Selection of texture-associated biomarkers in chilled and iced grass carp (Ctenopharyngodon idella) fillets via DIA-based proteomics.

Consumers care about the texture of fresh fish flesh, but a rapid quantitative analytical method for this has not been properly established. In this study, texture-associated biomarkers were selected by DIA-based proteomics for possible future application. Results indicated a significant decline in texture and moisture characteristics with extended storage under chilled and iced conditions, and flesh quality was categorized into three intervals. A total of 8 texture-associated biomarkers were identified in the chilled storage group, and 3 distinct ones in the iced storage group. Biomarkers were further refined based on their expression levels. Isobutyryl-CoA dehydrogenase, mitochondrial and [Phosphatase 2A protein]-leucine-carboxy methyltransferase were identified as effective texture-associated biomarkers for chilled fish, and Staphylococcal nuclease domain-containing protein 1 for iced fish. This study provided suitable proteins as indicators of fresh fish flesh texture, which could help establish a rapid and convenient texture testing method in future studies.

Effects of different protein cross-linking degrees on physicochemical and subsequent thermal gelling properties of silver carp myofibrillar proteins sol subjected to freeze-thaw cycles.

Knowledge regarding the denaturation process and control methods for depolymerized sol-state myofibrillar proteins (MPs) during freezing remains scant. This study investigated the effects of protein cross-linking treatment before freezing on physicochemical and subsequent gelation properties of MPs sol subjected to freeze-thaw (F-T) cycles. Results indicated that after five F-T cycles, cross-linked MPs sols showed increased high molecular weight polymers and bound water (T21a and T21b) mobility, suggesting enhanced protein-protein interactions at the expense of protein-water interactions. Upon heating after F-T cycles, gels formed from cross-linked sols exhibited significantly higher hardness, springiness, and cooking loss (P < 0.05), alongside more contracted gel networks. Correlation analysis revealed that the formation and properties of thermal gel after freezing closely relate to changes in molecular conformation and chemical bonds of cross-linked MPs sol during freezing. This study provides new insights into regulating the freezing stability and post-thawed thermal processing properties of sol-based surimi products.

Uncovering quality changes of surimi-sol based products subjected to freeze-thaw process: The potential role of oxidative modification on salt-dissolved myofibrillar protein aggregation and gelling properties.

Frozen surimi quality generally correlates with oxidation, but impacts of protein oxidation on salt-dissolved myofibrillar protein (MP) sol in surimi remain unclear. Hence, physicochemical and gelling properties of MP sol with different oxidation degrees were investigated subjected to freeze-thaw cycles. Results showed that mild oxidation (≤1 mmol/L) improved unfrozen MP gel quality with lowest cooking loss (3.29 %) and highest hardness (829.76 N). Whereas, oxidized sol suffering freeze-thawing degenerated severely, showing reduction of 23.85 % of salt soluble protein contents with H2O2 concentrations of 10 mmol/L. Shearing before heating influenced gelling properties of freeze-thawed sol, depending on oxidation levels. Oxidized gel with shearing displayed disorganized network structures, whereas gel without shearing displayed relatively complete appearances without holes under high oxidation condition (10 mmol/L). Overall, freeze-thaw process aggravated denaturation extents of MP sol subjected to oxidation, further causing high water loss and yellow color of heat-induced gel, especially to gel with shearing.

Effect of sugar reduction on sensory characteristics of dried fish mince product.

BACKGROUND: Excessive sugar consumption has been linked to type 2 diabetes, obesity and other diseases. Therefore, it is indispensable to reduce sugar of food. However, the sensory characteristics of food are affected after sugar reduction (SR). Currently, SR has been reported in drinks, jams, candies, and other fruit related or sweet foods; but salty or protein related foods have not been explored, therefore there is a big gap that needs to be filled. RESULTS: Sensory scores of initial sweetness and sweetness reduced by 0.26 and 0.12 in 10% SR dried fish mince product compared with control, and there was a small difference between 25% SR (3.33) and 40% SR (3.09) samples. It also showed that 10% SR sample had a small reduction in sweetness value and free sugar content by 3.5% (0.42/11.9) and 7.8% (2.12/27.06) compared with control; while values in 25% SR sample decreased sharply but were not much different from 40% SR sample. Electronic nose results showed that SR had a small effect on odor. Texture analysis showed that texture properties of 25% SR sample were significantly different from control. CONCLUSIONS: Dried fish mince product with below 10% SR had a small difference on the sensory characteristics and there was a big change when SR was more than 20%. Dried fish mince product with 25% SR and 40% SR had no significant difference. SR had a small effect on odor, but had a great effect on texture properties, especially over 20% SR. © 2023 Society of Chemical Industry.

