Search Results (69)

Lentinus edodes stipe is an under-utilised processing by-product that holds potential as a natural umami source. This study systematically compared nine enzymatic hydrolysis combinations, including single proteases, cellulase, and double enzyme systems comprising both protease–protease and protease–cellulase mixtures, to optimise the release of taste-active and volatile flavor compounds from the stipes. The Flavourzyme-only hydrolysate achieved the highest degree of hydrolysis (50.38%), the richest 5′-nucleotides (5.46 mg/g), free amino acids (59.06 mg/g) and volatile compounds (150.23 μg/kg), and an equivalent umami concentration of 288 g monosodium glutamate equivalent per 100 g matter. It also delivered the strongest umami taste and the lowest bitterness in sensory evaluation. The double-enzyme systems consistently underperformed due to competitive substrate binding, suboptimal pH for companion enzymes, and potential mutual protease digestion. Among the volatile compounds, 1,2,4-trithiolane and cedrol were identified as key aroma differentiators by multivariate analysis. These findings demonstrate that Flavourzyme alone constitutes the optimal enzymatic system for producing clean-tasting, umami-rich condiments from L. edodes stipes and caution against indiscriminate enzyme blending without prior compatibility assessment. The optimized hydrolysis process provides a scalable route for the industrial valorisation of mushroom processing by-products. Full article

Quinoa (Chenopodium quinoa) is a promising source of plant proteins with the potential to produce bioactive peptides through enzymatic hydrolysis. This study aimed to extract quinoa protein and produce bioactive peptides using two microbial proteases: Alcalase (from Bacillus licheniformis) and Flavourzyme (from Aspergillus oryzae). The protein was extracted through alkaline solubilization and isoelectric precipitation, achieving a 72% yield. Hydrolysis was conducted for 4 h, and enzymatic activity was measured using the TNBS method to determine the degree of hydrolysis, while SDS-PAGE was used to analyze protein breakdown. The reaction was performed at controlled pH and temperature (Alcalase: 9.5 and 55 °C; Flavourzyme: 7 and 37 °C). Both enzymes achieved maximum hydrolysis at 60 min. Consequently, the separation and inhibitory capacity of angiotensin-converting enzyme (ACE-I) were tested at the first four time points (0, 20, 40, and 60 min). A wider variety and higher concentration of peptides smaller than 2 kDa were found in hydrolysates treated with Flavourzyme, which is associated with antihypertensive activity. The ACE-I assay showed greater activity at the end of hydrolysis. Inhibition percentages of 87.5 ± 2.11 were observed in hydrolysates with Flavourzyme, and 94.1 ± 1.11 in those with Alcalase. These findings indicate that quinoa protein, hydrolyzed with microbial proteases, is a feasible source of peptides with potential antihypertensive effects for use in functional foods and nutraceuticals. Full article

Sea cucumber viscera by-products are abundant but remain underutilized. Although the development of umami peptides from marine by-products has been well-reported, sea cucumber viscera have received less attention. In this study, an umami-rich hydrolysate was prepared from sea cucumber viscera through synergistic dual-enzyme hydrolysis. Under optimal conditions, the co-hydrolysis using Flavourzyme and aminopeptidase yielded extraction rates of 69.38% for solids, 67.29% for protein, and 66.96% for total sugar, and produced a 1.75-fold higher umami signal intensity (electronic tongue) than the single-enzyme (Flavourzyme) hydrolysate. The target umami fraction was enriched through sensory-guided separation combined with ultrafiltration and ion-exchange chromatography. Thirty-three umami peptides, predominantly derived from actin hydrolysis, were identified in this fraction via peptidomics and virtual screening. Based on docking simulations against the umami receptor T1R1/T1R3, two peptides (DFLDDGPG and SDTGNFGF) with the lowest docking scores were selected. The predictions revealed that two peptides bind to the T1R3 subunit via hydrogen bonds and π-related interactions. The umami-enhancing effect of peptide DFLDDGPG in salty systems was demonstrated by a trained panel (n = 10) across concentration ranges of 0.1–1.0 mg/mL peptide and 0.1–1.0% NaCl, with a positive correlation validated by RSM and ANOVA (p < 0.05). This study identified novel umami peptides from sea cucumber by-products as promising candidates for natural, low-sodium flavor enhancers. Full article

