The guarana seed peel (GSP), an agroindustrial waste, was characterized for its chemical composition and evaluated for its antimicrobial activity as a potential source of antibacterial compounds for wastewater disinfection using UPLC-QTOF-MS analysis. UPLC-QTOF-MS analysis of GSP, its raw extract (GSPE), and its purified extract (GSPP) identified 36 compounds, including catechin and its oligomers: a B-type procyanidin dimer, pavetannin B6 (trimer), and cinnamtannin A2 (tetramer). Principal component analysis (PCA) separated GSPE from GSPP along PC2, and the purification step concentrated flavonoids—mainly catechin and its oligomers—in GSPP. Antimicrobial activity was assessed against the human pathogens Escherichia coli and Salmonella typhimurium, causing gastroenteritis and invasive disease, and Staphylococcus aureus, causing skin and invasive infections. In real wastewater, GSPE (500 mg L−1) reduced total coliforms by up to 81 ± 7% and E. coli by up to 74 ± 7% within 180 min. Confocal microscopy with the LIVE/DEAD® BacLight™ kit (Thermo Fhisher, Eugene, OR, USA) confirmed loss of membrane integrity in GSPE-treated Escherichia coli, and disc diffusion with GSPP produced inhibition zones of 9–11 mm against Staphylococcus aureus, Escherichia coli and Salmonella typhimurium at 5 mg per disc. These results show that guarana seed peel—currently discarded—is a viable source of catechin-rich antibacterial extracts for wastewater disinfection.
Glucoamylases are highly demanded by the industry for starch saccharification, which necessitates an increase in enzyme activity. To improve the activity of Aspergillus awamori VKPM F-1262 glucoamylase N181Q (AGAM), amino acid substitutions were made in the region of substrate binding. In the case of the variant D237G, there was a 1.3–1.4-fold increase in activity towards soluble starch for the individual enzyme and crude enzyme solution after cultivation in flasks. For the variants G57A and C319T, there was also a 1.1–1.4-fold increase in enzyme activity for the crude flask solution, while the increase in individual enzyme activity was minor. AGAM showed an increase in apparent molecular weight consistent with dimerization in the presence of soluble starch. A high T optimum of 70 °C was observed for AGAM and the variants in the hydrolysis of soluble starch. In further protein engineering, a high T optimum for hydrolysis, which may coincide with dimerization, may be considered as a hypothesis for the selection of thermostable glucoamylases, while an area of binding of oligo- and polysaccharides may be a site for mutagenesis.
The implementation of Quality Management Standards (QMS) in health laboratories is essential to ensuring the traceability, reliability, and reproducibility of results, thus supporting compliance and continuous improvement. This study identifies and compares the requirements adopted in four widely used four standards for health laboratoriesto evaluate their equivalence and contribute to the harmonization of normative terminology. A comparative analysis of the requirements and their applications in health laboratories was conducted. Subsequently, interrelationships and equivalence among the standards were assessed, followed by an evaluation of the terminology used and the grouping of requirements across the four normative documents. The results revealed a substantial overlap among the standards, particularly regarding quality management, risk management, personnel competence, documentation control, and continuous improvement. However, differences in terminology, structure, and scope were identified, which may hinder the uniform interpretation and implementation of quality systems. The harmonization of terms and equivalent requirements could facilitate compliance, integration of management systems, and accreditation processes. The findings demonstrate that standardizing normative terminology contributes to greater alignment among quality standards, improving their understanding, dissemination, and application in health laboratories while supporting more efficient and reliable quality management practices.
The cohesin complex is a highly conserved, ring-shaped protein assembly that plays fundamental roles in chromosome biology. Originally identified as the molecular glue that holds sister chromatids together from DNA replication until cell division, cohesin has since been recognized as a pleiotropic regulator of genome organization, gene expression, DNA repair, and cell fate determination. In embryonic stem cells (ESCs), cohesin’s functions extend beyond canonical sister chromatid cohesion to include the maintenance of three-dimensional (3D) chromatin architecture through DNA loop extrusion, regulation of pluripotency-associated transcriptional programs, and facilitation of homologous recombination-mediated DNA repair during the prolonged S phase. Recent discoveries have revealed that meiosis-specific cohesin components, particularly the α-kleisin subunit REC8 and its interacting partner STAG3, are expressed and functionally active in mitotic ESC chromosomes, where they contribute to chromosomal organization and sister chromatid cohesion in concert with mitotic RAD21-containing cohesin. Furthermore, the interplay between cohesin and condensin complexes at shared genomic binding sites has emerged as a critical determinant of chromosome topology, with cohesin depletion leading to aberrant condensin accumulation and chromosome hypercompaction. Importantly, perturbations in cohesin function not only impair ESC self-renewal but also direct lineage-specific differentiation, linking cohesin to stem cell fate determination. Germline mutations in cohesin and its regulators underlie a spectrum of developmental disorders termed cohesinopathies, while somatic mutations are frequently observed in various cancers. This review provides a comprehensive overview of the diverse roles of cohesin in chromosome structure, cell cycle regulation, and stem cell biology, with particular emphasis on recent findings in ESCs that illuminate the complex interplay between mitotic and meiotic cohesin complexes.
