Ancestral sequence reconstruction (ASR) offers a revolutionary approach to resurrect functional proteins, yet its potential in transporter engineering remains underexplored. Here, we pioneered the application of ASR to reconstructing ancestral xylose transporters, addressing the persistent challenge of glucose-mediated inhibition of xylose uptake in Saccharomyces cerevisiae during xylose co-fermentation. Through rigorous ASR analysis, we reconstructed ancestral xylose transporters (Xt) and selected two candidates—Xt3 (approximately 140 million years old) and Xt7 (approximately 40 million years old)—based on their phylogenetic positioning, degree of sequence divergence from extant homologs, and predicted structural integrity. Functional characterization demonstrated that both Xt3 and Xt7 significantly enhance xylose uptake efficiency and mitigate glucose-induced repression. In fermentation experiments with mixed sugars (40 g/L xylose and 40 g/L glucose) within 72 h, recombinant S. cerevisiae expressing Xt3 achieved 22.75 g/L xylose consumption, surpassing the benchmark N326F^Xltr1p (16.22 g/L) by 40.27% and outperforming Xt7 (21.36 g/L) by 6.51%, highlighting Xt3 as the most efficient transporter. Molecular docking suggested a potentially more favorable binding mode for xylose in the ancestral transporters (binding affinity: −3.68 kcal/mol for Xt3 vs. −3.15 kcal/mol for N326F^Xltr1p). Molecular dynamics simulations further demonstrated that the ancestral transporters formed complexes with xylose that exhibited faster convergence to a stable state and maintained significantly greater conformational stability throughout the simulation compared to the N326F^Xltr1p complex. These computational insights provide a plausible structural basis for their enhanced performance. This work contributes to the advancement of lignocellulosic biorefinery technology and provides a practical reference for resurrecting other valuable proteins using ASR’.

Natural exogenous additives (EA) suitable for the tobacco fermentation need to be developed to enhance the fermentation quality and economic value of low-grade cigar tobacco leaves (CTLs). This study analyzed the impacts of three compound Chinese herbal medicine (CHM) on metabolites and microorganisms during CTLs fermentation. The results manifested that EA facilitated the degradation of total sugar, starch and protein, while enhancing the accumulation of reducing sugar in CTLs. Furthermore, EA raised contents of free amino acids (FAAs), while Asp, Glu, Ser and His were found to be key differential FAAs of CTLs. During fermentation, the total contents of volatile flavor components (VFCs) initially increased and then declined. Furthermore, EA contributed to more harmonious compositions of VFCs by promoting the formation of neophytadiene, ketones, esters and aldehydes, as well as facilitating nicotine degradation. According to variable importance in the projection (VIP) > 1 and odor activity value (OAV) > 1, 7 key differential VFCs were identified. EA enhanced positive microbial interactions and led to a more stable and coordinated symbiotic network. Linear discriminant analysis effect size (LEfSe) identified 9 genera as differentially dominant microorganisms in CTLs, which were closely associated with chemical compositions and key differential flavor metabolites. In addition, EA promoted cigar tobacco characteristics (CTCs) by altering bacterial alpha diversity and influencing the assembly of dominant microbial communities. Overall, this study offered theoretical insights into the innovative applications of CHM in CTLs fermentation, and presented new perspectives for enhancing CTLs quality and customizing flavor profiles.

Escherichia coli, a bacterium indicating improper hygiene practices during food production, is commonly found in the intestines of humans and animals, while Salmonella spp. are dangerous bacteria that cause typhoid fever and severe diarrhea. These pathogens have been found in fresh vegetables. This study investigated how the vegetable surface characteristics influenced bacterial adhesion. The reduction of bacteria during the washing process was assessed using different concentrations and types of chemicals. The relationships between variables obtained from image analysis techniques and bacterial adhesion on vegetable surfaces were also evaluated. The most effective way to inhibit bacteria was by washing with 2.0% lactic acid, with bacterial reduction from an initial concentration of 8.74 to 2.92 log CFU/m². Pearson’s correlation with the highest r value was surface area (A) with values ranging from 0.764 to 0.993, followed by surface roughness (R) with values between 0.019 and 0.986, and Fractal dimension (FD) with values between − 0.510 and − 0.992. The correlation between A and the number of bacteria (E. coli and Salmonella) was the highest, with surface area influencing bacterial adhesion to the vegetable surface. Greater surface roughness was associated with a higher initial bacterial load, making the A value a good predictor of changes in bacteria during washing with organic acids at various concentrations.

Synthetic chemical seed treatments, while effective, often raise significant environmental and health concerns. These concerns stem from the use of hazardous chemicals such as fungicides and insecticides that, besides posing risks to workers, have broader environmental impacts. These hazardous chemicals can leach into the soil and water systems, disrupting ecosystems, harming beneficial organisms, and entering the food chain. Agro-industrial byproducts/wastes (AIBWs) represent an abundant, environmentally friendly resource with potential for seed treatments. We focused on AIBWs that are produced in enormous amounts and do not pose potential hazards since they are commonly used to feed animals as well as food additives for humans, including wheat bran (WB), wine pomace (WP), and brewer’s spent grain (BSG). We investigated the effects of imbibing wheat seeds in water-soluble extracts of AIBWs or coating seeds with a biopolymer supplemented with AIBW substances on wheat growth and reproduction. As controls, we used water-soaked (WS) and non-soaked (NS) seeds, as well as chemically Celest Top-coated seeds. Petri dish assays showed that seeds imbibed in AIBW extracts exhibited enhanced post-germination growth as compared to NS seeds. Thus, while 81% of NS seedlings produced up to 3 seminal roots (SRs), 84% of WB and 64% of Celest Top seedlings produced 4 and 5 SRs. Net-house experiments revealed that Celest Top and AIBW extracts had a positive effect on reproduction as compared to NS, displaying 17.4%, 14.5%, 30.3%, and 34.3% increases in grain weight per spike in Celest Top, WB, GP, and WP, respectively. Metabolic analysis of seeds derived from treated plants revealed variation in metabolite profiles with a notable increase in the amino acid tryptophan. We utilized the nature-sourced polysaccharide carboxymethylcellulose (CMC) to coat seeds with AIBW substances derived from GP, referred to as CMC-GP. The results indicated that CMC-GP and Celest Top enhanced root growth, displaying 2- and 1.5-fold increases in fresh and dry weight, respectively, as compared to NS and CMC-coated seeds. Thus, AIBWs appear to provide cost-effective, eco-friendly alternatives to the hazardous chemical seed coatings, whether applied via imbibition or coating, while aiding in waste valorization within the circular economy.

