Emulsifying and foaming properties of transglutaminase-treated wheat gluten hydrolysate as influenced by pH,temperature and salt

Hydrolyzed wheat gluten (GH, 77–85% protein) was prepared by limited hydrolysis with chymotrypsin at 37 °C for 4 h (degree of hydrolysis=6.4%) and 15 h (degree of hydrolysis=10.3%). The effect of microbial transglutaminase (MTGase) treatment (55 °C for 1 h, or 5 °C for 18 h) on the emulsifying and foaming properties of GH was evaluated under selected food processing conditions (pH 4.0 and 6.5, 0 and 0.6 M NaCl, and temperature 20 and 5 °C). At pH 4.0 and 0 M NaCl the MTGase treatment substantially increased foaming capacity (FC) of GH compared with their respective control GH samples, as a result of enhanced peptide adsorption to the air–water interface, but FC was similar for both control and MTGase-treated GH at pH 6.5. In contrast, foam drainage stability (FS) of MTGase-treated GH decreased at pH 4.0, but increased significantly (P<0.05) at pH 6.5 when compared with their respective control GH samples. The FC and FS were affected by 0.6 M NaCl in a pH-dependent manner. The MTGase treatments increased emulsion activity index up to 15-fold at pH 6.5, while emulsion stability index was influenced by emulsion temperature and ionic strength conditions. The MTGase-induced changes in functional properties of GH were attributed to pH-dependent solubility changes, the amphiphilic nature of gluten peptides and increased electrostatic repulsion resulting from deamidation.     

Effect of microbial transglutaminase treatment on thermal stability and pH-solubility of heat-shocked whey protein isolate

Whey protein isolate (WPI) dispersions (5% protein, pH 7.0) were subjected to heat-shock at 70 °C for 1, 5 and 10 min. The heat-shocked WPI dispersions were treated with microbial transglutaminase (MTGase) enzyme, and thermal properties and pH-solubility of the treated proteins were investigated. Heat-shocking of WPI for 10 min at 70 °C increased the thermal denaturation temperature (T_d) of β-lactoglobulin in WPI by about 1.5 °C. MTGase treatment (30 h, 37 °C) of the heat-shocked WPI significantly increased the T_d of β-lactoglobulin by about 6.3–7.3 °C when compared with heat-shocked only WPI at pH 7.0. The T_d increased by about 13–15 °C following pH adjustment to 2.5; however, the T_d of heat-shocked WPI was not substantially different from heat-shocked and MTGase-treated WPI at pH 2.5. Both the heat-shocked and the heat-shocked-MTGase-treated WPI exhibited U-shaped pH-solubility profiles with minimum solubility at pH 4.0–5.0. However, the extent of precipitation of MTGase-treated WPI samples at pH 4.0–5.0 was much greater than all heat-shocked and native WPI samples. The study revealed that while MTGase cross-linking significantly enhanced the thermal stability of β-lactoglobulin in heat-shocked WPI, it caused pronounced precipitation at pH 4.0–5.0 via decreasing the hydrophilic/hydrophobic ratio of the water-accessible protein surface.     

Hydrolyzed wheat gluten suppresses transglutaminase-mediated gelation but improves emulsification of pork myofibrillar protein

The influence of 15-h chymotrypsin-hydrolyzed wheat gluten (GH) on microbial transglutaminase (MTGase)-mediated interaction, gelation and emulsification of pork myofibrillar protein isolate (MPI) was investigated at two ionic strengths (0 M and 0.6 M NaCl) and pH 6.5. MTGase treatments in 0 M NaCl solution decreased the size of myosin heavy chain through deamidation, but this was inhibited by GH or in 0.6 M NaCl where myosin polymerization dominated. Stabilization of MPI (thermal transitions) by the MTGase treatment was also diminished (P < 0.05) by the presence of GH at both ionic strengths. These GH-induced MPI physicochemical changes greatly weakened the ability of MTGase to promote MPI thermal gelation (gel storage modulus, P < 0.05), especially at 0.6 M NaCl, which was shown to result from reduced protein aggregation. However, GH improved (P < 0.05) emulsifying properties of MPI, regardless of MTGase treatment.     