The invasive fish Atherina boyeri constitutes an ecologically disruptive yet underexploited biomass with strong potential for transformation into value-added biofunctional ingredients. This study investigates the functional, antioxidant, and antimicrobial properties of protein hydrolysates that were produced from fish collected in the Hirfanlı and Yamula reservoirs using three commercial proteases (alcalase, bromelain, and flavourzyme). Bromelain produced the highest degree of hydrolysis, yielding higher proportions of low-molecular-weight peptides and greater radical-scavenging activity. Flavourzyme hydrolysates exhibited the most favorable emulsifying properties, Alcalase hydrolysates produced the highest foaming capacity and stability. All hydrolysates showed high absolute zeta-potential values across pH 3–9, demonstrating strong colloidal stability. Protein solubility remained above 80% across most pH levels, indicating extensive peptide release and improved compatibility with aqueous media. The Oil-binding capacity (2.78–3.75 mL/g) was consistent with reported values for marine hydrolysates. Antioxidant and antimicrobial evaluations revealed clear enzyme-dependent patterns, with Bromelain exhibiting the strongest DPPH activity and Alcalase and Flavourzyme showing the most pronounced inhibition of major foodborne pathogens. Additionally, all hydrolysates exhibited measurable ACE-inhibitory activity, with flavourzyme-derived peptides showing the highest inhibitory activity, underscoring their potential relevance for antihypertensive applications. These findings highlight the strategic valorization of A. boyeri through enzymatic hydrolysis, demonstrating its potential as a sustainable, clean-label functional ingredient source. Full article

The growing demand for sustainable protein sources has increased interest in algae and aquatic plants as alternatives to animal-derived proteins. These resources are rich in protein, amino acids, and umami compounds but require suitable extraction methods to maximize yield and quality. This study compared three green extraction techniques—maceration (MAE, 80 °C, 2 h), ultrasound-assisted extraction (UAE, 750 W, 20 kHz, 50% amplitude, 35 °C, pH 12, 1 h), and enzyme-assisted extraction (EAE, 5% β-glucanase/flavourzyme, 55 °C, pH 6.5, 1 h)—on five raw materials: wakame (commercial seaweed), hair seaweed, sea lettuce, water silk algae, and Wolffia. The result revealed that both raw materials and extraction methods significantly (p < 0.05) affected protein yield, amino acid, physicochemical properties, and taste detection with e-tongue. Wolffia extracted by MAE yielded the highest protein overall, followed by UAE and EAE methods, when compared with commercial seaweed. The relationship between amino acid profiles and taste detection was investigated by principal component analysis (PCA) and hierarchical cluster analysis (HCA); the samples with higher glutamic and aspartic acids were linked with umami taste, while histidine contributed to bitter taste. Overall, the findings highlighted that extraction efficiency was influenced more by the extraction method–material compatibility than the raw material alone. Full article

To prepare and characterize low-bitter angiotensin-converting enzyme (ACE)-inhibitory peptides from sesame protein, a triple-enzyme hydrolysis system was optimized using mixture design and response surface methodology. The resulting hydrolysate was separated by ultrafiltration and medium-pressure chromatography, followed by identification through nano-liquid chromatography–electrospray ionization-tandem mass spectrometry. Finally, the mechanism of typical low-bitter ACE-inhibitory peptides was elucidated by molecular docking and molecular dynamics simulation. Results showed that the optimal enzyme activity ratio of 1:0.94:1.07 for Alcalase, trypsin, and Flavourzyme, combined with optimized hydrolysis conditions (E/S ratio of 126,793.03 nkat/g, pH 8.40, 4.82 h hydrolysis time, and 45 °C), resulted in a peptide yield of 93.19 ± 0.14%, ACE-inhibitory rate of 95.92 ± 0.23%, and bitter value of 3.15 ± 0.09. APQLGR and APWLR exhibited high ACE-inhibitory activity and minimal bitterness among the seventeen identified peptides. Although both peptides bound to the S1 pocket and Zn²⁺ catalytic site of ACE, APWLR exhibited an additional interaction with the S2 pocket. Both peptides were predicted to antagonize the bitter taste receptor T2R14 by forming stable complexes with key residues, but two complexes exhibited distinct mechanisms of stabilization. This work demonstrates a method for producing dual-functional peptides from sesame protein, paving the way for their application in functional foods. Full article