Baiying Juhua Decoction (BYJHD) is a well-established traditional Chinese herbal formula primarily composed of Solanum lyratum and chrysanthemum, which necessitates a thorough investigation to clarify its mechanisms in combating non-small cell lung cancer (NSCLC). This study employed a combination of network pharmacology predictions, serum pharmacochemistry analysis, and various machine learning algorithms (including LASSO, SVM-RFE, and RF) to identify 38 bioactive compounds that target 653 proteins associated with NSCLC. A cross-analysis of 2161 differentially expressed genes (DEGs) and 3124 functional modules led to the identification of 54 critical therapeutic targets. Following this, protein-protein interaction (PPI) and machine learning analysis pinpointed five key signaling regulators. Molecular docking studies demonstrated strong binding affinities between four representative compounds from BYJHD and these targets. Both in vitro and in vivo experiments confirmed that BYJHD inhibits the progression of NSCLC by exerting anti-angiogenic effects, specifically through the inhibition of the ACVRL-1/Smad/ID-1 signaling pathway and the downregulation of CD34. These findings effectively connect traditional clinical applications with contemporary mechanistic insights, positioning BYJHD as a promising multi-target therapeutic candidate for NSCLC.

Conventional soybean processing relies on hexane extraction and high-temperature treatments, which cause protein denaturation and hinder the separate recovery of oil, protein, and carbohydrate. To address this limitation, this study aimed to establish a sustainable enzymatic soybean processing (ESP) strategy for separate collection of all three major components: intact oil bodies, undenatured protein, and monomerized carbohydrate. This is the first demonstration that ESP efficiency can be significantly enhanced by integrating pulsed sonication. Multi-enzyme systems were produced via solid-state fermentation (SSF) of soyhull by Aspergillus niger and applied to cracked soybean particles. Screening of 15 SSF enzyme extracts revealed that pectinase, polygalacturonase, and invertase were the limiting carbohydrase activities for cell wall degradation. Effects of processing variables including protease activity, reaction media, and reaction time were evaluated to minimize protein loss. Using water instead of citrate buffer as reaction medium and limiting processing time to≤48 h reduced protein dissolution to below 20%. ESP was further enhanced through pulsed probe-sonication (12 W/mL, 1 s on/23 s off), which reduced processing time to 24 h while increasing carbohydrate solubilization to up to approximately 90% depending on enzyme loading. Simple centrifugation enabled efficient fractionation into intact oil bodies (100%), native proteins (≈70%), and hydrolysates containing soluble proteins (≈30%) and monomerized carbohydrate (≈90%). These findings demonstrate an integrated enzymatic-sonication approach that enables hexane-free, low-temperature soybean processing with minimally denatured, high-value products and offers a pathway for sustainable soybean biorefinery.

To explore the bioaugmentation mechanism of biogas-production promotion and risk reduction of antibiotic-resistance genes (ARGs) in the anerobic co-digestion of cattle manure and rice straw with biochar addition, the performances of digestion and productivity with different amounts of biochar additions (0, 1.25, 3.75 and 5 g·L⁻¹ slurry) were studied. The results indicated that, biochar addition could effectively promote biogas production, and the cumulative methane (CH₄) yields from the treatments with 3.75 g·L⁻¹ slurry and 5 g·L⁻¹ slurry biochar additions were18.7 times and 14.8 times of CK (0 g·L⁻¹ slurry), respectively. Combined with Fourier transform infrared spectroscopy (FTIR) analysis and high-throughput sequencing-based microbial community quantification, the methanogenesis was enhanced through three possible pathways: (1) the porous structure and aromatization characteristics of biochar could promote destruction of cellulose bundle structure, thereby promoting the hydrolysis of lignocellulosic substrates; (2) Biochar regulated volatile fatty acid (VFA) concentrations within an optimal range, enhancing the buffering capacity of the anaerobic digestion system; (3) The low-dose biochar (1.25 g·L⁻¹ slurry) achieving the optimal ARG risk mitigation by suppressing the primary ARG host Bacteroidota.

Daqu is a critical saccharifying and fermenting agent in the traditional solid-state production of strong-aroma Baijiu, with its physicochemical properties and microbial communities playing a vital role in both starter quality and final aroma. This study analyzed twelve Daqu samples from four key production regions in Sichuan Province—Luzhou (L), Suining (S), Yibin (Y), and Zigong (Z)—assessing physicochemical parameters such as moisture content, acidity, saccharification, liquefaction, and esterification activities. Microbial communities were characterized using Illumina high-throughput sequencing. Regional variations in physicochemical properties were observed, with Y showing the highest acidity, saccharification, and esterification activities. While no significant regional differences in alpha-diversity were found, β-diversity analysis (PCoA) revealed distinct microbial structures. Bacteria were predominantly represented by Bacillus, Lactobacillus, and Weissella, while Aspergillus, Saccharomycopsis, and Rhizopus dominated fungi. LEfSe analysis (LDA≥4.0) identified region-specific microbial shifts. PICRUSt2 and FUNGuild predictions indicated significant potential for amino acid and carbohydrate metabolism, with saprotrophic fungi being dominant. Correlation analysis highlighted Thermoactinomyces, Weissella, Bacillus, and Rhizopus as key microbes influencing saccharification, starch degradation, and esterification. These findings provide insights into microbial and biochemical factors driving regional differences in Daqu, offering a foundation for standardized production methods and quality control.

Heparin, mainly used as an anticoagulant, has also shown potential in the treatment of diseases such as inflammation and cancer. Currently, heparin is mainly extracted from the intestinal mucosa of pigs. However, due to concerns about disease transmission and contamination associated with animal-derived products, biomanufacturing techniques have been explored as alternative production methods. Through enzyme engineering, metabolic engineering, and synthetic biology approaches, the heparin biosynthetic pathways have been systematically optimized. The main biomanufacturing techniques include in vivo/in vitro combination strategy (microbial heparosan fermentation followed by chemoenzymatic modification) and de novo biosynthesis. This article comprehensively discusses the latest advancements, challenges, and future perspectives of these heparin biomanufacturing techniques.