Partial purification and characterization of trypsin-like proteinases in Indian anchovy (Stolephorus spp.)

Four fractions (P111, P21, P31, and P4) of proteinases were obtained from various purification steps including heat treatment (60 °C, 10 min), 30–60% ammonium sulfate precipitation, anion exchange, hydrophobic interaction, and gel filtration chromatography. Optimal temperature and pH of all fractions were 50–60 °C and 8.5, respectively. All partially purified proteinases preferably hydrolyzed substrates containing Arg at the P₁ position. All proteinases were inhibited by soybean trypsin inhibitor, leupeptin, and N-tosyl-l-lysine chloromethyl ketone. Partially purified proteinases were stable at 35 °C up to 12 h. However, their activity decreased about 40% when incubated at the optimal temperature (50–55 °C) for 2 h. Only P111 was stable at its optimal temperature (60 °C) up to 12 h. Molecular weight (MW) of P111, P21, and P31 was estimated to be 27, 33, 37, 43, 48, 55, 60, and 65 kDa, while MW of P4 was 39 kDa based on activity staining. All partially purified proteinases hydrolyzed washed anchovy mince at 4.0 M NaCl, pH 8.5, at 35 °C and at their optimal temperatures (50–60 °C).     

Functional properties of protein fractions obtained from commercial yellow field pea (Pisum sativum L.) seed protein isolate

Commercial pea protein isolate was separated into water-soluble (WS), salt-soluble (SS), alkaline-soluble (AS) and ethanol-soluble (ES) fractions. AS fraction was the most abundant, constituting about 87% of the proteins in PPI followed by WS, SS and ES fractions in decreasing order. ES fraction consistently formed emulsions with a narrow range of smaller oil droplet sizes (0.6–19 μm) at pH 4.0, 7.0 or 9.0 compared to a wider range of sizes for emulsions stabilised by WS, SS and AS fractions. Emulsions formed with ES fraction were also the most stable (p < 0.05) over the 3 h test period at all the pH values used in this work. The WS fraction had significantly highest (p < 0.05) protein solubility and foaming capacity at all the pH values when compared to solubility of PPI, SS, and ES. Except for AS and ES fractions, foaming capacities of the protein fractions were higher at pH 9.0 than at pH 4.0 or 7.0.     

Pacific whiting (Merluccius productus) underutilization in the Gulf of California: Muscle autolytic activity characterization

In México’s Northwest coast, Pacific whiting (Merluccius productus) is considered an under-utilized species because of its high tendency to become parasitized, thus promoting a high proteolytic activity present in muscle tissue. Sample fish for study were separated in lots by degree of parasitism in “apparent” parasitized (APP) and advanced parasitized (ADP). Thus, pH and temperature conditions of mayor endogenous proteolytic activity in muscle were determined. Maximum activity was detected at 50 °C (pH 3.5–4.0) and at 60 °C (pH 6.75–7.0). Parasitism degree had a significant effect in enzyme activity at acidic pH (p < 0.05) being higher in APP at low temperature (30 °C). Higher temperatures (40–50 °C) favored (p < 0.05) activity in ADP muscle (same pH range) with the highest (p < 0.05) observed at 50 °C at pH 3.5. No much difference was observed at pH 7.0–8.0. Results suggest that pH around physiological conditions at 60 °C could be used as an advantage in fish protein hydrolysate production or as processing aid where a protein hydrolysis is required.     

