Taguchi design-assisted immobilization of Candida rugosa lipase onto a ternary alginate/nanocellulose/montmorillonite composite: Physicochemical characterization, thermal stability and reusability studies

Biomass from oil palm frond leaves (OPFL) is an excellent reservoir of lignocellulosic material which full potential remains untapped. This study aimed to statistically optimize the covalent immobilization of Candida rugosa lipase (CRL) onto a ternary support comprised of OPFL derived nanocellulose (NC) and montmorillonite (MMT) in alginate (ALG) (CRL-ALG/NC/MMT). The coarser topology and the presence of characteristic spherical globules in the field emission scanning electron micrographs and atomic force micrographs, respectively, supported the existence of CRL on ALG/NC/MMT. In addition, amide peaks at 3478 and 1640 cm-1 in the fourier transform infrared spectra affirmed that CRL was covalently bonded to ALG/NC/MMT. The optimized Taguchi Design-assisted immobilization of CRL onto ALG/NC/MMT (7 h of immobilization, 35℃, pH 5, 7 mg/mL protein loading) gave a production yield of 92.89 % of ethyl levulinate (EL), as proven by gas chromatography-mass spectrometric ([M] +m/z 144, C7H12O3), FTIR and nuclear magnetic resonance (CAS-539-88-8) data. A higher optimal reaction temperature (50℃) and the reusability of CRL-ALG/NC/MMT for up to 9 esterification cycles substantiated the appreciable structural rigidification of the biocatalyst by ALG/NC/MMT, which improved the catalytic activity and thermal stability of the lipase.