Halotolerant plant growth-promoting bacteria offer a sustainable strategy to improve crop performance under saline conditions. This study evaluated Agave tequilana Weber var. Azul leaf juice was used as an alternative growth medium for Kocuria sediminis AS04, and the bacterium's ability to alleviate salt stress in chiltepin (Capsicum annuum) was evaluated. The chiltepin seedlings were grown in a specialized chamber and exposed to higher salt levels (200, 400, and 600 mM NaCl) for 10 days. During this time, the protective effect of K. sediminis AS04, which was held in a polymer film, was examined. K. sediminis AS04 grew well in a medium containing 25% agave juice and urea, reaching a density of 1.3 × 1010 CFU mL-1. Compared with conventional Tryptic Soy Broth, the medium formulated from agave leaf juice and urea could reduce the cost per kilogram of biomass by approximately 4-fold. Under severe salinity stress, plants inoculated with immobilized K. sediminis at 600 mM NaCl exhibited the highest shoot biomass (0.31 g-1 plant), root length (50.7 mm), and proline accumulation (11.25 µmol g-1 fresh weight), whereas uninoculated plants displayed reduced biomass (0.16 g plant-1) and shorter roots (34.5 mm). At 600 mM NaCl, inoculation increased shoot biomass, root length, and plant survival by approximately 94%, 47%, and 200%, respectively, compared with uninoculated seedlings. This method values agro-industrial waste and enhances chiltepin performance under high-salinity conditions. It promotes sustainable production and helps create affordable inoculants for agricultural biotechnology.
Polyamines and their acetylated derivatives are promising biomarkers for noninvasive cancer diagnostics, creating demand for robust enzymatic tools for their detection. In this study, we screened several phylogenetically diverse yeast polyamine oxidases and identified a new enzyme from Kluyveromyces marxianus (KmaPAO) as a promising candidate for analytical development. Among the tested candidates, only enzymes from K. marxianus and Lachancea thermotolerans were obtained in soluble active form, while only KmaPAO could be purified and characterized in detail. KmaPAO was produced in soluble active form in Escherichia coli at approximately 250 ± 40 mg of active enzyme per liter of culture, purified to near homogeneity in a single IMAC step, and showed a melting temperature of 66.6 ± 0.5 °C. The enzyme preferred acetylated polyamines and spermine, while showing lower activity toward spermidine. Kinetic analysis revealed sub-micromolar or low-micromolar Michaelis constants for several substrates and the highest catalytic efficiency toward spermine, 2.2 × 107 M-1 s-1. Due to its favorable expression level, stability, and substrate profile, KmaPAO represents a promising basis for the development of enzymatic assays for total polyamine determination.
Network pharmacology studies of medicine-food homology plants have identified broad injury response pathways and hubs that cannot support compound-level or Parkinson's disease (PD)-specific claims. We developed a traceable, non-weighted framework that separates regulatory provenance, PD-context evidence, structural support, and safety/developability liabilities. A ten-plant feasibility panel was locked before overlap with a 1631-gene PD union, yielding 382 plant-associated targets and 190 strict intersections. Leave-one-plant-out analysis retained 173-190 targets, whereas disease source and threshold stress tests showed curation dependence. Whole-blood classifiers showed modest five-fold discrimination (area under the curve, 0.606-0.682) and were excluded from candidate decisions. Redocking-validated AutoDock Vina and protein-ligand interaction fingerprints retained baicalein-MMP9, baicalein-AKT1, and baicalein-BCL2 as caution-tagged follow-up pairs. Quercetin-MMP9 was retained as a liability-tagged comparator, while KCNH2 relations were safety-only. Because no biological validation is presented, these pairs remain hypotheses for prospective MPP+-treated SH-SY5Y testing with orthogonal injury, dopaminergic phenotypes, target dependency, material confirmation and safety controls. Baicalein remains source-pending for the material chain.