Effect of chitosan on the stability and properties of modified lecithin stabilized oil-in-water monodisperse emulsion prepared by microchannel emulsification

The effect of chitosan (CHI) on the stability of monodisperse modified lecithin (ML) stabilized soybean oil-in-water (O/W) emulsion was investigated. Monodisperse emulsion droplets with particle size of 24.4 ± 0.7 μm and coefficient of variation below 12% were prepared by microchannel (MC) emulsification using a hydrophilic asymmetric straight-through MC silicon 24 × 24 mm microchip consisting of 23,348 microchannels. The stability of the ML stabilized monodisperse emulsion droplets was investigated as a function of CHI addition at various concentration, pH, ionic strength, thermal treatment and freezing-thawing treatment by means of particle size and ζ-potential measurements as well as microscopic observation. The monodisperse O/W emulsions were diluted with CHI solution at various concentrations to a final droplet concentration of 1 wt% soybean oil, 0.25 wt% ML and 0–0.5 wt% CHI at pH 3. Pronounced droplet aggregation was observed when CHI was present at a concentration range of between 0.01 and 0.04 wt%. Above this concentration range, flocculations were less extensive, indicating some restabilization. ML stabilized emulsions were stable at a wide range of NaCl concentrations (0–1000 mM) and pH (3–8). On the contrary, in the presence of CHI, aggregation of the emulsion droplets was observed when NaCl concentration was above 200 mM and when the pH started to approach the pKa of CHI (i.e. ∼6.2–7.0). Emulsions containing CHI were found to have better stability at high temperature (>70 °C) in comparison to the emulsion stabilized only by ML. With sucrose/sorbitol as cryoprotectant aids, emulsions with the addition of CHI were found to be more resistant to droplet coalescence as compared to those without CHI after freezing at −20 °C for 22 h and thawing at 30 °C for 2 h. The use of CHI may potentially destabilize ML-stabilized O/W emulsions but its stability can be enhanced by selectively choosing the appropriate CHI concentrations and conditions of preparation.     

Comparison of Atlantic menhaden gels from surimi processed by acid or alkaline solubilization

Heat-induced gelling abilities of surimis prepared by pH shifting (isoelectric precipitation following acid (AC) or alkaline (AL) solubilization) were compared to that of conventionally washed (CW) surimi. Greater endogenous transglutaminase activity (evidenced as enhanced strength of cooked gels subjected to 30–40 °C preincubation) was measured for CW and AL surimi than for AC surimi (all at pH 7). Upon addition of microbial transglutaminase (MTGase), increased crosslinking of myosin heavy chain and gel strengthening during 30–40 °C preincubation were apparent for all three types of surimi, most markedly in CW and AL surimi. Salt addition improved CW gels most, but seemed to adversely affect MTGase activity in AC and AL surimi. AC and AL surimi gels were of lower whiteness than were CW surimi gels.     

Effects of combining microbial transglutaminase and high pressure processing treatments on the mechanical properties of heat-induced gels prepared from arrowtooth flounder (Atheresthes stomias)

The objective of this study was to evaluate the effect of setting conditions (25 °C for 2 h or 40 °C for 30 min) and combining of microbial transglutaminase (MTGase) and high pressure processing (HPP) on the mechanical properties of heat induced gels obtained from paste from arrowtooth flounder (Atheresthes stomias). Treatments included fish paste control without added MTGase, fish paste incubated with MTGase but not pressurized (MTGase + cooking), fish paste incubated with MTGase and pressurized at 600 MPa for 5 min (MTGase + HPP + cooking) and fish paste pressurized at 600 MPa for 5 min and incubated with MTGase (HPP + MTGase + cooking). The controls and the treated samples were then subjected to one of two thermal treatments: 90 °C for 15 min or 60 °C for 30 min before cooking at 90 °C for 15 min. Samples of fish paste heated at 60 °C before cooking could not be used to prepare gels for texture profile analysis (TPA). TPA showed that pressurization improved the mechanical properties of gels made from paste treated with MTGase and set at 25 °C. The opposite was observed for samples set at 40 °C. Setting at 40 °C appeared to induce proteolytic degradation of myofibrillar proteins.     

