Pleurotus Tuber-regium Sclerotia Research Articles

Framework as a Service, FaaS: Personalized Prebiotic Development for Infants with the Elements of Time and Parametric Modelling of in vitro Fermentation.

We proposed a framework with parametric modeling to obtain biological relevant parameters from the total probiotic growth pattern and metabolite production curves. The lag phase, maximum increase rate, and maximum capacity were obtained via a 205-h exploratory In vitro fermentation of a library of 13 structural-characterized prebiotic candidates against an exclusively breastfed infant fecal inoculum. We also conducted 16S rRNA amplicon sequencing of the infant fecal inoculum. Moreover, we introduce a robust composite metabolite-based indicator that reflects the eubiosis/dysbiosis of microbiota to complement the conventional microbial markers. In terms of short-chain fatty acid, we discovered that polymeric beta-glucans from barley demonstrated potential as prebiotic candidates, while alpha-glucans as glycogen showed the least dissolved ammonia production. In terms of total probiotic, beta-glucans from oat and mushroom sclerotia of Pleurotus tuber-regium showed comparable sustainability when compared to alpha-glucans after 48 h. Being classical prebiotic, galacto-oligosaccharides gave the second-highest metabolite-based indicator, followed by lactose. While limited improvement could be made to lactose and oligosaccharides, polymeric beta-glucans from barley avails more capacity for novel prebiotic development, such as structural modification. We anticipate that more similar parallel screening with the element of time and parametric modeling will provide more novel insights.

Rheological and controlled release properties of hydrogels based on mushroom hyperbranched polysaccharide and xanthan gum

A hyperbranched polysaccharide, coded as TM3a, was extracted from the Pleurotus tuber-regium sclerotia. TM3a was hybridized with xanthan gum (XG) by chemical crosslinking using sodium trimetaphosphate (STMP) to obtain new hydrogels with self-healing and release-controlled properties. The oscillatory rheological measurements indicated that chemically crosslinking was happened immediately on mixing STMP solutions into the XG-TM3a solutions, and the crosslinked network developed slightly as time increased. The resultant hydroges were disturbed into the loose structures and regrouped the microstructure in 2 min when a large and a small amplitude oscillation were applied in turn, suggesting a self-healable property. The XG-TM3a-STMP hydrogels exhibited shear-thinning behavior with yield stress. The storage modulus of the XG5-TM3a-STMP hydrogel was 445.2 Pa at 1% strain and 243.3 Pa at 100% strain, and yield stress was 160.6 Pa, which was higher than the corresponding value of the XG5-STMP hydrogel. The morphological observation indicated the aggregates of double helical XG chains exhibited directional arrangement, and were combined with the TM3a aggregates to constitute a network of hierarchical structures. The hybrid hydrogels with enhanced mechanical properties displayed good drug loading efficiencies and sustained drug release properties.

In Vitro and In Vivo Photothermal Cancer Therapeutic Effects of Gold Nanorods Modified with Mushroom β-Glucan.

The photothermal cancer therapeutic effect of the AuNR-Glu nanohybrids produced by coating native gold nanorods (AuNRs) with a natural mushroom biopolymer from the Pleurotus tuber-regium sclerotia (Glu) were studied in the second near-infrared window (NIR-II). The AuNR-Glu exhibited low cytotoxicity and high biocompatibility due to the surface modification of Glu when compared with the native AuNRs. AuNR-Glu nanohybrids had a high photothermal transduction efficiency (η) of 43.12%, causing effective in vitro cell ablation in both HT-29 (94.2 ± 0.8% cell death) and SW480 (94.8 ± 1.1% cell death) colon cancer cells under 1064 nm NIR-II laser irradiation at 1.0 W/cm2. Intravenous injection of AuNR-Glu nanohybrids followed by irradiation from a NIR-II laser at a safe dose (1.0 W/cm2 for 5 min) in nude mice implanted with HT-29 tumors was effective in significantly reducing the tumor growth, with no obvious harmful side effects, as evidenced by histological analysis of major organs. The present results have shown that AuNRs modified by natural biopolymers from mushroom β-glucans are novel nanomaterials with low cytotoxicity and effective photothermal anticancer agents with potential biomedical applications.

