Study On The New Technology For Separation And Preparation Of Bioactive Compounds From Eucommia In High Pure Form

In this dissertation, modern technologies such as preparative high performance liquid chromatography (P-HPLC), flocculation by natural clarification and macroporous resin were used to purify the four bioactive compounds and two kinds of active mixtures from Eucommia (EU) under the guidance of relative theories. Some significative results were obtained as follow:1. Study on the extraction techniques. Base on the choice of various raw materials, fresh bark was chosen to extract Iridoids compounds such as geniposidic acid (GPA), aucubin (AU) and geniposide (GP). Leaves were used to extract chlorogenic acid (CA). Traditional and new extract technologies such as hot solvent extraction; ultrasonic wave extraction and microwave extraction were used to determine the suitable method to extract active mixtures. The experimental results show that ultrasonic-extraction is quick and efficient, but has low selectivity. Microwave extraction is the best one. It not only is a quick and efficient method but also has high selectivity, low energy consumed.2. Study on the separation and purification methods. The bark extraction was dealt with solvent extract, adsorption by macroporous adsorption resin or special adsorbent to enrich and purify Iridoids. So the materials used in preparative HPLC were obtained. Instead of ethanol sedimentation, natural clarificant combing with ultra-filtration technology were performed to get CA extract from the leaves of EU. It shows that these techniques are simple, low cost and easy to enlarge.3. GPA, AU, GP and CA were prepared by PHPLC method. A new technology was presented that three target compounds could be obtained in one step.Chromatographic behaviors of GPA, AU, GP and CA in PHPLC were investigated in the experiments. It was the first time to establish mathematics model to describe their retention behavior on PHPLC system, which can be used to guide the optimization of mobile phase. The relationship between the retention behavior and the structure of Iridoids was discussed.Suitable chromatography conditions were obtained to prepare GPA, AU and GP in one step or CA, such as the composition of mobile phase, flow rate and elution mode, et al. Flow rate gradient and stepwise gradient modes were presented originally. The products of GPA and AU can feed the necessity of our country. The recycle of the large amount of mobile phase, comprehensive utilization of the resources of EU and green chemical engineering were investigated in the whole process.4. Flexible analytical methods of effective components in EU were established in the dissertation. Simultaneous and quick determination methods of three main Iridoids in EU bark or GPA and CA in EU leaves or Chinese patent medicin by FIPLC were prestnted. HPLC-MS was adopted to analysize Iridoids in EU samples. It was proved to be repeatable, specific, accurate and easy to operate. A high sensitive spectrophotometry to determine AU was built.Uniform experimental design was used to optimize the analysis condition of AU. Back propagation artificial neural network (ANN) technique was applied to forecast the result.5.The structure elucidation of GPA, GP, AU and CA were carried out by spectroscopic techniques such as UV, IR, NMR (13C and 'H) and MS. The stabilities of the four compounds in the condition of different pH, temperature, light and organic solvents were discussed.6. It was the first time to study on the effect of CA on Alzheimer disease (AD) in this paper, and satisified results were obtained. And the related animal model has been built as well.