Sephadex G-100 Column Research Articles

Isolation, Purification, and Characterization of Lentinus edodes Polysaccharides Extracted With Subcritical Water Enhanced With Deep Eutectic Solvent.

The Lentinus edodes polysaccharide (LEP) was extracted with a new subcritical water extraction (SWE) enhanced with deep eutectic solvent (DES) method and then purified with a DEAE-52 cellulose column and a Sephadex G-100 column. Two purified polysaccharides (LEP1 and LEP2) were obtained, and their structure, antioxidant activity, and immunomodulatory activity were analyzed. LEP1 and LEP2 were composed of mannose, glucose, and galactose with a molar ratio of 1:12.97:7.84 and 1:51.18:5.29, respectively. The molecular weights were 9.878×104 and 1.976×104Da, respectively. Interestingly, both LEP1 and LEP2 were mainly composed of →4)-β-d-Glcp-(1→, →6)-β-d-Glcp-(1→ and →6)-α-d-Galp-(1→ with different molar ratio. Besides, both LEP1 and LEP2 had strong DPPH free radical scavenging activity and Fe2+ chelating capacity. Moreover, they could reduce the level of reactive oxygen species (ROS) and regulate the activities of malondialdehyde (MDA), catalase (CAT), and superoxide dismutase (SOD) in HepG2 cells, demonstrating strong antioxidant activity. Furthermore, both LEP1 and LEP2 could improve the phagocytic capacity, nitric oxide (NO) release, and the content of interleukin (IL)-6, IL-1β, and tumor necrosis factor-α (TNF-α) in RAW264.7 cells, exhibiting significant immunostimulatory activity. It was worth noting that LEP2 exhibited stronger biological activities than LEP1. Therefore, the SWE enhanced with DES is an ideal method for extracting polysaccharides, which can be further applied to extract other polysaccharides.

Production and purification of protease produced by UV-mutated Pseudomonas aeruginosa

Background: The enzyme protease accounts for approximately two-thirds to one-third of all enzymes used in different industries. Protease is produced in large quantities as a crude enzyme for food industries. Objectives: The aim of this study was to investigate the effect of UV radiation on the protease produced by Pseudomonas aeruginosa. Methods: Of a total of 149 samples collected from various clinical cases (wounds, burns, otitis media, and urinary tract infections) in four hospitals in Baghdad for the isolation of P. aeruginosa, 90 isolates were identified as belonging to P. aeruginosa after isolation on a specific selective media, and the Vitek 2 system was later used for final identification. Thirty-six P. aeruginosa isolates were screened for protease production on a skim milk medium. Results: This study found that most of the P. aeruginosa isolates were protease producers and the isolate p3 was the best, which had a hydrolysis zone of up to 29 mm and their specific activity was 0.134 units (U)/mg protein in the culture filtrate. Therefore, this isolate was selected to study the effect of UV light on the production of the protease enzyme. The results showed that protease was produced at high levels from mutant p3 isolates after exposure to UV irradiation for different durations (15, 25, 35, 45, 55, 65, 75, 85, and 95 seconds). The specific activity of the enzyme in a crude filtrate was 0.65 U/mg protein compared with the wild type (0.134 U/mg protein). Protease produced from the mutated isolate p3 was later purified in three purification steps: first, its precipitate with 70% ammonium sulfate, second through a DEAE-cellulose column, the specific activity of which reached 7 U/mg protein, and in the final step through a Sephadex G-200 column. The purified enzyme protease had a specific activity that gradually increased to 10.5 U/mg with an 18.23-fold purification and 84.1% enzyme recovery. Conclusion: UV radiation was an efficient method to induce the production of protease enzymes.

Structure elucidation and molecular mechanism of an immunomodulatory polysaccharide from Nostoc commune

