Search Results (114)

Allicin, the most critical bioactive compound of garlic (Allium sativum L.), is of significant industrial importance when extracted at high purity while preserving its structural integrity. In this study, the combined use of supercritical-CO₂ (SC-CO₂) extraction and molecular distillation (MD) techniques was investigated to obtain garlic extracts with high allicin content from Gaziantep (Araban) garlic. The SC-CO₂ extraction process was optimized using Response Surface Methodology (RSM) within a range of 150–300 bar pressure, 50–80% co-solvent concentration and 0.5–3.0 mL/min solvent flow rate. The obtained extracts were characterized by LC-ESI-DAD-MS/MS, and their biological activities were evaluated using a comprehensive in vitro digestion model. Allicin in vitro digestion was performed using models simulating gastrointestinal conditions of young adults (<65 years) and older adults (>65 years), and its bioactive properties were comparatively evaluated. In the antimicrobial analysis, for SC-CO₂, a strong activity was demonstrated against Staphylococcus aureus and Escherichia coli in the oral phase of the in vitro digestion model, with inhibition zones of 36.33 mm and 26.50 mm in young samples and 34.67 mm and 25.83 mm in older samples, respectively. Owing to the immediate nucleophilic attack triggered by the subsequent alkaline pH shift and pancreatic enzymatic stress, free allicin underwent total structural degradation, falling below detectable limits within the intestinal chyme. In terms of purification performance, allicin content increased from 45.77% after SC-CO₂ extraction to 67.10% after molecular distillation. Crucially, due to the immediate nucleophilic attack driven by the subsequent alkaline pH shift and pancreatic enzymatic stress, free allicin underwent complete structural degradation and was rendered strictly undetectable within the intestinal chyme. This approach provides a sustainable and environmentally friendly purification strategy that effectively limits the thermal degradation of allicin. The results present a practical framework for the scalable production of allicin-rich nutraceutical intermediates and functional food ingredients. Full article

Almonds (Prunus dulcis; family Rosaceae) contain 18–25% protein (dry weight). They are an important plant-based protein source in dairy alternatives and other functional foods. The hard and dense nature of almond kernels and the localization of proteins with lipid bodies in the cotyledons of almond seeds make it challenging to recover protein from the seed efficiently and preserve its function. Therefore, this review evaluates the influence of pretreatments, including blanching, grinding, and defatting, on almond protein recovery and functionality, and compares conventional and emerging technologies for almond protein. Traditional protein extraction techniques such as alkaline extraction–isoelectric precipitation (AE–IEP), aqueous extraction, and salt extraction provide moderate-to-high protein yields, but harsh processing conditions denature the proteins, decrease solubility, and cause functional properties to be lost. On the other hand, emerging protein extraction technologies (including enzyme-assisted aqueous extraction (EAE) ultrasound-assisted extraction (UAE), microwave-assisted extraction (MAE), high-pressure processing (HPP), and pulsed electric field (PEF) treatment) improve protein recovery, resulting in protein extract with superior functional properties and reduced allergenicity. However, their application in industry remain challenging. This review reveals that pretreatment approaches and conditions/parameters significantly influence protein extraction efficiency and the functional and structural properties of almonds, and that no single method is universally optimal. This review concludes that controlled enzymatic hydrolysis combined with physical pretreatment may be the best approach for producing high-value-added almond protein ingredients with specific techno-functional properties for use in plant-based beverages, hypoallergenic products, or nutraceuticals. More research is needed to develop an efficient, applicable, sustainable and eco-friendly almond protein extraction process, optimizing processing conditions to achieve high protein recovery while retaining desirable functional properties, and reduce operating costs. Full article

