Biotransformation of sucrose rich Maple syrups into fructooligosaccharides, oligolevans and levans using levansucrase biocatalyst: Bioprocess optimization and prebiotic activity assessment.

Maple syrup was investigated as a source to produce FOSs and ß-(2-6)-linked-oligolevans/levans. The modulation of this biotransformation was achieved through the control of Maple syrup °Bx and reaction conditions. Reaction time was identified as the most influential factor for the oligolevans/FOSs production in Maple syrup 30°Bx reaction system as well as for the oligolevans/levans synthesis in the 66°Bx one. In the predictive model of oligolevans/levans production in Maple syrup 60°Bx, the interactive effect between levansucrase unit and reaction time was significant (p-value of 0.0008). The optimal conditions for oligolevans/FOSs production (109.20 g/L) in Maple syrup 30°Bx were 3.73 U/mL, pH 6.60 and 23.12 h; while 5 U/mL, pH 6.04 and 29.92 h were identified as the optimal conditions for oligolevans/levans production (147.09 g/L) in Maple syrup 66°Bx. As compared to inulin-type commercial FOSs, the fermentation of oligolevans/FOSs from Maple syrup led to a higher count of Lactobacillus acidophilus and Bifidobacterium lactis and resulted in a higher production of lactic acid. This study lays the foundation for the biotransformation of Maple syrups into functional prebiotic ingredients.

Bacillus amyloliquefaciens levansucrase-catalyzed the synthesis of fructooligosaccharides, oligolevan and levan in maple syrup-based reaction systems.

Maple syrups with selected degree Brix (°Bx) (15, 30, 60) were investigated as reaction systems for levansucrase from Bacillus amyloliquefaciens. The enzymatic conversion of sucrose present in the maple syrup and the production of the transfructosylation products were assessed over a time course of 48h. At 30°C, the use of maple syrup 30°Bx led to the highest levansucrase activity (427.53µmol/mg protein/min), while maple syrup 66°Bx led to the highest converted sucrose concentration (1.53M). In maple syrup 30°Bx, oligolevans (1080%). In maple syrup 66°Bx, the most abundant products were oligolevans at 30°C and levans (DP≥30) at 8°C. The acceptor specificity study revealed the ability of B. amyloliquefaciens levansucrase to synthesize a variety of hetero-fructooligosaccharides (FOSs) in maple syrups 15°Bx and 30°Bx enriched with various disaccharides, with lactose being the preferred fructosyl acceptor. The current study is the first to investigate maple-syrup-based reaction systems for the synthesis of FOSs/oligolevans/levans.

Enzymatic Synthesis of Galactosylated Serine/Threonine Derivatives by ß-Galactosidase from Escherichia coli.

The transgalactosylations of serine/threonine derivatives were investigated using ß-galactosidase from Escherichia coli as biocatalyst. Using ortho-nitrophenyl-ß-D-galactoside as donor, the highest bioconversion yield of transgalactosylated N-carboxy benzyl L-serine benzyl ester (23.2%) was achieved in heptane:buffer medium (70:30), whereas with the lactose, the highest bioconversion yield (3.94%) was obtained in the buffer reaction system. The structures of most abundant galactosylated serine products were characterized by MS/MS. The molecular docking simulation revealed that the binding of serine/threonine derivatives to the enzyme's active site was stronger (-4.6~-7.9 kcal/mol) than that of the natural acceptor, glucose, and mainly occurred through interactions with aromatic residues. For N-tert-butoxycarbonyl serine methyl ester (6.8%) and N-carboxybenzyl serine benzyl ester (3.4%), their binding affinities and the distances between their hydroxyl side chain and the 1'-OH group of galactose moiety were in good accordance with the quantified bioconversion yields. Despite its lower predicted bioconversion yield, the high experimental bioconversion yield obtained with N-carboxybenzyl serine methyl ester (23.2%) demonstrated the importance of the thermodynamically-driven nature of the transgalactosylation reaction.

Interactions of caseins with phenolic acids found in chocolate.