Protective Effects of Extract from Sclerotium of the King Tuber Medicinal Mushroom, Pleurotus tuberregium (Higher Basidiomycetes) on Carbon Tetrachloride-Induced Hepatotoxicity in Wistar Albino Rats.

This study was undertaken to investigate the effect of 50% ethanol extract from the sclerotia of Pleurotus tuberregium on hepatocellular integrity of albino rats treated with carbon tetrachloride (CCl4). Activities of hepatic enzymes alkaline phosphatase (ALP), alanine transaminase (ALT), and aspartate transaminase (AST) were determined in serum. Antioxidant enzyme activities, total protein, and level of malondialdehyde (MDA) were assayed in liver homogenate from treated rats. Oral administration of CCl4 (2 ml/kg body weight/day) for 7 consecutive days caused hepatotoxicity. There was a significant increase in the activity of serum ALT, AST, and ALP and the level of MDA in the liver. However, the activities of catalase and superoxide dismutase, which are antioxidant enzymes, were significantly reduced (p < 0.05). Concomitant treatment with 250 mg/kg body weight of extract and CCl4 for 7 consecutive days resulted in a significant decrease in the activity of liver marker enzymes [46% (ALT), 67.25% (AST), and 68.51% (ALP)]. These activities were further decreased [68.24% (ALT), 78.39% (AST), and 75.38% (ALP)] when the extract was increased to 500 mg/kg body weight. The activities of antioxidant enzymes, total protein, and MDA content of the liver were improved to levels that are not significantly different from the control. These findings, therefore, suggest that ethanol extract from the sclerotia of P. tuberregium has a protective action against CCl4-induced hepatotoxicity.

Characterization of water and alkali-soluble polysaccharides from Pleurotus tuber-regium sclerotia

Pleurotus tuber-regium sclerotia, an edible and medicinal mushroom, is a rich source of polysaccharides used as a functional food to promote health and longevity. The crude polysaccharides were isolated from the P. tuber-regium by hot aqueous and alkali extraction and then further purified by DEAE-52 cellulose chromatography and Sephadex G-100 chromatography. Two polysaccharides, water and alkaline extraction of polysaccharides (W-PTR and A-PTR), were obtained and their extraction process were optimized through orthogonal array design. Structure characteristics (physicochemical property analysis, molecular weight, monosaccharide composition, sulfate and uronic acid contents, triple helical structures, ultraviolet spectrum and infrared spectroscopy) of the two polysaccharides were investigated. Results showed that the main difference between the two polysaccharides is reflected in color, solubility, molecular weight and monosaccharide composition. Conformational analysis showed that both W-PTP and A-PTP had triple-helix conformation. The 3D structure of the two polysaccharides and their structure–function relationship will be challenge in the future.

Dilute solution and rheological properties of hyperbranched polysaccharide from Pleurotus tuber-regium sclerotia

The dilute solution properties, critical coil overlap concentration, and rheological properties of the hyperbranched PTM3b polysaccharide were studied and compared. Flory factor (Φ) values of PTM3b in different solvents were smaller than those of linear chains as a result of the hyperbranched structure and wide polydispersity index. The relationships of ‘zero shear’ specific viscosity (ηsp,0) and coil overlap parameter (c[η]) were represented asηsp,0∝c1.05±0.03, ηsp,0∝c1.45±0.03 and ηsp,0∝c2.89±0.11 for the dilute, semi-dilute, and concentrated solution regions of PTM3b in 0.15 M aqueous NaCl, and coil overlap parameters were c*[η] = 0.07 and c**[η] = 0.29. Shear-thinning behavior was observed for PTM3b in 0.15 M aqueous NaCl in high concentration region, whereas Newtonian behavior in relatively low concentration region. The slope values of 2.34 and 12.8 were obtained from the double logarithmic plot of η0 versus c for PTM3b in 0.15 M aqueous NaCl. The slope value of 12.8 suggested that the hyperbranched polysaccharide resembled incompressible coils in the higher concentrated region. This work could provide some valuable and fundamental information to further understand the natural hyperbranched polysaccharide solutions from dilute to concentrated regions.

