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This study aims to analyze the differences in digestion properties and peptide profiles between the skim camel and bovine milk powder after static in vitro simulated infant gastrointestinal digestion. The hydrolysis degree of camel milk proteins exceeded by 13.18% that of bovine milk. The concentration and release rate of free amino groups in the camel milk digesta was higher than that of bovine milk powder, which was likely due to the higher β-/α s -casein ratio and larger casein micelle size in camel milk. Camel milk powder presented higher β-CN coverage and comparatively shorter bioactive peptides compared to bovine milk powder. The anti-inflammatory peptide KVLPVPQ displayed the highest abundance in camel milk powder. Outcomes of this study showed that camel milk proteins possessed superior digestibility and unique peptides, which outlined the potential nutritional implications of camel milk for infants., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier Ltd.)

An infant in vitro digestion model was utilized to investigate protein digestion characteristics in human and diverse mammalian milk (i.e., cow, goat, sheep, mare, and camel milk) using electrophoresis and chromatography. Digestive differences among milks were mainly manifested in the infant gastric phase, as evidenced by varying degrees of protein digestion. Notably, proteins (i.e., lactoferrin, serum albumin, and immunoglobulin G-heavy chain) remained partially intact in human milk, whereas these proteins in animal milk were exclusively degraded after gastrointestinal digestion. The peptide spectra of human, mare, and camel milk were highly similar, with a predominant formation of low-intensity small peptides, whereas the other three milk showed the opposite phenomenon. Heatmap cluster analysis indicated that camel milk was the most comparable to human milk before digestion, yet sheep milk was the most similar to human milk regarding protein digestion behaviors following infant gastric digestion.

The demand for plant-based meat analogs (PBMA) is on the rise as a strategy to sustain the food protein supply while mitigating environmental change. In addition to supplying essential amino acids and energy, food proteins are known sources of bioactive peptides. Whether protein in PBMA affords similar peptide profiles and bioactivities as real meat remains largely unknown. The purpose of this study was to investigate the gastrointestinal digestion fate of beef and PBMA proteins with a special focus on their potential as precursors of bioactive peptides. Results showed that PBMA protein showed inferior digestibility than that in beef. However, PBMA hydrolysates possessed a comparable amino acid profile to that of beef. A total of 37, 2420 and 2021 peptides were identified in the gastrointestinal digests of beef, Beyond Meat and Impossible Meat, respectively. The astonishingly fewer peptides identified from beef digest is probably due to the near-full digestion of beef proteins. Almost all peptides in Impossible Meat digest were from soy, whereas 81%, 14% and 5% of peptides in Beyond Meat digest were derived from pea, rice and mung proteins, respectively. Peptides in PBMA digests were predicted to exert a wide range of regulatory roles and were shown to have ACE inhibitory, antioxidant and anti-inflammatory activities, supporting the potential of PBMA as a source of bioactive peptides.

High salt (NaCl) consumption can impact on human health, and KCl is the most widely used replacement salt in meat products. This study investigated the effects of 0% NaCl (NS), 3% NaCl (HS), 1.95% NaCl (RS), 1.95% NaCl+1.05% KCl (RS + K) on protein digestibility of pork sausage in vitro. The results indicated that RS + K showed the highest gastrointestinal digestibility (GID) because of the structure of looser cross-linked strands and uniform cavities, while HS exhibited the lowest GID. RS + K released more peptides (2499) during gastrointestinal than NS (2301), RS (2130) and HS (2235), with a higher proportion of peptides with molecular weights <1000 Da, and more unique peptides. Meanwhile, the digestion product of RS + K exhibited excellent radical scavenging activity and improved the antioxidant abilities to reduce oxidative injury which was induced by H 2 O 2 in HepG2 cells. These results demonstrated that partial substitution with KCl can be an effective strategy for improving the digestibility of sodium-reduced gel-type meat products., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022. Published by Elsevier Ltd.)

