Synthesis and Characterization of a New Cryogel Matrix for Covalent Immobilization of Catalase

Abstract

The advantages of cryogels for enzyme immobilization applications include their mechanical and chemical robustness, ease of production, superior porosity, and low cost. Currently, many researchers are exploring porous material-based systems for enzyme immobilization that are more efficient and economically viable. Here, poly(2-Hydroxyethyl methacrylate-co-allyl glycidyl ether) (p(HEMA-co-AGE)) cryogel matrices were synthesized via the free radical cryopolymerization method to be employed as the support material. For the immobilization of the catalase enzyme onto the p(HEMA-co-AGE) cryogel matrix (catalase@p(HEMA-co-AGE), the best possible reaction conditions were determined by altering parameters such as pH, catalase initial concentration, and flow rate. The maximum catalase immobilization amount onto the p(HEMA-co-AGE) cryogel was found to be 48 mg/g cryogel. To determine the advantages of the cryogel matrix, e.g., the stability and reusability of the cryogel matrix, the adsorption–desorption cycles for the catalase enzyme were repeated five times using the same cryogel matrix. At the end of the reusability tests, it was found that the cryogel was very stable and maintained its adsorption capacity with the recovery ratio of 93.8 ± 1.2%. Therefore, the p(HEMA-co-AGE) cryogel matrix affords repeated useability, e.g., up to five times, without decreasing its catalase binding capacities significantly and has promising potential for many industrial applications. Cryogels offer clear distinctive advantages over common materials, e.g., micro/nano particles, hydrogels, films, and composites for these applications. At present, many researchers are working on the design of more effective and economically feasible, porous material-based systems for enzyme immobilization

Alpha‐Glucosidase Enzyme Immobilized Dextran‐Co Metal Nanoparticle Cryogel Composite Systems as Dual Catalyst with Enhanced Stability and Catalytic Activity

“…The details of the covalent linking of α‐Glu enzyme onto Dex cryogels were given in the experimental section. The epoxy groups as commonly used functional groups for enzyme immobilization were utilized [35,36,43] . Before use, the prepared α‐Glu C @Dex cryogels were also washed with water, dried with a freeze dryer, and stored in closed tubes.…”

“…This could be explained by the fact that upon the immobilization process, the active sites of the α‐Glu enzymes within the cryogel network are protected. As a result, the enzymes within cryogel are more stable at lower or higher temperatures in comparison to free enzymes allowing them to function somewhat more effectively in challenging reaction conditions [27,35,36] …”

“…These features provide cryogels a high surface area without restricting the activities of enzymes each other and higher exchange rates that enzyme-substrate interaction can be easily achieved. [28,[34][35][36] However, since cost reduction is one of the most important considerations in industry, studies were focused on the development of systems that can perform more than one reaction at the same time. [37][38][39][40] Therefore, more recently many studies concentrated on performing sequential enzymatic reactions by immobilizing more than one enzyme at the same time.…”

The specific biopanning of single‐domain antibody against haptens based on a functionalized cryogel

“…Blocking was a critical step in reducing non‐specific binding to avoid false positives 39 . Previous experiments revealed that unblocked AG cryogels have adsorption to proteins, which might lead to adsorption of non‐specific phages 40 . Comparing the phage output of once and double blocking during biopanning by AG cryogels, the experimental results showed that the number of non‐specific phages in double blocking was significantly lower at low wash solution volumes (<100 mL), suggesting that double blocking had a role in reducing the number of sites on cryogels induced to non‐specific adsorption (Figure 2A).…”

“…39 Previous experiments revealed that unblocked AG cryogels have adsorption to proteins, which might lead to adsorption of non-specific phages. 40 Comparing the phage output of once and double blocking during biopanning by AG cryogels, the experimental results showed that the number of non-specific phages in double blocking was significantly lower at low wash solution volumes (<100 mL), suggesting that double blocking had a role in reducing the number of sites on cryogels induced to non-specific adsorption (Figure 2A). However, it should be noted that the effect of double blocking on reducing non-specific phages was significantly reduced at high wash solution volumes, so a wash solution volume of 30 mL was finally selected in this experiment.…”