Preparation And In Vitro And In Vivo Evaluation Of Jujuboside B Nanoparticles

Ziziphus jujuba Mill.var.spinosa（Bunge）Hu ex H.F.Chow is the first batch of medicinal and edible plants issued by the Ministry of health.It is widely planted in North and Northwest China.Its leaves,flowers,fruits and roots can be used as medicine,and has high pharmacological effect and edible value.Jujube kernel,as an important medicinal part,has been proved to have the effects of sedation and hypnosis,anti anxiety,anti depression,calming the heart and mind,nourishing the liver and so on.Among them,the tetracyclic triterpenoid component Jujuboside B（JuB）plays an important role,but the bioavailability of JuB is low.Therefore,a JuB nanoparticle delivery system was designed and synthesized in this experiment In order to prepare an effective nano-system to improve the absorption and utilization of JuB in vivo.The research of this subject is mainly carried out from the following four aspects.（1）Optimization of preparation conditions of nanoparticles.On the basis of previous experiments,JuB-polylactic-co-glycolic acid copolymer（PEG-PLGA）nanoparticles（B-NPs）were prepared by nano precipitation method,and the entrapment efficiency of nanoparticles was determined by ultrafiltration centrifugation.Taking JuB as the model drug and PEG-PLGA as the carrier material,the model was established by single factor experiment and response surface experiment,and the optimum preparation conditions were obtained,that is,the concentration of Tween-80 was 0.75%,The volume ratio of aqueous phase to organic phase is 8,the proportion of acetone occupied the organic phase was 70%,the ultrasonic power was 320W,the ultrasonic time was 10 min,and the drug loading ratio was11.7%,Under the above optimized conditions,the particle size was 109.13±9.33 nm and the entrapment efficiency was 73.46±1.26%.The in vitro release characteristics of B-NPs were investigated,and it was found that it can play a significant sustained-release role.（2）Modification of nanoparticles with stearyl carnitine（LC-SA）.Next,the prepared B-NPs nanoparticles were modified with stearyl carnitine（LC-SA）.L-carnitine（LC）,as an important substance for energy conversion in human body,can target OCTN2 and ATB^（0,+）transporter of human intestinal cells.The ligation modification of nanoparticles with LC can promote the uptake of nanoparticles by cells.The experiment determined that the optimal modification ratio is 20%.At this time,the particle size of nanoparticles is 139.33±9.45 nm,and the encapsulation efficiency of nanoparticles reaches 76.01±2.10%,It has good dilution and placement stability.（3）Cellular uptake of nanoparticles.Caco-2 cells similar to human intestinal cells were used.The uptake of nanoparticles was simulated.Firstly,the cytotoxicity experiment of nanoparticles was investigated.The results showed that when the concentration of nanoparticles was large,the cytotoxicity to Caco-2 cells was still small,which proved its relative safety.Through the qualitative and quantitative analysis of cell uptake of nanoparticles,it was found that LC-SA modified nanoparticles could significantly improve cell uptake,and OCTN2 transporter played an important role.In addition,caveolin and clathrin were involved in the endocytosis of nanoparticles.Finally,the content changes of cell tight junction protein and efflux transporter by nanoparticles were studied.It was found that after nanoparticles cultured cells for a period of time,the contents of cell tight junction protein ZO-1 and Occludin did not change significantly,indicating that nanoparticles did not pass through the paracellular transport.（4）Pharmacokinetics of nanoparticles in rats.Plasma was established by HPLC-MS/MS.The content determination method of JuB in and the method was investigated.SD rats were selected as the experimental object,and the tail vein blood was taken for plasma sample processing.It was found that the specificity,precision,recovery and matrix effect of this method were good,all within the required range.The results showed that the Cmax of B-NPs group and LC-B-NPs group were both improved.In addition,the relative bioavailability of JuB was 134.33%and 159.04%respectively.After preparation and modification,the oral bioavailability increased significantly.