Insights into digestibility, biological activity, and peptide profiling of flaxseed protein isolates treated by ultrasound coupled with alkali cycling.

This study aims to investigate the effects of ultrasound coupled with alkali cycling on the structural properties, digestion characteristics, biological activity, and peptide profiling of flaxseed protein isolates (FPI). The digestibility of FPI obtained by ultrasound coupled with pH 10/12 cycling (UFPI-10/12) (74.56 % and 79.12 %) was significantly higher than that of native FPI (64.40 %), and UFPI-10 showed higher hydrolysis degree (35.76 %) than FPI (30.65 %) after intestinal digestion. The combined treatment induced transition from α-helix to ß-sheet with an orderly structure. Large FPI aggregates broke down into small-sized FPI particles, which induced the increase of specific surface area of particles. This might expose more cutting sites and contact area with enzymes. Furthermore, UFPI-10 showed high antioxidant activity (29.18 %) and lipid-lowering activity (70.52 %). Peptide profiling revealed that UFPI-10 exhibited a higher proportion of 300-600 Da peptides and significantly higher abundance of antioxidant peptides than native FPI, which might promote its antioxidant activity. Those results suggest that the combined treatment is a promising modification method to improve the digestion characteristics and biological activity of FPI. This work provides new ideas for widespread use of FPI as an active stabilizer in food systems.

Effects of airflow superfine pulverization on the structure, functional properties, and flavor quality of wheat bran.

BACKGROUND: Wheat bran (WB) is a byproduct of refined wheat flour production with poor edible taste and low economic value. Herein, the WB was micronized via airflow superfine pulverization (ASP), and the effects of the ASP conditions on its particle size, nutritive compositions, whiteness, hydration characteristics, moisture distribution, microstructure, cation exchange capacity, volatile flavor components, and other characteristics were investigated. RESULTS: Reducing the rotational speed of the ASP screw and increasing the number of pulverizations significantly decreased the median particle size Dx(50) of WB to a minimum of 12.97 ± 0.19 µm (P < 0.05), increased the soluble dietary fiber content from 55.05 ± 2.94 to 106.86 ± 1.60 mg g-1, and improved the whiteness and water solubility index. In addition, the water holding capacity and oil holding capacity were significantly reduced (P < 0.05), while the cation exchange and swelling capacities first increased and then decreased. Up to about 70% of water in WB exists as bound water. As the Dx(50) of WB decreased, the content of bound and immobile water increased, while the free water decreased from 14.37 ± 1.21% to 7.59 ± 1.03%. Furthermore, WB was micronized and the particles became smaller and more evenly distributed. Using gas chromatography-ion mobility spectrometry, a total of 37 volatile compounds in micronized WB (including 10 aldehydes, 9 esters, 7 alcohols, and several acids, furans, ethers, aldehydes, esters, and alcohols) were identified as the main volatile compounds of WB. CONCLUSION: Collectively, ASP improved the physicochemical properties of WB. This study provides theoretical references for the use of ASP to improve the utilization and edibility of WB. © 2024 Society of Chemical Industry.

Physicochemical characterization of a composite flour: Blending purple sweet potato and rice flours.

In this study, the physicochemical characterization of different ratios of purple sweet potato flour (PSPF) and rice flour was investigated to improve the nutritional value and enrich the variety of rice-based staple food. The results showed that adding PSPF increased total dietary fiber and anthocyanin content whereas decreased amylose content of the composite flours. Additionally, the composite flours exhibited lower thermodynamic parameters and displayed darker, redder, and bluer colors. There were no noticeable changes in the functional group structure of the composite flours. The addition of PSPF decreased the crystallinity and water-holding capacity of the composite flours, whereas increased the average particle size and iodine blue value. PSPF increased the pasting temperature of the flours whereas decreased the breakdown and setback values. Overall, the addition of PSPF significantly affects the nutrition, color and physicochemical properties of the composite flours.

Effect of pH, protein/polysaccharide ratio and preparation method on the stability of lactoferrin-polysaccharide complexes.

