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Commercial isolated yellow field pea protein isolate (IPP) was subjected to 200, 400 and 600 MPa high hydrostatic pressure (HHP) treatments followed by determination of some physicochemical and functional properties. Native polyacrylamide gel electrophoresis confirmed HHP-induced formation of high molecular weight protein aggregates. Intrinsic fluorescence showed most intense at 600 MPa where fluorescence intensity was less than half of the control IPP. The 600 MPa-treated IPP also showed more unfolded structure with an extensive red shift (384 nm) in wavelength of maximum tryptophan fluorescence. However, solubility profile was very similar and was not significantly ( p > 0.05) affected by HHP treatment. At pH 3.0, HHP-treated IPP formed significantly ( p < 0.05) higher quality emulsions with oil droplet sizes ( d 4,3 ) of 26–68 μm when compared to 52–92 μm for the control IPP. Foaming capacity was also higher at pH 3.0 with a maximum value of 81% when compared to maximum values of 38% and 62% obtained, respectively at pH 5.0 and 7.0. [ABSTRACT FROM AUTHOR]

Enzyme-assisted aqueous extraction processing (EAEP) is an environmentally-friendly alternative to solvent and mechanical oil extraction methods, and can achieve ∼97% oil recovery from soybeans. The present study utilized soy skim (protein rich) and insoluble fiber (IF; carbohydrate rich), both co-products of EAEP, in dry-grind corn fermentation. The effects of adding soy skim and untreated IF (UIF), either separately or together, and adding pretreated IF (TIF), on ethanol production were investigated. Maximum ethanol production was achieved when UIF and skim were slurried together (corn-to-UIF ratio 1:0.16; skim-to-UIF ratio 6.5:1) and when fiber-hydrolyzing enzymes were added to corn fermentation. This modification to corn fermentation increased ethanol yield by 20%, ethanol production rate by 3%, and decreased fermentation time by 38 h compared to corn-only fermentation. An attempt was also made to utilize pentoses (from soy skim and IF) in integrated corn-soy fermentation slurry by an additional Escherichia coli KO11 fermentation step. [ABSTRACT FROM AUTHOR]

A sublethally injured bacterial cell has been defined as a cell that survives a stress such as heating, freezing, acid treatment, or other antimicrobial intervention but can repair the cellular damage exerted by the stressor and later regain its original ability to grow. Consequently, sublethally injured cells are not likely to be included in conventional enumeration procedures, which could result in unrealistically low counts unless efforts are made to encourage recovery of the injured cells before enumeration. The objective of this study was to evaluate the use of the thin agar layer (TAL) method for the recovery of pressure-injured and heat-injured Listeria monocytogenes in a tryptic soy broth with 0.6% yeast extract system. Pressure injury consisted of treatment of a culture of mixed L. monocytogenes strains with high hydrostatic pressure at 400 or 600 MPa for 1 s, 2 min, 4 min, or 6 min at a process temperature of 12 ± 2°C. Heat injury consisted of treatment of a culture of mixed L. monocytogenes strains at 60 ± 1°C for 3, 6, or 9 min. Growth media were tryptic soy agar (TSA) with 0.6% yeast extract, modified Oxford medium (MOX), and TAL, which consisted of a 7-ml layer of TSA overlaid onto solidified MOX. Counts of viable L. monocytogenes on TAL were higher than those on MOX in the heat-injury experiment but not in the pressure-injury experiment. Therefore, the effectiveness of the TAL method may be specific to the type of injury applied to the microorganism and should be investigated in a variety of cellular injury scenarios. [ABSTRACT FROM AUTHOR]

