Proteinase inhibitory activity of sarcoplasmic proteins from threadfin bream (Nemipterus spp.)

BACKGROUND: Thailand is the second largest surimi producer in the world and 50% of surimi is produced from threadfin bream. During surimi processing, sarcoplasmic proteins are removed through water washing and discarded in the waste stream. This study was aimed at investigating the proteinase inhibitory activity of sarcoplasmic proteins. RESULTS: Sarcoplasmic proteins from threadfin bream (TBSP) exhibited inhibitory activity toward trypsin but did not inhibit papain and chymotrypsin. Sodium dodecyl sulfate–polyacrylamide gel electrophoresis under non‐reducing condition stained by trypsin inhibitory activity revealed three protein bands of molecular mass of 95, 41 and 37 kDa. Inhibitory activity of TBSP reached a maximum when subjected to 45 °C and completely disappeared at 60 °C. The breaking force and deformation of lizardfish surimi gel with added TBSP and pre‐incubated at 37° for 20 min increased with additional levels of TBSP (P < 0.05). Trichloroacetic acid–oligopeptide content of lizardfish surimi gel with added TBSP decreased with the addition of 4 g kg^?1 TBSP (P < 0.05). Retention of myosin heavy chain (MHC) increased when TBSP concentration was increased. TBSP effectively protected MHC from proteolysis at 37 °C to a similar extent as egg white powder, but efficacy of TBSP was not observed at 65 °C. CONCLUSION: TBSP could be applied to reduce proteolytic degradation of lizardfish surimi or other surimi associated with trypsin‐like proteinase, rendering an improvement in surimi gelation set at 37–40 °C. Copyright © 2009 Society of Chemical Industry     

Gel‐enhancing effect and protein cross‐ linking ability of tilapia sarcoplasmic proteins

BACKGROUND: Tilapia (Oreochromis niloticus) sarcoplasmic proteins contain substantial transglutaminase (TGase) activity. The enzyme catalyzes the protein cross‐linking reaction, resulting in a more elastic gel. The objective was to investigate the gel‐enhancing effect of sarcoplasmic proteins from tilapia as related to TGase activity. RESULTS: Total TGase activity of sarcoplasmic proteins concentrate (SpC) increased about 3.6‐fold after ultrafiltration using 30 kDa membrane, but specific activity remained unchanged, indicating minimal TGase purification by ultrafiltration. Addition of 1 mg mL^?1 SpC containing 40 units TGase activity induced cross‐linking of tilapia actomyosin, and the extent of cross‐linking increased with added level of SpC. Myosin heavy chain (MHC) and troponin were preferably cross‐linked by tilapia SpC, while actin and tropomyosin were not affected. Higher retention of MHC was observed concomitantly with greater content of cross‐linked protein when SpC was added to lizardfish surimi. Lizardfish surimi with 10 g kg^?1 SpC added and pre‐incubated at 37 °C for 1 h exhibited 91.6% and 26.7% increase in breaking force and deformation, respectively, when compared to the control. CONCLUSIONS: Residual TGase activity in SpC played an important role in catalyzing the protein cross‐linking and enhancing actomyosin gelation. SpC could be a potential ingredient for improving textural properties of fish protein gel. Copyright © 2007 Society of Chemical Industry     

Comparative study on protein cross-linking and gel enhancing effect of microbial transglutaminase on surimi from different fish

