Whey protein hydrolysates and infant formulas: Effects on physicochemical and biological properties.

Whey protein hydrolysates are recognized for their substantial functional and biological properties. Their high digestibility and amino acid composition make them a valuable ingredient to hydrolyzed whey infant formulas, enhancing both product functionality and nutritional values for infant growth. It is important to understand the functional and biological properties of whey protein hydrolysates for their applications in infant formula systems. This review explored preparation methods of whey protein hydrolysates for infant formula-based applications. The effects of whey protein hydrolysate on the physicochemical and biological properties of hydrolyzed whey infant formulas were summarized. The influences of whey protein hydrolysates on the functional and nutritional properties of formulas from manufacturing to infant consumption were discussed. Whey protein hydrolysates are crucial components in the preparation of infant formula, tailored to meet the functional and nutritional demands of the product. The selection of enzyme types and hydrolysis parameters is decisive for obtaining "optimal" whey protein hydrolysates that match the intended characteristics. "Optimal" whey protein hydrolysates offer diverse functionalities, including solubility, emulsification and production stability to hydrolyzed whey infant formulas during manufacturing processes and formulations. They simultaneously promote protein digestibility, infant growth and other potential health benefits, including reduced allergenic potential, as supported by in vitro, in vivo and clinical trials. Overall, the precise selection of enzymes and hydrolysis parameters in the production of whey protein hydrolysates is crucial in achieving the desired characteristics and functional benefits for hydrolyzed whey infant formulas, making them critical in the development of infant nutrition products.

Advances in Understanding the Antioxidant and Antigenic Properties of Egg-Derived Peptides.

Pepsin, trypsin and proteinase K were used in the present study to hydrolyse the proteins from whole eggs, yolks or whites, and the resulting hydrolysates were characterised in terms of antioxidant and IgE-binding properties, using a combination of in vitro and in silico methods. Based on the degree of hydrolysis (DH) results, the egg yolk proteins are better substrates for all the tested enzymes (DH of 6.2-20.1%) compared to those from egg whites (DH of 2.0-4.4%). The SDS-PAGE analysis indicated that pepsin and proteinase K were more efficient compared to trypsin in breaking the intramolecular peptide bonds of the high molecular weight egg proteins. For all the tested substrates, enzyme-assisted hydrolysis resulted in a significant increase in antioxidant activity, suggesting that many bioactive peptides are encrypted in inactive forms in the parent proteins. The hydrolysates obtained with proteinase K exhibited the highest DPPH radical scavenging activity (124-311 µM Trolox/g protein) and the lowest residual IgE-binding capacity. The bioinformatics tools revealed that proteinase K is able to break the integrity of the main linear IgE-binding epitopes from ovalbumin and ovomucoid. It can be concluded that proteinase K is a promising tool for modulating the intrinsic properties of egg proteins.

Enzymatic Hydrolysis Optimization of Yak Whey Protein Concentrates and Bioactivity Evaluation of the Ultrafiltered Peptide Fractions.

Yak whey protein concentrates (YWPCs) have good functional properties, but there is still a gap in the study of their peptides. In this study, peptides were obtained by enzymatic hydrolysis, and the bioactivity of each ultrafiltration fraction was evaluated using an optimal process. YWPCs were isolated and purified from yak milk as the raw material. Alkaline protease, trypsin, and papain were used to hydrolyze YWPCs. The protease with the highest degree of hydrolysis (DH) and peptide concentration was selected as the most suitable enzyme. The effects of pH, temperature, time, and the enzyme-to-substrate ratio (E/S) on the DH and peptide concentration were investigated, and response surface methodology was utilized to optimize the hydrolysis process. The hydrolysate was separated using ultrafiltration membranes with molecular weight cut-offs of 10 kDa, 5 kDa, 3 kDa, and 1 kDa. The bioactivity of each ultrafiltration component was analyzed, including the inhibition rates of α-amylase and xanthine oxidase (XOD) activities and the scavenging rates of 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) cation radicals. The results indicated that alkaline protease was the best enzyme for hydrolyzing YWPCs. The peptide concentration in the YWPC hydrolysate was the highest (17.21 mg/mL) at a pH of 8 and a concentration of 7500 U/g, after 2.5 h at 62 °C. The enzymatic hydrolysate was ultrafiltered to yield four peptide fractions, of which the <1 kDa peptides exhibited the highest α-amylase inhibitory activity (22.06%), XOD inhibitory activity (17.15%), and ABTS cationic free radical scavenging rate (69.55%). This demonstrates the potential of YWPC hydrolyzed peptides for hypoglycemic, uric acid-lowering, and antioxidant applications, providing a theoretical basis for the high-value utilization of YWPCs.

