Purification,Structural Modification Of Flaxseed Lignan And Biological Activity Of Its Derivatives

Lignans are phytoestrogens that are very similar to the physiological activity of human estrogens.The main plant source for this natural product is flaxseed.Studies have shown that lignans display a variety of biological activities and thus can act as netruceuticals.However,due to the presence of several hydrophilic hydroxyl groups,flaxseed lignans are poorly soluble in fat,meanwhile the high-purity lignans on the market are expensive,which limits their use in oils and related products.This thesis focuses on improving the fat solubility of purified flaxseed lignans,through modifying the hydrophilic groups of flaxseed lignan by chemical or enzymatic methods,thereby effectively improves its functional properties.Then the biological activity of flax lignan and its derivatives was evaluated.The method for improving the fat solubility of flaxseed lignan is achieved by two major means:viz.esterification of the the glycosidic part of flaxseed lignan with long-chain fatty acids;and removal of the glycoside part of flaxseed lignan.The main results obtained are as follows:1.By using macroporous resin method and silica gel column chromatography,AB-8macroporous resin was selected to separate and purify the flaxseedseed extract.The purity of flaxseed lignan SDG was determined as 80.2%,with a yield of 74.9%.The crude purified lignans were then purified via silica gel column chromatography.The final purity of the obtained SDG products was up to 94.7%,with a overall yield of 58.9%,which means the purification protocol is robust.2.The di-esterified flaxseed lignan SDG derivatives were prepared by treating the starting material with a series of medium or long chain fatty acid vinyl ester under the metal catalyst ZnTAC24,by such means,the esterification took place at the G6,6'position of the glycoside.The products were named as d-SDG8.d-SDG10,d-SDG12,d-SDG14,d-SDG16,d-SDG18;moreover,when SDG was treated with excess acetic anhydride,the associate acetate?SDGAA?was obtained.Meanwhile,an enzymatic selective esterification on SDG and medium or long chain fatty acids performed by Novozym435,the mono-esterified products?at G6'position?were then named as s-SDG8,s-SDG12 and s-SDG16,respectively.The structures of all above-mentioned products were confirmed by nuclear magnetic resonance spectroscopy?NMR?,low resolution mass spectrometry?MS?and Fourier transform infrared spectroscopy?IR?.3.The optimized condition for hydrolysis of SDG to associate SECO was determined as follows:dissolving SDG in hydrogen chloride dioxane solution,the favorable temperature was40°C.The reaction was carried out at 300 rpm for 4 h,and the yield can be up to 37.3%.The same process can also be completed via cellulose.The results showed that 30 U/mL of cellulase 1under 60°C is capable to hydrolyze the SDG in a 48 h period,with a yield of 43.6%.4.Three different in vitro antioxidant effects evaluation systems,namely,the DPPH free radical scavenging ability,ABTS free radical scavenging ability and oxygen free radical absorptive capacity?ORAC?,were applied to determine the antioxidant potencies of the obtained derivatives.The cytotoxicities of each products against the normal liver cell LO₂,breast cancer cell MCF-7?with hormone stimulation?,breast cancer cell MDA-MB-231?hormone-free stimulating?as well as hepatocellular carcinoma cell HepG2 were evaluated by MTT essay.The results showed that the in vitro antioxidant capacity of SDG and its derivatives were in order of:hydrolysate>SDG>di-substituted esterification product>SDG acetate;in cell model,SDG and its derivatives promoted cancer cell proliferation while the SECO inhibits the proliferation.In those esterified derivatives,the anti-proliferative potencies decreased when the alkyl side chain incrased.The current work has developed various structural modification methods for SDG and prepared several derivatives of it,the systemic biological evaluation has provided a basis for further expanding the application range of flaxseed lignan.