Sodium nitrite exerts an inhibitory effect on the growth of Listeria monocytogenes. The objective of this study was to investigate the effects of various nitrite concentrations from a vegetable source with and without high hydrostatic pressure (HHP) on the recovery and growth of L. monocytogenes on ready-to-eat restructured ham. A preconverted celery powder was used as the vegetable source of nitrite. Targeted concentrations of natural nitrite investigated were 0, 50, and 100 mg/kg. HHP treatments evaluated were 400 MPa for 4 min and 600 MPa for 1 or 4 min at 12 ± 2°C (initial temperature of the pressurization fluid). Viable L. monocytogenes populations were monitored on modified Oxford medium and thin agar layer medium through 98 days of storage at 4 ± 1°C. Populations on both media did not differ. The HHP treatment at 600 MPa for 4 min resulted in L. monocytogenes populations below the detection limit of our sampling protocols throughout the storage period regardless of the natural nitrite concentration. The combination of HHP at 400 MPa for 4 min or 600 MPa for 1 min with natural nitrite resulted in initial inhibition of viable L. monocytogenes. Ham formulations that did not contain natural nitrite allowed faster growth of L. monocytogenes than did those with nitrite, regardless of whether they were treated with HHP. The results indicate that nitrite from a vegetable source at the concentrations used in this study resulted in slower growth of this microorganism. HHP treatments enhanced the inhibitory effects of natural nitrite on L. monocytogenes growth. Thus, the combination of natural nitrite plus HHP appears to have a synergistic inhibitory effect on L. monocytogenes growth. [ABSTRACT FROM AUTHOR]

Abstract: Microalgae are a promising source of lipids for biofuel production. To improve the economic feasibility and sustainability of this biofuel feedstock, one should create value for co-products after lipid extraction. Thus, protein isolation from the defatted biomass presents an opportunity. To extract algae protein, temperature and pH were evaluated to maximize the extraction from Nannochloropsis biomass. Maximum quantity of protein was solubilized at 60°C and pH11 and recovered at pH3.2. The isolated protein fractions contained 56.9% and 40.5% protein when using isopropanol (IPA) defatted and non-defatted biomass as the starting materials, with protein yields being 16 and 30%, respectively. The IPA-defatting treatment significantly decreased the protein extraction yield. These values are low compared with soybean protein isolates (>90% protein and ~60% yield). The relatively high protein content (>34%) in the pH11 insoluble fraction indicates needs for further extraction optimization. The nitrogen and amino acid content of the initial materials and all the fractions were determined and the calculated nitrogen to protein conversion factor was in the range of 4.06–4.70. The possibility of the presence of conjugated protein, i.e., N-containing glycoproteins, is also discussed. [Copyright &y& Elsevier]

Abstract: The effect of high pressure treatment on whole egg white was examined using in vitro pepsin digestion and proteomic methods. Pepsin incubations conducted with an enzyme to protein ratio of 3:1 following high pressure treatment (400–800MPa and 9°C) resulted in increased hydrolysis of egg white proteins. Pressure treatment of egg white at 800MPa resulted in greater susceptibility to pepsin hydrolysis than did thermal treatment at 95°C. The effect of 800MPa pressure treatment on egg white proteins was additionally examined by incubation with pepsin at an enzyme to protein ratio of 1:20 followed by 2-D electrophoresis. Results of these experiments showed extensive hydrolysis of most egg white proteins. Subsequent LC-MS/MS investigation of the low Mr fraction (<3.0kDa) derived from pepsin digested 800MPa treated egg white contained numerous peptides previously shown to have bioactive and/or immunological properties. Industrial Relevance: Short time high pressure treatment of whole egg white at relatively low temperature (9°C) resulted in increased pepsin digestibility equivalent to or better than heat treatment at 95°C. Examination of the peptides resulting from high pressure treatment and pepsin digestion revealed sequences with known biological activities. Thus high pressure treatment represents a promising technology for enhancing egg white''s healthiness and contributing to its role as a functional food. [Copyright &y& Elsevier]