Nostoc commune Vaucher, a terrestrial and benthic blue-green alga, widely used in food and medicine worldwide. N. commune Polysaccharides (NCVP) have excellent biological activities, especially immunomodulatory, hypoglycemic and anti-tumor activities. However, the mechanism and structure-activity relationship of NCVP has been less studied. In this study, based on methylation and NMR results, a novel polysaccharide NCVP2 with 135 kDa, containing→4)-α-D-Galp-(1→, → 4)-β-D-Glcp-(1→, and →4)-α-D-Xylp-(1→ residues as the backbon, was sequentially purified from N.commune by DEAE-52 and Sephadex G-100 column. NCVP2 (50 μg/mL) exhibited the strong in vitro immunomodulatory activity by promoting the generation of nitric oxide (NO) and reactive oxygen species (ROS). A total of 2048 differentially expressed genes (DEGs) were identified by RNA-seq, including 1019 down-regulated genes and 1065 up-regulated genes. These DEGs were mainly enriched in the immune-related biological processes, involving in Mitogen-activated protein kinase (MAPK) and Toll-like receptor (TLR) signaling pathways by GO and KEGG enrichment analysis. Furthermore, Western blot results proved NCVP2 could recognize TLR2 and TLR4/MD2, and regulate TLR7/IRF7, MAPK and PI3K/AKT signaling pathways. In summary, a novel polysaccharide NCVP2 from N.commune was proposed to exhibit significant immunomodulatory effects with multiple-paths and targets, and has great potential in the development of healthy foods such as immunomodulators.

Improved exopolysaccharide production by Lactiplantibacillus plantarum Z-1 under hydrogen peroxide stress and its physicochemical properties

In this study, a strain with good exopolysaccharide (EPS)-producing ability was isolated from the fermented Benincasa hispida and identified as Lactiplantibacillus plantarum Z-1. Its EPS production was further improved by H2O2 stress under optimized culture conditions, increasing from 180 ± 0.45 mg/L to 409.52 ± 2.16 mg/L. Purification of EPS with DEAE-52 and subsequent Sephadex G-100 column chromatography provided three fractions, namely, EPS-0, EPS-1 and EPS-3, respectively. The molecular weight of EPS, EPS-0, EPS-1 and EPS-3 were 85.4, 25.7, 131.88 and 93.2 kDa, respectively. EPS, EPS-1 and EPS-3 were mainly composed of glucose, rhamnose, arabinose and galactose with molar ratios of 1:0.544:0.211:0.281, 1:1.279:0.807:0.704, and 1:1.459:0.759:0.75, respectively, along with small proportions of fucose, mannose and xylose. EPS-0 was composed of glucose, arabinose, galactose and xylose, with molar ratios of 1:0.618:0.206:0.275. The structural analysis indicated that EPS-3 was mainly consisted of (1,2,4)-β-Rhap, (1,2,3)-β-Araf, (1,4)-β-Galp, T-α-Glcp units. The three purified fractions showed typical characteristics of non-Newtonian fluids and good viscoelasticity. Congo red test revealed that irregular triple-helical conformation existed in EPS and EPS-3. These physicochemical properties of EPSs make them a potential candidate for the use as a health-beneficial food additive in the food processing industry.

Physicochemical properties, antioxidant and anti-inflammatory activities of polysaccharides from lotus leaves by pectinase-assisted extraction

In order to improve the extraction yield of lotus leaf polysaccharide (LLP) and offer scientific foundation for utilization of LLPs in the food and pharmaceutical industries, pectinase-assisted extraction method was employed for the preparation of LLP. The chemical composition, physicochemical properties (e.g. rheological and thermal properties etc.), antioxidant and anti-inflammatory effects of LLP and its two purified fractions (LLP1 and LLP3) obtained using DEAE-52 and subsequent Sephadex G-100 column chromatography were also investigated. Antioxidant assays indicated that the polysaccharides had remarkable DPPH• and ABTS+• scavenging activities. In H2O2-induced RAW264.7 cells, LLPs showed marked protective effect on oxidative stress, reducing the MDA and ROS contents and increasing the CAT and SOD activities. LLPs were also found to effectively modulate the expressions of CAT, SOD1, Nrf-2, HO-1, NQO1 and Keap1 mRNA. Thus, the potent antioxidant effects of LLPs were achieved possibly by regulating the Keap1/Nrf-2/HO-1 signaling pathways. In lipopolysaccharide-treated RAW264.7 macrophages, LLPs reduced phagocytic ability, decreased release of NO, TNF-α, IL-1β and IL-6, and increased release of IL-10 and TGF-β1, indicating their potent anti-inflammatory effect. LLPs downregulated expressions of iNOS, IL-1β, IL-6, TNF-α, COX-2, and TLR-4 mRNA. Therefore, modulation of NF-κB signaling pathway is possibly the important anti-inflammatory mechanism of LLPs.

