One- and two-dimensional F-19 NMR spectroscopy was used to investigate the conformational heterogeneity of two arylamine-modified DNA duplexes, d[CTTCTTG*ACCTC]. d[GAGGTCAAGAAG], in which G* is either N-(deoxyguanosin-8-yl)-4’-fluoro-4-aminobiphenyl (dG-C8-FABP) (I) or N-(deoxyguanosin-8-yl)-7-fluoro-2-aminofluorene (dG-C8-FAF (II). The F-19 NMR spectrum of I showed a single peak, while that of II revealed two prominent signals with a 55:45 ratio, in good agreement with previous H-1 NMR results (Cho et al. Biochemistry 1992, 31, 9587-9602; 1994, 33, 1373-1384). Slow interconversion between the two conformations of II was established by temperature-dependent two-dimensional F-19 NMR chemical exchange spectre. On the basis of magnetic anisotropy effects and isotopic solvent-induced shifts, the F-19 signals at -117.31 and -118.09 ppm in the F-19 NMR spectrum of II were assigned to a relatively undisturbed "B-type" conformer and a highly perturbed "stacked" conformer, respectively. Analysis of the temperature dependent (5-40 degrees C) line shapes by computer simulation yielded an interconversion barrier (Delta G(double dagger) of 14.0 kcal/mol with a chemical exchange time of 2 ms at 30 degrees C. This new F-19 approach should be very useful in investigating the sequence-dependent conformational heterogeneity of arylamine-modified DNA.