| Arisaema rhizomatum C.E.C. Fischer is a perennial herb of Arisaema genus in Araceae and just growing in China. The herb has been used as a traditional ethnic Chinese medicine for long in history because of it's remarkably activity to eliminate dampness, dispel wind, free the flow of network vessels, quicken blood, reduce toxin or poison and relieve pains. Folk, in Enshi Tujia and Miao autonomous prefecture of Hubei, are commonly used to remedy rheumatism numb pain, treat injuries from falls, contusions and strains, manage viper bite, and alleviate turgescence for a long history because of it's remarkably activity with weak side-effect. Based on traditional folk medicine and researches which some scholars have done, we think that Arisaema rhizomatum C.E.C. Fischer can be effective in the treatment for rheumatoid arthritis (RA). The purpose of our research is to investigate the pharmacological activity of the herb on collagen-induced arthritis in mice, ascertain active site, extract and separate (medicative) compounds, and explore the potential immunological mechanisms. In addition, our studies also provide some basis for protecting and developing medicinal resources of Arisaema rhizomatum C.E.C. Fischer.1. Screening of active part(s).According to the traditional medicine and pharmacodynamics experiment results that was done previously, in order to evaluate the activity of methanol extract and extracts from petroleum ether, ethyl acetate, n-butyl alcohol and water of Arisaema rhizomatum C.E.C. Fischer, the following researches were set about. (1) Acute toxicity tests and maximum tolerance tests. In acute toxicity tests, the maximum dose of acute toxicity tests in all groups is 5 g/Kg b.w (crude drug). In maximum tolerance tests, mice were taken methanol extracts for three times in 24 h, and the total dose was 60 g/Kg b.w. After treatment, all mice were observed for one week and no toxic symptom was found. (2) Anti-inflammatory and analgesic action researches. To investigate the anti-inflammatory and analgesic activity, acute ear swelling experiment and body torsion experiment were carry out. The results showed that extracts from ethyl acetate and n-butyl alcohol were good at reducing inflammation and alleviating pain. (3) Atenuate inflammatory response on CIA. During the experiment, edema of mice hind ankle and paws, spleen index, histological changes of hind ankles, and levels of TNF-α, IL-1β, IL-6, IL-33 and RF in the serum were detected and measured. The experimental results indicated that extracts from ethyl acetate and n-butyl alcohol were more effective than the methanol extract and other parts.2. Isolation of compounds from active partsAccording to the results of selecting the active parts,12 compounds were isolated and separated from the extracts from ethyl acetate and n-butyl alcohol by using extraction method, chromatography, re-crystallization, et al.10 of these compounds were identified by various modem spectral analysis, (such as'H-NMR, 13C-NMR, HSQC, H-HCOSY, HMBC) and other classical chemical methods. Their types were fatty acids, galactosyl ceramide, sterol kind, and pyranoid ketone. Their structures were elucidated as:β-sitosterol (xlj-1), (xlj-5),5,7,4'-Hrihydroxy-3'-methoxyflavone (xlj-8),2-[(2-hydroxybexadecanoyl) amido]-4,8-octadecadiene-1, 3-diol (xlj-9), Cinnamic acid (xlj-10), Tridecanoic acid (xlj-11), Lauric acid (xlj-13), Pentadecenoic acid (xlj-15), Myristic acid (xlj-16),β-daucosterol (xlj-17). Exceptβ-sitosterol andβ-daucosterol, all these compounds were isolated from this plant for the first time; especially compound xlj-5 and xlj-18 were new compounds.3. Exploration of active mechanism and screening the activity of compounds We studied the activities of the compounds xlj-5, xlj-8 and xlj-9, and explored the active mechanism of extracts of methanol, ethyl acetate and n-butyl alcohol. Experiment in vitro suggested that compounds of xlj-5 and xlj-8 significantly reduced the levels of TNF-α, IL-1βand IL-6 in inflammation cell model reduced by LPS, restrained synovial cell proliferation, reduced levels of TNF-α, IL-1βand PGE2 secreted by synovial cells obtained from AA rats model. Extracts of methanol, ethyl acetate and n-butyl alcohol displayed the same active in RAW264.7 cell and the original generation synovial cells from knee of AA rats model. Adjusting the physiological behavior of macrophages and joint synovial cells, and reducing pre-inflammatory and inflammatory cells factor levels eventually, may be one of the mechanisms by which the Arisaema rhizomatum C.E.C. Fischer play the role of anti-inflammatory and remedy the CIA mice model.
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