Search Results (472)

A chickpea-based milk beverage containing both plant and animal proteins represents an excellent substrate for the production of biologically active peptides through fermentation. Fermentation by lactic acid bacteria (LAB) increases its nutritional value compared to the unfermented beverage while improving the digestibility and bioavailability of essential nutrients via proteolytic enzyme activity. This study investigated the production of bioactive peptides in fermented chickpea water extract using ten bacterial strains isolated from plant and animal sources. The proteolytic activity of each strain was quantified using the trinitrobenzene sulfonic acid (TNBS) method, and the presence of proteolytic genes was confirmed via agarose gel electrophoresis. Peptides released during fermentation were identified through two-dimensional electrophoresis, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), and tandem mass spectrometry. To predict the potential biological activities of the studied peptide sequences, a series of in silico analyses were performed using specialized bioinformatics tools. The identified peptides were predicted to exhibit antioxidant, аntihypertensive, anticancer, antibacterial, antifungal, antituberculosis, and angiotensin-converting enzyme (ACE) inhibitory activities. Based on the results, L. fermentum SB-2 and L. sakei SD-8, were selected as promising candidates for bioactive peptide production in a chickpea water extract-based milk beverage and were subsequently applied in the beverage prototype. Full article

The gut microbiota (GM) has become a key mediator of host health, with dietary manipulations promising ways of modulating the microbiome. This review focuses on the role of dairy bioactive (DB) compounds as precision modulators of intestinal microecology, including the whey proteins (WPs), including lactoferrin (LF), α-lactalbumin (LA), β-lactoglobulin, lysozyme (LZ), lactoperoxidase, glycomacropeptide (GMP), milk oligosaccharides (MOs), and bioactive peptides (BPs). This review compiles the existing evidence illustrating their dual-action mechanism through direct prebiotic activity and the promotion of beneficial taxa (Bifidobacterium, Lactobacillus, Faecalibacterium), along with selective antimicrobial activity and pathogen suppression. These compounds improve intestinal barrier integrity through tight junction (TJ) protein regulation, regulating short-chain fatty acid production, and modulating immune signaling pathways. Clinical evidence shows significant benefits in metabolism and inflammation among various populations. However, individual responses vary according to host factors such as enterotypes, FUT2 genotype, and baseline microbiota composition, suggesting the need for personalized intervention strategies. This review addresses critical knowledge gaps in dose–response relationships, long-term efficacy, and mechanistic pathways and suggests future directions for precision nutrition. By modifying molecular mechanisms in clinical applications, we have identified DB compounds as promising candidates for targeted modulation of the microbiota to optimize health and disease management. The review also brings together molecular mechanistic and clinically implementable, personalized dietary strategies, which have not been fully captured by previous reviews. It pinpoints gaps in knowledge related to dose–response characterization, long-term trial design, and multi-omics stratification that collectively define a new precision nutrition framework. In this approach, dairy-based intervention is planned for each person based on their microbial, genetic, and metabolic characteristics. Full article

Proteases, enzymes that catalyze the hydrolysis of peptide bonds in peptides and proteins, have widespread industrial applications, particularly in milk coagulation for cheese production. Microbial enzymes have been employed as alternatives to animal rennet, offering advantages such as cost-effectiveness, availability, and compliance with dietary, cultural, and religious requirements. Solid-state fermentation (SSF) is widely employed for microbial enzyme production because of its low operational costs, reduced water and energy requirements, high product concentrations, and the ability to utilize agro-industrial residues as low-cost substrates, thereby contributing to both process sustainability and waste valorization. We report the production and characterization of a novel milk-clotting protease produced by Trichoderma koningii FFT13. The protease was produced by SSF using wheat bran as the substrate, an agro-industrial residue. It was classified as a chymotrypsin-like serine protease and exhibited a specific caseinolytic activity of 9861 U/mg. The enzyme coagulated both reconstituted skim milk and pasteurized whole milk in the presence or absence of calcium. Coagulation was enhanced by increasing temperature, reaction time, enzyme concentration, and calcium levels. Scanning electron microscopy revealed destabilization of casein micelles, their progressive aggregation, and the formation of a well-defined gel network, confirming the effectiveness of the protease in milk coagulation. Therefore, these results demonstrate that the chymotrypsin-like protease from T. koningii is a promising enzyme for milk coagulation, with potential application in cheese production. The enzyme obtained constitutes an alternative to traditional coagulants, overcoming limitations related to animal rennet while potentially offering additional advantages in terms of process sustainability and industrial scalability. Full article