Shrinking factors of hyperbranched polysaccharide from fungus

The branched structure properties of hyperbranched polysaccharides (TM3a and TM3b), extracted from sclerotia of Pleurotus tuber-regium, were studied by using laser light scattering and viscometry. The configurational shrinking factor ( g) and viscometric shrinking factor ( g′) of TM3a and TM3b were discussed, where curdlan and pullulan were taken as the linear references for derivation of g and g′. The dependences of g factor, g′ factor, and Flory factor ( Φ branched) on weight average molecular weight ( M w) were established to be g = 1.07 × 10 2 M w - 0.48 ± 0.09 , g′ = 3.63 × 10 1 M w - 0.43 ± 0.01 , and Φ branched = 7.08 × 10 20 M w 0.39 ± 0.1 for TM3a in 0.25 M LiCl/DMSO at 25 °C, when curdlan acted as the linear reference. A power law relationship g′ = 2.71 × 10 −1 g −0.61±0.1 for TM3a was found, and the exponent was approximately same to 0.60 established by Kurata et al. for polystyrene star molecules. The dependence of g factor on M w for TM3b was found to be g = 1.99 × 10 2 M w - 0.53 ± 0.02 , when pullulan was used as the linear reference. On the basis of Zimm–Stockmayer equation for tetrafunctional units, molecular weight of branching unit ( M 0) deduced from nonlinear curve fitting of g versus M w was 8739 ± 564 g/mol and 3961 ± 1245 g/mol for TM3a and TM3b, respectively. The effect of different linear reference curves and polydispersity was discussed. This work gave valuable information on branched structure characterization and insights into the biosynthetic pathways of the hyperbranched polysaccharide from fungus.

Chemical modification and antitumor activities of two polysaccharide–protein complexes from Pleurotus tuber-regium

Two water-soluble polysaccharide-protein complexes, extracted from Pleurotus tuber-regium sclerotia, were modified chemically to obtain their sulfated and carboxymethylated derivatives. While C6 position of glucan was fully substituted, C2, C3, and C4 were only partially substituted by sulfate groups. C3, C4, and C6 position of glucan were partially substituted during the carboxymethylation. Chain conformation and antitumor activity of the native samples and their derivatives were studied. The native samples and derivatives existed in sphere conformation, and showed potent in vitro antitumor activities. Water-solubility and introduction of ionic groups played important roles in enhancing the antitumor activities of the polysaccharide–protein complexes.

Microwave-assisted solubilization and solution properties of hyperbranched polysaccharide

A water-insoluble hyperbranched β- d-glucan (TM3a), extracted from sclerotia of Pleurotus tuber-regium, was treated by microwave exposure to improve its solubility in water. This method led to complete dissolution of the TM3a polysaccharide in 0.02 wt % aqueous NaN 3. Various treatment periods were tested, and optimal conditions corresponded to 35 s at 765 W. The solution properties of TM3a in water were studied systematically by using size-exclusion chromatography combined with laser light scattering, viscometry, and dynamic light scattering at 25 °C. The dependences of intrinsic viscosity ([ η]), radius of gyration ( 〈 S 2 〉 z 1 / 2 ), and hydrodynamic radius ( R h) on weight average molecular weight ( M w) for TM3a in 0.02 wt % aqueous NaN 3 at 25 °C were found to be [ η ] = 3.02 × 10 - 2 M w 0.44 ± 0.1 ( cm 3 g - 1 ) , 〈 S 2 〉 z 1 / 2 = 8.32 × 10 - 2 M w 0.38 ± 0.2 ( nm ) , and R h = 4.37 × 10 - 2 M w 0.46 ± 0.08 ( nm ) in the M w range from 8.20 × 10 5 to 4.88 × 10 6. The fractal dimension, ratio of ρ = 〈 S 2 〉 z 1 / 2 R h , and the < r 2> o/ M w value of TM3a were calculated and discussed. The results indicated that TM3a existed in a sphere-like conformation in 0.02 wt % aqueous NaN 3. Furthermore, by using transmission electron microscopy, we observed directly the spherical molecules of TM3a. This work gave valuable information on improvement of solubility and chain conformation characterization of the water-insoluble polysaccharide in water.