The structural features and anti-inflammatory properties of a glucogalactan from Holotrichia diomphalia Bates (Qi Cao)

Ethnopharmacological relevanceThe dried larvae of Holotrichia diomphalia Bates, named Qi Cao, is a traditional Chinese medicine treat for liver diseases and arthritis. Polysaccharides is a principal component in Qi Cao, which exhibiting antioxidant and anti-inflammatory effects. However, the structural characteristics and underlying mechanisms of the polysaccharides remain inadequately elucidated. Aim of the studyTo analyze the primary structure and elucidate the molecular anti-inflammatory mechanisms of the active polysaccharide in Qi Cao. Materials and methodsThe total polysaccharide was extracted by water extraction and alcohol precipitation, and further isolated and purified by DEAE Sephadex A-25 column and Sephadex G-100 column. The anti-inflammatory properties of four major fractions (HDPS-1, HDPS-2, HDPS-3, HDPS-4) and the pure homogeneous polysaccharides (HDPS-1I and HDPS-1II) were assessed using a RAW 264.7 cell model induced by lipopolysaccharide (LPS), and HDPS-1II was identified as the polysaccharide exhibiting significant anti-inflammatory activity in Qi Cao. The structural characteristics of HDPS-1II were subsequently analyzed using high-performance size-exclusion chromatography (HPSEC), fourier-transform infrared spectroscopy (FT-IR), gas chromatography-mass spectrometry (GC-MS), and nuclear magnetic resonance (NMR) spectroscopy. The TLR4, NF-κB, COX-2 and iNOS expressions were determined by Western blot analysis to investigate the anti-inflammatory mechanism of HDPS-1II in vitro. Finally, the in vivo anti-inflammatory activity of HDPS-1II were evaluated by measuring the serum levels of pro-inflammatory factors, inflammatory cell infiltration and organelle damage in the lung tissues of sepsis model mice. ResultsA homogeneous polysaccharide (HDPS-1II) with molecular weight of 1.7 × 104 Da was isolated from Holotrichia diomphalia Bates. HDPS-1II contains a backbone of α-T-Glcp-(1 → 6)-α-Glcp-(1 → 4)-α-Galp-(1 → 4)-α-Galp-(1 → 6)-α-Galp-(1 → 3)-α-Galp-(1 → . It inhibited activation of the TLR4/NF-κB signaling and reduced pro-inflammatory factors and NO in LPS-stimulated macrophage. Moreover, HDPS-1II increased the survival rate, inhibited inflammatory cells infiltration, and ameliorated the lung tissue damage in septic mice. ConclusionsHDPS-1II exhibits anti-inflammatory effects in vitro and in vivo, which is the active polysaccharide components of the anti-inflammatory activity of Qi Cao.

Extraction, separation and efficacy of yam polysaccharide

Yam is used as common herbal remedy in traditional Chinese medicine and is grown all over Asia. According to previous research, one of the primary bioactive components of yam is yam polysaccharide. To shed light on the mechanism of yam polysaccharide in ulcerative colitis (UC), a yam heteropolysaccharide named CYP-3a was isolated and purified using ultrasonic extraction, the trichloroacetic acid technique, DEAE cellulose-52 and a Sephadex G75 column. CYP-3a comprises rhamnus: arabinose:galactose:mannose:galacturonic acid glucuronic acid, with a molar ratio of 2.25:4.17:3.30:0.09:0.13:0.26. CCK-8 and ELISA analysis results showed that CYP-3a increased the number of dextran sodium sulphate (DSS)-induced Caco-2 cells and could reduce and inhibit their inflammatory response by lowering the amounts of secreted TNF-α and IL-6. Western blot data demonstrated that CYP-3a at various doses could suppress the endoplasmic reticulum stress–mediated apoptotic pathway generated by DSS-induced UC and down-regulate the protein levels of GRP78, CHOP and NF-κB.

Optimization of Ultrasonic-Assisted Extraction, Characterization and Antioxidant and Immunoregulatory Activities of Arthrospira platensis Polysaccharides.

