Reishi White Mushrooms (Ganoderma Applanatum)

STUDY 1

Exopolysaccharide from Ganoderma applanatum as a Promising Bioactive Compound with Cytostatic and Antibacterial Properties

Abstract

A new exopolysaccharide preparation isolated from stationary cultures of the white rot fungus Ganoderma applanatum (GpEPS) was tested in terms of its bioactive properties including its cytotoxic and immunostimulatory effect. The results indicate that the tested GpEPS (at concentrations above 22.85 µg/mL and 228.5 µg/mL) may exhibit selective activity against tumor cells (cell lines SiHa) and stimulate production of TNF-α THP-1-derived macrophages at the level of 752.17 pg/mL. The GpEPS showed antibacterial properties against Staphyloccoccus aureus and a toxic effect against Vibrio fischeri cells (82.8% cell damage). High cholesterol-binding capacity and triglycerides-binding capacity (57.9% and 41.6% after 24 h of incubation with the tested substances, resp.) were also detected for the investigated samples of GpEPS.

Conclusion

The weight of evidence suggests that exopolysaccharides isolated from the white rot fungus Ganoderma applanatum are characterized by a lot of important biomedical properties. The conducted experiments have evidently shown an anticancer, immunomodulating, and antibacterial effect. The results of our tests proved that the crude GpEPS preparation exhibited antitumor activity against carcinoma cells (lines SiHa) and stimulated production of Il-6 and TNF-α by macrophage line THP-1. At the same time, the antibacterial tests also indicated good antiseptic properties of the exopolysaccharides studied against S. aureus and V. fischeri strains. The biological properties described as well as the hypocholesterolemic effect of the tested substances suggest that the present studies should be continued in view of future pharmacological applications. However, it is worth noting that further studies comprising preventive and therapeutic actions of the GpEPS fraction are needed. In the promotion of the described preparation as a promising bioactive product, the simple and economical way of production and isolation thereof as well as the possibility of standardization and control of the production conditions should be emphasized.

STUDY 2

Metabolomics Investigation of an Association of Induced Features and Corresponding Fungus during the Co-culture of Trametes versicolor and Ganoderma applanatum

Abstract

The co-culture of Trametes versicolor and Ganoderma applanatum is a model of intense basidiomycete interaction, which induces many newly synthesized or highly produced features. Currently, one of the major challenges is an identification of the origin of induced features during the co-culture. Herein, we report a 13C-dynamic labeling analysis used to determine an association of induced features and corresponding fungus even if the identities of metabolites were not available or almost nothing was known of biochemical aspects. After the co-culture of T. versicolor and G. applanatum for 10 days, the mycelium pellets of T. versicolor and G. applanatum were sterilely harvested and then mono-cultured in the liquid medium containing half fresh medium with 13C-labeled glucose as carbon source and half co-cultured supernatants collected on day 10. 13C-labeled metabolome analyzed by LC-MS revealed that 31 induced features including 3-phenyllactic acid and orsellinic acid were isotopically labeled in the mono-culture after the co-culture stimulation. Twenty features were derived from T. versicolor, 6 from G. applanatum, and 5 features were synthesized by both T. versicolor and G. applanatum. 13C-labeling further suggested that 12 features such as previously identified novel xyloside [N-(4-methoxyphenyl)formamide 2-O-beta-D-xyloside] were likely induced through the direct physical interaction of mycelia. Use of molecular network analysis combined with 13C-labeling provided an insight into the link between the generation of structural analogs and producing fungus. Compound 1 with m/z 309.0757, increased 15.4-fold in the co-culture and observed 13C incorporation in the mono-culture of both T. versicolor and G. applanatum, was purified and identified as a phenyl polyketide, 2,5,6-trihydroxy-4, 6-diphenylcyclohex-4-ene-1,3-dione. The biological activity study indicated that this compound has a potential to inhibit cell viability of leukemic cell line U937. The current work sets an important basis for further investigations including novel metabolites discovery and biosynthetic capacity improvement.

Conclusion

The application of 13C-labeling analysis produced valuable insight into the role of individual partners in the co-culture in production of known or unknown induced metabolites. Moreover, 13C-labeling approach combined with molecular network analysis can reveal whether certain structural analogs were produced by the same fungus or through the similar activation mechanism. The current 13C-labeling information sets an important foundation for further studies in the basidiomycetes, including but not limited to novel metabolites discovery and biosynthetic capacity improvement.

STUDY 3

Hypouricemic Effect of 2,5-Dihydroxyacetophenone, a Computational Screened Bioactive Compound from Ganoderma applanatum, on Hyperuricemic Mice

Abstract

Searching novel hypouricemic agents of high efficacy and safety has attracted a great attention. Previously, we reported the hypouricemic effect of Ganoderma applanatum, but its bioactives, was not referred. Herein, we report the hypouricemic effect of 2,5-dihydroxyacetophenone (DHAP), a compound screened from Ganoderma applanatum computationally. Serum parameters, such as uric acid (SUA), xanthine oxidase (XOD) activity, blood urea nitrogen (BUN), and creatinine were recorded. Real-time reverse transcription PCR (RT-PCR) and Western blot were exploited to assay RNA and protein expressions of organic anion transporter 1 (OAT1), glucose transporter 9 (GLUT9), uric acid transporter 1 (URAT1), and gastrointestinal concentrative nucleoside transporter 2 (CNT2). DHAP at 20, 40, and 80 mg/kg exerted excellent hypouricemic action on hyperuricemic mice, reducing SUA from hyperuricemic control (407 ± 31 μmol/L, p < 0.01) to 180 ± 29, 144 ± 13, and 139 ± 31 μmol/L, respectively. In contrast to the renal toxic allopurinol, DHAP showed some kidney-protective effects. Moreover, its suppression on XOD activity, in vivo and in vitro, suggested that XOD inhibition may be a mechanism for its hypouricemic effect. Given this, its binding mode to XOD was explored by molecular docking and revealed that three hydrogen bonds may play key roles in its binding and orientation. It upregulated OAT1 and downregulated GLUT9, URAT1, and CNT2 too. In summary, its hypouricemic effect may be mediated by regulation of XOD, OAT1, GLUT9, URAT1, and CNT2.

Conclusion

In summary, we report the hypouricemic effect of DHAP, a constituent screened from G. applanatum in silico. Its hypouricemic effect was mediated by decreasing XOD activities, along with its upregulation of OAT1 and downregulation of GLUT9, URAT1, and CNT2. Also, it showed some anti-inflammatory actions and little toxicity. Due to its inhibition against XOD, we chose XOD as a target for molecular docking studies, and revealed that three hydrogen bonds may play key roles in its orientation and binding to XOD.

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