Myofibrillar proteins (MPs) readily aggregate in low-ionic-strength media, limiting their solubility and emulsifying performance. MPs were examined across pH 3.0-11.0 in deionized water (ultra-low ionic strength) and high-ionic-strength medium (HIS, 0.6 M KCl, 50 mM phosphate), using high internal phase emulsions (HIPEs) as a functional model. MPs showed medium-dependent pH responses: in water, pH 3.0 and 11.0 improved dispersion and supported HIPE formation, whereas in HIS, pH 3.0 caused severe precipitation and pH 11.0 yielded the clearest solutions; maximal precipitation occurred at pH 5.0 (water) and pH 3.0 (HIS). These results indicate a pH-ionic strength complementarity mechanism. Under favorable extreme-pH conditions, MPs showed reduced particle size, structural unfolding, increased hydration, lower viscosity, and rapid formation of rigid interfacial films, enabling HIPEs with semi-solid rheology and improved stability. These findings support the design of pH-tunable, low-salt meat protein colloids.

Background Foodborne bacterial pathogens present a severe and persistent threat to global public health, attributable to their extraordinary adaptability to both environmental stresses and host immune responses. Small non-coding RNAs (sRNAs) have been recognized as pivotal post-transcriptional regulators in bacterial pathogens, playing essential roles in modulating virulence, stress adaptation, biofilm formation, and antibiotic resistance. Their dynamic expression patterns reflect pathogen responses across varied environments, spanning from food processing facilities to human hosts, yet an integrative framework linking these contexts through sRNA biology is lacking. Scope and approach This review is the first to synthesize sRNA research within a novel tripartite framework connecting food-processing-induced stresses, the adaptive evolution of foodborne bacterial pathogens, and sRNA-mediated regulatory networks throughout the infection cascade. We categorize sRNA types based on their genomic origins and mechanisms of action, with detailed discussion of their stage-specific functions including biofilm development, host colonization, antimicrobial defense, and precise regulation of virulence factors. We further explore the translational potential of sRNAs as novel diagnostic biomarkers and as targets for synthetic RNA-based antimicrobial interventions, while critically addressing persistent challenges such as multi-sRNA crosstalk, in vivo delivery efficiency, and off-target effects. Key findings and conclusion sRNAs are central hubs in the regulatory circuitry that enable rapid bacterial adaptation across the entire transmission continuum. Their integration into detection platforms and antibacterial strategies represents a paradigm shift towards intelligent food safety management. Future research leveraging artificial intelligence-driven network modeling, engineered nanovesicle delivery systems, and functional characterization in the viable but non-culturable (VBNC) state will be crucial to fully exploit sRNAs for predictive control and targeted mitigation of foodborne bacterial pathogens.

Cultured meat represents a promising sustainable food source, yet the development of efficient serum-free media remains a key bottleneck. Small molecules offer a cost-effective approach to system optimization. Through a functional screening of proliferation activators in bovine muscle stem cells (bMuSCs), we identified forskolin and formulated the serum-free “Beefy-F” medium. Over six passages, Beefy-F yielded 1.9 times more bMuSCs than the basal serum-free control (Beefy-9), matching serum-controlled levels while maintaining bMuSC morphology, myogenic gene expression, and differentiation potential. Subsequent synergistic screening revealed that the p38 inhibitor SB202190 enhanced forskolin’s effects. The optimized “Beefy-F + S” medium significantly outperformed both the basal serum-free control and single-supplemented formulations after three passages, upregulated the stemness marker PAX7, and preserved differentiation capacity. Transcriptomic analysis revealed that the Beefy-F + S medium broadly altered the bMuSC transcriptome to maintain myogenic identity, upregulate cell cycle genes, and reshape extracellular matrix (ECM) pathways, with forskolin sustaining myogenic factors and the p38 inhibitor promoting proliferation and modulating ECM interactions. Overall, this study establishes a cost-effective, small molecule-based strategy for the robust serum-free expansion of bMuSCs.