This study aimed to enhance the ultrasonic-assisted extraction (UAE) yield of seawater Arthrospira platensis polysaccharides (APPs) and investigate its structural characteristics and bioactivities. The optimization of UAE achieved a maximum crude polysaccharides yield of 14.78%. The optimal extraction conditions were a liquid-solid ratio of 30.00 mL/g, extraction temperature of 81 °C, ultrasonic power at 92 W and extraction time at 30 min. After purification through cellulose DEAE-52 and Sephadex G-100 columns, two polysaccharide elutions (APP-1 and APP-2) were obtained. APP-2 had stronger antioxidant and immunoregulatory activities than APP-1, thus the characterization of APP-2 was conducted. APP-2 was an acidic polysaccharide consisting of rhamnose, glucose, mannose and glucuronic acid at a ratio of 1.00:24.21:7.63:1.53. It possessed a molecular weight of 72.48 kDa. Additionally, APP-2 had linear and irregular spherical particles and amorphous structures, which contained pyranoid polysaccharides with alpha/beta glycosidic bonds. These findings offered the foundation for APP-2 as an antioxidant and immunomodulator applied in the food, pharmaceutical and cosmetic industries.

Isolation, Characterization, Moisturization and Anti-HepG2 Cell Activities of a Novel Polysaccharide from Cyanobacterium aponinum.

Polysaccharides from cyanobacteria are extensively reported for their complex structures, good biocompatibility, and diverse bioactivities, but only a few cyanobacterial species have been exploited for the biotechnological production of polysaccharides. According to our previous study, the newly isolated marine cyanobacterium Cyanobacterium aponinum SCSIO-45682 was a good candidate for polysaccharide production. This work provided a systematic study of the extraction optimization, isolation, structural characterization, and bioactivity evaluation of polysaccharides from C. aponinum SCSIO-45682. Results showed that the crude polysaccharide yield of C. aponinum reached 17.02% by hot water extraction. The crude polysaccharides showed a porous and fibrous structure, as well as good moisture absorption and retention capacities comparable to that of sodium alginate. A homogeneous polysaccharide (Cyanobacterium aponinum polysaccharide, CAP) was obtained after cellulose DEAE-52 column and Sephadex G-100 column purification. CAP possessed a high molecular weight of 4596.64 kDa. It was mainly composed of fucose, galactose, and galacturonic acid, with a molar ratio of 15.27:11.39:8.64. The uronic acid content and sulfate content of CAP was 12.96% and 18.06%, respectively. Furthermore, CAP showed an in vitro growth inhibition effect on human hepatocellular carcinoma (HepG2) cells. The above results indicated the potential of polysaccharides from the marine cyanobacterium C. aponinum SCSIO-45682 as a moisturizer and anticancer addictive applied in cosmetical and pharmaceutical industries.

Novel glucomannan-like polysaccharide from Lycium barbarum L. ameliorates renal fibrosis via blocking macrophage-to-myofibroblasts transition

The macrophage to myofibroblasts transition (MMT) has been reported as a newly key target in renal fibrosis. Lycium barbarum L. is a traditional Chinese medicine for improving renal function, in which its polysaccharides (LBPs) are the mainly active components. However, whether the role of LBPs in treating renal fibrosis is related to MMT process remain unclear. The purpose of this study was to explore the relationship between the regulating effect on MMT process and the anti-fibrotic effect of LBPs. Initially, small molecular weight LBPs fractions (LBP-S) were firstly isolated via Sephadex G-100 column. Then, the potent inhibitory effect of LBP-S on MMT process was revealed on bone marrow-derived macrophages (BMDM) model induced by TGF-β. Subsequently, the chemical structure of LBP-S was elucidated through monosaccharide, methylation and NMR spectrum analysis. In vivo biodistribution characteristics studies demonstrated that LBP-S exhibited effectively accumulation in kidney via intraperitoneal administration. Finally, LBP-S showed a satisfactory anti-renal fibrotic effect on unilateral ureteral obstruction operation (UUO) mice, which was significantly reduced following macrophage depletion. Overall, our findings indicated that LPB-S could alleviate renal fibrosis through regulating MMT process and providing new candidate agents for chronic kidney disease (CKD) related fibrosis treatment.

