Influence of muscle type on rheological properties of porcine myofibrillar protein during heat-induced gelation

The gelation characteristics of myofibrillar proteins are indicative of meat product texture. Defining the performance of myofibrillar proteins during gelation is beneficial in maintaining quality and developing processed meat products and processes. This study investigates the impact of pH on viscoelastic properties of porcine myofibrillar proteins prepared from different muscles (semimembranosus (SM), longissimus dorsi (LD) and psoas major (PM)) during heat-induced gelation. Dynamic rheological properties were measured while heating at 1 °C/min from 20 to 85 °C, followed by a holding phase at 85 °C for 3 min and a cooling phase from 85 to 5 °C at a rate of 5 °C/min. Storage modulus (G′, the elastic response of the gelling material) increased as gel formation occurred, but decreased after reaching the temperature of myosin denaturation (52 °C) until approximately 60 °C when the gel strength increased again. This resulted in a peak and depression in the thermogram. Following 60 °C, the treatments maintained observed trends in gel strength, showing SM myofibrils produced the strongest gels. Myofibrillar protein from SM and PM formed stronger gels at pH 6.0 than at pH 6.5. Differences may be attributed to subtle variations in their protein profile related to muscle type or postmortem metabolism. Significant correlations were determined between G′ at 57, 72, 85 and 5 °C, indicating that changes affecting gel strength took effect prior to 57 °C. Muscle type was found to influence water-holding capacity to a greater degree than pH.     

Lessening of high-pressure-induced changes in Atlantic salmon muscle by the combined use of a fish gelatin–lignin film

Salmon muscle is considerably affected by cooking with the resulting loss of its appealing red colour. The combined use of high pressure with fish gelatin–lignin film is proposed as an alternative to the more aggressive thermal processing procedures, with the aim of improving the appearance and overall quality of salmon fillets in ready-to-eat or semi-prepared dishes. The effects of high pressure processing (300 MPa, 10 min, 5 °C or 40 °C) and conventional heating (90 °C, 10 min) were evaluated in terms of colour changes, protein denaturation, as well as protein and lipid oxidation, by comparison with raw muscle. The stability of the processed products was assessed by monitoring changes in microbial growth and total volatile basic nitrogen and thiobarbituric acid reactive substances during 23 days of chilled storage. Fourier transform infrared spectroscopy (FTIR), apparent viscosity and dynamic oscillatory studies revealed notable differences in the overall degree and nature of protein aggregation between high pressure and heating treatments, especially when performed at 5 °C instead of 40 °C. SDS–PAGE of the protein fraction solubilised in 0.8 M NaCl showed MHC and α-actinin to be the main myofibrillar proteins denatured by high pressure processing at 40 °C, while actin was more denatured when pressurised at 5 °C. The film attenuated colour changes associated with high pressure treatment, especially at 5 °C, where redness was more preserved without jeopardising the appearance of a ready-to-eat product. High pressure processing at 5 °C in combination with gelatin–lignin film was found to improve protein quality of salmon fillets. The film reduced the levels of carbonyl groups formed immediately after processing, and prevented lipid oxidation from taking place at advanced stages of chilled storage. However, the effect on microbial growth was negligible, since total counts were similar for muscle with or without the film.     

Tenderization effect of cold-adapted collagenolytic protease MCP-01 on beef meat at low temperature and its mechanism

The enzymes currently used to increase meat tenderness are all mesophilic or thermophilic proteases. This study provides insight into the tenderization effect and the mechanism of a cold-adapted collagenolytic enzyme MCP-01 on beef meat at low temperatures. MCP-01 (10 U of caseinolytic activity) reduced the meat shear force by 23% and increased the relative myofibrillar fragmentation index of the meat by 91.7% at 4 °C, and it also kept the fresh colour and moisture of the meat. Compared to the commercially used tenderizers papain and bromelain, MCP-01 showed a unique tenderization mechanism. MCP-01 had a strong selectivity for degrading collagen at 4 °C, showed a distinct digestion pattern on the myofibrillar proteins, and had a different disruption pattern on the muscle fibres under scanning electron micrograph. These results suggest that the cold-adapted collagenolytic protease MCP-01 may be promising for use as a meat tenderizer at low and moderate temperatures.     