BACKGROUND: Microbial transglutaminase (MTGase) has been used to increase the gel strength of surimi. Nevertheless, its effectiveness varies with fish species. The aim of this study was to elucidate the effect of MTGase at different levels on protein cross‐linking and gel property of surimi from threadfin bream, Indian mackerel and sardine in the presence and absence of endogenous transglutaminase. RESULT: Breaking force of all surimi gels increased as MTGase levels (0–0.6 U g^?1) increased except for threadfin bream surimi gel, where the breaking force decreased at 0.6 U g^?1 (P < 0.05). In the presence of EDTA, the gel strengthening effect was lower, suggesting the combined effect of endogenous transglutaminase with MTGase. With the addition of MTGase, the gel with the highest increase in breaking force showed highest decrease in myosin heavy chain. When cross‐linking activity of MTGase on natural actomyosin (NAM) was determined, the highest decreasing rate in ε‐amino group content with the concomitant increased formation of cross‐linked proteins was found in NAM from threadfin bream. The reactivity of muscle proteins toward MTGase‐induced cross‐linking was in agreement with surimi gel strengthening. CONCLUSION: The composition and properties of muscle proteins of varying fish species more likely determined protein cross‐linking induced by MTGase, thereby affecting their gel properties. Copyright © 2011 Society of Chemical Industry     

Thermal Gel Degradation (Modori) in Sturgeon (Acipenseridae) Surimi Gels

Sturgeon meat has been found to be suitable as surimi raw materials. The present study determined the modori phenomenon in sturgeon surimi gels and identified its relationship with cathepsins. In all heat‐treated gels (25 to 90 °C, at 5 °C intervals), the 40 °C‐incubated sturgeon surimi gel showed the weakest gel properties and water‐holding capacity (P < 0.05), a rough protein gel network under SEM, and the highest protein solubility and trichloroacetic acid‐soluble peptides content (P < 0.05). SDS‐PAGE indicated that the myosin heavy chain band of sturgeon surimi gels was almost completely degraded at 40 °C. Moreover, the highest cathepsin L activity was observed in 40 °C‐treated sturgeon surimi gels (P < 0.05). Our results suggested that the modori phenomenon in sturgeon surimi gels occurred at 40 °C, which was partially attributed to cathepsin L, thereby allowing for the better exploitation and utilization of sturgeon surimi.     

Effects of Bambara Groundnut Protein Isolates and Microbial Transglutaminase on Textural and Sensorial Properties of Surmi Gel from Sardine (Sardinella albella)

Effects of different bambara groundnut protein isolates (BGPIs) at a level of 6 % (w/w) in combination with microbial transglutaminase (MTGase) at a concentration of 0.6 U g^?1 surimi on gel properties of sardine (Sardinella albella) surimi were investigated. In the absence of MTGase, all BGPIs showed the adverse effect on gel-forming properties of surimi, as evidenced by the decreases in breaking force and deformation (P?<?0.05). When MTGase was incorporated, the increases in breaking force and deformation were found for all BGPIs used. Water-holding capacity of all gels was improved when BGPIs were added in combination with MTGase (P?<?0.05). Whiteness of gels slightly decreased with the addition of BGPIs; however, MTGase had no impact on whiteness (P?>?0.05). Surimi gel added with BGPI prepared from defatted flour with heat treatment in the presence of ethylenediaminetetraacetic acid (DF-BGPI-HE) and MTGase showed well-ordered network and exhibited the lowest peroxide value and thiobarbituric acid-reactive substances than those containing other BGPIs. Gel containing DF-BGPI-HE had negligible beany flavour. Additionally, DF-BGPI-HE had the lower amount of volatile compounds after storage of 30 days at room temperature than other BGPIs. Thus, the addition of DF-BGPI-HE and MTGase was an effective means to render sardine surimi gel with improved gel property and caused no beany flavour in resulting gel.     

Effect of formaldehyde on protein cross-linking and gel forming ability of surimi from lizardfish induced by microbial transglutaminase

Impact of formaldehyde (FA) at various levels (0–9 μmol/g surimi) on gel properties of surimi from lizardfish added with microbial transglutaminase (MTGase) was studied. During iced storage of 10 days, total and free FA in lizardfish flesh increased continuously (P < 0.05). In the presence of FA, breaking force of gels slightly increased, whilst the deformation decreased (P < 0.05). The addition of MTGase (0.4 units/g surimi) was able to increase gel strength and water holding capacity of resulting gel. Nevertheless, gel strengthening effect of MTGase was lowered when FA at higher level was present. Myosin heavy chain (MHC) dominantly underwent polymerisation to a higher extent when either MTGase or FA was added. The higher reduction in ε-amino group content was observed in natural actomyosin (NAM) when FA at higher levels (0–30 μmol/g protein) was incorporated. Acyl transfer reaction mediated by MTGase was impeded in NAM containing FA, especially at higher levels. Generally, FA had an adverse effect on cross-linking ability towards surimi proteins induced by MTGase. Therefore, cross-linking and gel-forming ability of lizardfish surimi could be maximised by MTGase when surimi contained no FA.     

