| ObjectiveCombined with network pharmacology and molecular docking technology,exploring the potential active ingredients and core targets of Pfaffia paniculata and Rubi Fructus,so as to provide basic and research direction for the later research.Through adenine modeling in rats,exploring the therapeutic effect of Pfaffia paniculata and Rubi Fructus on oligospermia preliminarily to verify the network pharmacology results.Through the multiple index determination and physical fingerprint evaluation of hot air drying,vacuum drying,spray drying and freeze drying,discussing the differences between drying methods,to provide valuable reference for the later study of solid preparations.Developing the Pfaffia paniculata and Rubi Fructus inervescent tablets based on the concept of QbD,geting the best taste feeling while reaching disintegration time limit and foaming amount,in order to achieve the quality goal of effervescent tablets product design.Methods1.The active ingredients of Pfaffia paniculata and Rubi Fructus were obtained by using the methods of TCMSP,TCMID and literature search.Determing the compound structure in Pub Chem database and establishing the composition database in China Natural Products Chemical Composition Bank.The Swiss ADME database was filtered for both Lipinski and Veber rules.Predicting and screening targets of the active components by Swiss Target Prediction.The disease targets associated with oligoasthenospermia were collected in Gene Cards,Pharm Gkb,OMIM and Uni Prot databases.Through the construction of medicinal materials-component-target network to predict the key targets and active components of drugs in the treatment of oligoasthenospermia.GOBP,GOCC and GOMF were analyzed for the core targets,and the enrichment results were used to speculate the mechanism of Pfaffia paniculata and Rubi Fructus against fewer asthenospermatosis.2.Fifty-six male rats were randomly divided into seven groups.They were normal group,model group,positive control group,PL group,PH group,PRL group and PRH group,with 8 mice per group.Except for the normal group,250mg kg-1adenine was given daily by continuous gavage for 4 weeks.Since the fifth week,the normal group and the model group were given equal amounts of pure water,the positive control group gave levocarnitine 100mg kg-1,PL group gave extract 1000 mg kg-1,PH group gave extract 1500mg kg-1,PRL group gave extract 450mg kg-1,and PRH group gave extract 860mg kg-1,once a day.Materials were taken after 4 weeks of continuous drug administration.The sperm number,sperm motility,SOD motility and MDA content of the subject rats were examined,and the tissue morphology of rat testis and epididymal sperm were stained.3.Pfaffia paniculata and Rubi Fructus powder samples were prepared by vacuum drying,spray drying,hot air drying and freeze drying.Characterizing the uniformity by median particle size,particle size distribution width and particle size distribution range;characterizing accumulation by loose packing density and vibration density;characterizing compressibility by intergranular porosity and Carr index;characterizing liquidity through the resting angle and Haunesby;characterizing stability by moisture content and hygroscopic rate.The physical fingerprint of the powder was constructed according to the above secondary indexes.The differences between the drying methods and the main factors were compared by similarity analysis,cluster analysis and principal component analysis.4.Determining the product quality objectives,key quality attributes and key material attributes.With the time limit of disintegration and foaming amount,the acid-base ratio,filler,sweeteners,lubricant and main addition amount of effervescent tablets were investigated separately.The Box-Behnken response surface test was optimized according to the results of the univariate investigation.The best prescription was obtained by second-order regression model,and verified by sensory evaluation and effervescent tablets quality.Results1.A total of 32 drug-like components were obtained,and the core targets were mainly included CDK1、CCNA2、CCNB1、CCNB2、CDC20、CDK2、CCNA1、CDC25C、BUB1、CCNE1.The GO BP,GO CC,and GO MF were mainly enriched in Cell cycle,p53 signaling pathway,Cellular senescence,AMPK signaling pathway and Fox O signaling pathway,which involving biological processes such as cell cycle regulation and steroid hormone metabolism.2.After four weeks of administration in the model of oligoasthenospermiathe rats,the testicular tissue of rats in the positive control group,PL group,PH group,PRL group and PRH group produced some degree of repair effect.Their testicular seminiferous tubules are relatively full and their epithelial structure is still intact,but the spermatogenic cells in all levels are slightly disordered,with more sperm cells visible in the lumen of the model group.The Pfaffia paniculata and Rubi Fructus extractive could improve the sperm concentration and vitality of rats,enhance the SOD vitality,reduce the MDA content in testis and reduce the damage of adenine to testis,which verified the results of network pharmacology by efficacy experiments.3.The correlation coefficients of hot air drying,vacuum drying,spray drying and freeze drying powder are 0.516,0.965,0.697 and 0.717,which compared with the control atlas.The powder properties of vacuum drying and freeze drying are most similar,but the freeze drying powder takes higher water content and longer drying time.The water content of hot air drying powder is the least,but it is easy to appear gelatinization phenomenon.The spray drying powder has the best fluidity,while the accumulation and pressure are poor.4.The filler acid-base ratio was in the range of 1.3:1 to 1.5:1.The filler is microcrystalline cellulose,the sweetener is erythritol,and the lubricant is PEG6000.The response surface optimization results show that the influence of each factor on the sensory score of effervescent tablets is successively:main drug addition>acid to base ratio>disintegration agent addition.Through the established second-order regression model,the best prescription of effervescent tablets is 20%of the main drug,50%of the disintegrating agent and the acid to base ratio of 1.4:1.The proved model is more reliable.ConclusionPfaffia paniculata and Rubi Fructus may improve the symptoms of rats through the anti-inflammatory and the antioxidant active components,and the mechanism of action may be the protective effect on rat testicular spermoepithelial cells,so as to reduce the damage of adenine in vivo,reduce the occurrence of oxidative stress and thus improve sperm quality.The spray drying powder could be well applied in the preparation of effervescent tablets.The hygroscopic phenomenon is not obvious in the process of effervescent tablets.The development of effervescent tablets could provide some reference for the development of Pfaffia paniculata related products. |
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