Effects of Bioactive Peptides from Atlantic Salmon Processing By-Products on Oxyntopeptic and Enteroendocrine Cells of the Gastric Mucosa of European Seabass and Gilthead Seabream.

The present study was designed to evaluate the effects of dietary levels of bioactive peptides (BPs) derived from salmon processing by-products on the presence and distribution of peptic cells (oxyntopeptic cells, OPs) and enteric endocrine cells (EECs) that contain GHR, NPY and SOM in the gastric mucosa of European seabass and gilthead seabream. In this study, 27 seabass and 27 seabreams were divided into three experimental groups: a control group (CTR) fed a control diet and two groups fed different levels of BP to replace fishmeal: 5% BP (BP5%) and 10% BP (BP10%). The stomach of each fish was sampled and processed for immunohistochemistry. Some SOM, NPY and GHR-IR cells exhibited alternating "open type" and "closed type" EECs morphologies. The BP10% group (16.8 ± 7.5) showed an increase in the number of NPY-IR cells compared to CTR (CTR 8.5 ± 4.8) and BP5% (BP10% vs. CTR p ≤ 0.01; BP10% vs. BP5% p ≤ 0.05) in the seabream gastric mucosa. In addition, in seabream gastric tissue, SOM-IR cells in the BP 10% diet (16.8 ± 3.5) were different from those in CTR (12.5 ± 5) (CTR vs. BP 10% p ≤ 0.05) and BP 5% (12.9 ± 2.5) (BP 5% vs. BP 10% p ≤ 0.01). EEC SOM-IR cells increased at 10% BP (5.3 ± 0.7) compared to 5% BP (4.4 ± 0.8) (5% BP vs. 10% BP p ≤ 0.05) in seabass. The results obtained may provide a good basis for a better understanding of the potential of salmon BPs as feed ingredients for seabass and seabream.

Mineral Bioaccessibility and Antioxidant Capacity of Protein Hydrolysates from Salmon (Salmo salar) and Mackerel (Scomber scombrus) Backbones and Heads.

Information on the bioaccessibility of minerals is essential to consider a food ingredient as a potential mineral fortifier. In this study, the mineral bioaccessibility of protein hydrolysates from salmon (Salmo salar) and mackerel (Scomber scombrus) backbones and heads was evaluated. For this purpose, the hydrolysates were submitted to simulated gastrointestinal digestion (INFOGEST method), and the mineral content was analyzed before and after the digestive process. Ca, Mg, P, Fe, Zn, and Se were then determined using an inductively coupled plasma spectrometer mass detector (ICP-MS). The highest bioaccessibility of minerals was found in salmon and mackerel head hydrolysates for Fe (≥100%), followed by Se in salmon backbone hydrolysates (95%). The antioxidant capacity of all protein hydrolysate samples, which was measured by Trolox Equivalent Antioxidant Capacity (TEAC), increased (10-46%) after in vitro digestion. The heavy metals As, Hg, Cd, and Pb were determined (ICP-MS) in the raw hydrolysates to confirm the harmlessness of these products. Except for Cd in mackerel hydrolysates, all toxic elements were below the legislation levels for fish commodities. These results suggest the possibility of using protein hydrolysates from salmon and mackerel backbones and heads for food mineral fortification, as well as the need to verify their safety.

Nutritional and Sensory Properties of Protein Hydrolysates Based on Salmon (Salmo salar), Mackerel (Scomber scombrus), and Herring (Clupea harengus) Heads and Backbones.

Protein hydrolysates based on salmon, mackerel, and herring heads and backbones were produced, and the sensory properties of the hydrolysates were evaluated by a highly trained sensory panel. The nutritional content of the products was evaluated, and the hydrolysates contained all the amino acids inherent to the raw material, including considerable levels of connective tissue amino acids glycine, proline, and hydroxyproline. Hydrolysates based on herring were the most flavor intense, whereas hydrolysates based on salmon were deemed more palatable. In this work, choice of fraction (heads vs. backbones) and enzyme had minor effects on sensory and nutritional properties, indicating that choice of raw material species was the major factor for flavor development in the produced protein hydrolysates. There were large variations in protein content and amino acid composition in the raw material fractions, but as expected, only minor variations were found in the final products.

