A new two-step enzymatic conversion process for the production of a novel mono-α-1,4-glucosylated rebaudioside A derivative (RA1G) was established via transglycosylation followed by hydrolyzation. In the transglycosylation reaction catalyzed by cyclodextrin glycosyltransferase, rebaudioside A (RA) was converted into glucosylated RA derivatives (RAGs), and the maximum conversion of 87.8% was obtained with the optimal conditions of 2 U/mL enzyme, 82.5 mg/mL β-cyclodextrin, and 82.5 mg/mL RA at 60 °C for 5 h. The obtained RAG solution was then directly used in hydrolyzation. Four amylases were screened for shortening the newly synthesized α-glucans of RAGs. A glucoamylase was found to produce RA1G as the single glucosylated product, and the maximum yield of 53.3% was achieved with the optimal conditions of adding 1.5 U/mL glucoamylase into RAG solution at 60 °C for 3 h. RA1G was identified as 13-[(2-O-β-D-glucopyranosyl-3-O-β-D-glucopyranosyl-β-D-glucopyranosyl) oxy] ent-kaur-16-en-19-oic acid-[(4-O-α-D-glucopyranosyl-β-D-glucopyranosyl) ester] by MS and NMR analysis and showed an improved sensory quality compared to RA.
Keywords: cyclodextrin glycosyltransferase; glucoamylase; mono-α-1,4-glucosylated rebaudioside A derivative; rebaudioside A; two-step enzymatic conversion.