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The combined effect of ascorbic acid (AsA) and hydrogen peroxide (H2 O2 ) on gel-forming ability and structural changes of lizardfish (LZ) and threadfin bream (TB) surimi were investigated. Addition of 0.15% AsA and 0.1% H2 O2 greatly increased breaking force and distance of LZ surimi by 300% and 55%, respectively. Combination of 0.2% AsA and 0.15% H2 O2 resulted in the maximum TB surimi gel improvement with 150% and 90% increase in breaking force and distance, respectively. Browning reaction obviously occurred when combined AsA and H2 O2 was added, due to ascorbic acid oxidation. Polymerization of myosin heavy chain via disulfide bonds was promoted, and the formation of disulfide bonds was involved through oxidation of sulfhydryl groups with increasing AsA and H2 O2 . Fourier-transform infrared (FT-IR) spectroscopy revealed a decrease in α-helix and an increase in β-sheet content as AsA and H2 O2 increased in both species. A decrease of band area of aliphatic (2,800 to 3,000 and 1,450 cm-1 ), aromatic (1,208, 757, and ratio 850/830 cm-1 ), and change of disulfide bonds (525 and 540 cm-1 ) suggested an increase in hydrophobic interactions and disulfide bonds with addition of these additives. Based on principal component analysis (PCA), textural characteristics were positively correlated with β-sheet content. Our study suggested that combination of AsA and H2 O2 greatly enhanced gelation of LZ and TB by increasing not only disulfide bonds but also hydrophobic interactions. PRACTICAL APPLICATION: The combined ascorbic acid and hydrogen peroxide can be used to improve gelation of two important tropical surimi species, namely threadfin bream and lizardfish surimi, without requirement of setting. The optimum concentration of each additive varied with fish species.

The effect of thermal treatments on chemical and cellular antioxidant activities of chicken breasts subjected to in vitro gastrointestinal digestion was investigated. Breast of Korat crossbred chicken (KC) and commercial broiler (BR) were cooked under various conditions, namely heating at 70 °C for 30 min (H-0.5) and 24 h (H-24), autoclaving (AC) at 121°C for 15 min (AC-15) and 60 min (AC-60). Protein digestibility decreased upon the extreme thermal treatment of AC-60. The H-0.5 improved metal chelating activity of KC digesta, FRAP, and anti-liposome oxidation of BR digesta. Digesta of BR/H-0.5 and KC/AC-15 at 50 μg/mL exhibited the highest cytoprotective effect against tert-butyl hydroperoxide (TBHP)-induced oxidative damage of HepG2 cells. In addition, the KC/AC-15 digesta at a concentration as low as 12.5 μg/mL inhibited intracellular TBHP-induced reactive oxyfen species (ROS) production (P < 0.05). Thus, the digesta of KC breasts subjected to AC-15 provides not only nutritional value but also antioxidant activity at the cellular level.

Staphylococcus sp. CMC5-3-1 and CMS5-7-5 isolated from fermented fish sauce at 3 to 7 mo, respectively, showed different characteristics on protein hydrolysis and volatile formation. These Gram-positive cocci were able to grow in up to 15% NaCl with the optimum at 0.5% to 5% NaCl in tryptic soy broth. Based on ribosomal 16S rRNA gene sequences, Staphylococcus sp. CMC5-3-1 and CMS5-7-5 showed 99.0% similarity to that of Staphylococcus piscifermentans JCM 6057(T) , but DNA-DNA relatedness was

The hydrolysis of milk proteins by the recombinant AprX-SK37 protease and the changes in the rheological properties of the milk gel generated with AprX-SK37 and glucono-δ-lactone (GDL) were investigated. The AprX-SK37 and rennet selectively hydrolysed κ-casein to yield a 16-kDa band, while subtilisin hydrolysed all of the casein components. Milk treated only with AprX-SK37 formed softer gel. Storage modulus (G′) values of the combined gels increased with GDL concentrations up to 7 g/L. High tan δ was observed in the combined gel at 8.75 g/L GDL alongside syneresis. AprX-SK37 is a promising milk-clotting enzyme when combined with an optimal GDL concentration.

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.

