Study On Chemical Constituents Of Schisandra Chinensis And Evaluation Of Neuroprotective Activity

Natural products play an important role in modern drug development as one of the sources of drugs or lead compounds for the treatment of major human diseases.Surveys show that more than 50%of new chemical drugs developed internationally are directly or indirectly derived from natural products between 1981 and 2019.Natural products provide us with compounds of rich types,diverse structures and significant biological activities,which provide an important resource for the development of drug lead molecules or drugs.In the present study,the active ingredients in natural products were explored,and Schisandra chinensis was selected as subject of the study.The active ingredients in Schisandra chinensis were extracted and separated using various column chromatographic methods,and the structures of 20 compounds obtained from the separation were identified by using wave spectrometry,combined with spinodal determination to resolve the compound conformations.Based on the network pharmacological investigation,the isolated lignans were evaluated for their anti-neuroinflammatory activity and neuroprotective activity under oxidative stress injury.the compounds with significant anti-neuroinflammatory activity were initially explored mechanistically using western blot technique.The identification of quality markers（Q-markers）of Schisandra chinensis was also predicted and analyzed in combination with phytopharmacology,compositional measurability,and network pharmacology.The main research results obtained are as follows:（1）20 compounds were isolated and purified from the petroleum ether,ethyl acetate and ethanol parts of the supercritical fluid extract of Schisandra chinensis and structurally characterized,including:10 biphenylcyclooctenyl lignans,namely:Schisandrin B（1）,Schisandrin C（2）,Gomisin K₁（3）,Arisanschinin F（4）,Gomisin F（5）,Schisphenin D（6）,Schisantherin D（7）,7（18）-dehydroschisandro A（8）,Sieverlignan E（9）,Gomisin D（10）;2,3-dimethyl-1,4-diarylbutane 2 lignans:meso-dihydroguaiaretic acid（11）,pregomisin（12）;2,5-diaryltetrahydrofurananyl lignans:magnolin（13）,epimagnolin（14）;1 steroidal compound:β-sitosterol（15）;3 organic acid derivatives:1,5-dibutyl-1’-methyl citrate（16）,shikimic acid n-butyl ester（17）,diethyl malate（18）,2 other types of compounds,namely:5-hydroxymethyl-2-furaldehyde（19）and Bis（2-ethylhexyl）phthalate（20）.Among them,compound 8 is the first natural compound isolated from nature,compound 4 is the first isolated from Schisandra chinensis,and compounds 9 is the first isolated from Schisandra Michx.（2）Based on the network pharmacology study,we searched for the association targets of the components and diseases.The GO enrichment analysis was carried out in terms of biological processes,cell composition and molecular functions,and the KEGG pathway analysis was also performed.The"component-target-pathway"network of Schisandra chinensis and its nodes were synthesized,and the analysis revealed that the lignan-like components of Schisandra chinensis are closely related to the treatment of Parkinson’s disease,and their targets involve neuroinflammation and oxidative damage,so the in vitro pharmacological activities of these two aspects were investigated.LPS-induced BV2 cells were selected as a model of neuroinflammation,and the isolated lignans were evaluated for their anti-inflammatory activity.5 compounds with significant anti-neuritis activity（1,7,8,9,10）were screened,and their NO inhibition rates at 10μM concentrations were above 50%.The compound 8 with the highest rate of inhibition,were determined by IC₅₀ and the results was 11.95±0.26μM.further preliminary studies on the anti-neuritis mechanism of compound 8 was conducted,and the results of Western blot experiments showed that compound 8 down-regulated the expression of i NOS and COX-2 to some extent.H₂O₂-induced PC-12 cells were selected as a model of oxidative stress,and the MTT method was used to detect the survival rate of the cells after compound action and to evaluate their neuroprotective activity under oxidative stress.The experimental results showed that compounds 3,4,6,11,12 had significant protective effects on H₂O₂-induced oxidative damage in PC-12 cell,and the cell survival rate was above 70%at10μM concentration.The most significant activity was found in compounds 11 and 4,with cell survival rates of 93.15%and 88.12%respectively.（3）The chemical components in Schisandra chinensis that can be used as Q-markers were predicted,and deoxyschisandrin,Schisandrin B,Schisandrin C,Schisandrol A,gomisin A,gomisin G were reasonably predicted as quality markers of Schisandra chinensis through plant affinities,component effectiveness,in vivo absorption,and network pharmacology,combined with qualitative analysis of the components,to improve the quality evaluation criteria of Schisandra chinensis.