Neuroprotective effect of chitosan oligosaccharide on alcohol-induced hippocampal injury using proteomic analysis.

Alcoholism is a serious public health problem, and the abuse of drinking seriously damages the health of people. Chitosan oligosaccharides (COSs) are small-molecule oligosaccharides with amino groups that have many unique properties. The neuroprotective effect of COS on alcohol-induced hippocampal injury in Sprague-Dawley (SD) rats was investigated. The discrimination ratio of the COS group in the Y-maze experiment was 59.3% higher than that of the ETOH group. Meanwhile, the discrimination index was less than 0 in the ETOH group but greater than 0 in the COS group during the object recognition test. The cells in the COS group were more tightly arranged than those in the ETOH group. Proteomics was used to identify differentially expressed proteins in the hippocampus. There were 27 differentially expressed proteins in the COS and ETOH group for further bioinformatic analysis. There are three enriched pathway categories, namely, primary immunodeficiency, hedgehog signaling, and sulfur relay system. Next, sonic hedgehog signaling pathway-related proteins were verified through western blotting. The protein expression level of ß-arrestin-2 in the COS group was 2.85 times higher than that in the ETOH group. This work may contribute to understanding the underlying mechanism of the neuroprotective effect of COS against alcohol-induced hippocampal injury in SD rats.

Comparison of the flavor qualities between two varieties of Mercenaria mercenaria.

The saltwater hard clam Mercenaria mercenaria (M. mercenaria) as a representative of low-value shellfish, enhancing its flavor quality, is the key to enter the high-end market. Nevertheless, there has not been reported research on the flavor quality of M. mercenaria. This study compared the flavor quality of selective and non-selective saltwater hard clams of M. mercenaria by using various indicators: proximate component, free amino acids, nucleotides, and metabolomic analysis. The results indicated that selective breeding contributed to the significant improvement contents of crude protein, flavor-associated free amino acids (glutamic acid, aspartic acid, proline, etc.), and nucleotides (AMP) (P < 0.05). Then, the metabolome was utilized to assess the metabolite changes in the pre/post-selective breeding of M. mercenaria and further understand the flavor characteristics and metabolic status. In the metabolomics assay, among the 3143 quantified metabolites, a total of 102 peaks were identified as significantly different metabolites (SDMs) between the selective and non-selective varieties of M. mercenaria (VIP > 1 and P < 0.05). These results can provide new insights for future research on improving the quality of saltwater bivalves through selective breeding.

The inhibition mechanism of nanoparticles-loading bilayer film on texture deterioration of refrigerated carp fillets from the perspective of protein changes and exudates.

Herein, the protective pattern of bilayer film on the texture stability of fillets was discussed in terms of endogenous enzyme activity, as well as protein oxidation and degradation. The texture properties of fillets wrapped with nanoparticles (NPs) bilayer film were greatly improved. NPs film delayed protein oxidation by inhibiting the formation of disulfide bond and carbonyl group as evidenced by the increase of α-helix ratio (43.02%) and the decrease of random coil ratio (15.87%). The protein degradation degree of fillets treated with NPs film was lower than that of control group, specifically with a more regular protein structure. The exudates accelerated the degradation of protein, while NPs film effectively absorbed exudates to delay protein degradation. Overall, the active agents in the film were released into the fillets to play an antioxidant and antibacterial roles, and the inner layer of film could absorb exudates, thus maintaining the texture characteristics of fillets.

The migration and loss of water in emulsified surimi gels prepared with different phase states of lipids: Effect of freeze-thawing treatments.

