The Study On Isolation Of Chlorogenic Acid Form Ilex Kudingcha C.J.Tseng And Their Anti-Obesity Efficacy

The population of people suffering from adiposity has been increasing rapidly in recent years. As a result, for the first time in history, the number exceeds both the population of people suffering from hypoalimentation and infection. Adiposity has been defined as a global epidemic by World Health Organization (WHO). It has been proved by medical research that obesity is related to a lot of diseases and it has become the prominent factor endangering the level of public health and standard of living in our country. However, diet pills have more or less some side-effects. Besides, the likelihood of rebounding is very high when stopping taking them. Consequently, the search for effective cure of adiposis from natural food source has been a hotspot in the research area of adiposis prevention and treatment.Kudingcha can be mainly divided into two species:Folium llicis Latifoliae and Ligustrum. With unique physiological functions, chlorogenic acid is a kind of active substance taken from Kudingcha, and it has become more and more popular for its outstanding function in lowering the level of blood lipid and blood glucose, and its effect of anti-oxidation. This paper mainly focuses on, first, the preparation of chlorogenic acid from Ilex kudingcha C.J.Tseng and its derivatives; second, the creation of a rapid method to measure the content of chlorogenic acid with the help of HPLC; last, the discussion on the effects of extracts from Ilex kudingcha C.J.Tseng on3T3-L1adipocytes. The article aims to be of some referential value for the study of its anti-obesity mechanism.Macroporous resin HP20was used to preliminarily separate and purify TP, the crude product of chlorogenic acid which was abstracted from Ilex kudingcha C.J.Tseng. The resulting components of elution by water and70%ethanol in turn were monosubstituted caffeoylquinic acid compounds (TP I) and disubstituted caffeoylquinic acid compounds (TP II). Next, through semi-preparative HPLC and mobile phase of methanol and water, six monomers were separated by gradient change. Identified by NMR. the six substances are3-O-CQA,4-O-CQA,5-O-CQA,3,4-O-diCQA,3,5-O-diCQA and4,5-O-diCQA. By adopting HPLC, standard curve can be depicted according to the linear relationship between sample quantity and peak area. The relationship between5-O-CQA,3,5-O-diCQA and peak area is quite linear when their injection quantity is within the range of0.1μg-0.2ug. The average recovery rate is99.33%and99.23%respectively, and RSD (n=5) is0.91%and0.35%. It is proved to be a simple, rapid and accurate quantitative analysis method. In this paper, the quantity of chlorogenic acid and its derivatives in Ilex kudingcha C.J.Tseng, Ilex lartifolia Thunb and Ligustrnm rabutum (Roxb)Blume is compared, which are119.95mg/g,104.87mg/g and50.5mg/g respectively.3T3-L1preadipocytes are used as target cells to explore the effect of TP, TP I、 TP II on their proliferation measured by MTT assay and we can determine the growth inhibition (IC50) of cells, respectively are180μg/mL,240μg/mL、200μg/mL. Besides, cell differentiation using differentiation cocktail, oil red O staining and spectrometric determination were carried out for investigating the effect of TP, TP I, TP II on the differentiation of3T3-L1preadipocytes.We can observe the changes between3T3-L1preadipocytes and adipocytes through inverted microscope.Results: TP、TP Ⅰ、TPⅡall could inhibit the proliferation and differentiation of3T3-L1preadipocytes in comparison with the blank groups,especially TP II. Conclusion:Ilex kudingcha C.J.Tseng appear to have potential for the prevention of obesity.