Jellyfish, as high-biomass marine resources, frequently exhibit explosive proliferation in coastal waters and possess both nutritional functional factors and potential medicinal value. This study investigated the enzymatic hydrolysis of two jellyfish species, Rhopilema hispidum (R. hispidum) and Nemopilema nomurai (N. nomurai), using Alcalase, Flavourzyme, and Protamex, with a specific focus on the molecular weight distribution and antioxidant activity. The optimal hydrolysis conditions were systematically determined via single-factor experiments followed by orthogonal test optimization. The Flavourzyme hydrolysates had the highest proportion of low-molecular-weight peptides (<3 kDa) and exhibited the most potent antioxidant activity, while Protamex hydrolysates had more high-molecular-weight peptides (>3 kDa, >40%) with comparatively weaker antioxidant activity. R. hispidum hydrolysates exhibited stronger DPPH• and O₂•⁻ scavenging activities and contained a higher proportion of low-molecular-weight peptides (<3 kDa), whereas N. nomurai hydrolysates showed a higher degree of hydrolysis, and its hydrolysates demonstrated superior •OH scavenging activity. Chromatographic analysis confirmed low-molecular-weight peptides positively correlated with antioxidant potential. This study highlights molecular weight distribution, together with enzyme type, as a pivotal determinant of the antioxidant activity of jellyfish hydrolysates, providing insights for antioxidant peptide development. Full article

As the global population continues to grow, so does the demand for alternative protein sources. Entomophagy, the consumption of insects, has long been practiced in many cultures worldwide and is now gaining increasing interest in Western countries. In this work, we developed novel, functional insect-based ingredients from the house cricket (Acheta domesticus) by utilizing optimized enzymatic hydrolysis, using two enzymes (Alcalase^® or Flavourzyme^®) coupled with spray drying. A Box–Behnken experimental design was used to optimize enzymatic treatments and maximize spray-drying performance and product solubility. Under optimized conditions, spray-dried hydrolyzed cricket protein (HCP) produced using Alcalase^® achieved a solids recovery of 51.44% and a solubility of 58.28 ± 0.5%. In comparison, Flavourzyme^®–HCP, under optimized conditions, exhibited a higher solubility of 61.25 ± 0.8%. Additional functional properties were improved for Alcalase^®–HCP and Flavourzyme^®–HCP, respectively, including foaming capacity at pH 4 (26.80 ± 4.0%, 36.27 ± 1.0%) and 10 (50.98 ± 2.8%, 47.06 ± 1.6%), and foaming stability in acidic conditions at pH 4 (24.18 ± 4.0%, 30.39 ± 2.9%). Moreover, the emulsion stability, especially at pH 7 (74.70 ± 3.5%, 52.04 ± 2.8%) and 10 (68.20 ± 11.3%, 69.72 ± 3.2%), was also enhanced. To the best of our knowledge, this is the first study to investigate optimized enzymatic hydrolysis coupled with spray drying to enhance the functional properties of A. domesticus protein powder. Overall, we established optimized processing conditions to produce spray-dried functional insect ingredients with desirable functional attributes. Full article

Plant-based bioactive compounds have been recognized as promising alternatives to conventional chemical treatments. Selenium (Se), a trace element, can be incorporated into proteins to enhance the bioactivity of plant-derived peptides. Perilla frutescens seeds are high-protein plants that have shown the ability to absorb Se and biosynthesize selenopeptides. This study examined Se biotransformation during the germination of perilla seeds to synthesize selenoprotein, investigating enzymatic hydrolysis using Alcalase and Flavourzyme as single enzymes, as well as their combinations. The results showed that Alcalase hydrolysates produced Se-peptides with the highest degree of hydrolysis and antioxidant activity. Hydrolysates were purified via ultrafiltration and size-exclusion chromatography, and Se-peptides were characterized by LC-MS/MS. Nine peptides containing Se-binding residues such as cysteine, methionine, and glutamic acid confirmed successful Se incorporation. The Se-peptides demonstrated strong antioxidant activity (ABTS: 66.30%, FRAP: 54.93%), ACE inhibition (83.87%), and cytotoxicity against A549 lung cancer cells (85.88% viability). Compared to non-Se-peptides, Se-enriched peptides showed superior bioactivity, highlighting their potential as functional ingredients in food and pharmaceutical applications. Full article

