The clinically useful anticancer drug mitoxantrone intercalates preferentially into 5'-(A/T)CG and 5'-(A/T)CA sites on DNA. The 5,8 hydroxyl substituents on its anthracenedione chromophore are available to interact with the double helix. Footprinting experiments with two anthraquinone derivatives structurally related to mitoxantrone and ametantrone have been undertaken to assess the influence of the hydroxyl groups on the DNA recognition process. The results confirm that they do play a role in the recognition of preferred nucleotide sequences and suggest that the binding of anthraquinones to a 5'-(A/T)CG site is dependent on the presence of the 5,8 hydroxyl substitutes whereas binding to 5'-(A/T)CA sites appears to proceed just as well without them.