This review aims to examine the immunological, anti-inflammatory, antiviral, and antibacterial activities of key whey and milk proteins, specifically lactoferrin, glycomacropeptide, β-lactoglobulin, α-lactalbumin and their derived peptides, particularly lactoferricin and lactoferrampin, highlighting their potential as preventive or therapeutic agents. Whey and dairy products represent complex biological matrices that, beyond their high nutritional value, serve as reservoirs of bioactive proteins and peptides with documented health-promoting properties. It has been reported that certain whey proteins (WPs) and whey-derived peptides may contribute to improvements in both innate and adaptive immunity, exert direct antiviral and antibacterial effects while also modulating host defenses through immunoregulatory, antioxidant, and anti-inflammatory activities. These mechanisms contribute not only to enhanced resistance against viral pathogens but also to maintaining intestinal homeostasis and microbiota balance, both of which are critical during infection. In recent years, particularly in the context of the COVID-19 pandemic, natural bioactive compounds derived from whey, and, more broadly, milk, have attracted increasing attention as potential adjuncts or alternatives to conventional antivirals, with reported activity not only against SARS-CoV-2, influenza but also other viral and microbial infections. Despite encouraging in vitro and in vivo evidence, clinical validation remains limited, and the antiviral and immunomodulatory effects of WPs still require deeper mechanistic clarification. Future research should focus on identifying molecular targets, as well as characterizing the pharmacokinetics and safety profiles of WPs and WP peptides across diverse clinical settings. At the same time, attention should be given to optimizing their application as nutraceuticals or functional dairy ingredients. Full article

Background: The translocation of diet-derived antigens from the maternal intestine to breast milk represents a primary gateway for neonatal immune priming, yet the structural basis for why certain proteins survive this transit while others do not remains poorly understood. This review introduces the “Survivor Peptide” hypothesis, proposing that specific food allergens possess intrinsic “stability architectures” that enable them to resist maternal digestion and navigate the gut–mammary axis to reach the infant in an immunologically active form. Methods: We analyzed the current literature regarding the detection and structural characteristics of food allergens in human milk. Integrating evidence from 26 major sources, we performed an in silico structural analysis of five representative “survivor” proteins: Gal d 1 (egg white), Bos d 5 (cow’s milk), Gal d 6 (egg yolk), Tri a 19 (wheat), and tropomyosin (Der p 10-mite/shellfish). High-resolution 3D models were retrieved from the Protein Data Bank and AlphaFold2, and then visualized in UCSF ChimeraX to map stability anchors, including disulfide bonds and hydrophobic clusters, against solvent-accessible IgE-binding epitopes. Results: We identified and categorized allergens into distinct Molecular Resilience Architectures: the “Covalent Cage” (Gal d 1), defined by dense disulfide stapling, the “Glycoprotein Shield” (Gal d 6), utilizing yolk-matrix structural anchors, the “Topological Shield” (Bos d 5), characterized by a stable β-barrel, and “Coiled-Coil Rigidity” (Der p 10). These frameworks protect large, immunogenic fragments that maintain the spatial arrangement required for IgE cross-linking. Conclusions: Allergen persistence in the gut–mammary axis is dictated by a protein’s intrinsic structural architecture. Identifying these stability fingerprints provides a unified theory for allergen persistence and offers a path for refining component-resolved diagnostics and neonatal oral tolerance strategies. Full article

