Traditionally, herbal medicines have played an important role in treating with several diseases over thousands of years in China and attract more and more attention with few or no side effect. Unlike synthetic drugs, herbal medicines are a quite complicated system of various mixtures. Their therapeutic effect is generally based on the synergic effect of the mass constituents. Recently, various modern analytical techniques, such as high performance liquid chromatography, gas chromatography and thin layer chromatography, have been also adopted for this purpose.
Gentiana straminea Maxim. & G. dahurica Fischer, belonging to the Gentianaceae family, were two famous herbaceous perennial medicine distributing in Qinghai-Tibetan Plateau, and the dried root was the most commonly used part employed in medicine. They have been widely utilized by Tibetan folk people to relieve rheumatoid arthritis, and to remove damp-heat for the intensely bitter properties residing in the root. They were also registered by Chinese Pharmacopoeia (version 2005) as traditional herbal medicine and gentiopicroside was their marker compound of quality assessment. Currently, most studies of their chemical constituents were focused on the determination of marker compound gentiopicroside, however, other chemical constituents, such as amino acids and fatty acids etc, were not involved in. Therefore, throughout the Qinghai-Tibetan Plateau, a series of mass samplings were performed and determined by all kinds of modern analytical technologies. Relative results were published in Chromatographia (2007, 2009) and listed as follows:
1. Concentrations of 20 free amino aicds in a famous Tibetan medicine Gentiana dahurica was first investigated using 1,2-benzo-3,4-dihydrocarbazole-9-ethyl chloroformate (BCEOC) as the pre-column fluorescence derivatization reagent by reversed-phase high performance liquid chromatography. 20 amino acid derivatives were separated on a Hypersil BDS C18 column with a good baseline resolution within 65 min. Identification of 20 amino acid derivatives was by online post-column mass spectrometry with an electrospray ionization source. Results of the present study proved that the established method was rapid and reproducible for further separation and determination of FAAs in more medicinal plants.
2. A simple and sensitive high-performance liquid chromatographic method with fluorescence detection and mass spectrometric identification has been developed for analysis of 30 long-chain and short-chain free fatty acids. The fatty acids were derivatized to their esters with 1-[2-(p-toluenesulfonate)ethyl]-2- phenylimidazole-[4,5-f]-9,10-phenanthrene (TSPP) in N,N-dimethylformamide at 90 C with anhydrous K2CO3 as catalyst. A mixture of C1–C30 fatty acids was completely separated within 60 min by gradient elution on a reversed-phase C8 column. Qualitative identification of the acids was performed by atmospheric-pressure chemical ionization mass spectrometry in positive-ion mode. Quantitative determination of the 30 acids in two Tibetan medicines Gentiana straminea and G. dahurica was performed. The results indicated that the medicines contained many free fatty acids. When the fatty acid derivatives were determined in the two real samples results were satisfactory and the sensitivity and reproducibility of the method were good.
(Studied by Dr. Sun Jing, Associate Professor, Research Center for Biological Resources in Qinghai-Tibet Plateau, NWIPB,CAS)