Chain Conformation of Water-Insoluble Hyperbranched Polysaccharide from Fungus

Water-insoluble polysaccharide (TM3a), extracted from sclerotia of Pleurotus tuber-regium, was identified as a hyperbranched beta-d-glucan from the results of one- and two-dimensional NMR and GC-MS analysis. The degree of branching of TM3a is 65.5%. TM3a was fractionated by using a non-solvent addition method into 14 fractions, and its solution properties in 0.25 M LiCl/dimethylsulfoxide (DMSO) solution were studied systematically by using static laser light scattering, dynamic light scattering, and viscometry at 25 degrees C. The dependences among the values of intrinsic viscosity ([eta]), radius of gyration (<S2>z 1/2), and hydradynamic radius (Rh) on weight-average molecular weight (Mw) were found as the following: [eta] = 0.46Mw0.30+/-0.01, <S2>z 1/2 = 4.79 x 10-2Mw0.43+/-0.04, and Rh = 5.01 x 10-2Mw0.41+/-0.02 in the Mw range from 1.94 x 105 to 2.06 x 107 for TM3a in a 0.25 M LiCl/DMSO solution at 25 degrees C. The current theory of polymer solution was applied to explain the relationship among the fractal dimension, ratio of geometric to hydrodynamic radius (rho = <S2>z 1/2/Rh), and MwA2/[eta] of TM3a. The results indicated that TM3a existed as a compact chain conformation with a sphere-like structure in LiCl/DMSO solution. Furthermore, by using transmission electron microscopy, we observed directly the spherical molecules with an average diameter of 23.0 +/- 1.8 nm.

Fractionation and characterization of a protein–polysaccharide complex fromPleurotus tuberregium sclerotia

A water-soluble sample (TM4b), extracted from sclerotia of Pleurotus tuberregium, was analyzed using elemental analysis, one- and two-dimensional 1 H and 1 3 C NMR. The results indicated that TM4b was protein-polysaccharide complex, and the polysaccharide moiety was hyperbranched β-D-glucan with residuals branched at C3, C2, C4, and C6 positions. A preparative size-exclusion chromatography (SEC) column combined with nonsolvent addition method was used to fractionate TM4b, and nine fractions were obtained. Solution properties of TM4b in 0.15 M aqueous NaCl were studied using static laser light scattering and viscometry at 25°C. The dependences of intrinsic viscosity ([η]) and radius of gyration ( z 1/2 ) on weight-average molecular weight (M w ) for TM4b in the M w range from 1.89 X 10 4 to 2.58 X 10 6 were found to be [η] = 0.21Mw 0.32±0.4 and z 1/2 = 3.63M w 0.21±0.02 . It indicated that TM4b existed as compact sphere conformation in the aqueous solution. Atomic force microscopy image further confirmed that the TM4b molecules exhibited globular shape in the solution. This work gave valuable information on fractionation and chain conformation characterization of the globular protein-polysaccharide complex.

Evaluation of sulfated fungal β-glucans from the sclerotium of Pleurotus tuber-regium as a potential water-soluble anti-viral agent

Six water-insoluble fractions of fungal β-glucans extracted by hot alkali (TM8-1 to TM8-6) from the sclerotia of Pleurotus tuber-regium (PTR) having different molecular weights ( M w) were sulfated to give their corresponding water-soluble derivatives (S-TM8-1 to S-TM8-6) with the degree of sulfation (DS) ranging from 1.14 to 1.74. The in vitro anti-viral activities of the native β-glucans (TM8s) and their sulfated derivatives (S-TM8s) were evaluated by the cytopathic effect assay (CPE) and the plaque reduction assay (PRA) against four kinds of viruses, including herpes simplex virus type 1 (HSV-1), herpes simplex virus type 2 (HSV-2), respiratory syncytial virus (RSV), and influenza A virus (Flu A). Although TM8s were inactive in inhibiting the viral replication in cell cultures, the S-TM8 fractions with the defined M w range had potent anti-viral activity against HSV-1 and HSV-2 as shown by the CPE assay. The PRA results suggested that S-TM8 fractions seemed to exert their anti-viral effect by binding to the viral particles, preventing the latter from infecting the host cells. It was plausible that the negative charges on the polymer chain of S-TM8 could interact with the positively charged glycoproteins on the surface of HSV, minimizing the interaction between the HSV and the negatively charged host cells. The anti-viral activity of the S-TM8s might also be explained by their more extended chain conformation in solution due to an increase in one of their molecular parameter, persistence length ( q), as compared to the native TM8s. The potential use of S-TM8s as a water-soluble anti-HSV agent is discussed.

