Dissociation of natural actomyosin from kuruma prawn muscle induced by pyrophosphate

Effect of pyrophosphate (PP) on the dissociation and stability of natural actomyosin (NAM) from kuruma prawn muscle was studied in comparison with adenosine 5′-triphosphate (ATP). In the presence of PP up to 5 mM, NAM exhibited lower Mg²⁺-ATPase activity (P < 0.05), while no marked change was observed in NAM treated with ATP at all concentrations tested (0.25–10 mM) (P > 0.05). Ca²⁺-ATPase activity of NAM treated with 5 mM PP decreased markedly when incubated at temperatures greater than 30 °C, suggesting lowered thermal stability of the liberated myosin molecule. Nevertheless, Ca²⁺-ATPase activity of ATP-treated NAM was similar to the control NAM. In the presence of 5–10 mM MgCl₂, NAM treated with 5 mM PP underwent dissociation effectively, as evidenced by a greater decrease in Mg²⁺-ATPase activity as well as an increased band intensity of actin released. Therefore, addition of PP in combination with MgCl₂ was more effective than was ATP in dissociating the actomyosin complex of prawn muscle.     

Effect of legume seed extracts on the inhibition of proteolytic activity and muscle degradation of fresh water prawn (Macrobrachiumrosenbergii)

Trypsin inhibitors in the extracts from soybean (Glycine max), adzuki bean (Vigna angularis), bambara groundnut (Vigna subterranea) and red kidney bean (Phaseoulus vulgaris) varied in amount and molecular weight. The soybean extract had the highest level of trypsin inhibitor with molecular weight (MW) of 21 kDa, followed by bambara groundnut extract possessing trypsin inhibitor with MW of 15 kDa. Both extracts showed a more effective inhibition towards crude protease extract (CE) from the hepatopancreas of fresh water prawn (Macrobrachium rosenbergii) than the extracts from adzuki and red kidney beans. Activity staining also reconfirmed the higher inhibitory activity on CE from hepatopancreas by the extracts from both soybean and bambara groundnut. The extracts from all seeds were able to inhibit the degradation of fresh water prawn meat containing CE in a concentration dependent manner. Based on inhibitor study, the extracts from soybean and bambara groundnut can be a potential aid to suppress the muscle softening of fresh water prawn, mediated by trypsin-like proteases released from hepatopancreas, during extended iced storage.     

Properties of protein-based film from round scad (Decapterus maruadsi) muscle as influenced by fish quality

The properties of film prepared from round scad (Decapterus maruadsi) stored in ice for different times were investigated. Degradation of myosin heavy chain (MHC) was more pronounced with the coincidental increase in total volatile base and trimethylamine contents as the storage time increased (P<0.05). Regardless of storage time, no changes in tensile strength (TS) and elongation at break (EAB) of resulting films prepared from unwashed mince were observed (P>0.05). For the films prepared from washed mince, TS decreased, whereas EAB increased when the storage time of fish increased (P<0.05). However, films prepared from washed mince showed the greater mechanical properties with the lower film solubility and protein solubility than did those from mince (P<0.05). Generally, films prepared from fish stored in ice for a longer time became less transparent, darker and more yellowish. The electrophoretic study revealed that similar protein patterns were observed between films, irrespective of storage time of fish and washing. Therefore, the quality of fish did not show the marked impact on the mechanical property of the resulting films, while washing likely affected the film forming ability.     

Inhibitory effect of cysteine and glutathione on phenoloxidase from kuruma prawn (Penaeus japonicus)

The inhibitory effect of cysteine and glutathione on phenoloxidase (PO) from kuruma prawn were investigated. Cysteine and glutathione inhibited the oxidation of 3-(3,4-dihydroxylphenyl)-l-alanine (l-DOPA) catalyzed by kuruma prawn PO. Those thiol compounds showed competitive inhibition with K_i values of 0.45–0.46 mM. The inactivated PO could be partially recovered by the addition of copper acetate (0.01–0.2 mM). Almost complete restoration of PO activity was achieved with the addition of N-ethylmaleimide (NEM) in the range 0.05–0.2 mM, suggesting the importance of sulfhydryl groups of cysteine and glutathione for their inhibitory activity. Additionally, both cysteine and glutathione prevented the colour development by trapping the colour intermediates or reducing o-quinone to colourless compounds.     

