Study On The Chemical Constituents And Bioactivities Of The Fruiting Bodies Of Ganoderma Cochlear

Ganoderma cochlear has a long history used both as medicine and edible mushroom.Its fruiting body has multiple biological activities,such as anti-aging,improving memory,anti neurasthenia,anti-cancer,protecting liver,reducing blood lipid,anti-inflammatory and so on.In order to further study the structural diversity and related biological activities of chemical components of G.cochlear,the liposoluble components of fruiting bodies were systematically studied in this paper.The extract was separated and purified by normal phase column chromatography,RP-18 column chromatography,Sephadex LH-20 and other conventional column chromatography combined with preparative high performance liquid chromatography（HPLC）.The structure of the monomer compound was identified by nuclear magnetic resonance（NMR）,ultraviolet（UV）,mass spectrometry（MS）,single crystal X-diffraction and other comprehensive spectroscopy methods and analytical techniques.Finally,29 compounds were isolated and identified from the fat soluble extract of G.cochlear fruiting body,and their structures were mainly divided into two categories:triterpenes and meroterpenoids.18 lanostane triterpenoids were isolated,of which 13（1,2,3,4,5,6,7,8,9,10,11,17,18）were new 3,4-seco-25,26,27-trinorlanostane triterpenoids,and compounds 4-6 were 3,4-seco-25,26,27,28-tetranorlanostane triterpenoids of new skeleton,and 4 was further rearranged,form a new carbon skeleto of tetrameric dialkyl triterpenes.11 Ganoderma meroterpenoids,including benzene heteromonoterpene single chain type,polycyclic type,and a new benzene heteromonoterpene dimer（20）According to the literature research and the previous research experience of the research group,we tested the neurotrophic activities of eight triterpenoids with a yield of more than 5mg.The results showed that these compounds had different degrees of NGF dependent activity to promote the growth of PC12 cells,of which 3,11,13 and 15 had the best activity.We selected 11 and 15（3μM,10μM,30μM）conduct further concentration gradient test.The results showed that compound 15 promoted the protrusion growth of PC12 cells in a dose-dependent manner,while compound 11 enhanced the activity of protrusion growth of PC12 cells in a concentration dependent manner at low concentration,but at high concentration（30μM）.It shows the effect of inhibiting the growth of protrusion.In order to explore the relationship between the two compounds and neurite growth,the expression of growth associated protein（GAP43）was measured by Western blot.The results showed that 15 up-regulated the expression of GAP43 in a dose-dependent mannerwhich was consistent with the law of neurite growth.The expression of GAP43was indeed up-regulated at low concentration,but it was inhibited at high concentration,which was consistent with the cell level experiment.In addition,meroterpenoid dimer 20 shows cardioprotective effect,which can effectively inhibit the increase of Fe²⁺in cardiomyocytes induced by hypoxia reoxygenation,reverse the decrease of GPX4 and FTH1 caused by hypoxia reoxygenation,and improve the survival rate of cardiomyocytes.To sum up,this study further enriched the species and compound diversity of lanostane triterpenes in the fruiting bodies of G.cochlear,resulting in three lanostane triterpenes with carbon reduction and further rearrangement,which is conducive to further understanding the biosynthetic pathways of lanostane triterpenes.Through the study of cell level and protein level,the dose effect relationship and time effect relationship of neurotrophic activity of related compounds were discussed.This study revealed the material basis of the neurotrophic and cardioprotective effects of G.cochlear,and pointed out the direction for the application and further development of this special drug and food homologous fungus in neurodegenerative diseases and heart related diseases.