Tomato processing generates large amounts of by-products, with seeds representing an underutilized yet protein-rich fraction. This study investigated direct enzyme-assisted protein extraction from non-defatted tomato seeds. Various enzymes, enzyme/substrate ratios, pre-treatments, and incubation temperatures were evaluated and optimized. An enzyme/substrate ratio of 5% (w/w) was found to be optimal, with proteases alone outperforming cell wall-degrading enzymes and two-step extraction strategies. Bromelain, Protamex, and Trypsin, for the first time applied directly to non-defatted tomato seeds, achieved the highest protein recoveries (average 110.56 mg BSA eq/g DW). Among them, Trypsin also produced the highest reducing sugar content (25.07 mg GLU eq/g DW), indicating effective cell wall disruption. Digestates obtained from defatted and non-defatted tomato seeds showed comparable protein contents, demonstrating that defatting was unnecessary. Avoiding the defatting step improved process sustainability by reducing solvent use and energy consumption without significantly affecting protein extraction efficiency. Incubation at 37 °C was preferred over 60 °C, as similar yields were achieved under milder conditions while also reducing energy consumption by approximately three-fold (54,340 kJ vs 150,480 kJ for a 1000 L water-based scale-up simulation). These digestates showed significantly higher antioxidant and, for the first time in tomato seed extracts, anti-tyrosinase activities compared with controls. Protamex-derived samples exhibited the highest bioactivities (7.40 mg AA eq/g DW; 101.36 μg KA eq/g DW). Compared to conventional alkaline–acid extraction followed by enzymatic digestion, the direct enzymatic approach provided higher protein recovery. Overall, this method represents a sustainable strategy for producing bioactive peptide-rich extracts for food and non-food applications. Full article

The industrial extraction of chitin from shrimp shell waste conventionally employs corrosive chemical treatments, which pose significant environmental hazards and compromise polymer integrity. This study introduces a sustainable and highly efficient microbial biorefining strategy for the recovery of α-chitin from Litopenaeus vannamei shells, utilizing a sequential fermentation framework. Two potent strains—Bacillus haynesii MGPUMGRI, known for its proteolytic capabilities, and Lactobacillus delbrueckii MGPUMGRI, which produces lactic acid—were isolated and optimized. A notable technical achievement was the purification of an approximately 40 kDa extracellular alkaline protease from B. haynesii, which demonstrated optimal activity at pH 9.0 and 37 °C. Under optimized conditions, the sequential process—emphasizing enzymatic deproteinization (72.30 ± 1.56%) followed by lactic acid-mediated demineralization (84.98 ± 1.96%)—achieved a high-purity chitin recovery of 61.33 ± 1.06%. Comprehensive characterization using SEM-EDX, FTIR, and XRD confirmed the successful preservation of the α-chitin polymorphic structure, which exhibited a fragmented fibrillar morphology and a crystallinity index (CrI) of 60.51%. These findings indicate that this dual-strain bioprocess offers a scalable and environmentally friendly alternative for the valorization of seafood waste into high-quality biogenic polymers, while minimizing the ecological impact of chitin production. Full article

Lignocellulosic biomass represents an abundant and renewable carbon source, and its valorization through microalgal cultivation offers a sustainable route to resource-efficient bioprocessing. This study examined the effects of various carbon and nitrogen sources on the growth and lipid metabolism of Desmodesmus subspicatus, with a focus on ryegrass enzymatic hydrolysates as an alternative carbon source. Cultures were supplied with glucose, xylose, or arabinose at different concentrations, along with sodium nitrate or yeast extract, under different carbon-to-nitrogen ratios. Additionally, the impacts of alkaline- and acid-pretreated enzymatic ryegrass hydrolysates were evaluated. Growth was assessed by optical density and gravimetric analysis, and fatty acid profiles by gas chromatography. Glucose supplementation enhanced lipid accumulation, yielding fatty acid profiles dominated by C16 and C18 fatty acids, which are favorable for the quality of the produced biodiesel. Nitrogen limitation further promoted lipid accumulation; cultures supplied with sodium nitrate achieved higher total lipid content, while yeast extract favored greater proportions of PUFAs. Alkaline-pretreated ryegrass hydrolysate supported dose-dependent biomass formation reaching approximately 12 g L⁻¹ at 50%, whereas the acid-pretreated hydrolysate exhibited inhibitory effects at the same concentration. Scale-up in a 1 L photobioreactor yielded lower biomass but higher lipid content with a fatty acid profile shifted to SFA. These results support ryegrass as a viable alternative carbon source and highlight cultivation parameters that influence growth and lipid quality relevant for biofuel applications. Full article