Enzymatic hydrolysis of Atlantic salmon processing discards, including heads and frames, was carried out using Alcalase, Flavourzyme, and their combination. The degree of hydrolysis varied depending on the enzyme and substrate, ranging from approximately 5% to 15%. Alcalase was more effective than Flavourzyme for the hydrolysis of salmon frames and heads. The results also indicated that 1% enzyme concentrations had slightly higher mean DH than 2%. The bioactivity testing indicated that the head hydrolysates had superior DPPH scavenging activity, while frame hydrolysates demonstrated higher ABTS and hydroxyl radical scavenging activity. The correlation between DH and bioactivities indicated that the degree of hydrolysis did not have any significant effect on the DPPH and hydroxyl radical scavenging activities while demonstrating a significant positive correlation against ABTS scavenging activity. Full article

Hypertension and type 2 diabetes are the major metabolic syndromes, often managed using synthetic ACE and DPP-IV inhibitors that may cause adverse effects on health. This study investigated Bambara groundnut protein hydrolysates as a natural source of dual ACE- and DPP-IV-inhibitory peptides. Protein isolates were hydrolyzed using Flavourzyme, and the resulting peptides were fractionated using membranes with different molecular weight cut-offs. Those fractions were then analyzed for enzyme inhibition. Peptides were identified by LC-MS/MS and screened using PeptideRanker and BIOPEP-UWM, followed by molecular docking against ACE (PDB: 1O8A) and DPP-IV (PDB: 1NU6). The >10 kDa and 5–10 kDa fractions showed the highest ACE- and DPP-IV-inhibitory activities, respectively. Some peptides such as YKDGLYSPHW, LPVSTPGKF, and EPWWPK displayed strong binding affinities (ΔG: −10.2 to −11.3 kcal/mol for ACE, −8.6 to −9.1 kcal/mol for DPP-IV) and interacted with key catalytic residues, including His387 and Glu411 in ACE, and Ser630, Glu205, and Phe357 in DPP-IV. These findings highlight the potential of Bambara groundnut hydrolysates or peptides as a source of natural ACE and DPP-IV inhibitors. Full article

The search for new alternatives for the revalorization of porcine blood is crucial due to the large quantities that are annually generated in slaughterhouses. In this study, a sequential enzymatic hydrolysis of pig blood was optimized using different combinations of the enzymes, namely, Alcalase 4.0 L and Protana™ Prime, Flavourzyme 1000 L, and Protamex^®, as a sustainable method for obtaining extracts rich in bioactive peptides. All the assayed hydrolysates exhibited different peptide profiles and showed in vitro antioxidant, hypoglycemic, and anti-inflammatory activity, although their values differed significantly depending on the type of hydrolysis in ABTS, FRAP, and ORAC assays, as well as in the determination of the inhibitory activity of DPP-IV, NEP, TACE, and MGL enzymes. The hydrolysate obtained by the combination of Alcalase 4.0 L, Flavourzyme 1000 L, and Protana™ Prime (AFPP) resulted in the highest hydrolysis degree (33.39 ± 0.98%), and its peptide profile reflected a higher amount of peptides < 3 kDa. This hydrolysate also obtained significantly higher values for ABTS and the inhibition of TACE and MGL. However, APP2 stood out in NEP inhibition (79.39 ± 3.91%), while APPP was notable for DPP-IV inhibition (43.02 ± 1.39%). The analysis of the hydrolysates using mass spectrometry in tandem allowed for the identification of those sequences that are potentially responsible for the biological activities determined, which were characterized using in silico bioinformatic tools. The results show the potential of using sequential enzymatic hydrolysis in porcine blood to obtain multifunctional peptides. Full article