Improvement of Gelling Properties of Lizardfish Mince as Influenced by Microbial Transglutaminase and Fish Freshness

ABSTRACT:  The effects of microbial transglutaminase (MTGase) at different levels (0 to 0.8 units/g sample) on the properties of gels from lizardfish ( Saurida undosquamis ) mince set 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 and deformation of gels increased with increasing MTGase amount added ( P < 0.05). At the same MTGase level used, gels with the prior setting at 40 °C for 30 min showed a higher breaking force compared with those subjected to prior setting at 25 °C for 2 h ( 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. Regardless of setting condition, microstructure of gel added with MTGase was finer with a smaller void compared with that of gel without MTGase. Therefore, setting temperature affected the property of gels added with MTGase. Gel properties of mince obtained from lizardfish stored in ice for different times (0 to 10 d) with and without MTGase at a level 0.6 units/g were determined. Irrespective of MTGase addition, breaking force and deformation of all gels decreased as the storage time of lizardfish increased ( P < 0.05). The addition of MTGase was able to increase both breaking force and deformation of the resulting gel produced from lizardfish kept in ice for all storage times used. Therefore, both freshness and MTGase addition had the direct impact on gel properties of lizardfish mince.     

Trypsin Inhibitory Activity and Gel‐Enhancing Effect of Sarcoplasmic Proteins from Common Carp

Abstract: Proteinase inhibitory activity of sarcoplasmic protein (SP) extracted from common carp (Cyprinus carpio) muscle and its gel‐improving ability were investigated. SPs displayed 89% and 54% inhibitory activity toward trypsin at 40 and 65 °C, respectively. Protein bands with molecular mass of 69, 50, 44, 41, and 35 kDa appeared on trypsin inhibitory activity staining under nonreducing condition when incubated at 40 °C, while 2 protein bands at 54 and 35 kDa were observed at 65 °C. Addition of SP at 0.18 g protein/100 g increased textural properties of threadfin bream surimi gel. However, when SP was added in combination with various CaCl₂ concentrations (0.1% to 0.5%) it did not further improve textural properties as compared to the addition of SP alone. Retention of myosin heavy chain of threadfin bream surimi was greater with the addition of SP. These results indicated that the gel‐enhancing effect of common carp SP was due to the inhibitory activity toward endogenous trypsin‐like proteinases in threadfin bream surimi. Practical Application: Sarcoplasmic protein from common carp muscle could be used as a functional protein ingredient that minimizes muscle proteolysis and improves textural properties of surimi containing trypsin‐like endogenous proteinases.     

Effect of dietary fibre and MTGase on the quality of mackerel surimi gels

BACKGROUND: The effects of microbial transglutaminase (MTGase; 5 g kg^?1) and dietary fibre (inner pea fibre and chicory root inulin, 20 and 40 g kg^?1) on gels from surimi made with Atlantic (Scomber scombrus) and chub mackerel (Scomber japonicus) were studied with the purpose of achieving a better understanding of the underlying phenomena and improving gels. RESULTS: MTGase addition improved textural properties (namely, cohesiveness increased from 0.19–0.41 to 0.59–0.72) and increased pH and water‐holding capacity (WHC). Moreover, MTGase reduced the elastic and viscous modules and darkened gels; protein solubility declined, meaning greater protein aggregation, according to electropherograms. MTGase had no unequivocal effect upon the gels' microstructure. Pea fibre increased hardness (an increase of almost 60% with 40 g kg^?1) and related parameters, the elastic and viscous modules and WHC. Pea fibre reduced extractable protein in sodium dodecyl sulfate and urea (in the absence of MTGase) as well. Scanning electron microscopy revealed different structures for gels containing pea fibre. Regarding inulin, it worsened textural quality and WHC. CONCLUSION: A combination of MTGase and pea fibre can improve a poor quality surimi. Copyright © 2009 Society of Chemical Industry     