The freeze-thawing (FT) stability generally correlates well with the economic value and acceptability of frozen surimi-based products. However, quality changes of emulsified surimi gels under FT conditions are still unclear. Therefore, the gel properties of samples with different phase states of lipids (lard, lard + soybean oil, and soybean oil) were investigated at FT conditions. Results showed that the soybean oil evidently improved the rheological behaviors of sols/gels compared to the lard group. The moisture content of samples with different lipids decreased by 2.40%-2.71% after 4 FT cycles. With increasing FT cycles, the water-holding capacity decreased accompanied by the increase of cooking loss. Spin-spin relaxation spectra and hydrogen proton density images proved the occurrence of water migration of gels during these processes. Better gel integrity was observed in samples consisting of soybean oil, where the proportion of pores was lower than those with lard regardless of FT treatments. Additionally, the intermolecular forces of gels also changed under FT treatments. There results suggested that the lipids with different phase states affected the migration and loss of water in emulsified surimi gels under FT cycles. PRACTICAL APPLICATION: The quality changes of heating-induced surimi gel products under frozen storage have been ignored, especially the emulsified surimi gels. This study discloses the changes of the gel properties in emulsified gel products with different phase states of lipids after FT treatments, which provides critical insights into the quality improvement of this novel emulsified surimi product during processing, storage, and transportation.

Influence of the Enzymatic Hydrolysis Using Flavourzyme Enzyme on Functional, Secondary Structure, and Antioxidant Characteristics of Protein Hydrolysates Produced from Bighead Carp (Hypophthalmichthys nobilis).

In the current study, bighead carp fish were used in conjunction with the flavourzyme enzyme to obtain (FPH) fish protein hydrolysates. The optimum conditions of the hydrolysis process included an enzyme/substrate ratio of 4% and a temperature of 50 °C and pH of 6.5. The hydrolysis time was studied and investigated at 1, 3, and 6 h, and the (DH) degree of hydrolysis was recorded at 16.56%, 22.23%, and 25.48%, respectively. The greatest yield value was 17.83% at DH 25.48%. By increasing the DH up to 25.48%, the crude protein and total amino acid composition of the hydrolysate were 88.19% and 86.03%, respectively. Moreover, more peptides with low molecular weight were formed during hydrolysis, which could enhance the functional properties of FPH, particularly the solubility property ranging from 85% to 97%. FTIR analysis revealed that enzymatic hydrolysis impacted the protein's secondary structure, as indicated by a remarkable wavelength of amide bands. Additionally, antioxidant activities were investigated and showed high activity of DDPH radical scavenging, and hydroxyl radical scavenging demonstrated remarkable activity. The current findings demonstrate that the functional, structural, and antioxidant characteristics of FPH might make it an excellent source of protein and suggest potential applications in the food industry.

Research on apoptotic mechanism and related pathways involved in postmortem grass carp (Ctenopharyngodon idellus) muscle.

BACKGROUND: Apoptosis activation is an essential research to reveal the triggering mechanism of flesh quality deterioration. This study was aimed at explaining apoptotic mechanism of postmortem fish in terms of caspases activation, cytochrome c (cyt-c) release, B-cell lymphoma 2 (Bcl-2) and Bcl2-associated X (Bax) protein levels, transcriptional levels of its molecules, and apoptosis-inducing factor (AIF) translocation at 4 °C for 5 days. RESULTS: Activation of caspase-9, caspase-8, caspase-3 and the release of mitochondrial cyt-c were observed during storage. The decreased Bcl-2 protein levels, increased Bax protein expressions and Bax/Bcl-2 ratio were major steps for inducing apoptosis. Collectively, transcriptional regulation of Fas ligand (FasL), apoptotic protease activating factor-1 (Apaf-1), inhibitors of apoptosis proteins (IAPs), myeloid cell leukemia-1 (Mcl-1), c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinase (MAPK) indicated that extrinsic apoptotic pathways (FasL/caspase-8/caspase-3) and intrinsic pathway [(JNK and p38 MAPK)/(Bcl-2, Bax and Mcl-1)/cyt-c/Apaf-1/caspase-9/caspase-3] were involved in apoptotic process. Mitochondrial AIF translocation to nuclear indicated that AIF mediated caspase-independent pathway. CONCLUSION: Therefore, transcriptional and translational alterations of multiple signaling molecules acted important roles in regulating apoptosis activation in postmortem process. © 2022 Society of Chemical Industry.