Endogenous proteinases in true sardine (Sardinops melanostictus)

Characterisation of the autolytic profile of true sardine (Sardinops melanostictus) indicated the involvement of heat-activated proteinases, active at both acidic and alkaline pH values. Autolytic activity decreased with increasing NaCl concentration (0–30%). When crude proteolytic activity in true sardine was studied, two activity peaks were observed, at pH 3.5 and 9.0. Crude proteinase extracts exhibited the highest activity at 55 °C and 60 °C when assayed at pH 3.5 and 9.0, respectively. The pH 3.5 peak activity was effectively inhibited by pepstatin A, while the pH 9.0 peak activity was mostly inhibited by soybean trypsin inhibitor, PMSF and TLCK, suggesting that the various proteinases were present in true sardine. The enzymes were stable for up to 8 h at 55 °C. The activities were also stable at a pH range of 2.0–4.0 and still retained high activity toward hemoglobin after incubation at pH 3.5 for 8 h. Activities of the crude extract continuously decreased as NaCl concentration increased. ATP showed no effect on enzyme activity, while CaCl₂ and MgCl₂ activated the proteinase activity. The results implied that pepsin is a predominant enzyme responsible for autolysis in true sardine during fish sauce fermentation.     

Microbial transglutaminase-induced structural and rheological changes of cationic and anionic myofibrillar proteins

This study investigated the effects of microbial transglutaminase (TG) on structural changes in porcine myofibrillar protein (MP) under varying pH (2.0–6.0) and two ionic strength conditions (0.1 M versus 0.6 M NaCl). Lowering the pH below the isoelectric point (pI) of myosin induced protein unfolding as revealed by surface hydrophobicity and differential scanning calorimetry. Although the MP solubility at the low ionic strength (0.1 M NaCl) was maximal at pH 3.0, both SDS-PAGE profiles and dynamic rheology indicated TG could not cross-link MP under this condition. Based on the carboxyl group content, the TG-catalyzed deamidation was dominant at a pH lower than the pI of myosin (pH 5.0) while cross-linking occurred at higher pH. Moreover, deamidation had no effect on rheological properties of MP. The results indicate that the TG reaction was governed by the pH of substrate protein, and the reaction intensity was related to the solubility of protein.     

Functionality of phosphorylated vicilin exposed to chemical and physical agents

The effects of phosphorylation with sodium trimetaphosphate (STMP) on functional and physicochemical properties of pea vicilin, as probed by high hydrostatic pressure and chemical denaturation were evaluated. The isoelectric point of unmodified and phosphorylated vicilin decreased in the presence of 0.5 M NaCl, resulting in a decrease of the solubility at pH 1.0. The gelation capacity of unmodified vicilin in the presence of NaCl decreased approximately 80% when compared with unmodified vicilin without NaCl. Increasing pressure from 0.1 MPa (atmospheric pressure) to 240 MPa significantly decreased the solubility of vicilin phosphorylated with 4% STMP at pH 1.0 and 4.0 by about 30%. Pressure had no effect on solubility of native vicilin. Pressure treatment at 240 MPa improved the gelation capacity of vicilin phosphorylated with 1% STMP. Glycerol decreased the gelation capacity of vicilin and its solubility in the acidic pH range.     

Isolation and characterization of acid and pepsin-solubilized collagens from the skin of balloon fish (Diodon holocanthus)

Acid-solubilized collagen (ASC) and pepsin-solubilized collagen (PSC) were successfully extracted from the skin of balloon fish (Diodon holocanthus) with yields of 4% and 19.5% respectively (based on dry weight). According to the electrophoretic patterns, both the ASC and PSC consisted of two different α chains (α1 and α2), were characterized to be type I, and contained imino acid of 179 and 175 residues/1000 residues, respectively. The PSC had a lower content of high-molecular weight cross-links than the ASC. The ultraviolet (UV) absorption spectrum of collagens showed that the distinct absorption was between 210 and 230 nm. A maximum solubility in 0.5 M acetic acid was observed at pH 1–5, and a sharp decrease in solubility above 4% (w/v) in both the ASC and PSC was observed in the presence of NaCl. The denaturation temperatures (T_d) of the ASC and PSC measured by viscometry were 29.01 °C and 30.01 °C, respectively. The maximum temperatures (T_max) of the ASC and PSC were 29.64 °C and 30.30 °C, respectively.     