Determination of complex pesticide residues in food matrices poses a considerable analytical challenge, primarily because the analytes exhibit diverse physicochemical properties. Monitoring pesticides across a wide range is essential to meet all regulatory requirements and safeguard consumer health. One of the most promising analytical approaches is the hydrolysis of compounds, particularly acidic hydrolysis, which enables the identification of a broad range of substances that pose significant analytical challenges. In addition, pesticide residues may interact with food matrix components, leading to the formation of conjugated forms such as ester- or glycoside-bound compounds. Therefore, the development of appropriate analytical strategies, including hydrolytic steps, is essential to release these bound residues and enable the determination of complex residue definitions comprising multiple related compounds. Furthermore, this study compares different hydrolysis strategies, including enzymatic, acidic, and alkaline hydrolysis, in order to assess their suitability for the determination of complex pesticide residue definitions using a QuEChERS-based (Quick, Easy, Cheap, Effective, Rugged, Safe) extraction approach. The given methodology meets all criteria listed in the Document SANTE 11312/2021 v2026. The procedure allows for good measurement precision relative standard deviation (RSD < 20%), and recovery in the scope ranging from 55.6% to 107.8% acidic hydrolysis and 51.7% to 100.8% for alkaline hydrolysis and 35.1–108.9% for enzymatic hydrolysis depending on the experimental variant, and limit of quantification (LOQ) as low as 10 to 100 µg/kg for the determination of ten complex definitions of pesticides with the use of liquid chromatography mass spectrometry (LC-MS/MS) and gas chromatography mass spectrometry (GC-MS/MS) analytical methods. Full article

Plum pomace (PP), a key by-product of plum juice processing, is a rich yet underutilized source of dietary fiber. However, its high-value exploitation is severely limited by the lack of efficient extraction and modification technologies. This study optimized the extraction of soluble dietary fiber (SDF) and insoluble dietary fiber (IDF) from plum pomace (PP) via Design of Experiments (DOE), and evaluated their modification effects. Alkaline extraction was screened as the optimal method for IDF, and orthogonal experiments determined the optimal conditions: solid-to-liquid ratio 1:20 g/mL, 14 g/L NaOH, 60 °C, and 80 min, achieving a high extraction yield of 62.18%. For SDF, enzymatic extraction was superior, and response surface methodology (RSM) optimized the process to a solid-to-liquid ratio of 1:15.5, 1.0% enzyme dosage, 61.5 °C, and 92 min, with a yield of 29.3%. Physical, chemical, and biological modifications all significantly enhanced SDF’s water/oil-holding capacity, cholesterol/glucose adsorption capacity, and cation exchange capacity. Biologically modified SDF showed the most significant enhancement, with WHC of 5.58 ± 0.05 g/g, OHC of 4.38 g/g, CAC of 7.68 mg/g, and CEC of 3.28 mmol/g. These results provide technical support for the high-value utilization of PP and lay a foundation for its application in functional foods and nutraceuticals. Full article

