A surface forces apparatus (SFA) was used to measure the intermolecular forces between a biodegradable demulsifier (ethyl cellulose, EC) and asphaltenes immobilized individually on molecularly smooth mica surfaces in an organic solvent. A steric repulsion on approach between the immobilized EC layers and asphaltenes was measured despite strong adhesion (F-ad/R approximate to -2 mN/m; W-ad = 0.42 mJ/m(2)) during retraction. The measured adhesion was attributed to the interpenetration and tangling of aliphatic branches of swollen asphaltenes and solvated chains of EC macromolecules. Competitive adsorption of EC on/in immobilized asphaltene layers was confirmed by combining SFA force measurements and atomic force microscopy (AFM) imaging. Following the injection of EC-in-toluene solution, an immediate (<5 min) increase in the confined layer thickness of the immobilized asphaltene layers was measured. Irreversibly adsorbed asphaltenes were displaced by EC macromolecules through binding with unoccupied surface sites on mica, followed by the spreading of EC across the mica substrate due to increased surface activity governed by the higher number of hydroxyl groups per EC molecule. AFM imaging confirmed that the increase in confined layer thickness resulted from the formation of larger asphaltene aggregates/clusters protruding from the mica substrate. Molecular level topographical images showed that the asphaltenes were not resolvated in the organic phase but self-associated as the EC macromolecules spread across the hydrophilic mica substrate. The results from this study provide not only fundamental insights into the basic interaction mechanisms of asphaltenes with EC macromolecules as a demulsifier in organic media but also directions toward enhancing demulsification of water-in-oil emulsions.