Mushroom residues, especially Flammulina velutipes residues (FVR) rich in bioactive polysaccharides, are under exploited due to inefficient extraction methods conflicting with green chemistry. This study developed an ultrasonic-assisted three-phase partitioning (UAE-TPP) approach to extract crude polysaccharides (cTP-FVRP) from FVR, combined with structural and functional characterization. An integrated optimization strategy combining response surface methodology (RSM), artificial neural network (ANN), and genetic algorithm (GA) identified global optimum conditions (18 min, 1:1 crude extract to n-Butanol, 44 % (NH₄)₂SO₄), yielding cTP-FVRP at 3.49 ± 0.05 %-significantly higher than that of UAE method (2.38 ± 0.04 %). Purified TP-FVRP was a 108.13 kDa non-starch polysaccharide with a triple helix, α-linkages, and 9 monosaccharides, stable under physiological conditions. It exhibits potent antioxidant activity, biocompatibility, and immunomodulatory potential. UAE-TPP reduces solvents while preserving polysaccharide integrity. This green technique enables high-value utilization of FVR, aligning with sustainability goals and providing a scalable model for industrial by-product valorization.

By integrating waste valorization with green extraction, in the current study, the impacts of distinct extraction methods on the extraction yield, structural characterization, in vitro antioxidant abilities and in vitro immunomodulatory activity of polysaccharides from Pholiota nameko residue (PNRP) were determined, providing assistance for the resource utilization of Pholiota nameko. Six PNRPs were obtained by hot water extraction, ultrasonic-assisted extraction, acid-assisted extraction, base-assisted extraction, acid–base extraction and hot water–alkaline-assisted extraction, named PNRP-HWE, PNRP-UAE, PNRP-AE, PNRP-BE, PNAP-ABE and PNRP-HAE, respectively. PNRPs were heteropolysaccharides with similar functional groups, abundant branched chains and a triple helix conformation, but varied monosaccharide molar ratios and molecular weights (382.6–601.7 kDa). PNRP-HAE exhibited the highest yield (3.92%) and superior antioxidant activities, including DPPH, ABTS and hydroxyl radical scavenging capacities, attributed to its low molecular weight and high xylose content. Additionally, PNRP-HAE and PNRP-UAE demonstrated potent immunomodulatory effects by enhancing macrophage phagocytosis and cytokine secretion (NO, IL-1β, IL-6, TNF-α). These findings highlight HAE as an optimal method for extracting high-quality PNRPs, offering a sustainable strategy for valorizing mushroom residue in functional foods and nutraceuticals.

The medicinal fungus Oudemansiella raphanipies (O. raphanipies) attracts significant interest for its broad-spectrum bioactivities, with its polysaccharide demonstrating particularly notable pharmacological properties. This study systematically investigated the extraction, physicochemical properties, structural, functional traits, biological activities, and oral absorption of O. raphanipies polysaccharide. Response surface methodology optimized the extraction condition of O. raphanipie polysaccharides by UAE method, achieving a yield of 210.35 ± 0.20 mg/g (55 °C, 90 min, 250 W, 1: 60 g/mL). The purified O. raphanipie polysaccharide (UAE-ORP) (568.57 kDa), an α-pyranose mainly composed of glucose (35.48%) and galactose (28.51%), showed high thermal stability (322 °C). In addition, UAE-ORP showed interesting water retention capacity and oil retention capacity, and good foaming and emulsion properties. UAE-ORP demonstrated potent antioxidant activity, with DPPH scavenging rate reaching 90.43% and hydroxyl radical scavenging rate at 57.13%, significant inhibition of BSA denaturation (97.49%) and NO secretion, and potential prebiotic effects. UAE-ORP was significantly distributed in the gastrointestinal tract based on NIR imaging result, retaining in intestines for 24 h, indicating its potential as a prebiotic agent. Thus, UAE-ORP is a promising functional food additive with integrated advantages.

Escherichia coli displays strong adaptability for growth and reproduction at low temperatures, with ribosome biogenesis being a critical process for its growth in cold environments. The cold-shock proteins (CSPs) encompass a protein family that can assist bacterial growth at low temperatures by acting as molecular chaperones. In this study, we investigated whether CSP CspA, CspE, and CspI affect ribosomal RNA (rRNA) transcription. Deletion of the single genes encoding these proteins had only a very marginal effect on cellular growth at low temperatures, and rRNA synthesis was hardly affected. Double and triple deletion of the genes encoding these proteins resulted in a much stronger phenotype providing evidence that CspA, CspE, and CspI play an essential role in maintaining 16S rRNA synthesis and enabling optimal cellular growth at low temperatures. These findings suggest the existence of efficient backup mechanisms able to compensate for the absence of a single CSP.

The antimicrobial peptide LR_GG (LLRLLRRGGRRLLRLL-NH2) was designed and chemically synthesized in a study conducted by Jia et al. Gram-negative bacteria were found to be sensitive to LR_GG and exhibited a high therapeutic index. Genetic engineering methods were used to create the prokaryotic fusion expression vector pQE-GFP-LR_GG, and the resulting corresponding fusion protein GFP-LR_GG was subsequently expressed and purified. The precursor GFP was then removed by TEV proteolysis, and pure LR_GG was obtained after another round of purification and endotoxin removal. The prokaryotic-expressed antimicrobial peptide LR_GG displays a broad-spectrum antibacterial effect on Gram-negative bacteria, and its minimum inhibitory activity (MIC) against Escherichia coli can reach 2 μg/mL. Compared to the chemically synthesized LR_GG, the prokaryotic-expressed LR_GG exhibits similar temperature, pH, salt ion, serum stability, and cell selectivity. Furthermore, prokaryotic-expressed LR_GG showed excellent therapeutic effects in both the infection model of cell selectivity and no embryotoxicity in a Galleria mellonella infection model. The mechanism by which LR_GG causes bacterial death was found to be the disruption of the Gram-negative cell membrane.

In recent years, multidrug resistance in pathogenic bacteria has become increasingly serious, causing serious harm to the livestock and poultry breeding industries and posing severe challenges to its clinical prevention and treatment; therefore, the development of new antibacterial agents is urgently needed. We previously isolated Bacillus velezensis CB6, which exhibits broad-spectrum antibacterial activity, from Changbaishan in China. In this study, multidrug-resistant Escherichia coli B2(MDR E. coli B2) was used as an indicator bacterium. Ammonium sulfate precipitation, dextran gel chromatography, and Diethylaminoethyl Bestarose High Performance was used to isolate antibacterial protein with strong activity against MDR E. coli B2. SDS–PAGE combined with liquid chromatography-mass spectrometry was used to obtain the antibacterial protein CB6-E, which has a molecular weight of 54.537 kDa. Our study found that CB6-E has a strong inhibitory effect on Gram-negative bacteria such as Pseudomonas aeruginosa Z1, Salmonella H9812, and Shigella castellani Z1; among them, the minimum inhibitory concentration for MDR E. coli B2 was 32 µg/mL. In addition, CB6-E is stable under various conditions including exposure to various temperatures, organic reagents, pH values, and proteolytic enzymes. The hemolytic activity test and cytotoxicity test also showed that CB6-E is safe. Research on antibacterial mechanisms showed that CB6-E destroys cell membranes in a dose-dependent manner and can inhibit the growth of MDR E. coli B2 by targeting lipopolysaccharides on the cell membrane, showing good therapeutic effects in model animals. In summary, CB6-E is a newly discovered antibacterial protein with a high therapeutic index that is safe, nontoxic, and stabile, and is expected to be an effective antibacterial agent.