The INFOGEST protocol is a standardised in vitro digestion model widely utilised to evaluate the digestibility and bioaccessibility of nutrients in diverse food matrices. This review focuses on its application since 2020 (after the publication of the INFOGEST 2.0 model) to milk and dairy products, which often serve as a suitable food matrix in digestion studies. By analysing 50 studies selected using a semi-automated method, this review highlights its strong performance in reproducing general digestive trends, including peptide fingerprint profiling, consistent high-protein digestibility, and matrix-dependent lipid and mineral bioaccessibility. The model is particularly effective in evaluating structural modifications of dairy products and their impact on digestive behaviour. However, its application remains skewed toward bovine systems, limiting broader relevance to other dairy matrices. Methodological variability, including protocol modifications and emerging semi-dynamic adaptations, poses challenges to reproducibility. Furthermore, reliance on simplified downstream models constrains the physiological interpretation of bioactivity and nutrient absorption. Future progress requires harmonised dynamic extensions, expanded use of advanced biological systems, and inclusion of diverse dairy matrices. Collectively, these advances will support a shift from descriptive bioaccessibility toward more predictive assessments of nutrient bioavailability. This six-year, non-topic-dependent bibliometric analysis contextualises the expanding adoption of INFOGEST 2.0 as reflected in its versatility and evolving scope, positioning it as a cornerstone tool for advancing our understanding of dairy nutrition, digestion-derived bioactivity, and ultimately, the relationship between dairy consumption and human health. Full article

β-Casein A1 and A2 (β-CN-A1 and β-CN-A2) are the two predominant β-CN proteoforms in bovine milk. β-CN-A1 has been associated with a greater propensity to release opioid peptides, such as β-casomorphin-7 (β-CM-7) and β-casomorphin-5 (β-CM-5), during gastrointestinal (GI) digestion, which may have adverse biological effects. This has stimulated growing interest in milk from cows carrying the β-CN A2A2 genotype (A2 milk), which requires reliable characterization methods. In this work, we developed a rapid, selective, and sensitive capillary electrophoresis–mass spectrometry (CE-MS) method for the accurate identification and quantification of β-CM-7 and β-CM-5 in milk hydrolysates from in vitro GI digestion of bovine milk. The method showed good linearity (R² > 0.99, over 0.5–100 mg/L for β-CM-7 and 0.25–100 mg/L for β-CM-5), limits of detection (0.25 and 0.10 mg/L), and repeatability (<0.2% for times and <1.4% for areas), and tandem mass spectrometry (MS/MS) allowed confirmation. The method was applied to A1A1 and A2A2 milk digested using the standardized INFOGEST protocol, followed by solid-phase extraction. β-CM-7 was detected and quantified only in A1A1 digests (0.98 mg/L), whereas β-CM-5 was not detected (<0.10 mg/L). These results indicate a differential release of β-CMs from A1 and A2 milk and support the method’s suitability for β-CM profiling, which may help assess A2 milk quality control and β-CM health impact. Full article

Buffalo milk is a rich source of various nutritional components and bioactive peptides, offering significant health benefits. Endogenous peptides, which occur naturally in milk, represent a valuable source of bioactive peptides with potential nutraceutical applications. However, research on endogenous peptides in buffalo milk remains limited. This study employed a quantitative peptidomic approach to characterize endogenous peptides across different lactation stages. A total of 2099, 2946, and 4418 peptides were identified in colostrum, transitional milk, and mature milk, respectively. The majority of these peptides were derived from β-casein, followed by α_S1-casein, κ-casein, and other proteins. Notably, variations in precursor proteins contributing to peptide production were observed throughout lactation. Phosphorylation levels of endogenous peptides were highest in mature milk, with serine residues predominating. Enzymatic cleavage analysis identified cathepsin D as the key enzyme involved in endogenous peptide production, while proline endopeptidase and plasmin exhibited stage-specific activities. Bioinformatics analysis revealed differentially expressed precursor proteins linked to complement cascades and NF-κB signaling, emphasizing the immune protective role of colostrum. Furthermore, 54 potentially bioactive peptides with favorable water solubility were identified in colostrum, of which 17 were predicted to possess anti-inflammatory properties. These findings contribute to a deeper understanding of the molecular basis of buffalo milk’s functional properties, highlighting its potential as a source of bioactive peptides for both nutritional and pharmaceutical applications. Full article

