Incorporating PDADMAC/PAA coacervates into biphasic all-aqueous systems for the preparation of multiphase coacervate emulsions

Liquid-liquid phase separation (LLPS) is the phenomenon in which a liquid mixture spontaneously segregates in two or more liquid phases. LLPS can occur in aqueous mixtures of polycations and polyanions, which electrostatically bind forming a coacervate that phase separates, yielding a coacervate-rich phase and a dilute phase. Coacervates are promising systems for the encapsulation of biomolecules, such as enzymes.  In this work, we systematically studied how the polycation/polyanion charge ratio affects the droplet size, zeta potential and emulsion stability in polydiallyldimethyl ammonium chloride (PDADMAC)/polyacrylic acid (PAA) coacervate droplets in water. Varying PAA molecular weights were evaluated. It was observed that the coacervates formed at charge-stoichiometry phase-separated around the 4^th day, but by slightly shifting to a close-to-stoichiometric charge ratio (ca. <0.45 or >0.55) produced coacervates that were stable for over a month. Zeta potential measurements confirmed that electrostatic repulsion occurs in these conditions. Furthermore, we investigated the addition of some selected PDADMAC/PAA coacervates to biphasic all-aqueous systems, such as a second coacervate or polymer/polymer water-in-water emulsions. This triggered the formation of complex triphasic systems with droplets exhibiting core-shell and asymmetric snowman-like structures. Optical and confocal microscopy were used to characterize these systems. We aim to further study and control the encapsulation of enzymes and other biomolecules selectively in a particular phase of the multiphasic systems.