Low‐salt restructured fish products using microbial transglutaminase as binding agent

Low‐salt restructured silver carp products were obtained using mechanically deboned fish meat from filleting wastes of silver carp (Hypophthalmichthys molitrix). The additives used were NaCl at three levels (0 (control), 10 and 20 g kg^?1) and microbial transglutaminase (MTGase) also at three levels (0 (control), 3 and 6 g kg^?1). The fish meat was massaged with the additives at <15 °C for 1 h. The massaged fish paste was then packed into steel stainless tubes and cooked at 40 °C for 30 min followed by 90 °C for 15 min. Changes in mechanical properties (texture profile analysis and punch test), solubility, electrophoretic profile and expressible water were evaluated. Hardness was in the range from 26.3 to 52.4 N, cohesiveness varied from 0.185 to 0.318 and springiness varied from 0.418 to 0.768. Increasing the amount of both additives improved the mechanical and functional properties of the restructured silver carp products. MTGase activity was associated with a decrease in protein solubility and a decrease in the myosin band (SDS‐PAGE). Increasing NaCl decreased the amount of expressible water. The results indicated that it is feasible to obtain low‐salt restructured silver carp products with improved mechanical and good functional properties using 3 g kg^?1 MTGase and 10 g kg^?1 NaCl. © 2002 Society of Chemical Industry     

Microbial Transglutaminase and ε-(γ-Glutamyl)lysine Crosslink Effects on Elastic Properties of Kamaboko Gels

Kamaboko gels from Alaska pollock surimi (SA? and 2nd? grades) were prepared by setting at 10 or 45°C with Microbial transglutaminase (MTGase) and its effect on gel properties was investigated. At 10 and 45°C, gels from 2nd? grade surimi paste showed increases in breaking strength, without decline in deformation. Gel from SA? graded surimi paste showed an increase in breaking strength with no changes in deformation in 45°C setting, up to 0.03% MTGase. Crosslinking of myosin heavy chains through ε-(γ-glutamyl)lysine bonds was observed and a possible correlation was shown between ε-(γ-glutamyl)lysine content and gel strength (breaking strength X strain). ε-(γ-Glutamyl)lysine content up to 3 μmol/100g or MTGase 0.03% or higher improved gel properties.     

Protein cross-linking ability of sarcoplasmic proteins extracted from threadfin bream

Sarcoplasmic proteins extracted from threadfin bream (TB, Nemipterus sp.) contained transglutaminase (TGase) activity exhibiting a pH optima at 7.5. Activity increased with CaCl₂ and reached the maximum at 5 mmol/L, showing the Ca²⁺-dependent characteristic. TGase activity staining on native-polyacrylamide gel electrophoresis (native-PAGE) showed two fluorescent bands catalyzing incorporation of monodansylcadavarine (MDC) into di-methylated casein (DMC). However, both fluorescent bands showed only one major protein band with a molecular weight of about 66 kDa on sodium dodecylsulfate-PAGE (SDS-PAGE), suggesting the presence of isozymes. The TB sarcoplasmic protein (TBSP) concentrated using a 30-kDa ultrafiltration membrane induced cross-linkings of bovine serum albumin when incubated at 25 °C for 6 h. Addition of the concentrated TBSP at 1.6 g protein/100 g along with 0.1 g CaCl₂/100 g resulted in a 1.8-fold increase in the breaking force of the proteinase-laden lizardfish surimi pre-incubated at 37 °C for 20 min. TBSP effectively minimized proteolysis of lizardfish surimi at 37 °C. TBSP could be a promising protein additive that improves textural properties of proteinase-laden surimi.     