Reduction in flavor-intense components in fish protein hydrolysates by membrane filtration.

Enzymatic protein hydrolysates based on side stream materials from the fish-filleting industry are increasingly explored as food ingredients. However, intense sensory properties, and high salt contents, are often a limiting factor. Most of the sensory attributes, such as fish flavor and salty taste, can be ascribed to low-molecular-weight, water-soluble components, whereas bitterness is associated with small hydrophobic peptides. In this study, protein hydrolysates based on head and backbone residuals from Atlantic salmon (Salmo salar) and Atlantic cod (Gadus morhua) were produced using two different enzymes. The effects of micro- and nanofiltration on the chemical composition, protein recovery, and sensory properties of the final products were investigated. The choice of raw material and enzyme had negligible effects, whereas nanofiltration caused a considerable reduction in metabolites, ash, and the intensity of several sensory attributes. The intensity of bitterness increased after nanofiltration, indicating that small peptides associated with bitter taste were retained by the membrane. Total protein yield after microfiltration was 24%-29%, whereas 19%-24% were recovered in the nanofiltration retentate. PRACTICAL APPLICATION: Enzymatic protein hydrolysates can be included in food products to increase the protein content, and as a nutritional supplement and/or functional ingredient; however, unpalatable and intense flavors limit applications. This study investigated the use of membrane filtration to improve flavor quality and reduce salt content in fish protein hydrolysates. Although some protein loss is unavoidable in micro- and nanofiltration, this study demonstrates the production of fish protein hydrolysates with >90% protein and peptide content, which is suitable for inclusion in foods.

Collagen-Containing Fish Sidestream-Derived Protein Hydrolysates Support Skin Repair via Chemokine Induction.

Restoring homeostasis following tissue damage requires a dynamic and tightly orchestrated sequence of molecular and cellular events that ensure repair and healing. It is well established that nutrition directly affects skin homeostasis, while malnutrition causes impaired tissue healing. In this study, we utilized fish sidestream-derived protein hydrolysates including fish collagen as dietary supplements, and investigated their effect on the skin repair process using a murine model of cutaneous wound healing. We explored potential differences in wound closure and histological morphology between diet groups, and analyzed the expression and production of factors that participate in different stages of the repair process. Dietary supplementation with fish sidestream-derived collagen alone (Collagen), or in combination with a protein hydrolysate derived from salmon heads (HSH), resulted in accelerated healing. Chemical analysis of the tested extracts revealed that Collagen had the highest protein content and that HSH contained the great amount of zinc, known to support immune responses. Indeed, tissues from mice fed with collagen-containing supplements exhibited an increase in the expression levels of chemokines, important for the recruitment of immune cells into the damaged wound region. Moreover, expression of a potent angiogenic factor, vascular endothelial growth factor-A (VEGF-A), was elevated followed by enhanced collagen deposition. Our findings suggest that a 5%-supplemented diet with marine collagen-enriched supplements promotes tissue repair in the model of cutaneous wound healing, proposing a novel health-promoting use of fish sidestreams.

Cytoprotective Effects of Fish Protein Hydrolysates against H2O2-Induced Oxidative Stress and Mycotoxins in Caco-2/TC7 Cells.

