Studies On Chemical Constituents From Pogostemon Cablin And Their NO Inhibitory Activities

Pogostemonis herba,as a famous traditional Chinese medicine,is the dry aerial parts of the plant Pogostemon cablin（Blanco）Benth.It is a characteristic Chinese medicine in Lingnan region and is distributed in Guangdong,Guangxi,Taiwan,Hainan,Fujian and other provinces.Pogostemonis herba is commonly used as an aromatic herb for resolving dampness in clinic,such as vomiting,summer heat-dampness,febrile diseases,acute and chronic sinusitis,headache and so on.It is effective in the treatment of gastrointestinal discomfort and fever caused by summer heat-dampness.Besides,it was found in various recommended formulas and Chinese patent medicines of Chinese Clinical Guidance for COVID-19 Pneumonia Diagnosis and Treatment(Trial 8^th Edition)based on its function of supporting the normal and dispelling the evil aspect.The volatile oil of P.cablin has already been used for a long history,and has been widely and deeply studied on its chemical components and pharmacological effects.On the contrary,there are relatively few studies reported on non-volatile components.Modern pharmacological studies show that P.cablin has good anti-inflammatory action,and its anti-inflammatory substance has always been considered related to its essential oil,but some studies have shown that its aqueous extract also has anti-inflammatory activity,which indicate the possible anti-inflammatory activity of the aqueous extract.In order to further explore the anti-inflammatory pharmacological substance basis of P.cablin and improve the types of its chemical constituents,we plan to systematically separate and identify components from the aqueous layer of P.cablin 95%ethanol extract extracted by ethyl acetate.Then we will search for natural small molecule compounds that can significantly inhibite NO production through the test in vitro.A total of 45 compounds were isolated and identified from the aqueous layer after the extraction of ethyl acetate of Pogostemon cablin by means of various chromatographic methods and spectral analysis techniques,which were as follows:crenatoside（1）,3′-methoxy crenatoside（2）,isocrenatoside（3）,3′-methoxy isocrenatoside（4）,acteoside（5）,leucosceptoside A（6）,osmanthuside B（7）,jionoside C（8）,isoacteoside（9）,plantainoside C（10）,martynoside（11）,3′′-O-acetylmartynoside（12）,2′′-O-acetylmartynoside（13）,4′′-O-acetyl-martynoside（14）,2′′,4′′-O-acetylmartynoside（15）,2′′,3′′-O-acetylmartynoside（16）,（R）-campneosideⅡ（17）,（S）-campneosideⅡ（18）,（R）-campneosideⅠ（19）,（S）-campneosideⅠ（20）,decaffeoyl isocrenatoside（21）,lariketoester（22）,isolariketoester（23）,（+）-isolariciresinol（24）,icariol A₂（25）,（+）-（7R,7′R,8S,8′S）-neo-olivil（26）,（-）-（7′S,8S,8′R）-4,4′-dihydroxy-3,3′,5,5′-tetramethoxy-7′,9-epoxylignan-9′-ol-7-one（27）,glycosmisic acid（28）,tortoside F（29）,erythro-guaiacylglycerol-β-ferulic acid ether（30）,threo-guaiacylglycerol-β-ferulic acid ether（31）,（7R,8R,8′R）-4′-guaiacylglyceryl-evofolin B（32）,cistanoside F（33）,isocistanoside F（34）,guaiane A（35）,30β-hydroxyarjungenin（36）,29α-hydroxyarjungenin（37）,cytosporone W（38）,cytosporoneⅤ（39）,cytosporoneⅥ（40）,naringenin-7-O-β-D-glucopyranoside（41）,luteoloside（42）,3,4-dihydroxybenzaldehyde（43）,tianshic acid（44）,（9S,12S,13S）-trihydroxyoctadeca-（10E,15Z）-dienoic acid（45）.Among them,compounds 1～21 are phenylethanol glycosides,22～34 are penylpropanoids,35 is sesquiterpene,36～37 are triterpenes,38～40are octaketides,41～42 are flavonoid glycosides,43 is phenol,44～45 are fatty acids.Compounds 22～23,35～38 are six new compounds that had not been reported.Compounds 17～18 have been separated for the first time.Compounds 4,7～8,10～16,24～28,30～32,34,41～42,44～45 are isolated from Pogostemon cablin for the first time,and compounds 17～18 have been separated from natural products for the first time.Compounds 1～18 and 22～45 were evaluated for their anti-inflammatory activities by the inhibitory effects on LPS-induced NO production in RAW 264.7 cells.The results showed that compounds 22,32,37,42 and 43 had significantly inhibitory effects at a dose of 25μM.Subsequent MTT assay results showed that the above five compounds at 25μM had relatively little effect on cell viability except compound 37.The structure-activity relationship analysis showed that all these five compounds had free phenolic hydroxyl groups except for compound 37,and that pyrocatechol groups could improve inhibitory effects significangtly,which preliminarily indicated that free phenolic hydroxyl groups may be the active group to inhibit the production of NO.In order to search for anti-inflammatory active ingredients,the above compounds need to be screened for other anti-inflammatory models.