In a continuing analysis of substituent effects in intercalator-DNA interactions, an unsubstituted naphthalene monoimide, I [N-[3-(dimethylamino)propyl]-1,8-naphthalene-dicarboximide], with a 3-(dimethylamino)propyl group on the imide N was prepared along with 3- and 4-nitro- (2 [N-[3-(dimethylamino)propyl]-3-nitro-1,8-naphthalenedicarboximide] and 3 [N-[3-(dimethylamino)propyl]-3-amino-1,8-naphthalenedicarboximide]) and 3- and 4-amino- (4 [N-[3-(dimethylamino)propyl]-3-amino-1,8-naphthalenedicarboximide] and 5 [N-[3-(dimethylamino)propyl]-4-amino-1,8-naphthalenedicarboximide]) substituted derivatives. These derivatives allow an evaluation of the importance of the Hammett substituent constant and of the substituent position on the binding of naphthalene monoimides to DNA. Viscosity and spectrophotometric analyses indicate that all 5 compounds bind to DNA by intercalation. The 4-nitro compound gives a smaller viscosity increase and binds only approximately 1/3 as strongly as the 3-nitro derivative. This difference is apparently due to the significant angle that the 4-nitro group makes with the intercalated monoimide ring system. The 3-NO2 group can assume a coplanar configuration with the monoimide ring system, allowing more favorable interactions with DNA base pairs, larger viscosity increases and stronger binding to DNA. The binding constants of the 3-substituted monoimides are in the order 2 > 4 > 1 and do not follow a substituent constant pattern. The Tm values from thermal melting of DNA are in the order 2 > 1 > 4, suggesting that the enthalpy contributions are significantly different for the binding of the 3 compounds to DNA. Van''t Hoff plots support this finding and indicate that both enthalpy and entropy contribute significantly to the binding free energy of 1 and 2 while the binding of 4 is primarily an enthalpic process. Plots of Tm and 65.degree. C log K values as a function of substituent constant for 1, 2 and 4 are linear. CPK [Cory-Pauling-Koltun] model building studies suggest that 4 can form a H bond with the 5'' diester O2 of the sugar-phosphate backbone of DNA in an intercalation complex. This would lead to more favorable energetics of binding, but a loss of mobility and/or available binding configurations with a resulting enthalpy-entropy compensation in the binding free energy of 4. This series of compounds dramatically illustrates the steric and H bonding complexity that can arise in attempts to design drugs to favorably interact with a DNA intercalation site as a potential bioreceptor. [3-Nitronaphthalene monoimides with cationic substituents on the imide nitrogen have significant antitumor activity and the mode of action of these compounds apparently involves interaction with DNA.].