Abstract
Although enzymatic microbioreactors have recently gained lots of attention, reports on the use of whole cells as biocatalysts in microreactors have been rather modest. In this work, an efficient microreactor with permeabilized Saccharomyces cerevisiae cells was developed and used for continuous biotransformation of fumaric into industrially relevant L-malic acid. The immobilization of yeast cells was achieved by entrapment in a porous structure of various hydrogels. Copolymers based on different ratios of sodium alginate (SA) and polyvinyl alcohol (PVA) were used for hydrogel formation, while calcium chloride and boric or phenylboronic acid were tested as crosslinking agents for SA and PVA, respectively. The influence of hydrogel composition on physico-chemical properties of hydrogels prepared in the form of thin films was evaluated. Immobilization of permeabilized S. cerevisiae cells in the selected copolymeric hydrogel resulted in up to 72% retained fumarase activity. The continuous biotransformation process using two layers of hydrogels integrated into a two-plate microreactor revealed high space time yield of 2.86 g/(L·h) while no activity loss was recorded during 7 days of continuous operation.
Highlights
Microreactor technology has recently been attracting increasing attention due to many advantages such as better heat and mass transfer, as well as more efficient mixing and overall better process control [1,2]
Various techniques have been developed for biocatalyst immobilization, including adsorption or covalent linking to insoluble materials, entrapment in polymeric hydrogels and encapsulation in membranes [10,11,12,13]
In the case of biocatalyst immobilization, the suspension of permeabilized S. cerevisiae cells was added at this stage to yield final cell concentration of 1.64 × 108 cells/mL, followed by a thorough mixing
Summary
Microreactor technology has recently been attracting increasing attention due to many advantages such as better heat and mass transfer, as well as more efficient mixing and overall better process control [1,2]. Immobilization of enzymes or whole cells has been an essential part of many biocatalytic process developments since it enables the reusability of the often-expensive biocatalyst and, thereby, prolongs its lifetime. There are many reports on enzyme entrapment inside porous structures of alginate and PVA beads, but very few have tried to combine the polymers to form a hydrogel with better overall properties [22]. Whole cells offer distinct advantages over the isolated enzymes, if complex transformations involving cofactors are considered. Using a whole-cell biocatalyst has drawbacks in higher mass transfer limitations of the substrate and the product through the cell membrane. The aim of this study was to develop a microreactor with whole-cell biocatalyst immobilized in a copolymer hydrogel matrix. Thin hydrogel films with yeast cells were integrated in a two-plate microreactor and used for L-malic acid production. System stability was studied during several days of continuous biotransformation
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