Gardeniae Fructus (GF), the dried fruit of Gardenia jasminoides J. Ellis, has been used in East Asian medicine for centuries. Its carbonized form, Gardeniae Fructus Carbonisatus (GFC), is produced through processing, yet the effects of this transformation on active constituents and neuroprotective mechanisms remain unclear. This study aims to elucidate the key compositional changes induced by processing and explore their relevance to neuroprotective activity.

After obtaining GF and GFC extracts via CO₂ supercritical fluid extraction (SFE), UPLC-Q-TOF-MS/MS was employed for qualitative analysis of differential compounds. A pathology-specific network pharmacology screening approach, combined with UPLC-UV-DAD, was applied to quantify major bioactive differential components. Finally, in vitro models and molecular pharmacology techniques were utilized to validate the neuroprotective effects of key compounds.

We identified 23 differential compounds and quantified 10 key bioactive constituents. Integrated network pharmacology and quantitative analysis implicated neuroinflammation and ferroptosis in GF’s neuroprotection, with geniposide and crocetin as pivotal compounds. Mechanistic studies confirmed roles for TLR4/NF-κB and Nrf2 pathways.

Geniposide and Crocetin were identified as key compounds responsible for the neuroprotective effects of GF and GFC, primarily through the inhibition of neuroinflammation and ferroptosis. Crocetin is highlighted as a potential marker for GFC.

Processing transforms Gardeniae Fructus into GFC, enhancing glycoside–aglycone conversion and markedly increasing crocetin. Integrated network pharmacology and quantitative analysis reveal geniposide and crocetin as core neuroprotective agents. In vitro analysis, these compounds inhibit neuroinflammation and ferroptosis via TLR4/NF-κB suppression and Nrf2 activation, supporting crocetin as a characteristic marker of GFC.

This study aimed to extract chitosan (CS) from cuttlefish (Sepia pharaonis) bones (CB) and then chemically modify it to produce carboxymethyl chitosan (CMC). Marine cuttlefish skin collagen peptide (MCP) was then cross-linked with CMC to form a novel CMC-MCP complex. The physicochemical properties and biological effects of CS, CMC, MCP, and CMC-MCP were evaluated using human keratinocyte (HaCaT) cell lines. All materials showed cytotoxicity at high concentrations (100–1600 µg/mL), negatively affecting cell viability. At a lower concentration of 50 µg/mL, the materials were used to assess cell migration. Among them, the CMC-MCP complex significantly promoted cell migration. Additionally, CMC-MCP treatment led to increased expression levels of matrix metalloproteinases (MMP-2 and 9) and tissue inhibitors of metalloproteinases (TIMP-1 and 2), which are key regulators in the wound healing process. These findings suggest that the CMC-MCP complex has potential as an economical, safe, and effective biological dressing for promoting wound healing. Further studies are recommended to explore its interaction with other healing-related factors, such as nutrients and growth factors, to better understand its influence on various stages of tissue repair.

Microorganisms, particularly filamentous fungi, have become the dominant platforms for industrial enzyme production due to their rapid growth, low cost, and adaptability. However, current production technologies face limitations in yield and cost-efficiency, prompting the need for innovative enhancement strategies. Non-thermal atmospheric-pressure plasma has emerged as a promising tool for stimulating microbial enzyme production. In this study, we have employed micro-surface dielectric barrier discharge (MS-DBD) plasma, which operates in a completely different manner from jet plasma, and evaluated its potential for enhancing the production of cellulolytic enzymes in Neurospora crassa. The extracellular activity of cellulases increased (maximum 10.41±3.44% increase) after MS-DBD plasma treatment. The transcription levels of the four cellulase genes were significantly elevated (highest in the 120 s treatment). The fungal hyphal membrane was depolarized and chemically altered after plasma treatment. The levels of intracellular Ca²⁺ and nitric oxide (NO) were elevated, and a high-affinity Ca²⁺ influx system was activated after plasma treatment. Ca²⁺ channel inhibitors reduced fungal cellulase production by downregulating intracellular NO levels. Plasma-mediated enhancement of enzyme production seemed to occur at plasma energies below 500–600 J. However, the combination of the plasma source type and treatment time can affect the efficiency of enzyme production. We also observed the promotion of fungal cellulase production when jet plasma was applied to larger volume of fungal hyphae. Our results suggest that plasma may be a genetically and environmentally safe tool for fungal enzyme production on an industrial scale and can be applied to bioreactors.

L-(+)-Tartaric acid is a valuable organic acid with broad applications in the food, pharmaceutical, and chemical industries. Its eco-friendly synthesis typically relies on the enzymatic hydrolysis of cis-epoxysuccinate (CES) catalyzed by cis-epoxysuccinate hydrolases (CESHs), but conventional single-batch processes suffer from low space–time yields and poor continuity. To address these challenges, we devised two complementary fed-batch strategies to simplify the enzyme–product separation by exploiting differences in their solubilities. Strategy A employs carrier-free cross-linking immobilization of whole cells using 0.02% glutaraldehyde and 0.1% polyethylenimine. In this system, both the substrate sodium cis-epoxysuccinate (CESNa) and the product sodium L-(+)-tartrate remain soluble, while the enzyme is retained in the insoluble cell matrix. Under fed-batch operation, this configuration achieves a space–time yield of 150 g L⁻¹ h⁻¹. Strategy B uses cell-free extract of CESH to hydrolyze calcium cis-epoxysuccinate (CESCa) with inherently low solubility. Here, the enzyme is fully soluble but the L-(+)-tartrate formed precipitates as an insoluble calcium salt, allowing easy separation of the product from the reaction mixture. This approach overcomes potential substrate inhibition and minimizes sodium-ion discharge, delivering a space–time yield of 136 g L⁻¹ h⁻¹ and a specific productivity of 484 g_product/g_catalyst. Both the soluble-product/insoluble-enzyme system (A) and the insoluble-product/soluble-enzyme system (B) represent effective strategies to streamline downstream processing and markedly enhance productivity. Together, they offer a viable route to scalable and cost-effective industrial production of L-(+)-tartaric acid.

