Search Results (68)

The prediction of polypeptide chain fragmentation during digestion (proteolysis) of protein substrates by trypsin was carried out for globular β-lactoglobulin (β-LG) and micellar β-casein (β-CN). Despite significant differences in the protein structures of these substrates, the concentrations of peptide fragments are calculated as functions of time or degree of hydrolysis using the same equations derived from the general proteolysis model. This model considers the opening of protein substrates in the course of proteolysis, the so-called demasking process, and the subsequent hydrolysis of specific peptide bonds at different rates determined by the amino acid sequence of hydrolyzed sites. The use of this model for in silico prediction of proteolysis is discussed. An algorithm for calculating demasking rate constants based on the experimental distribution of peptide fragments is presented. The calculated concentration dependence on the degree of hydrolysis of peptide bonds was compared with the experimental data for the intermediate and final peptide fragments of β-LG and β-CN. The predicted and experimental concentration curves for the final products were compared based on their curvatures. For both substrates, the predicted redistribution of peptide fragments in the course of proteolysis was found to be consistent with the experimental one. Full article

This study aimed to formulate a carrier system to improve the oral bioaccessibility of goat casein peptides (GCAPS). Goat casein was hydrolyzed with papain and subsequently purified to obtain bioactive peptide fractions (GCAPS) with potent hypoglycemic activity. On this basis, spherical GCAPS-loaded nanocarrier systems were developed, including liposomes (GCAPS-LS) and niosomes (GCAPS-NS). Among them, GCAPS-NS exhibited higher encapsulation efficiency (94.98 ± 3.01%) and a smaller particle size (89.81 ± 8.97 nm) than GCAPS-LS. FT-IR analysis confirmed successful peptide encapsulation. Simulated gastrointestinal digestion experiments demonstrated that GCAPS-NS significantly improved GCAPS retention and DPP-IV inhibition. In vivo results from high-fat diet-induced (HFD) insulin-resistant mice demonstrated that GCAPS-NS effectively ameliorated metabolic abnormalities by including adiposity, enhancing GLP-1 levels and suppressing hsCRP expression, thereby contributing to improved glycemic homeostasis. Moreover, GCAPS-NS intervention resulted in a significant enrichment of Akkermansia and a reduced Firmicutes/Bacteroidetes ratio, suggesting its beneficial role in alleviating HFD gut dysbiosis. These findings indicated that goat casein peptides held great potential as a functional food for the management of type 2 diabetes. Full article

Background: Skin aging is mainly associated with oxidative stress and enzymatic degradation of collagen and elastin by protease activity. Peptides have antioxidant capacity and inhibitory effects on protease enzymes. Objective: The purpose of this study was to obtain peptides with in vitro anti-aging activity from the enzymatic hydrolysis of bovine casein with actinidin, a protease extracted from the green kiwi fruit (Actinidia deliciosa) Methodology: The enzyme actinidin was extracted from the pulp of the kiwi fruit, purified by ion exchange chromatography and characterized by polyacrylamide electrophoresis (SDS-PAGE). Subsequently, the extracted enzyme was used to hydrolyze commercial bovine casein at 37 °C for 30 min, precipitating the peptide fraction with trichloroacetic acid (TCA), and centrifuged. To determine the anti-aging potential of the peptides in vitro, antioxidant activity was evaluated using the ABTS (2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)) radical. Additionally, the inhibitory capacity of the peptides against collagenase and elastase enzymes was also studied. To complement the in vitro results, the enzymatic hydrolysis of casein with actinidin was simulated. The binding energy (ΔG) of each of the hydrolysates with the collagenase and elastase enzymes was calculated using molecular docking to predict the peptide sequences with the highest probability of interaction. Results: Actinidin was extracted and purified exhibiting a molecular weight close to 27 kDa. The enzyme hydrolyzed the substrate by 91.6%, and the resulting hydrolysates showed moderate in vitro anti-aging activity: antioxidant (17.5%), anticollagenase (18.55%), and antielastase (28.6%). In silico results revealed 66 peptide sequences of which 30.3% consisted of 4–8 amino acids, a suitable size to facilitate interaction with structural targets. The sequences with the highest affinity were FALPQYLK and VIPYVRYL for collagenase and elastase, respectively. Conclusions: Despite the modest inhibition values, the use of a fruit-derived enzyme and a food-grade substrate is in line with current trends in sustainable and natural cosmetics. These findings highlight the great potential for laying the groundwork for future research into actinidin-derived peptides as multifunctional and eco-conscious ingredients for the development of next-generation anti-aging formulations. Full article

