Saframycins Yd-1 and Y3, which have an amino functional group in the side chain, were recently obtained by a directed biosynthesis. Twenty-eight side chain-modified chemical derivatives were prepared from these saframycins, and their in vitro and in vivo antitumor activities were studied. Among these new derivatives, three saframycins, namely, two N-acyl derivatives, pivaloyl- and n-caproylsaframycin Y3, and one water-soluble type, saframycin Yd-1.HCl, were found to show marked antitumor activity against L1210 mouse leukemia cells. Further studies of these saframycin derivatives using B16-F10 melanoma and Lewis lung carcinoma indicated that all three saframycins are also active against B16-F10 melanoma; saframycin Yd-1.HCl showed the greatest prolongation of survival time. Marked inhibition of spontaneous metastasis of Lewis lung carcinoma was observed in mice treated with these new derivatives. Structure-activity relationships among these semisynthetic saframycins are discussed.