Production of Active Pharmaceutical Ingredients Encapsulating Ferritin Using Rapid Reassembly Technology via a Flow Microreactor

The encapsulation of active pharmaceutical ingredients (APIs) within ferritin represents a promising approach for drug delivery systems (DDS) due to ferritin’s unique self-assembling hollow structure. However, encapsulating nucleic acids such as DNA and siRNA remains challenging due to molecular size, charge balance, and the risks of protein aggregation and misassembly during the reassembly process. To address these limitations, this study developed and evaluated a novel ferritin disassembly and reassembly process using a sequential-mode flow microreactor (FMR), which offers precise control over mixing and reaction conditions. The FMR system demonstrated superior performance over traditional batch methods by ensuring uniform protein concentrations and reducing misassembly levels across varying scales. By optimizing flow rates and solution conditions, the misassembly ratio was significantly reduced to 3–4%, even at higher flow rates, while maintaining high yields. Moreover, the FMR system successfully encapsulated model DNA, within ferritin, achieving encapsulation efficiencies unattainable with batch processing. These findings establish the sequential-mode FMR as a robust and scalable platform for the production of ferritin-based therapeutics, paving the way for innovative applications in nucleic acid delivery and large-molecule biopharmaceuticals. The study underscores the potential of FMR technology to revolutionize protein reassembly techniques, offering transformative solutions for bio- and nanomedicine.