As a part of our project directed at the search of new chemotherapeutic agents against American trypanosomiasis (Chagas' disease), several drugs possessing the 4-phenoxyphenoxy skeleton and other closely related structures employing the thiocyanate moiety as polar end group were designed, synthesized, and evaluated as antiproliferative agents against Trypanosoma cruzi, the parasite responsible for this disease. These thiocyanate analogues were envisioned bearing in mind the potent activity shown by 4-phenoxyphenoxyethyl thiocyanate (compound 8) taken as lead drug. This compound had previously proved to be an extremely active growth inhibitor against T. cruzi with IC(50) values ranging from the very low micromolar level in epimastigotes to the low nanomolar level in the intracellular form of the parasite. Of the designed compounds, the ethyl thiocyanate drugs connected to nonpolar skeletons, namely, arylthio, 2,4-dichlorophenoxy, ortho-substituted aryloxy, and 2-methyl-4-phenoxyphenoxy (compounds 15, 34, 47, 52, 72, respectively), were shown to be very potent antireplicative agents against T. cruzi. On the other hand, conformationally restricted analogues as well as branched derivatives at the aliphatic side chain were shown to be moderately active against T. cruzi growth. The biological activity of drugs bearing the thiocyanate group correlated quite well with the activity exhibited by their normal precursors, the tetrahydropyranyl ether derivatives, when bonded to the same nonpolar skeleton. Compounds having the tetrahydropyranyl moeity as polar end were proportionally much less active than sulfur-containing derivatives in all cases. Drugs 47 and 72 also resulted to be very active against the amastigote form of the parasite growing in myoblasts; however, they were slightly less active than the lead drug 8. On the other hand, compounds 34 and 52 were almost devoid of activity against myoblasts. Surprisingly, the dithio derivative 15 was toxic for myoblasts.