Development of green and efficient technologies for valorizing crayfish shell waste is crucial for enhancing industrial value. This study presents an integrated strategy for the extraction and structural engineering of chitin using a novel alkaline deep eutectic solvent (DES) system composed of lysine and monoethanolamine (LysMEA), which enables the simultaneous deproteinization and architectural modification of chitin. Following mild demineralization, the optimized process yielded chitin with 97.1% purity and a high molecular weight of 209.3 kDa. DES demonstrated considerable reusability and decolorization capability. Structural characterization revealed that the LysMEA system effectively engineered the chitin architecture, resulting in lower crystallinity and a larger surface area compared to conventional methods. This engineered structure rendered the chitin highly accessible to enzymes. Consequently, the chitin extracted by LysMEA exhibited superior reactivity, achieving a deacetylation degree of 63.7% when catalyzed by Bacillus aryabhattai chitin deacetylase, significantly outperforming chitin obtained via acid-alkali or acidic DES methods. Molecular dynamics simulations elucidated the mechanism, showing that lysine and monoethanolamine molecules penetrated the chitin fiber bundles at high temperatures, weakening interchain hydrogen bonds and partially separating the chains. This work provides a green route for producing enzymatically reactive chitin, demonstrating the potential of solvent-based structural engineering in biocatalytic valorization. Full article

Quinoa (Chenopodium quinoa) is a promising source of plant proteins with the potential to produce bioactive peptides through enzymatic hydrolysis. This study aimed to extract quinoa protein and produce bioactive peptides using two microbial proteases: Alcalase (from Bacillus licheniformis) and Flavourzyme (from Aspergillus oryzae). The protein was extracted through alkaline solubilization and isoelectric precipitation, achieving a 72% yield. Hydrolysis was conducted for 4 h, and enzymatic activity was measured using the TNBS method to determine the degree of hydrolysis, while SDS-PAGE was used to analyze protein breakdown. The reaction was performed at controlled pH and temperature (Alcalase: 9.5 and 55 °C; Flavourzyme: 7 and 37 °C). Both enzymes achieved maximum hydrolysis at 60 min. Consequently, the separation and inhibitory capacity of angiotensin-converting enzyme (ACE-I) were tested at the first four time points (0, 20, 40, and 60 min). A wider variety and higher concentration of peptides smaller than 2 kDa were found in hydrolysates treated with Flavourzyme, which is associated with antihypertensive activity. The ACE-I assay showed greater activity at the end of hydrolysis. Inhibition percentages of 87.5 ± 2.11 were observed in hydrolysates with Flavourzyme, and 94.1 ± 1.11 in those with Alcalase. These findings indicate that quinoa protein, hydrolyzed with microbial proteases, is a feasible source of peptides with potential antihypertensive effects for use in functional foods and nutraceuticals. Full article

Arabinoxylans (AX) and their oligosaccharides (AXOS) have potential as functional ingredients. The emergence of biorefineries, leading to more Distillers Dried Grains with Solubles (DDGS) entering the animal feed market, encourages commercial production of AX products. Extracting AX from the two components of DDGS offers the opportunity to increase the biorefinery’s product portfolio and reduce costs. This paper explores AX extraction from solubles and wet grain, using a Gunt pilot-scale bioethanol plant to produce the two streams. After fermentation and distillation, solids were separated from the liquid to give Wet Distillers Grain (WDG), from which alkaline hydrogen peroxide extraction of water-unextractable AX (WUAX) was performed. The water-extractable AX (WEAX) was recovered from the solubles by ultrafiltration and ethanol precipitation. Both extracts were tested for suitability for AXOS production and characterised for their functionality. 10 kg of wheat yielded 3.2 litres of ethanol at 90% purity, 85 g of WUAX (51.6% purity, 110 kDa) and 92 g of WEAX (74.2% purity, 70 kDa). Enzymatic conversion of WEAX into oligosaccharides was 53%, whereas WUAX was unsusceptible to enzyme hydrolysis. Both AX fractions showed interactions with starch that could increase the shelf life of bakery products. AX-based products could be produced from a range of agricultural and biorefinery waste or low value streams, with the global market potentially > £1 billion per annum. Full article