Synergistic action of transglutaminase and high pressure on chicken meat and egg gels in absence of phosphates

The effects of simultaneous application of microbial transglutaminase (MTGase) and high pressure (HP) (500, 700 and 900 MPa/40 °C/30 min), only pressure under the same conditions or heat (75 °C/30 min) were investigated on chicken batters with the addition of egg components and without phosphates. MTGase gels (700 and 900 MPa) showed marked increases in textural parameters compared to gels without enzyme (NE) or those obtained by heat. The addition of enzyme did not show differences between gels obtained at 700–900 MPa; however, gels obtained at 500 MPa were darker and more reddish than those obtained by heat. MTGase gels were more homogeneous and compact. Thermal analysis revealed that pressure levels above 700 MPa caused as much denaturing as did heat. The microstructure and texture of MTGase gels suggest that a higher amount and heterogeneity of crosslinks was produced when meat and egg proteins were treated in the presence of MTGase under specific conditions of pressure.     

Protein Oxidation at Different Salt Concentrations Affects the Cross‐Linking and Gelation of Pork Myofibrillar Protein Catalyzed by Microbial Transglutaminase

In a fabricated then restructured meat product, protein gelation plays an essential role in producing desirable binding and fat‐immobilization properties. In the present study, myofibrillar protein (MFP) suspended in 0.15, 0.45, and 0.6 M NaCl was subjected to hydroxyl radical stress for 2 or 24 h and then treated with microbial transglutaminase (MTGase) in 0.6 M NaCl (E : S = 1 : 20) at 4 and 15 °C for 2 h. Protein cross‐linking and dynamic rheological tests were performed to assess the efficacy of MTGase for mediating the gelation of oxidized MFP. MTGase treatments affected more remarkable polymerization of myosin in oxidized MFP than in nonoxidized, especially for samples oxidized at 0.6 M NaCl. Notably, the extent of MTGase‐induced myosin cross‐linking at 15 °C in oxidized MFP improved up to 46.8%, compared to 31.6% in nonoxidized MFP. MTGase treatment at 4 °C for MFP oxidized in 0.6 M NaCl, but not MFP oxidized in 0.15 M NaCl, produced stronger gels than nonoxidized MFP (P < 0.05). The final (75 °C) storage modulus (G′) of oxidized MFP gels was significantly greater than that of nonoxidized, although the G′ of the transient peak (～44.5 °C) showed the opposite trend. Overall, oxidation at high salt concentrations significantly improved MTGase‐mediated myosin cross‐linking and MFP gelation. This might be because under this condition, MTGase had an increased accessibility to glutamine and lysine residues to effectively initiate protein–protein interactions and gel network formation.     

Effect of extraction conditions on the yield and chemical properties of pectin from cocoa husks

Different extraction conditions were applied to investigate the effect of temperature, extraction time and substrate–extractant ratio on pectin extraction from cocoa husks. Pectin was extracted from cocoa husks using water, citric acid at pH 2.5 or 4.0, or hydrochloric acid at pH 2.5 or 4.0. Temperature, extraction time and substrate–extractant ratio affected the yields, uronic acid contents, degrees of methylation (DM) and degrees of acetylation (DA) of the extracted pectins using the five extractants differently. The yields and uronic acid contents of the extracted pectins ranged from 3.38–7.62% to 31.19–65.20%, respectively. The DM and DA of the extracted pectins ranged from 7.17–57.86% to 1.01–3.48%, respectively. The highest yield of pectin (7.62%) was obtained using citric acid at pH 2.5 [1:25 (w/v)] at 95 °C for 3.0 h. The highest uronic acid content (65.20%) in the pectin was obtained using water [1:25 (w/v)] at 95 °C for 3.0 h.     

