Droplet microfluidics for double lipase immobilisation using TiO2 and alginate microbeads

A double immobilisation technique was developed for C. antarctica lipase (CALB) that improved its enzymatic activity and solved difficulties frequently observed in reactions catalysed by lipases. The first immobilisation consisted of CALB adsorption onto a TiO2 nanoparticle surface (CALB(TiO2)). The adsorption was carried out by an oriented monolayer formed by CALB's hydrophilic amino acids and the TiO2 surface, leaving the CALB's active site accessible for reaction. As a result, an increase in enzymatic activity was achieved. The Relative Enzymatic Activity ((REA) obtained was 289%. The second immobilisation consisted of CALBTiO(2) entrapment into calcium alginate microbeads [(CALBTiO(2))(EDTA-Ca)], obtained by an internal crosslinking ion-exchange mechanism and using microfluidic droplet technique. The microbiocatalyst obtained, (CALB(TiO2))(EDTA-Ca), retained a high enzymatic activity (REA(t=0) = 232%) and stability (REA(t=30days) = 263%) at a size (diameter, O=8.9-10(4) nm) that enabled easier recovery than CALB(TiO2) (O = 2.3-102 nm) or CALB(Free) (O=5.0 nm), and showed a favourable porosity for diffusion without releasing CALB(TiO2.) Although the microbeads showed CALBFree leaching, as demonstrated by the loss of REA after a mechanical resistant test, (CALB(TiO2)) EDTA-Ca maintained an almost constant REA. (C) 2022 The Korean Society of Industrial and Engineering Chemistry. Published by Elsevier B.V. All rights reserved.

Automated summary

A double immobilisation technique was developed to improve the enzymatic activity of CALB and address difficulties in lipase-catalysed reactions. The first immobilisation involved adsorption of CALB onto a TiO2 nanoparticle surface, resulting in increased enzymatic activity. The second immobilisation used calcium alginate microbeads to trap CALB(TiO2), providing high enzymatic activity and stability with easier recovery. This study presents a promising approach for enzyme immobilisation in industrial applications.