The Preparation And Application Of Taxifolin Molecular Imprinted Polymer

Background:Chinese medicine is a complex systerm,which contains lots of natural compounds with rich structural diversities and great difference of contents.How to capture the active compounds from the complicated traditional Chinese medicine?TCM?has hindered the further development of TCM.Moreover,it is difficult to evaluate the contribution of an effective ingredient or multiple effective ingredients to the efficacy of TCM.The development of Molecular Impringting Technology?MIT?has provided new ideas and methods for the study of TCM.The use of molecularly imprinted polymer?MIP?for the specific recognition of template molecule allows the enrichment or removal of template molecule from TCM extract for pharmacodynamic experiments.The"cross-reactivity"of MIPprovides a novel approach to separate structural analogues from complex TCM system.At present,the separation of taxifolin and its structural analogs from fructus polygoni orientalis by MIP has not been reported in literature.Purpose:Taxifolin which has anti-cancer activity was chosed as the template for MIP.The preparation conditions of the Taxifolin-MIP were optimized.By using the specific recognition ability of MIP for its template,taxifolin and its structural analogs were enriched or knocked out from the extraction of fructus polygoni orientalis.Then a negative drug solution with a lack of taxifolin and its structural analogs was obtained.The anti-cancer activity experiment was conducted to explore the difference in effecacy on the tumor suppression of of taxifolin and its structural analogs.Method:1.The synthesis of MIP:Bulk Polymerization was adopted to prepare Taxifolin-MIP and non-imprinted polymer?NIP?.The preparation conditions of the Taxifolin-MIP were optimized by single factor inspection.2.Removal of template molecules:Soxhlet extraction was used.3.The evaluation of recognition performance of MIP and NIP was carried out by using binding assay and HPLC method.4.Separation and enrichment of taxifolin from fructus polygoni orientalis and their structural analogs were carried out by solid-phase extraction by using Taxifolin-MIP as the absorbant.5.Anti-tumor activity experiment:MTT assay was used to detect the inhibitory effects of different samples on Hela cells.Result:1.The optimum conditions for preparing the Taxifolin-MIP are optimized as follow:The template molecule,functional monomer and crosslinker are taxifolin,4-VPY and EDMA,respectively,and the ratio of the three is 1:3:9.2.The solvent for Soxhlet extraction for template from MIP was methanol:acetic acid?6:4,V/V?.The recovery was 92.90%?theoretical value?.3.The Taxifolin-MIP has good selectivity for taxifolin and its structural analogs in acetonitrile.4.The Taxifolin-MIP can directly extract taxifolin and its structural analogs from the extract of fructus polygoni orientalis.5.The aqueous extract of fructus polygoni orientalis has no inhibitory effect on hela cells.The IC₅₀value of fructus polygoni orientalis extraction in methanol for Hela cells was less than that of Taxifolin.The IC₅₀value of the negative drug solution with a lack of taxifolin which was obtained by MIP-SPE was 46.27%higher than that of fructus polygoni orientalis extraction.Conclusion:The experimental results show that MIP is effective for separating and concentrating specific components of TCM.MIP can also directly remove template and structural analogs from the complex system of herbal extract.Taxifolin is an active component of the fructus polygoni orientalis which can inhibit Hela cells.The removel of Taxifolin leads to reduction of inhibitory effect for Hela cells.It also indicates that the efficacy of TCM is a comprehensive manifestation of the multi-component interaction.Using a known biologically active compound as an imprinting molecule to prepare a corresponding MIP can not only separate and enrich template molecule from TCM,but also obtain other similar structures or unknown active compounds.