Hybrid dry-fermented sausage analogues with texturized pea proteins (TPPs) are emerging, yet flavor formation mechanisms remain unclear. We combined quantitative descriptive analysis with complementary HS-SPME-GC-MS/HS-GC-IMS volatilomics, UHPLC-MS/MS untargeted metabolomics, and marker-gene microbiome sequencing across sausages with different fermentation and ripening stages to map key aroma and their potential microbial and metabolic drivers. Sensory data showed rising fruity, cocoa-chocolate and nutty notes. In total, 47 volatiles were identified by GC-MS and 40 by GC-IMS. Screening of odorants based on relative odor activity value (rOAV) consistently highlighted seven odorants, with a shift from hexanal-dominated raw profiles to linalool-dominated processed profiles, indicating suppression of aldehyde-derived off-notes and enrichment of terpene/ester notes. Metabolomics detected 2467 metabolites, dominated by lipids and organic acids, and short-peptide enrichment suggested intensified proteolysis supplying aroma precursors. Bacterial succession exceeded fungal variation, with Latilactobacillus and Staphylococcus as core taxa. The integrated dataset provides practical markers and microbial/process cues to enhance flavor quality of sustainable hybrid fermented meats.

Rapid identification of Listeria monocytogenes (L. monocytogenes) is critical to preventing outbreaks of foodborne listeriosis. Conventional methods required immunoassay or nucleic acid sequence recognition for indirect/labeled detection or identification of L. monocytogenes, separately. Herein, a facile label-free fingerprinting method was developed for direct identification of L. monocytogenes using immunomagnetic enrichment coupled with droplet-shrinkage-assisted surface-enhanced Raman spectroscopy (SERS). Immunomagnetic beads were used for specific enrichment of L. monocytogenes from the complex matrix. Silver nanoparticles were shrunk to bacterial cell surface, which excited bacterial Raman fingerprints under radiation. The enhanced Raman signal at 477 cm-1 was selected for quantification, and the limit of detection was 7 CFU/mL in real meat sample juice. The consistent results between the proposed method and the plate counting method demonstrated its applicability. High accuracy, speed, and sensitivity endowed this method with a great potential for monitoring L. monocytogenes contamination in meat and meat products.

Cultured meat offers substantial advantages over conventional livestock production, including slaughter-free production and enhanced food safety. However, current understanding of dynamic nutrient consumption and metabolite accumulation in large-scale bioreactors remains inadequate. To address this gap, nontargeted metabolomics was employed to characterize the dynamic alterations in conditioned medium from an 80 L stirred-tank bioreactor (STR) and to identify key metabolites regulating muscle stem cell proliferation. A total of 1761 metabolites were detected; leucine (Leu) and arginine (Arg) were identified as having high variable importance in projection (VIP) scores and pathway enrichment analysis. Functional validation in both 2D monolayer and 3D suspension shake flask systems confirmed that targeted supplementation with these two amino acids significantly enhanced muscle stem cell proliferation and dose-dependently upregulated the stemness marker PAX7. This study elucidates dynamic metabolic shifts in a scaled cultured meat bioreactor, providing a crucial theoretical and data foundation for medium optimization and dynamic bioprocess control in industrialized cultured meat production.

This study aimed to investigate the effect of different proportions of mycoprotein addition (0%, 0.5%, 1%, 2%, 3%, and 5%) on the quality attributes of pork emulsified sausages. The results showed that the addition of 1% and 2% mycoprotein significantly enhanced the hardness and chewiness of emulsified sausages, which reduced the porosity and increased water retention capacity. GC-IMS analysis showed that the additions of 1% and 2% mycoprotein enriched the volatile flavor profile. FTIR and interaction force analysis demonstrated that mycoprotein addition promoted the formation ofβ-sheet secondary structures of myofibrillar protein, strengthening intermolecular hydrogen bonds, ionic bonds, and non-specific associations, which further stabilized the emulsified network. However, excessive mycoprotein addition (3% and 5%) might disrupt the stability of the emulsification system, resulting in reducing texture properties and water-holding capacity, and unbalancing flavor profiles. As a functional ingredient, mycoprotein added at levels of 1% and 2% can significantly enhance the quality attributes of pork emulsified sausages, improving texture stability and flavor. This study would provide a theoretical foundation and practical guidance for the application of mycoprotein in meat products, so as to offer an innovative approach to improve product quality while promoting health and sustainability in the meat industry.

