Total Synthesis And Optimized Preparations Of An Isoflavone C-glycoside Geonone

Isoflavone C-glycosides,in which the sugar moiety is attached by a C–C bond directly to the isoflavone ring,are not easily hydrolyzed in acidic gastric juices compared with O-glycosides and aglycone.These glycosides exhibit various biological activities such as radioprotective,anti-myocardial ischemic and antidiabetic activities.The total synthesis of isoflavone C-glycosides only involves the chalcone pathway starting from C-glucosylacetophenone and employes the highly toxic thallium?III?nitrate in the oxidative rearrangement of the chalcone.Numerous aryl C-glycosides without acetyl groups are not used in synthesizing isoflavone C-glycosides.Hence,development of efficient synthetic method is a significant task for drug discovery of C-glucosylisoflavonesPuerarin which is found mainly in Pueraria radix shows strong anti-myocardial ischemic effects;it expands the coronary artery and cerebrovascular system,significantly reduces myocardial oxygen consumption,and improves cardiac systolic function.Puerarin has hemolytic side effects,poor permeability and poor water-solubility,which restricts its clinical application greatly.3'-?dimethylamino?methyl puerarin hydrochloride?geonone,G20?with good water solubility was developed and synthesized by our research group.Its anti-myocardial ischemic and anti-brain ischemia activities were better than those of puerarin.More importantly,we observed no hemoglobinuria in rats when G20 was administered intravenously.Three problems exist in the current study of G20:1)G20,semisynthesized from puerarin,lacks a reasonable synthetic method.2)G20 has a poor oral bioavailability and should be frequently administered for maintaining an effective blood drug concentration or effective local content.Therefore,an oral dosage forms of G20 should be developed.3)Target proteins of G20 on anti-myocardial ischemic are not clear.The thesis has made such main progress:1.In view of the fact that some shortcomings exist in total synthesis of isoflavone C-glycoside.Two synthetic routes were developed to totally synthesize an simple isoflavone C-glycoside?6-tert-butylpuerarin?:an improved chalcone pathway and a novel deoxybenzoin pathway.Firstly,the first total synthesis of 6-tert-butylpuerarin was achieved through an improved chalcone pathway with an overall yield of 2.8%.We developed a cyclization method using DIB,an environmentally friendly reagent,instead of thallium trinitrate in the oxidative rearrangement of the key intermediate chalcone.Secondly,two isoflavone C-glycosides?6-tert-butylpuerarin and6-tert-butyl-4'-methoxy-puerarin?was totally synthesized through a simple deoxybenzoin route in five steps for overall yields of 14.6%and 14.2%.Compared with the traditional chalcone pathway,this pathway represents a novel synthetic pathway based on Vilsmeier-Haack cyclization to achieve isoflavone C-glycosides in high yields.Two synthetic routes were beneficial for the total synthesis of G20 and promoted the research and development of isoflavone C-glycosides.2.The total synthesis of G20 was achieved through the improved chalcone pathway and deoxybenzoin pathway for overall yields of 0.5%and 1.6%.The alkyl substituents?ethyl/4-CH₃OC₆H₄CH₂CH₂?with a mild electron donating ability were played on the aromatic ring as a key rule for the enhancement of reactivity and regioselectivity of O-C rearrangement,which made it more available for the next de-tert-butylation and oxidation.The isoflavone ring of G20 could be constructed by the chalcone pathway of acetophenone 21 or deoxybenzoin pathway of deoxybenzoin16 to obtain puerarin.Finally,the Mannich reaction was used to convert puerarin to G20.This ethyl substitution strategy was suitable for the total synthesis of various C-glucosyisoflavones.3.In order to develop G20-phospholipid complex to enhance its oral bioavailability.Solvent evaporation method was used for preparing G20-phospholipid complex.Recombination rate was served as indicating index,the reaction conditions were optimized by orthogonal test to investigate the key factors,such as reaction temperature,reaction time,solvent styles and drug-lipid ratio.The obtained complex was analyzed by X-ray diffraction.SD rats were intragastrically administered with G20 and its phospholipid complex,respectively.The blood concentration of G20 was detected by HPLC,after which the pharmacokinetic parameters were calculated.The results showed that the optimum reaction conditions were as follows:at the temperature of 50?for 2 h,the dosage of ethanol was 40mL,the Moore ratio of G20and phospholipid was 1:2.The recombination rate was 100%.G20 existed in an amorphous state in the phospholipid complex,which was a new phase rather than a physical mixture.The relative bioavailabilities of G20 in rats after oral administration of G20-phospholipid complex were enhanced by 1.60-fold.The mean absolute bioavailability of G20 solution,G20-phospholipid complex were 3.02%,4.84%by comparison of G20 intravenous injection,respectively.4.A microemulsion technique was developed for enhancing the oral bioavailability of G20.The good mulsification region was identified by constructing pseudo-ternary phase diagrams and the average droplet size of selected microemulsions was characterized by a zetasizer.Finally,two microemulsion formulations?F3?W/O microemulsion?,F6?O/W microemulsion??were selected as research subject.Compared with the G20 solution,G20 microemulsion showed a higher bioavailability.The relative bioavailabilities of G20 in rats after oral administration of O/W microemulsion and W/O microemulsion were enhanced by 5.78-fold and 4.38-fold,respectively.The mean absolute bioavailability of G20 solution,O/W microemulsion and W/O microemulsion were 3.02%,17.3%and 13.21%by comparison of G20intravenous injection,respectively.5.The synthesis of G20 resin using commercially available merrifield resin was achieved in three steps with an overall yield of 22.0%.G20 resin has a connecting arm of 13 atoms to facilitates the affinity of the cardiac protein,which lays the foundation for the discovery of the target protein of G20.In the thesis,Taking an simple 6-tert-butylpuerarin for research object,Two synthetic routes of isoflavone C-glycosides were developed:an improved chalcone pathway and a novel deoxybenzoin pathway.The total synthesis of G20 was achieved through these two synthetic routes.A microemulsion technique and a phospholipid complex technique can improve oral bioavailability of G20.The synthesis of G20resin was achieved for the discovery of the target protein of G20.