The Study On The Anti-tumor Active Constituents From The Seeds Of Vitex Negundo

Objective Vitex negundo L. is large aromatic shrub distributed throughout the provinces in the south of China. The stem of this plant is used as a Traditional Chinese medicine with its primary health care needs including catarrhal rhinitis, cough, asthma, malaria, calenture, rtheumatism and arthritis. Phytochemical investigation of this plant indicates that it contains flavonoids, diterpenoids, triterpenoids and lignans. It has been reported to possess potent pharmacological properties such as anti-inflammatory, anti-rheumatic, antibiotic, hepatoprotective, antioxidant, oxidative stress, anti-androgen, snake venome neutralization and anti-allergic activities.Bioactive-targeted extraction techniques were used to screen the anti-tumor fraction from Vitex negundo L. in early stage. The 40% EtOH extract was subjected to 7 fractions, which were evaluated for the fraction with anti-tumor activity by MTT assay. The result showed that the growth of MCF-7cells were inhibited by all fractions, particularly the EtOAc fraction (Evn-50) and the fraction precipitated by EDTA disodium salt (fractionⅠ). The EtOAc fraction was investigated profoundly and systemically in early stage, so we focused on fractionⅠalso with significant growth inhibition.Method To separate and purify the chemical constituents of the fractionⅠ, several methods had been used in the process including silca-gel chromatography column (CC), ODS CC, sephadex LH-20 CC and recrystallization etc; to elucidate the structures of these compounds, spectroscopic methods and chemical evidences had been used. And then anti-tumor activity of compounds 1,2,3,4,5,6,14,15 was investigated on suppressing the proliferation of breast cancer MCF-7 cell by SRB assay.Result As a result, sixteen compounds (including eight triterpenoids, four lignans and four compounds with different types) had been isolated. The structures of these compounds were elucidated unambiguously as:2α, 3α,19α-trihydroxy- urs-12-en-28-oic acid (1),2α, 3β-dihydroxyolean-12-en-28-oic acid (2),2α,3β,19α-trihydroxy-urs-12-en-28-oic acid (3),2α,3β,19α,23-tetrahydroxyolean-12-en-28-oic acid (4),2α,3β,23-trihydroxyolean-12-en-28-oic acid (5),2α,3α,24-trihydroxyolean-12-en-28-oic acid (6), betulinic acid (14), ursonic acid (15); Vitrofolal F(7),6-hydroxy-4-(3,4-dimethoxy-phenyl)-3-hydroxymethyl-5-methoxy-3,4-dihydro-2-naphthaledehyde(8),6-hydroxy-4-(3,4-dimethoxy-phenyl)-3-hydroxymethyl-7-methoxy-3,4-dihydro-2-naphthaledehyde (10),1-sesamin (16); methyl 4-hydroxybenzoate (9), heptadecanoic acid (11),4,4'-thiobis (2-t-butyl-5-methylphenol) (12),β-sitosterol (13). Among them, compound 10 was a new compound and compound 1,3,4,5,6,16 were firstly separated from Vitex L..Study on the anti-tumor activity in vitro indicated that compounds 1, 2,3 and 15 exhibited strong activity against the proliferation of MCF-7 cells. The structure-activity relationship analysis of these eight triterpenoid compounds indicated that urs-12-en-28-oic acid triterpenoids and olean-12-en-28-oic acid triterpenoids without 23-OH or 24-OH substituent groups were the chemical constituents with anti-tumor activity from the seeds of Vitex negundo L..