Magnaporthe oryzae (M. oryzae) is a phytopathogenic fungus that inflicts damage on vital crops, particularly rice. Its asexual reproduction leads to the generation of numerous conidia, which is a critical factor contributing to the prevalence of rice blast disease. However, the molecules regulating the asexual reproduction of M. oryzae are unknown. In our study, to identify the molecules capable of regulating the asexual reproduction of M. oryzae, compositions of the complete medium (CM) were screened. Results showed that acid-hydrolyzed casein (AHC) could remarkably promote conidial production. One M. oryzae conidiation inducer was isolated from AHC using high-performance liquid chromatography (HPLC) under the guidance of bioassay. Its structure was further elucidated as a decapeptide compound (pyroGlu-EQNQEQPIR) by LC-MS/MS, chemical synthesis, and conidium-inducing assays, named M. oryzae conidiation inducer decapeptide (MCIDP). MCIDP could significantly promote the conidiation of M. oryzae and two other filamentous ascomycetes (Botrytis cinerea and Fusarium graminearum). The Mps1 MAPK cascade signaling pathway is crucial for conidiation, and the effect of MCIDP on this pathway was investigated to elucidate the mechanism underlying conidiation enhancement. qRT-PCR analysis demonstrated that MCIDP could remarkably upregulate the gene expression within the Mps1 MAPK cascade signaling pathway, especially the WSC2, WSC3, PKC1, MKK1, MPS1, and MIG1. Furthermore, the ΔMowsc1, ΔMowsc2, ΔMowsc3, and ΔMomid2 mutant strains were constructed. Bioassay results showed that MCIDP failed to promote conidial formation and hyphal growth in these mutant strains. These findings indicate that MCIDP promotes conidiation of M. oryzae by modulating the Mps1 MAPK signaling pathway. Full article

Caragana intermedia is a perennial shrub species in the genus Caragana (Fabaceae), demonstrating remarkable stress resistance and adaptability. However, research on its somatic embryogenesis (SE) and genetic transformation techniques remains limited. In this study, we established an SE system by utilizing immature cotyledons isolated from young C. intermedia seeds. Our findings demonstrated that the immature cotyledons at 6–7 weeks after flowering (WAF) were the best explants for SE. The optimal embryo induction medium consisted of an MS basal medium supplemented with 5 mg/L α-naphthaleneacetic acid (NAA), 3 mg/L 6-benzylaminopurine (6-BA), 30 g/L sucrose, 7 g/L agar, and 500 mg/L hydrolyzed casein. Cotyledon-stage embryos germinated on a half-strength MS medium, exhibiting a 34.36% germination rate. Based on the SE system, we developed a preliminary genetic transformation system using the RUBY reporter gene, which successfully generated transgenic calli and cotyledon-stage embryos. The establishment of the SE system is expected to shorten breeding cycles, facilitate propagation of superior cultivars, and support large-scale industrial applications in C. intermedia. Furthermore, the stable transformation system provides a platform for molecular breeding and gene function verification. Full article

Background. Cow’s Milk Protein Allergy (CMPA) is a prevalent pediatric food allergy affecting 2–5% of infants globally. CMPA presents significant clinical and economic challenges, requiring specialized diagnostic procedures, dietary management with hypoallergenic formulas, and multidisciplinary care. The financial burden on families and healthcare systems includes direct costs (e.g., diagnostic tests, medical consultations, and formula expenses), indirect costs (e.g., caregiver absenteeism and productivity loss), and intangible costs (e.g., psychological distress and impaired quality of life). Economic disparities further exacerbate these challenges, particularly in low-resource settings where access to specialized care is limited. Methods. A systematic review was conducted following PRISMA guidelines using PubMed, CINAHL, Scopus, and Cochrane databases to identify studies on the economic and psychosocial burden of CMPA. Studies published between 2010 and 2024 were analyzed. From an initial search of 11,565 articles, 802 duplicates were removed, leaving 10,763 articles for title and abstract screening. Twenty full-text articles meeting the inclusion criteria were included in the final analysis. Thematic categories focused on direct, indirect, and intangible costs, with findings synthesized to highlight global disparities and policy gaps. Results. CMPA management imposes substantial economic burdens, with hypoallergenic formulas alone accounting for up to 15% of household income in some regions. Indirect costs, including lost workdays and additional childcare expenses, further strain families. Intangible costs, such as heightened caregiver anxiety and reduced social participation, are significant yet frequently overlooked. Healthcare system disparities, particularly regarding insurance coverage for diagnostic tests and therapeutic formulas, contribute to financial inequities. The use of extensively hydrolyzed casein formulas with probiotics has demonstrated cost-effectiveness in promoting immune tolerance while reducing healthcare utilization. Conclusions. Addressing the economic impact of CMPA would benefit from standardized cost assessment frameworks and equitable access to cost-effective therapeutic options. Further analysis of reimbursement policies across diverse healthcare systems may provide insights into optimizing support for essential treatments. Integrating clinical and economic strategies can alleviate the burden on affected families and optimize healthcare resource allocation. Future research should prioritize longitudinal analyses and cross-regional comparisons to guide sustainable and equitable management strategies. Full article