Mathematical modeling of the heat transfer process and protein denaturation during the thermal treatment of Patagonian marine crabs

Southern Ocean swimming crab Ovalipes trimaculatus and the Patagonian stone crab Platyxanthus patagonicus are fishing resources with commercial value. Thermal treatment of crabs is necessary to denature muscle proteins, facilitating meat detachment from the crab shell (picking procedure). The proximal composition, protein patterns of crab muscle, thermophysical properties and heat transfer coefficients were determined. Heat transfer during thermal processing of body (i.e., cephalothorax) and claws of both crab species was simulated using a finite element computational code; the simulations were experimentally validated. Color changes in crab muscle during the heating process were measured. Thermal denaturation kinetics of myofibrillar proteins was determined using Differential Scanning Calorimetry (DSC) in small samples previously heated in water under controlled conditions. DSC thermograms of raw crab muscle showed two peaks at 49.0 ± 0.4 and 77.5 ± 0.6 °C corresponding to myosin and actin respectively. Activation energies for the denaturation of myosin (145.70 kJ/mol) and actin (156.42 kJ/mol) were calculated from Arrhenius equation. The degree of denaturation achieved by the myofibrillar proteins at the coldest point of the muscle in body and claws during the heating process was established by considering the protein denaturation kinetics determined by DSC, the activation energies and the heat penetration curves. Adequate conditions for the detachment of meat from the crab exoskeleton were established. The obtained results may help in determining the optimal heating times during the industrialization of these crustaceans.     

Kinetics of protein physicochemical changes induced by heating in meat using mimetic models: (2) Effects of fibre type,peroxides and antioxidants

Heating-induced changes in meat proteins were investigated using models made of aqueous suspensions of myofibrils according to muscle fibre types and cellular compounds (oxidants and antioxidants). These changes were evaluated by measurements of carbonyl groups and protein surface hydrophobicity. Model results were compared to trial results obtained on pork meat (M. Longissimus dorsi) heated under the same conditions (45 and 75 °C, from 5 to 120 min). Myofibrillar proteins from α-white fibres were more sensitive to oxidation and thermal denaturation than those from β-red fibres. At 45 °C, there were negligible differences due to peroxide or antioxidant types. At 75 °C, organic peroxides (ROOH) were less oxidative than hydrogen peroxide (H₂O₂), and antioxidant enzymes were less efficient than vitamin E and carnosine at protecting proteins against oxidation. Protein oxidation observed in meat is lower than in the mimetic models and the increase in hydrophobicity remained limited in meat.     

Myosin heavy chain degradation during post mortem storage of Atlantic cod (Gadus morhua L.)

Post mortem proteolytic degradation of fish fillets leads to textural changes like muscle softening and gaping. In this study proteolytic degradation of myosin heavy chain (MHC) was monitored during storage of muscle and of isolated myofibrils at different temperatures and pH-values by the use of MHC-specific antibodies. The ability of cathepsin D to associate to myofibrillar proteins was also studied. Muscle stored at 6 °C and isolated myofibrils stored at 0 °C, 6 °C and 20 °C were degraded at pH 6.3 or lower. Cathepsin D could be found associated with extensively washed myofibrils. Inhibition of cathepsin D during storage affected the observed MHC-degradation at pH 5.5, but not at pH 6.3. This indicates that cathepsin D to a less extend than formerly believed, is responsible post mortem degradation of MHC.     

High pressure induced changes in beef muscle proteome: Correlation with quality parameters

The relationship between pressure induced changes on individual proteins and selected quality parameters in bovine longissimus thoracis et lumborum (LTL) muscle was studied. Pressures ranging from 200 to 600 MPa at 20 °C were used. High pressure processing (HPP) at pressures above 200 MPa induced strong modifications of protein solubility, meat colour and water holding capacity (WHC). The protein profiles of non-treated and pressure treated meat were observed using two dimensional electrophoresis. Proteins showing significant differences in abundance among treatments were identified by mass spectrometry. Pressure levels above 200 MPa strongly modified bovine LTL proteome with main effects being insolubilisation of sarcoplasmic proteins and solubilisation of myofibrillar proteins. Sarcoplasmic proteins were more susceptible to HPP effects than myofibrillar. Individual protein changes were significantly correlated with protein solubility, L*, b* and WHC, providing further insights into the mechanistic processes underlying HPP influence on quality and providing the basis for the future development of protein markers to assess the quality of processed meats.     

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.     