Characterisation and beneficial effects of a Lupinus angustifolius protein hydrolysate obtained by immobilisation of the enzyme alcalase®.

Bioactive peptides have been considered potential components for the future functional foods and nutraceuticals generation. The enzymatic method of hydrolysis has several advantages compared to those of chemical hydrolysis and fermentation. Despite this fact, the high cost of natural and commercial proteases limits the commercialization of hydrolysates in the food and pharmacological industries. For this reason, more efficient and economically interesting techniques, such as the immobilisation of the enzyme, are gaining attention. In the present study, a new protein hydrolysate from Lupinus angustifolius was generated by enzymatic hydrolysis through the immobilisation of the enzyme alcalase® (imLPH). After the chemical and nutritional characterization of the imLPH, an in vivo study was carried out in order to evaluate the effect of 12 weeks treatment with imLPH on the plasmatic lipid profile and antioxidant status in western-diet-fed apolipoprotein E knockout mice. The immobilisation of alcalase® generated an imLPH with a degree of hydrolysis of 29.71 ± 2.11%. The imLPH was mainly composed of protein (82.50 ± 0.88%) with a high content of glycine/glutamine, arginine, and aspartic acid/asparagine. The imLPH-treatment reduced the amount of abdominal white adipose tissue, total plasma cholesterol, LDL-C, and triglycerides, as well as the cardiovascular risk indexes (CRI) -I, CRI-II, and atherogenic index of plasma. The imLPH-treated mice also showed an increase in the plasma antioxidant capacity. For the first time, this study demonstrates the beneficial in vivo effect of a lupin protein hydrolysate obtained with the alcalase® immobilised and points out this approach as a possible cost-effective solution at the expensive generation of the hydrolysate through the traditional batch conditions with soluble enzymes.

Anti-diabetic properties of brewer's spent yeast peptides. In vitro, in silico and ex vivo study after simulated gastrointestinal digestion.

Brewer's spent yeast (BSY) hydrolysates are a source of antidiabetic peptides. Nevertheless, the impact of in vitro gastrointestinal digestion of BSY derived peptides on diabetes has not been assessed. In this study, two BSY hydrolysates were obtained (H1 and H2) using ß-glucanase and alkaline protease, with either 1 h or 2 h hydrolysis time for H1 and H2, respectively. These hydrolysates were then subjected to simulated gastrointestinal digestion (SGID), obtaining dialysates D1 and D2, respectively. BSY hydrolysates inhibited the activity of α-glucosidase and dipeptidyl peptidase IV (DPP-IV) enzymes. Moreover, although D2 was inactive against these enzymes, D1 IC50 value was lower than those found for the hydrolysates. Interestingly, after electrophoretic separation, D1 mannose-linked peptides showed the highest α-glucosidase inhibitory activity, while non-glycosylated peptides had the highest DPP-IV inhibitory activity. Kinetic analyses showed a non-competitive mechanism in both cases. After peptide identification, GILFVGSGVSGGEEGAR and IINEPTAAAIAYGLDK showed the highest in silico anti-diabetic activities among mannose-linked and non-glycosylated peptides, respectively (AntiDMPpred score: 0.70 and 0.77). Molecular docking also indicated that these peptides act as non-competitive inhibitors. Finally, an ex vivo model of mouse jejunum organoids was used to study the effect of D1 on the expression of intestinal epithelial genes related to diabetes. The reduction of the expression of genes that codify lactase, sucrase-isomaltase and glucose transporter 2 was observed, as well as an increase in the expression of Gip (glucose-dependent insulinotropic peptide) and Glp1 (glucagon-like peptide 1). This is the first report to evaluate the anti-diabetic effect of BSY peptides in mouse jejunum organoids.

