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Polymersomes (PL), vesicles formed by self-assembly of amphiphilic block copolymers, have been described as promising nanosystems for drug delivery, especially of biomolecules. The film hydration method (FH) is widely used for PL preparation, however, it often requires long hydration times and commonly results in broad size distribution. In this work, we describe the challenges of the self-assembly of poly (ethylene glycol)-poly(lactic acid) (PEG-PLA) into PL by FH exploring different hydrophilic volume fraction ( f ) values of this copolymer, stirring times, temperatures and post-FH steps in an attempt to reduce broad size distribution of the nanostructures. We demonstrate that, alongside f value, the methods employed for hydration and post-film steps influence the PEG-PLA self-assembly into PL. With initial FH, we found high PDI values (&gt;0.4). However, post-hydration centrifugation significantly reduced PDI to 0.280. Moreover, extrusion at higher concentrations resulted in further improvement of the monodispersity of the samples and narrow size distribution. For PL prepared at concentration of 0.1% ( m / v ), extrusion resulted in the narrower size distributions corresponding to PDI values of 0.345, 0.144 and 0.081 for PEG 45 -PLA 69 , PEG 114 -PLA 153 and PEG 114 -PLA 180 , respectively. Additionally, we demonstrated that copolymers with smaller f resulted in larger PL and, therefore, higher encapsulation efficiency (EE%) for proteins, since larger vesicles enclose larger aqueous volumes.

Challenges for the Self-Assembly of Poly(Ethylene Glycol)–Poly(Lactic Acid) (PEG-PLA) into Polymersomes: Beyond the Theoretical Paradigms