Carboxymethylated β-glucans from mushroom sclerotium of Pleurotus tuber-regium as novel water-soluble anti-tumor agent

Six hot alkali extracts (HAE-1 to HAE-6) of mushroom (1→3)-β-glucans from the sclerotia of Pleurotus tuber-regium having different molecular weight ( M w) ranging from 1×10 4 to 42×10 4 were carboxymethylated to give their corresponding water-soluble derivatives (CMHAE-1 to CMHAE-6) with M w ranged from 2.08×10 4 to 53.2×10 4. In general, the CMHAE β-glucans had higher water solubility and higher in vivo (Sarcoma 180 solid tumor implanted on BALB/c mice) as well as in vitro (HL-60 tumor cell culture) anti-tumor activity than the native HAE β-glucans. From ELISA assay of cytokines, CMHAE fractions could increase production of tumor necrosis factor alpha (TNF-α) in mouse plasma stimulated by lipopolysaccharide (LPS). It was postulated that a more expanded flexible chain of these novel CMHAE β-glucans might account for their in vivo and in vitro anti-tumor activities.

Chain conformation of sulfated derivatives of β-glucan from sclerotia of Pleurotus tuber-regium

Six water-insoluble (1→3)-β- d-glucan fractions TM8-1 to TM8-6 with weight-average molecular mass M w ranging from 5.76 to 77.4×10 4 obtained from the sclerotia of Pleurotus tuber-regium were sulfated to produce the water-soluble fractions S-TM8-1 to S-TM8-6 with M w from 6.0 to 64.8×10 4. The degree of substitution (DS) of S-TM8 fractions was analyzed by elemental analysis (EA) to be 1.14–1.74. The 13C NMR results indicated that the C-6 was fully substituted, and C-2, C-4 were partially substituted by the sulfo-groups. The M w and the intrinsic viscosity [ η] of the S-TM8 fractions were measured, respectively, by size-exclusion chromatography combined with laser light scattering (SEC-LLS), LLS and viscometry in phosphate buffer solution (PBS) at 37 °C. The dependences of [ η] and radius of gyration 〈 s 2〉 z 1/2 on M w for the S-TM8 samples were found to be [ η]=1.89×10 −2 M w 0.70 (cm 3/g) and 〈 s 2〉 z 1/2=1.12×10 −4 M w 0.81 (nm) in the M w range tested. Based on current theories for a wormlike chain model, the molar mass per unit contour length M L and persistence length q of the S-TM8 were calculated to be 990 nm −1 and 8.5 nm, respectively. The relatively higher q value suggested a more expanded flexible chain of S-TM8 in PBS. The water-solubility and relatively expanded chain conformation of the STM8 fractions were considered to be significant to their antiviral activity.

Fractionation and characterization of a Polysaccharide from the sclerotia of Pleurotus tuber-regium by preparative size-exclusion chromatography

A kind of regenerated cellulose gel (RCG) particles were treated by toluene to obtain particles with smaller mean pore size, then was mixed with the cellulose gel with pore size of 370 and 525 nm. A preparative size-exclusion chromatography (SEC) column (700×20 mm) packed with three gel particles was used to fractionate water-soluble polysaccharide (WEP) extracted from the sclerotia of Pleurotus tuber-regium by aqueous solution. The exclusion limit and fractionation range of the stationary phase of the preparative SEC were molecular mass 8×10 5 and 5×10 3 to 8×10 5, respectively. The calibration curve of the preparative SEC was represented as: log M=13.96–0.53 Ve. The WEP sample (weight-average molecular mass M w=2.2×10 4, polydispersity=2.4) was divided into three fractions with M w ranging from 1.4×10 4 to 3.4×10 4 by the preparative SEC column, and the fractions were characterized by gas chromatography GC, SEC combined with laser light scattering (SEC-LLS) and viscometry. The unfractionated WEP exhibited triple peaks due to different molecular mass, but each fraction exhibited single peak with the polydispersity of 1.1–1.8 in the SEC patterns. The results indicated that the preparative SEC was efficient for fractionation of polysaccharides having low molecular weight and for determination of their molecular mass.

