S. T. Chang

Free radical scavenging activities of mushroom polysaccharide extracts

The superoxide and hydroxyl radical scavenging activities of eight mushroom antitumor polysaccharide extracts were investigated using phenazin methosulphate-NADH-nitroblue tetrazolium system and ascorbic acid-Cu(2+)-cytochrome C system respectively. The results showed that six of eight mushroom polysaccharide extracts had superoxide and hydroxyl radical scavenging activities. The protein content of the polysaccharide extracts appeared to contribute a direct effect on free radical scavenging activity. However, none of the mushroom polysaccharide extracts had antioxidative activity as measured by detecting malondialdehyde (MDA) contents of liver microsomes.

The immunomodulatory and antitumor activities of lectins from the mushroom Tricholoma mongolicum

TML-1 and TML-2 were two lectins isolated from the mushroom Tricholoma mongolicum. They did not differ appreciably in their pH stability and cationic requirement for hemagglutinating activity. They both stimulated the production of nitrite ions and activated the macrophages in mice. The two lectins were able to inhibit the growth of implanted sarcoma 180 cells by 68.84% and 92.39% respectively. The growth of tumor cells in the mouse peritoneal cavity was also inhibited by the two lectins with TML-2 expressing a greater potency.

Polysaccharide—peptide complexes from the cultured mycelia of the mushroom coriolus versicolor and their culture medium activate mouse lymphocytes and macrophages

The aim of this study was to investigate the effects of the mushroom Coriolus versicolor on cells of the immune system. The cultured mycelia of the mushroom Coriolus versicolor and their culture medium were separately extracted with boiling water. The resulting polysaccharopeptide preparations were designated intramycelial (IM) and extramycelial materials (EM), and were separated by gel filtration before determining their effects on lymphocytes and macrophages in vitro and in vivo. After gel filtration on Sepharose 6B, only a single peak with a molecular weight of 13-19 KDa was obtained. Gel filtration of IM and EM on Sephadex G-50 revealed the presence of a larger peak of 28 KDa (from IM) and 15 KDa (from EM) and a smaller peak of 3.5 KDa. IM, EM and their large molecular peaks enhanced the mitogenic response of T-cells from BALB/c mice in vitro. Splenocytes from C57BL/6 mice pre-treated by force-feeding with IM and EM demonstrated an augmented mitogenic response to Con A. The macrophages of C57BL/6 mice that had been pre-treated with IM or EM showed an enhanced production of nitrite ions. The results indicate that both mouse lymphocytes and macrophages were activated by preparations of polysaccharopeptide from cultured mycelia and culture medium of C. versicolor. However, no direct cytotoxic activity against fibroblasts, hepatoma cells and choriocarcinoma cells could be demonstrated.

Lignocellulolytic enzyme profiles of edible mushroom fungi

One of the most economically-viable processes for the bioconversion of many types of lignocellulosic wastes is represented by edible mushroom cultivation. Lentinula edodes, Volvariella volvacea and Pleurotus sajor-caju are three important commercially cultivated mushrooms which exhibit varying abilities to utilise different lignocellulosics as growth substrate. Examination of the lignocellulolytic enzyme profiles of the three species show this diversity to be reflected in qualitative variations in the major enzymic determinants (i.e. cellulases, ligninases) required for substrate bioconversion. For example, L. edodes, which is cultivated on highly lignified substrates such as wood or sawdust, produces two extracellular enzymes which have been associated with lignin depolymerisation in other fungi, (manganese peroxidase and laccase). Conversely, V. volvacea, which prefers high cellulose-, low lignin-containing substrates produces a family of cellulolytic enzymes including at least five endoglucanases, five cellobiohydrolases and two β-glucosidases, but none of the recognised lignin-degrading enzymes.

β-Glucosidase components of the cellulolytic system of the edible straw mushroom, Volvariella volvacea

The edible straw mushroom, Volvariella volvacea (V-14), produced β-glucosidase when grown in liquid culture on a variety of carbon sources including cellulose, cellobiose, salicin, sorbose, lactose, esculin, cotton wool, and filter paper. Two cell-associated β-glucosidases, BGL-I and BGL-II, were purified 32-fold and 23-fold, respectively, from extracts of cellulose-grown mycelium. The purification procedure included DEAE-Sepharose chromatography, Sephacryl S-200 chromatography, and fast protein liquid chromatography (FPLC) using Mono-Q and Mono-P anion-exchange columns. The enzymes were found to be homogeneous and to have native molecular weights of 158 kDa (BGL-I) and 256 kDa (BGL-II) by gel filtration. The isoelectric points for BGL-I and BGL-II were 5.6 and 5–5.2, respectively. Both isozymes displayed relatively broad pH optima with maximum reaction velocities recorded at pH 7.0 for BGL-I and pH 6.2 for BGL-II, and were rapidly denatured at temperatures of 60°C and above. Purified BGL-I and BGL-II were both active against p-nitrophenyl-β-d-glucopyranoside, p-nitrophenyl-α-l-arabinofuranoside, p-nitrophenyl-α-d-glucopyranoside, cellobiose, salicin, and esculin, but only BGL-I was active against p-nitrophenyl-β-d-xylopyranoside. BGL-I and BGL-II exhibited Km values for p-nitrophenyl-β-d-glucopyranoside of 90 and 500 μm, respectively. Isozyme activities were adversely affected by several reported β-glucosidase inhibitors, various metal ions, and lignin-derived aromatic acids and aldehydes. Glucose production from microcrystalline cellulose by a commercial cellulase preparation was enhanced by 9.7% in reaction mixtures supplemented with BGL-II.