This present study aimed to investigate the effects of various cooking times (1 h, 1.5 h, 2 h, 2.5 h, named as JHBP-1, JHBP-1.5, JHBP-2, JHBP-2.5) on the antioxidant activity and peptide profile of Jinhua ham broth peptides (JHBP). The peptides extracted from uncooked ham were used as an uncooked group with the name of JHBP-0. The results revealed that the antioxidant efficacy in the four cooked groups changed dramatically compared to JHBP-0. After cooking, the DPPH radical scavenging activity, hydroxyl radical scavenging activity and superoxide anion radical scavenging activity decreased, except for the Fe 2+ chelation and ABTS + scavenging capacity which increased significantly. However, the cooked groups still showed a strong antioxidant capacity. In particular, the superoxide anion radical scavenging ability and the Fe 2+ chelation action were significantly stronger compared to glutathione (GSH) and butylated hydroxytoluene (BHT) ( p < 0.05). JHBP-1.5 also displayed stronger antioxidant capacity than the other three cooked groups, and its secondary structure and mass distribution changed significantly after cooking, specifically with an increased proportion of helix and <1 kDa peptides. Moreover, the constitution of free amino acids (FAAs) and the types of peptides released in the broth increased significantly with a longer cooking time. In total, 1306 (JHBP-0), 1352 (JHBP-1), 1431 (JHBP-1.5), 1500 (JHBP-2), and 1556 (JHBP-2.5) peptide sequences were detected using LC-MC/MC. The proportion of <1 kDa peptides also gradually increased as the cooking time extended, which is consistent with the molecular weight distribution measurements.

Milk protein concentrate was hydrolyzed using one-step enzymatic hydrolysis. Both the peptide profiles and antioxidant activities of the resulting extensive hydrolysates of milk protein concentrate (EMPH) were analyzed using a peptidomics approach based on liquid chromatography-tandem mass spectrometry. The results demonstrated that the degrees of hydrolysis of the 4 EMPH by Alcalase-Protamex, Alcalase-Protease A 2SD, Alcalase-Flavorzyme, and Alcalase-ProteAXH were 12.02%, 16.85%, 15.87%, and 15.77%, respectively. Using size exclusion chromatography, 99.85% of the peptides in the Alcalase-Protease A 2SD hydrolysate were shown to have a molecular weight of <3 kDa. A total of 33 common peptides were identified in the EMPH by liquid chromatography-tandem mass spectrometry, 16 of which were identified as bioactive peptides using bioinformatics. The peptide profiles and the coverage of master proteins of the 4 EMPH were different. The EMPH also exhibited strong free radical scavenging capacity, as indicated by the results of the 1,1-diphenyl-2-picrylhydrazyl radical, 2,2'-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid), hydroxyl radical, and reducing power assays. The results of this study provided useful information on the peptide profiles and antioxidant activity of EMPH., (The Authors. Published by Elsevier Inc. and Fass Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

Rubing cheese is a traditional Chinese Protected Designation of Origin (PDO) cheese consumed for more than six hundred years, but to date, the digestion properties and peptide profiling during simulated gastrointestinal digestion are still uncertain. This study aimed to investigate the effects of traditional direct acidification technology (TRB) and fermentation acidification technology on digestion properties and peptide profiling of rubing cheese (FRB) proteins after simulated gastrointestinal digestion by protein digestomics, coupled with bioinformatic in silico analyses to identify potential bioactive peptides. The results demonstrated that FRB could significantly improve the in vitro digestibility, protein degradation, and polypeptide content than TRB (P ＜ 0.05). Furthermore, a total of 369 and 332 peptides were identified in FRB- and TRB-pancreatic digests, respectively, using LC-MS/MS. FRB could release more low molecular weight peptides of 400-1200 Da from α-casein and β-casein after digestion. These low peptides included 16 reported potential ACEIPs (angiotensin I-converting enzyme inhibitory peptides), 11 dipeptidyl peptidase-IV (DPP-IV) inhibitory peptides, and 6 antioxidant peptides, while TRB contained more than the reported potential antimicrobial peptides (10). In vitro activity determination showed that FRB had significantly higher ACEI, α-glucosidase inhibitory, and antioxidant activities than TRB during the entire digestion time (P ＜ 0.05), which was correlated to the reported potential bioactive peptides released during the digestion of FRB. Our study is the most comprehensive protein digestomic analysis of Chinese rubing cheese to date and provides a new positive outlook on rubing cheese consumption., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