The platelet collagen receptor glycoprotein VI (GPVI) has been suggested to function as a dimer, with increased affinity for collagen. Dissociation constants (Kd) obtained by measuring recombinant GPVI binding to collagenous substrates showed that GPVI dimers bind with high affinity to tandem GPO (Gly-Pro-Hyp) sequences in collagen, whereas the markedly lower affinity of the monomer for all substrates implies that it is not the collagen-binding form of GPVI. Dimer binding required a high density of immobilized triplehelical (GPO)10-containing peptide, suggesting that the dimer binds multiple, discrete peptide helices. Differential inhibition of dimer binding by dimer-specific antibodies, m-Fab-F and 204-11 Fab, suggests that m-Fab-F binds at the collagen-binding site of the dimer, and 204-11 Fab binds to a discrete site. Flow cytometric quantitation indicated that GPVI dimers account for ∼29% of total GPVI in resting platelets, whereas activation by either collagen-related peptide or thrombin increases the number of dimers to ∼39 and ∼44%, respectively. m-Fab-F inhibits both GPVI-dependent static platelet adhesion to collagen and thrombus formation on collagen under low and high shear, indicating that pre-existing dimeric GPVI is required for the initial interaction with collagen because affinity of the monomer is too low to support binding and that interaction through the dimer is essential for platelet activation. These GPVI dimers in resting circulating platelets will enable them to bind injury-exposed subendothelial collagen to initiate platelet activation. The GPVI-specific agonist collagen-related peptide or thrombin further increases the number of dimers, thereby providing a feedback mechanism for reinforcing binding to collagen and platelet activation. [ABSTRACT FROM AUTHOR]

Abstract: Oil, protein and solid extraction yields obtained during aqueous extraction processing (AEP) of full fat soybean flakes (FFSF), FFSF extruded at a die temperature of 100°C and FFSF pressurised at 200 and 500MPa for 15min at 25°C, were compared to those obtained during enzyme-assisted aqueous extraction processing (EAEP) using 0.5% of protease Protex 7L. Without enzyme addition, pretreatment of the FFSF with extrusion and 500MPa increased and decreased, respectively, the oil extraction yield while protein extraction yield was significantly decreased after both treatments. The best treatment in terms of oil and protein recovery was EAEP of extruded flakes with 90% and 82% of oil and protein extraction yield, respectively, and 17% of free oil. Addition of protease during extraction significantly decreased the yield of isolated soy protein (ISP) due to an increased solubility of the proteins at pH 4.5. ISP from extruded EAEP had higher solubility at pH 7.0 and better functionality. The DSC results, combined with the protein extraction yields, showed that a proportion of the proteins became insoluble after extrusion and 500MPa treatment, while only those extracted from 500MPa FFSF had a reduced native state. [Copyright &y& Elsevier]

Abstract: High hydrostatic pressure was applied to hydrated soybeans (100–700MPa, 25°C) and soymilk (400–750MPa; 25 and 75°C) to assess its effect on isoflavone content, profile and water-extractability. Neither pressure level nor initial treatment temperature affected soymilk isoflavone content. However, combined pressure and mild thermal treatment modified the isoflavone distribution. At 75°C, the isoflavone profile shifted from malonylglucosides toward β-glucosides, which was correlated to the effect of adiabatic heating. When pressure was applied to the hydrated soybeans, the soymilk isoflavone concentration varied between 4.32 and 6.06μmol/g. The content of protein decreased and fat increased in soymilks prepared from pressurized soybeans with increasing pressure level. [Copyright &y& Elsevier]

Abstract: The abundant Rap1 in platelets becomes activated when these cells are stimulated by various agonists, but its function has remained unknown. In view of this, we developed an assay to quantitatively measure activated Rap1 and used it to determine relationships between Rap1 activation and several platelet functions: integrin α2β1 activation, tyrosine phosphorylation, and the release reaction. We looked at how these processes are affected by the protein kinase C inhibitor BIMI, tyrosine kinase inhibitor PP2, PI 3-kinase inhibitor wortmannin, and ADP scavenger apyrase. In CRP (collagen related peptide)-activated platelets, all the inhibitors severely inhibited Rap1 activation, but had little effect on integrin α2β1 activation, indicating that the integrin activation mechanism is different from the Rap1 activation mechanism, at least in GPVI-dependent activation. With p85α-null mouse platelets, we demonstrated that Rap1 activation involves PI 3-kinase p85α-dependent tyrosine phosphorylation. All the inhibitors similarly decreased Rap1 activation and the serotonin release reaction, and the inhibition of Rap1 activation was not due to the lack of released ADP. Our results indicate that platelet Rap1 activation is closely related to the release reaction and not to integrin α2β1 activation in GPVI-mediated platelet activation. [Copyright &y& Elsevier]

