Iris Joye

Wheat Gluten Functionality as a Quality Determinant in Cereal-Based Food Products

The unique properties of wheat reside primarily in its gluten-forming storage proteins. Their intrinsic viscoelastic behavior is responsible for the characteristics of different wheat-based foods and for the use of wheat gluten proteins in different food products. Wheat-based food processing generally develops and sets the gluten protein network. Heat-induced gluten aggregation proceeds through cross-linking within and between its protein fractions. Prominent reactions include sulfhydryl (SH) oxidation and SH-disulfide (SS) interchange, which lead to SS cross-links. Other covalent bonds are also formed. Gluten functionality can be (bio-) chemically impacted. We focus on bread making, in which gluten proteins contribute to dough properties, bread loaf volume, and structure, and on pasta production, in which gluten proteins generate the desired cooking quality. Furthermore, it is speculated that the structure and texture of soft wheat products are also, at least to some degree, shaped by the heat-induced changes in the gluten protein fraction.

Fluorescence quenching study of resveratrol binding to zein and gliadin: towards a more rational approach to resveratrol encapsulation using water-insoluble proteins

Several health benefits have been ascribed to consumption of resveratrol, a polyphenol that can be extracted from grape skins. However, its use as a nutraceutical ingredient is compromised by its low water solubility, chemical stability, and bioavailability. Encapsulation of resveratrol in protein nanoparticles can be used to overcome these issues. Fluorescence quenching experiments were used to study the interaction of resveratrol with gliadin and zein. Resveratrol interacted with both proteins, but the binding constant was higher for zein than for gliadin at 35 °C. Furthermore, binding between resveratrol and gliadin increased at higher temperatures, which was not observed for zein. Analysis of the thermodynamic parameters suggested that resveratrol-gliadin binding mainly occurs through hydrophobic interactions while the binding with zein is predominantly mediated through hydrogen bonds. These results help rationalise ingredient selection and production of protein nanoparticles and microparticles for encapsulation, protection and release of resveratrol and potentially other bioactive compounds.

Computational design-based molecular engineering of the glycosyl hydrolase family 11 B. subtilis XynA endoxylanase improves its acid stability

Rational protein engineering was applied to improve the limited stability of the glycosyl hydrolase family 11 (GH11) endo-beta-1,4-xylanase from Bacillus subtilis under acidic conditions. Since the pH dependence of protein stability is governed by the ionisation states of the side chains of its titrable amino acid residues, we explored the strategy of changing pH-stability profiles by altering pK(a) values of key residues through in silico designed mutations. To this end, computational predictions and molecular modelling were carried out using the recently developed pKD software package. Four endoxylanase variants, in which the pK(a) values of either Asp4 and Asp11 or His149 were targeted to shift downwards through incorporation of three to five point mutations, were generated and recombinantly expressed in the cytoplasm of Escherichia coli. All four mutants showed considerably increased functional stability at acid pH levels. They retained approximately 30-70% and approximately 75-95% of their activity after incubation at pH 3 and 4, respectively, in comparison with only approximately 23% and approximately 57%, respectively, for the wild-type enzyme under the experimental conditions. No acidophilic adaptation of the catalytic activity had occurred. In addition, their functional stability and catalytic activity profiles under different temperature and ionic strength conditions were significantly altered. These findings contribute to general understanding of the molecular mechanisms governing the pH-dependent stability of GH11 proteins, and hence they can be applied to enhance the stability and effectiveness of many GH11 endoxylanases used in industry today.

Food-Grade Protein-Based Nanoparticles and Microparticles for Bioactive Delivery: Fabrication, Characterization, and Utilization

Proteins can be used to fabricate nanoparticles and microparticles suitable for use as delivery systems for bioactive compounds in pharmaceutical, food, cosmetic, and other products. Food proteins originate from various animal or vegetal sources and exhibit a wide diversity of molecular and physicochemical characteristics, e.g., molecular weight, conformation, flexibility, polarity, charge, isoelectric point, solubility, and interactions. As a result, protein particles can be assembled using numerous different preparation methods, from one or more types of protein or from a combination of a protein and another type of biopolymer (usually a polysaccharide). The final characteristics of the particles produced are determined by the proteins and/or polysaccharides used, as well as the fabrication techniques employed. This chapter provides an overview of the functional properties of food proteins that can be used to assemble nanoparticles and microparticles, the fabrication techniques available to create those particles, the factors that influence their stability, and their potential applications within the food industry.

Effect of Maillard Conjugates on the Physical Stability of Zein Nanoparticles Prepared by Liquid Antisolvent Coprecipitation

Protein nanoparticles are often not very stable in a complex food matrix because they are primarily stabilized by electrostatic repulsion. In this study, we envisaged the stabilization of zein nanoparticles through Maillard conjugation reactions with polysaccharides of different molecular mass. Zein nanoparticles (0.5% w/v) containing resveratrol (0.025% w/v grape skin extract) were produced by liquid antisolvent precipitation and coated with Maillard conjugates (MC) of sodium caseinate and different molecular mass carbohydrates during particle production. Zein nanoparticles coated with conjugated polysaccharides of 2.8, 37, and 150 kDa had diameters of 198 ± 5, 176 ± 6, and 180 ± 3 nm, respectively. The encapsulation efficiency (∼83%) was not affected by conjugation, but the conjugates significantly improved particle stability against changes in pH (2.0-9.0), CaCl2 addition (up to 100 mM), and heat treatment (30-90 °C, 30 min). Zein nanoparticles coated by MC may therefore be suitable delivery systems for hydrophobic bioactive molecules in a wide range of commercial products.

Biopolymer-Based Delivery Systems: Challenges and Opportunities.