Microalgae are recognized as sustainable biofactories for metabolites relevant to nutraceutical, pharmaceutical, and marine drug applications. Nannochloropsis salina is notable for its production of bioactive lipids, including the pharmaceutically relevant omega-3 eicosapentaenoic acid (EPA). Here, we evaluated orange peel extract (OPE), a citrus by-product, as a substrate for mixotrophic cultivation of N. salina within a bioeconomy framework. OPE supplementation triggered dose-dependent physiological responses. Among the tested OPE concentrations, 5% supplementation resulted in the highest total fatty acid content, increasing fatty acids from 24.4% (control) to 30.6% and EPA from 6.4% to 9.8%, whereas 10% OPE maintained biomass pigmentation. Higher OPE levels enhanced antioxidant potential, raising total antioxidant capacity from 4.6 (control) to 6.7 mg/g vitamin C equivalents (with 10% OPE), while the highest concentrations induced metabolic stress, reducing biomass and altering lipid composition. Fourier transform infrared spectroscopy analyses revealed biochemical adjustments consistent with metabolic reorganization, including increased intensities of the amide I and II bands associated with protein-rich structures. Overall, OPE emerges as a cost-effective supplement capable of modulating the biochemical quality of N. salina while valorizing agro-industrial residues. The increases in EPA and antioxidant capacity support the potential of OPE-supplemented cultures as a platform for the production of marine-derived bioactive compounds.
Filter-feeding bivalves can accumulate fecal bacteria and antimicrobial resistance determinants, yet routine regulatory monitoring relies mainly on quantitative Escherichia coli criteria. This study provides a whole-genome characterization of an ExPEC-like E. coli ST536 (O21) isolate recovered from commercially depurated retail mussels originating from a Class B harvesting area in Portugal. The isolate was phenotypically identified by MicroScan WalkAway Plus and subjected to antimicrobial susceptibility testing and Oxford Nanopore long-read sequencing, followed by genome assembly, annotation, antimicrobial resistance and virulence screening, plasmid and prophage detection, and phylogenomic analysis. Phenotypically, only resistance to tobramycin was detected using the MicroScan WalkAway system. Genomic analysis revealed a complex mobile element-rich architecture, including multiple resistance-associated determinants, 116 virulence-associated loci, prophage regions, and a mobilizable IncY plasmid carrying CTX-like class A β-lactamase-associated hits detected by ABRicate. Despite its ST536 assignment and ExPEC-associated traits, phylogenomic analysis placed the isolate within a phylogroup A/commensal-like background rather than among classical B2 ExPEC lineages, suggesting acquisition of pathoadaptive modules by an atypical environmental lineage. These findings highlight the potential utility of WGS as a complementary tool and suggest that depurated bivalves may serve as sentinels, warranting further investigation.
A previous study demonstrated that quorum sensing (QS) genes from the psychrophilic bacterium Aliivibrio logei can be used in Escherichia coli to obtain bacterial cultures with controlled amounts of a target protein. However, the QS system may be induced non-uniformly across the cell population. In this study, we used an expression vector containing luxR/luxI regulatory genes from A. logei carrying sfGFP as a reporter gene. Reporter expression was regulated by the autoinducer 3OxoC6-HSL, activated at 22 °C, and terminated at 37 °C. Flow cytometry was used to assess the GFP fluorescence distribution at the single-cell level. The system provided dose-dependent unimodal expression lacking formation of distinct ON/OFF subpopulations, while the robust coefficient of variation decreased with increasing autoinducer concentration. The autoinducer synthase LuxI enabled autoinduction, but under the tested conditions, no substantial effect on expression homogeneity was detected. Raising the temperature to 37 °C effectively halted expression, allowing intermediate target protein values to be fixed at the single-cell level. Overall, the developed system represents a promising tool for biotechnological applications requiring precise and uniform control of expression.