Changes in the ultrastructure and texture of prawn muscle (Macrobrachuim rosenbergii) during cold storage

Changes in the ultrastructure and proteolytic activity of prawn muscle were determined during storage at 5 °C, in order to better understand changes in physical and sensory texture measurements. Progressive deterioration of myofibril structure was observed during refrigeration of prawn for 14 d. The loss of density and order in Z-line alignment was first detected after 3 d of storage. Progressive disruption of Z-line, I-bands and M-lines was observed after 4-6 d of storage. Muscle degradation included pronounced disruption of the mitochondria as revealed by swollen cristae, loss of cristae material, and membrane breakage. Along with ultrastructural changes, decreased shear force values and mean textures scores were measured. An initial shear force value of 18.21 N/g decreased to 14.50, 12.46, and 10.79 N/g on days 3, 6, and 14, respectively. Mean texture scores indicated that prawn muscle maintained firm texture during 0-3 d of storage, and became soft during 4-6 d of storage. After 6 d of storage, the prawn texture was very soft. Increased deterioration of the muscle ultrastructure coincided with the increase of proteolytic microorganisms and salt soluble muscle proteins found by Sodium dodecyl sulfate-polyacrylamide gel electrophoresis. It is concluded that the weakening of the ultrastructure is related to proteolytic activity and results in a more soft texture in prawns.     

Physicochemical changes of tilapia (Oreochromis niloticus) muscle during salting

The effect of wet and dry saltings on the physicochemical changes of tilapia (Oreochromis niloticus) muscle was investigated. Dry salting resulted in the higher rate of salt uptake into tilapia muscle facilitating the faster decrease in A_w (p < 0.05). The pH of both dry and wet salted fish muscles tended to decrease throughout the salting time and the lower pH was found in dry salted fish (p < 0.05). The increase in the protein content in the salting medium was found during wet salted tilapia production (p < 0.05). The TCA-soluble peptide content tended to decrease with increasing the salting time in both salting methods (p < 0.05), suggesting a leaching effect of the salting medium or the exudative loss occurred in salted tilapia. Wet salting caused the greater formation of metmyoglobin in tilapia muscle when compared to dry salting at all time points (p < 0.05) and the content of metmyoglobin increased as salting time increased in both salting methods (p < 0.05). A lowered metmyoglobin with a lowered redness index of dry salted tilapia muscle was found, indicating the continuous oxidation of metmyoglobin to other hypervalent derivatives and hence the discolouration of salted tilapia. Lipid hydrolysis and oxidation of tilapia meat occurred with varying degrees in both salting methods and these changes depended on salting time. Dry salting resulted in a higher oxidation of tilapia muscle lipid as indicated by the higher PV and TBARS throughout the salting period when compared with that of wet salting (p < 0.05). In conclusion, the physicochemical changes of tilapia muscle during salting are governed by the salting method and the salting time applied.     

Characterisation of proteolytic enzymes from muscle and hepatopancreas of fresh water prawn (Macrobrachium rosenbergii)

BACKGROUND: Fresh water prawn in Thailand is widely consumed due to its delicacy. During postmortem handling and storage, prawn meat becomes soft and mushy, probably as a result of indigenous proteases. Therefore, an understanding of prawn proteases associated with the degradation of muscle proteins from fresh water prawn could pave the way for prevention of such a phenomenon during extended storage. RESULTS: Proteolytic enzymes in the crude extract (CE) from muscle and hepatopancreas of fresh water prawn (Macrobrachium rosenbergii) were characterised. CE from muscle exhibited the highest hydrolytic activities towards haemoglobin at pH 5 and 50 °C, while that from hepatopancreas had the highest activity on casein at pH 7 and 60 °C. Based on inhibitor study, cysteine protease and serine protease were dominant in CE from muscle and hepatopancreas, respectively. CE from muscle rarely hydrolysed natural actomyosin (NAM), but could not degrade pepsin‐soluble collagen (PSC). Conversely, NAM and PSC were susceptible to hydrolysis by CE from hepatopancreas as evidenced by the marked decreases in band intensity. Activity staining using haemoglobin, casein and gelatin as substrates revealed that no proteolytic or gelatinolytic activity was observed in CE from prawn muscle, while CE from hepatopancreas exhibited pronounced hydrolytic activities towards all substrates. CE from muscle showed calpain and cathepsin L activities but CE from hepatopancreas mainly exhibited tryptic and chymotryptic activities. CONCLUSION: Serine proteases, mainly trypsin‐like or chymotrypsin‐like, from hepatopancreas were probably responsible for the softening of prawn meat during postmortem storage via the degradation of both muscle and connective tissues. Copyright © 2010 Society of Chemical Industry     

