Isolation, Purification And Identification Of Compounds From Centella Asiatica

Traditional Chinese medicine (TCM) has a long history of thousands of years, which is built on rich experience of herb users. Efficiency of these herbs was extensively confirmed not only by Chinese scientists but also by researchers worldwide. The concept of treating both symptoms and causes of diseases of TCM makes it especially important in modern drug development in recent years. Other advantages of TCM include low toxicity, all natural and have good disease coverage. However, TCM system has not become a major drug system in modern medicine due to the following reasons:(1) active compounds have not been identified for most herbs or herbal formula;(2) Molecular targets are also not defined for most used herbs;(3) Relashionships between active compounds and molecular targets have not been established. Part of the reasons is due to the time-consuming process in single compounds isolation. There is an urgent need in establishing high speed, high efficient and easy to operate methods for the isolation and characterization of compounds from herbs.Centella asiatica is the dry grass of Centella asiatica (L.) Urb. which widely distribute in the area of Yangtse River and south. Main chemical constituents of Centella asiatica include triterpenes andflavonoid compounds, alkenes compounds, flavonoids compounds and volatile oils etc. It has been well confirmed that those compounds have many potential uses in anticancer, antibacterial therapy. Many attentions have been paid in isolation and purificaion of active components from natural products these years. Aim of the present paper is to systematically isolate chemical compounds from Centella asiatica, which will provide basic support for deciphering active compounds from this herbal medicine.In the present paper, Continous ethanol heat reflux method was used in the extraction of crud extract from Centella asiatica. The resulting extract was subjected to a macroporous resin chromatographic column, and was fractionated into three group accoriing to polarity of the compounds contained. Each fractionation was then further fractionated by using HPGPC, HSCCC and HPLC techniques to isolate single compounds. Chemical structures of the compounds were characterized by using mass spectrometry (MS),1H-nuclear magnetic resonance (NMR) and13C NMR methods.So far, we have isolated4compounds from the strongly polar group. These compounds include3,4-dihydroxybenzoic acid, chlorognic acid,5-Di-O-caffeoylqu-inic acid,3,5-Di-O-ca -ffeoylqu-inic acid. Purities of the compounds are98.46%、100、99.23%and99.23%, respectively. The middle polar group was fractionated by HSCCC method by using a two-phase solvent system that composed of petroleum ether-ethyl acetate-methanol-water (5:5:4.8:5, v/v/v/v) at a rotate speed of850r/min and flow rate of2.0mL/min. We have isolated3compounds. Purities of the compounds are98.89%、98.28%and96.22%, respectively. Two compounds are quercitin, kaempferol respectively, another compound seem to be hydroxyl phenolic compounds. This is the first time to isolate the compounds from the middle polar group of Centella asiatica by HSCCC.Three compounds were isolated from the weakly polar group by using HPLC technique, purities of one compound is98.56%,seem to be ursane triterpenoids compound,other two compounds are belong to the naphtha, caryophyllene and longifolene respectively, purities are100%.