Structural characterization of Astragalus polysaccharide-D1 and its improvement of low-dose metformin effect by enriching Staphylococcus lentus

To explore the adjuvant therapy drugs of low-dose metformin, one homogeneous polysaccharide named APS-D1 was purified from Astragalus membranaceus by DEAE-52 cellulose and Sephadex G-100 column chromatography. Its chemical structure was characterized by molecular weight distribution, monosaccharide composition, infrared spectrum, methylation analysis, and NMR. The results revealed that APS-D1 (7.36 kDa) consisted of glucose, galactose, and arabinose (97.51 %:1.56 %:0.93 %). It consisted of →4)-α-D-Glcp-(1→ residue backbone with →3)-β-D-Galp-(1→ residue and terminal-α/β-D-Glcp-(1→ side chains. APS-D1 could significantly improve inflammation (TNF-α, LPS, and IL-10) in vivo. Moreover, APS-D1 improved the curative effect of low-dose metformin without adverse events. APS-D1 combined with low-dose metformin regulated several gut bacteria, in which APS-D1 enriched Staphylococcus lentus to produce l-carnitine (one of 136 metabolites of S. lentus). S. lentus and l-carnitine could improve diabetes, and reduction of S. lentusl-carnitine production impaired diabetes improvement. The combination, S. lentus, and l-carnitine could promote fatty acid oxidation (CPT1) and inhibit gluconeogenesis (PCK and G6Pase). The results indicated that APS-D1 enhanced the curative effect of low-dose metformin to improve diabetes by enriching S. lentus, in which the effect of S. lentus was mediated by l-carnitine. Collectively, these findings support that low-dose metformin supplemented with APS-D1 may be a favorable therapeutic strategy for type 2 diabetes.

Structural analysis, anti-inflammatory activity of the main water-soluble acidic polysaccharides (AGBP-A3) from Panax quinquefolius L berry

BackgroundPanax quinquefolius L, widely recognized for its valuable contributions to medicine, has aroused considerable attention globally. Different from the extensive research has been dedicated to the root of P. quinquefolius, its berry has received relatively scant focus. Given its promising medicinal properties, this study was focused on the structural characterizations and anti-inflammatory potential of acidic polysaccharides from the P. quinquefolius berry. Materials and methodsP. quinquefolius berry was extracted with hot water, precipitated by alcohol, separated by DEAE-52-cellulose column to give a series of fractions. One of these fractions was further purified via Sephadex G-200 column to give three fractions. Then, the main fraction named as AGBP-A3 was characterized by methylation analysis, NMR spectroscopy, etc. Its anti-inflammatory activity was assessed by RAW 264.7 cell model, zebrafish model and molecular docking. ResultsThe main chain comprised of α-L-Rhap, α-D-GalAp and β-D-Galp, while the branch consisted mainly of α-L-Araf, β-D-Glcp, α-D-GalAp, β-D-Galp. The RAW264.7 cell assay results showed that the inhibition rates against IL-6 and IL-1β secretion at the concentration of 625 ng/mL were 24.83 %, 11.84 %, while the inhibition rate against IL-10 secretion was 70.17 % at the concentration of 312 ng/mL. In the zebrafish assay, the migrating neutrophils were significantly reduced in number, and their migration to inflammatory tissues was inhibited. Molecular docking predictions correlated well with the results of the anti-inflammatory assay. ConclusionThe present study demonstrated the structure of acidic polysaccharides of P. quinquefolius berry and their effect on inflammation, providing a reference for screening anti-inflammatory drugs.

Purple sweet potato polysaccharide ameliorates concanavalin A-induced hepatic injury by inhibiting inflammation and oxidative stress

BackgroundAutoimmune hepatitis (AIH) is a prevalent liver disease that can potentially lead to hepatic fibrosis and cirrhosis. The prolonged administration of immunosuppressive medications carries significant risks for patients. Purple sweet potato polysaccharide (PSPP), a macromolecule stored in root tubers, exhibits anti-inflammatory, antioxidant, immune-enhancing, and intestinal flora-regulating properties. Nevertheless, investigation into the role and potential mechanisms of PSPP in AIH remains notably scarce. PurposeOur aim was to explore the possible protective impacts of PSPP against concanavalin A (Con A)-induced liver injury in mice. MethodsPolysaccharide was isolated from purple sweet potato tubers using water extraction and alcohol precipitation, followed by purification through DEAE-52 cellulose column chromatography and Sephadex G-100 column chromatography. A highly purified component was obtained, and its monosaccharide composition was characterized by high performance liquid chromatography (HPLC). Mouse and cellular models induced by Con A were set up to investigate the impacts of PSPP on hepatic histopathology, apoptosis, as well as inflammation- and oxidative stress-related proteins in response to PSPP treatment. ResultsThe administration of PSPP significantly reduced hepatic pathological damage, suppressed elevation of ALT and AST levels, and attenuated hepatic apoptosis in Con A-exposed mice. PSPP was found to mitigate Con A-induced inflammation by suppressing the TLR4-P2X7R/NLRP3 signaling pathway in mice. Furthermore, PSPP alleviated Con A-induced oxidative stress by activating the PI3K/AKT/mTOR signaling pathway in mice. Additionally, PSPP demonstrated the ability to reduce inflammation and oxidative stress in RAW264.7 cells induced by Con A in vitro. ConclusionPSPP has the potential to ameliorate hepatic inflammation via the TLR4-P2X7R/NLRP3 pathway and inhibit hepatic oxidative stress through the PI3K/AKT/mTOR pathway during the progression of Con A-induced hepatic injury. The results of this study have unveiled the potential hepatoprotective properties of purple sweet potato and its medicinal value for humans. Moreover, this study serves as a valuable reference, highlighting the potential of PSPP-1 as a drug candidate for the treatment of immune liver injury.

