Purpose <p>The aim of this study was to develop a water-soluble form of umifenovir (UMF) through the synergistic action of the non-ionic surfactant Brij35 and choline bitartrate (ChB), and to investigate the solubility, aggregation and diffusion.</p> Methods <p>UMF solubilization was assessed using the saturation shake-flask method. The hydrodynamic radii and aggregation behavior of Brij35 micelles were characterized by light scattering. UMF diffusion rate was investigated using a Franz diffusion cell and an artificial membrane.</p> Result <p>Introduction of ChB into the UMF/Brij35 system reduced the degree of association with Brij35 micelles and the micelle/water partition coefficient, increased the solubilizing capacity, and modulated the aggregation behavior of Brij35 and permeability. The increase in total (micelle-associated and freely dissolved) drug solubility of UMF in buffer рН 7.4 in the system with the minimal Brij35 concentration (0.45%) was 18.4%, whereas upon addition of ChB (1.0%) - 35.2%. However, calculation of the molecularly dissolved fractions (f_free) of the compound showed that the increase in solubility of the freely dissolved drug in the system with ChB was only 10.6%. This finding is of fundamental importance because only the molecularly dissolved form of a drug can cross biological membranes. Optimization of membrane permeability in the UMF/Brij35 (0.45%)/ChB (1.0%) system was achieved by increasing the fraction of molecularly dissolved UMF molecules in the presence of ChB.</p> Conclusion <p>This study highlights the advantage of simultaneously using low concentrations of Brij35 and ChB to achieve a synergistic action for maximal improvement in UMF solubility with a minimal reduction in permeability. </p> Graphical Abstract <p></p>