The protective pattern of chitosan-based active coating on texture stabilization of refrigerated carp fillets from the perspective of proteolysis.

Herein, the protective pattern of chitosan coating enriched with green tea extract on texture stabilization of refrigerated grass carp fillets was explored. In general, higher shear force and lower endogenous enzyme activities were observed in coated fillets, with the max. level of cathepsins and calpain decreasing by 30.2 âˆ¼ 39.6 % when compared to the control during storage. Meanwhile, the coating reduced protein composition changes and accumulation of protein degradation products. According to label-free proteomic analysis, the proteome closer to fresh sample was observed in coated fillets than that of the control, as supported by PCA and hierarchical cluster analysis. Meanwhile, 33 differential proteins involved in tissue structure, protein phosphorylation and protein turnover were further screened out, and most DAPs showed up-regulation in coated fillets compared to the control on day 12. Presumably, the coating modulated endogenous enzyme-induced myofibrillar protein degradation and protein phosphorylation level, thereby stabilizing the texture properties of refrigerated fillets.

Effects of actomyosin dissociation on the physicochemical and gelling properties of silver carp myofibrillar protein sol during freeze-thaw cycles.

Myofibrillar protein (MP) system with different dissociation degrees of actomyosin was constructed by addition of ATP and the effects of actomyosin dissociation on the physicochemical and gelling properties of MP sol during freeze-thaw cycles were investigated. The results showed that the salt soluble protein content of the dissociated sample with 5 mM ATP (5-pH 6.2 group) was lower than that of other groups under unfrozen state, while the fluorescence intensity and hardness of all dissociated groups were significantly lower (P＜0.05) than that of the control group. After five freeze-thaw cycles, the low-field nuclear magnetic resonance (LF NMR) revealed that the immobile water (T22b) in MP sol with 5 mM ATP (pH 6.9) has lower fluidity compared with the control group. In addition, rheological studies revealed that the G' (storage modulus) value at 90℃ of MP sol with 5 mM ATP (pH 6.9) showed less decrease than that of the other groups after five freeze-thaw cycles. Meanwhile, after five freeze-thaw cycles, scanning electron microscope (SEM) showed that the gel of the control group has large holes and rough structure, while the microstructure of sample with 5 mM ATP (pH 6.9) was more compact and uniform. The heat-induced gel of 5-pH 6.9 group had highest hardness and lowest cooking loss among the groups after five freeze-thaw cycles. Briefly, the dissociation of actomyosin before freezing could slow down the rate of MP denaturation and improve the gelling properties after freeze-thaw cycle.

Effects of deheading and rinsing pretreatment on the quality of white leg shrimp (Litopenaeus vannamei) surimi based on endogenous proteases.

In this study, the effects of different pretreatments on the quality of white leg shrimp surimi were investigated based on shrimp endogenous proteases. The results showed that removing the head and rinsing significantly (P < 0.05) improved the gel strength, texture, whiteness, water distribution, and microstructure of the shrimp surimi gels. Headless shrimp surimi (HSS) had higher salt-soluble protein and lower water-soluble protein than whole shrimp surimi (WSS). The shrimp heads had high cathepsin B, L, D, and serine protease activities. Electrophoretic analysis revealed significant degradation of the myofibrillar proteins in the WSS during cold storage and thermal gelation. Moreover, the myosin heavy chains almost disappeared after thermal gelation, and new bands appeared at about 270 kDa and 100 kDa. However, rinsing reduced the endogenous proteases, water-soluble proteins, and concentrated salt-soluble proteins in the shrimp surimi; thus, the quality of the shrimp surimi gel improved after rinsing. These results suggest that the quality of the surimi gel was damaged by the endogenous proteases, and that removal of the shrimp heads and rinsing significantly (P < 0.05) improved the quality of the shrimp surimi gel. The gel properties of WSS were similar after the second rinse to those of unrinsed HSS. The choice of headless shrimp or whole shrimp as the raw material for production needs to be comprehensively considered according to the planned cost and the quality required for the shrimp surimi product. The recommended number of rinses is 1-2.

