Chemical Constituents From Leaves Of Callicarpa Arborea And Their In Vitro Anti-Rheumatoid Arthritis Activity

Callicarpa arborea Roxb Also known as"Bai-Ye-Zi-Shu"and"Nan-Yang-Zi-Zhu",it is a plant of the genus Callicarpa in Verbenaceae.It has a long history of medicine.In traditional medicine,it has the effect of treating bleeding,fever and headache,insect and scorpion bite,rheumatism and other diseases.Callicarpa arborea has folk medicinal records at home and abroad.Modern pharmacological research shows that it has many biological activities such as inhibiting inflammation,inhibiting cancer,bacteriostasis,anti-diabetes and anti-oxidation.The chemical constituents of Callicarpa arborea are diverse,but the material basis of its efficacy needs to be further explored.Objective:Through the systematic study on the chemical composition and anti-rheumatoid arthritis activity of the ethyl acetate extract from the leaves of Callicarpa arborea,in order to explore the material basis of its anti-rheumatoid arthritis effect.Methods:In this paper,methanol was used to extract the leaves of Callicarpa arborea;Silica gel column,Sephadex LH-20 gel column,MCI column,semi-preparative high performance liquid chromatography and recrystallization were used to separate and purify ethyl acetate extract from the leaves of Callicarpa arborea.According to the physical and chemical attributes of the compound,its structure was identified by ESI-MS,¹H-NMR,¹³C-NMR,HSQC,HMBC and other technologies.CCK-8 method was used to determine the cytotoxic activity of partial compounds isolated from Callicarpa arborea on Human Fibroblast-Like Synoviocytes（HFLS-RA）.Results:In this experiment,27 compounds were isolated from the ethyl acetate part of the leaves of Callicarpa arborea,21 of them were analyzed and identified as（3R,5S,8S,9R,10R）-3-hydroxycleroda-4（18）,13（14）-dien-15,16-olide（1）,6-acetoxy-cleroda-3,13-dien-15,16-olide（2）,（4→2）-abeo-16（R）-2,13Z-kolavadien-16,15-olide-3-al（3）,（4→2）-abeo-16（S）-2,13Z-kolavadien-16,15-olide-3-al（4）,cathayanalactone A（5）,oleanolic acid（6）,ursolic acid（7）,betulinic acid（8）,betulin（9）,α-amyrin（10）,β-amyrin（11）,（+）2,6-dimethyl-6（S）hydroxyl-2-trans-2,7-octadienoic acid（12）,p-hydroxybenzaldehyde（13）,eupatorin（14）,casticin（15）,1H-indazole（16）,erucylamide（17）,phytol（18）,methylα-hydroxypalmitate（19）,methyl linolenate（20）,sesamin-2,2’-diol（21）.Among them,compound 1 is a new diterpene.Except for compounds 6,7 and 9,all the other compounds were isolated from Callicarpa arborea for the first time.The cytotoxic activity of HFLS-RA showed that compounds 3 and 5 were can significantly inhibit the proliferation of HFLS-RA at 25μM～200μM;Compound 1 and compound9 had significant HFLS-RA cytotoxic activity at a concentration greater than100μM;Compound 12 at the concentration of 200μM showed significant HFLS-RA cytotoxic activity.Conclusion:Compound 1 is a new diterpene.Compounds 2～5,8 and10～21 were isolated from Callicarpa arborea for the first time.According to the screening results of cell activity,diterpenes 3 and 5 have better research value of anti-rheumatoid arthritis drugs.This paper not only enriched the research on the secondary metabolites of Callicarpa arborea,but also provides some references for excavating new anti-rheumatoid arthritis drugs from Callicarpa arborea.