Using biocatalysts for chemical manufacturing is a highly developed and applied science, used widely for production and processing of pharmaceuticals and food items. Both enzymes and whole cells can be used to catalyse organic reactions, with the most general example of biocatalysis being the yeast-mediated transformation of sugars into alcohols, acids or gases. However, the most frequently employed group of biocatalysts is lipases. These enzymes are very versatile, facilitating the hydrolysis of lipids into fatty acids as well as the synthesis of esters and amides. Areas of lipase-catalysed reactions include food processing, synthesis of fine chemicals and production of biofuel. Other regularly used enzymatic biocatalysts are proteases, cellulases and amylases, used for the hydrolysis of proteins, cellulose and starch, respectively.
The technique of using enzymes to mediate organic synthesis is widely endorsed due to their high selectivity and specificity, which ensures a more satisfactory yield and purity of the transformed products. Furthermore, by the use of protein engineering, the biocatalysis can be made even more efficient by having the enzyme customized to fit a specific need.
When utilizing isolated enzymes for organic synthesis, a typical approach is to keep the enzymes immobilized on the surface of an insoluble material. This is preferred to using soluble enzymes due to both technical and economic benefits. The immobilization will allow for easier recycling, manipulation and separation of the enzyme, as well as an increase in its thermal and operational stability. Additionally, immobilization of the enzyme will ensure minimized protein contamination during the experimental procedure, and improved control over the overall reaction process.
Rotating bed reactors
A rotating bed reactor (RBR) is designed to prevent disintegration of solid phase particles by retaining them inside a rotating cylinder during the reaction span. This minimizes solid phase debris even at high speeds, which would normally occur due to mechanical forces when using stirred tank reactors. Due to these properties of the RBR, reaction rates can be improved by increasing the speed at which an RBR is rotated, without shortening the lifespan of the solid phase particles. This ensures easy in-line monitoring of the process, as well as a more efficient recycling of the solid phase particles, making the RBR a cost and time efficient alternative to conventional methods for immobilized enzyme biocatalysis.
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Navam S. Hettiarachchy, Danis Jesus Feliz, J. Stricklin Edwards and Ronny Horax, “The use of immobilized enzymes to improve functionality” in Proteins in Food Processing, Second Edition, Rickey Y. Yada (ed.), Woodhead Publishing, 2018.
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Anup Ashok, Santhosh Kumar Devarai. “L-Asparaginase production in rotating bed reactor from Rhizopus microsporus IBBL-2 using immobilized Ca-alginate beads”, 3Biotech, (2019) 9:349.
Time lapse video showing how straightforward it is to use immobilized enzymes in a rotating bed reactor. A substrate giving a yellow coloured product was used to follow the reaction progress of an ester hydrolysis by an immobilized lipase. This substrate is commonly used to screen and characterize lipases.
Conditions: 4-nitrophenyl octanoate (30 mg) in aqueous phosphate buffer (150 mL, 0.1 M, pH 7.4, 0.1% Triton X100) was hydrolysed at 30 °C using Purolite® CalB immoplus™ (150 mg) filled into a SpinChem® rotating bed reactor (RBR) S221 rotating at 500 rpm within a SpinChem® flower-baffled reaction vessel V211. The reaction was monitored for 30 min by filming the formation of the yellow 4-nitrophenoxide product (abs. max. ca 405 nm).
Poster describing the screening of resins with immobilized enzymes for esterification reactions. The screening was performed with prepacked cartridges inside a SpinChem® rotating bed reactor (RBR) and compared to parallel screening with prototype disposable magnetic RBR (MagRBR) in 10 mL volume. Both approaches were very quick, required minimum preparation and no filtration. Up-scaled processes enabled quick enzyme recycling, opening for the possibility of semi-continuous processes with attractive production economy.
Conditions: This poster contains data from several reactions with immobilized enzymes performed at different conditions. The details of each reaction are displayed under each figure in the poster. This poster was presented at BioTrans2017 - the 13th international symposium on biocatalysis and biotransformations, in Budapest, Hungary, the 9th to 13th of July, 2017.
Poster on a case study of applying the rotating bed reactor for the lipase-mediated stereoselective acetylation of a racemate amine as a model reaction for the manufacturing of pharmaceutical building blocks. The results showed that enzyme recycling and synthesis scale up was easy to achieve with preserved yield, enantioselectivity and catalytic activity.
Conditions: This poster contains data from several reactions with immobilized enzymes performed at different conditions. The details of each reaction are displayed under each figure in the poster. This poster was presented at BioTrans2019 - the 14th international symposium on biocatalysis and biotransformations, in Groningen, Netherlands, the 7th to 11th of July, 2019.
Poster on study of catalyst recycling during esterification and transesterification reactions with immobilized lipases in rotating bed reactors. Data from several laboratories showed that no attrition or grinding occurred and that no filtration was necessary between reaction cycles. The production capacity was estimated to 50 kilograms per gram of catalyst thanks to the high catalyst stability.
Conditions: This study comprised several different laboratories, immobilized enzymes, and conditions for performing esterification and transesterification reactions using a SpinChem® rotating bed reactor (RBR). The details of every reaction are displayed in connection to each figure on the poster. This poster was displayed at Biocat2016 - the 8th international congress on biocatalysis, at Hamburg University of Technology, Germany, August 28th to September 1st, 2016.
Comparison of SpinChem® rotating bed reactor (RBR) with traditional reaction set-ups for a demanding biotransformation. SpinChem® RBR matched or outperformed the other systems and gave a 10 to 25-fold more time-efficient recycling of the encapsulated cells.