Glycoprotein (GP) VI is a platelet membrane protein with a molecular weight of 62 kDa that was identified as a physiological collagen receptor from studies of patients deficient in this protein. GPVI-deficient platelets lacked specifically collagen-induced aggregation and the ability to form thrombi on a collagen surface under flow conditions, suggesting that GPVI makes an indispensable contribution to collagen-induced platelet activation. On the platelet surface, GPVI is present as a complex with the Fc receptor (FcR) γ-chain, probably composed of two GPVI molecules and one FcR γ-chain dimer. GPVI must form such a dimeric complex to exhibit high affinity binding to collagen. The GPVI-induced activation mechanism is initiated by tyrosine phosphorylation of the immunoreceptor tyrosine-based activation motif (ITAM) of the FcR γ-chain, and then this signal is transduced to many related proteins, mainly by tyrosine phosphorylation. GPVI is widely recognized as a requisite factor for the formation of platelet aggregates on a collagen surface under blood flow. However, individuals with GPVI-deficient or null platelets do not exhibit any strong bleeding tendency. Analyzing this apparent dichotomy should provide us with a more precise understanding of the mechanism of thrombus formation. [Copyright &y& Elsevier]

Integrated corn-soy fermentation, utilizing co-products of soybean enzyme-assisted aqueous extraction process (EAEP) of soybeans in corn fermentation, has shown potential for enhancing bioethanol production compared to corn only fermentation. To maximize economic returns, oil may be recovered. In the present study, the effect of skim and insoluble fiber, and oil extraction aids on ethanol yield, oil partition, and oil recovery, and quality of Distillers Dried Grains (DDG) was investigated. Two fiber hydrolyzing enzymes (pectinase and cellulase), an acid protease (Fermgen ® ), and a surfactant (Tween 80) were evaluated. Addition of skim, mixture of skim and insoluble fiber, or Fermgen ® to corn fermentation resulted in a ∼32 h decrease in fermentation time. Addition of soy co-products also resulted in ∼10–28% increase in oil partition in thin stillage with no additional enzyme or surfactant treatment. Addition of insoluble fiber alone resulted in ∼19% decrease in solids partition in thin stillage. Maximum free oil recovery, 22.5 ± 4.5%, was achieved from corn-insoluble fiber thin stillage with a combined treatment of enzymes (pectinase, cellulase, and Fermgen ® ) and surfactant (Tween 80). Maximum extractable oil recovery, 67 ± 3.2%, was achieved with the enzyme treatment alone. Corn-soy DDG has ∼11% higher protein, ∼2% lower fiber, and ∼2% lower fat contents compared to corn DDG. The fiber content was further reduced to ∼2% after enzyme treatment. This study demonstrates an efficient use of soy EAEP co-products and enzymes to maximize oil partition in thin stillage, and produce a high quality corn-soy DDG. [ABSTRACT FROM AUTHOR]

Commercial isolated pea protein dispersion (IPP, 1%, w/v) was pretreated with high pressure (200–600MPa, 5min at 24°C) or heat (100°C, 30min) prior to hydrolysis using 1–4% (w/w) alcalase concentrations. Fluorescence spectroscopy showed that heat pretreated IPP had a 35% higher level of exposed hydrophobic groups (measured as fluorescence intensity, FI) than the untreated protein. In contrast, the 200MPa pressure pretreatment produced a 15% increase in FI while 400 and 600MPa pretreatments, respectively, caused 5 and 60% decreases in FI. Heat pretreatment of IPP enhanced hydrolysis into smaller peptide sizes when compared to peptides from the 24°C pretreated protein. The 200MPa pretreatment enhanced IPP hydrolysis into smaller peptides, especially at lower (1–2%) alcalase concentrations. Protein hydrolysates produced from heat-pretreated IPP were less active against angiotensin converting enzyme (ACE) when compared to those from the 24°C pretreated protein. In general, heat or high pressure pretreatment of IPP favored production of ACE- and renin-inhibitory enhanced protein hydrolysates at a lower (1%) alcalase concentration. [ABSTRACT FROM AUTHOR]