Riceberry rice milk (RBRM) is rich in phytochemicals, particularly anthocyanins, which are known for their potential in managing type 2 diabetes (T2D). This study aimed to develop a novel RBRM-based yogurt derived from its polysaccharide and protein components and to evaluate the effects of supplementation with W. globosa powder (WGP) at 0% (F1, control), 5% (F2), 10% (F3), and 15% (F4) on nutritional and functional properties. Among all formulations, F4 exhibited the highest nutritional values, including dietary fiber (41.25%), curd protein (21.34%), and carbohydrate (starch) content (25.25%), with a lower fat content (2.13%) compared to other groups. In terms of antioxidant activity, F4 showed high total phenolic content (33.70 mg GAE/g) and total flavonoid content (25.2 mg QUE/g), along with strong radical scavenging activities, with DPPH and ABTS inhibition values of 41.52% and 78.18%, respectively. Furthermore, F4 demonstrated notable antidiabetic potential through α-amylase and α-glucosidase inhibition, with IC₅₀ values of 0.89 and 1.32 mg/mL, respectively. Widely targeted metabolomics analysis identified 88 differential metabolites between F4 (potent condition) and F1 (control group). Twelve selected compounds from RBRM–WGP yogurt contributed to increased levels of amino acids, peptide derivatives, saccharides, organic acids, polyphenols, and flavonoids. Molecular docking analysis revealed that key metabolites, including vignatic acid B, glimepiride, and indoramin, exhibited strong binding affinities with the active sites of α-amylase (PDB: 2GVY, Aspergillus niger) and α-glucosidase (PDB: 3A4A, Saccharomyces cerevisiae). These findings indicate that phytonutrient compounds, particularly indoramin, play a significant role in enhancing the nutritional composition and functional properties of RBRM–WGP yogurt for potential applications in food processing. Full article

Hypertension is a severe global public health issue. Food-derived angiotensin-converting enzyme (ACE)-inhibitory peptides have shown great potential as safe and effective alternatives to synthetic antihypertensive drugs. Camel milk is rich in bioactive peptides. This study aimed to screen for ACE-inhibitory peptides from hydrolyzed camel casein, explore their inhibitory mechanisms and endothelial protective effects in vitro, and reveal their potential antihypertensive pathways using network pharmacology. This study screened three peptides with angiotensin-converting enzyme (ACE) inhibitory activity from enzymatically hydrolyzed camel casein components: MVPFLQPK, VPFLQPKVM, and QKWKFL, with IC₅₀ values of 277.1, 396.9, and 486.9 μmol/L, respectively. Enzyme inhibition kinetics analysis indicated that MVPFLQPK exhibited a non-competitive inhibition pattern, VPFLQPKVM exhibited a mixed inhibition pattern, and QKWKFL exhibited a competitive inhibition pattern. Molecular docking revealed that all three peptides formed hydrogen bond interactions with ACE, and QKWKFL and VPFLQPKVM directly bound to the enzyme’s active site to inhibit substrate catalysis. Molecular dynamics simulation further confirmed the high stability of the three peptide–ACE complexes, with binding free energies from −34.24 to −51.19 kcal/mol. The primary contributing forces include hydrogen bonds, van der Waals interactions, electrostatic forces, and nonpolar solvation effects. Network pharmacology analysis suggested that these peptides may exert synergistic antihypertensive effects by regulating multiple blood pressure-related pathways, including the renin–angiotensin system, renin secretion, and calcium signaling pathways, by acting on key targets such as ACE, REN, SRC, and MMP9. Cell experiments demonstrated that all three peptides exhibited no cytotoxicity in the Ang II-induced HUVEC injury model, significantly promoted NO release, inhibited ET-1 secretion, and possessed endothelial protective potential. This study investigated the in vitro ACE-inhibitory mechanism of peptides derived from camel milk and their potential role in blood pressure regulation, providing experimental evidence for subsequent in vivo activity validation and the development of functional camel milk protein products. Full article