This study explored how combining supercritical fluid extraction (SFE) and enzymatic hydrolysis influences the structure and functionality of peptides recovered from filter-pressed shrimp waste. Freeze-dried press cake (PC) was defatted via SFE and hydrolyzed using Alcalase (ALC) and trypsin (TRYP). ALC-treated PC achieved the highest protein recovery (63.49%), extraction yield (24.73%), and hydrolysis degree (18.10%) (p < 0.05). SFE-treated hydrolysates showed higher zeta potential (−47.23 to −49.93 mV) than non-SFE samples (−25.15 to −38.62 mV) but had larger droplet sizes, indicating lower emulsion stability. SC-ALC displayed reduced fluorescence intensity and a red shift in maximum wavelength. TRYP hydrolysates reduced interfacial tension (20 mN/m), similar to sodium caseinate (Na-Cas, 13 mN/m), but with lesser effects. Dilatational rheology showed TRYP hydrolysates formed stronger, solid-like structures. These results emphasize protease efficacy over SFE for extracting functional compounds, enhancing shrimp waste valorization. Full article

This study optimizes the process conditions for preparing angiotensin-converting enzyme (ACE) inhibitory peptides from skimmed goat milk (SGM) hydrolyzed by multi-enzymes using response surface methodology. When the enzymatic hydrolysis time was 90 min, the optimal hydrolysis conditions were a pH of 8.49, enzyme-to-substrate ratio (E/S ratio) of 8.04%, and temperature of 61.54 °C. The hydrolysis degree and ACE inhibitory activity were 65.39% ± 0.01% and 84.65% ± 0.03%, respectively. After purification by ultrafiltration, macroporous resin, and gel filtration, the ACE inhibitory activity of F2-2 in the two components of F2 was higher, with the ACE inhibitory rate of 93.97% ± 0.15% and IC₅₀ of 0.121 ± 0.004 mg/mL. The content of hydrophobic amino acids, fatty amino acids, and aromatic amino acids in component F2-2 accounts for 73.17%, 33.86%, and 33.72%, respectively. Eleven peptides were isolated and identified from the F2-2 components of the enzymatic hydrolysate of SGM, including two peptides without an established database. The peptides mainly came from β casein, α_S1 casein, and α_S2 casein. Full article

Alzheimer’s disease (AD), a leading neurodegenerative disorder, is closely associated with the accumulation of amyloid-beta (Aβ) peptides in the brain. The enzyme β-secretase (BACE1), pivotal in Aβ production, represents a promising therapeutic target for AD. While bioactive peptides derived from food protein hydrolysates have neuroprotective properties, their inhibitory effects on BACE1 remain largely unexplored. In this study, we evaluated the inhibitory potential of protein hydrolysates from gliadin, whey, and casein proteins prepared using bromelain, papain, and thermolysin. Through in vitro and cellular assays, bromelain-hydrolyzed gliadin (G-Bro) emerged as the most potent BACE1 inhibitor, with an IC₅₀ of 0.408 mg/mL. G-Bro significantly reduced BACE1 expression and amyloid precursor protein (APP) processing in N2a/PS/APP cell cultures, suggesting its potential to attenuate Aβ aggregation. The unique peptide profile of G-Bro likely contributes to its inhibitory effect, with proline residues disrupting β-sheets, lysine residues introducing positive charges that hinder aggregation, hydrophobic residues stabilizing binding interactions, and glutamine residues enhancing solubility and stability. These findings highlight gliadin hydrolysates, particularly G-Bro, as potential natural BACE1 inhibitors with applications in dietary interventions for AD prevention. However, further studies are warranted to elucidate specific peptide interactions and their bioactivity in neural pathways to better understand their therapeutic potential. Full article