Collagenolytic serine protease in fresh water prawn (Macrobrachiumrosenbergii): Characteristics and its impact on muscle during iced storage

Proteolytic activity of crude protease extract (CPE) from the hepatopancreas of fresh water prawn (Macrobrachium rosenbergii) was studied. Optimal activity of CPE was found at pH 7 and 60 °C when casein was used as a substrate. The activity was strongly inhibited by 10 mM N-ρ-tosyl-L-lysine chloromethylketone (TLCK), suggesting that trypsin-like protease was dominant. CPE also showed the collagenolytic activity toward pepsin soluble collagen extracted from prawn muscle. During extended iced storage of 4 days, proteolytic and trypsin activities were found in the first segment of prawn abdomen. These activities were detected in the second segment after 4 days of storage. Heat soluble collagen content was continuously increased during the storage. Nevertheless, no changes in proteolytic activity and heat soluble collagen content were obtained in the abdomen of prawn with the removal of hepatopancreas. Therefore, the release of trypsin-like collagenase from hepatopancreas was most likely responsible for the softening of prawn meat during iced storage.     

Retardation of post-mortem changes of freshwater prawn (Macrobrachium rosenbergii) stored in ice by legume seed extracts

Meat quality of freshwater prawn (Macrobrachium rosenbergii) treated with soybean and bambara groundnut extracts at different concentrations was monitored during 10 days of iced storage. During storage, the control sample (without treatment) had a higher pH, TCA-soluble peptide content, heat soluble collagen content, proteolytic activities and psychrophilic bacterial count than did samples treated with soybean and bambara groundnut extracts. Conversely, shear force value and likeness scores of the control sample decreased (p<0.05), more likely associated with softening of muscle. The decrease in myosin heavy chain in the control sample was found after 6 days of storage. However, no changes in protein patterns of samples treated with soybean extracts at 2.5 mg/mL were found after 10 days of storage. Therefore, the injections of legume seed extracts, especially soybean extract, at a sufficient concentration, could be a means to retard muscle softening and maintain the qualities of freshwater prawn during iced storage.     

Properties of a protein-based film from round scad (Decapterus maruadsi) as affected by muscle types and washing

The effects of muscle types and washing on the properties of a protein-based film from round scad (Decapterus maruadsi) mince were investigated. Washing resulted in an increase in the protein content with a coincidental decrease in the fat content of mince, especially from whole muscle and dark muscle. Among all types of muscle (ordinary, dark and whole muscle), the ordinary muscle rendered the film with the highest tensile strength (TS) (p < 0.05). TS of films from washed mince was greater than that of films prepared from unwashed mince for the same type of muscle used (p < 0.05). Nevertheless, the water vapour permeability (WVP) of films from unwashed mince was higher than that of films prepared from washed mince (p < 0.05). Films from washed mince had higher solubility but lower protein solubility than those from unwashed mince (p < 0.05). Regardless of washing, films from ordinary muscle showed the highest L^∗-value (p < 0.05). However, films prepared from dark muscle were more yellowish than those prepared from other muscles, as evidenced by the greater b^∗-value. Films from round scad mince and washed mince had excellent barrier properties to UV light at the wavelength of 200–280 nm. Generally, films from round scad mince had a lower preventive effect on visible light transmission than had those from unwashed mince. Among films from all muscles, those prepared from dark muscle exhibited the highest barrier to visible light transmission (p < 0.05). Therefore, the properties of films from round scad meat were governed by muscle type as well as by washing.     

Physicochemical changes of pepsin-solubilized and insoluble collagen in jumbo squid (Dosidicus gigas) muscle after cooking process

Collagen is the major connective tissue (CT) protein and one of the main constituents of the jumbo squid (Dosidicus gigas). Therefore, physicochemical changes of pepsin-solubilized collagen (PSC) and insoluble collagen (IC) were studied after cooking (100°C/30 min) of muscle (mantle, fins, and arms). Different pyridinoline (Pyr) contents (the major cross-linking molecule in collagen fibers) were found in the fresh muscle of the three anatomical regions. After the cooking process, a decrease from 10 to 30% in the thermal resistance of collagen was observed, depending on the anatomical region and fraction evaluated. Furthermore, the electrophoretic profile, Fourier transform infrared (FTIR) spectroscopy, and the amino-acid profile revealed that structural changes occurred in the two different collagen fractions caused by the thermal process, and the changes were greater in the mantle. Under the conditions applied in this study, collagen fractions from the squid arms showed more stability during the cooking process due to the high cross-linking degree of their fibers.     

