Anti-inflammatory Constituents And Quality Evaluation Of Vitex Negundo Fruits

With the development of industry and the rapid aggravation of our ecological environment, the incidences of inflammation-related diseases are globally increasing. Therefore, it is necessary to investigate and develop new anti-inflammatory drugs from natural source. Fructus Viticis Negundo named Huang-Jing-Zi is the ripe fruits of Vitex negundo L. (Verbenaceae) and is taken to dispel the wind, relieve cough and dyspnea, regulate qi, and relieve pains in Traditional Chinese Medicine. It is commonly used in clinical practice for curing asthma, chronic tracheitis, gastrelcosis, chronic gastritis and rheumatoid arthritis in folk medicine. However, the insufficient research of Fructus Viticis Negundo and the ambiguity of its bioactive components resulted in unable quality control of this crude drug. So the confusion of species and irregular quality will surely bring adverse effect to its application. The main contents of our study are as following three aspects:1 Study on bioactive components of Vitex negundo fruits1.1 Bioactivity guided isolation of anti-inflammatory fractions and compounds was undertaken by various chromatographic methods. The active fractions–dichlormethane fraction (DF) and acetoacetate fraction (AF) were obtained from the 80% ethanol extract of Vitex negundo fruits. A total of 21 new with 58 known compounds were isolated from these active fractions and their structures were elucidated by detailed spectroscopic analyses on the basis of NMR, IR, and MS data. The major types of the isolated compounds were lignans, diterpenoids and triterpenoids, flavones, steroids and phenolic small molecular compounds. Compound VN-1 was proved to be the active compound with anti-inflammatory activity in xylene induced ear edema in mice. Phenylnaphthalene-type lignans were found to be the principal anti-inflammatory constituents in Vitex negundo fruits.1.2 The lipophilic components of petroleum ether fraction of Vitex negundo fruits were analyzed through GC-MS method. A total of 11 compounds were identified, comprised of (un)saturated fatty acids, of which linoleic acid was the major component with the relative content of 57.22%.2 Pharmacological research of Vitex negundo fruitsEthanol extract of Vitex negundo fruits, DF and AF showed significant anti-inflammatory and analgesic effects in mice xylene induced ear edema test, writhing test and formalin test. They exhibited significant inhibition on acute inflammatory edema and chemical-stimulated inflammatory pain. Among the pure compounds, phenylnaphthalene-type lignans including VN-12, VN-50, VN-52 and VN-53 showed high NO production inhibitory and antioxidant activities. The introduction of free hydroxy groups plays a vital role in the potency of these compounds. Moreover, VN-1 showed significant anti-inflammatory and analgesic effects in vivo. In addition, diterpenoids including VN-38 and VN-64 also possessed notable nitric oxide scavenging activity, which may be related to their inhibitory effects on the protein expression of iNOS in LPS-stimulated Raw 264.7 cells.3 Quality evaluation of Vitex negundo fruits3.1 Samples of 13 populations of Vitex negundo fruits were collected from 9 provinces and manicipalities of China. Studies on chemical fingerprint of of Vitex negundo fruits were performed by HPLC-UV-DAD. The HPLC data showed considerable variation of chemical constituents among the samples collected from different populations, with chemical profiles varied from each other. The hierarchical clustering analysis (HCA) further revealed that 13 samples were classified to three types (Ⅰ,ⅡandⅢ-type) based on their varied HPLC data, which were consistent with three chemotypes divided intuitively according to the appearance of those 13 HPLC chromatograms.3.2 Studies on the simultaneous determination of 4 phenylnaphthalene-type lignans by HPLC-UV method were performed for quality evaluation. Thirteen samples from different populations and sites of China were qualitified and the difference between them was proved significant. Compound VN-1 (6-hydroxy-4β- (4-hydroxy-3-methoxyphenyl)-3α-hydroxymethyl-7-methoxy-3,4-dihydro-2-naphthalde-hyde) is suggested to be one of the main indices for quality evaluation due to its abundance in most samples and significant antiinflammatory properties both in vitro and in vivo. Thus our findings provided scientific basis for the quality control, further research and development and clinical use of Vitex negundo fruits.