In this study, carrageenan (CG), xanthan gum (XG) and locust bean gum (LBG), which can be used in infant formulas in China national standards, were selected to prepare LF-polysaccharide complexes to improve the stability of lactoferrin. The results showed that LF interacted more strongly with polysaccharides and did not affect the LF structure to a large extent when the pH and protein/polysaccharide mass ratio were 7 and 10:1 for LF-CG, 8 and 5:1 for LF-XG, 7 and 15:1 for LF-LBG. The zeta potential and fluorescence intensity of the LF-polysaccharide complexes displayed a decreasing trend with the increase in pH. When pH < 6, LF-CG and LF-XG exhibited precipitation and increased UV absorbance. Complexation between LF and CG/XG mainly attributed to electrostatic interactions, while LF and LBG form complexes based on hydrogen bonding or hydrophobic interactions. This study could provide a reference for the practical application of LF in infant formula.

Soft gliadin nanoparticles at air/water interfaces: The transition from a particle-laden layer to a thick protein film.

HYPOTHESIS: Protein-based soft particles possess a unique interfacial deformation behavior, which is difficult to capture and characterize. This complicates the analysis of their interfacial properties. Here, we aim to establish how the particle deformation affects their interfacial structural and mechanical properties. EXPERIMENTS: Gliadin nanoparticles (GNPs) were selected as a model particle. We studied their adsorption behavior, the time-evolution of their morphology, and rheological behavior at the air/water interface by combining dilatational rheology and microstructure imaging. The rheology results were analyzed using Lissajous plots and quantified using the recently developed general stress decomposition (GSD) method. FINDING: Three distinct stages were revealed in the adsorption and rearrangement process. First, spherical GNPs (∼105 nm) adsorbed to the interface. Then, these gradually deformed along the interface direction to a flattened shape, and formed a firm viscoelastic 2D solid film. Finally, further stretching and merging of GNPs at the interface resulted in rearrangement of their internal structure to form a thick film with lower stiffness than the initial film. These results demonstrate that the structure of GNPs confined at the interface is controlled by their deformability, and the latter can be used to tune the properties of prolamin particle-based multiphase systems.

Understanding the mechanism of high viscosity food delaying gastric emptying.

Controlling the structure and viscosity of food can influence the development of diet-related diseases. Food viscosity has been linked with health through its impact on human digestion and gastrointestinal transit, however, there is limited understanding of how the viscosity of food regulates gastric emptying. Here, we used model food preparations with different viscosities using guar gum, to explore the mechanism underlying the influence of viscosity on gastric motility, gastric emptying and postprandial blood glucose. Based on experiments in human volunteers and animals, we demonstrated that high viscosity meals increased gastric antrum area and gastric retention rate. Viscosity also affected gut hormone secretion, reduced the gene expression level of interstitial cells of Cajal, resulting in a delay of gastric emptying and limiting the increase in postprandial glucose. This improved mechanistic understanding of food viscosity during gastric digestion is important for designing new foods to benefit human health.

Regulation of interfacial mechanics of soy protein via co-extraction with flaxseed protein for efficient fabrication of foams and emulsions.

Enrichment of plant proteins with functionality is of great importance for expanding their application in food formulations. This study proposed an innovation to co-enrich soy protein and flaxseed protein to act as efficient interfacial stabilizers for generating foams and emulsions. The structure, interfacial properties, and functionalities of the soy protein-flaxseed protein natural nanoparticles (SFNPs) obtained by alkali extraction-isoelectric precipitation (AE) and salt extraction-dialysis (SE) methods were investigated. Overall, the foamability of AE-SFNPs (194.67 %) was 1.45-fold that of SE-SFNPs, due to their more flexible structure, smaller particle size, and suitable surface wettability, promoting diffusion and adsorption at the air-water interface. AE-SFNPs showed higher emulsion stability (140.89 min), probably because the adsorbed AE-SFNPs with smaller size displayed soft particle-like properties and stronger interfacial flexibility, and therefore could densely and evenly arrange at the interface, facilitating the formation of a stiff and solid-like interfacial layer, beneficial for more stable emulsion formation. The findings may innovatively expand the applications of SFNPs as food ingredients.

