Salvianolic Acids-Phospholipids Complex And Its Self-emulsifying Drug Delivery Systems

Water-soluble components are the main active ingredients from traditional Chinese medicine.However,many of them are found to be poor membrane permeability through the gastrointestinal wall after oral administration,which leads to low bioavailability and efficacy.Many natural active compounds can form complexes with phospholipid at a certain ratio with significant changes of the physical and chemical properties,as well as the biological properties.There are many unknown areas in the present study about photosomes,such as the influence of preparation conditions,the structure of phospholipid complex and the mechanisms of bioactivity.Salvianolic Acids(SAs)is the effective water-soluble part from Radix Salvia miltiorrhiza and salvianolic acid B(SalB)is one of main element and the strongest pharmacological activity salvianolic acids.It was reported that the bioavailability of SalB is extremely low after oral administration of SAs.SAs was selected as a typical model of water soluble compounds and was been made into phytosome to enhance it biomembrane permeability and improve its oral bioavailability.First of all,the method of determination of salvianolic acid components in dog plasma by HPLC was established.It was proved the low bioavailability of SalB(1.07±0.43%)after oral administration of SAs in beagle dogs.The pharmacological effects of SAs after single oral administration were preliminarily evaluated by parameters of hemorheology.The low bioavailability was verified on the point of pharmacodynamics.In order to elucidate why the oral bioavailability of salvianolic acids is low,the mechanism of SAs oral absorption were deeply researched.The vitro results showed that the SAs was stable in the acidic environment and the most optimal stable pH value range lies at 2.5～5.0 and SAs was degradated rapidly in the alkaline environment.After oral administration,SAs was showen relatively stablity in the stomach and easy degradation in small intestine and colon.In rat gastric in situ perfusion model,SalB,RA,LA from SAs showed a slow absorption mainly through passive diffusion in the stomach The mechanism of intestinal absorption of SAs was studied using everted intestinal sac and Caco-2 cell model.The results showed that SalB possessed poor permeation through the intestinal membrane with influence by pH and there is no efflux pump-out by P-glycoprotein.The other components in SAs maybe inhibited the intestinal absorption of SalB to some extent.In the isolated rat liver perfusion model,it was showen that the liver can uptake a majority of SalB, which result in a serious liver first-pass effect.In sum,the study basical illustrated that the low oral bioavailability of SalB was due to the poor stability and membrane permeability in digestive tract and severe hepatic first-passeffect as well.In order to improve the stability and permeability of SAs,a detailed study of SAs phospholipid complex preparation was carried out on focus of the type of reaction solvent,drug lipid ratio,phospholipid type,reaction temperature,reactant concentration in the formation of phospholipid complex.It was the first time that the type of solvent was a decisive factor in the process.The ratio of drugs and lipid was another important factor with 1:1～3 as the successful range.The preparation method of phospholipid complex was established:to use methanol-tetrahydrofuran(2/18) as a reaction solvent,stirring at 40℃till the solution becomes clear,then to remove solvent by heating and vacuum,to get the solid powder that is SPC.The preparation conditions were optimized according to combination rate and the stability of the phospholipid complex.It was proven to be of high combination rate close to 100% and good stability.According to diversity in solubility,the method of identification of the phospholipid complex was established the first time.Chloroform and N-hexane were chosen to remove free SAs and phospholipids.The content of SalB in the left pure phospholipid complex was determined to get the real drug-phospholipid ratio.Variety of methods and indicators were used to identify the phospholipid complex. The results of UV,IR and H~1NMR spectrum show that the UV absorption spectrum behavior of SAs,based on the chromophores,was not changed in the phospholipid complex.It was shown in IR and NMR results that there were no covalent combination between SAs and phospholipid although some polar interaction was found.But the complex was not simplily a physical mixture.The main connection between them was strong hydrogen bond.The complex was shown to be amorphous, completely different from simple mixture,measured by DSC and X-ray diffraction analysis.Compared with the original drug,SaLB represented a significant higher solubility in chloroform and n-octanol after made into phospholipid complex.Its distribution coefficient of n-octanol/water and chloroform / water increased significantly indicating that phospholipid complex significantly improved lipotropism of SAs.The increased stability in aqueous environment as the result..The stability of the impact of factors investigation showed that the SPC was very sensitive to humidity and should be kept in dry place.To improve the absorption properties of phospholipid complex further more, self-emulsifying agent was prepared.The prescription was optimized using Pseudo-ternary phase diagram with the best one was(Imwitor 742-Miglycol 812(1:1): Labrasol:Tween80:SPC=55:25:20:12).The SPC-SEDDS can not only enhance permeability but also increased drug stability to some extent.The model of everted intestinal sac and Caco-2 cell were used to study the mechanism of enhanced intestinal adsorption of SPC,SPC-SEDDS on SalB.It was found that single SPC could not enhance the membrane permeability of SalB.But its intestinal adsorption and transmembrane ability were significantly increased after made into SPC-SEDDS. So it was confirmed that the increase in membrane permeability was the co-effect of SPC and SEDDS.Four methylation metabolites of SalB were separated and purified at first to comprehensively evaluate the vivo process of SalB.The pharmacokinetics in animal and tissue distribution of SPC and its SEDDS were researched.The result showed that SPC-SEDDS could increase the content of SalB and its metabolites in various tissues, especially in liver,which can significantly enhance the efficiency of SAs in treatment of liver fibrosis and other diseases.The pharmacodynamics results showed that the, SPC-SEDDS can significantly rehabilitate the CC14-induced acute liver injury. SPC-SEDDS can significantly enhance the protective effects of SAs on the liver.The research results show that,the stability and solubility of water-soluble salvianolic acids will be significantly improved after made into phospholipid complex. When made into SEDDS,its permeability and adsorption can be greatly enhanced, thus to improve its oral bioavailability and pharmacological effects.At the same time, the the comprehensive quality evaluation system for phospholipid complex formulations was established.This research provided ideas and technical methods for development of the third category of pharmacy drug,which is of water-solublity and poor membrane permeability.