D-Pantothenic acid (DPA), also known as vitamin B₅, is a water-soluble organic acid, widely applied in foods, feeds, cosmetics, and medicines. Although numerous and rapidly developing cell factories have been established for DPA biosynthesis, there has been no report of any attempts to engineer Yarrowia lipolytica to synthesize DPA. To explore further possibilities in DPA biosynthesis, we tried to employ systematic metabolic engineering strategies to identify and break the potential bottlenecks in DPA biosynthesis by Y. lipolytica. By improving the rate-limiting steps of the DPA biosynthesis pathway, weakening the strongly competitive pathways, and enhancing the multiple cofactor supplies, a robust Y. lipolytica cell factory for DPA biosynthesis was successfully constructed. Consequently, the resulting strain DPA34 produced 2.18 g/L DPA in a 5-L bioreactor, representing the first report of DPA production to date in Y. lipolytica. This work is believed to facilitate the development of Y. lipolytica for sustainable manufacturing of vitamin B₅ and its derivatives.

This work investigates the green synthesis of silver nanoparticles (AgNPs) using mixed aqueous extracts of Azadirachta indica leaves and roots as natural reducing and stabilizing agents. The synthesis was optimized by varying extract concentration, pH, and temperature, and nanoparticle formation was confirmed by UV–Vis spectroscopy showing a characteristic surface plasmon resonance between 350 and 450 nm. Structural and morphological analyses {X-ray diffraction (XRD), scanning electron microscope (SEM), Fourier-Transform Infrared (FT-IR), particle size analysis} revealed predominantly crystalline, spherical AgNPs capped by phytochemicals like flavonoids, phenols, amide- and carbonyl-containing compounds. The phytochemical profile of the extract was further validated by Gas Chromatography-Mass Spectrometry (GC–MS) analysis. The biosynthesized AgNPs exhibited strong colorimetric sensing capability for heavy metals, showing noticeable spectral and visible color changes particularly in the presence of Hg²⁺, Pb²⁺, and Cd²⁺ ions. Antibacterial evaluation indicated significant inhibitory activity against Staphylococcus aureus (33±0.13 mm) and Escherichia coli (45±0.21 mm), outperforming standard gentamycin controls. These findings highlight neem-derived AgNPs as low-cost, eco-friendly nanomaterials with dual applications in environmental monitoring of heavy metals and antimicrobial therapy.

The employment of plant extracts for green production of bimetallic nanoparticles (BNPs) has gotten significant consideration because of its cheap, ecological, single–step, and easily scalable procedures. This methodology enables the manufacture of biocompatible nanoparticles (NPs) with improved activity. In this study, an environmentally friendly approach was utilized to biosynthesize manganese oxide–silver BNPs (MnO–Ag BNPs) using Cucumis melo (C. melo) peel extract (CPE), which served as the source of the required reducing and stabilizing materials. Several spectroscopic analytical methods, including ultraviolet–visible (UV–vis) spectroscopy, Fourier transform infrared (FTIR) spectroscopy, energy–dispersive X–ray (EDX) spectroscopy, X–ray diffraction (XRD), scanning electron microscopy (SEM), and transmission electron microscopy (TEM), were applied for careful confirmation and characterization of successful MnO–Ag BNPs assembly. This work introduces a novel green route employing CPE for MnO–Ag BNPs synthesis, providing distinct phytochemical efficiency and multifunctional bioactivity compared with previously reported plant–based systems. The biosynthesized MnO–Ag BNPs bacterial inhibitory capability as well as free radical scavenging effect were evaluated. Also, human kidney normal epithelial–derived cells (Vero cell line CCL–81) was employed for assessment of the cytotoxic outcome of MnO–Ag BNPs at various concentrations. Regarding the elemental composition, the manganese (Mn) and Ag contents were detected by the UV–vis, XRD, and EDX studies with consequent validation of MnO–Ag BNPs biosynthesis. The range of the assessed BNPs size was 2 to 10 nm with average diameter of 5.8 ± 1.7 nm and an average area of 22.7 nm². Analysis based on EDX technique revealed the presence of Mn and Ag metals with 23.7–46.6% of the atomic percentages and 32.2–28.0% of the weight percentages, respectively. The biosynthesized NPs showed strong free radical scavenging, achieving 85–90% inhibition at higher concentrations. The cytotoxic activity findings indicated no significant harmful effects, at concentration range of 31.25–250 µg/mL, on Vero cell line. Additionally, the viability of the tested cell line infected with herpes simplex virus type–1 (HSV–1) significantly increased from 43% (untreated) to 78–99% when treated with 125 µg/mL MnO–Ag BNPs and acyclovir, respectively. Moreover, the inhibition rates achieved against the tested virus were 73% for MnO–Ag BNPs and 99% for acyclovir. These outcomes highlight the potential of MnO–Ag BNPs as promising candidates for biomedical and antiviral applications.

Nemadectin, a milbemycin-class macrocyclic lactone antibiotic produced by Streptomyces cyaneogriseus, is a potent broad-spectrum insecticide with excellent environmental compatibility. Its derivative moxidectin, featuring a C-23 methoxime modification, demonstrates enhanced insecticidal activity and has become a commercially successful agrochemical. This study reveals ammonium regulation effectively boosts nemadectin biosynthesis in S. cyaneogriseus, with mechanistic insights gained through integrated multi-omics analysis. Transcriptomic profiling showed ammonium sulfate supplementation significantly upregulates the nemadectin biosynthetic gene cluster, including polyketide synthase (PKS) genes, backbone modification genes, and pathway-specific transcription factors, while also enhancing the expression of Avenolide-like signaling molecules and global transcription factor Afskne. Metabolomic dynamics revealed reinforced precursor biosynthesis through coordinated metabolic reprogramming: enhanced acetyl-CoA production, reinforced Embden–Meyerhof–Parnas pathway and amino acid/acyl-CoA metabolism, coupled with reduced tricarboxylic acid cycle activity. Systematic integration of physiological phenotyping, metabolite profiling, and transcriptional regulation data comprehensively elucidated the ammonium-driven overproduction mechanism, providing critical insights for developing advanced fermentation strategies and genetic engineering approaches in industrial antibiotic production.