Effect of pH at heating on the acid-induced aggregation of casein micelles in reconstituted skim milk

Reconstituted skim milk samples at pH between 6.5 and 7.1 (heating pH) were heated at 80°C, 90°C or 100°C for 30 min (heating temperature). The particle size of the casein micelles was measured at pH 4.75-7.1 (measurement pH) and at temperatures of 10°C, 20°C and 30°C (measurement temperature) using photon correlation spectroscopy. The particle size of the casein micelles, at a measurement pH of 6.7 and a measurement temperature of 20°C, was dependent on the heating pH and heating temperature to which the milk was subjected. The casein micelle size in unheated milk was about 215 nm. At a heating pH of 6.5, the casein micelle size increased by about 15, 30 and 40 nm when the milk was heated at 80°C, 90°C or 100°C, respectively. As the heating pH of the milk was increased, the size of the casein micelles decreased so that, at pH 7.1, the casein micelles were ∼20 nm smaller than those from unheated milk. Larger effects were observed as the heating temperature was increased from 80°C to 100°C. The size differences as a consequence of the heating pH were maintained at all measurement temperatures and at all measurement pH down to the pH at which aggregation of the micelles was observed. For all samples, size measurements at 10°C showed no aggregation at all measurement pH. Aggregation occurred at progressively higher pH as the measurement temperature was increased. Aggregation also occurred at a progressively higher measurement pH as the heating pH was increased. The particle size changes on heating and the aggregation on subsequent acidification may be related to the pH dependence of the association of whey proteins with, and the dissociation of κ-casein from the casein micelles as milk is heated.     

Kinetics of protein physicochemical changes induced by heating in meat using mimetic models: (1) Relative effects of heat and oxidants

Optimizing the nutritional quality of cooked meat needs a better understanding of the mechanisms responsible for protein changes induced by heating. The relative contributions of chemical and thermal effects on protein physicochemical changes were studied using meat models. Two models were tested: a basic model made of an aqueous suspension of myofibrillar proteins, and a complex model, in which oxidants were added in physiological concentrations. Various heating time–temperature combinations were applied to both models in the ranges 45–90 °C and 5–120 min. Protein oxidation was evaluated by carbonyl and free thiol contents. Conformational changes of proteins were assessed by measurements of surface hydrophobicity and aggregation. Carbonyl formation was weakly affected by the thermal process alone but exacerbated by oxidants. A synergistic effect of oxidants and heat treatments on protein oxidation was noted. Changes in protein hydrophobicity and aggregation were dominated by the thermal process.     

The effect of active caspase-3 on degradation of chicken myofibrillar proteins and structure of myofibrils

The objective of this study was to investigate the potential contribution of caspase-3 to meat postmortem tenderisation by examining the role of caspase-3 in the degradation of myofibrillar proteins and disruption of myofibril structure in vitro. Myofibrillar protein prepared from chicken muscle was incubated with EDTA or EDTA plus caspase-3 at 25 °C for 16 h and used for detecting muscle protein degradation and ultrastructure of myofibril. Results revealed that caspase-3 reproduced the degradation patterns of titin, nebulin and α-actinin during postmortem storage of meat, but caused little proteolysis of desmin and no appearance of 28–30 kDa peptides. Meanwhile, caspase-3 also induced the weakening in the I band adjacent to Z-lines, which occurred during meat postmortem ageing. Therefore, caspase-3 could account only for a part of the myofibrillar protein degradation observed in naturally aged meat and is likely involved in postmortem tenderisation of meat together with other endogenous proteases.     

Microemulsions as nanoreactors to produce whey protein nanoparticles with enhanced heat stability by thermal pretreatment

Native whey proteins (NWPs) may form gels or aggregates after thermal processing. The goal of this work was to improve heat stability of NWPs by incorporating protein solutions in nanoscalar micelles of water/oil microemulsions to form whey protein nanoparticles (WPNs) by thermal pretreatment at 90 °C for 20 min. The produced WPNs smaller than 100 nm corresponded to a transparent dispersion. The WPNs produced at NWP solution pH of 6.8 had a better heat stability than those produced at pH 3.5. The salt concentration (0–400 mM NaCl) in NWP solutions did not significantly change the size of corresponding WPNs. Compared to NWPs, the 5% (w/v) dispersion of WPNs at pH 6.8, 100 mM NaCl did not form a gel after heating at 80 °C for 20 min. The improved heat stability and reduced turbidity of WPNs may enable novel applications of whey proteins in beverages.     

