Potential of Subcritical Water Hydrolysis to Valorize Low-Valued Ray-Finned Fish (Labeobarbus nedgia): Effects of Hydrolysis Temperature and Pressurization Agent.

Subcritical water (SCW) hydrolysis was applied to valorize the low-valued ray-finned fish (Labeobarbus nedgia) into valuable protein hydrolysates, employing N2 and CO2 as pressurization agents at varying temperatures (140, 160, 180, and 200 °C). The degree of hydrolysis (DH) and total free amino acid content increased with temperature for both pressurizing agents. The highest DH (54.5 ± 0.4%) and total free amino acid content (210 ± 1 mg/gprot) were observed at 200 °C when CO2 gas was used as the pressurizing agent. Predominantly, glycine and alanine were released for both pressurizing agents. The antioxidant activity, evaluated through three different assays, increased with temperature and was found to be the highest at 200 °C. This study illustrated the advantages of the intensified SCW technology by using CO2 as a pressurization agent in valorizing low-valued ray-finned fish (Labeobarbus nedgia), as animal residue rich in proteins, for the production of valuable protein hydrolysates with a high fraction of valuable free amino acids, which could offer potential applications as a functional ingredient in the food industry.

Green fractionation and hydrolysis of fish meal to improve their techno-functional properties.

A green strategy employing water as solvent has been adopted to obtain protein hydrolysates from fish meal (FM), its water-soluble fraction (WSP), and its non-water-soluble fraction (NSP). The techno-functional properties of the hydrolysates have been investigated and compared to hydrolysates obtained with Alcalase®. In general, SWH hydrolysates presented higher content of free amino acids and higher degree of hydrolysis, which reflected on the molecular size distribution. However, Alcalase® hydrolysates presented better solubility (from 74 ± 4% for NSP at pH = 2 up to 99 ± 1% for WSP at pH = 4-7). According to fluorescence experiments, FM and NSP hydrolysates showed the highest surface hydrophobicity, which has been related to better emulsifying properties and higher emulsion stability. The emulsions stabilized with 2%wt. of SWH-treated NSP showed the smallest particle sizes, with D[4,3] = 155 nm at day 0, and good stability, with D[4,3] = 220 nm at day 7, proving that water fractionation followed by SWH treatment is a good method to improve the techno-functional properties of the hydrolysates.

Cosmeceutical Potential of Extracts Derived from Fishery Industry Residues: Sardine Wastes and Codfish Frames.

The fishery industry generates large amounts of waste (20-75% (w/w) of the total caught fish weight). The recovery of bioactive compounds from residues and their incorporation in cosmetics represents a promising market opportunity and may contribute to a sustainable valorisation of the sector. In this work, protein-rich extracts obtained by high-pressure technologies (supercritical CO2 and subcritical water) from sardine (Sardina pilchardus) waste and codfish (Gadus morhua) frames were characterized regarding their cosmeceutical potential. Antioxidant, anti-inflammatory and antibacterial activities were evaluated through chemical (ORAC assay), enzymatic (inhibition of elastase and tyrosinase), antimicrobial susceptibility (Klebsiella pneumoniae, Staphylococcus aureus and Cutibacterium acnes) and cell-based (in keratinocytes-HaCaT) assays. Sardine extracts presented the highest antibacterial activity, and the extract obtained using higher extraction temperatures (250 °C) and without the defatting step demonstrated the lowest minimum inhibitory concentration (MIC) values (1.17; 4.6; 0.59 mg/mL for K. pneumoniae, S. aureus and C. acnes, respectively). Codfish samples extracted at lower temperatures (90 °C) were the most effective anti-inflammatory agents (a concentration of 0.75 mg/mL reduced IL-8 and IL-6 levels by 58% and 47%, respectively, relative to the positive control). Threonine, valine, leucine, arginine and total protein content in the extracts were highlighted to present a high correlation with the reported bioactivities (R2 ≥ 0.7). These results support the potential application of extracts obtained from fishery industry wastes in cosmeceutical products with bioactive activities.

Valorization of brewer's spent grain by consecutive supercritical carbon dioxide extraction and enzymatic hydrolysis.

The double effect of supercritical carbon dioxide, sc-CO2, in a biorefinery concept applied to brewer's spent grain (BSG) was assessed in this work. Extraction conditions to remove and valorize the lipophilic fraction were studied (20-40 MPa and 40-80 °C) obtaining a maximum yield of 5.70 ± 0.07 g/100 gBSG at 80 °C and 40 MPa. High pressures and temperatures resulted in higher content of total phenolic and flavonoid compounds, as well as higher antioxidant capacity. It was observed an improvement of the enzymatic hydrolysis yield by cellulase in the sc-CO2 treated BSG compared to the non-treated. This improvement could be partially attributed to the removal of the lipid fraction and to morphological changes of BSG after sc-CO2. Based on this double benefit, sc-CO2 can play an important role on biomass valorization.