Dynamic oscillatory rheological measurement and thermal properties of pea protein extracted by salt method: Effect of pH and NaCl

The effects of two important factors, pH (3.0-10.0) and NaCl (0-2.0 M), on pea protein gelation properties were studied using dynamic oscillatory rheometer and differential scanning calorimeter (DSC). The strongest gel stiffness was achieved at 0.3 M NaCl; higher or lower salt concentrations lead to weakening of the gel. The gelation temperature was also influenced by ionic strength; salt had a stabilization effect which inhibited pea protein denaturation at higher salt concentrations resulting in higher gelling points (p < 0.05). At a NaCl concentration 2.0 M, pea protein gelation was completely suppressed at temperatures ?100 °C. The pH also played an important role in gel formation by pea protein isolates since acid and base cause partial or even total protein denaturation. In this paper the maximum gel stiffness occurred at pH 4.0 in 0.3 M NaCl; higher or lower pH values resulted in reduced gel stiffness (p < 0.05). pH also altered the denaturation temperature of the pea protein; higher pH values resulted in higher denaturation temperatures and higher enthalpies of denaturation (p < 0.05). At pH 3 pea proteins seem like completely denatured by acid as the DSC curve showed a straight line. The gelation temperature (gelling point) peaked at pH ∼6.0 (89.1 °C). Careful adjustment of pH and NaCl concentration would enable the food industry to effectively utilize the salt-extracted pea protein isolate as a gelling agent.     

Extraction and characterisation of pectin methylesterase from black carrot (Daucus carota L.)

This study was carried out to determine some of the biochemical properties of pectin methylesterase (PME) from black carrot. The enzyme showed very high activity in a broad pH range of 6.5–8.5, with the optimum pH occurring at 7.5. The optimum temperature for maximal PME activity was found to be 55 °C. NaCl enhanced PME activity, particularly at 0.2 M. K_m and V_max values for black carrot PME using apple pectin as substrate were found to be 2.14 mg/ml (r² = 0.988) and 3.75 units/ml, respectively. The enzyme was stable between the temperatures of 30–50 °C/5 min whereas it lost nearly all of its activity at 70 °C/5 min. E_a and Z values were found to be 196.8 kJmol⁻¹ (r² = 0.996) and 2.16 °C (r² = 0.995), respectively.     

Solubility of sodium and potassium iodates in saturated salt solutions

The determination of the solubilities of KIO₃ and NaIO₃ in saturated NaCl solution at 0, 10, 20 °C were carried out in two ternary systems KIO₃–NaCl–H₂O and NaIO₃–NaCl–H₂O. Saturated KIO₃ and NaIO₃ solutions were prepared and NaCl was added to saturation. Iodate concentrations were measured using standardized sodium thiosulphate. The solubilities of KIO₃ and NaIO₃ are temperature sensitive over the range 0 °C to 20 °C (unlike NaCl). Iodate solubility was much lower in saturated NaCl than that of deionised water. In saturated NaCl at 10 °C, for example, NaIO₃ was 19% and KIO₃ was 15% the solubility of that found in deionised water at the same temperature.     

Gelling properties of white shrimp (Penaeus vannamei) meat as influenced by setting condition and microbial transglutaminase

The properties of white shrimp (Penaeus vannamei) gel added with different levels of microbial transglutaminase (MTGase) and subjected to setting at 25 °C for 2 h or 40 °C for 30 min, prior to heating at 90 °C for 20 min were studied. Breaking force of gels with and without setting increased with increasing MTGase amount added (P<0.05). However, no changes in deformation in all samples were noticeable (P>0.05). Directly heated gels showed the lower breaking force than those with prior setting at all MTGase levels added (P<0.05). Generally, gels prepared by setting at 25 °C exhibited the greater breaking force than those set at 40 °C, possibly associated with the appropriate protein structure for cross-linking at 25 °C and greater degradation at 40 °C as evidenced by a greater trichloroacetic acid soluble peptide content (P<0.05). Sodium dodecyl sulfate polyacrylamide gel electrophoretic study revealed that myosin heavy chain (MHC) underwent polymerization to a higher extent in the presence of MTGase, but the strengthening effect on gel was dependent on setting temperature. Regardless of setting condition, microstructure of gel added with MTGase was finer with a smaller void, compared with those of gel without MTGase. Therefore, setting temperature played an essential role in gel property of white shrimp meat added with MTGase.