This study investigates the antioxidant properties of several synthetic peptides derived from milk proteins previously identified in milk permeate, a by-product of the dairy industry. The aim of the research is to identify which peptides present in milk permeate are responsible for its antioxidant activity. A comprehensive experimental strategy was employed to evaluate their antioxidant potential, including in silico selection, in vitro free radical scavenging assays and cellular models using Caco-2 and HCT116 cell lines. The peptides were screened using a molecular docking approach for their potential interaction with the Kelch-like ECH-associated protein 1/nuclear factor erythroid 2-related factor 2 (Keap1/Nrf2) pathway, and eight out of twenty-eight were selected and synthesized for further analyses. In vitro, six of the selected peptides demonstrated significant direct antioxidant activity in the DPPH scavenging assay, and two in the ABTS scavenging test. In cellular environments, three peptides (LPAPELGPRQA, LPIIQKLEPQI and NGQVWEESLKRL) effectively protect cells from oxidative stress induced by tert-butyl hydroperoxide, reducing reactive oxygen species production and partially mitigating lipid peroxidation. Further investigation showed that two of them (LPAPELGPRQA and LPIIQKLEPQI) effectively induce the Keap1/Nrf2 pathway, as evidenced by a ∼1.5-fold increase in Nrf2 levels and overexpression of downstream proteins. Permeability studies revealed that these peptides can cross the intestinal monolayer (2–3% in 2 h), suggesting potential systemic effects. Overall, these findings highlight the multifunctional antioxidant properties of the investigated peptides and support their potential application as nutraceuticals or therapeutic agents for oxidative stress-related conditions. Full article

The rapid global spread of antimicrobial resistance among pathogenic microorganisms poses a serious challenge to both human and animal health, underscoring the urgent need for new strategies to combat resistance. Antimicrobial peptides (AMPs), key components of the innate immune system, are promising candidates because they disrupt the membranes of bacteria, fungi, and viruses, thereby reducing the risk of resistance development. Lactoferrin (LF), a multifunctional iron-binding glycoprotein abundant in mammalian milk, is a rich source of AMPs. Cationic peptide fragments such as lactoferricin and lactoferrampin exhibit more potent direct antimicrobial activity than the intact protein. Our previous studies have shown that peptides derived from Equine milk lactoferrin exhibit antihypertensive, anti-inflammatory, and anti-oncogenic activity in silico, highlighting their multifunctional bioactive potential. Building on these results, the present study aims to investigate the antimicrobial properties of these peptides. We used an integrated approach combining computer modeling and in vitro studies to identify and validate novel antimicrobial peptides from equine milk lactoferrin. Bioinformatics tools, including AMPScanner and CAMP, were used to predict antimicrobial domains, followed by experimental testing against Escherichia coli, Staphylococcus aureus, and Pseudomonas aeruginosa. The results showed that equine milk lactoferrin peptides possess potent and selective antimicrobial activity, with efficacy varying across bacterial species. These data expand the functional profile of lactoferrin-derived peptides, demonstrating their multifunctionality, and suggest that equine milk lactoferrin represents a promising natural source of antimicrobial agents, supporting alternative strategies to reduce antibiotic use in human and veterinary medicine. Full article

Fermented milk is rich in probiotics, peptides, vitamins, and minerals, which are used as routine food supplements and are of great benefit for regulating human health. This study explored the mechanism of Lactobacillus delbrueckii ssp. bulgaricus CGMCC 21287 or Lacticaseibacillus casei CGMCC 15956 fermented milk for alleviating acute alcoholic liver injury. We found that fermented milk was associated with reduced activation of TLR4/NF-κB pathways, alleviating alcohol-induced liver inflammation. Meanwhile, the two probiotics regulated different intestinal microbial communities in mice. The LC group specifically increased the abundance of probiotics such as Roseburia, unidentified_Lachnospiraceae, and Allobaculum, and decreased the abundance of pathogenic bacteria such as Enterococcus and Shigella. The LB group increased the abundance of Adlercreutzia and Ruminococcus, thereby increasing butyric acid, acetic acid, and valeric acid levels and decreasing lipopolysaccharide (LPS) production. These results suggest that daily intake of fermented milk can attenuate alcohol-induced acute liver injury in mice via the gut–liver axis, though differences exist in the mechanisms of action and areas of emphasis. Full article