Redesign of air/oil-water interface via physical fields coupled with pH shifting to improve the emulsification, foaming, and digestion properties of plant proteins.

The application of plant proteins in food systems is largely hindered by their poor foaming or emulsifying properties and low digestibility compared with animal proteins, especially due to the aggregate state with tightly folded structure, slowly adsorbing at the interfaces, generating films with lower mechanical properties, and exposing less cutting sites. Physical fields and pH shifting have certain synergistic effects to efficiently tune the structure and redesign the interfacial layer of plant proteins, further enhancing their foaming or emulsifying properties. The improvement mechanisms mainly include: i) Aggregated plant proteins are depolymerized to form small protein particles and flexible structure is more easily exposed by combination treatment; ii) Particles with appropriate surface properties are quickly adsorbed to the interfacial layer, and then unfolded and rearranged to generate a tightly packed stiff interfacial layer to enhance bubble and emulsion stability; and iii) The unfolding and rearrangement of protein structure at the interface may result in the exposure of more cutting sites of digestive enzymes. This review summarizes the latest research progress on the structural changes, interfacial behaviors, and digestion properties of plant proteins under combined treatment, and elucidates the future development of these modification technologies for plant proteins in the food industry.

Reporter Coxsackievirus A5 Expressing iLOV Fluorescent Protein or Luciferase Used for Rapid Neutralizing Assay in Cells and Living Imaging in Mice.

Coxsackievirus A5 (CV-A5) is a re-emerging enterovirus that causes hand, foot, and mouth disease in children under five years of age. CV-A5-M14-611 is a mouse-adapted strain that can infect orally and lead to the death of 14-day-old mice. Here, recombinants based on CV-A5-M14-611 were constructed carrying three reporter genes in different lengths. Smaller fluorescent marker proteins, light, oxygen, voltage sensing (iLOV), and nano luciferase (Nluc) were proven to be able to express efficiently in vitro. However, the recombinant with the largest insertion of the red fluorescence protein gene (DsRed) was not rescued. The construction strategy of reporter viruses was to insert the foreign genes between the C-terminus of VP1 and the N-terminus of 2A genes and to add a 2A protease cleavage domain at both ends of the insertions. The iLOV-tagged or Nluc-tagged recombinants, CV-A5-iLOV or CV-A5-Nluc, exhibited a high capacity for viral replication, genetic stability in cells and pathogenicity in mice. They were used to establish a rapid, inexpensive and convenient neutralizing antibody assay and greatly facilitated virus neutralizing antibody titration. Living imaging was performed on mice with CV-A5-Nluc, which exhibited specific bioluminescence in virus-disseminated organs, while fluorescence induced by CV-A5-iLOV was weakly detected. The reporter-gene-tagged CV-A5 can be used to study the infection and mechanisms of CV-A5 pathogenicity in a mouse model. They can also be used to establish rapid and sensitive assays for detecting neutralizing antibodies.

Aggregation kinetics of green tea nanoparticles: Effects of pH, metal ions, and temperature.

Colloidal nanoparticles in tea infusion are the link connecting micromolecular mechanism and macro-aggregation process of tea cream formation. In order to elucidate, the kinetics mechanism of green tea nanoparticles (gTNPs) aggregation, zeta-potentials, total average aggregation (TAA) rates, and critical coagulation concentration (CCC) in the presence of various pH and metal ions were investigated. Additionally, the effect of temperature on gTNPs aggregation was further explored. The results revealed that the TAA rate of gTNPs increased with decreasing pH values, the CCC of gTNPs increased in the order Mg2+  ≈ Ca2+  < Na+  ≈ K+ . The reason was that different positive ions changed the surface electric field strength of gTNPs to a different extent. Furthermore, it was indicated that low temperature could promote gTNPs aggregation in indirect way. Low temperature promoted the binding of epigallocatechin gallate (EGCG) and caffeine, and the combination between gTNPs and EGCG-caffeine complexes weakened the stability of gTNPs resulting from reduction in electrostatic repulsion. PRACTICAL APPLICATION: Tea is a popular beverage all over the world. This research revealed the mechanism of green tea nanoparticles aggregation and laid a theoretical foundation for the regulation of tea cream formation in tea beverage.

