Probing the inner structure and dynamics of pH-sensitive block copolymer nanoparticles with nitroxide radicals using scattering and EPR techniques

This research emphasizes the crucial role of electron paramagnetic resonance (EPR) spectroscopy in nanoparticle analysis, showcasing its distinctive ability to probe molecular-level details. We demonstrate the synthesis and self-assembly of a new class of pH-responsive amphiphilic diblock copolymers, specifically poly[N-(2-hydroxypropyl)-methacrylamide]-block-poly[2-(diisopropylamino)ethyl methacrylate] (PHPMA-b-PDPA), incorporating 2,2,6,6-tetramethylpiperidine-1-oxyl (TEMPO) radicals covalently bound to the hydrophilic PHPMA segment. The copolymer synthesis involved a three-step process, combining reversible addition − fragmentation chain transfer (RAFT) polymerization with carbodiimide (DCC) chemistry. TEMPO radical-containing nanoparticles (RNPs) were created using microfluidic (MF) nanoprecipitation, a technique essential for generating uniform nanoparticles with predictable biodistribution and cellular uptake. Adjusting MF protocol parameters allowed fine-tuning of RNP sizes. EPR spectroscopy confirmed the formation of core–shell RNPs, with features aligning closely with those obtained from scattering techniques like dynamic light scattering (DLS), static light scattering (SLS), small-angle X-ray scattering (SAXS), and cryo-transmission electron microscopy (cryo-TEM). EPR spectroscopy emerges as a potent tool for molecular-level analysis of colloidal polymer systems. This study highlights the EPR-spin label method’s efficacy in probing the internal structural features and dynamics of core–shell nanoparticles, especially their response to pH-triggered disassembly. The findings open new avenues for the use of pH-responsive TEMPO-labeled diblock copolymers in controlled drug delivery applications.