We applied in vitro models of gastrointestinal (GI) digestion simulating the conditions of the GI tract of healthy adults and elderly individuals with achlorhydria (EA) to investigate differences in the digestibility of meat (chicken, beef and pork) and soy proteins. Digestibility was significantly affected by EA alterations. Peptidomics analyses revealed significant differences in peptide profiles between control and EA conditions, including number, length distribution, clustering, and differentially abundant peptides (DAPs). Our results revealed that the differences in meat peptide profiles diminished going from the gastric to intestinal phase. For soy protein, the marked differences between control and EA conditions were maintained in the gastric and intestinal phases. Higher numbers of potentially bioactive peptides were generated under the control condition compared to the EA condition. The present study provides insight into the distinct peptide profiles generated by in vitro digestion of meat and soy proteins under adult and EA GI conditions., (Copyright © 2022. Published by Elsevier Ltd.)

This study aimed to investigate the effect of steam cooking on the proteolysis of Pacific oysters (Crassostrea gigas) using the simulated oral-gastrointestinal digestion model and a NCM460 cell monolayer. Steam cooking changed the peptide profile of the digests of oysters considerably and induced more thorough hydrolysis. However, the heat-stable allergen, Cra g 1, still had remnant fragments in the intestinal phase, which could be allergenic epitopes. Two regions of Cra g 1 (residues 224-228 and 245-248) were digestion-tolerant. Furthermore, more oligopeptides were derived from raw proteins than from steamed proteins. After molecular docking and in vitro determination, six novel angiotensin I-converting enzyme inhibitory (ACEi) peptides were finally identified in the hydrolysates (WIS, WLS, LSL, SGPF, LGPI, and IGLP). Among them, LSL exhibited the highest ACEi activity (IC 50  = 107.17 nM). Our findings provide supportive information on the effective utilization of oyster proteins., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Background: Degenerative mitral valve disease is a common heart problem in dogs. The aims are to evaluate the relationships between right and left ventricular function, and blood lactate concentrations, assess prognostic contribution, and investigate whether serum peptidomics profile could reveal markers or determine the stage in dogs with valve degeneration., Materials and Methods: Ninety-three dogs were evaluated in this study. Thirty-nine dogs' serum was collected and analyzed using matrix-assisted laser desorption/ionization-time of flight mass spectrometry. The Kaplan-Meier curve was used to predict the outcomes of mitral valve disease. Follow-up was obtained by a questionnaire or telephone to determine a survival time., Results: The BUN/creatinine ratio, vertebral heart score, and left atrium/aorta ratio were the independent predictors of cardiac mortality. Right ventricular systolic dysfunction was found in 50% of dogs with mitral valve disease. Dogs with right ventricular dysfunction had a significantly higher incidence of lower fractional shortening and larger right ventricular dimensions. The occurrence of right-sided dysfunction is proportionate to age and the degree of left ventricular dysfunction. High blood lactate concentrations were investigated in dogs with mitral valve disease stage C compared with stage B. The peptides such as mitogen-activated protein kinase, kallikrein, and tenascin-C appeared in the heart disease progression group., Conclusion: Right-hearted function assessment, blood lactate levels, and peptidomics analysis may help early detection and prognosis of this disease in dogs. Peptidomics profiles from this study demonstrate the possibility for prognosis indicators of heart valve degeneration., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Petchdee, Yalong, Kaewnet, Ithisariyanont and Padawong.)

Abstract: This paper shows the potential of dual enzyme approach on antioxidant activity of casein hydrolysates. Casein was hydrolysed using the proteolytic enzymes alcalase, flavourzyme in isolation and in sequential order. Casein hydrolysates were evaluated for the degree of hydrolysis, antioxidant activity, molecular weight distribution patterns and peptide sequence. Casein hydrolysate produced by the sequential hydrolysis of alcalase and flavourzyme showed higher degree of hydrolysis and antioxidant activity as compared to hydrolysate obtained by individual enzymes. In size exclusion chromatograph of casein hydrolysate S3, peptides with molecular weight of 0.57 kDa share 12% area in total area of chromatogram which was 10 times higher than that of hydrolysate S1 and nearly half of that of hydrolysate S2. On subjecting to HPLC-TOF-ESI separation potential antioxidant peptides were identified. The peptide sequence VLPVPQ along with potential fragments was identified in hydrolysate S1 and S2 and HPHPHLS along with its potential sequence was identified in hydrolysate S1, S2 and S3. Sequential hydrolysis of casein showed better antioxidant activity and peptide profile in less duration as compared to the casein hydrolysate obtained by individual enzyme., (© Association of Food Scientists & Technologists (India) 2020.)

