Structure-dependent incorporation of terpenes into amphiphilic Poly(2-oxazoline) micelles

Poly(2-oxazoline) (POx) amphiphilic block copolymers are highly effective carriers for poorly water-soluble small molecules, yet the structural features governing drug incorporation remain incompletely understood. Motivated by the unusually high loading of diterpene taxanes in POx micelles, we systematically examined whether shared terpene motifs support micellization with POx using natural compounds with established safety profiles. Representative mono-, di-, and triterpenes-α-pinene, carnosic acid (CA), carnosol (CARN), squalene, lupeol, betulin, and ursolic acid-were evaluated with a well-defined POx triblock copolymer. Only the diterpenes CA and CARN showed efficient solubilization, forming small, stable, and uniform micelles, whereas α-pinene produced polydisperse assemblies and all triterpenes were insoluble. CA exhibited the best performance, forming reproducible 22-29 nm micelles at a 2/10 (w/w) CA/polymer feed ratio with ~ 17% loading capacity and quantitative efficiency; higher feed ratios generated large, polydisperse aggregates by DLS and TEM. CA-loaded micelles were colloidally stable for at least 48 h at room temperature, although CA chemical stability varied across batches. Lyophilization markedly improved stability, enabling 67-89% recovery upon reconstitution. Experimental solubilization data were further used to benchmark existing computer-aided prediction models, which failed to capture the behavior of several terpene classes, underscoring the need to expand their chemical space and incorporate environmental effects. Overall, a rigid diterpene scaffold combined with ortho-hydroxyl functionality emerges as a key determinant of POx incorporation, identifying CA as a robust and reproducible formulation candidate for further POx-based delivery and pharmaceutical development.