The amino acid (AA) content of multiple samples of various dairy powders was determined, providing a comprehensive evaluation of the differences in AA profiles attributable to distinct manufacturing processes. Products examined included whole milk powder (WMP), skim milk powder (SMP), cheese whey protein concentrate (WPC-C), lactic acid casein whey protein concentrate (WPC-L), high-fat whey protein concentrate (WPC-HF), hydrolyzed whey protein concentrate (WPH), whey protein isolate (WPI), and demineralized whey protein (D90). WMP and SMP exhibited broadly similar AA profiles, with minor differences likely due to the minimal milk fat protein content, which is nearly absent from SMP. Comparative analysis of WPC-C and WPC-L indicated higher levels of threonine, serine, glutamic acid, and proline in WPC-C but lower levels of tyrosine, phenylalanine, and tryptophan, attributed to the different methods of separation from casein proteins. WPI and WPC-HF originate from similar sweet whey streams but follow divergent processing methods; consequent on this were variations in the levels of all AAs except histidine. The nanofiltration step in D90 production retains its non-protein nitrogen content and affects its AA profile; consequently, D90 consistently exhibited lower AA levels than WPC-C. These findings underscore the significant impact of manufacturing processes on dairy powder AA composition. Full article

Proteases hydrolyze proteins and reduce them to smaller peptides or amino acids. Besides many biological processes, proteases play a crucial in different industrial applications. A 792 bp protease gene (nprB) from the thermophilic bacterium Streptomyces thermovulgaris was cloned and expressed in E. coli BL21 using pET 50b (+). Optimal recombinant protease expression was observed at 1 mM IPTG, 37 °C for 4 h. The resulting protease was observed in soluble form. The molecular mass estimated by SDS-PAGE and Western blot analysis of the protease (NprB) fused with His and Nus tag is ~70 KDa. The protease protein was purified by Ammonium sulfate precipitation and immobilized metal ion affinity chromatography. The optimum pH and temperature for protease activity using casein as substrate were 7.2 and 70 °C, respectively. The mature protease was active and retained 80% of its activity in a broad spectrum of pH 6–8 after 4 h of incubation. Also, the half-life of the protease at 70 °C was 4 h. EDTA (5 mM) completely inhibited the enzyme, proving the isolated protease was a metalloprotease. NprB activity was enhanced in the presence of Zn²⁺, Mn²⁺, Fe²⁺ and Ca²⁺, while Hg²⁺ and Ni²⁺ decreased its activity. Exposure to organic solvents did not affect the protease activity. The recombinant protease was stable in the presence of 10% organic solvents and surfactants. Further characterization showed that zinc-metalloprotease is promising for the detergent, laundry, leather, and pharmaceutical industries. Full article

The global protein demand is constantly on the increase, requiring sustainable and healthier protein alternatives for animal and human nutrition. Yeast-based proteins (YBPs) represent a non-negligible environmentally friendly fermentation-based solution with high nutritional quality and bioavailability. Although in vitro studies cannot reflect the full complexity of in vivo digestion, they are considered a useful alternative to animal models in assessing protein digestibility. Herein, TIM-1 (TNO gastro-intestinal model) was used to assess the digestibility profile of a proprietary edible YBP according to INFOGEST guidelines. We characterized the YBP’s digestibility and amino acid bio-accessibility and compared the YBP with milk-based references (casein and whey proteins). Each treatment was evaluated in triplicate during 5 h of digestion with hourly collection from jejunum and ileum compartments and final residual stomachal samples. Total nitrogen and free amino acid (FAA) were quantified. Size-exclusion chromatography and SDS-PAGE were also applied to assess the fate of protein hydrolysis over time. This study showed that all proteins were fully hydrolyzed upon one hour of digestion. YBPs were as good as milk-based references in terms of digestibility and small intestine absorption, reaching up to 60% of total bio-accessible protein after 5 h. Noteworthy, total YBP absorption followed a kinetics closer to that of whey protein in jejunum and ileum compartments. Our results are in line with a previous in vivo evaluation of YBPs where fecal N digestibility, PDCAAS, and DIAAS were evaluated. Altogether, our results suggest that YBPs could be a nutritionally relevant animal protein alternative. Full article