High pressure-assisted enzymatic hydrolysis potentiates the production of quinoa protein hydrolysates with antioxidant and ACE-inhibitory activities.

The impact of different pressure levels in the HHP-assisted hydrolysis by Alcalase of quinoa proteins on the catalytic efficiency, peptide release, phenolic compounds content, and biological activities was investigated. The protein profile (SDS-PAGE) showed a more extensive peptide breakdown for the HHP-assisted proteolysis at 300-400 MPa, which was confirmed by the higher extent of hydrolysis and peptide concentration. Quinoa protein hydrolysates (QPH) produced at 200 and 300 MPa exhibited higher total phenolic contents and antioxidant activities (methanol-acetone and aqueous extracts) when compared to the non-hydrolyzed (QPI) and non-pressurized hydrolyzed samples. Kaempferol dirhamnosyl-galactopyranoside was the prevalent phenolic compound in those samples, increasing total flavonoids by 1.8-fold over QPI. The QPH produced at 300 MPa inhibited ACE more effectively, exhibiting the greatest anti-hypertensive potential, along with the presence of several ACE-inhibitory peptides. The peptide sequences GSHWPFGGK, FSIAWPR, and PWLNFK presented the highest Peptide Ranker scores and were predicted to have ACE inhibitory, DPP-IV inhibitory, and antioxidant activities. Mild pressure levels were effective in producing QPH with enhanced functionality due to the effects of bioactive soluble phenolics and low molecular weight peptides.

Assessment of functional efficacy of sheep plasma protein hydrolysates and their utilization in mutton sausage.

The present study examines the bioactive potential of sheep plasma protein hydrolysates (SPPH) produced by in-vitro gastrointestinal digestion as antioxidants, antimicrobials, anti-obesity agents, and inhibitors of lipid oxidation in sausage to address the oxidative stability and shelf-life issues of mutton. The antioxidant and antimicrobial activities, indicate a positive relationship between the degree of hydrolysis and digestion duration. The study finds that SPPH has a potent inhibitory effect on pancreatic lipase and cholesterol esterase. It has higher oil holding capacity than sheep plasma protein, observed at one hour of hydrolysis time. SPPH exhibit an improved behavior in foaming properties along alkaline pH and digestion time while display lower emulsifying activity and stability with hydrolysis advancement. The SPPH act as a natural preservative in developing functional mutton sausage by inhibiting lipid-oxidation. This study showed that the recovery of SPPH can be a cost-effective and sustainable strategy for generating available ingredients for enhanced shelf-life of meat products.

Investigation of the peptides with calcium chelating capacity in hydrolysate derived from spent hen meat.

Calcium peptide chelates are developed as efficient supplements for preventing calcium deficiency. Spent hen meat (SHM) contains a high percentage of proteins but is generally wasted due to the disadvantages such as hard texture. We chose the underutilized SHM to produce peptides to bind calcium by proteolysis and aimed to investigate chelation between calcium and peptides in hydrolysate for a sustainable purpose. The optimized proteolysis conditions calculated from the result of response surface methodology for two-step hydrolysis were 0.30% (wenzyme/wmeat) for papain with a hydrolysis time of 3.5 h and 0.18% (wenzyme/wmeat) for flavourzyme with a hydrolysis time of 2.8 h. The enzymatic hydrolysate (EH) showed a binding capacity of 63.8 ± 1.8 mg calcium/g protein. Ethanol separation for EH improved the capacity up to a higher value of 68.6 ± 0.6 mg calcium/g protein with a high association constant of 420 M-1 (25°C) indicating high stability. The separated fraction with a higher amount of Glu, Asp, Lys, and Arg had higher calcium-binding capacity, which was related to the number of ─COOH and ─NH2 groups in peptide side chains according to the result from amino acid analysis and Fourier transform infrared spectroscopy. Two-step enzymatic hydrolysis and ethanol separation were an efficient combination to produce peptide mixtures derived from SHM with high calcium-binding capacity. The high percentage of hydrophilic amino acids in the separated fraction was concluded to increase calcium-binding capacity. This work provides foundations for increasing spent hen utilization and developing calcium peptide chelates based on underutilized meat.