Defatted grape seed meal (DGSM) is a residue obtained from grape pomace and is an important source of protein. The aim of this study was to select peptides with optimal antioxidant and colour properties, obtained using enzymes of different origins and proteolytic character, for application in winemaking. For this purpose, the assay was performed using novo-ProD (NP), alcalase (AL), novozym (NZ), pepsin (PE), flavourzyme (FZ), and papain (PA) enzymes. The peptide percentage, peptide yield, molecular size of the peptide fractions, total amino acid, peptide content, antioxidant activity, and CIELAB colour coordinates of the hydrolysates were determined. The peptide hydrolysates obtained using PE showed the significantly (p < 0.05) highest percentages of peptides (93%), amino acid content (188 mg aa/g hydrolysate), and lightness (L*, 70.3). On the other hand, NP peptide hydrolysates displayed the significantly (p < 0.05) highest antioxidant activity (154 µmol TE/g hydrolysate) and peptide yield (39%). Regarding molecular weight (MW), PE led to hydrolysates with a lower proportion of low-MW peptides (MW < 1 kDa). In conclusion, the peptide hydrolysates obtained by NP and PE exhibited the greatest chemical characteristics for further application, both separately and combined in targeted hydrolysis, as colour stabilisers and antioxidant capacity enhancers in warm climate winemaking. Full article

Protein constitutes the primary nutrient in soy, and its modifications are intricately linked to the properties of the soy milk powder. This study employed six main commercial enzymes (bromelain, neutrase, papain, trypsin, flavourzyme, and alcalase) to investigate the impact of enzymatic hydrolysis on the structural and functional properties of soy protein isolate (SPI), as well as its influence on the physicochemical properties of soy milk powder. The findings indicated that each of enzymes exhibits distinct specificity, with the degree of hydrolysis following the order: alcalase > flavourzyme > papain > bromelain > neutrase > trypsin. Enzymatic hydrolysis facilitates the unfolding of SPI, leading to the exposure of chromogenic fluorophores and hydrophobic amino acid residues, which in turn promotes an increase in free sulfhydryl content. Concurrently, this process induces the transformation of α-helix and β-sheet into β-turn and random coil. The enzyme modification enhances the solubility, emulsification, and foaming activities of SPI and significantly augment its antioxidant properties (p < 0.05). However, this enzymatic treatment adversely affects the stability of its emulsification and foaming properties. Subsequent to enzymatic hydrolysis, soy milk powder demonstrated a reduction in particle size and an improvement in solubility, which significantly enhanced its flavor profile. In summary, alcalase offers substantial advantages in augmenting the functional properties of SPI and increasing the solubility of soy milk powder. However, this process adversely affects the flavor profile of soy milk powder, a consequence attributed to the broad hydrolysis specificity of alcalase. Full article

Pumpkin seed proteins (PSPs) are a promising resource for obtaining bioactive peptides but their low solubility hinders enzymatic hydrolysis, reducing yield and bioactivity. In addition, enzymatic processes require specific conditions and long processing times; improving the efficiency of this process is essential to expand its industrial applications. In this context, using a high-frequency, low-intensity ultrasound (US) has proven to be an effective strategy for optimizing the hydrolysis of plant protein. This study evaluated the US-assisted (38 W/L, 40 kHz) and conventional hydrolysis of pumpkin seed proteins (PSPs) for 180 min at 25 °C, 40 °C, and at the optimum temperature condition for each enzyme studied (60 °C for Brauzyn^®, 55 °C for Flavourzyme^®, and 50 °C for Neutrase^®), as well as the impact of this process on the macrostructural and functional characteristics of the hydrolysates obtained. The degree of hydrolysis (DH) was significantly higher in US-assisted reactions, reaching increases of up to 57.7% with Neutrase^® at 40 °C. The US also positively influenced the protein solubility of the hydrolysates, especially at pH levels close to the isoelectric point, with improvements of up to 100%, compared to the hydrolysates obtained from the conventional reaction. The antioxidant activity was also enhanced by the US, compared to the conventional reaction, emphasizing the hydrolysates obtained through the action of Flavourzyme^®, which showed increases of 52.4% and 42.6% in the scavenging of DPPH and ABTS radicals, respectively. The analysis of the mean particle size revealed significant reductions with the US (<26.2%). Consequently, the polydispersity index (PDI) demonstrated greater uniformity in the particles obtained from the US-assisted reactions (reductions of up to 20.3%). UV-Vis spectroscopy and intrinsic fluorescence also indicated possible alterations in the tertiary structure of the peptides obtained, mainly in US-assisted reactions. Therefore, US-assisted PSP hydrolysis resulted in better enzymatic performance and produced protein hydrolysates with bioactive potential for food applications. Full article