Abstract: The effectiveness of several pretreatments [high-power ultrasound, sulfuric acid (H2SO4), sodium hydroxide (NaOH), and ammonium hydroxide (NH3OH)] to enhance glucose production from insoluble fractions recovered from enzyme-assisted aqueous extraction processing of extruded full-fat soybean flakes (FFSF) was investigated. Sonication of the insoluble fraction at 144μmpp (peak-to-peak) for 30 and 60s did not improve the saccharification yield. The solid fractions recovered after pretreatment with H2SO4 [1% (w/w), 90°C, 1.5h], NaOH [15% (w/w), 65°C, 17h], and NH3OH [15% (w/w), 65°C, 17h] showed significant lignin degradation, i.e., 81.9%, 71.2%, and 75.4%, respectively, when compared to the control (7.4%). NH3OH pretreatment resulted in the highest saccharification yield (63%) after 48h of enzymatic saccharification. A treatment combining the extraction and saccharification steps and applied directly to the extruded FFSF, where oil extraction yield and saccharification yield reached 98% and 43%, respectively, was identified. [Copyright &y& Elsevier]

Abstract: This research focused on the use of high-power ultrasound prior to soy protein extraction to simultaneously enhance protein and sugar release in the extract. Defatted soy flakes dispersed in water were sonicated for 15, 30, 60 and 120s using a bench-scale ultrasound unit. The ultrasonic amplitudes used were: 0, 21, 42, 63 and 84μmpp (peak-to-peak). The respective power densities were 0.30, 0.87, 1.53 and 2.56W/ml. Scanning electron micrographs of sonicated samples showed the structural disruption of soy flakes. The particle size decreased nearly 10-fold following ultrasonic treatment at high amplitudes. Sonication at high amplitude for 120s gave the highest increase in total sugar released (50%) and protein yield (46%) when compared with non-sonicated samples (control). Ultrasonic pretreatment was also carried out with and without cooling for temperature moderation. The heat generated during sonication had no significant effect on protein and sugar release from defatted soy flakes. The use of ultrasound can significantly improve protein yield and reduce the overall cost of producing soy protein from flakes. [Copyright &y& Elsevier]

Isolated pea protein (IPP) dispersions (1%, w/v) were pretreated with high pressure (HP) of 200, 400, or 600 MPa for 5 min at 24 °C or high temperature (HT) for 30 min at 100 °C prior to hydrolysis with 1% (w/w) Alcalase. HP pretreatment of IPP at 400 and 600 MPa levels led to significantly ( P < 0.05) improved (>40%) oxygen radical absorption capacity (ORAC) of hydrolysates. 2,2-Diphenyl-1-picrylhydrazyl, superoxide radical and hydroxyl radical scavenging activities of pea protein hydrolysates were also significantly ( P < 0.05) improved (25%, 20%, and 40%, respectively) by HP pretreatment of IPP. Protein hydrolysates from HT IPP showed no ORAC, superoxide or hydroxyl scavenging activity but had significantly ( P < 0.05) improved (80%) ferric reducing antioxidant power. The protein hydrolysates had weaker antioxidant properties than glutathione but overall, the HP pretreatment was superior to HT pretreatment in facilitating enzymatic release of antioxidant peptides from IPP. [ABSTRACT FROM AUTHOR]

Background: Glycoprotein VI (GPVI) is the essential platelet collagen receptor in atherothrombosis, but its inhibition causes only a mild bleeding tendency. Thus, targeting this receptor has selective antithrombotic potential.Objectives: This study sought to compare compounds interfering with platelet GPVI-atherosclerotic plaque interaction to improve current antiatherothrombotic therapy.Methods: Human atherosclerotic plaque-induced platelet aggregation was measured in anticoagulated blood under static and arterial flow conditions (550/s, 1,100/s, and 1,500/s). Inhibition by dimeric GPVI fragment crystallizable region of IgG (Fc) masking GPVI binding sites on collagen was compared with that of 3 anti-GPVI antibodies: BLO8-1, a human domain antibody; 5C4, a fragment antigen-binding (Fab fragment) of monoclonal rat immunoglobulin G; and m-Fab-F, a human recombinant sFab against GPVI dimers.Results: GPVI-Fc reduced plaque-triggered platelet aggregation in static blood by 51%, BLO8-1 by 88%, and 5C4 by 93%. Under arterial flow conditions, BLO8-1 and 5C4 almost completely inhibited platelet aggregation while preserving platelet adhesion on plaque. Inhibition by GPVI-Fc, even at high concentrations, was less marked but increased with shear rate. Advanced optical imaging revealed rapid persistent GPVI-Fc binding to collagen under low and high shear flow, upstream and downstream of plaque fragments. At low shear particularly, platelets adhered in plaque flow niches to GPVI-Fc-free segments of collagen fibers and recruited other platelets onto aggregates via ADP and TxA2 release.Conclusions: Anti-GPVI antibodies inhibit atherosclerotic plaque-induced platelet aggregation under static and flow conditions more effectively than GPVI-Fc. However, potent platelet inhibition by GPVI-Fc at a higher shear rate (1,500/s) suggests localized antithrombotic efficacy at denuded or fissured stenotic high-risk lesions without systemic bleeding. The compound-specific differences have relevance for clinical trials targeting GPVI-collagen interaction combined with established antiplatelet therapies in patients with spontaneous plaque rupture or intervention-associated plaque injury. [ABSTRACT FROM AUTHOR]