Quality Improvement of Mackerel Surimi with NADPH-Sulfite Reductase from Escherichia coli

The application of NADPH‐sulfite reductase, produced from Escherichia coli, on mackerel surimi was investigated. The activity of this reductase was very stable. There was about 70%, 80%, and 90% activity left even after 8–wk storage at 25 °C and 12–wk storage at 4 or ‐30 °C, respectively. Increase in mackerel‐surimi gel properties containing reductase was consistent with the amounts of reactive sulfhydryl groups detected. SDS‐PAGE indicated that NADPH‐sulfite reductase could reconstitute myosin heavy‐chain and increase soluble actomyosin. From the data obtained, E. coli NADPH‐sulfite reductase effectively improved the gel properties of mackerel surimi.     

Gel Strength Enhancement by Addition of Microbial Transglutaminase during Onshore Surimi Manufacture

Surimi from Alaska pollock flesh was manufactured onshore with Microbial transglutaminase (MTGase). Effect of MTGase was investigated by evaluating breaking strength and deformation of gels from MTGase-treated surimi with and without setting at 30°C. Quantitative analysis of ε-(γ-glutamyl)lysine (GL) crosslink was also carried out to monitor the MTGase reaction. In set gels, breaking strength and GL crosslink increased, and myosin heavy chain decreased correspondingly with MTGase concentration. These changes were smaller in gels prepared without setting. Results suggest that surimi gel could be improved through the formation of GL crosslinks by added MTGase in surimi.     

Effects of Thermal Sensitivity of Fish Proteins from Various Species on Rheological Properties of Gels

ABSTRACT: Thermal sensitivity of myofibrillar proteins from various fish species were compared at various preincubation (setting) treatments and chopping temperatures. There was a significant species effect on gel texture by both pre‐incubation and chopping temperatures. Whereas Alaska pollock had the highest shear stress values at 5 °C or lower temperatures, big eye, lizardfish, and threadfin bream had higher fracture shear stress values at 25 °C or higher temperatures. Decreased intensity of myosin heavy chain (MHC) for warm‐water species set at 40 °C clearly revealed higher thermal stability of these particular species.     

Influence of texture of suwari gels on kamaboko gels made from sardine (Sardina pilchardus) surimi

The textural characteristics and water holding capacity of suwari (set) and kamaboko (set and cooked) sardine surimi gels were examined in order to clarify the influence of the initial network formed in setting conditions (25, 35 and 40°C for 30 and 60 min) on the texture of the kamaboko gels. Although the texture of suwari gels set at 35°C improved with longer setting, both setting times ensured kamaboko gels with the highest gel strength. Suwari gels set at 25°C also improved with longer setting but the gel strength of both suwari and kamaboko gels was lower than at 35°C. For gels set at 40°C prolonged setting weakened the suwari networks formed, leading to kamaboko gels with poorer textural characteristics. ©1997 SCI     

Quality of dried white salted noodles affected by microbial transglutaminase

To examine the potential application of microbial transglutaminase (MTGase) in oriental noodle making, the effects of various MTGase addition levels on the rheological, textural and structural properties of noodles were investigated using good quality (‘Red Bicycle’) and poor quality (‘Sandow’) wheat flours. Addition of MTGase at 5–20 g kg^?1 levels, but not at 1 g kg^?1 level, to the two different wheat flours decreased rapid visco‐analyser (RVA) parameters of hot paste viscosity and final viscosity while increasing breakdown. For fresh white salted noodle dough sheets, the storage modulus (G′) and loss modulus (G″) increased significantly at 1 g kg^?1 MTGase addition for both types of flour, but there was no clear trend with higher levels of MTGase. For dried white salted noodles, textural parameters (tensile force, hardness and gumminess) generally increased, cooking loss was little affected and the yield of the cooked noodle was significantly decreased by MTGase. Color was slightly adversely affected. Scanning electron microscopy (SEM) results indicated that physical properties of dry noodles were improved through the formation of cross‐links [ε‐(γ‐glutamyl)lysine] by MTGase. Copyright © 2005 Society of Chemical Industry     