Due to its very early introduction, cow’s milk is one of the first foods that can cause adverse reactions in human beings. Lactose intolerance (LI) and cow’s milk allergy (CMA) are the most common adverse reactions to cow’s milk. While LI is due to insufficient small intestinal lactase activity and/or a large quantity of ingested lactose, CMA is an aberrant immune reaction to cow’s milk proteins, particularly casein or β-lactoglobulin. However, the clinical manifestations of LI and CMA, particularly their gastrointestinal signs and symptoms, are very similar, which might lead to misdiagnosis or delayed diagnosis as well as nutritional risks due to inappropriate dietary interventions or unnecessary dietary restriction. Formula-fed infants with LI should be treated with formula with reduced or no lactose, while those with CMA should be treated with formula containing extensive hydrolyzed cow’s milk protein or amino acids. This review is therefore written to assist clinicians to better understand the pathophysiologies of LI and CMA as well as to recognize the similarities and differences between clinical manifestations of LI and CMA. Full article

Bacillus subtilis is an important part of the gut microbiota and a commonly used probiotic. In the present study, to assess the biological characteristics and probiotic properties of B. subtilis derived from mink, we isolated B. subtilis MG-1 isolate from mink fecal samples, characterized its biological characteristics, optimized the hydrolysis of casein by its crude extract, and comprehensively analyzed its potential as a probiotic in combination with whole-genome sequencing. Biological characteristics indicate that, under low-pH conditions (pH 2), B. subtilis MG-1 can still maintain a survival rate of 64.75%; under the conditions of intestinal fluid, gastric acid, and a temperature of 70 °C, the survival rate was increased by 3, 1.15 and 1.17 times compared with the control group, respectively. This shows that it can tolerate severe environments. The results of hydrolyzed casein in vitro showed that the crude bacterial extract of isolate MG-1 exhibited casein hydrolyzing activity (21.56 U/mL); the enzyme activity increased to 32.04 U/mL under optimized reaction conditions. The complete genome sequencing of B. subtilis MG-1 was performed using the PacBio third-generation sequencing platform. Gene annotation analysis results revealed that B. subtilis MG-1 was enriched in several Kyoto Encyclopedia of Genes and Genomes (KEGG) metabolic pathways, and most genes were related to Brite hierarchy pathways (1485–35.31%) and metabolism pathways (1395–33.17%). The egg-NOG annotation revealed that most genes were related to energy production and conversion (185–4.10%), amino acid transport and metabolism (288–6.38%), carbohydrate transport and metabolism (269–5.96%), transcription (294–6.52%), and cell wall/membrane/envelope biogenesis (231–5.12%). Gene Ontology (GO) annotation elucidated that most genes were related to biological processes (8230–45.62%), cellular processes (3582–19.86%), and molecular processes (6228–34.52%). Moreover, the genome of B. subtilis MG-1 was predicted to possess 77 transporter-related genes. This study demonstrates that B. subtilis MG-1 has potential for use as a probiotic, and further studies should be performed to develop it as a probiotic additive in animal feed to promote animal health. Full article

Vitamin A is an essential micronutrient that is readily oxidized. In this study, the encapsulation of vitamin A palmitate (AP) within a core–shell carbohydrate matrix by co-axial electrospray and its oxidative stability was evaluated. The electrosprayed core–shell microcapsules consisted of a shell of octenyl succinic anhydride (OSA) modified corn starch, maltose (Hi-Cap), and a core of ethyl cellulose–AP (average diameter of about 3.7 µm). The effect of different compounds (digestion-resistant maltodextrin, soy protein hydrolysate, casein protein hydrolysate, and lecithin) added to the base core–shell matrix formulation on the oxidative stability of AP was investigated. The oxidative stability of AP was evaluated using isothermal and non-isothermal differential scanning calorimetry (DSC), and Raman and Attenuated Total Reflectance–Fourier Transform Infrared (ATR-FTIR) spectroscopy methods. The core–shell carbohydrate matrix minimizes the amount of AP present at the microparticle surface, thus protecting AP from oxidation. Furthermore, the most effective oxidation protection was achieved when casein protein hydrolysate was added to the core of the microcapsule due to hydrophobic and hydrogen bond interactions with AP and by the resistant maltodextrin in the shell, which acted as a filler. The utilization of ethanol as a solvent for the dispersion of the core compounds increased the hydrophobicity of the hydrolyzed proteins and contributed to the enhancement of their antioxidant ability. Both the carbohydrate core–shell microcapsule prepared by co-axial electrospray and the addition of oxidation protection compounds enhance the oxidative stability of the encapsulated AP. Full article