| Research BackgroundTaxus chinensis seeds are a kind of abundant medicinal resources,but there are few related research reports.According to the current research,the yew seeds contain a variety of chemical substances,including taxanes,terpenes,flavonoids and phenolic compounds,and its pharmacological activities also include anti-cancer,anti-inflammatory,anti-oxidation,liver protection and analgesic effects.Asthma is a chronic airway inflammatory disease with high incidence,which is mediated by various types of immune inflammatory cells and structural cells such as airway smooth muscle cells,and is usually induced by allergens.According to the‘Global Initiative for Asthma Prevention and Control’in 2020,there are more than 300 million people with asthma in the world,of which more than 40 million people in China suffer from asthma.Due to factors such as the decline in air quality and the increase in obesity,the number of patients is still increasing.At present,asthma has become one of the global public health risks that need to be solved urgently.Research purposesThis subject aims to extract and isolate the parts with anti-asthma activity from Taxus chinensis seeds,and prepare them into liposome preparations by means of pharmacy to increase their water solubility,and further investigate the parts and the therapeutic effect of the preparation on asthma and the lung targeting ability of the preparation provide a new idea for the medicinal development of Taxus chinensis seeds and the clinical treatment of asthma.Research methodsThe extraction process of Taxus chinensis seeds was investigated by single factor experiment;egg albumin（OVA）was used to induce mouse asthma model,and interleukin（IL）-4 and IL-5 and hematoxylin-eosin（HE）stained pathological tissue sections were used as indicators to screen and extract the anti-asthmatic activity of Taxus chinensis seeds;identification by thin layer chromatography（TLC）,liquid chromatography-mass spectrometry（LC-MS）analysis and High performance liquid chromatography（HPLC）analysis is used to characterize the chemical components in the parts and determine the in vitro HPLC analysis method of the parts.Preparation of liposomes with entrapped sites by solvent injection method;screening of the optimal formulation of liposomes for sites by single factor experiment and characterization in vitro;investigation of stability and safety of liposomes for sites;determination of sites and lipids Cumulative release of plastids in different dissolution media.The HPLC analysis method of the site in the mouse was established;the tissue distribution characteristics of the site administered by intragastric administration and the liposome in the mouse were studied.The mouse asthma model was induced by OVA,and the spleen index,IL-4,IL-5 and tumor necrosis factor-alpha（TNF）-αcytokine concentrations and HE stained pathological tissue sections were used as indicators to investigate the site and Anti-asthmatic activity of liposomes.Research resultsDichloromethane extraction（DET）,ethyl acetate extraction（EET）,n-butanol extraction（BET）and water solution extraction（WET）in Taxus chinensis seeds were extracted and separated,and the average extraction rate of DET was 6.78±0.26%.In the screening of site anti-asthma activity,the concentrations of IL-4 and IL-5 in the model group were significantly higher than those in the blank group,which indicated that the asthma model was successfully established;It is smaller than other sites（P<0.01）,and the results of histopathological studies show that DET is the best in improving the morphology of mouse lung tissue,so it is believed that DET has the best anti-asthma activity compared with other sites,and it will be used in future formulation studies.Through TLC identification,LC-MS analysis and HPLC analysis,it was proved that DET contains paclitaxel（PTX）,the content of PTX in det was 2.58±0.67%,and PTX was used as the representative substance of DET in HPLC analysis,and the HPLC analysis method of DET in-vivo and in-vitro was established,and the inspection method was specific.,the linear relationship is good,and the intra-day and inter-day precision,recovery rate,repeatability and stability meet the methodological requirements,and can be used for qualitative and quantitative analysis of DET in vivo and in vitro;The equilibrium solubility is 846.02±9.34μg/m L.Through the single factor experiment,the optimal prescription of dichloromethane liposome（DETL）is 10 mg of DET,20mg of egg yolk lecithin,and 10 mg of cholesterol;the average particle size of DETL measured by particle size analyzer is 133.29±0.63 nm,more the polydispersion index（PDI）was 0.097±0.023,theζpotential was -31.53±0.72 m V,the drug loading was 19.32±0.23%,and the encapsulation efficiency was 92.59±0.77%.DETL is stable for 7 days at room temperature.According to the results of the zebrafish toxicity experiment,DETL significantly increased（P<0.001）the half lethal concentration(LC50)of DET,which proved the safety of DETL.The in vitro dissolution test showed that the dissolution rate and dissolution rate of DETL were higher than that of DET.In the mouse tissue distribution experiment,DETL increased the concentration of DET in vivo,and maintained a high drug concentration in the mouse lung tissue for 2 hours.In the asthma drug efficacy experiment,the spleen index,IL-4,IL-5 and TNF-αconcentrations of the treatment group were lower than those of the model group（P<0.001 or P<0.01）,indicating that DET and DETL can effectively relieve asthmatic mice.The histopathological study of the inflammatory response also showed that DETL significantly improved the morphology of lung tissue in asthmatic mice.At the same dose,the therapeutic effect of DETL was better than that of DET（P<0.01 or P<0.05）,and the therapeutic effect of high dose was better,which indicated that DETL had better anti-asthma activity,and it was dose-dependent.ConclusionDET is the part with anti-asthma activity in Taxus chinensis seeds,and DETL greatly improves the in vitro dissolution rate,in vitro dissolution rate,lung targeting ability and anti-asthma activity of DET,providing a new method for the utilization of Taxus chinensis seeds resources new ideas. |