Salt effect on heat-induced physical and chemical changes of salmon fillet (O. gorbuscha)

This research studied the effect of salt on kinetics for quality changes in pink salmon fillet, during commercial sterilisation. Sample cuts from salmon fillets were placed in sealed aluminum containers and heated at 121.1 °C for 10, 30 and 60 min. Samples with 1.5% (w/w) salt addition were compared with those without added salt. Salt addition reduced cook loss, area shrinkage and shear force of the heated fillet and resulted in a slightly darker colour. Effect of salt addition in thiamin loss, degree of lipid oxidation and fatty acid profile was not significant. Peroxide (PV) and thiobarbituric acid (TBA) values slightly increased within the first 10 min of heating, followed by a significant reduction as heating progressed. No measurable loss of polyunsaturated fatty acids (PUFA) was observed. Thermally processed shelf-stable salmon investigated should be a valuable source of ω-3 PUFA, with EPA values ranging from 52 to 71 mg/100 g product and DHA ranging from 258 to 340 mg/100 g of product.     

Heating of milk alters the binding of curcumin to casein micelles. A fluorescence spectroscopy study

Curcumin, a polyphenolic compound present in turmeric, is a hydrophobic molecule that has been shown to bind to casein micelles. The present work tested the hypothesis that surface changes in the casein micelles caused by heat-induced interactions with the whey proteins would affect the binding of curcumin. Binding was quantified by direct and tryptophan quenching fluorescence spectroscopy. Curcumin binds to the hydrophobic moieties of the casein proteins, with a 10 nm blue shift in its fluorescence emission peak, and causes quenching of the intrinsic fluorescence spectra of the proteins. The fluorescence intensity of curcumin increased after heating of milk at 80 °C for 10 min; a similar trend in the binding constants was also observed with casein micelles separated from the soluble proteins by centrifugation. There was an increase in the non-specific interactions with heating milk at 80 °C for 10 min, both in milk as well as in casein micelles separated from the serum proteins. The increased capacity of milk proteins to bind curcumin after heat treatment can be attributed to whey protein denaturation, as whey proteins bind to the surface of casein micelles with heating.     

Study on formation of acrylamide in asparagine–sugar microwave heating systems using UPLC-MS/MS analytical method

Microwave heating can be regarded as a possible way to produce a considerable amount of acrylamide. The present study investigated the formation of acrylamide in asparagine–glucose, asparagine–fructose and asparagine–sucrose microwave heating systems by the response surface methodology (RSM) and the orthogonal array methodology (OAM). The acrylamide content was rapidly quantified by a validated ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method. Results of RSM study indicated that in the asparagine–glucose system, the acrylamide content increased in the combined condition of high temperature accompanying with short heating time (>190 °C, <20 min) or low temperature accompanying with long heating time (<180 °C, >30 min). In the asparagine–fructose system, the similar conclusion was made in the combined condition of high temperature accompanying with short heating time (>175 °C, <20 min) or low temperature accompanying with long heating time (<170 °C, >25 min). In the asparagine–sucrose system, the amount of acrylamide enhanced with the increase of both heating temperature and heating time. The fitted mathematic models were successfully applied to the quantification of acrylamide formation when the heating temperature and heating time fell into the ranges of 120–240 °C and 5–35 min simultaneously. OAM study showed that acrylamide is readily formed via heating binary precursors 5 min at 180 °C in the asparagine–glucose and asparagine–fructose systems. However, acrylamide is readily generated when the binary precursors are heated 15 min at 180 °C in the asparagine–sucrose system.     

A proposed mechanism of tenderising post-rigor beef using high pressure–heat treatment

Tenderness of beef M. Sternomandibularis was tough when cooked from both raw, and when previously heated (60 °C, 20 min), whereas a significant improvement in tenderness was achieved when pressure–heat (P–H) treated muscle (200 MPa, 60 °C, 20 min) was cooked. In order to determine the mechanism for this improvement, connective tissue, myofibrillar and sarcoplasmic proteins, were separated into three fractions and studied with regard to their solubilisation, denaturation and aggregation, degradation and strengthening of protein structures for the three treatments (raw, heated and H–P treated). Measurements included DSC, SDS–PAGE, surface hydrophobicity, and the appearance, length and width of myofibres (light microscopy). For the connective tissue fraction, heat solubility was determined.     

