Preparation and evaluation of dual-enzyme microreactor with co-immobilized trypsin and chymotrypsin

Meller, Kinga; Pomastowski, Paweł; Grzywiński, Damian; Szumski, Michał; Buszewski, Bogusław

doi:10.1016/j.chroma.2016.02.070

Cited by 42 publications

(26 citation statements)

References 43 publications

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“…However, most open‐tubular CE‐integrated IMERs are limited to the immobilization of single enzyme, which significantly confines their broad practical application. In the past two decades, there have been several attempts to develop the CE‐integrated IMERs for multienzyme immobilization . Yang et al.…”

Section: Introductionmentioning

confidence: 99%

Controllable and high‐performance immobilized enzyme reactor: DNA‐directed immobilization of multienzyme in polyamidoamine dendrimer‐functionalized capillaries

Shen

Zhou

et al. 2020

Electrophoresis

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In recent years, CE‐integrated immobilized enzyme reactors (IMERs) for single‐enzyme immobilization have attracted considerable attention. However, there has been little research on multienzyme immobilization in CE. Here, we introduce a method for fabricating a CE‐integrated IMER, using DNA‐directed immobilization to fix glucose oxidase and horseradish peroxidase in the capillary, which had been functionalized with polyamidoamine dendrimer (PAMAM). Owing to the reversibility of DNA hybridization, the reactor is capable of dynamic immobilization. Moreover, by introducing the PAMAM, the loading capacity of the IMER is greatly enhanced, and the PAMAM can spontaneously form complexes with DNA and then contribute to the efficiency and stability of the reactor. After 25 days storage, the prepared IMER ultimately retained approximately 70% of its initial activity. We also used the IMER to detect glucose, and the favorable linearity was obtained over the concentration range of 0.78–12.5 mM, with an LOD of 0.39 mM, demonstrating that the CE‐integrated IMER can be applied to actual samples. We believe that this strategy can be extended to other multienzyme immobilization systems, and CE‐integrated IMERs are potentially useful in a wide range of biochemical research applications.

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Section: Introductionmentioning

confidence: 99%

Controllable and high‐performance immobilized enzyme reactor: DNA‐directed immobilization of multienzyme in polyamidoamine dendrimer‐functionalized capillaries

Shen

Zhou

et al. 2020

Electrophoresis

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“…Within this particular approach for enzyme immobilization, glutaraldehyde is one of the most disseminated cross-linking agents to bind the enzyme to the support, mostly through the formation of both Schiff bases and Michael-type additions [24,25]. Accordingly, it has also been used when enzymatic microreactors are addressed, in monolith [26], packed bed [13,20,27] and wall-coated configurations [16,17,21]. The developed microreactor assembly methodology proved to be highly reproducible, which led to the establishment of optimal operational conditions under varying conditions of pH and temperature.…”

Section: Introductionmentioning

confidence: 99%

Sucrose Hydrolysis in a Bespoke Capillary Wall-Coated Microreactor

2017

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Abstract:Microscale technology has been increasingly used in chemical synthesis up to production scale, but in biocatalysis the implementation has been proceeding at a slower pace. In this work, the design of a low cost and versatile continuous flow enzyme microreactor is described that illustrates the potential of microfluidic reactors for both the development and characterization of biocatalytic processes. The core structure of the developed reactor consists of an array of capillaries with 450 µm of inner diameter with their inner surface functionalized with (3-aminopropyl)triethoxysilane (APTES) and glutaraldehyde where Saccharomyces cerevisiae invertase was covalently bound. The production of invert sugar syrup through enzymatic sucrose hydrolysis was used as model system. Once the microreactor assembly reproducibility and the immobilized enzyme behavior were established, the evaluation of the immobilized enzyme kinetic parameters was carried out at flow rates ranging from 20.8 to 219.0 µL·min −1 and substrate concentrations within 2.0%-10.0% (w/v). Despite the impact of immobilization on the kinetic parameters, viz. K m(app) was increased two fold and K cat showed a 14-fold decrease when compared to solution phase invertase, the immobilization proved highly robust. For a mean residence time of 48.8 min, full conversion of 5.0% (w/v) sucrose was observed over 20 days.

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“…Furthermore, enzymes are specific to the substrate and produce protein hydrolysates which have better quality chemical and nutritional characteristics (Castro et al, 2011;Tavano, 2013). It is important to notice that the application of proteolytic enzymes for protein digestion is not limited only to the qualitative analysis in proteomic research involving the use of one, highly specific enzyme (Balcão & Vila, 2015;Meller et al, 2016).…”

Section: Introductionmentioning

confidence: 99%

Improvement of the stability and activity of immobilized trypsin on modified Fe3O4 magnetic nanoparticles for hydrolysis of bovine serum albumin and its application in the bovine milk

2016

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Preparation and evaluation of dual-enzyme microreactor with co-immobilized trypsin and chymotrypsin

Cited by 42 publications

References 43 publications

Controllable and high‐performance immobilized enzyme reactor: DNA‐directed immobilization of multienzyme in polyamidoamine dendrimer‐functionalized capillaries

Controllable and high‐performance immobilized enzyme reactor: DNA‐directed immobilization of multienzyme in polyamidoamine dendrimer‐functionalized capillaries

Sucrose Hydrolysis in a Bespoke Capillary Wall-Coated Microreactor

Improvement of the stability and activity of immobilized trypsin on modified Fe3O4 magnetic nanoparticles for hydrolysis of bovine serum albumin and its application in the bovine milk

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