| Pholiota nameko,belonging to the basidiomycota,hymenomycetes,agaricales,Strophariaceae,Pholiota,,is a mushroom with both medicinal and dietary uses.P.nameko is planted widely in China and Japan and contains considerable amounts of minerals,dietary fiber,proteins,phenols,sugars,and vitamins.Each 100 g of dried P.nameko contains more than 60%carbohydrates.Polysaccharide is an important active ingredient in P.nameko.It has been reported that the polysaccharide from P.nameko has anti-inflammatory,immune and antioxidant activities.In this paper,the water extraction residue of Pholiota nameko was used as the research object,and the alkali extraction polysaccharide was extracted by the alkali extraction alcohol precipitation method,and the separation and purification and structural analysis were carried out,and study its rheological properties and biological activity.In-depth excavation of the functional components and development value of Pholiota nameko residue can provide a theoretical basis for the comprehensive utilization of Pholiota nameko.The main research contents and results of this paper are as follows:Study on extraction process of water-soluble polysaccharides extracted from Pholiota nameko residue.The basic nutrient components of the residue after hot water extraction of Pholiota nameko were determined,and the results showed that the nutrient content in the residue was moisture 10.78±0.09%,ash 0.78±0.01%,fat 3.56±0.71%,protein 28.65±0.27%,crude fiber 6.12±0.09%and total sugar 25.86±0.27%.It can be seen that the total sugar content is still high and the residue can still be further studied.The alkali extraction method was chosen for the extraction of polysaccharides.The yield of water-soluble polysaccharides from Pholiota nameko residue as an index for investigation.The best extraction process was determined using single-factor and response surface experiments:the material-to-liquid ratio was 1:20(g/mL),2.0 mol/L NaOH concentration and 2h extraction time,and the extraction rate was 19.28±0.05%,which was not significantly different from the predicted value.Study on the rheological characteristics of water-soluble polysaccharides extracted from the pulp of Pholiota nameko residue.APNPs was obtained by precipitation(80%),protein removal,pigment removal and dialysis.The intrinsic viscosity of APNPs in distilled waterwas 328 mL/g.In the steady-state shear test,APNPs solution showed shear thinning characteristics,and the shear thinning phenomenon became more obvious with the increase of polysaccharide concentration and shear rate.The zero-shear viscosity(η0)was obtained by fitting the Cross model,and the critical transition concentration was obtained by fitting the change trend of η0 with concentration,which was 3.5 mg/mL,which was consistent with 1/[η].The thixotropic ring becomes progressively larger with increasing polysaccharide concentration.Dynamic frequency scans show that the storage modulus(G’)and loss modulus(G")increase with frequency,elasticity and viscosity increase with concentration,and fluidity deteriorates.In the creep-recovery test,the Burger model was fitted to the creep data and the correlation coefficient R2>0.99 showed that the model fitted the data very well.The Burger model data showed that with increasing concentration,the values of J0,Ji,Jmax and λ decreased,while η0 increased.The compliance recovery percentage value increased with increasing concentration,indicating that the gel network structure of the APNPs system enhanced with increasing concentration and that gel flexibility increased.Temperature has a strong influence on viscosity,which decreases with increasing temperature.It is thermally reversible at a concentration of 10 mg/mL and not at concentrations up to 15 mg/mL.Microrheological experiments showed that APNPAs solutions at a concentration≥15 mg/mL can form a gel at room temperature.Isolation,purification and structural characterization of water-soluble polysaccharides extracted from Pholiota nameko residue.Pure polysaccharide APNP was obtained by grading alcohol precipitation(10%-70%)and dextran gel G-200,with a total sugar content of 92.35%and a molecular weight of 3.56×103 kDa.The UV showed that APNP did not contain proteins and nucleic acids;the monosaccharide composition showed that APNP was composed of galactose,glucose and mannose in a molar ratio of 0.09:1:0.15.FT-IR and NMR analysis showed that APNP contained both α-and β-type glycosidic bonds.Methylation analysis showed that APNP existed in the linked forms of(1→3),(1→6)and(1→3,6)-β-D-glucose,(1→)-α-D-glucose,(1→)-α-D-galactose and(1→3,6)-β-D-mannose.In addition,the specific rotation of APNP is+80°,the surface morphology of APNP under SEM is a sheetlike accumulation with interwoven filaments attached;the TGA results show that the pyrolysis process of APNP goes through three main stages;the results of Congo red experimental analysis show that APNP does not have chain helical structure in weak base solutions.Study on the activity of water-soluble polysaccharides extracted from Pholiota nameko residue.The hypoglycemic activity of APNP was studied by colorimetric method and the establishment of HepG2 cell insulin resistance model(HepG2-IR).α-glucosidase inhibition assays showed that APNP inhibited α-glucosidase at a range of concentrations with an IC50 value of 0.165 mg/mL.Establishment of a HepG2-IR model of insulin resistance(0.6 mmol/L palmitic acid for 24 h),APNP can improve insulin resistance in HepG2 cells by increasing glucose consumption and glycogen content.Western blotting experiments revealed that APNP promoted the activation of key proteins in PI3K/AKT in HepG2 insulin-resistant cells,increased the rapid transfer of PI3k protein,promoted Akt phosphorylation,and in turn promoted the reduction of Gsk-3 expression,alleviated insulin resistance and improved glucose metabolism.The in vitro antioxidant results of APNP showed that APNP exhibited a dose-dependent effect on the scavenging of DPPH,ABTS and hydroxyl groups.The protective effects of APNP against H2O2-induced oxidative stress injury in PC 12 cells were investigated by CCK-8,kit,Western blotting,Hoechst 33342/PI and Annexin V/PI doublestaining assays.The results showed that APNP had no toxic effect on PC 12 cells,which were induced by 1200 μM H2O2 for 2 h.APNP protected PC12 cells from H2O2 induction by increasing the levels of SOD and GSH and decreasing the levels of LDH.The results showed that APNP attenuated apoptosis induced by oxidative damage.The results showed that APNP reduced apoptosis induced by oxidative stress by regulating PI3K/Akt signaling pathway and apoptosis-related pathway proteins(Bcl2,Bax.Caspase3).Effect of ultrasound on rheological properties and activity of water-soluble polysaccharides extracted from Pholiota nameko residue.The effect of different ultrasound powers on the structure of APNPs was investigated by HPGPC,nanoparticle size analyzer and FT-IR,Ultrasound reduces the average molecular weight and average particle size of polysaccharides.Higher ultrasound intensities give access to lower molecular weight fractions and do not alter their primary structure.The effect of different ultrasonic powers on the rheological properties of APNPs was investigated using a dynamic rheometer:the results showed that ultrasonic treatment significantly reduced the apparent viscosity of APNPs,weakened their shear-thinning fluid behaviour and enhanced their elasticity and thermal reversibility.The effect of different ultrasound powers on the in vitro antioxidant activity of APNPs was investigated by measuring the ability of the samples to scavenge DPPH and ABTS:the results showed that the degraded APNPs fractions exhibited stronger DPPH and ABTS scavenging activity. |
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