Synthesis and structural characterization of raffinosyl-oligofructosides upon transfructosylation by Lactobacillus gasseri DSM 20604 inulosucrase

Abstract

A new process based on enzymatic synthesis of a series of raffinose-derived oligosaccharides or raffinosyl-oligofructosides (RFOS) with degree of polymerization (DP) from 4 to 8 was developed in the presence of raffinose. This process involves a transfructosylation reaction catalyzed by an inulosucrase from Lactobacillus gasseri DSM 20604 (IS). The main synthesized RFOS were structurally characterized by Nuclear Magnetic Resonance (NMR). According to the elucidated structures, RFOS consist of β-2,1-linked fructose unit(s) to raffinose: α-D-galactopyranosyl-(1→6)-α-Dglucopyranosyl-(1↔2)-β-D-fructofuranosyl-((1←2)-β-D-fructofuranoside)n (where n refers to the number of transferred fructose moieties). The maximum yield of RFOS was 33.4% (in weight respect to the initial amount of raffinose) and was obtained at the time interval of 8-24 h of transfructosylation reaction initiated with 50% (w/v) of raffinose. Results revealed the high acceptor and donor affinity of IS towards raffinose, being fairly comparable to that of sucrose for the production of fructooligosaccharides (FOS), including when both carbohydrates coexisted (sucrose:raffinose mixture, 250 g L -1 each). The production of RFOS was also attempted in the presence of sucrose:melibiose mixtures; in this case, the predominant acceptor-product formed was raffinose followed by a minor production of a series of oligosaccharides with varying DP. The easiness of RFOS synthesis and the structural similarities with both raffinose and fructan series of oligosaccharides warrant the further study of the potential bioactive properties of these unexplored oligosaccharides.

Structure and function of non-digestible carbohydrates in the gut microbiome

“…FOS have been also efficiently separated using classical HILIC conditions, i.e. on amine modified silica columns under isocratic conditions with acetonitrile and water as mobile phase and a refractive index detector (Díez-Municio et al, 2016;Rodríguez-Gómez et al, 2015). Similar column and chromatographic conditions have been successfully applied for the efficient separation of hetero-fructooligosaccharides, such as maltosyl- (Díez-Municio et al, 2013), lactosyl-(Díez-Municio et al, 2015 and raffinosyl-oligofructosides (Díez-Municio et al, 2016).…”

“…on amine modified silica columns under isocratic conditions with acetonitrile and water as mobile phase and a refractive index detector (Díez-Municio et al, 2016;Rodríguez-Gómez et al, 2015). Similar column and chromatographic conditions have been successfully applied for the efficient separation of hetero-fructooligosaccharides, such as maltosyl- (Díez-Municio et al, 2013), lactosyl-(Díez-Municio et al, 2015 and raffinosyl-oligofructosides (Díez-Municio et al, 2016). Different types of FOS together with raffinose have been also determined and characterised by different HILIC-MS methods with gradient elution in complex food matrices, such as wheat flour (Liu and Rochfort, 2015) and artichoke industrial waste (Machado et al, 2015).…”

Fructan Biosynthesis by Yeast Cell Factories

“…For inulin-type fructan, inulosucrase (ISRase, EC 2.4.1.9, for inulin) was used. Functional analysis of ISRase was conducted using enzymes from the Lactobacillus genus [ 45 - 49 ], but has not been applied in the industrial production of fructan due to low productivity. Therefore, the industrial production of inulin-type fructan has been performed using FTases from Aspergillus species, ensuring superior fructosylation activity.…”

Fermentation by Probiotic Lactobacillus gasseri Strains Enhances the Carotenoid and Fibre Contents of Carrot Juice

“…Recently, Lactobacillus gasseri strains have attracted attention due to their probiotic properties [12][13][14][15] and their ability to produce fructosyltransferase enzymes that catalyze the conversion of sucrose into prebiotic fructans [16][17][18]. Genomic and experimental evidences indicate that L. gasseri strains possess probiotic traits including tolerance to low pH, resistance to bile salts, adhesion to host epithelium and production of antimicrobial bacteriocins with inhibitory activity against pathogens [18,19].…”