Immunomodulation and antitumor activity of polysaccharide-protein complex from the culture filtrates of a local edible mushroom, Tricholoma lobayense

1. The polysaccharide-protein complex (PSPC) isolated from the culture filtrate of Tricholoma lobayense showed antitumor activity in mice. 2. PSPC had the ability to restore the phagocytic function of peritoneal exudate cells (PEC) and T-cell mitogenic activity of tumor-bearing mice. 3. An increase in the production of reactive nitrogen intermediates and tumor necrosis factor (TNF) was determined in PEC of mice that had received PSPC. 4. The present study showed that PSPC both induced the various immune responses in vivo and exhibited cytotoxicity against tumor cell lines in the presence of PSPC at doses of 30 and 60 micrograms/ml in vitro.

Induction in the mouse of gene expression of immunomodulating cytokines by mushroom polysaccharide-protein complexes

Two antitumor polysaccharide-protein complexes, PSPC and PSK from mushrooms, were compared for their modulating effect on cytokine and cytokine receptor gene expression. RNA samples were isolated from the splenocytes and peritoneal exudate cells of the untreated or treated mice. Reverse transcription-polymerase chain reaction (RT-PCR) was used to analyze the cytokine gene expression. Nine out of 17 cytokine mRNAs and 5 out of 6 cytokine receptor mRNAs were detected in the splenocytes and peritoneal exudate cells from both untreated and treated mice. However, IL-4 was only detected in the splenocytes while IL-7 and IL-1R(typeI) were only detected in the peritoneal exudate cells. Among the 9 cytokine genes, the expression level of M-CSF was up-regulated in splenocytes and peritoneal exudate cells of the mice by PSPC and PSK. The expression level of TNF-alpha was only up-regulated in the peritoneal exudate cells by PSK, but not by PSPC.

Production of cellulases and hemicellulases by the straw mushroom, Volvariella volvacea

Carboxymethylcellulase (CMCase), Avicelase and β-glucosidase production was monitored in submerged cultures of two strains (V 14 and V 34) of Volvariella volvacea grown on Avicel. All three cellulolytic enzymes were detectable in culture supernatants, although levels of CMCase and Avicelase were considerably higher in cultures of V. volvacea V 14. β-Glucosidase activity was also observed in mycelial extracts of both fungal strains. Addition of 0·2% Tween 80 to cultures markedly enhanced extracellular cellulase activity, especially in V. volvacea V 34. Cellulases, together with xylanase and both intra- and extracellular β-xylosidase, were also recorded in cultures of V. volvacea V 34 grown on paddy straw.

Anti-tumour components of the culture filtrates from Tricholoma sp.

The anti-tumour component (Fraction A) found in the culture filtrates of a local mushroom, Tricholoma sp. (strain STC20-T1), was purified using DEAE-cellulose ion exchange chromatography and Sepharose CL-4B gel filtration. Of its two purified sub-fractions, A-1 and A-2, only the latter showed anti-tumor activity. When Fraction A-2 was treated with cetyltrimethylammonium hydroxide and chloroform/butanol (24:1, v/v), it behaved as a proteinbound polysaccharide, with a molecular mass estimated to be 154 kDa by gel filtration. It contained 40% polysaccharide, which consisted of fucose, galactose, glucose, arabinose and mannose, and 30.5% protein, the amino-acid composition of which was determined. Fraction A-2 could inhibit the growth of Sarcoma-180 implanted in mice intraperitoneally and subcutaneously, with no sign of toxicity, at a dose of 20 mg/kg.day for 10 days.

Effect of lignin-derived phenolic monomers on the growth of the edible mushrooms Lentinus edodes, Pleurotus sajor-caju and Volvariella volvacea

Pleurotus sajor-caju was generally more tolerant to lignin-related phenolic monomers and tannin derivatives than Lentinus edodes and the straw mushroom, Volvariella volvacea. Several phenols, at up to 5 mM, enhanced mycelial growth of P. sajor-caju. No clear pattern was evident when the effects of phenols and tannins on the growth of V. volvacea and L. edodes were compared, but the lower concentrations of 4-hydroxybenzaldehyde and vanillin which were tested were markedly more toxic to the straw mushroom. The distribution of phenolic monomers and tannin derivatives in the agricultural wastes used for mushroom cultivation may be an important growth determinant. However, the differences in the growth inhibition profiles of L. edodes, P. sajor-caju and V. volvacea suggest that, alone, the effect of these compounds on fungal growth is unlikely to account for the varying abilities of the three mushroom species to grow and fruit on a particular lignocellulosic substrate.