyme properties, nature of the process and of the substrate. In some occasions the selection will be compulsory, for example due to the nature of the substrate. In other cases the optimal biocatalyst may depend on the company requirements (e.g., volumetric activity, enzyme stability, etc).

1,378 citations

Journal ArticleDOI
TL;DR: This tutorial review focuses on the understanding of enzyme immobilisation, which can address the issue of enzymatic instability.
Abstract: 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.

1,115 citations