Dynamic Formation of Green Tea Cream and the Identification of Key Components Using the "Knock-Out/Knock-In" Method.

The composition of green tea cream is extremely complex, and identification of key components is a prerequisite for elucidating its microstructure formation mechanism. This study examined the dynamic changes in the content of components and properties of colloid particles during the formation process of tea cream by chemical analysis and dynamic laser scattering (DLS). A "knock-out/knock-in" method was developed and used to further explore the relationship between the interaction of these components and the microstructure formation of tea cream. The results revealed that polysaccharides, proteins, epigallocatechin gallate (EGCG), and caffeine were the main components involved in tea cream formation. These components participated in the formation process in the form of polysaccharide-protein and EGCG-caffeine colloidal particles. Consequently, there were synchronized dynamic changes in the levels of polysaccharides, proteins, EGCG, and caffeine. The "knock-out/knock-in" experiment revealed that the interactions between EGCG or caffeine and macro-molecule components were not the key factors in tea cream microstructure formation. However, it was found that the complexation between EGCG and caffeine played a crucial role in the formation of tea cream. The findings suggested that decreasing the concentrations of EGCG and caffeine could be useful in controlling tea cream formation during tea beverage processing and storage.

Effects of epigallocatechin gallate, caffeine, and their combination on fat accumulation in high-glucose diet-fed Caenorhabditis elegans.

Epigallocatechin gallate (EGCG) and caffeine are inevitable to be ingested together in the process of drinking green tea. This study used Caenorhabditis elegans as an organism model to examine whether the binding of EGCG and caffeine could influence the fat-reduction effect. The results revealed that EGCG significantly reduced the Nile Red fluorescence intensity and the triglyceride/protein ratio of the C. elegans obesity model by 14.7% and 16.5%, respectively, while the effect of caffeine was not significant. Moreover, the degree of reduction in fluorescence intensity and triglyceride/protein ratio by EGCG + caffeine was comparable to that of EGCG. In the exploration of underlying mechanism, we found that EGCG and EGCG + caffeine treatments had no influence on food intake and energy expenditure of C. elegans. Their fat-reduction effects were dependent on the regulation of lipogenesis, as shown by the decreased expression of the sbp-1, fat-7, and daf-16 genes.

Identification of a Conserved, Linear Epitope on VP3 of Enterovirus A Species Recognized by a Broad-Spectrum Monoclonal Antibody.

Outbreaks of hand, foot and mouth disease (HFMD) have occurred frequently in the Asian-Pacific region over the last two decades, caused mainly by the serotypes in Enterovirus A species. High-quality monoclonal antibodies (mAbs) are needed to improve the accuracy and efficiency of the diagnosis of enteroviruses associated HFMD. In this study, a mAb 1A11 was generated using full particles of CV-A5 as an immunogen. In indirect immunofluorescence and Western blotting assays, 1A11 bound to the viral proteins of CV-A2, CV-A4, CV-A5, CV-A6, CV-A10, CV-A16, and EV-A71 of the Enterovirus A and targeted VP3. It has no cross-reactivity to strains of Enterovirus B and C. By mapping with over-lapped and truncated peptides, a minimal and linear epitope 23PILPGF28 was identified, located at the N-terminus of the VP3. A BLAST sequence search of the epitope in the NCBI genus Enterovirus (taxid: 12059) protein database indicates that the epitope sequence is highly conserved among the Enterovirus A species, but not among the other enterovirus species, first reported by us. By mutagenesis analysis, critical residues for 1A11 binding were identified for most serotypes of Enterovirus A. It may be useful for the development of a cost-effective and pan-Enterovirus A antigen detection for surveillance, early diagnosis and differentiation of infections caused by the Enterovirus A species.

