Muscle Protein Hydrolysates and Amino Acid Composition in Fish

Fish muscle, which accounts for 15%–25% of the total protein in fish, is a desirable protein source. Their hydrolysate is in high demand nutritionally as a functional food and thus has high potential added value. The hydrolysate contains physiologically active amino acids and various essential nutrients, the contents of which depend on the source of protein, protease, hydrolysis method, hydrolysis conditions, and degree of hydrolysis. Therefore, it can be utilized for various industrial applications including use in nutraceuticals and pharmaceuticals to help improve the health of humans. This review discusses muscle protein hydrolysates generated from the muscles of various fish species, as well as their amino acid composition, and highlights their functional properties and bioactivity. In addition, the role of the amino acid profile in regulating the biological and physiological activities, nutrition, and bitter taste of hydrolysates is discussed.     

Amino Acid Profiles and Biopotentiality of Hydrolysates Obtained from Comb Penshell (Atrina pectinata) Viscera Using Subcritical Water Hydrolysis

The recovery of amino acids and other important bioactive compounds from the comb penshell (Atrina pectinata) using subcritical water hydrolysis was performed. A wide range of extraction temperatures from 140 to 290 °C was used to evaluate the release of proteins and amino acids. The amount of crude protein was the highest (36.14 ± 1.39 mg bovine serum albumin/g) at 200 °C, whereas a further increase in temperature showed the degradation of the crude protein content. The highest amount of amino acids (74.80 mg/g) was at 230 °C, indicating that the temperature range of 170–230 °C is suitable for the extraction of protein-rich compounds using subcritical water hydrolysis. Molecular weights of the peptides obtained from comb penshell viscera decreased with the increasing temperature. SDS-PAGE revealed that the molecular weight of peptides present in the hydrolysates above the 200 °C extraction temperature was ≤ 1000 Da. Radical scavenging activities were analyzed to evaluate the antioxidant activities of the hydrolysates. A. pectinata hydrolysates also showed a particularly good antihypertensive activity, proving that this raw material can be an effective source of amino acids and marine bioactive peptides.     

Sunflower Protein Hydrolysates Reduce Cholesterol Micellar Solubility

Plant protein hydrolysates are a source of bioactive peptides. There are peptides that decrease the micellar cholesterol solubility from bile acids and therefore may reduce in vivo cholesterol absorption. The presence of these peptides in sunflower protein hydrolysates has been studied. Sunflower protein hydrolysates produced with alcalase plus flavourzyme or with pepsin plus pancreatin inhibited in some degree the cholesterol incorporation to micelles. Protein hydrolysates generated after 30 min of hydrolysis with alcalase, and after 30 min of hydrolysis with pepsin, were the inhibitoriest of the cholesterol incorporation to micelles. The average amino acid hydrophobicity of inhibitory peptides in cholesterol micelles was higher than the observed in the corresponding protein hydrolysates. This high hydrophobicity probably favours their inclusion in the lipid micelles. In vivo, this inhibition may translate in a decrease of cholesterol absorption. Reported results show that a combination of different characteristics such as peptide size or hydrophobicity may be responsible of the inhibitory activity of generated peptides.     

In Vitro Bioactivity of Astaxanthin and Peptides from Hydrolisates of Shrimp (Parapenaeus longirostris) By-Products: From the Extraction Process to Biological Effect Evaluation,as Pilot Actions for the Strategy “From Waste to Profit”

Non-edible parts of crustaceans could be a rich source of valuable bioactive compounds such as the carotenoid astaxanthin and peptides, which have well-recognized beneficial effects. These compounds are widely used in nutraceuticals and pharmaceuticals, and their market is rapidly growing, suggesting the need to find alternative sources. The aim of this work was to set up a pilot-scale protocol for the reutilization of by-products of processed shrimp, in order to address the utilization of this valuable biomass for nutraceutical and pharmaceuticals application, through the extraction of astaxanthin-enriched oil and antioxidant-rich protein hydrolysates. Astaxanthin (AST) was obtained using “green extraction methods,” such as using fish oil and different fatty acid ethyl esters as solvents and through supercritical fluid extraction (SFE), whereas bioactive peptides were obtained by protease hydrolysis. Both astaxanthin and bioactive peptides exhibited bioactive properties in vitro in cellular model systems, such as antioxidant and angiotensin I converting enzyme (ACE) inhibitory activities (IA). The results show higher astaxanthin yields in ethyl esters fatty acids (TFA) extraction and significant enrichment by short-path distillation (SPD) up to 114.80 ± 1.23 µg/mL. Peptide fractions of <3 kDa and 3–5 kDa exhibited greater antioxidant activity while the fraction 5–10 kDa exhibited a better ACE-IA. Lower-molecular-weight bioactive peptides and astaxanthin extracted using supercritical fluids showed protective effects against oxidative damage in 142BR and in 3T3 cell lines. These results suggest that “green” extraction methods allow us to obtain high-quality bioactive compounds from large volumes of shrimp waste for nutraceutical and pharmaceutical applications.     

