Hetero-association of caffeine and aromatic drugs and their competitive binding with a DNA oligomer

NMR spectroscopy has been used to elucidate the molecular basis of the acti on of caffeine (CAF) on the complexation with DNA of mutagens such as ethid ium. bromide, propidium iodide, proflavine and acridine orange, and antican cer drugs such as actinomycin D and daunomycin. The hetero-association of C AF and each of the aromatic ligands in 0.1 mol L-1 phosphate buffer (pD=7.1 ) has been investigated as a function of concentration and temperature by 5 00 MHz H-1 NMR spectroscopy and analysed in terms of a statistical-thermody namic model, in which molecules form indefinite aggregates for both self-as sociation and hetero-association. The analysis leads to determination of th e equilibrium constants of hetero-association and to the values of the limi ting chemical shifts of the heteroassociation of CAF with each of the aroma tic molecules. The hetero-association constants between CAF and each of the aromatic drugs/dyes are found to be intermediate in magnitude between thos e for self-association of CAF and the corresponding drug/dye. The most prob able structures of the 1:1 CAF + ligand hetero-association complexes have b een, determined from the calculated values of the induced limiting chemical shifts of the drug protons. Knowledge of the equilibrium constants for sel f-association of CAF and the aromatic ligands, for their hetero-association and their complexation with a DNA fragment, the deoxytetranucleotide 5'-d( TpGpCpA), enabled the relative content of each of the CAF-ligand and CAF-li gand-d(TGCA) complexes to be calculated as a function of CAF concentration in mixed solutions. It is concluded that, on addition of CAF to the solutio n, the decrease in binding of drug or mutagen with DNA is due both to compe tition for the binding sites by CAF and the aromatic molecules, and to form ation of CAF-ligand hetero-association complexes in the mixed solution; the relative importance of each process depends on the drug or mutagen being c onsidered.