Driven by waste resource utilization and carbon neutrality imperatives, this study synthesized cyanobacterial growth elicitor (CGE) and cyanobacterial-bamboo growth elicitor (CBGE) via acid-hydrothermal hydrolysis. The coupling potential of cyanobacterial biochar (CB) for improving rhizosphere soil and crop quality was investigated through four pot trial treatments: (1) CK (control), (2) BR (rhizospheric CB), (3) LBR (rhizospheric CB with CGE), (4) LZBR (rhizospheric CB with CBGE). Integration of phenotypic analyses, microbiome profiling, functional gene predictions, and risk assessment elucidated biostimulant mechanisms. Compared to CK, all treatments elevated soil nutrient levels. BR exhibited superior nitrogen enrichment (15±3 g/kg), while LBR and LZBR—particularly LZBR—enhanced phosphorus/potassium bioavailability and maximized soil organic carbon (SOC). LZBR treatment markedly increased Chryseobacterium abundance (an organic matter-decomposing genus). Functional verification confirmed enhanced C-N-P cycling activity, minimized environmental nutrient leakage, and improved plant nutrient assimilation. Specifically, LZBR increased soybean grain protein content by 37.0 g/kg and plant nitrogen accumulation by 5.4 g/kg compared to CK, and risk assessments indicated no detectable ecotoxicological effects. Consequently, the coupled application of CGE and CBGE derived from cyanobacteria and bamboo powder simultaneously improves soil quality and crop performance. This approach establishes a novel waste valorization pathway, suitable for partial replacement of chemical fertilizers and carbon emission reduction.

The dual crises of antimicrobial resistance and cancer demand innovative therapeutic platforms that overcome conventional treatment limitations. This study uniquely combines systematic Box-Behnken optimization of green-synthesized copper oxide nanoparticles from Thymus vulgaris with comprehensive evaluation of their synergistic antimicrobial and anticancer activities. HPLC profiling identified quercetin (55.92%), chlorogenic acid (15.33%), and gallic acid (12.28%) as principal phytochemical reducing and capping agents. Statistical optimization (R²=0.9886) established copper acetate concentration (F=670.48, p<0.0001) and incubation time (F=124.11, p<0.0001) as critical synthesis determinants, yielding monodisperse spherical nanoparticles (19–25 nm TEM; Z-average 119.2 nm, PDI 0.22; ζ-potential−45.8 mV). XRD confirmed a crystalline monoclinic CuO phase, while FTIR validated phytochemical surface functionalization. TE-CuONPs exhibited concentration-dependent bactericidal activity (MIC 250–950 μg/mL; MBC/MIC≤0.58) against Staphylococcus aureus, Pseudomonas aeruginosa, Escherichia coli, and Enterococcus faecalis as well as inhibition of biofilm formation in S. aureus and P. aeruginosa, with BIC₅₀ of 299 and 315 μg/mL, respectively. Critically, checkerboard assays revealed strong synergy with gentamicin (FICI 0.13–0.28), achieving eightfold dose reduction for both agents against S. aureus and P. aeruginosa. Time-kill kinetics demonstrated accelerated bacterial eradication, with combination therapy achieving≥3-log₁₀ reduction 8–12 h faster than monotherapies, a clinically significant advantage for acute infections. Furthermore, TE-CuONPs showed moderate antiproliferative activity (IC₅₀=117.26 μg/mL) against MCF-7 breast cancer cells, with limited selectivity over normal fibroblasts (SI=1.85), representing a sixfold enhancement over the crude extract. Additionally, Flow cytometric analysis revealed profound apoptotic induction, with 77.25% of cancer cells undergoing cell death (29.73% early apoptosis, 47.52% late apoptosis/necrosis). DPPH radical scavenging (IC₅₀=55 μg/mL) demonstrated a threefold superior antioxidant capacity versus plant extract alone. These findings advance the reproducible botanical nanoparticle synthesis and translational potential of plant-mediated nanomedicine for infectious disease management.

Oil palm trunks (OPT) represent an underutilized agricultural byproduct that poses significant environmental challenges. The effectiveness of OPT as feedstock for biochar production will be depends on carbonization conditions, yet the relationship between process parameters and biochar properties remains insufficiently explored. This study investigates the potential of converting OPT into micropores bioadsorbent through controlled carbonization. Biochar was produced at temperatures of 300, 400, and 500 °C, with residence times of 2, 3, and 4 h, and subsequently characterized for its physicochemical properties and adsorption capacity. The results indicate that biochar produced at 300 °C for 2 h exhibited the highest surface area (10.24 m²/g), while the carbon content peaked at 79.9% in biochar synthesized at 500 °C for 4 h. Notably, although the maximum surface area was observed at 300 °C for 2 h, superior MB removal (52.5%) at longer residence time (4 h) indicates that adsorption performance was governed primarily by surface functional chemistry and pore accessibility rather than surface area alone. The enhanced adsorption at mild carbonization was attributed to the preservation of oxygen-containing surface functional groups rather than surface area alone. Langmuir isotherm analysis provided the best fit (R²>0.9), yielding a maximum monolayer adsorption capacity of 3.57 mg g⁻¹ and a favourable separation factor (R_L<1). These results demonstrate that adsorption performance of OPT-derived biochar is governed by surface chemistry controlled through carbonization severity, positioning OPT as a promising low-cost precursor for sustainable dye adsorption applications.

Enzymatic synthesis is currently the primary method for preparing 1,3-medium chain- 2-long-chain triacylglycerols (MLM-TAGs), which serve as both a dietary component and clinical nutrient for specific populations. The application of MLM-TAGs is obviously constrained by the high cost of catalysts. Hence, a novel approach was proposed for MLM-TAGs production by engineered yeast. Overexpressing the mutated fas1^R1834K increased the production of medium-chain fatty acids (C8–C12) and resulted in an MLM content of 0.41 mol% of the total TAGs. The introduction of RnACSM4 enabled the recombinant to produce MLM-TAGs at a level of 4.2 mol% when supplemented with 0.2 mM sodium laurate. Further deletion of GAT2 and LRO1 increased the content of MLM-TAGs to 6.7 mol%. Iterative optimization involving sodium laurate dosage, culture temperature, and amino acid addition elevated the MLM-TAGs content to 34.4 mol%. Under the optimized conditions, the maximum yield of MLM-TAGs reached 18.5 mg/g DCW, representing a 135-fold improvement over the original strain. This research presents a promising and sustainable alternative for MLM-TAGs production and demonstrates the feasibility of tailoring the acyl composition of intracellular TAGs.