Many studies report the potent antioxidant capacity for fish protein hydrolysates, including radical scavenging activity and inhibition ability on lipid peroxidation (LPO). In this study, the in vitro cytotoxicity of protein hydrolysates from different salmon, mackerel, and herring side streams fractions was evaluated in the concentration range from 1 to 1:32 dilution, using cloned human colon adenocarcinoma cells TC7 (Caco-2/TC7) by MTT and PT assays. The protein hydrolysates' antioxidant capacity and oxidative stress effects were evaluated by LPO and reactive oxygen species (ROS) generation, respectively. The antioxidant capacity for pure and bioavailable hydrolysate fraction was also evaluated and compared. Additionally, mycotoxin levels were determined in the fish protein hydrolysates, and their cytoprotective effect against T-2 toxin was evaluated. Both hydrolysates and their bioavailable fraction induced similar cell viability rates. The highest cytoprotective effect was obtained for the salmon viscera protein hydrolysate (HSV), which increased the cell viability by 51.2%. ROS accumulation induced by H2O2 and LPO was suppressed by all pure hydrolysates. The cytoprotective effect of hydrolysates was observed against T-2. Moreover, the different fish fraction protein hydrolysates contain variable nutrients and unique bioactive peptide composition showing variable bioactivity, which could be a useful tool in developing dietary supplements with different target functional properties.

Fish Sidestream-Derived Protein Hydrolysates Suppress DSS-Induced Colitis by Modulating Intestinal Inflammation in Mice.

Inflammatory bowel disease is characterized by extensive intestinal inflammation, and therapies against the disease target suppression of the inflammatory cascade. Nutrition has been closely linked to the development and suppression of inflammatory bowel disease, which to a large extent is attributed to the complex immunomodulatory properties of nutrients. Diets containing fish have been suggested to promote health and suppress inflammatory diseases. Even though most of the health-promoting properties of fish-derived nutrients are attributed to fish oil, the potential health-promoting properties of fish protein have not been investigated. Fish sidestreams contain large amounts of proteins, currently unexploited, with potential anti-inflammatory properties, and may possess additional benefits through bioactive peptides and free amino acids. In this project, we utilized fish protein hydrolysates, based on mackerel and salmon heads and backbones, as well as flounder skin collagen. Mice fed with a diet supplemented with different fish sidestream-derived protein hydrolysates (5% w/w) were exposed to the model of DSS-induced colitis. The results show that dietary supplements containing protein hydrolysates from salmon heads suppressed chemically-induced colitis development as determined by colon length and pro-inflammatory cytokine production. To evaluate colitis severity, we measured the expression of different pro-inflammatory cytokines and chemokines and found that the same supplement suppressed the pro-inflammatory cytokines IL-6 and TNFα and the chemokines Cxcl1 and Ccl3. We also assessed the levels of the anti-inflammatory cytokines IL-10 and Tgfb and found that selected protein hydrolysates induced their expression. Our findings demonstrate that protein hydrolysates derived from fish sidestreams possess anti-inflammatory properties in the model of DSS-induced colitis, providing a novel underexplored source of health-promoting dietary supplements.

Emulsion and Surface-Active Properties of Fish Solubles Based on Direct Extraction and after Hydrolysis of Atlantic Cod and Atlantic Salmon Backbones.

The focus on natural foods and "clean" labeled products is increasing and encourages development of new biobased ingredients. Fish solubles derived from downstream processing of side stream materials in the fish filleting industries have potential as emulsifiers based on their surface-active and emulsion stabilizing properties. The aim of this study was to evaluate and compare emulsion properties and critical micelle concentration (CMC) of direct protein extracts and protein hydrolysates based on fish backbones, and to identify associations between molecular weight distribution and process yield with the studied physicochemical properties. Protein extracts and enzymatic protein hydrolysates were produced based on two raw materials (cod and salmon backbones), two enzymes with different proteolytic specificity, and varying hydrolysis time. Emulsion activity index (EAI), emulsion stability index (ESI) and CMC were measured and compared with casein as a reference to protein-based emulsifiers. Protein hydrolysis was found to have negative impact on EAI and CMC, likely due to generation of small peptides disrupting the amphiphilic balance. The direct protein extracts had comparable EAI with casein, but the latter had superior ESI values. Protein hydrolysates with acceptable EAI could only be obtained at the expense of product yield. The study emphasizes the complexity of physicochemical properties of protein hydrolysates and discusses the challenges of achieving both good surface-active properties and high product yield.