Label-free quantification proteomics reveals potential proteins associated with the freshness status of crayfish (Procambarus clarkii) as affected by cooking.

In order to characterize the freshness status of raw material of cooked crayfish products, protein and physicochemical changes were investigated in raw and cooked crayfish from fresh-live state to the early postmortem using Label-free quantification proteomics, total volatile basic nitrogen (TVB-N) and biogenic amines (BAs) analysis. Results showed that the TVB-N and BAs contents in both raw and cooked crayfish remained low levels and no obvious changes were observed within 24 h of postmortem storage at 4 °C. Altogether, six differentially expressed proteins (DEPs) were detected in both C6/C0 and C24/C0 groups by proteomics. Furthermore, five among the six DEPs were verified both in cooked and raw crayfish samples by parallel reaction monitoring. Among the six DEPs, "Unigene990_S_0_Gene.8754" changed most significantly with the increasing of postmortem time and not obviously influenced by heating, indicating that "Unigene990_S_0_Gene.8754" may be highly associated to the freshness status of crayfish.

Effect of starter cultures and spices on physicochemical properties and microbial communities of fermented fish (Suanyu) after fermentation and storage.

Suanyu is one of the traditional Chinese fermented fish products with a pleasant flavor. The present study sought to evaluate the effects of starter cultures and spices on the physicochemical properties and microbial flora of Suanyu after fermentation and storage. The results indicated that fermentation with mixed starter cultures could lower the pH and inhibit TVB-N formation in Suanyu after fermentation and during storage. Moreover, inoculation of starter cultures could promote free amino acids production during storage. Additionally, inoculation of mixed starter cultures and spices mixture could effectively inhibit the formation of most biogenic amines in Suanyu, revealing the significant role of mixed starter cultures. Staphylococcus, Macrococcus, and Lactobacillus were the core microbial genus in Suanyu. The inoculation of starter cultures could inhibit Enteroccoccus and other microorganisms, while spices introduced Bacillus into Suanyu. The study results provide a theoretical basis for the industrial production and storage of Suanyu.

A comparison of eating safety and quality of live and dead freshwater crayfish (Procambarus clarkii) at different stages.

The risk of Procambarus clarkii eating safety attracts consumers' big concern, but it has not been addressed properly. Therefore, this study was aimed to investigate eating safety and quality of live and dead Procambarus clarkii at different stages by total volatile basic nitrogen (TVB-N), biogenic amines (BAs), total aerobic plate counts (TPC) and microbiota. The results showed that in live Procambarus clarkii, TVB-N and TPC values were below the limit despite vitality, while cadaverine in gills, intestines, and glands (GIG) exceeded in articulo-mortis Procambarus clarkii. For the dead, it showed that Procambarus clarkii posed a high risk in eating safety within one to two days after death; and BAs of high risk were putrescine and cadaverine. The dominant microorganisms threatening eating safety and quality were potentially pathogenic bacteria of Citrobacter and Acinetobacter from the environment; and spoilage bacteria of Shewanella from viscera.

Characterization of key aroma compounds in low-salt fermented sour fish by gas chromatography-mass spectrometry, odor activity values, aroma recombination and omission experiments.

In this study, key aroma compounds of low-salt fermented sour fish were characterized using headspace solid-phase micro extraction (HS-SPME) coupled with gas chromatography-mass spectrometry (GC-MS), odor activity values (OAV) and aroma recombination and omission experiments. Eighty-eight volatile compounds, including esters, aldehydes, alcohols, acids, furans and pyrazines, were identified by HS-SPME-GC-MS. Eighteen aroma-active compounds were quantified by employing calculation of OAV greater than 1. A recombination aroma model prepared using aroma-active compounds based on the odorless fish matrix sensorially matched the aroma of fermented sour fish with a score of 4.5 out of 5. The omission experiment showed that 7 out of 18 compounds had a significant contribution to the overall aroma (P < 0.05). The key aroma compounds of fermented sour fish were concluded to be ethyl acetate (OAV = 189), ethyl hexanoate (OAV = 66), isoamyl acetate (OAV = 424), ethyl butyrate (OAV = 26), hexanal (OAV = 49), 1-hexadecanal (OAV = 14) and 2-pentylfuran (OAV = 13).