Co-Extraction of Flaxseed Protein and Polysaccharide with a High Emulsifying and Foaming Property: Enrichment through the Sequence Extraction Approach.

A new focus with respect to the extraction of plant protein is that ingredient enrichment should target functionality instead of pursuing purity. Herein, the sequence aqueous extraction method was used to co-enrich five protein-polysaccharide natural fractions from flaxseed meal, and their composition, structure, and functional properties were investigated. The total recovery rate of flaxseed protein obtained by the sequence extraction approach was more than 80%, which was far higher than the existing reports. The defatted flaxseed meal was soaked by deionized water to obtain fraction 1 (supernatant), and the residue was further treated to get fraction 2 (supernatant) and 3 (precipitate) through weak alkali solubilization. Part of the fraction 2 was taken out, followed by adjusting its pH to 4.2. After centrifuging, the albumin-rich supernatant and precipitate with protein content of 73.05% were gained and labeled as fraction 4 and fraction 5. The solubility of fraction 2 and 4 exceeded 90%, and the foaming ability and stability of fraction 5 were 12.76 times and 9.89 times higher than commercial flaxseed protein, respectively. The emulsifying properties of fractions 1, 2, and 5 were all greater than that of commercial sodium caseinate, implying that these fractions could be utilized as high-efficiency emulsifiers. Cryo-SEM results showed that polysaccharides in fractions were beneficial to the formation of network structure and induced the formation of tighter and smoother interfacial layers, which could prevent emulsion flocculation, disproportionation, and coalescence. This study provides a reference to promote the high-value utilization of flaxseed meals.

Identification of specific and shared epitopes at the extreme N-terminal VP1 of Coxsackievirus A4, A2 and A5 by monoclonal antibodies.

Hand, foot and mouth disease (HFMD) is caused by a variety of serotypes in species A of the Enterovirus genus, including recently re-emerged Coxsackievirus A2 (CV-A2), CV-A4 and CV-A5. For development of diagnostic reagents, for surveillance, and the development of multivalent vaccines against HFMD, the antigenicity of HFMD-associated enteroviruses warrants investigation. The purified virions of CV-A4 were inoculated into Balb/c mice and hybridomas were obtained secreting monoclonal antibodies (mAbs) directed against CV-A4 and cross-reacting with other closely related species A enteroviruses. The mAbs were characterized by ELISA, Western blotting and in vitro neutralizing assays. The majority of mAbs was non-neutralizing, with only 2% of the mAbs neutralizing CV-A4 specifically. Most of mAbs bound to linear VP1 epitopes of CV-A4. Interestingly, four types of mAbs were obtained which bound specifically to CV-A4 or were broadly to CV-A4/-A2, CV-A4/-A5 and CV-A4/-A2/-A5, respectively. Mapping with overlapping or single-amino-acid mutant peptides revealed that the four types of mAbs all bound to the first 15 amino acids at the N-terminus of the VP1. This region of picornaviruses is functionally important as it is involved in uncoating and releasing of viral RNA into the cytosol. The binding footprints of four type mAbs are composed of conserved and variable residues and are different from each other. The newly discovered broadly cross-reactive mAbs reflect the high homology of CV-A4/ CV-A2/CV-A5. The results also demonstrate that it is possible and beneficial to develop the diagnostic reagents to detect rapidly the main pathogens of enteroviruses associated with HFMD cause by CV-A4/CV-A2/CV-A5.

Formation, digestion properties, and physicochemical stability of the rice bran oil body carrier system.