Physicochemical and Antioxidant Properties of Gelatin and Gelatin Hydrolysates Obtained from Extrusion-Pretreated Fish (Oreochromis sp.) Scales

Fish gelatin and its hydrolysates exhibit a variety of biological characteristics, which include antihypertensive and antioxidant properties. In this study, fish gelatins were extracted from extrusion-pretreated tilapia scales, and then subjected to analyses to determine the physicochemical properties and antioxidant activity of the extracted gelatins. Our findings indicate that TSG2 (preconditioned with 1.26% citric acid) possessed the greatest extraction yield, as well as higher antioxidant activities compared with the other extracted gelatins. Hence, TSG2 was subjected to further hydrolyzation using different proteases and ultrafiltration conditions, which yielded four gelatin hydrolysates: TSGH1, TSGH2, TSGH3, and TSGH4. The results showed that TSGH4 (Pepsin + Pancreatin and ultrafiltration < 3000 Da) had a higher yield and greater antioxidant activity in comparison with the other gelatin hydrolysates. As such, TSGH4 was subjected to further fractionation using a Superdex peptide column and two-stage reverse-phase column HPLC chromatography, yielding a subfraction TSGH4-6-2-b, which possessed the highest 2,2-diphenyl-1-picrylhydrazyl (DPPH) scavenging activity compared with the other fractions. Further LC-ESI/MS/MS analysis of TSGH4-6-2-b suggested two novel peptides (GYDEY and EPGKSGEQGAPGEAGAP), which could have potential as naturally-occurring peptides with antioxidant properties. These promising results suggest that these antioxidant peptides could have applications in food products, nutraceuticals, and cosmetics.     

Characterization of Protein Hydrolysates from Fish Discards and By-Products from the North-West Spain Fishing Fleet as Potential Sources of Bioactive Peptides

Fish discards and by-products can be transformed into high value-added products such as fish protein hydrolysates (FPH) containing bioactive peptides. Protein hydrolysates were prepared from different parts (whole fish, skin and head) of several discarded species of the North-West Spain fishing fleet using Alcalase. All hydrolysates had moisture and ash contents lower than 10% and 15%, respectively. The fat content of FPH varied between 1.5% and 9.4% and had high protein content (69.8–76.6%). The amino acids profiles of FPH are quite similar and the most abundant amino acids were glutamic and aspartic acids. All FPH exhibited antioxidant activity and those obtained from Atlantic horse mackerel heads presented the highest 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity, reducing power and Cu²⁺ chelating activity. On the other hand, hydrolysates from gurnard heads showed the highest ABTS radical scavenging activity and Fe²⁺ chelating activity. In what concerns the α-amylase inhibitory activity, the IC₅₀ values recorded for FPH ranged between 5.70 and 84.37 mg/mL for blue whiting heads and whole Atlantic horse mackerel, respectively. α-Glucosidase inhibitory activity of FPH was relatively low but all FPH had high Angiotensin Converting Enzyme (ACE) inhibitory activity. Considering the biological activities, these FPH are potential natural additives for functional foods or nutraceuticals.     

Purification of Antioxidant Peptides by High Resolution Mass Spectrometry from Simulated Gastrointestinal Digestion Hydrolysates of Alaska Pollock (Theragra chalcogramma) Skin Collagen

