| Tea is abundant in polyphenols, which accounting for18%-360of the total dry weight. Tea polyphenols include flavanols, flavonols and their glycosides, anthocyanidins and leucoanthocyanidins, as well as phenolic acids and depsides. Catechins, a kind of flavanols, account for120～24%of the total dry weight of fresh tea leaves and70%～80%of total tea polyphenols. Theaflavins, oxidized from a pair of catechins during black tea fermentation, are major compounds in black tea which account for1%～5%of its dry weight. Both catechins and theaflavins are the important compounds which not only contribute to tea taste, but also provide the health effects. It have been shown that catechins and theaflavins are excellent natural phytochemicals with many biological activities and pharmacological actions including anti-oxidation, free radical scavenging, anti-lipid, anti-bacteria, anti-allergy, preventing atherosclerosis, and anti-radiation. Along with the deepening of the pharmacological reasearches of catechins and theaflavins, separation and preparation of catechin and theaflavin monomers become more prominent. So, it is important to explore a simple but efficient method with low-cost to prepare those monomers. Based on this purpose, the preparative medium pressure liquid chromatography (MPLC) have been explored and successfully applied to obtain the monomers of catechins and theaflavins in this thesis. The major content and accomplished results are as following:1. The methods for isolating and preparing catechins and theaflavins were reviewed based on the retrieved references. The current common methods include silica gel column chromatography, Sephadex LH-20column chromatography, adsorption column chromatography, preparative high pressure liquid chromatography (HPLC) and high-speed counter-current chromatography (HSCCC). As a kind of liquid chromatography between traditional column chromatography and preparative HPLC, MPLC featured with large sample loading, high effeciency, time saving and low cost, is a popular way to separate and prepare pure phytochemicals in a small-scale preparation.2. A MPLC system coupled with a C18(ODS-AQ) column has been applied to separate catechin monomers from tea polyphenols with the purity of92.6%, and consequently four major catechin monomers as epigallocatechin (EGC), epigallocatechin gallate (EGCG), epicatechin (EC) and epicatechin gallate (ECG) were obtained just in95min separation. Their purity are up to91.8%,97.6%,97.7%and99.3%, respectively. The yield and recovery of total four catechin monomers are up to56.0%and68.2% respectively.3. In this thesis, a selected MPLC system with C18silica columns has also been successful to separate theaflavins from the tea pigments containing65.2%of theaflavins. After190min separation, three kinds of theaflavins as theaflavin (TF1), theaflavin monogallate (TF2a,2b) composing of theaflavin-3-gallate (TF2a) and theaflavin-3’-gallate (TF2b), and theaflavin digallate (TF3) were prepared. Their purity are up to92.6%,98.9%, and98.7%, respectively. The total yield and recovery of three theaflavins are up to51.5%and79.0%, respectively.4. In order to separate TF2a and TF2b, an optimal MPLC separation condition has been used to obtain four theaflavin monomers after550min separation. The purity of four theaflavin monomers TF1、TF2a、TF2b、TF3are92.6%,96.5%,94.8%and98.7%respectively. The total yield and recovery of the four theaflavins are up to53.5%and82.1%respectively.5. Finally, the structural and molecular information of the harvested catechins have been confirmed by wavelength scanning, H-NMR (Nuclear magnetic resonance),13C-NMR and mass spectrometry, while the theaflavin monomers have been confirmed by wavelength scanning,’H-NMR and13C-NMR. |
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