Immobilization of pullulanase from Bacillus licheniformis on magnetic multi-walled carbon nanotubes for maltooligosaccharide production

In this study, Fe₃O₄-coated multi-walled carbon nanotubes (MWCNT-Fe₃O₄) or nickel oxide-coated multi-walled carbon nanotubes (MWCNT-NiO) were activated with 3-Glycidyloxypropyl)trimethoxysilane (3-GPTMS) to create oxirane groups. Pullulanase from Bacillus licheniformis was covalently immobilized on these magnetic MWCNTs to obtain magnetically separable immobilized pullulanase preparations (MWCNT-Fe₃O₄@Pul or MWCNT-NiO@Pul) for producing maltooligosaccharides (MOSs) from pullulan. The highest recovered activity values were 78% and 85% respectively, for MWCNT-Fe₃O₄@Pul and MWCNT-NiO@Pul after 24 h of immobilization at pH 7.0. The optimal pH and temperature were found to be 5.5 and 45 °C for free pullulanase, whereas the corresponding values were 5.5 and 50 °C for both immobilized pullulanase preparations. The thermal stabilities of MWCNT-Fe₃O₄@Pul and MWCNT-NiO@Pul increased by 6.2- and 8.2-fold, respectively, at 50 °C. The catalytic efficiencies of MWCNT-Fe₃O₄@Pul and MWCNT-NiO@Pul were calculated to be 0.8- and 1.1-fold that of free pullulanase, respectively. After 24 h of hydrolysis, MOS yields were determined to be 470 and 490 mg MOS/g pullulan for MWCNT-Fe₃O₄@Pul and MWCNT-NiO@Pul, respectively. The remaining activities were 86% and 85% for MWCNT-Fe₃O₄@Pul and MWCNT-NiO@Pul after 10 reuses, respectively.