The study aimed to analyze pH and heat treatment's effect in modulating the release of peptides with antioxidant activity after simulated gastrointestinal (GI) digestion of Egg white powder (EWP). EWP samples with neutral (EWPN) and alkaline (EWPA) pH were heat-treated at 20, 60, and 90 °C and analyzed for protein aggregation, solubility, and GI digestibility. Heat treatment decreased solubility and induced protein aggregation, which was higher for EWPN as compared to EWPA. The unfolding of EWPA proteins at 60 °C exhibited a higher GI digestibility and antioxidant activity via Oxygen Radical Absorbance Capacity (ORAC) assay as compared to EWPN. Interestingly, a reverse trend was observed in the cellular antioxidant assay, and the GI-digest of EWPN exhibited a higher antioxidant activity. The LC-MS/MS analysis are in concordance with cellular antioxidant activity assay and showed a higher intensity for peptides with potential antioxidant activity in the GI-digest of EWPN. The results indicate that heat treatment but not the pH is a critical factor in improving the protein digestibility and releasing peptides with antioxidant activity after GI digestion.

The performances of protein hydrolysates highly depend on their peptide composition (amount, size and diversity), which itself closely depends on raw material origin and the hydrolysis parameters of the manufacturing process. The current analyses that characterize protein hydrolysates provide information on the level of hydrolysis (degree of hydrolysis, DH). However, they need additional describers to better characterize peptide profiles and product standardization. To reach this objective, we developed a fast and standardized method to characterize the abundance and the diversity of low-molecular-weight peptides in protein hydrolysates. This method innovatively combines classical HPSEC and nLC-ESI-MS analytical tools to characterize any kind of hydrolysate, whether solid or liquid, in terms of peptide level and diversity, and then merge peptides into 2D diagrams to visualize comparisons between protein hydrolysates. The targeted applications of this new tool for characterizing complex protein hydrolysates are (i) verifying the standardization of the produced products across batches, and (ii) analyzing and understanding the consequences of the modifications of the hydrolysis process on the molecular profiles of the generated peptides. The sample standardization described in this study is therefore an essential prerequisite for the functional characterization of hydrolysates in vitro ., (© 2020 The Authors.)

Effects of high-pressure treatments (HPT, 100-300 MPa, 9 min, 25 °C) on the in vitro digestibility of gel-type meat products were studied using a simulated digestion-model. In vitro digestibilities of the cooked rabbit meat batters throughout the simulated oral-, gastric-, and intestinal-phases were determined. Peptides in the intestinal digesta were identified via Mass Spectrometer. Results revealed that in vitro digestibilities of HPT-samples were higher than the control (1.98%, 6.13% and 61.31% for oral-, gastric- and intestinal-phase respectively) throughout the digestion (P < 0.05). Alterations of the peptide profiles were induced by HPT, showing HPT-specific patterns of mutual peptides in the digestive products. Coupled with the identifications of salt-soluble proteins from raw batters, it was confirmed that myofibrillar proteins account for the major contribution to the HPT-induced changes. The results indicated that HPT can potentially be an effective technology to improve the digestibility of meat products., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

The chemical and biological characterization of peptide and protein components of the paralyzing venom from three Pompilidae solitary spider wasps (Pepsis mexicana, Pepsis terminata, and Anoplius nigritus) is described for the first time. The molecular masses of the most abundant peptides were determined. The N-terminal sequences of two cysteine-rich peptides were obtained from Pepsis. Metalloproteinase and hyaluronidase activities were identified in the venom of P. mexicana. A novel non-lethal method to collect venom is described., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Health-promoting effects of kefir may be partially caused by bioactive peptides. To evaluate their formation or degradation during gastrointestinal digestion, we monitored changes of the peptide profile in a model of (1) oral, (2) gastric, and (3) small intestinal digestion of kefir. Matrix-assisted laser desorption/ionization time-of-flight mass spectroscopy analyses revealed clearly different profiles between digests 2/3 and kefir/digest 1. Subsequent ultraperformance liquid chromatography-electrospray ionization-tandem mass spectrometry identified 92 peptides in total (25, 25, 43, and 30, partly overlapping in kefir and digests 1, 2, and 3, respectively), including 16 peptides with ascribed bioactivity. Relative quantification in scheduled multiple reaction monitoring mode showed that many bioactive peptides were released by simulated digestion. Most prominently, the concentration of angiotensin-converting enzyme inhibitor β-casein 203-209 increased approximately 10 000-fold after combined oral, gastric, and intestinal digestion. Thus, physiological digestive processes may promote bioactive peptide formation from proteins and oligopeptides in kefir. Furthermore, bioactive peptides present in certain compartments of the gastrointestinal tract may exert local physiological effects.