Cryoprotective effect of soluble soybean polysaccharides and enzymatic hydrolysates on the myofibrillar protein of Nemipterus virgatus surimi.

Cryoprotective effect and potential mechanism of soluble soybean polysaccharides (SSPS) and enzymatic hydrolysates on surimi was investigated. After hydrolysis, the molecular weight of SSPS significantly decreased, and the hydrolysates prepared by endo-polygalacturonase (EPG-SSPS) was the lowest (154 kDa). Infrared spectrum analysis revealed that enzymatic hydrolysis didn't alter the functional groups of SSPS, but it did augment the exposure to hydroxyl groups. Surimi containing 5 % EPG-SSPS had the lowest freezable water after 20 days of frozen storage. Furthermore, the 5 % EPG-SSPS group manifested the highest metrics in total sulfhydryl (8.0 × 10-5 mol/g), active sulfhydryl content (6.7 × 10-5 mol/g), Ca2+-ATPase activity, and exhibited the lowest level in carbonyl content, surface hydrophobicity (153 µg). Notably, the 5 % EPG-SSPS maintained the stability of protein structure. Conclusively, SSPS enzymatic hydrolysate using endo-polygalacturonase imparted superior cryoprotective effect on the myofibrillar protein of surimi, and the mechanism might be a decrease in molecular weight and exposure of hydroxyl groups.

The novel angiotensin-I-converting enzyme inhibitory peptides from Scomber japonicus muscle protein hydrolysates: QSAR-based screening, molecular docking, kinetic and stability studies.

Food-derived angiotensin-converting enzyme-inhibitory (ACE-I) peptides have attracted extensive attention. Herein, the ACE-I peptides from Scomber japonicus muscle hydrolysates were screened, and their mechanisms of action and inhibition stability were explored. The quantitative structure-activity relationship (QSAR) model based on 5z-scale metrics was developed to rapidly screen for ACE-I peptides. Two novel potential ACE-I peptides (LTPFT, PLITT) were predicted through this model coupled with in silico screening, of which PLITT had the highest activity (IC50: 48.73 ± 7.59 µM). PLITT inhibited ACE activity with a mixture of non-competitive and competitive mechanisms, and this inhibition mainly contributed to the hydrogen bonding based on molecular docking study. PLITT is stable under high temperatures, pH, glucose, and NaCl. The zinc ions (Zn2+) and copper ions (Cu2+) enhanced ACE-I activity. The study suggests that the QSAR model is effective in rapidly screening for ACE-I inhibitors, and PLITT can be supplemented in foods to lower blood pressure.

Effect of bi-enzyme hydrolysis on the properties and composition of hydrolysates of Manchurian walnut dreg protein.

Walnut dreg (WD) active peptides are an important source of dietary antioxidants; however, the products of conventional hydrolysis have limited industrial output owing to poor flavour and low bioactivity. To this end, in this study, we aimed to employ bvLAP, an aminopeptidase previously identified in our research, as well as commercially available Alcalase for bi-enzyme digestion. The flavour, antioxidant activity, and structures of products resulting from various digestion methods were compared. The results showed that the bi-enzyme digestion products had enhanced antioxidant activity, increased ß-sheet content, and reduced bitterness intensity from 9.65 to 6.93. Moreover, bi-enzyme hydrolysates showed a more diverse amino acid composition containing 1640 peptides with distinct sequences. These results demonstrate that bi-enzyme hydrolysis could be a potential process for converting WD into functional food ingredients. Additionally, our results provide new concepts that can be applied in waste processing and high-value utilisation of WD.

Novel antioxidant peptides from sheep plasma protein hydrolysates: Purification, identification and cytoprotective effects against H2O2-induced oxidative stress.