Effect of the Degree of Hydrolysis on Nutritional, Functional, and Morphological Characteristics of Protein Hydrolysate Produced from Bighead Carp (Hypophthalmichthys nobilis) Using Ficin Enzyme.

The production of fish protein hydrolysates from bighead carp (Hypophthalmichthys nobilis) using ficin enzymes was achieved in optimal conditions of 3% enzyme/substrate ratio, 40 °C temperature, and pH 6. Three different hydrolysis times, 1, 3, and 6 h, were investigated, and their degree of hydrolysis (DH) values were 13.36%, 17.09%, and 20.15%, respectively. The hydrolysate yield values increased with DH increase, and the highest yield was obtained at DH 20.15%. The crude protein content increased from 80.58% to 85.27%, and amino acid compositions increased from 78.33% to 83.07%. The peptides formed during hydrolysis indicated low molecular weight that might improve functional characteristics of fish protein hydrolysates, including protein solubility, which ranged from 84.88% to 95.48% for all hydrolysates. The thermal degradation of hydrolysates occurred from 160 to 168 °C with intensive endothermic peaks. Results revealed that oil holding capacity was higher at DH 13.36%; water holding capacity was higher when DH increased. Hence, fish protein hydrolysates (FPH) from bighead carp have improved functional properties, and can be utilized as supplements and excellent protein sources in various uses in food applications.

The role of cathepsin L on structural changes of collagen fibers involved in textural deterioration of chilled grass carp (Ctenopharyngodon idella) fillets.

BACKGROUND: Textural deterioration is a serious problem in chilled fish flesh. Cysteine proteinases are proposed to participate in disintegration of collagen fibers during this process, while its mechanism remains elusive. In the present study, a cysteine proteinase was purified from grass carp muscle and identified by mass spectrometry, and its effect on structural changes of collagen fibers was investigated. RESULTS: During storage at 4 °C, cysteine proteinase activity in fillets increased to 1.53-fold at day 5 and maintained a high level later, and this variable was related to a decline in shear force and an increase in drip loss. A 29 kDa cysteine proteinase was purified through ammonium sulfate precipitation and column chromatography, and identified as cathepsin L. Cathepsin L caused collagen fibers to partly disintegrate into fibril bundles and individual fibrils at 48 h, while the triple helical structure of collagen molecules remained stable. Release of soluble proteins and glycosaminoglycans from cathepsin L-treated collagen fibers was time dependent, coinciding with a release of 4.12 ± 0.13% and 8.57 ± 0.03% at 48 h respectively. However, 0.85 ± 0.02% of hydroxyproline was freed from cathepsin L-treated collagen fibers at 48 h. Furthermore, scanning electron microscopy revealed that the inhibitory effect of cathepsin L could retard the destruction of intramuscular connective tissues (IMCTs). CONCLUSION: These results indicated that cathepsin L might be involved in collagen fiber breakdown by degrading collagen-associated proteoglycans during textural deterioration of grass carp. © 2022 Society of Chemical Industry.

Identification of characteristic flavor and microorganisms related to flavor formation in fermented common carp (Cyprinus carpio L.).

Microbial metabolism is crucial for the flavor development of traditional Suanyu. A total of 21 characteristic volatile compounds of Suanyu were identified according to GC-MS combined with GC-O analysis. HCA results indicated that the whole fermentation process could be divided into three stages: raw material (fresh), initial fermentation (0-2 week) and the late fermentation (2-4 week). Moreover, nine flavor markers in different fermentation stages were revealed using the OPLS-DA model and VIP values. Furthermore, metagenomic sequencing was used to analyze the possible formation pathways of the characteristic flavor compounds and identify the key microorganisms related to flavor formation at the gene level. The top 10 bacterial genera associated with flavor formation were Staphylococcus, Enterococcus, Vibrio, Kocuria, Lactobacillus, Macrococcus, Streptococcus, Enterobacter, Lactococcus and Pediococcus, accounting for large parts of flavor formation of fermented Suanyu. This study is helpful to elucidate the different metabolic roles of microorganisms in flavor formation during fermentation.