Studies On The Chemical Constituents From Tibetan Medicine Compound Ruteng Capsule And Their Bioactivities

In modern medicine,rheumatoid arthritis is a common chronic autoimmune disease.It has become a worldwide problem due to its etiology and pathogenesis remaining unclear.Tibetan medicine has a unique view on rheumatoid arthritis,which belongs to the category of“yellow water disease”in Tibetan medicine.Due to the alpine region,large temperature difference between day and night,and severe climate change in Tibetan areas,rheumatoid arthritis,“yellow water disease”,become more widespread.Tibetan medicine has accumulated a wealth of clinical experience in the treatment for rheumatoid arthritis,and many unique and effective compound preparations have been developed.Compound Ruteng Capsules（CRTC）is composed of seven kinds of Tibetan medicine,including Boswellia carterii Birdw.,Terminalia chebula Retz.,Cassia obtusifolia L.,Abelmoschus Manihot（L.）Medic.,Tinospora sinensis（Lour.）Merr.,Lamiophlomis rotate（Benth.）Kudo and Pyrethrum tatsienense（Bur.Et Franch.）Ling..According to many years of clinical experience,it is a preparation for the treatment of“yellow water disease”,but the active constituents are not clear.In this study,the active constituents of CRTC was studied by the“whole prescription”method.Tibetan medicine CRTC was successively extracted by petroleum ether,ethyl acetate and methanol,and three different effective parts were obtained.The chemical composition of the ethyl acetate extract was mainly studied.29 compounds were separated by chromatography on silica gel,MCI column chromatography and semi-preparation HPLC.Their structures were identified by spectral data analysis(¹H-NMR,¹³C-NMR,2D NMR,MS,UV,etc.),these compounds were identified as:3-keto-tirucall-8,24-dien-21-oic acid（1）,3-keto-tirucall-8,24-dien-21-al（2）,3α-hydroxy-tirucall-8,24-dien-21-oic acid（3）,3β,25-dihydroxy-tirucall-8,23E-dien-21-oic acid（4）,3α,25-dihydroxy-tirucall-8,23E-dien-21-oic acid（5）,3-keto-25-hydroxy-tirucall-8,23E-dien-21-oic acid（6）,ruteng A（7）,ruteng B（8）,3α-acetoxy-tirucall-24,25-dihydroxy-8-en-21-oic acid（9）,3-keto-tirucall-8,24-dien-21,23-olide（10）,3-keto-tirucall-7,9（11）,24-triene-21,23-olide（11）,3α-acetoxy-tirucall-8,24-dien-21,23-olide（12）,acety-11-keto-β-boswellic acid（13）,β-boswellic acid（14）,9,11-dehydro-β-boswellic acid（15）,α-boswellic acid（16）,nephthenol（17）,acetyl incensole oxide（18）,incensole（19）,populeuphrines A（20）,isonincensole acetate（21）,13-keto-verticilla-4（20）,7,11-triene（22）,6α-hydroxy-15-（3-methyl-2-butenyl）alloaromadendrane（23）,aurantio-obtusin（24）,obtusin（25）,chrysophanol（26）,physcion（27）,gallic acid（28）and bakuchiol（29）.The main types of compounds are triterpenoids（1-16）,diterpenoids（17-23）,anthraquinones（24-27）,and others（28-29）.Among them,compounds 2,4-9,12,22-23 are new compounds.In order to evaluate the anti-inflammatory activities of 29 compounds,CCK-8method was used to determine the cytotoxicity.RAW264.7 cells induced by lipopolysaccharide（LPS）were used as the inflammatory model,NO content in the culture supernatant was measured by Griess method.The results showed that at the concentration of 20μM,29 compounds had no cytotoxicity to RAW264.7 cells,the survival rate was more than 80%,and the inhibition rates of NO were less than 50%.29 isolated compounds were screened for anti-tumor activities in vitro.The antiproliferative activities of 29 compounds against human pleomorphic malignant glioma T98,human liver cancer cell Hep G2 and human breast cancer cell MCF-7 were investigated by the CCK-8 method.The results showed that at a concentration of 20μM,29 compounds had no significant anti-proliferative activities against the tested tumor cells,and their inhibition rates were less than 50%.