Sensory Assessment of Fish and Chicken Protein Hydrolysates. Evaluation of NMR Metabolomics Profiling as a New Prediction Tool.

Nuclear magnetic resonance (NMR) metabolomics profiling was evaluated as a new tool in sensory assessment of protein hydrolysates. Hydrolysates were produced on the basis of different raw materials (cod, salmon, and chicken), enzymes (Food Pro PNL and Bromelain), and hydrolysis time (10 and 50 min). The influence of raw material and hydrolysis parameters on sensory attributes was determined by traditional descriptive sensory analysis and 1H NMR spectroscopy. The raw material had a major influence on the attribute intensity and metabolite variation, followed by enzyme and hydrolysis time. However, the formation of bitter taste was not affected by the raw material. Partial least-squares regression (PLSR) on 1H NMR and sensory data provided good models (Q2 = 0.55-0.89) for 11 of the 17 evaluated attributes, including bitterness. Significant metabolite-attribute associations were identified. The study confirms the potential prediction of the sensory properties of protein hydrolysates from cod, salmon, and chicken based on 1H NMR metabolomics profiling.

Valorization of Proteins from Co- and By-Products from the Fish and Meat Industry.

Large volumes of protein-rich residual raw materials, such as heads, bones, carcasses, blood, skin, viscera, hooves and feathers, are created as a result of processing of animals from fisheries, aquaculture, livestock and poultry sectors. These residuals contain proteins and other essential nutrients with potentially bioactive properties, eligible for recycling and upgrading for higher-value products, e.g. for human, pet food and feed purposes. Here, we aim to cover all the important aspects of achieving optimal utilization of proteins in such residual raw materials, identifying those eligible for human consumption as co-products and for feed applications as by-products. Strict legislation regulates the utilization of various animal-based co- and by-products, representing a major hurdle if not addressed properly. Thorough understanding and optimization of all parts of the production chain, including conservation and processing, are important prerequisites for successful upgrading and industrial implementation of such products. This review includes industrially applied technologies such as freezing/cooling, acid preservation, salting, rendering and protein hydrolysis. In this regard, it is important to achieve stable production and quality through all the steps in the manufacturing chain, preferably supported by at- or online quality control points in the actual processing step. If aiming for the human market, knowledge of consumer trends and awareness are important for production and successful introduction of new products and ingredients.

A Systematic Approach to the Comparison of Cost Efficiency of Endopeptidases for the Hydrolysis of Atlantic Salmon (Salmo salar) By-Products.

The hydrolytic and cost efficiencies of five endopeptidases (Alcalase 2.4L, Corolase 7089, Neutrase 0.8L, Promod 671L and Protex 7L) to hydrolyze Atlantic salmon by-products were compared at standardized activity levels based on a casein assay. The substrate was characterized prior to the hydrolytic experiments (pH=6.5, t=50 °C) to obtain substrate--specific constants for nitrogen to protein mass (in g) ratio, i.e. conversion factor fN=5.23 and total amount of peptide bonds (htot)=9.3 mmol per g of protein. At low enzyme activity to substrate ratio, all enzymes were equally efficient in hydrolyzing the substrate. At highest enzyme activity to substrate ratio, Protex 7L, Alcalase 2.4L and Promod 671L gave higher degree of hydrolysis (DH=14.2-14.6%) than Corolase 7089 (13.2%) and Neutrase 0.8L (11.6%) after 120 min of hydrolysis. No differences were observed in protein recovery (yield of solubilized protein) relative to DH. Determination of DH was followed by the pH-STAT and o-phthaldialdehyde methods. Based on pH-STAT data, response surface regression models were established based on the combined effects of hydrolysis time and enzyme activity to substrate ratio on DH and protein recovery. The modelling approach was combined with enzyme cost to identify the most cost-efficient enzyme (Protex 7L).