Participation of cysteine protease cathepsin L in the gel disintegration of red bulleye (Priacanthus macracanthus) surimi gel paste

BACKGROUND: Endogenous proteases, among them cysteine‐type proteases, are reported to contribute to gel disintegration, resulting in kamaboko of poor quality. Severe gel disintegration occurs in red bulleye surimi gel paste. The objective of this study was to clarify the participation of cysteine protease cathepsin L in the gel disintegration of red bulleye surimi. The surimi was made into kamaboko with and without cathepsin L inhibitors. To confirm its hydrolysis action, crude cathepsin L was also extracted and added to the surimi to make kamaboko. RESULTS: The gel strength of kamaboko obtained by both one‐step (50 °C, 2 h) and two‐step (50 °C, 2 h + 80 °C, 20 min) heating was very low in the absence of inhibitors. Protease inhibitors E‐64 and leupeptin were found to enhance the gel strength considerably. Sodium dodecyl sulfate polyacrylamide gel electrophoresis revealed that the hydrolysis of kamaboko was promoted by crude cathepsin L and inhibited by E‐64 and leupeptin. The gel strength of two‐step heated kamaboko was increased from 12 to 110 and 130 g cm^?2 by E‐64 and leupeptin respectively at a concentration of 0.2 g kg^?1 surimi. CONCLUSION: Endogenous cathepsin L of red bulleye surimi participates in gel disintegration during kamaboko processing. It does so by degrading the myosin heavy chain of actomyosin and consequently hindering the gelation of red bulleye surimi. Copyright © 2009 Society of Chemical Industry     

The effect of modified starches on the gel properties and protein conformation of Nemipterus virgatus surimi

The study investigated the effect of native cassava starch (NCS), hydroxypropylated starch, acetic acid esterification starch (AAES), acetylated distarch phosphate on gel properties and protein conformation of Nemipterus virgatus surimi. Addition of 10 g kg^?1 NCS or 20 g kg^?1 AAES could significantly promote the gel strength and textural profiles of the surimi gels (p < .05). The water holding ability and whiteness of surimi were remarkably increased when the four types of starch were added at all concentrations (p < .05). In rheological test, the lower G′ was observed in surimi samples added with starch at low temperature, suggesting starch played an inactive filler role in surimi. Along with the increase of starch additive amount, ionic bond and hydrophobic interaction first increased and then decreased, while hydrogen bond first decrease and then increased. According to Raman spectroscopy data, small content of starch promoted the heat‐induced conformational transition of surimi protein from α‐helix to β‐sheet, leading to the change in gel properties of surimi gels. Scanning electron microscopy photographs showed surimi gels added with 20 g kg^?1 starch had the finer and denser network structure. Therefore, 20 g kg^?1 AAES or 10 g kg^?1 NCS or 10 g kg^?1 HS could be proposed to a potential modifier to effectively improve the quality of surimi products.     

Effect of microbial transglutaminase on rheological properties of oxidised and non‐oxidised natural actomyosin from two species of bigeye snapper

The contribution of microbial transglutaminase (MTGase) to the thermal gelation of non‐oxidised and oxidised natural actomyosin (NAM) from two species of bigeye snapper was studied by dynamic rheological testing. Under two‐step heating, the addition of MTGase increased the gel‐forming ability of both species. During setting at 40 °C, Priacanthus macracanthus NAM seemed to show higher reactivity to MTGase than P tayenus NAM, resulting in a higher rate and magnitude of G′ development. An increase in final G′ of NAM from P macracanthus and P tayenus by 13.7–24 and 6.6–10.8 times respectively was observed compared with NAM alone. Iron‐catalysed oxidation substantially decreased the gel‐forming ability of P tayenus NAM but enhanced that of P macracanthus NAM. However, oxidation decreased the reactivity of both NAM types to added MTGase. The results suggested that the native NAM structure was essential for MTGase to maximise the setting response. The gel‐forming ability and setting response of denatured NAM were partially recovered by the addition of MTGase. © 2002 Society of Chemical Industry