This study sought to purify and identify antioxidant peptides from sheep (Ovis aries) plasma protein hydrolysates and assess their protective impacts on H2O2-induced Caco-2 cells. The purification process involved reversed high-performance liquid chromatography, anion-exchange chromatography, and Sephadex G-25. Three peptides, namely Trp-Glu-Glu-Pro-Ala-Met (WEEPAM), Ser-Leu-His-Phe-Met-Glu (SLHFME), and His-Cys-Thr-Thr-Phe-Met-Ile, with molecular weights of 761.84, 762.87, and 852.03 Da, respectively, were identified by liquid chromatography with tandem mass spectrometry. Among the three antioxidant peptides, superoxide radical (O2 -) radical scavenging capacity of WEEPAM and SLHFME was not significantly different from glutathione (GSH) (p > 0.05), while their 1,1-diphenyl-2-picrylhydrazyl radical scavenging capacity was greater than GSH (p < 0.05). WEEPAM revealed increased antioxidant activity after pepsin and trypsin hydrolysis under an in vitro digestion model. In addition, WEEPAM inhibited oxidative damage in Caco-2 cells by significantly reducing reactive oxygen species accumulation, early apoptosis, malondialdehyde formation, and increasing intracellular superoxide dismutase, glutathione peroxidase, and catalase activities.

Integrated extraction, structural characterization, and activity assessment of squid pen protein hydrolysates and ß-chitin with different protease hydrolysis.

Squid pen (SP) is a valuable source of protein and ß-chitin. However, current research has primarily focused on extracting ß-chitin from SP. This study innovatively extracted both SP protein hydrolysates (SPPHs) and SP ß-chitin (SPC) simultaneously using protease hydrolysis. The effects of different proteases on their structural characteristics and bioactivity were evaluated. The results showed that SP alcalase ß-chitin (SPAC) had the highest degree of deproteinization (DP, 98.19 %) and SP alcalase hydrolysates (SPAH) had a degree of hydrolysis (DH) of 24.47 %. The analysis of amino acid composition suggested that aromatic amino acids accounted for 17.44 % in SPAH. Structural characterization revealed that SP flavourzyme hydrolysates (SPFH) had the sparsest structure. SPC exhibited an excellent crystallinity index (CI, over 60 %) and degree of acetylation (DA, over 70 %). During simulated gastrointestinal digestion (SGD), the hydroxyl radical scavenging activity, ABTS radical scavenging activity, Fe2+ chelating activity, and reducing power of the SPPHs remained stable or increased significantly. Additionally, SPFC exhibited substantial inhibitory effects on Staphylococcus aureus and Escherichia coli (S. aureus and E. coli), with inhibition circle diameters measuring 2.4 cm and 2.1 cm. These findings supported the potential use of SPPHs as natural antioxidant alternatives and suggested that SPC could serve as a potential antibacterial supplement.

Influence of sonication pretreatment on antiradical and anti-ACE activity of protein hydrolysates from fermented pork loins.

The aim of this study was to assess whether ultrasound treatment (sonification time: 5, 15, and 30 min; constants: ∼40 kHz, ∼2.5 W cm2) can be applied prior to hydrolysis to enhance the anti-radical and angiotensin converting enzyme inhibiting (anti-ACE) effect of the hydrolysates from fermented pork loins. Enzymatic hydrolysis was performed using pepsin, followed by pancreatin. The influence of meat matrix on the course of hydrolysis, shaped using a lactic acid bacteria (LAB)-based starter culture, was also analyzed. It was found that proteases caused a systematic increase in the content of peptides, while pancreatin limited the peptide content in the protein hydrolysate from the loins subjected to spontaneous fermentation. Moreover, for these tests, sonication time had a negligible effect on the peptides content of the hydrolysates. On the other hand, for the sample of LAB-fermented products, both sonication time and stage of hydrolysis promoted the biological activity of the hydrolysates. Samples from the LAB-fermented meat had more peptides at the stage of digestion with pepsin and pancreatin, exhibiting much faster antiradical and anti-ACE activity compared to the control sample. The obtained results suggest that the use of LAB promotes the release of antiradical peptides during the two-step enzymatic hydrolysis, the duration of which can be shortened to achieve satisfactory biofunctionalities. Additional application of sonication pretreatment allows controlling the course of the hydrolysis, as the pro-health, biological effect of some protein-derived sequences is associated with the content of peptides.