Antioxidant and cytotoxic effects of isolated polysaccharides from cultured Ophiocordyceps sinensis

The Ophiocordyceps sinensis is well-known as a traditionally valued medicine. Recently, the cultured O. sinensis biomass have been producing and commercializing in Vietnam. Besides, the O. sinensis - isolated polysaccharides have been revealed possessing great biological activities, especially the potential antioxidant. Hence, this study focused on antioxidant activity and cytotoxicity of polysaccharides isolated from cultured O. sinensis biomass.Methods: Firstly, crude polysaccharides (PS) including IPS (endo-polysaccharide extracted from biomass) and EPS (exo-polysaccharide extracted from broth) were isolated from O. sinensis, respectively. Then, the crude IPS and EPS were separated into fractions using a Sephadex G-200 column with a flow rate of 0.6 ml/min. The PSs and their fractions including IPS-1, IPS-2 and EPS-1, EPS-2 were screened for antioxidant (ABTS·+ free radical scavenging assay) and cytotoxic (HepG2 assay) activites.Results: Both EPSs and IPSs were mixtures of peptide-polysaccharide. In the ABTS assay, the IC50 value of EPS and EPS-2 fractions were 2,688.80 ± 19.83 µg/ml and 1,885.90 ± 2.90 µg/ml, respectively. This result indicated that the antioxidant activity of EPS and EPS-2 fractions were higher than that of IPSs as well as EPS-1 fraction. Furthermore, only crude EPS exhibited HepG2 cell growth inhibitory ability (IC50 = 15.59 ± 0.17 µg/ml) among the samples. Taken together, the EPSs exhibited greater antioxidant and cytotoxic activity than the IPSs. Thus, the EPSs could be considered as an antioxidant source.

Inhibition of Proliferation and Induction of Apoptosis in Prostatic Carcinoma DU145 Cells by Polysaccharides from Yunnan Rosa roxburghii Tratt.

This study aimed to investigate methodologies for the extraction and purification of polysaccharides from Rosa roxburghii Tratt fruits and their impact on various cellular processes in prostate cancer DU145 cells, including survival rate, migration, invasion, cell cycle, and apoptosis. Compared to the control group, the polysaccharide exhibited a significant reduction in the viability, migration, and invasion rates of DU145 cells in a time- and dose-dependent manner within the polysaccharide-treated groups. Additionally, it effectively arrested the cell cycle of DU145 cells at the G0/G1 phase by downregulating the expressions of CDK-4, CDK-6, and Cyclin D1. Furthermore, it induced apoptosis by upregulating the expressions of Caspase 3, Caspase 8, Caspase 9, and BAX. Polysaccharides were extracted from Rosa roxburghii Tratt sourced from Yunnan, China. Extraction and decolorization methods were optimized using response surface methodology, based on a single-factor experiment. Polysaccharide purification was carried out using DEAE-52 cellulose and Sephadex G-100 column chromatography. The optimal dosage of R. roxburghii Tratt polysaccharide affecting DU145 cells was determined using the CCK-8 assay. Cell migration and invasion were assessed using transwell and scratch assays. Flow cytometry was employed to analyze the effects on the cell cycle and apoptosis. Western blotting and Quantitative real-time PCR were utilized to examine protein and mRNA expressions in DU145 cells, respectively. Rosa roxburghii Tratt polysaccharides, consisting of D-mannose, L-rhamnose, N-acetyl-D-glucosamine, D-galacturonic acid, D-glucose, D-galactcose, D-xylose, L-arabinose, and L-fucose, possess the ability to hinder DU145 cell proliferation, migration, and invasion while inducing apoptosis through the modulation of relevant protein and gene expressions.

