Functionalized mesoporous biosilica for immobilizing D-allulose 3-epimerase: A multienzyme cascade strategy for calorie reduction in fruit tea drinks

Abstract

D-allulose, a low-calorie rare sugar, can be biosynthesized by D-allulose 3-epimerase (DAE). Here, DAE was covalently immobilized on polyethyleneimine-modified diatomite using two methods: a biopolymer coating approach (DAE@PEI-GA@PDP) and a direct attachment method (DAE-GA@PEI@PDP). Both immobilized DAE enzymes exhibited enhanced thermal stability and broader pH tolerance compared to the free DAE. They also exhibited excellent reusability and maintained more than 70% of their activity after 10 cycles. Additionally, a multienzyme cascade system was developed to produce D-allulose from low-cost D-glucose using immobilized glucose isomerase (GI) and immobilized DAE enzymes. This system obtained 76 g/L D-allulose from 500 g/L D-glucose. Notably, the application was extended to fruit tea drinks, successfully transforming high-calorie sugars (D-glucose and D-fructose) into D-allulose. This research offers a promising immobilization strategy for DAE on silica-based materials and provides robust biocatalysts for creating functional fruit tea drinks aimed at diabetics and individuals with specific dietary needs.