In this study, the stable collagen hydrolysate was prepared by alcalase hydrolysis and twice simulated gastrointestinal digestion from Alaska pollock skin. The characteristics of hydrolysates and antioxidant activities in vitro, including 2,2′-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) radical (ABTS^•+) scavenging activity, ferric-reducing antioxidant power (FRAP) and hydroxyl radical (OH·) scavenging activity, were determined. After twice simulated gastrointestinal digestion of skin collagen (SGI-2), the degree of hydrolysis (DH) reached 26.17%. The main molecular weight fractions of SGI-2 were 1026.26 and 640.53 Da, accounting for 59.49% and 18.34%, respectively. Amino acid composition analysis showed that SGI-2 had high content of total hydrophobic amino acid (307.98/1000). With the simulated gastrointestinal digestion progressing, the antioxidant activities increased significantly (p < 0.05). SGI-2 was further purified by gel filtration chromatography, ion exchange chromatography and high performance liquid chromatography, and the A_1a3c–p fraction with high hydroxyl radical scavenging activity (IC50 = 7.63 μg/mL) was obtained. The molecular weights and amino acid sequences of key peptides of A_1a3c–p were analyzed using high resolution mass spectrometry (LC-ESI-LTQ-Orbitrap-MS) combined with de novo software and UniProt of MaxQuant software. Four peptides were identified from A_1a3c–p, including YGCC (444.1137 Da) and DSSCSG (554.1642 Da) identified by de novo software and NNAEYYK (900.3978 Da) and PAGNVR (612.3344 Da) identified by UniProt of MaxQuant software. The molecular weights and amino acid sequences of four peptides were in accordance with the features of antioxidant peptides. The results indicated that different peptides were identified by different data analysis software according to spectrometry mass data. Considering the complexity of LC-ESI-LTQ-Orbitrap-MS, it was necessary to use the different methods to identify the key peptides from protein hydrolysates.     

Biofunctionality of Enzymatically Derived Peptides from Codfish (Gadus morhua) Frame: Bulk In Vitro Properties,Quantitative Proteomics,and Bioinformatic Prediction

Protein hydrolysates show great promise as bioactive food and feed ingredients and for valorization of side-streams from e.g., the fish processing industry. We present a novel approach for hydrolysate characterization that utilizes proteomics data for calculation of weighted mean peptide properties (length, molecular weight, and charge) and peptide-level abundance estimation. Using a novel bioinformatic approach for subsequent prediction of biofunctional properties of identified peptides, we are able to provide an unprecedented, in-depth characterization. The study further characterizes bulk emulsifying, foaming, and in vitro antioxidative properties of enzymatic hydrolysates derived from cod frame by application of Alcalase and Neutrase, individually and sequentially, as well as the influence of heat pre-treatment. All hydrolysates displayed comparable or higher emulsifying activity and stability than sodium caseinate. Heat-treatment significantly increased stability but showed a negative effect on the activity and degree of hydrolysis. Lower degrees of hydrolysis resulted in significantly higher chelating activity, while the opposite was observed for radical scavenging activity. Combining peptide abundance with bioinformatic prediction, we identified several peptides that are likely linked to the observed differences in bulk emulsifying properties. The study highlights the prospects of applying proteomics and bioinformatics for hydrolysate characterization and in food protein science.     

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.     

Fishery Wastes as a Yet Undiscovered Treasure from the Sea: Biomolecules Sources,Extraction Methods and Valorization

The search for new biological sources of commercial value is a major goal for the sustainable management of natural resources. The huge amount of fishery by-catch or processing by-products continuously produced needs to be managed to avoid environmental problems and keep resource sustainability. Fishery by-products can represent an interesting source of high added value bioactive compounds, such as proteins, carbohydrates, collagen, polyunsaturated fatty acids, chitin, polyphenolic constituents, carotenoids, vitamins, alkaloids, tocopherols, tocotrienols, toxins; nevertheless, their biotechnological potential is still largely underutilized. Depending on their structural and functional characteristics, marine-derived biomolecules can find several applications in food industry, agriculture, biotechnological (chemical, industrial or environmental) fields. Fish internal organs are a rich and underexplored source of bioactive compounds; the fish gut microbiota biosynthesizes essential or short-chain fatty acids, vitamins, minerals or enzymes and is also a source of probiotic candidates, in turn producing bioactive compounds with antibiotic and biosurfactant/bioemulsifier activities. Chemical, enzymatic and/or microbial processing of fishery by-catch or processing by-products allows the production of different valuable bioactive compounds; to date, however, the lack of cost-effective extraction strategies so far has prevented their exploitation on a large scale. Standardization and optimization of extraction procedures are urgently required, as processing conditions can affect the qualitative and quantitative properties of these biomolecules. Valorization routes for such raw materials can provide a great additional value for companies involved in the field of bioprospecting. The present review aims at collecting current knowledge on fishery by-catch or by-products, exploring the valorization of their active biomolecules, in application of the circular economy paradigm applied to the fishery field. It will address specific issues from a biorefinery perspective: (i) fish tissues and organs as potential sources of metabolites, antibiotics and probiotics; (ii) screening for bioactive compounds; (iii) extraction processes and innovative technologies for purification and chemical characterization; (iv) energy production technologies for the exhausted biomass. We provide a general perspective on the techno-economic feasibility and the environmental footprint of the production process, as well as on the definition of legal constraints for the new products production and commercial use.     

