AbstractBenzoporphyrins, chlorin‐type compounds, are efficient new drugs in photodynamic therapy. In this work, some aspects of the benzoporphyrin‐1,3‐diene dimethyl ester (DiesterB) aggregation were investigated in water–organic solvent mixtures, using visible spectrophotometry. The effects of solvent mixtures, water with acetonitrile, dioxane, methanol and ethanol, were compared taking the calculated aggregation equilibrium constant (K) and the aggregation number. The K dependence on temperature, DiesterB concentration and water content suggest that, in water with acetonitrile, the predominant species is a dimer, whereas in dioxane, methanol and ethanol systems, there are multiple equilibria, with a pre‐equilibrium between the monomer and dimer (small amount), followed by trimer formation. Not only the solvent polarity but also the nature of the organic solvent are important for the aggregation process, driving the aggregate size and the magnitude of the equilibrium constant. Copyright © 2004 John Wiley & Sons, Ltd.