Identification of the Bioavailable Peptidome of Chia Protein Hydrolysate and the In Silico Evaluation of Its Antioxidant and ACE Inhibitory Potential.

The incorporation of novel, functional, and sustainable foods in human diets is increasing because of their beneficial effects and environmental-friendly nature. Chia (Salvia hispanica L.) has proved to be a suitable source of bioactive peptides via enzymatic hydrolysis. These peptides could be responsible for modulating several physiological processes if able to reach the target organ. The bioavailable peptides contained in a hydrolysate obtained with Alcalase, as functional foods, were identified using a transwell system with Caco-2 cell culture as the absorption model. Furthermore, 20 unique peptides with a molecular weight lower than 1000 Da and the higher statistical significance of the peptide-precursor spectrum match (-10 log P) were assessed by in silico tools to suggest which peptides could be those exerting the demonstrated bioactivity. From the characterized peptides, considering the molecular features and the results obtained, the peptides AGDAHWTY, VDAHPIKAM, PNYHPNPR, and ALPPGAVHW are anticipated to be contributing to the antioxidant and/or ACE inhibitor activity of the chia protein hydrolysates.

Novel Multitarget ACE Inhibitory Peptides from Bovine Colostrum Immunoglobulin G: Cellular Transport, Efficacy in Regulating Endothelial Dysfunction, and Network Pharmacology Studies.

In this study, we used traditional laboratory methods, bioinformatics, and cellular models to screen novel ACE inhibitory (ACEI) peptides with strong ACEI activity, moderate absorption rates, and multiple targets from bovine colostrum immunoglobulin G (IgG). The purified fraction of the compound proteinase hydrolysate of IgG showed good ACEI activity. After nano-UPLC-MS/MS identification and in silico analysis, eight peptides were synthesized and verified. Among them, SFYPDY, TSFYPDY, FSWF, WYQQVPGSGL, and GVHTFP were identified as ACEI peptides, as they exhibited strong ACEI activity (with IC50 values of 104.7, 80.0, 121.2, 39.8, and 86.3 µM, respectively). They displayed good stability in an in vitro simulated gastrointestinal digestion assay. In a Caco-2 monolayer model, SFYPDY, FSWF, and WYQQVPGSGL exhibited better absorption rates and lower IC50 values than the other peptides and were thereby identified as novel ACEI peptides. Subsequently, in a H2O2-induced endothelial dysfunction (ED) model based on HUVECs, SFYPDY, FSWF, and WYQQVPGSGL regulated ED by reducing apoptosis and ROS accumulation while upregulating NOS3 mRNA expression. Network pharmacology analysis and RT-qPCR confirmed that they regulated multiple targets. Overall, our results suggest that SFYPDY, FSWF, and WYQQVPGSGL can serve as novel multitarget ACEI peptides.

Targeted Affinity Purification and Mechanism of Action of Angiotensin-Converting Enzyme (ACE) Inhibitory Peptides from Sea Cucumber Gonads.

Protein hydrolysates from sea cucumber (Apostichopus japonicus) gonads are rich in active materials with remarkable angiotensin-converting enzyme (ACE) inhibitory activity. Alcalase was used to hydrolyze sea cucumber gonads, and the hydrolysate was separated by the ultrafiltration membrane to produce a low-molecular-weight peptide component (less than 3 kDa) with good ACE inhibitory activity. The peptide component (less than 3 kDa) was isolated and purified using a combination method of ACE gel affinity chromatography and reverse high-performance liquid chromatography. The purified fractions were identified by liquid chromatography-tandem mass spectrometry (LC-MS/MS), and the resulting products were filtered using structure-based virtual screening (SBVS) to obtain 20 peptides. Of those, three noncompetitive inhibitory peptides (DDQIHIF with an IC50 value of 333.5 µmol·L-1, HDWWKER with an IC50 value of 583.6 µmol·L-1, and THDWWKER with an IC50 value of 1291.8 µmol·L-1) were further investigated based on their favorable pharmacochemical properties and ACE inhibitory activity. Molecular docking studies indicated that the three peptides were entirely enclosed within the ACE protein cavity, improving the overall stability of the complex through interaction forces with the ACE active site. The total free binding energies (ΔGtotal) for DDQIHIF, HDWWKER, and THDWWKER were -21.9 Kcal·mol-1, -71.6 Kcal·mol-1, and -69.1 Kcal·mol-1, respectively. Furthermore, a short-term assay of antihypertensive activity in spontaneously hypertensive rats (SHRs) revealed that HDWWKER could significantly decrease the systolic blood pressure (SBP) of SHRs after intravenous administration. The results showed that based on the better antihypertensive activity of the peptide in SHRs, the feasibility of targeted affinity purification and computer-aided drug discovery (CADD) for the efficient screening and preparation of ACE inhibitory peptide was verified, which provided a new idea of modern drug development method for clinical use.

