Formation of Multiphase Complex Coacervates and Partitioning of Biomolecules within them.

Biological systems employ liquid-liquid phase separation to localize macromolecules and processes. The properties of intracellular condensates that allow for multiple, distinct liquid compartments and the impact of their coexistence on phase composition and solute partitioning are not well understood. Here, we generate two and three coexisting macromolecule-rich liquid compartments by complex coacervation based on ion pairing in mixtures that contain two or three polyanions together with one, two, or three polycations. While in some systems polyelectrolyte order-of-addition was important to achieve coexisting liquid phases, for others it was not, suggesting that the observed multiphase droplet morphologies are energetically favorable. Polyelectrolytes were distributed across all coacervate phases, depending on the relative interactions between them, which in turn impacted partitioning of oligonucleotide and oligopeptide solutes. These results show the ease of generating multiphase coacervates and the ability to tune their partitioning properties via the polyelectrolyte sharing inherent to multiphase complex coacervate systems.

Design and formulation of nanoemulsions using 2-(poly(hexafluoropropylene oxide)) perfluoropropyl benzene in combination with linear perfluoro(polyethylene glycol dimethyl ether).

This is the first report where PFPAE aromatic conjugates and perfluoro(polyethylene glycol dimethyl ether) are combined and formulated as nanoemulsions with droplet size below 100 nm. A perfluoropolyalkylether (PFPAE) aromatic conjugate, 2-(poly(hexafluoropropylene oxide)) perfluoropropyl benzene, was used as fluorophilic-hydrophilic diblock (FLD) aimed at stabilizing perfluoro(polyethylene glycol dimethyl ether) nanoemulsions. Its effects on colloidal behaviors in triphasic (organic/fluorous/aqueous) nanoemulsions were studied. The addition of FLD construct to fluorous phase led to decrease in PFPAE nanoemulsion droplet size to as low as 85 nm. Prepared nanoemulsions showed high colloidal stability. Our results suggest that these materials represent viable novel approach to fluorous colloid systems design with potential for biomedical and synthetic applications.

Practical considerations for generation of multi-compartment complex coacervates.

We discuss preparation of experimental models for multi-compartment membraneless organelles in which distinct compositions are maintained indefinitely for macromolecule-rich phases in contact with each other. These model systems are based on the physical chemistry phenomenon of complex coacervation. In complex coacervation, liquid-liquid phase separation occurs due to ion pairing interactions between oppositely charged polyelectrolytes. This mechanism can drive the associative phase separation of proteins and nucleic acids, the major macromolecular components of membraneless organelles. Here we provide examples, advice and practical considerations for the design, generation, and analysis of multi-compartment complex coacervates. These structures are of interest to compartmentalize the interior of artificial cells and as models for the intracellular membraneless organelles of biological cells.