Rice bran is a major by-product of rice processing with abundant nutrient content. Oil bodies (OBs), which are fat particles with unique physicochemical stability, are specialized organelles for the storage of oils and fats in plant tissues. In this study, we extracted OBs from rice bran, to evaluate the function of hydrophobic nutrients efficiently delivered by OBs. The carrier system was prepared by sonicating curcumin with medium chain triglycerides (MCT) into rice bran oil bodies (RBOBs). Emulsions comprising different RBOB mass fractions were characterized. The results showed that the highest encapsulation efficiency (EE, 87.67%), optimal particle size (190 nm), and best storage stability were achieved with the 1.5 wt% RBOBs. Based on activity evaluation data, the carrier system can achieve sustained oil release in the intestine and shows high bioaccessibility (61.04%; IC50 in Caco-2 cells was 77.21 µg/mL), which is important for promoting grain by-product utilization.

Improve stability and application of rice oil bodies via surface modification with ferulic acid, (-)-epicatechin, and phytic acid.

Rice bran oil bodies (RBOBs) are one of the most exploited functional components from rice bran by-products and are predominantly based on oleosin stabilization. In this study, we explored the effects of different concentrations of added (-)-epicatechin, ferulic acid, and phytic acid on the RBOBs stability. The results revealed that the incorporation of all three natural phytoconstituents could reduce the RBOBs particle size and increase emulsifying properties, demonstrating increasing surface hydrophobicity (p < 0.05), and a good antioxidant effect, which was especially obvious with (-)-epicatechin incorporation. Fourier transform infrared (FT-IR) spectroscopy data demonstrated that these three small molecule substance classes can modify with oleosin on RBOBs surface by covalent and noncovalent effects. Raman spectroscopic analysis illustrated that the vibrational modes of disulphide bonds in oleosin were modified by these three plant natural ingredients. The interactions between the three phytoconstituents and the model protein were investigated by molecular docking experiments.

Modulating interfacial structure and lipid digestion of natural Camellia oil body by roasting and boiling processes.

Oil body (OB) is the lipid-storage organelle in oilseed, and its stability is crucial for oilseed processing. Herein, effects of roasting and boiling on the structure, stability, and in vitro lipid digestion of Camellia OB were studied. The interfacial structure and physical stability of the extracted OB were investigated by electrophoresis, confocal-Raman spectroscopy, zeta-potential, and surface hydrophobicity, etc. Boiling caused protein loss on the OB surfaces, forming a stable phospholipid interface, which resulted in coalescence of the droplets (d > 100 µm) and negative ζ-potential (-3 âˆ¼ -8 mV) values at a pH of 2.0. However, roasting partially denatured the proteins in the seeds, which were adsorbed on the OB surfaces. The random coil structure of interfacial protein increased to ∼20 % after thermal treatment. Besides, heating decreased the surface hydrophobicity of OB and improved lipid digestion. After boiling 60 min, the extent of lipolysis increased from 41.7 % (raw) to 57.4 %.

Structural Transitions of Alpha-Amylase Treated with Pulsed Electric Fields: Effect of Coexisting Carrageenan.

Pulsed electric field (PEF) is an effective way to modulate the structure and activity of enzymes; however, the dynamic changes in enzyme structure during this process, especially the intermediate state, remain unclear. In this study, the molten globule (MG) state of α-amylase under PEF processing was investigated using intrinsic fluorescence, surface hydrophobicity, circular dichroism, etc. Meanwhile, the influence of coexisting carrageenan on the structural transition of α-amylase during PEF processing was evaluated. When the electric field strength was 20 kV/cm, α-amylase showed the unique characteristics of an MG state, which retained the secondary structure, changed the tertiary structure, and increased surface hydrophobicity (from 240 to 640). The addition of carrageenan effectively protected the enzyme activity of α-amylase during PEF treatment. When the mixed ratio of α-amylase to carrageenan was 10:1, they formed electrostatic complexes with a size of ~20 nm, and carrageenan inhibited the increase in surface hydrophobicity (<600) and aggregation (<40 nm) of α-amylase after five cycles of PEF treatment. This work clarifies the influence of co-existing polysaccharides on the intermediate state of proteins during PEF treatment and provides a strategy to modulate protein structure by adding polysaccharides during food processing.