Research On The Fabrication Of Liposomes By Microfluidics

Liposomes are artificial carriers with biological membrane structure.The phospholipid molecules are amphiphilic,with the hydrophilic end outward and the hydrophobic end inward to form spherical vesicles with bilayer membranes when dissolve in water.Liposomes can be used for drug carriers to achieve sustained release,targeting or to improve drug stability.There have been some successful cases of liposomes used for chemical drug carriers,in recent years,with the development of biotechnological drugs,liposomes have also begun to be used as protein drug carriers.However,conventional liposome preparation methods have certain defects in the loading of proteins,such as low encapsulation efficiency,poor controllability and reproducibility,which brings many difficulties to the practical application of liposomes.The introduction of microfluidic chips provides a new approach for the preparation of liposomes.Many reports have shown that micron-sized liposomes been prepared by microfluidic droplet technology.In the present study,we designed and optimized the structure of microfluidic chip in order to prepare liposomes with good controllability and repeatability and also increase the encapsulation efficiency of protein drugs.First of all,we designed step emulsification and flow focusing microfluidic chip structure.Through the preparation of water-in-oil emulsions,we found that step emulsification microfluidic chip had higher resistance when the viscosity of the internal aqueous phase was high,considering its two layer lithography design had a very high requirement on the equipment,which increased the difficulty of operation.Therefore,we chose flow focusing microfluidic chip.Through structural optimization,we added the buffer and filter structure to reduce pressure fluctuations and balance the flow resistance,enhancing the stability of water-in-oil droplets.Owing to the hydrophobic character of PDMS material,we need to do the local hydrophilic treatment during the preparation of double emulsion.We use trichlorosilane to hydrophobize part of the chip and use SDS+PVA to do hydrophilic treatment for the rest of chip.We successfully prepared double emulsions,however,PVA treatment formed an uneven surface,so the prepared double emulsions were polydisperse and had poor reproducibility.In order to improve the above situation,we next adopted a step-by-step method.We connected two chips in series,the first chip was not treated to maintain its hydrophobicity property,while the second chip was treated by oxygen plasma method to achieve the overall hydrophilicity property.The W/O emulsion prepared by the first chip further flowed into the second chip to form W/O/W double emulsion.This method avoids the use of PVA,which could maintain a flat surface of PDMS and also improve the stability of the emulsion.Through the optimization of chip connection,prescription,and oleic acid removal method,we prepared double emulsions of uniform size and good morphology.The formation of suspected liposomes was also observed.At the same time,we also studied the glass capillary microfluidic device to improve the monodisperse property of double emulsion,and investigated the influence of tapered end size and flow rate on the particle size of W/O emulsions.Through investigating and optimizing of the double emulsion chip establishing method,we finally prepared monodisperse double emulsions.The size and morphology of suspected liposomes were also observed.In summary,we applied microfluidic technology to the preparation of liposomes based on double emulsion method.Through investigation and optimization of PDMS and capillary microfluidic chip,we have improved the homogeneity and stability of double emulsions and laid the foundation for subsequent protein-loaded liposome preparation.