Analysis of polyphenols in cereals may be improved performing acidic hydrolysis: A study in wheat flour and wheat bran and cereals of the diet

Most analytical studies on polyphenols in cereals refer to compounds determined in aqueous-organic extracts and alkali hydrolysates, but an appreciable amount of polyphenols bound to cell wall constituents may remain insoluble in the residues of extraction and alkali hydrolysis. The main objective of this work was to determine if sulphuric acid hydrolysis may release significant amounts of polyphenols to be considered for analytical and nutritional studies. HPLC/MS analyses of polyphenols were performed in methanol–acetone extracts, alkali and sulphuric acid hydrolysates of wheat flour, bran and a pool of cereals of the diet. The amount of polyphenols found in the acidic hydrolysates (200–1600 mg/100 g) was higher than in alkali hydrolysates (0.2–372 mg/100 g). Lower amount of polyphenols were found in the methanol–acetone extracts (44–160 mg/100 g). Hydroxybenzoic, caffeic, cinammic, ferulic and protocatechuic acids were the main constituents of the hydrolysates. The contribution of cereals to the intake of dietary polyphenols in Spain was estimated around 360 mg/person/day (65 mg of extractable and 295 mg nonextractable polyphenols).     

Proteases Production and Chitin Preparation from the Liquid Fermentation of Chitinous Fishery By-Products by Paenibacillus elgii

Chitinous fishery by-products have great application in the production of various bioactive compounds. In this study, Paenibacillus elgii TKU051, a protease-producing bacterial strain, was isolated using a medium containing 1% squid pens powder (SPP) as the sole carbon/nitrogen (C/N) source. P. elgii TKU051 was found to produce at least four proteases with molecular weights of 100 kDa, 57 kDa, 43 kDa, and 34 kDa (determined by the gelatin zymography method). A P. elgii TkU051 crude enzyme cocktail was optimally active at pH 6–7 and 60 °C. The 2,2-diphenyl-1-picrylhydrazyl radical scavenging activity and α-glucosidase inhibitory activity of the hydrolysates obtained from the hydrolysis of shrimp shell powder, shrimp head powder, shrimp meat powder, fish head powder and soya bean powder catalyzed by the P. elgii TkU051 crude enzyme cocktail were also evaluated. P. elgii TKU051 exhibited a high deproteinization capacity (over 94%) on different kinds of shrimp waste (shrimp heads and shells; fresh and cooked shrimp waste; shrimp waste dried by oven and lyophilizer), and the Fourier-transform infrared spectroscopy profile of the chitin obtained from the deproteinization process displayed the characteristic of chitin. Finally, the obtained chitin exhibited an effect comparable to commercial chitin in terms of adsorption against Congo Red (90.48% and 90.91%, respectively). Thus, P. elgii TKU051 showed potential in the reclamation of chitinous fishery by-products for proteases production and chitin extraction.     

Physicochemical,Nutritional and In Vitro Antidiabetic Characterisation of Blue Whiting (Micromesistius poutassou) Protein Hydrolysates

Protein hydrolysates from low-value underutilised fish species are potential sources of high-quality dietary protein and health enhancing peptides. Six blue whiting soluble protein hydrolysates (BW-SPH-A_F), generated at industrial scale using different hydrolysis conditions, were assessed in terms of their protein equivalent content, amino acid profile and score and physicochemical properties in addition to their ability to inhibit dipeptidyl peptidase IV (DPP-IV) and stimulate the secretion of insulin from BRIN-BD11 cells. Furthermore, the effect of simulated gastrointestinal digestion (SGID) on the stability of the BW-SPHs and their associated in vitro antidiabetic activity was investigated. The BW-SPHs contained between 70–74% (w/w) protein and all essential and non-essential amino acids. All BW-SPHs mediated DPP-IV inhibitory (IC₅₀: 2.12–2.90 mg protein/mL) and insulin secretory activity (2.5 mg/mL; 4.7 to 6.4-fold increase compared to the basal control (5.6 mM glucose alone)). All BW-SPHs were further hydrolysed during SGID. While the in vitro DPP-IV inhibitory and insulin secretory activity mediated by some BW-SPHs was reduced following SGID, the activity remained high. In general, the insulin secretory activity of the BW-SPHs were 4.5–5.4-fold higher than the basal control following SGID. The BW-SPHs generated herein provide potential for anti-diabetic related functional ingredients, whilst also enhancing environmental and commercial sustainability.     