Fibroblasts replicate rapidly, maintain stable phenotypes, and require minimal nutrients, making them valuable for cultured meat production. However, research on porcine fibroblasts is limited and has revealed significant heterogeneity across different ages and anatomical sites in pigs. In this study, fibroblasts were isolated from the ear margins, forelegs, backbone, and hind legs of neonatal and mature pigs, and their proliferation, migration, and extracellular matrix (ECM) secretion were compared. Optimal culture conditions were established using type I collagen-coated dishes containing 10% fetal bovine serum and 5.0 ng/mL basic fibroblast growth factor. Porcine neonatal fibroblasts exhibited superior proliferative and migratory capacities compared to mature fibroblasts, with neonatal ear fibroblasts (Ne) showing the highest proliferation (1.25-fold compared to mature foreleg fibroblasts). Neonatal foreleg fibroblasts (Nf) displayed the strongest ECM secretion, approximately twice that of Ne cells. These findings delineate the intrinsic functional heterogeneity among porcine fibroblasts and provide a basis for selecting optimal seed cells for cultured meat applications.

Dietary lipopolysaccharide (LPS) is naturally occurring bacterial toxins found in food. High-fat diet promoted its absorption in the gut, leading to chronic inflammation and being a major cause of metabolic endotoxemia. This study investigated how dietary protein digests regulate the physicochemical state and intestinal absorption of LPS during gastrointestinal digestion. Results showed that protein digests remodel LPS through mechanisms such as electrostatic neutralization, hydrophobic anchoring, and conformational encapsulation. Digest of chicken myofibrillar protein, with its excellent binding affinity (binding constant K = 4.66 × 104 M-1), exhibited the strongest LPS masking effect. Digest of casein, by slowing LPS release kinetics, significantly inhibited transmembrane absorption of LPS in the Caco-2 model, accumulating approximately 78% release, with a 4-h absorption rate of approximately 3.5%, the lowest among all protein groups. Conversely, while soy protein digest induced significant conformational changes in LPS, the resulting complex was unstable, resulting in limited masking. We conclude that dietary proteins and their digests, acting as key gatekeepers, regulate systemic endotoxin exposure by remodeling the bioavailability of LPS. These findings provide a theoretical basis for using specific protein matrices to alleviate diet-induced inflammation.

Animal adipose tissue consists of clusters of fat cells surrounded by intercellular fibrous networks. In this study, soybean protein isolate (SPI) and bacterial cellulose (BC) co-assemblies are utilized to simulate the intercellular fibrous networks in animal adipose tissue, stabilizing 3D-printed high internal phase emulsion (HIPE) gels with suitable processing properties. Experimental results reveal that under pH = 2 conditions, the compact spherical structure of SPI unfolds and intertwines with the high-aspect-ratio BC. Upon neutralizing the pH, SPI partially recovers its folded conformation, embedding with the intertwined BC to form stable SPI/BC co-assemblies. The SPI/BC co-assemblies in the molten-globule state exhibit improved surface sulfhydryl content and hydrophobicity. During emulsification, the HIPE system stabilized by SPI/BC co-assemblies maintains consistent interfacial layer thickness, smaller oil droplet size, and excellent rheological properties, ensuring long-term system stability even after extended storage. In the SPI/BC co-assemblies, the SPI in its molten globular state enhances emulsification and extensibility, while the high-aspect-ratio bacterial cellulose forms a three-dimensional network structure with effective thickening properties. Microstructural images of the 3D-printed HIPE gel systems demonstrate that the dense and uniform gel networks formed by the SPI/BC co-assemblies exhibit exceptional resistance to extrusion and shear forces. The SPI/BC co-assemblies show a homogeneous structure, whereas the SPI/BC mixtures display phase separation. Consequently, the 3D-printed HIPE gels stabilized by SPI/BC coassemblies (15:1) show no signs of imbalance or oil phase separation, even after 48 h of storage.

