Study On The Chemical Constituents And The Anti-inflammatory Activity Of Quassinoids In Brucea Javanica

Brucea javanica（L.）Merr.（Simaroubaceae）.The dried and ripe fruits are mainly distributed in Guangdong,Guangxi and other regions of China.There are 6 species of Brucea in the world and 2 species in China,which are Brucea javanica and Brucea javanica,all of which are used as folk traditional medicines in varying degrees and have a variety of pharmacological activities.According to the records of the 2020 edition of Chinese Pharmacopoeia,Brucea javanica has good effects of clearing away heat and detoxification,intercepting malaria,stopping dysentery,corroding warts and so on.It is used internally to treat all kinds of malaria and dysentery,and externally to treat warts and corns.Modern pharmacological activity studies have shown that Brucea javanica has anti-inflammatory,anti-feeding,anti-malaria,anti-virus,anti-ulcer,hypoglycemic and herbicidal effects.It is found that Brucea javanica seeds are rich in chemical constituents,including quassinoids,triterpenes,alkaloids,flavonoids,fatty acids,which determine the anti-inflammatory effects of Brucea javanica.Therefore,the chemical constituents and anti-inflammatory activities of Brucea javanica were studied from the following aspects.We systematically studied the chemical constituents of Brucea javanica.67compounds were isolated by macroporous adsorption resin HP-20,silica gel column chromatography,Sephadex LH-20 and semi-prepared HPLC,and their structures were identified by HRESIMS,¹H-NMR,¹³C-NMR and other techniques.The name is as follows:Bruceine A（1）,Bruceine B（2）,Bruceine C（3）,Bruceine D（4）,Bruceine H（5）,Bruceoside A（6）,Bruceoside B（7）,Bruceoside C（8）,Bruceoside D（9）,Bruceoside E（10）,Brusatol（11）,Dehydrobruceine B（12）,Dehydrobruceantinol（13）,Desmethyl-brusatol（14）,Deacetylated isobrucein B（15）,Javanicoside A（16）,Javanicoside B（17）,Javanicoside G（18）,Javanicoside I（19）,Javanicoside J（20）,Yadanzioside A（21）,Yadanzioside E（22）,Yadanzioside F（23）,Yadanzioside G（24）,Yadanzioside I（25）,Yadanzioside J（26）,Yadanzioside L（27）,Yadanzioside K（28）,Yadanziolide C（29）,Yadanziolide S（30）Demethyl-dehydrobrusatol（31）,Deacetyl-Yadanzioside I（32）,Javanicoside G（33）,Yadanziolide E（34）,4-quinolinecarboxylic acid（35）,3-（hydroxyacetyl）indole（36）,11-hydroxycanthin-6-one（37）,Indole-2-carboxylic acid（38）,Thymidine（39）,Excavatine B（40）,3-hydroxy-6-（hydroxymethyl）-5-methyl-1H-indole-2-carboxylic acid（41）,Bi-4-methoxycarbonyl-2-quinolone（42）,Cleomiscosin A（43）,Cleomiscosin B（44）,Cleomiscosin C（45）,Protocatechuic acid（46）,Gallic acid（47）,Vanillic acid（48）,Ferulic acid（49）,1-phenyl-1,2-ethanediol（50）,Methyl-4-hydroxybenzoate（51）,p-Hydroxybenzoic acid（52）,（S）-3-Hydroxy-3-methylglutarate（53）,（6S,9S）-roseoside（54）,Blumenol B（55）,Blumenol A（56）,Undatuside B（57）,5-hydroxymethylfurfural（58）,Luteolin（59）,3’-methoxy-luteolin（60）,Apigenin（61）,Luteolin-7-O-β-D-glucoside（62）,Kaempferol-3-O-α-L-rhamnoside（63）,Luteolin-3’-O-β-D-glucoside（64）,Quercetin-3-O-α-L-rhamnoside（65）,Apigennin-7-O-β-D-glucoside（66）,7-methoxy-luteolin（67）.Among them,compounds 31-34 and 41-42were new compounds,and compounds 35-36,38-40,47,49-55,57-58,61-67 were isolated from Brucea javanica seed for the first time.On the basis of the separation test,we found that the chemical constituents of Brucea javanica were mainly quassinoids,which were used to screen the compounds with significant anti-inflammatory activity and their anti-inflammatory activity mechanism in vitro.In this experiment,34 quassinoids compounds were screened for anti-inflammatory activity.The cellular inflammation model was established by using mouse alveolar macrophages（MH-S）induced by LPS,and the content of NO was determined by Griess reagent to determine the anti-inflammatory activity of the compounds.The anti-inflammatory mechanism of the compounds with low toxicity and the best anti-inflammatory activity in vitro was studied by ELISA kit and Western blot technique.The results showed that compounds 1-5,7,11,13,15 showed significant NO inhibitory activity,and its IC₅₀ value was 0.11-45.56μM.The other compounds have weak anti-inflammatory activity.Bruceoside B with low toxicity and good anti-inflammatory activity was selected to detect inflammatory factors in ELISA kit.The results showed that Bruceoside B could significantly inhibit the inflammatory cytokines TNF-αand IL-6,IL-1βin the supernatant of MH-S cells induced by LPS.Western blot test showed that,Bruceoside B could effectively regulate the expression of proteins related to NF-κB signal pathway and play an anti-inflammatory role.