(+)-Bicyclogermacrene and its derivatives, with promising antimicrobial, anticancer, and insecticidal properties, hold significant potential for applications in pharmaceuticals, agriculture, and industry. However, traditional extraction methods from plant essential oils are unsustainable. In this study, we achieved the de novo biosynthesis of (+)-bicyclogermacrene using a metabolically engineered Escherichia coli strain. The biosynthetic pathway of (+)-bicyclogermacrene was partitioned into upstream and downstream modules to enable precise regulation. This was accomplished through the genome-integrated overexpression of the endogenous methylerythritol phosphate pathway to ensure an adequate supply of terpenoid precursors, which pulled the titer from the initial 11.3 mg/L to 50.1 mg/L. Production was further enhanced to 96.9 mg/L by fusion of downstream key genes to facilitate precursor channeling, along with expression level optimization to improve pathway efficiency. Additionally, NADPH supply was fine-tuned through overexpressing dehydrogenases to improve the overall metabolic balance and this approach achieved a titer of 119 mg/L. Following site-directed of (+)-bicyclogermacrene synthase, the engineered E. coli strain M6-36 produced 565 mg/L of (+)-bicyclogermacrene in a 5-L bioreactor, an approximately 50-fold increase from the initial. To the best of our knowledge, the obtained titer in this study represents the highest level ever reported for the production of (+)-bicyclogermacrene. This study demonstrates an effective approach for the heterologous biosynthesis of sesquiterpenoids in E. coli and provides a scalable platform for the sustainable production of terpenoid-derived valuable chemicals.

This study aims at improving the undissociated lactic acid production by Lactobacillus helveticus using a whey-based fermentation. It first describes the effect of pH on the ability of this bacterium to produce lactic acid, by considering final lactic acid concentration, production rate, volumetric productivity and sugar consumption. As a low performance was achieved at pH 4.3, an adaptive evolution of Lb. helveticus LH-B01 to acidic conditions was performed during continuous cultures of sweet hydrolysed whey. Two mutants have been isolated, which exhibited different characteristics. The mutant Lb. helveticus LH-B01-B4 displayed the higher maximal total lactic acid concentration (37.9 g/L), sugar consumption (82%) and volumetric productivity (0.39 g/L/h), when compared to the parental strain and the mutant Lb. helveticus LH-B01-A4. This performance was explained by the higher critical undissociated lactic acid concentration (10.1 g/L) of Lb. helveticus LH-B01-B4, compared with those of the parental strain (8.7 g/L) and the mutant Lb. helveticus LH-B01-A4 (7.5 g/L). From these results, the mutant strain Lb. helveticus LH-B01-B4 was the most promising option to produce undissociated lactic acid during low pH fermentation, thus making it suitable for industrial use as a descaling agent and biocide in detergents.

Based on the self-assembling properties of the SpyCatcher/SpyTag system and the structural advantages of Dps protein, this study successfully constructed a three-dimensional nano-enzyme cascade reactor (3DNECR) through the covalent self-assembly of SpyTag-ADH and SpyCatcher-Dps-ATA117 fusion proteins. The 3DNECR exhibited significantly enhanced catalytic efficiency compared to the two-dimensional control, attributed to optimized spatial organization promoting substrate channeling. The reactor exhibited remarkable storage, pH, and thermal stability. It maintained over 80% activity after 9 days of storage, showed superior pH tolerance across pH 8–10, and remained stable in the temperature range of 4–40 ℃. Molecular docking confirmed strong interfacial binding (− 17.3 kcal/mol) between assembly components and favorable substrate binding (− 7.4 kcal/mol) within the active site. Furthermore, the 3DNECR was applied to the asymmetric synthesis of (R)-1-[3,5-bis(trifluoromethyl)phenyl]ethanamine (R-BPA) in an oil-water biphasic system. Under optimized conditions, the 3DNECR consistently achieved high yields (99.9%) and excellent enantioselectivity (99.9%). The 3DNECR maintained a relative enzyme activity as high as 92% even after six cycles of reuse. This integrated platform showcases substantial potential for efficient and sustainable biocatalytic applications.

Filamentous algae, characterized by high cellulose content and absence of lignin, present a promising sustainable alternative to conventional plant and synthetic fibers. The present study systematically evaluated the suitability of freshwater filamentous algae as a new resource for textile fibers, targeting applications in moisture-absorbent textiles. Among twelve strains screened, the isolate Rhizoclonium sp. emerged as the most promising candidate due to its high biomass yield (1.04 g dry weight L^− 1) after 21 days of cultivation. In addition, it showed superior visible fiber flexibility following air-drying, an essential prerequisite for textile processing. Cultivation conditions were optimized (using WHM medium, pH 8, and thiamin supplementation) to maximize fiber quality, resulting in 8.6% increase in biomass productivity. Biochemical profiling of the optimized biomass revealed a significant enhancement of total carbohydrates (+ 18.0%), alongside reductions in protein (-18.4%) and ash content (-14.9%), supporting improved fiber durability and flexibility. Comparative FTIR analysis showed a strong cellulose signature and marked similarity to cotton, while also revealing high native starch content, further supporting their applicability as bio-based binders in nonwoven products. Functional characterization demonstrated that optimized Rhizoclonium sp. fibers exhibited exceptional moisture regain (~ 12%), surpassing conventional fibers such as cotton and lyocell. Overall, this study establishes native Rhizoclonium sp. as a highly versatile and renewable bioresource for innovative aquatic fibers, underpinning the development of an environmentally responsible algae-derived textile value chain.

As the terminal management for evaluating the engineering effectiveness of antibiotics production and utilization, the toxic effects of moxifloxacin (MOX) and trace concentration of Cu²⁺ (MOX-Cu) on Caenorhabditis elegans (C. elegans) were investigated at physiological, biochemical, and molecular level. Although the stimulate effects were observed after prolonged exposure (72 h) to MOX (0.2-2.0 mg/L), the expressions of HSPs, ace genes, and daf-16 were inhibited, indicating its adverse impact on cellular health, locomotion behaviors, and antioxidant defense of C. elegans. Similarly, the down-regulation of oxidative stress (sod-1 and daf-16) and cell damage (HSPs) related genes and the up-regulation of apoptosis-related genes (cep-1 and ape-1) indicated the oxidative stress and genotoxicity after prolonged exposure to MOX-Cu. For the chronic exposure (10 days) to MOX, the level of ROS was reduced due to the increased expressions of daf-16, sod-3, and hsp-16, accompanied with and the down-regulation of cep-1. Meanwhile, at the exposure to MOX-Cu, the levels of ROS and lipofuscin were decreased due to the up-regulation of sod-1 and daf-16, and the antioxidant defense was promoted and confirmed by the increase of amino acids and their related metabolic pathways. These results can provide a theoretical basis for the toxicity evaluation of typical antibiotics (MOX) that co-existing with trace heavy metals in natural environment media and bioresources processes.