Conventional alkaline extraction of plant proteins typically requires highly alkaline conditions (pH ≥ 11) and extended extraction times (~1 h). Although protease addition can lower extraction pH and improve functionality, it often requires prolonged hydrolysis. In this study, enzymatic reactive extrusion (eREX) using Alcalase, followed by a short duration alkaline extraction (5 min, pH 9), was evaluated as an alternative approach for producing protein-rich extracts from canola meal. The eREX process increased protein recovery by 48% and 42% compared with alkaline extraction conducted without and with Alcalase, respectively. The resulting powdered extracts reached a protein content of up to 49% and consisted primarily of partially hydrolyzed proteins (10–23 kDa) with increased surface hydrophobicity. Amino acid analysis showed substantial enrichment of essential amino acids, particularly histidine and sulfur-containing amino acids. Functional properties were improved, including enhanced solubility across pH 2–10, high foaming stability (88%), and increased oil-binding capacity (~5.5 g g⁻¹), while in vitro digestibility remained comparable (~85%). Techno-economic analysis indicated reductions in water use (~11%), energy consumption (~48%), and production cost (16–25%). Overall, eREX provides a rapid, higher-throughput, and cost-effective strategy for producing premium canola protein ingredients. Full article

Docosahexaenoic acid (DHA), an omega-3 fatty acid essential for human health, is highly prone to oxidation in nanoemulsions due to their large interfacial area and presence of transition metal ions. This study investigated macroalgal chelators for stabilizing DHA-rich nanoemulsions. Sequential enzymatic–alkaline extraction using Alcalase^® produced an extract with the strongest Fe²⁺-chelating activity (IC₅₀ = 1.22 mg/mL), protein content of 10.11 ± 0.15%, and total phenolics ≈ 17 µg GAE/mL. This extract was incorporated into nanoemulsions (5 wt% DHA oil, 1 wt% Tween^® 20) at 0.61, 1.22, and 2.44 mg/mL and compared with controls containing EDTA (0.025 mg/mL) or no antioxidant. Droplet size remained stable (D_3,2 ≈ 77–80 nm; D_4,3 ≈ 199–215 nm) and zeta potential averaged −17 to −19 mV, confirming physical stability. Confocal microscopy revealed concentration-dependent interfacial adsorption of extract components. During iron-accelerated storage, extract-treated nanoemulsions slowed hydroperoxide formation and delayed tocopherol depletion compared to the control, while reducing volatile oxidation markers such as 1-penten-3-ol by up to 40%. However, EDTA consistently provided superior protection against oxidation. These findings highlight the potential of macroalgal extracts as clean-label, natural chelators for mitigating metal-driven oxidation in DHA nanoemulsions, though synthetic chelators remain more effective under severe prooxidant conditions. Full article

Macroalgae are increasingly recognized as a valuable source of nutrients and bioactive compounds for animal nutrition, including for aquatic species. However, the complex structure of the macroalgal cell wall limits the accessibility of intracellular components, restricting their use in feeds. To overcome this limitation, macroalgal hydrolysis using various technological treatments has been tested, often employing a low solid-to-water ratio, which complicates downstream processing due to phase separation. In contrast, high-solids loading hydrolysis has the advantage of producing a single and consolidated fraction, simplifying subsequent processing and application. The present study assessed the effectiveness of high-solids loading water or alkaline (0.5 and 1N NaOH) autoclaving for 30 or 60 min, applied alone or followed by sequential enzymatic hydrolysis, using a xylanase-rich enzymatic complex aimed at promoting cell wall disruption and increasing the extractability of intracellular components in the red macroalga Palmaria palmata with minimal free water. The 1N NaOH treatment for 30 min decreased neutral and acid detergent fiber while increasing Folin–Ciocalteu total phenolic content (GAE) (expressed as gallic acid equivalent) and the water-soluble protein fraction and decreased crude protein, indicating enhanced extractability of these components. Microscopic examination showed relatively mild structural changes on the surface of P. palmata after high-solids loading alkaline (1N NaOH) autoclaving for 30 min. Following alkaline or water treatment, the enzymatic complex hydrolysis further increased the Folin–Ciocalteu total phenolic content (GAE), with minimal effects on NDF, ADF, or crude protein. Overall, these results showed that high-solids loading alkaline autoclaving, with or without subsequent enzymatic hydrolysis, effectively disrupts P. palmata cell walls and induces substantial modifications while simplifying processing by avoiding phase separation. Full article

