Screening Of Nano Supramolecular Carrier In Ligustrum Lucidum And Application In ?-carotene Encapsulation

The triterpenoids in Ligustrum lucidum have a variety of active effects such as anti-inflammatory and hepatoprotective,and have the potential of self-assembly to form nano supramolecules based on amphiphilic properties.However,it is difficult to screen triterpenoids from complex plant extracts.Studies have shown that superparamagnetic iron oxide(SPIO)nanoparticles were easy to be used in the extraction and separation of compounds due to their magnetic response.Inspired by this technology,SPIO was embed into triterpenoids by the method of ultrasonic emulsification.With the help of magnetic adsorption method,the self-assembled nanocarrier material oleanolic acid(OA)was identified and the preparation method of oleanolic acid nanoparticles(OA NPs)was optimized.On this basis,the hepatoprotective effect of OA NPs and the inclusion properties of?-carotene(CAR)were investigated.The main results are as follows:(1)Screening of nano supramolecular carrier materials in Ligustrum lucidum:Ligustrum lucidum powder as raw material is extracted by refluxing with 95%ethanol,concentrated,and then extracted from dichloromethane.After the extraction solution is obtained,the nanoparticles carrying SPIO are prepared by the method of ultrasonic emulsification,and the nanoparticles are screened by a magnetic separation method.Then,the nanoparticles obtained were dissolved and purified,and further analyzed by thin layer chromatography,nuclear magnetic resonance hydrogen spectroscopy,and carbon spectroscopy.The results determined that the nano supramolecular material in the extract was mainly oleanolic acid.(2)Preparation and characterization of OA-NPs:The effects of organic solvent,OA concentration,polyvinyl alcohol(PVA)concentration and phacoemulsification time on the particle size,polymer dispersion index(PDI)and yield of OA-NPs were investigated by single factor experiment.The results showed that the four factors had significant influence on the preparation of OA-NPs,among which the particle size of OA-NPs prepared by dichloromethane was the smallest;Furthermore,three factors and three levels of orthogonal test were used to further optimize the parameters.The optimal preparation process of OA-NPs was as follows:OA concentration 10.0 mg/m L,PVA concentration 2.0%,phacoemulsification time 120 s.Under these conditions,OA-NPs were spherical particles with smooth and complete appearance,average particle size of 260.3 nm,PDI of 0.19±0.02 and yield of 81.04±2.23%.(3)Hepatoprotective activity of OA-NPs:OA has low cytotoxicity and no significant effect on cell survival rate at 2?g/m L.Besides,OA and OA-NPs could significantly increase the survival rate of Hep G2cells induced by 160?L/m L alcohol.It may be related to the significant decrease of reactive oxygen species(ROS),the increase of mitochondrial membrane potential(MMP)and DNA content,but not the change of calcium concentration.Among these mechanisms of action,the effect of OA-NPs on DNA content and MMP was significantly higher than that of free oleanolic acid,but there was no significant difference between them on ROS level.(4)The nanoparticles,OA-CAR-NPs,were prepared by embedding CAR in OA-NPs,which of the CAR's embedding rate was 52.62±2.18%.Its stability and drug loading properties under different conditions were investigated.The results showed that the stability of CAR was improved by OA-CAR-NPs,and the retention rate of CAR in OA-CAR-NPs was significantly higher than that in free CAR under the conditions of heating,light,Ca²⁺solution and room temperature storage;However,The CAR contained in OA-NPs will still be degraded in a large amount during long-term storage at room temperature,but the OA-CAR-NPs's lyophilized powder has good resolvability and strong stability and is suitable for long-term storage.In vitro simulated release test,results show that the release rate of OA-CAR-NPs was 46.02±0.82%in the simulated gastric juice for 3 hours and 69.13±1.04%in the simulated intestinal juice for 4 hours.Intestinal fluid is the main release site.In conclusion,this project has successfully established a method for screening triterpenoid nano carrier materials from Ligustrum lucidum extract.The method of"ultrasonic emulsification magnetic separation"provides a demonstration for the screening of natural triterpenoids in the extraction of other plants,and the nanoparticles prepared by oleanolic acid provide a new scheme for the embedding of?-carotene,which provides a reference for the application of triterpenoid nano carrier materials in food field.