F1 fraction isolated from Mesobuthus eupeus scorpion venom induces macrophage polarization toward M1 phenotype and exerts anti-tumoral effects on the CT26 tumor cell line

Scorpion venoms identified as agents with anti-tumor and anti-angiogenic features. Tumor microenvironment (TME) plays a pivotal role in the process of tumorigenesis, tumor development, and polarization of M2 phenotype tumor associated macrophages (TAMs). M2 polarized cells are associated with tumor growth, invasion, and metastasis. The fractionation process was performed by gel filtration chromatography on a Sephadex G50 column. To elucidate whether scorpion venom can alter macrophage polarization, we treated interleukin (IL)-4-polarized M2 cells with isolated fractions from Mesobuthus eupeus. Next, we evaluated the cytokine production and specific markers expression for M2 and M1 phenotype using enzyme linked immunosorbent assay (ELISA) and real-time polymerase chain reaction (PCR), respectively. The phagocytic capacity of macrophages was also assessed. In addition, the migration assay and MTT analysis were performed to investigate the effects of reprogrammed macrophages on the CT-26 colon cancer cells. The results indicated that F1 fraction of venom significantly upregulated the levels and expression of M1-associated cytokines and markers, including tumor necrosis factor-alpha (TNF-α) (p < 0.001), IL-1 (p < 0.01), interferon regulatory factor 5 (IRF5) (p < 0.0001), induced nitric oxide synthase (iNOS) (p < 0.0001), and CD86 (p < 0.0001), and downregulated M2-related markers, including transforming growth factor-beta (TGF-β) (p < 0.05), IL-10 (p < 0.05), Fizz1 (p < 0.0001), arginase-1 (Arg-1) (p < 0.0001), and CD206 (p < 0.001). The macrophage phagocytic capacity was enhanced after treatment with F1 fraction (p < 0.01). Moreover, incubation of CT-26 cell line with conditioned media of F1-treated macrophages suppressed migration (p < 0.0001) and proliferation (p < 0.01) of tumor cells. In conclusion, our findings demonstrated the potential of Mesobuthus eupeus venom in M2-to-M1 macrophage polarization as a promising therapeutic approach against proliferation and metastasis of colon cancer cells.

Purification and Structural Characterization of Polysaccharides from Polygonum multiflorum Thunb. and Their Immunostimulatory Activity in RAW264.7 Cells.

Polygonum multiflorum Thunb. (PM) and derived products are broadly utilized in Chinese traditional medicine. According to our previous research, PM mostly contains polysaccharides, which display a wide range of biological activities. Two water-soluble polysaccharides (PMPs-1 and PMPs-2) were obtained from PM by DEAE-Cellulose and Sephadex G-100 column chromatography. Colorimetry, HPGPC-MALLS-RID, HPLC-PDA, methylation, FT-IR, NMR, and SEM were used to characterize these polysaccharides. PMPs-1 and PMPs-2 had average molecular weights of 255.5 and 55.7 kDa, respectively. PMPs-1 consisted of Man, Glc, Gal, and Ara at 0.9:78.6:1.0:1.6 and was a glucan with → 4)-Glcp-(1 → as a backbone. Meanwhile, PMPs-2, an acidic polysaccharide, comprised Rha, GalA, Glc, Gal, and Ara at 3.2:20.3:2.7:1.0:8.3. PMPs-1 and PMPs-2 significantly improved the proliferation of RAW 264.7 cells and induced NO, TNF-α, and IL-6 release. This study reveals that these two polysaccharides can be explored as novel immunomodulators and provide a basis for further development of PM in food and pharmaceutical industries.

Enzyme-Assisted Ultrasonic Extraction and Antioxidant Activities of Polysaccharides from Schizochytrium limacinum Meal.