This study aimed to evaluate the effects of alternating magnetic field treatment on the mass transfer rate, microstructure, water retention, and texture of beef during the marination process. Results demonstrated that both Sodium chloride (NaCl) content and moisture content in beef were significantly increased after alternating magnetic field-assisted marination compared to conventional static marination. Mass transfer was modeled using Fick's second law, which provided a good fit to the experimental data in the marination process. The diffusion coefficients (D-values) of NaCl and moisture were found to increase significantly with increasing magnetic field intensity, respectively. Specifically, alternating magnetic field-assisted marination with an intensity of 5 mT for 90 min resulted in a maximum increase of 21.77 % in NaCl content and a 1.98 % improvement in moisture content. In addition, microstructural analysis suggested that the accelerated marination induced by the alternating magnetic field might be due to the fact that the alternating magnetic field promoted the expansion of myofibrillar gaps in muscles, thereby enhancing the permeation rate of NaCl and moisture. The cooking loss reduced by marination assisted with a 5 mT magnetic field in beef tenderloin, although no significant negative impact in tenderness was observed compared to static marination. This study demonstrates that a low-frequency alternating magnetic field can simultaneously enhance marination efficiency and maintain myofibrillar structural integrity, providing a potential alternative to conventional physical methods.

Cultured meat presents significant potential as a sustainable supplement to traditional livestock farming. Addressing the critical challenge of residual chemical substances from the culture media, this study established a rapid method combining protein precipitation (PP) with liquid chromatography-tandem mass spectrometry (LC-MS/MS) for the quantitative analysis of 12 risk substances in both porcine cultured meat biomass and washing solutions. All validation parameters, including linearity, sensitivity, precision, accuracy, recovery, matrix effect, dilution integrity and stability, met the predefined acceptance criteria. The results demonstrated that consecutive centrifugal washing cycles significantly reduced residue levels in porcine cultured meat biomass, with the majority of the 12 target substances effectively removed after four cycles. Analysis of 15 production batches revealed that 9 out of the 12 target substances were below the limit of detection (LOD) in the final product. For the remaining three substances, the mean concentrations were 307.91 μg/kg for forskolin, 568.04 μg/kg for thymidine, and 397.61 μg/kg for N-acetylcysteine. A target hazard quotient (THQ) assessment confirmed that none of the 12 residues posed a health risk. These findings provide the technical groundwork and theoretical rationale needed to optimize quality and safety specifications in the cultured meat industry.

Cultured meat is a promising approach to sustainable food production but is limited by inefficient serum-free media. Food-derived small molecules provide a cost-effective strategy for medium optimization. In this study, based on the serum-free medium Beefy-9, we performed high-throughput screening using a small-molecule library consisting of 480 food-grade compounds in bovine muscle stem cells (bMuSCs) and identified eight candidate compounds to promote cell proliferation. Through combinatorial optimization, we further established a cocktail (4C) consisting of hesperetin, hesperidin, oleanolic acid, and naringin dihydrochalcone that exhibited a sustained pro-proliferative effect on bMuSCs under serum-free conditions. Compared with the Beefy-9 control, bMuSCs cultured in the 4C medium exhibited 1.73- and 3.63-fold increases in cell yield after three and six passage culture, respectively, with significant increases in EdU-positive cells by 73.2% and 99.04%. Moreover, bMuSCs cultured in the 4C medium showed the lowest proportion of senescent cells, significant upregulated expression of myogenic factors PAX3, PAX7, MYOD, and cell cycle-related genes CCND1, CCNE1, with downregulated inhibitory regulators CDKN1C, CDKN2C, and TP53. Additionally, the 4C medium maintained the myogenic differentiation potential of bMuSCs and displayed advantages in supporting long-term cell expansion. Overall, this study establishes a cost-efficient, food-derived small molecule-based strategy for the serum-free expansion of bMuSCs.