Effect of thermal process on connective tissue from jumbo squid (Dosidicus gigas) mantle

The effect of two thermal treatments (fast freezing at −40 °C and vapor cooking at 100 °C) on connective tissue extract (CTE) from jumbo squid (Dosidicus gigas) was investigated. Samples of CTE frozen at −40 °C were taken at 0, 3, 5 and 12 min. Also CTE was cooked at 100 °C and samples were taken at 0, 1, 2.5 and 5 min. Light microscopic observations of CTE after 12 min of freezing showed rupture of fibres. The CTE fibres showed agglutination during cooking time. The CTE insoluble fraction increased with freezing and cooking time. Maximum zeta potential value of untreated CTE was detected at pH 5.0 at +30 mV, meanwhile in the frozen CTE it was detected at pH 7.0 at +30 mV and two peaks (at pH 5.5 and 9.0) were observed at +20 mV in the cooked CTE. One endothermic peak was found at 105.9 °C in the untreated CTE, while in the frozen and cooked CTE the endothermic peaks were found at lower temperatures and enthalpies. Electrophoresis analysis of untreated CTE showed three bands. In the frozen CTE two bands appeared above 200 kDa, and in the cooked CTE, a 45 kDa band disappeared. These results suggest that during freezing and cooking processes there were modifications to molecular bonds that hold the integrity of the structure of the connective tissue of the jumbo squid mantle.     

Analysis and formation of trans fatty acids in hydrogenated soybean oil during heating

Hydrogenated oil has been widely used for production of shortenings or margarine, however, the presence of trans fatty acids may be detrimental to human health. The objectives of this study were to develop an improved method for analysis of trans fatty acids and evaluate their formation in both unhydrogenated and hydrogenated soybean oil during heating at 160, 180 and 200 °C for varied length of time. Results showed that among the four columns tested, an Agilent HP-88 column (100 × 0.25 mm I.D., 0.2-μm film thickness) could resolve eight trans fatty acids and nine cis fatty acids simultaneously within 31 min with injector temperature 240 °C, detector temperature 250 °C, and column temperature 170 °C in the beginning, maintained for 24 min, increased to 220 °C at 7.5 °C/min, 230 °C at 10 °C/min, and maintained for 5 min. The contents of both cis and trans fatty acids showed a decreased trend for the increase of heating time or temperature. No trans fatty acid formation was observed even after extensive heating of unhydrogenated and hydrogenated soybean oil for 24 h. This phenomenon demonstrated that trans fatty acids can only be formed under severe conditions.     

Effect of thermal pretreatment of raw soymilk on the gel strength and microstructure of tofu induced by microbial transglutaminase

The influence of thermal pretreatment of raw soymilk on the gel hardness and microstructure of tofu, induced by microbial transglutaminase (MTGase), was investigated in this paper. Modulated differential scanning calorimetry analysis showed that individual proteins in soymilk were to a various extent denatured by different thermal pretreatments. The viscosity of the soymilk and the gel hardness of MTGase-induced tofu were more highly related with the heating rate (up to 90 °C) than the mode of heating. At any enzyme concentration of MTGase, the tofus prepared from soymilk heated at 75 °C for 10 or 30 min showed highest gel hardness among all tested ones (P?0.05). Scanning electron microscopy analysis indicated that the microstructure of the tofu from soymilk heated at 75 °C for 30 min had a unique coral-like structure, much more continuous and homogenous than that from soymilk at 95 °C for 5 min. These results confirmed that the appropriate heat pretreatment (e.g. in the present, at 75 °C for 10-30 min) remarkably improved the gel strength of tofu by means of MTGase, and strengthened the tofu gel structure.     

High early post-mortem temperature induces activation of AMP-activated protein kinase and development of pale,soft and exudative characteristics in turkey muscles

This study investigated the effect of early post-mortem temperature on development of a turkey muscle's pale, soft and exudative (PSE) characteristics. Muscles obtained at 20 min post-mortem were incubated at 0, 20 and 40 °C until 4 h post-mortem and then stored at 4 °C. During incubation, the 40 °C group had greater rate of pH decline, lactate accumulation and R-values increase than other groups. Moreover, AMP-activated protein kinase (AMPK) activation at 1 h post-mortem was higher in the 40 °C group than the other groups. At 24 h post-mortem, the 40 °C group had higher L* values and drip loss; SDS-PAGE and western blotting indicated that lower protein solubility (sarcoplasmic, myofibrillar) in the 40 °C group resulted from phosphorylase denaturation and further adhere to myofibrillar fraction. These results suggest that high temperature early post-mortem could induce AMPK activation, which results in rapid glycolysis, thus affecting protein solubility and generating PSE characteristics.