Enzyme-assisted ultrasonic extraction (EAUE) was utilized and optimized for extracting polysaccharides from Schizochytrium limacinum meal (SLMPs) via the response surface methodology. The optimal EAUE conditions were determined as follows: enzyme concentration at 5.18%, ultrasonic temperature at 53 °C, ultrasonic duration of 40 min, ultrasonic power at 60 W, and a liquid-to-material ratio of 34 mL/g, achieving a polysaccharide extraction yield of 11.86 ± 0.61%. The purified polysaccharide component, SLMP1-1, isolated using DEAE Sepharose Fast Flow and Sephadex G-100 columns, exhibited potent antioxidant activity. SLMP1-1, with a molecular weight of 25.5 kDa, comprises glucose, mannose, arabinose, and galactose in a molar ratio of 16.39:14.75:1:693.03. 1H NMR analysis revealed the α configuration of SLMP1-1. Antioxidant assessments, including DPPH, ABTS, and ferric ion reduction assays, were detected with inhibitory values at 21.82-82.98%, 38.21-98.46%, and 3.30-20.30% at 0.2-1.0 mg/mL. This confirmed the effective antioxidant capacity of SLMP1-1, which is notably enhanced post oral and gastric digestion. The findings suggest that polysaccharides extracted from Schizochytrium limacinum meal hold significant promise as natural antioxidants.

Purification, kinetic characterization of thermostable multicopper oxidase from the oyster mushroom and its versatility for greener agro-pulp bio bleaching in the paper industry.

Production of a thermostable laccase from Pleurotus florida was reported for the first time, both in submerged and solid-state fermentation using agro-industrial residues. This enzyme was purified using ammonium sulphate precipitation (60-90%), Sephadex G-100 and DEAE column ion exchange chromatography, respectively. The laccase was purified to 21.49 fold with an apparent molecular weight of 66 kDa and had an optimal pH of 5 with temperature stability at 60°C. Metal ions such as Cu2+ (91.26 µmole/mL/min), Mg2+ (68.15 µmole/mL/min), and Fe2+ (1.73 µmole/mL/min) enhanced the laccase activity, but Fe2+ (1.73µmole/mL/min) inhibited the enzyme activity. The purified laccase had Km and Vmax of 16.68 mM and 26.73 µmole/mL/min for guaiacol as a substrate. The isolated enzyme was characterized by FT-IR which revealed bands at 3655.0 cm-1, 2894.7 cm-1, and 1151.7 cm-1 corresponding to primary amines, C-H stretch, and amide -III, respectively. The enzymatic bio bleaching of paddy straw pulp was found to be most effective which resulted in a lowering of kappa number and yellowness by 19.47% & 17.84% whereas an increase in brightness and whiteness by 41.92%. & -19.61%. Thus, this might be stated that the crude laccase from P. florida can be exploited to reduce the toxic waste load for managing environmental pollution and helps in enhancing the yield and quality of the paper.

Synovial Fluid Proteomics and Serum Metabolomics Reveal Molecular and Metabolic Changes in Osteoarthritis

Background: Osteoarthritis (OA) is a common joint disorder with a complex and multifactorial pathogenesis. Proteomics analysis using two-dimensional gel electrophoresis (2DE) and mass spectrometry (MS) enables high-throughput identification of differentially expressed proteins related to OA. However, the etiology, pathophysiology, and early diagnostic markers of OA are still poorly understood. Methods: Synovial fluid protein biomarkers were compared between OA patients and healthy controls. It was fractionated using DEAE cellulose and Sephadex G-200 columns, followed by SDS‒PAGE and 2D-PAGE for visualization and identification. Mass spectrometry and Mascot were used for protein analysis, and serum metabolite profiles were also investigated using 1D 1H CPMG NMR spectra. Multivariate data analysis, including PCA and PLS-DA, was performed to detect metabolic differences between groups. Results: Proteomics analysis revealed differential expression of synovial fluid proteins, such as serine protease inhibitors, complement components, and apolipoproteins, which may be involved in inflammation and cartilage breakdown. Additionally, serum metabolite profiles differed significantly between OA patients and controls, involving amino acid, lipid, glucose, and energy metabolism. The pathway analysis indicated disruption of the metabolic pathways associated with these metabolites. Conclusions: This study provides insights into the molecular and metabolic changes in OA. Protein biomarkers and serum metabolite alterations enhance the understanding of OA pathogenesis and offer potential opportunities for early diagnosis and disease management. Further validation and translation of these findings into clinical applications are needed for improved OA detection and intervention strategies.