Microfluidic Assembly of Poly(glutamic acid) Nanogels Through SPAAC Click Chemistry

Background/Objectives: Nanogels (NGs) are promising carriers for drug delivery due to their tunable size, biocompatibility, and capability to encapsulate sensitive molecules. However, conventional batch synthesis often lacks control over key parameters, such as size distribution and encapsulation efficiency. This study aimed to develop a microfluidic platform for the reproducible synthesis of poly(α-glutamic acid) (PGA)-based NGs using strain-promoted azide-alkyne cycloaddition (SPAAC) click chemistry and to investigate the effects of flow parameters on the physicochemical properties of nanogels. Methods: Functionalized PGAs (with azide and DBCO) were co-injected into a microfluidic system within a flux of acetone to form NGs via SPAAC. Flow rate ratios (FRR) and total flow rates were systematically screened at 25 °C, with tests at 50 °C. We evaluated the particle size, polydispersity index (PDI), zeta potential, and encapsulation efficiency (EE%) of doxorubicin-loaded NGs. Results: NGs with tunable sizes ranging from ~50 nm to >170 nm and low PDI (<0.1 in optimal conditions) were obtained. Higher FRR and total flow rates yielded smaller and more uniform NGs. Doxorubicin loading did not affect the nanogel size and uniformity, and in some cases, it improved them. The EE% reached up to ~65%, and ~40% for the best formulations. Elevated temperature improved the characteristics of drug-loaded nanogels at intermediate solvent ratios. Compared to batch synthesis, the microfluidic process offers enhanced reproducibility and size control. Conclusions: Microfluidic SPAAC synthesis enables precise and scalable fabrication of PGA NGs with controllable size and drug loading. This platform supports future integration of on-chip purification and monitoring for clinical nanomedicine applications.