Subcritical Water Extraction and Hydrolysis of Cod (Gadus morhua) Frames to Produce Bioactive Protein Extracts.

The valorization of Atlantic cod (Gadus morhua) frames from a filleting industry was investigated using subcritical water extraction and hydrolysis (SBW) at different temperatures (90, 140, 190 and 250 °C) and 100 bar to obtain extracts rich in proteins, peptides and amino acids. Up to 57.7 g of extract per 100 g of codfish frames were obtained, with nearly total recovery of the protein fraction. At each temperature, protein extracts of decreasing molecular weight were obtained, according to SEC-GPC results. Most of the protein present in the raw material and extracts was collagen and collagen fragments, as suggested by the amino acid profile. Codfish SBW extracts did not show cytotoxicity in the range of concentrations tested and the protein extract obtained at the lowest temperature (90 °C) showed the highest anti-inflammatory potential in human intestinal epithelium cell model. The mineralized residue left after SBW treatment of cod frames was identified as practically pure, crystalline, hydroxyapatite, that may find applications in biomedical field and hard-tissue engineering. This study shows the possible valorization of cod frames using green extraction methods such as SBW process to obtain protein extracts for food and nutraceutical applications.

Omega-3 encapsulation by PGSS-drying and conventional drying methods. Particle characterization and oxidative stability.

Particles from Gas-Saturated Solutions (PGSS)-drying has been used as a green alternative to encapsulate omega-3 polyunsaturated fatty acids (n-3 PUFAs) at mild, non-oxidative conditions. PGSS-dried particles have been compared to those obtained by conventional drying methods such as spray-drying and freeze-drying, finding encapsulation efficiencies (EE) up to 98% and spherical morphology for PGSS- and spray-dried particles. Freeze-dried powders showed irregular morphology and EE from 95.8 to 98.6%, depending on the freezing method. Differential scanning calorimetry (DSC) analysis revealed glass-transition and melting peaks of OSA-starch and a cold-crystallization peak corresponding to the encapsulated n-3 PUFA concentrate. Compared to conventionally dried powders, PGSS-dried microparticles showed lower primary and secondary oxidation after 28 days of storage at 4 °C. Ascorbic acid addition combined with the mild processing conditions of PGSS-drying yielded particles with a maximum peroxide value of 2.5 meq O2/kg oil after 28 days of storage at 4 °C.

Substrates emulsification process to improve lipase-catalyzed sardine oil glycerolysis in different systems. Evaluation of lipid oxidation of the reaction products.

Mono- and diacylglycerols rich in omega-3 have a great interest due to their good bioavailability and oxidation stability compared with other kind of omega-3 concentrates. The main drawback in mono- and diacylglycerols production by glycerolysis is the immiscibility of the substrates, oil and glycerol. To improve mass transfer rates, avoiding the use of organic solvents, emulsification of both reactants as reverse micelles (glycerol-in-oil) was carried out previous to lipase-catalyzed sardine oil glycerolysis. Substrate emulsification yielded higher reaction rates compared to kinetics with no previous emulsification, but still lower than in organic solvents. To avoid the use of organic solvent, SC-CO2 was used as reaction medium but no kinetic advantages were demonstrated in the pressure range from 15 to 25 MPa. By increasing temperature, from 40 to 90°C, reaction rates increased both in a solvent-free system and in SC-CO2 medium. It was also found that an increase in temperature does not lead to an increase in the final oxidation status of the reaction products. This behavior was due to the adsorption capacity of the Lipozyme 435 support, giving lower oxidation status at the highest temperature, 80-90°C.

Kinetic study for the ethanolysis of fish oil catalyzed by lipozyme(®) 435 in different reaction media.

The ethanolysis of fish oil in various reaction medium (tert-pentanol, n-hexane and solvent free system) catalyzed by the immobilized commercial lipase Lipozyme(®) 435 (Candida Antarctica) at atmospheric pressure has been studied in this work. The effect of some kinetic parameters, such as the amount of lipase, temperature and the initial reactant molar ratio ethanol:oil on monoacyglyceride and ethyl ester yield has been analyzed. Experimental data were successfully correlated by a simple kinetic model based on the elementary reactions proposed in this work. At high initial reactant molar ratio the three elementary steps can be considered as irreversible. However the reaction rate constants ratio for the deacylation of monoglyceride to glycerol decreased by decreasing the molar ratio ethanol:oil. The reaction rates are slower in n-hexane as reaction medium compared to tert-pentanol and a solvent-free system, at the experimental conditions essayed in this work. In this last case, ethanol acts as solvent for reaction and as reactant.