This study evaluated the effects of low-frequency alternating magnetic field (AMF) on mass transfer, myofibrillar protein (MP) structure, and quality of beef during marination. AMF significantly enhanced NaCl and moisture diffusion, reducing marination time by over 50%. The improvement was linked to AMF-induced MP conformational unfolding, increased surface hydrophobicity, reduced solubility, and promoted protein aggregation via strengthened hydrophobic interactions. Unlike conventional treatments, AMF-driven aggregation widened inter-myofibrillar spaces, facilitating ion and water migration. Additionally, AMF reduced cooking loss while maintaining shear force and texture, indicating improved water-holding capacity without compromising tenderness. These effects were attributed to microstructural remodeling that preserved myofibrillar integrity. Overall, AMF-assisted marination enhances processing efficiency and technological quality, representing a promising non-thermal strategy for meat processing.

Genetically engineered cell lines hold promise for cultured meat production, yet are hindered by safety concerns, regulatory issues, and unproven serum-free adaptability. Here, we establish the porcine myogenic cell line PM166 via co-overexpression of telomerase reverse transcriptase (TERT) and cyclin-dependent kinase 4 (CDK4) to assess its proliferative and myogenic capacity, genomic stability and serum-free culture performance. The PM166 cell line achieved a doubling time of less than 24 h, sustained expansion beyond 200 population doublings, and robust myogenic differentiation after extensive passaging. Critically, PM166 maintained a normal diploid karyotype and genomic stability during prolonged culture, with whole-genome sequencing revealing a conserved mutational landscape and intact key cancer-associated loci. The absence of anchorage-independent growth further underscored its non-tumorigenic phenotype. Notably, PM166 was successfully adapted to serum-free conditions, maintaining a stable proliferation rate and myogenic identity. Moreover, following differentiation in 3D hydrogels, serum-free expanded PM166 cells formed myotubes, demonstrating the capacity to generate muscle tissues for cultured meat. This study presents PM166 as a genomically stable, serum-free compatible cell resource potentially for cultured meat production and safety assessment.

The industrialization of cultured meat urgently requires the establishment of a scalable seed cell expansion platform. However, adherent cells commonly used in cultured meat production face substantial challenges when adapting to serum-free suspension culture. In this study, using a porcine kidney epithelial cell line PK15 with cultured meat application potential, we systematically compared sequential and direct serum reduction strategies, and found that sequential serum reduction was more effective in acclimating adherent PK15 cells to low-serum culture conditions, maintaining robust cell proliferation, normal cell cycle progression, and stable epithelial phenotypes. On this basis, low-serum adherent PK15 cells were further adapted to serum-free suspension via stepwise serum reduction, and displayed a trend toward a more energy-efficient metabolic state. Importantly, these adapted cells retained partial phenotypic reversibility. When returned to adherent culture conditions, the adapted cells generally exhibited a clear recovery toward the state of parental adherent cells in terms of morphology, proliferation-related phenotypes, and oxidative stress-related indicators. Transcriptomic analysis revealed that genes with reversible expression patterns during the adherent-suspension-re-adherent transition were mainly enriched in cell adhesion and metabolic regulation pathways, indicating adaptive remodeling of adhesion- and metabolism-related programs during cellular adaptation and recovery. Overall, this study establishes an effective and stable serum-free suspension culture strategy for PK15 cells, and provides a methodological framework and valuable reference for the suspension adaptation of adherent seed cells in cultured meat research.