Molecular mass and chain conformation of carboxymethylated derivatives of beta-glucan from sclerotia of Pleurotus tuber-regium.

Seven water-insoluble (1 --> 3)-beta-D-glucan fractions TM8-1 to TM8-7 with weight-average molecular mass M(w) ranged from 2.22 to 77.4 x 10(4) obtained from the sclerotia of Pleurotus tuber-regium were carboxymethylated to produce the water-soluble fractions CTM8-1 to CTM8-7 with M(w) ranged from 3.87 to 87.8 x 10(4). The degree of substitution (DS) of CTM8 fractions was analyzed by ir and elemental analysis (EA) to be 0.3-0.68. The M(w) and the intrinsic viscosity [eta] of the CTM8 fractions were measured by size-exclusion chromatography combined with multiangle laser light scattering (SEC-MALLS), MALLS, and viscometry in phosphate buffer solution (PBS) at 37 degrees C. The dependencies of [eta] and radius of gyration <s(2)>(z) (1/2) on M(w) for the CTM8 samples were found to be [eta] = (8.82 +/- 0.03) x 10(-3) M(w)(0.78 +/- 0.04) (cm(3) g(-1)) and <s(2)>(z) (1/2) = (3.09 +/- 0.05) x 10(-3) M(w)(0.75 +/- 0.06) (nm) in the M(w) range from 3.87 x 10(4) to 53.2 x 10(4). Based on current theories for wormlike chain model, the conformational parameters of the CTM8 were obtained to be 790 (nm(-1)) for M(L), 9.6 (nm) for q, which were higher than those of the native TM8 fractions, suggesting a more extended flexible chain of CTM8 in PBS. On the whole, the CTM8 fractions showed higher antitumor activity than their corresponding TM8 fractions. In view of data from molecular parameters and bioactivity, the antitumor activity of the CTM8 fractions may be correlated to its water solubility and relatively extended chain.

Evaluation of mushroom dietary fiber (nonstarch polysaccharides) from sclerotia of Pleurotus tuber-regium (Fries) singer as a potential antitumor agent.

Mushroom dietary fiber or nonstarch polysaccharides (NSPs) that were soluble in hot alkali and belonged to the beta-glucan type were isolated from the sclerotia of an edible mushroom, Pleurotus tuber-regium. The mushroom NSPs were further separated into a number of fractions [hot alkali extracts (HAEs)] with weight-average molecular weights (M(w)) ranging from 1 x 10(4) to 42.2 x 10(4). The HAE fractions [with M(w) of (5.8-17.1) x 10(4)] administered intraperitoneally at a dose of 20 mg/kg of body weight to BALB/c mice implanted with solid tumor Sarcoma 180 were found to be effective in inhibiting tumor proliferation with an inhibition ratio of > or =50%. In vitro experiments using human tumor cell lines HL-60 and HepG2 had shown that HAE fractions with M(w) of (5.8-42.2) x 10(4) also had antiproliferative activity at three different concentrations (50, 100, and 200 microg/mL) toward the tumor cell lines tested. All HAE fractions did not inhibit the growth of a normal kidney cell line (Vero) from monkey. It is therefore postulated that the antitumoral effect of NSPs from the sclerotia of P. tuber-regium is probably host-mediated and cytocidal.

Fractionation and characterization of mushroom dietary fiber (nonstarch polysaccharides) as potential nutraceuticals from sclerotia of Pleurotus tuber-regium (Fries) singer.

The nonstarch polysaccharides (NSPs) in the total dietary fiber (TDF) from the sclerotia of Pleurotus tuber-regium (tiger milk mushroom) were fractionated by the sequential use of chemical solvents. About half of the TDF was solubilized and two major alkali-soluble fractions (1 and 4 N sodium hydroxide) that contained 126 and 293 g/kg TDF were obtained. Sugar analysis and infrared spectroscopy indicated that the NSPs in these alkali-soluble fractions were mainly beta-glucans and chitin. These alkali-soluble NSPs were further purified by anion-exchange chromatography followed by gel permeation chromatographic separation. Methylation analysis revealed that these purified glucans were highly branched and contained a mixture of sugar linkages of beta-1,3, beta-1,6, and beta-1,4. The potential use of these sclerotial beta-glucans as nutraceuticals was discussed.