Structural Elucidation of Enzymatically Synthesized Galacto-oligosaccharides Using Ion-Mobility Spectrometry–Tandem Mass Spectrometry

Abstract

Galacto-oligosaccharides (GOS) represent a diverse group of well-characterized prebiotic ingredients derived from lactose in a reaction catalyzed with β-galactosidases. Enzymatic transgalactosylation results in a mixture of compounds of various degrees of polymerization and types of linkages. Because structure plays an important role in terms of prebiotic activity, it is of crucial importance to provide an insight into the mechanism of transgalactosylation reaction and occurrence of different types of β-linkages during GOS synthesis. Our study proved that a novel one-step method, based on ion-mobility spectrometry-tandem mass spectrometry (IMS-MS/MS), enables complete elucidation of GOS structure. It has been shown that β-galactosidase from Aspergillus oryzae has the highest affinity toward formation of β-(1→3) or β-(1→6) linkages. Additionally, it was observed that the occurrence of different linkages varies during the reaction course, indicating that tailoring favorable GOS structures with improved prebiotic activity can be achieved by adequate control of enzymatic synthesis.

Whey valorization using transgalactosylation activity of immobilized β‐galactosidase

“…Namely, the reaction mixture consisted of GOS (tri‐saccharides ‐GOS3 and tetra‐saccharides ‐ GOS4), lactose, glucose and galactose. Such composition was rather expected, bearing in mind that β‐galactosidase from A. oryzae exhibits a low tendency towards creating other disaccharides, as well as the higher oligosaccharides (Carević et al ., ). Additionally, it can be seen that obtained results (total GOS concentrations) using both substrates are quite similar (Fig.…”

“…Throughout this study, the possibility of economically viable whey utilization for GOS synthesis using enzyme β‐galactosidase from A. oryzae was examined. This enzyme was chosen on the basis of its availability, relatively low prices, and more importantly, our previous findings claiming its outstanding transgalactosylation activity (Carević et al ., ,b).…”

Optimization of galacto-oligosacharides synthesis using response surface methodology

“…Products of transgalactosylation are depicted by trisaccharide (GOS3) peak (t R =8.733 min) and tetrasaccharide (GOS4) peak (t R =8.213 min). In our previous study (Carević et al, 2016a) was confirmed by IMS/MS-MS analysis that other disaccharides, except the lactose, are not pre-sent in the reaction mixture and that GOS3 and GOS4 peaks exclusively with β(1→6) and β(1→3) glycosidic linkages are the only transgalactosylation products. …”

“…GOS represent a diverse group of water soluble carbohydrates consisting of several galactose and usually one terminal glu-cose unit, that can notably vary in polymerization degree, type of linkages, and more importantly in their properties (Urrutia et al, 2013, Carević et al, 2016a. They are marked as non-digestible compounds, recognized as prebiotics owing to the fact that are selectively metabolized by some favorable intestinal microbiota, particularly bifidobacteria and lactobacilli, providing their proliferation and promotion of their activity (Arreola et al, 2014;Hong et al, 2016).…”

Galactooligosaccharide Production from Pantoea anthophila Strains Isolated from “Tejuino”, a Mexican Traditional Fermented Beverage

“…The most studied β-galactosidases for GOS synthesis from lactose are those from strains of Kluyveromyces [15,16], Aspergillus [17,18], Bacillus [19,20], and Bifidobacterium [5,21]. GOS production with microbial β-galactosidases could be performed using crude, purified, recombinant, or immobilized enzymes and also whole cells [2,8].…”