Cross-linking activity of sarcoplasmic fraction from bigeye snapper (Priacanthus tayenus) muscle

Addition of sarcoplasmic fraction from bigeye snapper (Priacanthus tayenus) into natural actomyosin in combination with setting at 40°C resulted in the cross-linking of myosin heavy chain (MHC). Higher amount of sarcoplasmic fraction and extended setting time resulted in a higher cross-linking, indicating the presence of endogenous transglutaminase (TGase) in bigeye snapper muscle. TGase activity was activated by calcium ion and reducing agents (β-mercaptoethanol and dithiotreitol), but was inhibited by N-ethylmaleimide (NEM), NH₄Cl and EDTA. TGase in the sarcoplasmic fraction was not stable when heated at temperature above 40°C, particularly with an increasing heating time. TGase was stable at pH ranging from 5.0 to 7.0, in which more than 70% activity was retained. Therefore, sarcoplasmic fraction possessed a cross-linking activity caused by TGase and its recovery for further uses should be considered.     

Identification of xyloglucan endotransglucosylase/hydrolase genes (XTHs) and their expression in persimmon fruit as influenced by 1-methylcyclopropene and gibberellic acid during storage at ambient temperature

Xyloglucan endotransglucosylase/hydrolase (XTH) is thought to contribute to fruit softening by degrading xyloglucan that is a predominant hemicellulose in the cell wall. In this study, two full-length XTH genes (DKXTH1 and DKXTH2) were identified from ‘Fupingjianshi’ persimmon fruit, and the expression level of both XTH genes was investigated during softening for 18–24 d using RT-qPCR. Sequence analysis showed that DKXTH1 and DKXTH2 contained a putative open reading frame of 861 and 876 bp encoding polypeptides of 287 and 292 amino acid residues, respectively, which contained the conserved DEIDFEFLG motif of XTH, a potential N-linked glycosylation signal site. RT-qPCR analysis showed that DKXTH1 and DKXTH2 in untreated fruit had different expression patterns during fruit softening, in which maximum expression occurred on days 3 and 12 of ripening, respectively. 1-Methylcyclopropene (1-MCP) and gibberellic acid (GA₃) treatments delayed the softening and ethylene peak of persimmon fruit, as well as suppressed the expression of both XTH genes, especially DKXTH1. These results indicated that the expression of both XTH genes might be ethylene dependent action, and closely related to softening of persimmon in the early (DKXTH1) and later (DKXTH2) ripening stages.     

Decaffeination of fresh green tea leaf (Camellia sinensis) by hot water treatment

Hot water treatment was used to decaffeinate fresh tea leaf in the present study. Water temperature, extraction time and ratio of leaf to water had a statistically significant effect on the decaffeination. When fresh tea leaf was decaffeinated with a ratio of tea leaf to water of 1:20 (w/v) at 100 °C for 3 min, caffeine concentration was decreased from 23.7 to 4.0 mg g⁻¹, while total tea catechins decreased from 134.5 to 127.6 mg g⁻¹; 83% of caffeine was removed and 95% of total catechins was retained in the decaffeinated leaf. It is considered that the hot water treatment is a safe and inexpensive method for decaffeinating green tea. However, a large percentage of tea catechins was lost if rolled leaf and dry tea were decaffeinated by the hot water treatment and so the process is not suitable for processing black tea.     

Antioxidant activity of hydrolysates obtained from scallop (Patinopecten yessoensis) and abalone (Haliotis discus hannai Ino) muscle

Scallop (Patinopecten yessoensis) and abalone (Haliotis discus hannai Ino) muscle were hydrolysed with commercially available food-grade proteases. The resulting hydrolysates showed DPPH and hydroxyl radicals scavenging abilities, reducing power, and ferrous ion chelating capacity. The antioxidant activities of hydrolysate of abalone foot muscle (HAFM) increased with increasing incubation time during the whole hydrolysis process in 180 min. Whereas, the antioxidant activities of hydrolysate of scallop adductor muscle (HSAM) increased at initial stage and peaked after 25-30 min of hydrolysis, and then gradually decreased thereafter. Compared with HAFM, HSAM with comparable hydrolysis time contained more free amino acids (FAA) and small-sized peptides (below 500 Da), which may account for the differences in antioxidant activities versus hydrolysis time curves of the two hydrolysates. The above results indicate that limited hydrolysis of proteins can increase their antioxidant activity, whereas extensive hydrolysis can decrease it.     