Chilli peppers (Capsicum species) have been widely used around the world because of their economic value and distinctive sensory characteristics. They contain abundant functional metabolites, especially a group of vanillylamide compounds belonging to the family of capsaicinoids, which have been exploited for medicinal, nutritional, agricultural, and cosmetic uses. The demand for capsaicinoid molecules is increasing day by day due to their high economic value and wide range of applications. Therefore, increasing bioactive metabolites, especially capsaicinoids in chilli peppers, is a major priority in the current scenario. Multi-omics approaches such as genomics, transcriptomics, proteomics, and metabolomics have substantially contributed to understanding the complex regulatory networks governing capsaicinoid biosynthesis. Key structural genes, transcription factors, and signaling pathways involved in the phenylpropanoid and branched-chain fatty acid pathways have been identified, providing valuable targets for metabolic engineering in chilli pepper. Despite these advances, the integration of genetic modification approaches for the targeted enhancement of capsaicinoid production remains limited in chilli pepper. Recent developments in biotechnology, particularly CRISPR/Cas-mediated genome-editing, enable the precise genetic modification of metabolic pathways and regulatory networks in plants. Therefore, it can contribute to the precise modification of key genes involved in the capsaicinoid biosynthesis pathway, offering potential strategies to enhance the capsaicinoid content in chilli pepper. However, CRISPR/Cas-mediated genome editing in chilli pepper is still in its early stages. There are currently no reports available on the successful enhancement of capsaicinoid content in chilli peppers through CRISPR/Cas-mediated genome editing. To date, no comprehensive review has evaluated the CRISPR-Cas-mediated genome-editing approaches for capsaicinoid metabolic engineering in chilli pepper. This review critically evaluates the recent advances in CRISPR/Cas-mediated metabolic engineering in chilli peppers, with particular emphasis on regulatory genes involved in capsaicinoid biosynthesis. Furthermore, multi-omics approaches are expected to complement these strategies by enabling the identification of key regulatory genes, the optimization of genome-editing targets, and the prediction of metabolic outcomes for enhanced capsaicinoid production. Overall, this review provides insights into improving capsaicinoid accumulation in chilli peppers through advanced genome-editing technologies.
Microalgae-fungal pellets have been studied as a versatile and robust biotechnological platform, offering significant advantages for microalgal biomass harvesting, wastewater treatment, biofuels production and/or obtaining of value-added products. This review presents an integrated analysis of the mechanisms governing the formation, stability, and functionality of these systems, combining physicochemical, biological, and mathematical modelling approaches and aims to describe the current state of the art and main research needs. The aggregation process is strongly influenced by the complementarity of the surface properties of microalgae and filamentous fungi, including electrostatic interactions, production of extracellular polymeric substances (EPSs), and modifications in surface roughness. Recent advances in multiscale characterization techniques, such as confocal microscopy, micro-computed tomography, atomic force microscopy, and X-ray photoelectron spectroscopy, have allowed a more precise elucidation of the internal architecture and surface chemistry of the pellets. In parallel, biological characterization through enzymatic assays, oxidative stress biomarkers, and photosynthetic activity analyses has provided relevant information on the metabolic responses and functional resilience of the consortium. Additionally, the incorporation of mathematical flocculation models can contribute to the prediction of pellet growth, density, and stability, supporting process optimization and application. The understanding of these interaction phenomena is important for the design of high-yield and efficient systems, including their development and validation, to expand the use of microalgae-fungal pellets in bioprocesses, as evidenced by this review.
Helicoverpa armigera is one of the most destructive polyphagous pests, yet the transcriptional organization underlying its digestive capacity remains poorly resolved. Here, we compiled 579 publicly available RNA-seq libraries representing 54 independent experiments and quantified transcript abundance across tissues and developmental stages. This complete dataset was used to support broader tissue-level expression profiling. After metadata harmonization and quality filtering, a subset of 130 biologically comparable libraries from five tissue/developmental categories was retained for weighted gene co-expression network analysis. WGCNA identified four biologically informative modules, among which the turquoise module was positively associated with fourth- and fifth-instar larval midgut samples. Independent expression profiling revealed strong midgut-biased expression of several trypsin- and chymotrypsin-like serine proteases, although only a subset of these genes was assigned to the turquoise module. Descriptive functional annotation of this module identified 202 co-expressed loci, including digestive enzymes, nutrient transporters, detoxification-related proteins, epithelial components and putative transcriptional or signaling-associated genes. Phylogenetic analyses and manual inspection of genomic locations further showed that several digestive protease genes occur in local clusters and have closely related counterparts in H. zea, suggesting partial conservation of local genomic organization. Collectively, these results describe a midgut-associated co-expression module containing genes associated with digestive, absorptive and protective functions and provide candidate genes for future functional studies.