Via efficient, scalable, and climate resilient processes, seaweed biorefineries can advance cleaner production, delivering nutrient-rich ingredients with relatively lower land and freshwater requirements. Here, a land-based Ulva ohnoi platform that integrates optimized outdoor tank cultivation, pH-shift protein extraction with dual-product valorization, scale-aware techno-economic analysis (TEA), and a climate-scenario emulator was developed and evaluated. Year-round cultivation yielded a pooled feedstock with carbohydrate and protein contents at 46.6% and 26.8%, respectively, and under optimal conditions, protein yield was 10.30% DW. Further, the protein content of cultivated Ulva protein (CUP) was 47.64%, with bound amino acids showing predominance (96%), implying food-grade utility. pH-shift extraction also improved in-vitro pepsin digestibility to 68.08%. The total dietary fiber content of CUP residue (CUPR) was 40.59%, with insoluble dietary fiber (31.59%) showing predominance, supporting the applicability of CUPR in improving gastrointestinal tract function. TEA revealed strong economies of scale for the platform. The 10,000-kg production model outperformed the 100- and 1,000-kg production models, with gross margin, Return On Investment (ROI), and Internal Rate of Return (IRR) at 78.56%, 110%, and 66.02%, respectively, and a payback time of only 0.91 years. Based on site-specific regression analysis (R² = 0.884), protein yield increased with temperature but decreased with rainfall, and further warming increased mean protein yield, while higher-emission pathways introduced rainfall-driven volatility, necessitating strategies for sustaining performance under climate variability. Overall, the use of the Ulva platform showed a broad growth temperature window as well as rapid acclimation for U. ohnoi, implying resilience even under global warming conditions and increasing weather variability.

Gut microbiota regulation is a key strategy for treating metabolic dysfunction-associated fatty liver disease (MAFLD). Arbutin (ARB) is a natural hydroquinone active agent with anti-inflammatory and antioxidant effects, as well as regulatory effects on the gut microbiota. However, its therapeutic effect on MAFLD and the responsible mechanisms remain unclear.

This study explored the therapeutic effect and mechanisms of ARB in MAFLD treatment.

High-fat diet (HFD)-fed mice served as the in vivo MAFLD model, and ARB treatment was given simultaneously. The extent of liver injury was assessed through histopathological staining. AML12 cells treated with free fatty acids served as the in vitro model. The effects of ARB were evaluated via oil red O staining and biochemical assays. Subsequently, we utilized bioinformatics techniques to predict the potential mechanisms and targets of ARB. The expression of liver apoptosis-related genes was detected using molecular biology techniques. Alterations in the gut microbiota were analyzed by 16S rRNA sequencing. Ultrahigh-performance liquid chromatography–high-resolution mass spectrometry was used to analyze the changes in fecal metabolite levels.

ARB treatment effectively improved liver injury in mice with MAFLD. Its mechanism was associated with anti-apoptotic effects mediated by signal transducer and activator of transcription 3. Meanwhile, ARB effectively reversed gut microbiota imbalance in mice with MAFLD and altered the composition of gut microbes and fecal metabolites.

ARB displayed potential effects in alleviating the pathology of MAFLD, exerting anti-apoptotic actions, and restoring the gut microbiota balance.

The enzymatic esterification of 5-hydroxymethylfurfural (HMF) with long-chain fatty acids offers a sustainable route for producing biolubricants and other high-value chemicals. This work evaluates the synthesis of 5-hidroxymethylfurfural stearate catalyzed by immobilized lipases in both batch and continuous packed-bed bioreactors, combining molecular dynamics (MD) simulations with experimental validation to identify suitable green solvents. Four solvents were tested: 2-methyl-3-buten-2-ol (2-MB), tert-butanol (TB), 2-methyltetrahydrofuran (2-MeTHF), and cyclopentyl methyl ether (CPME). MD simulations revealed that CPME increased hydrophobic surface exposure and flexibility near the catalytic site, favoring substrate accessibility. In preliminary experimental tests, CPME provided the highest conversion (50%). In batch bioreactor at 40 °C, 30 mM HMF and 250 mM stearic acid achieved 67% conversion with Candida antarctica lipase B (CALB), maintaining full activity (100%) over four reuse cycles. In continuous operation, using a single packed-bed bioreactor at 0.02 mL min⁻¹ yielded conversions above 50% (residence time ≈ 55 min), while connecting two packed-bed bioreactors in series increased conversion to over 90% and productivity to 0.094 h⁻¹, compared with 0.076 h⁻¹ for one column and 0.003 h⁻¹ in batch mode. Deviations from ideal plug flow were observed over time, attributed to substrate or product deposition and in-situ water formation shifting the reaction equilibrium. Overall, CPME proved to be an efficient and sustainable solvent for the enzymatic synthesis of 5-hydroxymethylfurfural stearate, demonstrating the feasibility of continuous operation and highlighting pathways for further optimization through improved immobilization or reactor design.

Agricultural residues like sugarcane bagasse and rice straw are rich in cellulose and xylan. Their efficient conversion into (oligo)saccharides and value-added products requires microbial cellulases and xylanases, but low enzyme yields and high production costs hinder industrial application. This study isolated Penicillium oxalicum UNN1, a high-xylanase-producing strain with an initial activity of 51.63 U/mL. Submerged fermentation conditions were optimized using different carbon/nitrogen sources to enhance enzyme production. The optimized xylanase activity reached 191.22 U/mL (sugarcane bagasse xylan as sole carbon source) and 142.32 U/mL (combined with Avicel), with filter paper cellulase activity of 0.76 U/mL. The crude enzymes exhibited optimal activity at pH 5.0 and 50 °C. Cellulase retained over 75% activity after 7 h at pH 4.0–6.0 (4 °C) or 40 °C (pH 5.0), while xylanase activity remained nearly unchanged, even after over 21 days of storage at 4 °C (pH 5.0). However, the half-life of xylanase was less than 1 h at 50 °C, though it exceeded 72 h at 40 °C (pH 5.5). 3–5 mM Ca²⁺ and Cu²⁺ strongly inhibited both enzymes. Crude enzyme addition (about 7 U cellulase and 1,400 U xylanase) effectively enhanced reducing sugar production from agricultural residues. Single-factor and response surface optimization yielded optimal hydrolysis conditions: 480 U/g sugarcane bagasse xylan of xylanase, hydrolysate pH of 5.5, hydrolysis temperature of 40 °C, achieving a maximum reducing sugar yield of 0.355 g/g dry biomass. This work demonstrates the potential of P. oxalicum UNN1 enzymes for efficient and stable saccharification of agricultural residues, offering a viable approach for their valorization and environmental management.