Changes of lipids in sardine (Sardinella gibbosa) muscle during iced storage

Changes in lipids of sardine (Sardinella gibbosa) muscle during 15 days of iced storage were investigated. Lipid deterioration, lipolysis and lipid oxidation, occurred throughout the storage. The progressive peroxide formation was monitored by the increase in the absorbance band at 3600–3200 cm⁻¹ in Fourier transform infrared (FTIR) spectra and increased peroxide values were observed in sardine muscle up to 6 days of iced storage, followed by a continuous decrease from then for 9 days (P < 0.05). The increase in thiobarbituric acid reactive substances (TBARS) was noticeable throughout the iced storage (P < 0.05). However, no difference in conjugated diene (CD) of sardine muscle was found within the first 12 days of iced storage (P > 0.05). Marked decreases in unsaturated fatty acids, especially eicosapentaenoic acid (EPA; C20:5(n − 3)) and docosahexaenoic acid (DHA; C22:6(n − 3)) were observed as the storage time increased. Those changes indicated that lipid oxidation occurred in sardine muscle. A gradual increase in free fatty acid formation, with decreases in triglyceride and phospholipid contents, was found during iced storage (P < 0.05), suggesting hydrolysis induced by lipases and phospholipases.     

Physicochemical characteristics,proximate analysis and mineral composition of ostrich meat as influenced by muscle

The influence of muscle on the physicochemical characteristics, proximate analysis, and mineral composition of meat from 10 ostriches (10–12 months old), slaughtered according to commercial abattoir procedures, were evaluated. Muscle had no influence (p > 0.05) on L*-values (32.5), a*-values (11.9), water-holding capacity (11.9%), final pH (pH₂₄) values (6.07), and ash contents (1.12 g/100 g edible meat). However, intramuscular lipid contents varied (p < 0.05) from 0.88 (M. fibularis longus) to 1.44 (M. flexor cruris lateralis) g/100 g edible meat, at a mean value of 1.16 g/100 g edible meat for 10 different muscles. Sodium (34.7 mg/100 g edible meat) and iron (3.14 mg/100 g edible meat) contents, both influenced (p < 0.05) by muscle, possessed substantially lower and higher values, respectively, than values reported for beef and chicken.     

Isolation and characterization of collagen from the cartilages of brownbanded bamboo shark (Chiloscyllium punctatum) and blacktip shark (Carcharhinus limbatus)

Acid soluble collagen (ASC) and pepsin soluble collagen (PSC) from the cartilages of brownbanded bamboo shark (BBS; Chiloscyllium punctatum) and blacktip shark (BTS; Carcharhinus limbatus) were isolated and characterized. ASC and PSC extracted from BBS cartilage showed the yields of 1.27 and 9.59 g/100 g (Based on dry weight), respectively, while yields of ASC and PSC from BTS cartilage were 1.04 and 10.30 g/100 g (based on dry weight), respectively. All collagens had protein as a major constituent with the trace amount of ash and fat. They contained glycine as the major amino acid with high contents of alanine, proline and hydroxyproline. Based on sodium dodecyl sulfate-polyacrylamide gel electrophoretic patterns and subunit compositions, all collagens more likely comprised 2 types of collagen, type I and II, and contained α- and β-chains as the major components. Peptide maps of those collagens from both species digested by V8-protease and lysyl endopeptidase were different and were completely different from those of type I collagen from calf skin. Thermal transition temperature of ASC from those collagens (36.28-36.73 °C) was slightly higher than their corresponding PSC (34.56-35.98 °C). From zeta potential analysis, isoelectric points (pI) of collagen from the cartilages of BBS and BTS were estimated to be from 6.53 to 7.03 and from 6.96 to 7.26, respectively. Fourier transform infrared (FTIR) spectra of both ASC and PSC were quite similar, suggesting that pepsin hydrolysis did not affect the secondary structure of collagen, especially triple-helical structure.     

Characterization of acid-soluble collagen from the skin of walleye pollock (Theragra chalcogramma)

Acid-soluble collagen (ASC) was extracted from the skin of walleye pollock (Theragra chalcogramma) and partially characterized. It exhibited a maximum absorbance at 220 nm, but little absorbance near to 280 nm. Amino acid composition and SDS-PAGE suggested that the collagen might be classified as type I collagen. Moreover, FTIR investigations showed the existence of helical arrangements of collagen. The denaturation temperature (T_d) and shrinkage temperature (T_s) were 24.6 °C and 47 °C, respectively, both lower than those of mammalian collagens. However, T_d of walleye pollock skin collagen was higher than that of cod skin collagen reported previously. These results indicate that walleye pollock skin is a potential source of collagen and provide the theoretical basis for further research.