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

A means to accelerate fish sauce fermentation without adversely affecting fish sauce quality was investigated. Starter cultures prepared from Virgibacillus sp. SK33, Virgibacillus sp. SK37, and Staphylococcus sp. SK1-1-5 were added separately to anchovy that was hydrolyzed by 0.25% Alcalase at 60 degrees C for 2 h followed by 0.5% Flavourzyme at 50 degrees C for 4 h. The mixtures were then adjusted to contain 25% solar salt and incubated at 35 degrees C for 4 mo. alpha-Amino contents of all inoculated samples were higher than the control (without the addition of starter culture) during the course of fermentation. After 4-mo fermentation, the samples inoculated with Staphylococcus sp. SK1-1-5 contained the highest alpha-amino content of 733.37 +/- 13.89 mM while that of the control was 682.67 +/- 3.33 mM. Amino acid profiles of inoculated samples showed similar patterns to that of commercial product fermented for 12 mo, with glutamic, aspartic, and lysine being predominant amino acids. Virgibacillus sp. SK33 appeared to decrease histamine content of fish sauce by 50% when compared to the control. Volatile compounds analyzed by GC-MS of all inoculated samples fermented for 4 mo exhibited a similar pattern to those of the 12-mo-old commercial product. Samples inoculated with Staphylococcus sp. SK1-1-5 produced higher levels of volatile fatty acids and showed similar sensory characteristics to the commercial fish sauce fermented for 12 mo. Staphylococcus sp. SK1-1-5 is a potential strain that can be applied to produce fish sauce with overall sensory characteristics of traditional fish sauce in shorter time.

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

Solubility of rockfish whole muscle and actomyosin was minimum at pH 5 and gradually increased as the pH was shifted to acidic or alkaline pH. Acidic and alkaline solubilization was followed by isoelectric precipitation induced degradation of myosin heavy chain, resulting in a protein band of about 120 kDa. Both myofibrillar and sarcoplasmic proteins underwent denaturation after acidic and alkaline treatment, exhibiting minimal solubility and absence of endothermic peaks. Acid- and alkali-treated muscle proteins readily aggregated upon heating, showing different dynamic rheological patterns compared with whole muscle and washed mince. Disulfide linkages occurred at a greater extent in gel prepared by alkaline solubilization, resulting in higher breaking force and deformation.

Trypsin-like proteinases exhibited the highest activity in viscera and muscle of Indian anchovy (Stolephorus spp.). The molecular weights (MWs) of proteinases in viscera were estimated to be 31, 35, 44, 49 and 57 kDa by activity staining in the presence of 4 mol L−1 NaCl. The MW of proteinase extracted from muscle was 56 kDa. This reflects that only one proteinase was extracted from muscle whereas five proteinases were extracted from viscera. Trypsin-like, chymotrypsin-like and cathepsin L-like proteinases were found in commercial fish sauce samples throughout 12 months of fermentation, suggesting that these proteinases were stable at high salt concentrations (250–300 g kg−1 NaCl) and under acidic conditions (pH 5.2–5.8). In contrast, leucine aminopeptidase activity was detected only in the first month. The MWs of proteinases found in fish sauce after various fermentation periods were estimated to be 37, 47 and 53 kDa, which coincided with the MWs of proteinases found in Indian anchovy. Copyright © 2006 Society of Chemical Industry

The effect of Ca2+ on physicochemical and conformational changes of threadfin bream (TB) myosin and actin during setting at 25 and 40°C was investigated. Ca2+ ion at 10 to 100 mM induced the unfolding of myosin and actin as evident by an increase of surface hydrophobicity (So ANS) at 40 °C. Total SH groups also decreased with an increased Ca2+ concentration, suggesting that Ca2+ promoted the formation of disulfide bonds during setting at 40 °C. Both hydrophobic interactions and disulfide linkages were involved in formation of myosin aggregates at 40 °C and were enhanced by addition of 10 to 100 mM Ca2+. Myosin Ca-ATPase activity decreased when Ca2+ was greater than 50 mM, indicating conformational changes of myosin head. Circular dichroism spectra demonstrated that Ca2+ reduced the α-helical content of myosin and actin incubated at either 25 or 40 °C. Ca2+ induced conformational changes of TB myosin and actin incubated at 40 °C to a greater extent than at 25 °C.