Sophorolipids (SLs) produced from Starmerella bombicola using four different secondary substrates such as refined, bleached, and deodorized palm olein (RBD PO), RBD palm kernel olein (RBD PKO), RBD coconut olein (RBD CO) and fatty acid methyl ester (FAME) waste are reported. Their interfacial characteristics at medium-chain triglyceride (MCT) oil-water interface and ability to form nano/submicron emulsions were studied. The effects of SLs from different sources, SL concentrations and blend ratios of SLs and soybean lecithin on characteristics of emulsions produced by ultrasonication were examined. Initially, emulsion formed using SLs coded (from F2 to F5) showed large droplets (d32 > 1000 nm) and poor stability. They were then blended with soybean lecithin at a ratio of 3:1 to produce emulsions coded F6 to F9 with smaller droplets (d32 < 400 nm) and great stability over a range of temperatures (from 40 °C to 90 °C) and pH values (from 3 to 9). However, highly acidic (pH 2) and low ionic strength (1 mM NaCl) processing caused the separation of the emulsions. These emulsions also displayed potential antimicrobial activities towards Bacillus cereus and Pseudomonas aeruginosa, as well as cytotoxic effects against the human epithelial colorectal adenocarcinoma cell line (Caco-2). These results illustrated that stable emulsions required a mixture of SLs and soybean lecithin.
Cellular identity determination and lineage tracing is a pivotal technique in modern biological research. Conceptually simple yet efficient cell labeling techniques offering broader applicability are warranted despite many methods requiring intricate construction procedures. One such technique is ADAR (adenosine deaminase acting on RNA)-mediated RNA sensing, a live cell labeling technique that is based on the expression and abundance of cell-type-specific RNAs. Here, we utilized the optimized version, CellREADR (Cell access through RNA sensing by Endogenous ADAR), to establish a feasible tracing system for mouse spermatogonial stem cells (mSSCs) which are refractory to CRISPR-based reporter gene knock-in. We identified several previously unreported features of CellREADR, including RNA interference induced by double-strand RNA formation that is obligatorily generated during the normal operation of the CellREADR system. More importantly, we established its application for long-term labeling of mSSCs and monitoring mSSC differentiation induced by retinoic acid (RA) treatment in vitro. This work offers a practical solution for dynamic monitoring of mSSC self-renewal and differentiation and supports that CellREADR can be developed into more versatile and efficient tools in stem cell research.
Betalains are water-soluble pigments containing nitrogen, and they exist naturally in the plants of the order Caryophyllales. They have gained increasing attention in recent years because of their intense colours, antioxidant activity, and safety, thus making them suitable replacements for artificial dyes. The increasing interest in natural pigments has led to intensified research on betalain biosynthesis and optimization of pigment production. Nonetheless, their application in industry faces limitations, such as their low natural occurrence, sensitivity to environmental conditions, and instability during manufacturing and storage. Unlike previous reviews that primarily focused on betalain chemistry, biosynthesis pathways, or biological activity, the present review highlights recent developments in the engineering of the biosynthesis pathways, synthetic biology, elicitation approaches, omics-based pathway identification, and nanobiotechnology for betalain pigments. Special attention is paid to the comparison of plant, plant cell, yeast, and bacterial production systems, as well as recent advancements towards industrial production of betalain pigments and bottlenecks in the commercialization of sustainable betalain bio-factories.
Background: Adeno-associated virus (AAV) vector-mediated gene transfer is an emerging treatment strategy for severe cardiac disorders with genetic etiology. We refined the AAV toolkit to achieve efficient and selective expression of the therapeutic transgene in mouse hearts. Methods: Using vectors with a reporter transgene, we evaluated AAV administration routes, AAV serotype tropism to the myocardium, and cardiospecific promoters. Results: We showed that systemic AAV administration provides potent delivery and uniform transduction of cardiac tissue, outperforming localized injection techniques. The MyoAAV 2A capsid variant enabled an improved heart-to-liver transduction ratio compared to the parental AAV9 serotype. Screening a panel of cardiac and pan-muscular promoters in vitro and in vivo verified the superiority of the cardiac troponin T (cTnT) promoter for robust heart-specific transgene expression. Finally, we demonstrated that the cumulative properties of systemic AAV delivery, the MyoAAV 2A serotype, and the cTnT promoter allowed for efficient cardiac synthesis of the therapeutic transgene—an artificial miRNA designed for the gene suppression strategy of FLNC-related cardiomyopathy. Conclusions: Our findings establish an effective AAV approach for transgene transfer into the mouse heart and promote the development of gene therapy for cardiac disorders.