Mycoprotein, with a balanced amino acid profile and potential benefits for muscle maintenance, is a promising protein source for elderly-friendly high-protein foods. Yet, rigid fungal cell walls and entangled hyphal networks enclosing intracellular proteins may restrict enzymatic accessibility under elderly digestive conditions. In this study, high-pressure homogenization (HPH) was applied as a food-grade structural regulation strategy to improve the digestive adaptability of mycoprotein. Native mycoprotein (MYC), HPH-treated mycoprotein (HMYC), pork (PORK), and commercial plant-based meat (PLANT) were evaluated using a static in vitro oral-gastric-intestinal digestion model simulating elderly physiological conditions. Structural disintegration, hydrolysis behavior, free amino acid release, multiple light scattering, and microrheological properties were analyzed to elucidate digestive adaptation mechanisms. HPH markedly disrupted the cell-wall-associated and hypha-entangled structure of mycoprotein without significantly altering its major nutrient composition, thereby improving enzyme accessibility and digestion-induced disintegration. During gastric and intestinal digestion, the particle size D[4,3] of HMYC decreased by over 80%, indicating enhanced structural breakdown efficiency. HMYC exhibited a significantly higher hydrolysis degree than MYC, while its free amino acid release approached that of PORK and exceeded those of both MYC and PLANT. Multiple light scattering revealed improved enzyme-substrate interaction and greater physical structural transformation after HPH treatment. Microrheological analysis further confirmed improved restructuring behavior and reduced structural resistance during digestion. Overall, HPH effectively improved the digestive adaptability of mycoprotein under elderly digestive conditions, making its digestive behavior closer to that of PORK, while the distinct digestion behavior of PLANT was mainly associated with matrix effects from added lipids and hydrocolloids, supporting the development of elderly-friendly mycoprotein-based foods.

Mycoprotein, with high nutritional value and intrinsic fibrous morphology, is a promising ingredient for meat analogs. Yet, achieving stable and well-aligned muscle-like fibers in complex formulations remains challenging. In this study, a full-formulation mycoprotein system-comprising mycoprotein with egg white protein, wheat gluten, dietary fiber, and flavoring agents-was employed to better simulate realistic matrices. The effects of sodium alginate (SA) on the rheological behavior and printability of mycoprotein inks were examined, and subsequent freezing treatment was assessed for its impact on structural stability, texture, and water distribution. Molecular interactions and protein secondary structures were further analyzed to elucidate the underlying mechanisms, including hydrogen bonding enhancement, hydrophobic interaction strengthening, ionic interaction rearrangement, and secondary structure reorganization. Moderate SA addition (0.6-0.8%) significantly enhanced viscoelasticity and structural recovery, improving printing precision and stacking stability. Extrusion induced alignment of mycoprotein hyphae, while SA-supported gelation facilitated the formation of parallel muscle-like fibers. Freezing exerted dual effects: in low-support groups (characterized by low sodium alginate levels and weak gel-supported structural integrity), it highlighted the macroscopic appearance of fibrous structures, whereas in high-SA groups it primarily modulated water distribution while preserving the preestablished fiber continuity and textural properties. Molecular analysis showed SA strengthened hydrogen bonding and hydrophobic interactions and reconstructed ionic bonds, stabilizing the protein network. Correlation analysis confirmed close linkages among molecular forces, water states, and texture. SA combined with 3D printing effectively promoted muscle-like fiber construction in full-formulation mycoprotein, while freezing introduced both structural benefits and textural drawbacks. This study provides theoretical insights and practical guidance for optimizing mycoprotein-based meat analogs under cold-chain conditions.

Hot fresh pork is highly preferred by Chinese consumers for its desirable flavor and color. However, its quality deteriorates rapidly during ambient-temperature transportation, leading to unappealing meat color and shortened shelf life. This study investigated the effects of different transportation temperature setpoints (5 °C, 10 °C, 15 °C Setpoint groups, and ambient temperature) on pork carcass quality. Transportation at the lower setpoints (5 °C, 10 °C) reduced carcass center temperature, attenuated pH decline, minimized cooking and drip losses, suppressed microbial proliferation, and curtailed TVB-N accumulation (p < 0.05). These conditions also shortened the duration of high temperatures in vehicles, decelerated glycogenolysis, and moderated energy metabolism, collectively preserving meat quality. Regarding color, 5 °C Setpoint group inhibited myoglobin oxidation, yielding lower oxygenated myoglobin content and reduced a* values compared with 10 °C Setpoint group over 150 km (p < 0.05). High-throughput sequencing revealed that temperature setpoint transportation significantly influenced bacterial community succession, with highly similar profiles between the 5 °C and 10 °C Setpoint groups, yet clear divergence from the ambient control. Therefore, transportation at 10 °C Setpoint represents a balanced approach to preserving color, delaying spoilage, and extending shelf life.