Screening Of Potential Anti-inflammatory Components In Polygonum Cuspidatum And Study On Resveratrol Phospholipid Complex Loaded Self-microemulsion Drug Delivery System

The traditional Chinese medicine Polygonum cuspidatum Sieb.et Zucc.,belongs to the polygonaceae,which is the dried root and rhizome of Polygonum cuspidatum.It is widely used clinically because it has a variety of pharmacological activities,such as anti-tumor,anti-bacterial,anti-inflammatory and anti-viral.At present,a large number of studies have shown that Polygonum cuspidatum extract(PCE)has significant anti-inflammatory effects.However,there are few reports on the method of screening potential anti-inflammatory active ingredients from the complex multi-component system of traditional Chinese medicine.In addition,as the main anti-inflammatory active ingredient in PCE,resveratrol(RES)has poor water solubility(0.03 mg/m L),chemical instability,and short half-life in vivo,resulting in low oral bioavailability,which limits its clinical application.Therefore,it is particularly important to choose a suitable dosage form to improve the solubility and stability of RES and prolong its circulation time in vivo,so as to further improve the bioavailability of RES.In this study,macrophages were used as the biological extraction medium,combined with the HPLC-ESI-MS/MS to screen the potential anti-inflammatory active components in PCE.Then,combining the phospholipid complex(PC)technique and self-microemulsion drug delivery system(SMEDDS)technique to prepare resveratrol-phospholipid complex loaded self-microemulsion drug delivery system(RPC-SMEDDS).It is expected that the bioavailability of RES will be further improved on the basis of PC.The main study results of this article are as follows:(1)Using macrophages to bio-specifically extract the potential anti-inflammatory active components in PCE.The optimized conditions of extraction are as follows: the drug concentration was 1.00 mg/m L,the co-incubation time was 45 min,an extraction solvent was 75% ethanol,and an extraction time was 60 min.After HPLC-ESI-MS/MS analysis,it was found that there were 5 components that can specifically bind to macrophages,they were polydatin,RES,emodin-1-O-β-D-glucoside,and emodin-8-O-β-D-glucoside and emodin,respectively.It is further confirmed by literature that these 5 ingredients are the anti-inflammatory active ingredients of Polygonum cuspidatum.This indicated the feasibility and reliability of using this method to screen potential anti-inflammatory active ingredients in PCE.(2)The solvent evaporation method was used to prepare resveratrol-phospholipid complex(RPC),and the preparation technics of RPC were optimized through single factor experiment and orthogonal experiment design.The best preparation conditions were determined as follows: the reaction solvent was ethyl acetate,the reaction temperature was 40 ℃,the drug concentration was 10.00 mg/m L,the molar ratio of drug to phospholipid(PL)was 1:1,and the reaction time was 1 h.The analysis results of UV,FT-IR,DSC,XRPD and SEM showed that RES and PL formed a complex.The results of solubility experiments revealed that the RPC could significantly enhance the solubility of RES in water and n-octanol.(3)On the basis of RPC,through solubility experiment,determination of blank microemulsion particle size and PDI,compatibility experiment and drawing of pseudo-ternary phase diagram,the composition and dosage range of RPC-SMEDDS prescription were initially screened out.Then the best prescription was optimized by the central composite design and the results are as follows: 25%(w/w)triacetin,56.25%(w/w)Tween 80 and 18.75%(w/w)anhydrous alcohol.The physical and chemical properties of RPC-SMEDDS were measured,and the results showed that the prepared prescription had small particle size,high dispersion and good stability.The results indicated that RPC-SMEDDS could enhance the solubility of RES and promote the rapid release of drugs in vitro dissolution study.(4)The Caco-2 cell uptake experiment and Ussing chamber technique were used to study the intestinal absorption and transport of drugs and their preparations.The results showed that both RPC and RPC-SMEDDS can promote the intestinal absorption of RES.In addition,Ussing chamber studies revealed that RES was the substrate of MRP2 and BCRP rather than P-gp,and RPC-SMEDDS could significantly prevent the efflux mediated by MRP2 and BCRP.(5)The results of in vivo pharmacokinetic experiments in rats proved that compared with RES suspension,the relative bioavailability of RPC and RPC-SMEDDS were 132% and 253% at the same dosage,respectively,which indicated that RPC-SMEDDS could further improve the bioavailability of RES on the basis of RPC.