Application of the Gadidae Fish Processing Waste for Food Grade Gelatin Production

Waste from fish cutting (heads, swim bladders, fins, skin, and bones) is a high-value technological raw material for obtaining substances and products with a wide range of properties. The possibility of using waste from cutting fish of the Gadidae family: the Alaska pollock (Gadus chalcogrammus) and the Pacific cod (Gadus macrocephalus), processed in the coastal zone, is scientifically substantiated. In this work, a technology has been developed for processing accumulated waste from fish cutting in order to obtain fish gelatin, which is characterized by high protein content (more than 80.0%) and a full set of essential and nonessential amino acids. We studied the quality of fish gelatin obtained from wastes from cutting the fish of the Gadidae family. The possibility of using fish gelatin as a component of fish products is shown; the dose of its introduction into the fish products is substantiated. The data obtained made it possible to recommend the use of fish processing waste products as a gelling component and a source of amino acids in multicomponent food systems.     

Development and evaluation of rheological and bioactive properties of rice protein-based gels

The rice industry produces a large amount of by-products daily. In fact, it is estimated that approximately 100 million tons of rice residues and by-products are generated each year. However, new protein sources are demanded for human food because of the continuous increase in the global population coupled with the almost total inability to increase global food production.A rice protein concentrate (RPC), which is a by-product of the rice industry, and two different hydrolysates (RPH₂₅ and RPH₁₂₀) obtained from RPC, have been used in order to evaluate the potential of RPC to form a gel-like food product at three different pH values (2.0, 6.5 and 8.0). The gelation process was monitored and subsequently, mechanical spectra were obtained. In addition, protein interactions (ionic interactions, hydrophobic interactions, hydrogen bonds and disulphide bonds) were also determined in order to understand the gel structure. Finally, an antioxidant characterisation was carried out using three different reagents: DDPH, ABTS and Folin-Ciocalteu.A proper degree of hydrolysis and an optimal pH value seem to be key factors in the manufacturing of gels, showing a remarkable influence on both rheological properties and antioxidant activities.     

Preparation and Characterization of Thermally Stable Collagens from the Scales of Lizardfish (Synodus macrops)

Marine collagen is gaining vast interest because of its high biocompatibility and lack of religious and social restrictions compared with collagen from terrestrial sources. In this study, lizardfish (Synodus macrops) scales were used to isolate acid-soluble collagen (ASC) and pepsin-soluble collagen (PSC). Both ASC and PSC were identified as type I collagen with intact triple-helix structures by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and spectroscopy. The ASC and PSC had high amino acids of 237 residues/1000 residues and 236 residues/1000 residues, respectively. Thus, the maximum transition temperature (T_max) of ASC (43.2 °C) was higher than that of PSC (42.5 °C). Interestingly, the T_max of both ASC and PSC was higher than that of rat tail collagen (39.4 °C) and calf skin collagen (35.0 °C), the terrestrial collagen. Solubility tests showed that both ASC and PSC exhibited high solubility in the acidic pH ranges. ASC was less susceptible to the “salting out” effect compared with PSC. Both collagen types were nontoxic to HaCaT and MC3T3-E1 cells, and ASC was associated with a higher cell viability than PSC. These results indicated that ASC from lizardfish scales could be an alternative to terrestrial sources of collagen, with potential for biomedical applications.     

Characterisation and functional properties of Australian rice protein isolates

Albumin, globulin, glutelin and prolamin fractions were isolated from an Australian rice variety (cv. Langi) and characterised by yield, protein content and molecular weight profile using both capillary electrophoresis (SDS-CE) and sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE). The influence of pre-extraction enzymatic hydrolysis of starch and heating to 70 °C was also investigated, as was the extraction of the glutelin fraction without prior removal of the albumin and globulin fractions. Pre-extraction treatment affected mainly the albumin fraction, increasing dry matter yield but reducing protein content. SDS-CE was able to separate the protein fractions over a wider molecular weight range than SDS-PAGE, and the peaks from SDS-CE showed slightly higher molecular weight compared to equivalent bands from SDS-PAGE. The glutelin fraction extracted without prior removal of albumin and globulin fractions had different characteristics compared to those obtained by conventional extraction methods. Pre-extraction hydrolysis of starch did not significantly affect the emulsifying, foaming and gelling properties of extracted protein. Although rice glutelin had poor solubility, emulsifying and foaming properties in aqueous systems, it had good gelling properties which could be important for food applications.     

