Harnessing the hydrolytic stability of poly(2-oxazine) living chain-ends for direct aqueous bioconjugations

Abstract

Poly(2-oxazine)s (PAOzi) have recently emerged as attractive alternatives to poly(2-oxazoline)s (PAOx) for biomedical applications. In this work, we exploit the slower hydrolysis of the oxazinium chain-ends in PAOzi, synthesized by cationic ring-opening polymerization, enabling direct, one-step nucleophilic conjugation with amines in aqueous environments. On the other hand, analogous PAOx showed exclusively chain-end hydrolysis. We show that the PAOzi hydrolysis rate strongly depends on pH, guiding us to optimal conjugation conditions. By employing buffer/DMSO mixtures and precisely tuning pH, we achieved good conjugation efficiencies with model primary amine and bovine serum albumin (BSA). Efficient bioconjugation under these mild, biocompatible conditions was confirmed by MALDI-TOF mass spectrometry, SEC, and diffusion-ordered (DOSY) NMR. Such aqueous conjugation is unique for living polymers arising from ionic polymerizations, which are notoriously known for their moisture sensitivity. This streamlined, scalable approach offers a versatile platform for next-generation polymer-protein conjugates and advanced drug delivery systems.