The growing concern over plastic pollution and the widespread presence of micro- and nanoplastics has renewed interest in polyhydroxybutyrate (PHB) as a biodegradable alternative; however, its industrial deployment remains constrained by costly recovery operations with a high environmental burden. This study examines how PHB biosynthesis and intracellular organization, physicochemical properties, and the characteristics of the producing microorganism influence the performance of conventional recovery routes, including extraction with organic solvents, alkaline/oxidative chemical digestion, and enzymatic–physical schemes coupled with mechanical disruption. Based on this foundation, quantitative data are analyzed for PHB content in bacteria, mixed microbial cultures, cyanobacteria, and microalgae, along with extraction yields, polymer purity, and solvent recyclability in processes employing chlorine-free solvents, green solvents, and hydrophobic natural deep eutectic solvents (NaDESs) formulated with terpenes and organic acids. The analysis integrates mechanistic perspectives on NaDES–cell and NaDES–PHB interactions with solvent design criteria, biorefinery configurations, and preliminary evidence from technoeconomic and life cycle assessments. The findings identify NaDES as an up-and-coming platform capable of reconciling biopolymer quality with the principles of green chemistry while delineating critical gaps in recovery efficiency, viscosity management, solvent recycling, and pilot-scale validation. Full article

Invasive species are a recurring global problem, and the water hyacinth (Pontederia crassipes) is a well-known example. Various strategies have been explored to manage its spread, including its use as an agricultural amendment. However, when P. crassipes biomass is incorporated into soil and undergoes degradation, it may increase soil conductivity and promote metal leaching, potentially affecting soil biota, particularly microbiota. Saprophytic fungi play a key role in the decomposition and renewal of organic matter, and their resilience to stressors is crucial for maintaining soil function. Thus, the aim of this study was to evaluate the effects of P. crassipes biomass extracts on the saprophytic fungus Trametes versicolor by evaluating fungal growth and metabolic changes [including sugar content, phosphatase enzymatic activity, and reactive oxygen species (ROS) production]. The fungus was exposed for 8 days to a dilution series of extracts (100%—undiluted, to 3.13%) prepared from P. crassipes biomass collected at five locations in Portuguese wetlands. Two sites were in the south, within a Mediterranean climate (Sorraia and Estação Experimental António Teixeira), and three were in the north, within an Atlantic climate (São João de Loure, Pateira de Fermentelos, and Vila Valente), representing both agricultural-runoff–impacted areas and recreational zones. Extracts were used to simulate a worst-case scenario. All extracts have shown high conductivity (≥15.4 mS/cm), and several elements have shown a high soluble fraction (e.g., K, P, As, or Ba), indicating substantial leaching from the biomass to the extracts. Despite this, T. versicolor growth rates were generally not inhibited, except for exposure to the São João de Loure extract, where an EC₅₀ of 45.3% (extract dilution) was determined and a significant sugar content decrease was observed at extract concentrations ≥25%. Possibly due to the high phosphorous leachability, both acid and alkaline phosphatase activities increased significantly at the highest percentages tested (50% and 100%). Furthermore, ROS levels increased with increasing extract concentrations, yet marginal changes were observed in growth rates, suggesting that T. versicolor may efficiently regulate its intracellular redox balance under stress conditions. Overall, these findings indicate that the degradation of P. crassipes biomass in soils, while altering chemical properties and releasing soluble elements, may not impair and could even boost microbiota, namely saprophytic fungi. This resilience highlights the potential ecological benefit of saprophytic fungi in accelerating the decomposition of invasive plant residues and contribution to soil nutrient cycling and ecosystem recovery. Full article