Cellulase Immobilization on Poly(methyl Methacrylate) Nanoparticles by Miniemulsion Polymerization

Abstract

Cellulases are efficient enzymes for the conversion of cellulose into glucose. Their use in immobilized form enables them to be reused in successive cycles in many biotechnological processes. Unlike conventional methods of immobilization by covalent bonding, in miniemulsion polymerization the immobilization of enzyme and the synthesis of polymer nanoparticles (support) occur simultaneously. Based on these aspects, the immobilization of cellulose on poly(methyl methacrylate) (PMMA) nanoparticles by miniemulsion polymerization was studied. The surfactant type (non-ionic and ionic) and latex pH showed great influence on cellulase activity. High activity values were obtained only when non-ionic surfactant (Lutensol AT50) and buffering agent (NaHCO 3 ) were used simultaneously. MMA polymerization rate and final monomer conversion were not affected by the presence of cellulase. The maximum immobilization efficiency (60%) was obtained when 6 wt.% of cellulase was used and stable PMMA nanoparticles (133 nm) were obtained. The relative activity profile of immobilized cellulase, for pH as well as temperature, was similar to that reported for the free form. Immobilized enzyme keeps its activity throughout seven days when stored at 4 ºC and phosphate buffer pH 6.0. Based on the results obtained in this work, miniemulsion polymerization as a method for cellulase immobilization on PMMA nanoparticles showed to be a promising technique with high possibility of industrial application.

Enzyme immobilization: polymer–solvent–enzyme compatibility

“…98 Occasionally, physical enzyme adsorption is reported as part of a co-immobilization mode. [99][100][101][102] For example, CALB was immobilized on magnetic poly(urea-urethane) nanoparticles (MNPs-PUU) by single-step mini-emulsion polymerization using diisocyanate and 1,6-hexanediol monomers. 103 Diisocyanate monomer and organic coated MNPs were in the dispersed phase (organic), and free enzyme and polyol monomer were in the continuous phase (aqueous).…”

“…Enzymes are co-immobilized in/on the synthesized polymer by single-step mini-emulsion polymerization. [99][100][101][102][103]300 For instance, Candida antarctica lipase B (CALB) was immobilized using PMMA nanoparticles by mini-emulsion polymerization. 102 The organic phase comprised methyl methacrylate monomers, Crodamol (a fully saturated triglyceride), and an soluble initiator (2,2′azobisisobutyronitrile (AIBN)).…”

Immobilization of Aspergillus Niger cellulase onto Lifetech TM carriers and its application in the hydrolysis of sunflower seed meal lignocellulosic fraction

“…Related immobilization yield was constant within entire examined rangearound 80%. For example, when poly(methyl methacrylate) nanoparticles obtained by miniemulsion polymerization method were used for cellulase immobilization, maximum of 59.1% immobilization yield was achieved (Simon, Lima et al 2018), while by applying SiO 2 /alginate gel biocomposites as carriers for cellulase immobilization, 86% enzyme loading was accomplished (Lin, Chen et al 2008). The most descriptive output regarding catalytic efficacy of enzyme molecules attached to the carrier is specific activity (IU/g of bound proteins) of immobilized enzyme which revealed that, al-though carrier with larger pores was used, there is possibility that catalyst molecules immobilized inside the pores within deeper layers of the carrier particles (usually filled at higher protein concentrations) are more difficult for substrate molecules to access.…”

Hydrolysis of sunflower seed meal lignocellulosic fraction by free and immobilized cellulases

“…A convenient method for overcoming these issues is immobilization on adequate solid supports, providing increased operational and storage stability and reusability (Vaz et al, 2016). During the last decades, various microbial cellulases were immobilized onto different nanoparticles (Xu et al, 2011;Gokhale et al, 2013;Lima et al, 2017;Han et al, 2018;Simon et al, 2018), magnetic chitosan microspheres (Miao et al, 2016), kaolin (Lima et al, 2019), etc. However, most of these immobilized preparations are not applicable for the saccharification of natural lignocellulosic substrates due to their high prices.…”