Liquid infant formula has garnered increasing attention due to its mild thermal processing and superior retention of bioactive nutrients. Within such matrices, the lipid source is a critical determinant of protein digestion behavior, yet its influence on peptide bioavailability and intestinal homeostasis remains undefined. Given that efficient peptide absorption is vital for the systemic delivery of bioactivity in infants, understanding the lipid–protein synergy is essential for formula optimization. Moreover, excessive oxidative stress is closely associated with impaired intestinal health and developmental disorders in infants, making the regulation of oxidative stress crucial for maintaining intestinal function. The present study evaluated the effects of three distinct lipid sources—soybean oil (SM), bovine milk fat (BM), and goat milk fat (GM)—on the physicochemical stability, proteolytic digestion, peptide release, intestinal absorption, and oxidative stress modulation of goat-milk-based infant formula. An integrated approach combining physicochemical characterization, in vitro simulated infant digestion, and a Caco-2 intestinal epithelial cell model was employed. we demonstrate that all three lipids (3% w/w) formed stable emulsions with uniform spherical structures and mean particle diameters of 117–300 nm, as visualized by laser confocal microscopy. Following in vitro simulation of infant gastrointestinal digestion, the SM group exhibited the most extensive protein hydrolysis, yielding the highest total peptide content (4.28 ± 0.10 mg/mL) and generated the highest number of peptides identified by LC-MS/MS (474 types). Bioinformatic analysis predicted that peptides from all groups possess potential antihypertensive, hypoglycemic, and immunomodulatory activities. The Caco-2 monolayer cell model demonstrated that although the GM group produced fewer identified peptide species than the SM group (365 types), it achieved significantly higher intestinal peptide absorption rate (55.34 ± 1.05%). Furthermore, the GM digests provided superior protection against H₂O₂-induced oxidative stress in Caco-2 cells, markedly reducing reactive oxygen species levels and suppressing the expression of pro-inflammatory cytokines TNF-α and IL-6. Collectively, these findings reveal that while soybean oil promotes more extensive proteolysis, the use of homologous goat milk lipid enhances peptide bioaccessibility and confers potential cytoprotective effects on intestinal epithelial cells, underscoring its potential as a preferred lipid source in infant formula formulations. Full article

Insulin resistance is a multifactorial cellular state involving coordinated alterations in protein homeostasis and organelle function; however, its proteome-wide organization and response to bioactive peptides remain incompletely defined. In this study, we employed DIA-based quantitative proteomics to characterize global protein abundance changes associated with insulin resistance in HepG2 cells and to examine proteomic remodeling following treatment with a camel milk-derived peptide (P2). Comparative proteomic profiling revealed that insulin-resistant cells exhibit extensive reorganization of protein networks linked to redox regulation, endoplasmic reticulum protein processing, mitochondrial metabolism, lysosomal function, and extracellular matrix-associated components. Gene Ontology, KEGG pathway, protein domain, and subcellular localization enrichment analyses consistently indicated disruption of organelle-associated proteomic architecture rather than isolated pathway perturbations. Peptide TYYPPQ treatment was associated with selective, rather than global, proteomic shifts, prominently affecting mitochondrial and peroxisome-associated protein groups as well as extracellular and secretory proteins. Enrichment and localization analyses suggest that peptide exposure reshapes organelle-linked protein representation patterns without implying direct activation of signaling pathways or physiological restoration. Collectively, these results define insulin resistance and peptide responsiveness at a systems-level proteomic resolution and establish an organelle-resolved framework for interpreting peptide-induced proteomic remodeling in insulin-resistant hepatocyte models. This dataset provides a foundation for future targeted functional validation of candidate pathways identified through proteomic association. Full article