An improved method for the preparation of the heme octapeptide acetyl-MP8, obtained by proteolysis of horse heart cytochrome c, is described. AcMP8 obeys Beer’s law at pH 7.0 in aqueous solution up to a concentration of 3 x 10(-5) M. The self-association constant measured at 25 degrees C (log K-D = 4.04) is an order of magnitude lower than that for MP8, reflecting the role of the N-acetyl protecting group in abolishing intermolecular coordination. However, AcMP8 does form pi-stacked dimers in aqueous solution with increasing ionic strength. A more weakly packed pi-pi dimer reaches a maximum abundance at similar to 3 M ionic strength, but a more tightly packed dimer is favored at mu > 3 M.