The etiology of the paramagnetic relaxation enhancement seen in malignant melanoma on proton magnetic resonance (MR) images has been the subject of many recent investigations and has been ascribed to iron from associated hemorrhage or chelated metal ions, rather than directly due to melanin. The purpose of this study was to correlate proton relaxation times on MR images in malignant melanomas with histopathologic features (i.e., degree of pigmentation, iron deposition, and necrosis), water content, and electron paramagnetic resonance (EPR) spectra to elucidate the etiology of the relaxation behavior demonstrated by these neoplasms. Cultured cells derived from human malignant melanoma metastases were implanted subcutaneously into nude mice. Twelve separate lesions were evaluated in 10 mice. Magnetic resonance imaging was performed in vivo at 1.9 T using spin echo and inversion recovery acquisitions for the purposes of calculating T1, T2, and proton density [N(H)]. Histopathologic examination was performed on specimens resected immediately after imaging, using hematoxylin/eosin, Prussian blue, and Fontana stains to assess tumor necrosis, and iron and melanin content. Dry/wet weight ratios and EPR spectra were also obtained on resected specimens. Our results indicate that T1 shortening correlates with increasing melanin content and not with increasing iron deposition, EPR-active metallic cations, necrosis, or water content. In fact, a presumably unrelated statistical correlation was found between increased iron and T1 prolongation. The T2 relaxation times did not correlate with the presence of any single factor other than proton density. Although the unique relaxation behavior of nonhemorrhagic malignant melanoma in vivo cannot be traced to a single cause, our data suggest that, contrary to previous investigations, it is strongly influenced by the presence of melanin rather than iron or other naturally occurring paramagnetic ions.