Analysis of flavor changes in Huangshan floral mushroom hydrolysates obtained by different enzyme treatments.

This study aimed to investigate the flavor changes in Huangshan floral mushroom by different enzyme treatments. Seven enzyme groups were used to hydrolyze its protein to obtain protein hydrolysates (FPHs). Flavourzyme composite with dispase hydrolysates (FDHs) were selected for ultrafiltration to obtain peptides (FPs) with different molecular weights (Mw). Changes in flavor were investigated using HPLC, LC-MS, GC-MS, amino acid analysis and sensory evaluation. Color parameters and DPPH-scavenging activity were also determined. The results revealed that flavor characteristics of FPHs obtained from different enzyme treatments varied. FDHs presented the highest degree of hydrolysis (DH) (58.61 ± 1.55) %, rich 5'-nucleotides (8.61 ± 0.43 mg/mL), volatile compounds (28.54 ± 0.11 µg/g) and free amino acids (FAAs) (7.73 ± 0.51 mg/g). Further tests suggested that FPs with small Mw (<1K, 1-3 K) were optimal for the development of novel flavors, thus providing application value for rational utilization of Huangshan floral mushroom.

Preparation and identification of antioxidant peptides from Quasipaa spinosa skin through two-step enzymatic hydrolysis and molecular simulation.

Frog skin, a by-product of Quasipaa Spinosa farming, is rich in protein and potentially a valuable raw material for obtaining antioxidant peptides. This study used papain combined with acid protease to digest frog skin in a two-step enzymatic hydrolysis method. Based on a single factor and response surface experiments, experimental conditions were optimized, and the degree of hydrolysis was 30 %. A frog skin hydrolysate (QSPH-Ⅰ-3) was obtained following ultrafiltration and gel filtration chromatography. IC50 for DPPH, ABTS, and hydroxyl radical scavenging capacities were 1.68 ± 0.05, 1.20 ± 0.14 and 1.55 ± 0.11 mg/mL, respectively. Peptide sequences (17) were analyzed and, through molecular docking, peptides with low binding energies for KEAP1 were identified, which might affect the NRF2-KEAP1 pathway. These findings suggest protein hydrolysates and antioxidant peptide derivatives might be used in functional foods.

Comparison of prediction models for soy protein isolate hydrolysates bitterness built using sensory, spectrofluorometric and chromatographic data from varying enzymes and degree of hydrolysis.

The bitterness of soy protein isolate hydrolysates prepared using five proteases at varying degree of hydrolysis (DH) and its relation to physicochemical properties, i.e., surface hydrophobicity (H0), relative hydrophobicity (RH), and molecular weight (MW), were studied and developed for predictive modelling using machine learning. Bitter scores were collected from sensory analysis and assigned as the target, while the physicochemical properties were assigned as the features. The modelling involved data pre-processing with local outlier factor; model development with support vector machine, linear regression, adaptive boosting, and K-nearest neighbors algorithms; and performance evaluation by 10-fold stratified cross-validation. The results indicated that alcalase hydrolysates were the most bitter, followed by protamex, flavorzyme, papain, and bromelain. Distinctive correlation results were found among the physicochemical properties, influenced by the disparity of each protease. Among the features, the combination of RH-MW fitted various classification models and resulted in the best prediction performance.