Fractionation of tryptic gliadin hydrolysates based on proline levels

The central domain (CD) and terminal domains (TDs) of wheat gliadins contain high and low levels of the amino acid proline (Pro), respectively. The CD is rather hydrophilic while the TDs are rather hydrophobic and contain most of the ionisable amino acids, although present only in low levels. Therefore, peptides derived from the CD or TDs may strongly differ in their physico–chemical properties. Trypsin cleaves peptide bonds at lysine and arginine residues which are mainly present in the TDs and was used in the present study. Several fractionation methods were examined to isolate CD and TD related peptides from tryptic hydrolysates. Pro was used as a marker for CD and TD related peptides. Both semi-preparative reversed-phase HPLC separation and fractionation of the water-soluble peptides of the tryptic gliadin hydrolysates by graded ethanol precipitation resulted in peptide fractions with different Pro levels. Whereas the fractions precipitating up to 90% ethanol were Pro rich, a Pro poor fraction consisting of small peptides was soluble in 90% ethanol solution. The water insoluble peptides of the tryptic hydrolysates also showed a low Pro level. Ultrafiltration of the water-soluble peptides using a 5-k membrane resulted in small Pro poor peptides and larger Pro rich peptides.     

Evidence of Anti-Proliferative Activities in Blue Mussel (Mytilus edulis) By-Products

Shellfish waste components contain significant levels of high quality protein and are therefore a potential source for biofunctional high-value peptides. The feasibility of applying a pilot scale enzymatic hydrolysis process to whole Mytilus edulis and, by fractionation, recover hydrolysates presenting a biological activity of interest, was evaluated. Fractions were tested on four immortalized cancerous cell lines: A549, BT549, HCT15 and PC3. The 50 kDa fraction, enriched in peptides, presented anti-proliferative activity with all cell lines and results suggest a bioactive molecule synergy within the fraction. At a protein concentration of 44 µg/mL, the 50 kDa fraction induced a mortality of 90% for PC3, 89% for A549, 85% for HCT15 and of 81% for BT549 cell lines. At the low protein concentration of only 11 µg/mL the 50 kDa fraction still entails a cell mortality of 76% for A549 and 87% for PC3 cell lines. The 50 kDa fraction contains 56% of proteins, 3% of lipids and 6% of minerals on a dry weight basis and the lowest levels detected of taurine and methionine and highest levels of threonine, proline and glycine amino acids. The enzymatic hydrolysis process suggests that Mytilus edulis by-products should be viewed as high-valued products with strong potential as anti-proliferative agent and promising active ingredients in functional foods.     

Enzyme-Assisted Extraction of Bioactive Material from Chondrus crispus and Codium fragile and Its Effect on Herpes simplex Virus (HSV-1)

Codium fragile and Chondrus crispus are, respectively, green and red seaweeds which are abundant along the North Atlantic coasts. We investigated the chemical composition and antiviral activity of enzymatic extracts of C. fragile (CF) and C. crispus (CC). On a dry weight basis, CF consisted of 11% protein, 31% neutral sugars, 0.8% sulfate, 0.6% uronic acids, and 49% ash, while CC contained 27% protein, 28% neutral sugars, 17% sulfate, 1.8% uronic acids, and 25% ash. Enzyme-assisted hydrolysis improved the extraction efficiency of bioactive materials. Commercial proteases and carbohydrases significantly improved (p ≤ 0.001) biomass yield (40%–70% dry matter) as compared to aqueous extraction (20%–25% dry matter). Moreover, enzymatic hydrolysis enhanced the recovery of protein, neutral sugars, uronic acids, and sulfates. The enzymatic hydrolysates exhibited significant activity against Herpes simplex virus (HSV-1) with EC₅₀ of 77.6–126.8 μg/mL for CC and 36.5–41.3 μg/mL for CF, at a multiplicity of infection (MOI) of 0.001 ID₅₀/cells without cytotoxity (1–200 μg/mL). The extracts obtained from proteases (P1) and carbohydrases (C3) were also effective at higher virus MOI of 0.01 ID₅₀/cells without cytotoxity. Taken together, these results indicate the potential application of enzymatic hydrolysates of C. fragile and C. crispus in functional food and antiviral drug discovery.