Biopolymer-based nanostructures or microstructures can be fabricated with different compositions, structures, and properties so that colloidal delivery systems can be tailored for specific applications. These structures can be assembled using various approaches, including electrospinning, coacervation, nanoprecipitation, injection, layer-by-layer deposition, and/or gelation. A major application of biopolymer-based particles is to encapsulate, protect, and release active molecules in the agricultural, food, supplements, personal care, and pharmaceutical sectors. The inherent variability and complexity of biopolymers (proteins and polysaccharides) often makes it challenging to produce particles with well-defined physicochemical and functional attributes. In this review, we discuss the properties of biopolymers, common particle fabrication methods, and some of the major challenges and opportunities associated with developing biopolymer-based particles for application as food-grade delivery systems.

In situ production of γ-aminobutyric acid in breakfast cereals

Breakfast cereals are an important part of an equilibrated diet in the Western world, making them extremely suited for carrying health benefits. Intake of γ-aminobutyric acid (GABA), a major inhibitory neurotransmitter of the nervous system, has been related to blood pressure lowering in hypertensive individuals. In vivo, GABA is formed from glutamic acid (GA) by glutamic acid decarboxylase (GAD), a widely distributed enzyme in prokaryotic and eukaryotic species. We here enriched breakfast cereals with GABA by recipe and process optimisation. The dynamics of GA and GABA were monitored throughout the production process. Addition of exogenous recombinantly produced GAD of Yersinia intermedia increased GABA levels by 2- to 5-fold. As only trace levels of GABA (<15ppm) and relatively low levels of its precursor (GA, <100ppm) are present in the wheat and rice flour used, a well-thought ingredient choice (inclusion of quinoa flour (ca. 90ppm GABA and 700ppm GA) or bran enrichment (ca. 66ppm GABA and 500ppm GA)) also significantly increases the GABA content in the final flakes. Finally, a strict control of the heating steps during the production process reduces GA and GABA losses. Consumption of one portion (30g) of the here produced enriched breakfast cereals can even meet up to 55% of the daily intake earlier reported to lower blood pressure (ca. 10mg).

Emulsifying and Emulsion-Stabilizing Properties of Gluten Hydrolysates

Gluten is produced as a coproduct of the wheat starch isolation process. In this study, gluten was hydrolyzed to degrees of hydrolysis (DH) of 3-6-10 and 1-2-3 with alcalase and trypsin, respectively. These peptidases have a clearly distinct substrate specificity. Corn oil-in-water emulsions (10 wt % oil) were prepared by high-pressure homogenization at pH 7.5. Gluten peptides with DH 3 proved to be the most effective in producing peptides displaying emulsifying properties. Higher levels of alcalase hydrolysates (2.0 wt %) than of trypsin hydrolysates (1.0 wt %) were required to produce stable emulsions with small droplet sizes, which is attributed to differences in the nature of the peptides formed. The emulsions had small mean droplet diameters (d32 < 1000 nm). Emulsions produced with trypsin hydrolysates (stable after 9 days at 55 °C) displayed better thermal stability compared to those produced with alcalase hydrolysates (destabilized after 2 days at 37 °C). The hydrolysate-containing emulsions, however, were quickly destabilized by salt addition (≤100 mM NaCl) and when the pH approached the isoelectric point of the coated droplets (pH ~5.5). Microscopic analysis revealed the formation of air-in-oil-in-water emulsions at lower hydrolysate concentrations, whereas at higher concentrations (≥3.0 wt %) extensive flocculation occurred. Both phenomena contributed to creaming of the emulsions. These results may be useful for the utilization of gluten hydrolysates in food and beverage products.

The Bread Dough Stability Improving Effect of Pyranose Oxidase from Trametes multicolor and Glucose Oxidase from Aspergillus niger: Unraveling the Molecular Mechanism

Glucose oxidase (GO) and pyranose oxidase (P2O) improve dough stability and bread quality. We here studied whether their mode of action resides in cross-linking of proteins and/or arabinoxylan (AX) molecules through the production of H2O2. Evidence for both was deduced from a decrease in extractability of protein and AX from dough made with P2O, GO, or H2O2, using sodium dodecyl sulfate containing buffer and water, respectively. The addition of H2O2, P2O, or GO to a glutathione solution sharply decreased its sulfhydryl (SH) content. P2O or GO can trigger protein cross-linking through the formation of disulfide (SS) bonds. As a result thereof, SH/SS interchange reactions between low molecular mass SH containing compounds and gluten proteins can be hampered. Furthermore, a decrease in the level of monomeric ferulic acid (FA) esterified to AX in dough points to a role of FA bridges in cross-linking of AX molecules. Our results indicate that the molecular mechanism of dough and bread improvement by P2O and GO resides in cross-linking of gluten proteins and AX by formation of H2O2. They furthermore show that the extent of cross-linking upon addition of P2O or GO strongly depends on the concentration (and production rate) of H2O2.

Monitoring Molecular Oxygen Depletion in Wheat Flour Dough Using Erythrosin B Phosphorescence: A Biophysical Approach

During wheat flour dough mixing air is incorporated. However, soon after mixing, the dough’s molecular oxygen (O2) disappears. O2 is vital in several oxidation reactions which affect, and, in many cases, improve dough and bread quality. We here for the first time monitored the O2 level in fresh dough as a function of time using erythrosin B, a phosphorescent probe. O2 is a very efficient phosphorescence quencher. Upon its depletion, erythrosin B phosphorescence lifetime is significantly extended. The O2 depletion in time in unyeasted dough substantially depends on flour characteristics. As expected, yeast in the recipe significantly accelerated the rate of O2 consumption in dough. Furthermore, little if any diffusion of O2 into dough occurs. Overall, monitoring phosphorescence lifetimes is very valuable for measuring O2 depletion and can contribute to the development of and search for potential O2 dependent bread improving agents.