Abstract: Interaction of bromophenol blue (BPB) with serum albumins from different mammalian species, namely, human (HSA), bovine (BSA), goat (GSA), sheep (SSA), rabbit (RbSA), porcine (PSA) and dog (DSA) was studied using absorption and absorption difference spectroscopy. BPB-albumin complexes showed significant differences in the spectral characteristics, i.e., extent of bathochromic shift and hypochromism relative to the spectral features of free BPB. Absorption difference spectra of these complexes also showed variations in the position of maxima and absorption difference (ΔAbs.) values. Absorption difference spectra of different bilirubin (BR)-albumin complexes showed a significant blue shift accompanied by decrease in ΔAbs. values in presence of BPB which were indicative of the displacement of bound BR from its binding site in BR-albumin complexes. These changes in the difference spectral characteristics of BR-albumin complexes were more marked at higher BPB concentration. However, the extent of these changes was different for different BR-albumin complexes. Taken together, all these results suggest that BPB partially shares BR binding site on albumin and different mammalian albumins show differences in the microenvironment of the BR/BPB binding site.