To investigate the interactions between caseins and phenolic acids, such as the ones present in chocolate, casein was incubated with protocatechuic acid or p-coumaric acid at 55°C. In addition, casein was isolated from chocolate and the phenolic compounds within these caseins were quantified. Electrophoresis results revealed that casein-phenolic interactions were induced by incubation; minor aggregation of casein subunits was observed after incubation of casein with protocatechuic acid. Minor aggregation of casein isolated from milk chocolate was also observed. In vitro hydrolysis of casein control, casein-protocatechuic acid, casein-p-coumaric acid, caseins isolated from milk chocolate and white chocolate using trypsin showed degree of hydrolysis of 19.3, 18.6, 17.7, 10.4 and 17.8% respectively. The presence of protocatechuic acid and p-coumaric acid in the model system and the presence of phenolic compounds in milk chocolate, in addition to the structural changes occurring during processing, affected the peptide profiles of casein hydrolysates.

Investigation of the use of Maillard reaction inhibitors for the production of patatin-carbohydrate conjugates.

Selected Maillard reaction inhibitors, including aminoguanidine, cysteine, pyridoxamine, and sodium bisulfite, were evaluated for their effect on the production of carbohydrate conjugated proteins with less cross-linking/browning. Patatin (PTT), a major potato protein, was glycated with galactose, xylose, galactooligosaccharides, xylooligosaccharides, galactan, and xylan under controlled conditions. The effectiveness of the inhibitors to control the glycation reaction was assessed by monitoring the glycation extent, the protein cross-linking, and the formation of dicarbonyl compounds. Sodium bisulfite was the most effective inhibitor for PTT-galactose and PTT-xylan reaction systems (reaction control ratios of 210.0 and 12.8). On the other hand, aminoguanidine and cysteine led to the highest reaction control ratios for the PTT-xylose/xylooligosaccharide (160.0 and 143.0) and PTT-galactooligosaccharides/galactan (663.0 and 71.0) reaction systems, respectively. The use of cysteine and aminoguanidine as inhibitors led to 1.7-99.4% decreases in the particle size distribution of the PTT conjugates and to 0.4-9.3% increases in their relative digestibility, per 5% blocked lysine.

Production and characterisation of potato patatin-galactose, galactooligosaccharides, and galactan conjugates of great potential as functional ingredients.

Potato proteins are of high interest because of their high nutritional quality and multiple health benefits, but they are currently undervalued due to their limited solubility and stability. Glycated patatin (PTT) with galactose, galactooligosaccharides (GOSs) and galactan were produced through the Maillard reaction and characterised structurally and functionally. Fourier-transform infrared and fluorescence spectroscopy data revealed important changes in total secondary structures through glycation with GOSs (61.2%) and galactan (36.7%) and also significant tertiary structural changes leading to an exposure of tryptophan residues. These structural changes led to more heat stable forms of PTT with a higher unfolding temperature (70-90 °C) than the unmodified protein (50-70 °C) and with higher antioxidant activity. PTT:galactose conjugates exhibited similar thermal stability and pH-structural behaviour to native PTT. However, the high level of galactose conjugation to PTT and increased exposure of hydrophobic residues led to a significant increase in its emulsifying stability at pH 3.

Allergenicity of potato proteins and of their conjugates with galactose, galactooligosaccharides, and galactan in native, heated, and digested forms.

The effect of glycation of potato proteins on their immunoreactivity was studied by using a pool of human sera with specific IgE to potato proteins. Patatin conjugates were more immunoreactive than protease inhibitors ones. To better understand this behavior, the changes in patatin structure upon glycation and heat treatment were investigated. Patatin demonstrated an increase in total immunoreactivity when glycated with galactose and galactooligosaccharides. However, galactan conjugation to patatin resulted in a decrease in immunoreactivity by restricting IgE's access to the epitopes. Although the heat treatment resulted in a decrease in patatin's immunoreactivity through aggregation, it was less effective when patatin conjugates were used due to the decrease in aggregation and the secondary structural changes. Upon digestion, native patatin exhibited the largest decrease in immunoreactivity resulting from the disruption of both conformational and sequential epitopes. Patatin conjugates were less digested and had higher IgE-immunoreactivity as compared to the digested patatin.