Reuse of Immobilized Komagataella phaffii Cells for the Elimination of d‑Glucose in Syrups of Bioactive Carbohydrates

During the synthesis of prebiotic carbohydrates such as fructooligosaccharides (FOS), galactooligosaccharides (GOS), or isomaltooligosaccharides (IMOS), d-glucose is released as a side-product of the transglycosylation process. It is desirable to remove glucose from these sugar mixtures due to its caloric contribution and its effect on caries and diabetes. In this work, we have investigated the use of immobilized Komagataella phaffii (formerly Pichia pastoris) for elimination of d-glucose and d-fructose in several sugar syrups. K. phaffii cells were immobilized in calcium alginate beads to facilitate the separation of the yeast cells from the reaction medium and reuse of the biocatalyst. The immobilized yeasts were successfully reutilized for at least 20 cycles (of 195 min) to remove d-glucose (62.3 g/L) and d-fructose (5.5 g/L) in a FOS syrup, without affecting the concentration of oligosaccharides. Excellent selectivity was also found for elimination of d-glucose (57.2 g/L) in IMOS syrups. The methodology is versatile and easy to scale-up, as demonstrated in the removal of d-glucose (97.5 g/L) and d-fructose (142 g/L) for the purification of heteroglucooligosaccharides synthesized by Metschnikowia reukaufii α-glucosidase. In addition, d-glucose (50 g/L) was selectively removed by K. phaffii beads in the presence of d-galactose (50 g/L) for at least 20 cycles of 150 min and applied to GOS purification.