[Isolation and identification of endophytic fungi from Huperzia serrata and their metabolites' inhibitory activities against acetylcholinesterase and anti-inflammatory activities].

A total of 27 endophytic fungal strains were isolated from Huperzia serrata,which were richly distributed in the stems and leaves while less distributed in roots. The 27 strains were identified by Internal Transcribed Spacer( ITS) r DNA molecular method and one of the strains belongs to Basidiomycota phylum,and other 26 stains belong to 26 species,9 general,6 families,5 orders,3 classes of Ascomycota Phylum. The dominant strains were Colletotrichum genus,belonging to Glomerellaceae family,Glomerellales order,Sordariomycetes class,Ascomycota Phylum,with the percentage of 48. 15%. The inhibitory activities of the crude extracts of 27 endophytic fungal strains against acetylcholinesterase( ACh E) and nitric oxide( NO) production were evaluated by Ellman's method and Griess method,respectively. Crude extracts of four fungi exhibited inhibitory activities against ACh E with an IC50 value of 42. 5-62. 4 mg·L~(-1),and some fungi's crude extracts were found to inhibit nitric oxide( NO) production in lipopolysaccharide( LPS)-activated RAW264. 7 macrophage cells with an IC50 value of 2. 2-51. 3 mg·L~(-1),which indicated that these fungi had potential anti-inflammatory activities.The chemical composition of the Et OAc extract of endophytic fungus HS21 was also analyzed by LCMS-IT-TOF. Seventeen compounds including six polyketides,four diphenyl ether derivatives and seven meroterpenoids were putatively identified.

Polyketides from Alternaria alternata MT-47, an endophytic fungus isolated from Huperzia serrata.

Four new polyketides, alternatains A-D (1-4), along with 17 known compounds (5-21) were obtained from the solid substrate fermentation cultures of Alternaria alternata MT-47, an endophytic fungus isolated from the medicinal plant of Huperzia serrata. Their structures were elucidated by extensive spectroscopic and spectrometric techniques (1D and 2D NMR, IR, and HRESIMS) and calculated electronic circular dichroism (ECD) method. Compounds 4, 6, 15, and 21 exhibited inhibitory activities on ATP release of thrombin-activated platelets with IC50 values in the range of 18.2-68.8 µM.

Lycopodium alkaloids from Huperzia serrata.

Five new Lycopodium alkaloids, huperzine Y1 (1), huperzine Y2 (2), huperzine Y3 (3), (+)-huperzine Z (4a) and (-)-huperzine Z (4b) as well as ten known alkaloids (5-14) were isolated from Huperzia serrata. The structures of the new compounds were established using extensive spectroscopic (1D and 2D NMR, IR, and HRESIMS) and calculated electronic circular dichroism (ECD) methods. Compounds 4a and 4b were a pair of enantiomers successfully separated by chiral HPLC resolution. Compounds 2 and 3 indicated inhibitory activities against acetylcholinesterase with IC50 value of 57.1 ±â€¯1.6 and 32.7 ±â€¯1.0 µΜ, respectively. However, no compound showed inhibitory effect on butyrocholinesterase at the concentration of 100 µΜ.

Tanshinone I selectively suppresses pro-inflammatory genes expression in activated microglia and prevents nigrostriatal dopaminergic neurodegeneration in a mouse model of Parkinson's disease.

ETHNOPHARMACOLOGICAL RELEVANCE: Radix Salviae Miltiorrhizae, known as Danshen, is a well-known traditional Chinese herb which has been used extensively for the treatment of various diseases, including cardiovascular and cerebrovascular disease and neurodegenerative diseases for thousands of years. Tanshinone I is one of major bioactive flavonoids of Radix Salviae Miltiorrhizae. Modulation of microglial over-reaction may represent a therapeutic target to alleviate the progression of neurodegenerative diseases. Here, we tested the effect of Tanshinone I on neuro-inflammation and whether it can provide neuroprotection through inhibition of neuro-inflammation. MATERIALS AND METHODS: The effects of Tanshinone I on the production and/or mRNA expression of pro-inflammatory and anti-inflammatory factors in lipopolysaccharide(LPS)-induced BV-2 microglia cells were tested by Griess reaction, enzyme-linked immunosorbent assay (Elisa) or real time polymerase chain reaction. Activation of nuclear factor κ B (NF-κB) was measured by the nuclear translocation p65 and DNA binding activity. A model of Parkinson׳s disease was established by treatment of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) in C57BL/6 mice. The effect of Tanshinone I on the behavioral changes, dopamine and its metabolites levels, expression of tyrosine hydroxylase (TH) and IBA-1, production of cytokines in the midbrain were investigated by the rotarod test, high-performance liquid chromatography (HPLC)-ECD, immunohistochemistry and Elisa. 1-methyl-4-phenylpyridinium (MPP+) concentration was tested by HPLC. Liver toxicity was determined by biochemical assay and histochemistry. RESULTS: We found that the productions and/or expressions of several pro-inflammatory M1 factors such as nitric oxide (NO), tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß) and IL-6 were highly suppressed by Tanshinone I in LPS-induced microglia. Interestingly, it did not affect the enhancement of expression of some anti-inflammatory M2 microglia markers, including IL-10, IL-1 receptor antagonist (IL-1Ra) and Cox-2. But it could significantly inhibit LPS-induced granulocyte colony-stimulating factor (G-CSF) expression. Tanshinone I could also inhibit LPS-induced NF-κB activation in microglia. Furthermore, it improved motor functions, normalized striatal neurotransmitters, and provided dopaminergic neuronal protection in MPTP-intoxicated mice. In vivo results also indicated that Tanshinone I could modulate MPTP-induced microglial activation, attenuated the increase of TNF-α, reserved the increase of IL-10 concentrain of MPTP-intoxicated mice. Tanshinone I does not alter MPTP toxic metabolite (MPP+) concentration. Oral administration of Tanshinone I at 10mg/kg daily for 2 weeks did not show liver toxicity. CONCLUSIONS: Tanshinone I selectively suppressed pro-inflammatory M1 genes expression in activated microglia, interestingly, partially reserved anti-inflammatory M2 genes expression. It also could provide neuroprotection in a mouse model of Parkinson׳s disease. These data indicated that Tanshinone I could make the most of the beneficial side and minimize the detrimental side of activated microglia simultaneously, and provide neuroprotection by modulating the immune response of microglia.