In the present study, the antipyretic activity of Bubali Cornu (water buffalo horn) fraction and its metabolomics were investigated. The fraction decreased rat rectal temperature, and 13 endogenous metabolites were identified as potential biomarkers. Selected metabolites were involved in arachidonic acid metabolism and glycerophospholipid metabolism etc. Following treatment with the fraction, four metabolites, pyroglutamic acid, palmitelaidic acid, leukotriene A4, and prostaglandin A2 were reversed. In addition, the levels of interleukin-1β, tumor necrosis factor-α, prostaglandin E 2 , and cyclic adenosine monophosphate in plasma were also reversed after treatment as determined by enzyme linked immunosorbent assay. Furthermore, nano-flow liquid chromatography with orbitrap mass spectrometry detection was used to analyze the peptides in the fraction. In total, 824 peptide sequences mainly from keratins were determined, with Keratin 14, Keratin 34, and Keratin 86 representing the three main types of keratin hydrolysis in water buffalo horn based on peptide heat maps. Of the identified peptides, 81.2% were hydrophilic and the molecular weight of 70.3% of identified peptides was lower than 2000 Da. According to the metabolomics- and peptidomics-based approach used in the present study, it is feasible to identify and analyze the active peptide matrix from animal-horn-derived traditional Chinese medicines., (© 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Enzymatic hydrolysis of whey protein (WP) was carried out under pH-controlled and non pH-controlled conditions using papain and a microbial-derived alternative (papain-like activity). The impact of such conditions on physicochemical and bioactive properties was assessed. WP hydrolysates (WPH) generated with the same enzyme displayed similar degree of hydrolysis. However, their reverse-phase liquid chromatograph mass spectrometry peptide profiles differed. A significantly higher oxygen radical absorbance capacity (ORAC) value was obtained for WP hydrolysed with papain at constant pH of 7.0 compared to the associated WPH generated without pH regulation. In contrast, there was no significant effect of pH regulation on dipeptidyl peptidase IV (DPP-IV) properties. WP hydrolysed with papain-like activity under pH regulation at 7.0 displayed higher ORAC activity and DPP-IV inhibitory properties compared to the associated WPH generated without pH regulation. This study has demonstrated that pH conditions during WPH generation may impact on peptide release and therefore on WPH bioactive properties., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

In recent years, mass spectrometry imaging (MSI) has been shown to be a promising technique in oncology. The effective application of MSI, however, is hampered by the complexity of the generated data. Bioinformatic approaches that reduce the complexity of these data are needed for the effective use in a (bio)medical setting. This holds especially for the analysis of tissue microarrays (TMA), which consist of hundreds of small tissue cores. Here we present an approach that combines MSI on tissue microarrays with principal component linear discriminant analysis (PCA-LDA) to predict treatment response. The feasibility of such an approach was evaluated on a set of patient-derived xenograft models of triple-negative breast cancer (TNBC). PCA-LDA was used to classify TNBC tumor tissues based on the proteomic information obtained with matrix-assisted laser desorption ionization (MALDI) MSI from the TMA surface. Classifiers based on two different tissue microarrays from the same tumor models showed overall classification accuracies between 59 and 77%, as determined by cross-validation. Reproducibility tests revealed that the two models were similar. A clear effect of intratumor heterogeneity of the classification scores was observed. These results demonstrate that the analysis of MALDI-MSI data by PCA-LDA is a valuable approach for the classification of treatment response and tumor heterogeneity in breast cancer.