(3dm) High-Throughput Synthesis of Polymeric Nanoparticles Using 3D Flow Focusing in Parallel Microchannels

Use of microfluidics for synthesis of nanoparticles has been of great interest due to controllability and reproducibility in their physicochemical properties. Recently, we reported microfluidic synthesis of poly(lactide-co-glycolide)-b-polyethyleneglycol (PLGA-PEG) nanoparticles for drug delivery using rapid nanoprecipitation by mixing polymer solution in acetonitrile with water in a 3D hydrodynamic flow focusing (3D HFF) geometry that isolates the precipitating polymers from channel walls, eliminating microchannel fouling. However, applications of the microfluidic synthesis were limited to in vitro studies due to low throughput. Here, we report multilayer microfluidic devices with 8 and 32 parallel channels for synthesis of nanoparticles using 3D HFF that enable sufficient throughput for in-vivo studies. In addition, sub-30 nm PLGA-PEG nanoparticles with various size distributions could be synthesized reproducibly using 3D HFF.

Master molds for the bottom and upper layers were patterned in SU-8 photoresist via conventional photolithography. Post arrays for interconnecting holes were patterned in SU-8 on top of the bottom layer with precise alignment using an UV aligner. A thin PDMS membrane was spin-coated on the master mold; spin speed was tuned to precisely control the membrane thickness to be slightly lower than the interconnect post height. The upper PDMS microchannel was prepared using conventional replica molding of PDMS in SU-8. The upper PDMS microchannel was aligned under a microscope and bonded on top of bottom PDMS membrane and peeled off. The multilayer PDMS microchannel was bonded to glass by oxygen plasma treatment to obtain the final device.

Because of the multilayer architecture, only 4 interconnecting tubes were required per microfluidic device. The PLGA-PEG in acetonitrile solution and pure acetonitrile could flow from the upper layer to the bottom layer through the interconnecting holes. The PLGA-PEG in acetonitrile solution was focused vertically by two distinct vertical acetonitrile sheath flows using the three sequential holes as confirmed in COMSOL simulations and confocal microscope images. It is noteworthy that stable vertical focusing could be achieved even at the high fraction of polymer flow in organic flow due to the optimal shapes of interconnecting holes.

At the junction between vertically focused PLGA-PEG in acetonitrile stream and lateral water sheath stream, the PLGA-PEG block copolymers self-assemble into the PLGA-PEG nanoparticles by nanoprecipitation. By using the multilayer PDMS microfluidic device, PLGA-PEG nanoparticles with various sizes could be synthesized with polymers of different molecular weights and concentrations. The nanoparticles synthesized from the microfluidic 3D HFF were smaller compared to the nanoparticles synthesized from the bulk synthesis method because the mixing time is shorter than the characteristic aggregation time scale in the microfluidic rapid mixing. Since the mixing time for hydrodynamic flow focusing is function of flow ratio of polymer stream to the total flow rate of water, the size of nanoparticles could be controlled in 3D HFF simply by changing the flow ratio of polymer stream to the total flow rate of water. Sub-30 nm PLGA-PEG nanoparticles with various size distributions could be synthesized when small molecular weight PLGA-PEG was used as polymeric precursor in the 3D HFF. Since the pharmacokinetics of nanoparticles highly depends on the physicochemical properties of nanoparticles, the reproducible and robust synthesis of PLGA-PEG nanoparticles by 3D HFF will be beneficial for in-vivo screening to find the optimal biodistribution and therapeutic efficacy.