In the absence of microbial transglutaminase (MTGase), the textural properties of lizardfish surimi (Saurida spp) improved when pre-incubated at 4 and 25 degrees C for 24 and 4 h, respectively. MTGase optimally catalyzed incorporation of monodansylcadaverine (MDC) into surimi at 40 degrees C. Addition of MTGase appeared to reduce autolytic activity at 25 and 40 degrees C, but had no effect on autolytic activity at 65 degrees C. Breaking force and deformation of lizardfish surimi significantly improved when 0.1 unit MTGase g(-1) surimi (1.8 g kg(-1)) was added and pre-incubated at either 25 or 40 degrees C. Textural properties improved concomitant with cross-linked polymers of myosin heavy chain and tropomyosin, but not actin. Addition of MTGase also improved the storage modulus (G'). The gel network of surimi mixed with MTGase and pre-incubated at 40 degrees C readily formed during the pre-incubation period, while formation of the gel network began at 48.1 degrees C in the absence of MTGase.

The formation of biogenic amines in fish sauce made from fresh and temperature-abused (left at 35°C for 8 and 16 h) Indian anchovy (Stolephorus indicus) was investigated. Histamine, cadaverine, putrescine, and tyramine were the predominant biogenic amines found in anchovy left at 35°C for 16 h and its fish sauce product. Changes of biogenic amines were subtle during the course of fermentation at room temperature (RT) and at 40°C, suggesting that the main source of biogenic amines was associated with raw material, rather than with the fermentation process. Soluble peptide and total nitrogen of fish sauce prepared from temperature-abused anchovy were higher at the initial stage of fermentation at RT and 40°C and became comparable to those prepared from fresh anchovy at the end of fermentation. Total free amino acid contents of samples fermented at RT for 52 wk (7208.3 to 8473.6 mg/100 mL) were higher than those fermented at 40°C for 13 wk (4560.9 to 5730.9 mg/100 mL). Fish sauce prepared from temperature-abused anchovy contained less free histidine and arginine. Fish sauce of a good quality was obtained using fresh anchovy fermented at RT. Besides total nitrogen content, biogenic amines should be considered as quality indicators of fish sauce.

Transglutaminase(TGase) activity of threadfin bream mince was 99.6 units/g of dry weight. After washing and screw-pressed dewatering, 44% residual activity was retained. Covalent cross-linking of myosin heavy chain (MHC) was observed at both 25 and 40°C and supported by increased gel strength. When pre-incubation at 40°C was prolonged to 4 h, breaking force and MHC decreased due to endogenous proteinase(s). TGase activity towards MHC and synthetic substrates was effectively inhibited by iodoacetic acid (IAA). Autolytic activity and degradation of MHC was inhibited by phenylmethanesulfonyl fluoride (PMSF). Addition of 0.2% Ca(2+) significantly improved breaking force and increased MHC cross-linking of surimi gels pre-incubated at 40°C for 2 h. Keywords: transglutaminase, myosin heavy chain, cross-linking, threadfin bream.

Proteolytic degradation of tropical tilapia surimi was biochemically and rheologically characterized to identify a group of proteinase(s) responsible for its textural degradation. Proteolysis of tilapia surimi occurred as the temperature increased and attained the highest activity at 65 °C. Smaller proteins with molecular weight of 116-97 kDa were noted as a result of myosin heavy chain (MHC) degradation. MHC completely disappeared when incu- bated at 65 °C for 4 h. Proteolysis was significantly inhibited by soybean trypsin inhibitor (SB) and leupeptin (LE). Storage modulus (G) of surimi gels mixed with either SB or LE was higher than other inhibitors indicating that serine type proteinase(s) were involved in proteolysis of tropical tilapia surimi.