The yeast Komagataella phaffii is an emerging microbial host for the production of functional recombinant proteins. However, proteolytic degradation during fermentation often compromises product yield and stability, posing a major hurdle for industrial-scale applications. This study presents a strategy to enhance the production of recombinant humanized type I collagen (rhColI) by engineering a host strain with reduced protease activity. An initial production strain, CL1, was engineered using post-transformational vector amplification (PTVA), achieving a titer of 558.86 ± 20.05 mg/L in flask culture. Subsequent scale-up fermentation, however, revealed significant rhColI degradation. To address this, we systematically deleted 11 candidate endogenous protease genes. The knockout of a serine protease gene, KpSub2, resulted in the most pronounced improvement, elevating the rhColI titer to 1039.06 ± 34.08 mg/L, a 23.47% increase over the parent strain CL1. Furthermore, the purified KpSub2 protein, obtained from inclusion bodies in Escherichia coli, demonstrated broad proteolytic activity against various types of recombinant humanized collagens. This broad substrate specificity was consistent with the observation that KpSub2 deletion also mitigated the degradation of recombinant humanized type III collagen (rhColIII). Our findings establish KpSub2 as a key mediator of collagen degradation in K. phaffii and provide an effective engineering strategy for optimizing the production of collagen and other degradation-susceptible functional proteins in this host.

The aqueous phase (AP) generated during biomass pyrolysis is often considered a waste product due to its dilute and toxic nature, making it difficult to upgrade. This study explores the potential of using AP as a laccase inducer for the white-rot fungus, Pleorotus ostreatus, and as a mediator in laccase-catalyzed reactions. As an inducer, AP increased laccase production from P. ostreatus to 570 U/g, outperforming copper, a common inducer, by almost 180%. A maximum laccase yield of 955 U/g was achieved when P. ostreatus was co-induced by both AP and copper. Characterization of the AP-induced laccase revealed greater pH tolerance relative of this enzyme compared to copper-induced laccase. The AP-induced laccase was further evaluated for various applications. Laccase alone was effective in decolorizing coomassie blue dye, increasing saccharification yield from prairie biomass, and detoxifying tetracycline. When laccase was mediated with AP, the enzyme was also capable of decolorizing crystal violet dye, demonstrating additional benefit of AP to mediate laccase-based oxidation reactions with certain substrates. Overall, these findings suggest that using AP to induce laccase production, and potentially mediate the laccase-based reactions, could be a promising method to valorize this byproduct from biomass pyrolysis.

Climate change and environmental pollution are among the most pressing global challenges today, with water pollution standing out as a particularly critical issue. Industrial wastewater discharge, especially from distilleries, significantly contributes to the degradation of aquatic and terrestrial ecosystems. Molasses-based distilleries are major perpetrators, producing vast quantities of dark brown effluent known as spent wash. This colouration is largely due to the presence of melanoidin, a recalcitrant compound formed via the Maillard reaction. Although many distilleries now utilize anaerobic digestion to convert this organic-rich waste into biogas, the resultant biomethanated spent wash remains highly coloured and environmentally hazardous. Direct discharge of untreated or partially treated spent wash into rivers, lakes, or soil severely disrupts ecological balance and poses risks to biodiversity. Existing disposal practices, such as lagoon storage or composting with press mud, offer limited solutions to the colour problem. Fungi, particularly those producing laccase and other oxidative enzymes, have demonstrated promising potential for decolourizing spent wash in laboratory studies. However, the enzymatic pathways involved in melanoidin degradation are still not fully understood. To address the persistant colour challenge, integrated treatment strategies combining fungal systems with complementary physical or chemical processes (eg, adsorption or advanced oxidation) may be required to achieve effective decolourisation. Such advancements are vital for creating effective, eco-friendly solutions to mitigate the environmental impact of the distillery industry and promote a circular bioeconomy.

Allergic rhinitis (AR) is a prevalent inflammatory disorder of the upper respiratory tract, affecting 20–40% of the global population and severely impairing quality of life. Given the limitations and adverse effects associated with conventional pharmacotherapy, naturally derived bioactives with low toxicity are gaining prominence as alternative interventions. In this study, we developed a bioresource-based nanoemulsion (NE) by integrating Sargassum polysaccharides (SP) into algal oil (AO) to enhance intranasal delivery and therapeutic efficacy against AR. Structural analysis confirmed that SP comprised sulfated polysaccharides enriched in fucose, glucose, and galactose. The optimized SP–AO NE, formulated with Tween 80 and prepared via ultrasonic emulsification, exhibited uniform spherical droplets (53.4 ± 1.8 nm), a low polydispersity index (0.3 ± 0.1), and a negative zeta potential (− 29.1 ± 2.8 mV), indicating high colloidal stability and effective oxidative protection of AO during refrigerated storage. In an ovalbumin-induced AR mouse model, intranasal administration of SP–AO NE significantly alleviated nasal rubbing, epithelial hypertrophy, goblet cell hyperplasia, mast cell infiltration, and pulmonary inflammation. Intranasal SP–AO NE treatment decreased IgE levels in serum, nasal lavage, and bronchoalveolar lavage fluids, while enhancing mucosal IgA. In addition, SP–AO NE downregulated IL-4 and TNF-α expression and upregulated TGF-β1, demonstrating a robust immunomodulatory effect. Overall, this work presents a stable, biocompatible, and functional NE that improves intranasal delivery of algal bioactives, offering a promising natural therapeutic strategy for the management of AR.