Vital pulp therapy aims to preserve pulp vitality by stimulating reparative processes. However, conventional approaches often result in incomplete tissue regeneration. Extracellular vesicles (EVs) have emerged as promising cell-free therapeutic agents because of their ability to regulate angiogenesis, odontogenesis, and immune responses through the transfer of bioactive molecules. Despite their significant regenerative potential, the clinical application of EVs remains limited by rapid clearance, insufficient local retention, and uncontrolled release following administration. To address these challenges, various extracellular matrix (ECM)-based and ECM-bioinspired scaffolds have been developed as delivery platforms. These scaffolds can provide structural support and enable controlled, localized release of EVs. This narrative review critically evaluates the current evidence regarding scaffold systems as EV delivery platforms for dental pulp regeneration, comparing their biological performance, methodological quality, and translational potential. Across the available studies, scaffold-assisted EV delivery consistently enhanced angiogenesis, odontogenic differentiation, mineralization, and immunomodulation; however, the evidence remains preclinical and is characterized by substantial heterogeneity regarding EV source, isolation and characterization methods, scaffold composition, experimental models, and outcome assessment. Current findings support the feasibility of scaffold-assisted EV delivery for regenerative endodontics, but important challenges remain, including standardization of EV production and characterization, scalable manufacturing, regulatory approval, and demonstration of long-term safety and functional pulp-dentin complex regeneration. Further well-designed translational and clinical studies will be essential before routine clinical implementation can be considered.
Biosafety Level 3 (BSL-3) laboratories are essential for handling high-risk pathogens and strengthening global health security. This study presents a patent landscape analysis of BSL-3-related technologies using the Derwent World Patents Index (DWPI) to identify technological trends, geographic distribution, patent classifications, and temporal evolution. Patent documents associated with laboratory infrastructure, ventilation systems, containment devices, and biosafety procedures were screened and analyzed. A total of 58 patent documents filed between 2009 and 2024 met the inclusion criteria. The results showed that China and the United States are the leading contributors to BSL-3 patent development, reflecting continued investments in biosafety and biosecurity infrastructure. The most frequent International Patent Classification (IPC) categories were C12M (microbiological devices), E04H (specialized construction infrastructure), and F24F (ventilation and air control systems), highlighting the multidisciplinary nature of innovations in laboratory containment and safety. The temporal trends revealed increases in patent activity following major public health emergencies, including SARS, Ebola, and particularly the COVID-19 pandemic. Furthermore, a significant increase in patent expirations is expected by 2029, creating opportunities for technology transfer, open innovation, and broader access to critical biosafety technologies. These findings emphasize the strategic importance of continued investment in BSL-3 technologies, especially in developing countries with growing biosafety demands.
Global food waste management necessitates circular bioeconomy solutions to transform organic residues into high-value nutrients to address nutritional demands. This study investigated the valorization of two abundant waste streams, stale bread and sunflower oil through solid state fermentation using food-grade filamentous fungi. Three strains, Neurospora intermedia, Aspergillus oryzae and Rhizopus oryzae were evaluated for the bioconversion of stale bread. Oil supplementation levels of 10, 20 and 30% (g/100 g dry matter) using both fresh and spent sunflower oil were tested to assess changes in proximate composition, characterizing fungal growth dynamics and mycelial development. Furthermore, modifications in fatty acid profiles and hydrolytic enzyme activities were analyzed to determine species responses to oil source and concentration. The results demonstrated that N. intermedia achieved peak protein levels of 36% (g/100 g) alongside efficient starch catabolism, while 10% fresh oil supplementation induced a significant protein increase (26%) in A. oryzae. Regarding lipid accumulation, 10% spent oil supported higher fat content in R. oryzae (19%) compared to fresh oil (17%). PUFA/SFA ratio reached its maximum in A. oryzae with the highest of 5.91 ± 0.56 under 10% fresh oil. Enzymatic analysis identified A. oryzae as the most efficient lipase producer, reaching a maximum activity of approximately 0.10 U/g at 10% spent oil supplementation. Conversely, R. oryzae lipase activity peaked at 20% supplementation (0.08 U/g), reflecting its high capacity for lipid accumulation. These findings establish a potent bioprocess for upcycling mixed food wastes into enhanced functional ingredients for sustainable food and feed systems.