Thermal aggregation of cod myosin and Pacific whiting myosin were compared at different heating rates. Turbidity of Pacific whiting started to increase at 30°C and decreased at >50°C, and fluorescence intensity of ANS-myosin of both species was greater at slower heating rates at 20-80°C. Storage modulus (G') of cod myosin gradually increased as temperature increased resulting in more elastic gel at slower heating rates. G' of whiting myosin rapidly increased at 30°C, maximized at 45-48°C and decreased afterward. The onset temperature of a decreased G' shifted from 45°C to 49°C as heating rate increased from 0.5° to 2°C/min. Whiting myosin heated at 2°C/min retained more myosin heavy chain.

Degradation kinetics of whiting surimi gel texture were examined over a temperature/time range (40-85C, 0.5-35 min). Changes in textural properties of whiting surimi gel were mainly affected by proteolytic activity of endogenous proteinase. A decrease of failure shear stress and shear strain followed first order kinetics. The kinetic parameters developed using either isothermal or nonisothermalprinciples were similar. Degradation rate of gel texture increased with temperature, reaching a maximum at 55C. It then decreased to a minimum at 70C. E a values for the activation and inactivation temperature range were 138.6-162.6 and 13.5-35.0 kJ/mol, respectively.

Pacific whiting surimi paste was ohmically heated to investigate degradation of myosin heavy chain (MHC) caused by endogenous proteinase over a range of 40–85°C and 0.5–35 min. Degradation was best described with an apparent reaction order of 1.4. Changes of degradation rate increased with temperature and reached a maximum at 57°C. Then, rate of MHC degradation decreased with higher temperature and reached a minimum at 75°C. Ea values of activation and inactivation zone were 142.3 and 83.1 kJ/mol, respectively. Generally, failure shear stress and shear strain increased linearly with MHC content. Proteolytic degradation of actin exhibited the same trend as that of MHC but at a slower rate. The synergistic effect of actin in the gelation of whiting surimi was predominant at ≥75°C.

Shear stress of Alaska pollock surimi gels with and without beef plasma protein (BPP) increased as heating rate decreased, but shear strain was unaffected. An increase in shear stress was accompanied by an increase of cross-linked myosin heavy chain. Slow heating rates increased proteolysis in Pacific whiting surimi as shown by degradation of myosin heavy chain and low shear stress and shear strain. Proteolysis of whiting surimi was lessened by BPP to a greater extent at rapid heating rates (20 and 30°C/min) than at slow heating rates (1 and 5°C/min).

Electrical conductivities of Pacific whiting surimi paste with various moisture contents (75, 78, 81, and 84%) and added salt (1, 2, 3, and 4%) were measured using ohmic heating at alternating current of 3.3, 6.7, and 13.3 V/cm. Electrical conductivity of surimi increased with temperature and salt content and slightly increased with moisture content. Electrical conductivity correlated linearly with temperature (r2 approximately equal to 0.99). Generally, voltage gradient did not affect conductivity. However, variations of conductivity with voltage gradient observed in surimi containing 3-4% salt, were probably caused by electrochemical reactions at electrode surfaces. The empirical model of electrical conductivity predicted values +/- 16% of independent experimental results.

Surimi without enzyme inhibitors containing 78% moisture and 2% NaCl was heated conventionally and ohmically to 90 o C after holding at 55 o C for 0, 1, 3 and 5 min. Gels heated slowly in a water bath exhibited poor gel quality, while the ohmically heated gels without holding at 55 o C showed more than a twofold increase in shear stress and shear strain over conventionally heated gels. Degradation of myosin and actin was minimized by ohmic heating, resulting in a continuous network structure. Ohmic heating with a rapid heating rate was an effective method for maximizing gel functionality of Pacific whiting surimi without enzyme inhibitors

Gelation behavior and potential cross-linking of Pacific whiting (Merluccius productus) surimi were affected by setting temperatures and an enzyme inhibitor. Gels of Pacific whiting surimi with salt and beef plasma protein were compared with those containing guanidine hydrochloride, sodium dodecyl sulfate, and β-mercaptoethanol. The strongest gels were formed at 25 o C setting followed by 90 o C heating. Hydrogen and hydrophobic bonds appeared to strongly influence gel formation, while the influence of disulfide bonds was moderate. Viscosity scanning during setting at different temperatures was also useful to estimate effects of enzymes and inhibitors