The interaction of aristololactam beta-D-glucoside (ADG) with different polymorphic structures of poly(dG-me5dC).poly(dG-me5dC), poly(dG-dC).poly(dG-dC), and poly(dI-dC). poly(dI-dC) has been studied by spectrophotometric, spectrofluorimetric, circular dichroism, UV melting profiles, and thermodynamic analysis. The binding of ADG to B-form duplexes is characterized by the typical bathochromic and hypochromic effects in the absorption spectrum, quenching of steady-state fluorescence intensity, a decrease in fluorescence quantum yield of ADG, an increase in fluorescence polarization anisotropy, an increase of thermal transition temperature, and perturbation in circular dichroic spectrum. Scatchard analysis indicates that ADG binds to the right-handed form of each polymer in a noncooperative manner. Comparative binding parameters determined from absorbance titration by Scatchard analysis, employing the excluded site model, indicate a stronger binding of ADG to the B-form of poly(dG-me5-dC). poly(dG-me5dC) than to the B-form of poly(dG-dC).poly(dG-dC) or poly(dI-dC). poly(dI-dC). Thermodynamic parameters (delta G degree, delta H degree, and delta S degree) obtained by van't Hoff analysis of the data show that the process of binding to all B-form duplexes is exothermic and enthalpy driven as characterized by a favorable negative enthalpy change (delta H degree). The binding is opposed by a negative entropy change (delta S degree) contribution. Conformational changes indicate that the alkaloid converts the left-handed form of poly(dG-dC). poly(dG-dC), and its methylated analogue and high salt form of poly(dI-dC). poly(dI-dC) to a bound right-handed form, while it inhibits both the rate and extent of the B to Z transition. These studies reveal that ADG binds strongly to B-form polymers while it does not bind to polymers of Z-form.