Abstract: The objective of this study was to investigate natural antimicrobials including cranberry powder, dried vinegar and lemon juice/vinegar concentrate, and post-lethality interventions (lauric arginate, octanoic acid, thermal treatment and high hydrostatic pressure) for the control of Listeria monocytogenes on alternatively-cured frankfurters. Lauric arginate, octanoic acid, and high hydrostatic pressure (400MPa) reduced L. monocytogenes populations by 2.28, 2.03, and 1.88log10CFU per g compared to the control. L. monocytogenes grew in all post-lethality intervention treatments, except after a 600MPa high hydrostatic pressure treatment for 4 min. Cranberry powder did not inhibit the growth of L. monocytogenes, while a dried vinegar and a vinegar/lemon juice concentrate did. This study demonstrated the bactericidal properties of high hydrostatic pressure, octanoic acid and lauric arginate, and the bacteriostatic potential of natural antimicrobial ingredients such as dried vinegar and vinegar/lemon juice concentrate against L. monocytogenes. [Copyright &y& Elsevier]

Abstract: High pressure processing of ovotransferrin was carried out to study the structural and physiochemical changes of ovotransferrin under various pressure levels. At pH 8 and pressures higher than 200MPa, a decrease in total sulfhydryl groups and an increase in surface hydrophobicity were observed along with a partial aggregation. A gradual shift of denaturation peak towards higher temperature was noticed up to 500MPa, leading to a total loss of the enthalpy of denaturation at pressures of 600 and 700MPa, where a significant decrease in intrinsic fluorescence was also observed. At pH 3, the ovotransferrin adopted a molten globule state, associated with a significant increase in surface hydrophobicity and reactive sulfhydryl content; structurally, no clear denaturation peaks in differential scanning calorimetry (DSC) were detected at any level of pressure treatment whereas a noticeable decrease in intrinsic fluorescence was evidenced up to 600MPa and then increased at 700MPa pressure treatment. Fourier transform infrared spectroscopy (FT-IR) revealed that the conformational structure were changed from helices, sheets, turns, and aggregated strand to mostly intermolecular β-sheets or aggregated strands at pH 8 at 200MPa but switched back to original structure at higher pressures. [Copyright &y& Elsevier]

Abstract: Soybean insoluble fractions recovered from aqueous extraction processing (AEP) and enzyme-assisted AEP (EAEP) of full-fat soybean flakes (FFSF) and extruded FFSF were evaluated as a feedstock for the production of fermentable sugars using enzymes. Among the four insoluble fractions (AEP FFSF, EAEP FFSF, AEP extruded FFSF and EAEP extruded FFSF), the composition analysis revealed that the one recovered from EAEP of extruded FFSF had the highest glucan content, 16% [dry basis (db)], as compared to about 10% (db) for the other fractions. Thirty-three percent of the initial glucan of the insoluble recovered from AEP and EAEP of FFSF were converted into glucose using 33 FPU of Accellerase 1000/g-glucan. This saccharification yield was increased to 44% with extruded fibers. The higher saccharification yield of 49% was obtained at 45°C, 1% glucan loading, and 101 FPU/g-glucan enzymes loading after 27h of hydrolysis. [ABSTRACT FROM AUTHOR]