The Fuchuan ophiolite is located in the northeasternmost segment of the Neoproterozoic Jiangnan orogen and consists mainly of harzburgites, with minor dunites, pyroxenite and gabbro veins and dykes. In order to investigate the genesis and tectonic setting of the Fuchuan ophiolite and chromitites, in situ analyses of unaltered chromites and silicates were carried out. Trace element analyses of unaltered chromites from the Fuchuan chromitites indicate the parental magma is of mid-ocean ridge basalt (MORB)-like origin, with the Ti/Fe3+#–Ga/Fe3+# diagram of chromites showing that the chromitites are a result of melt/rock interaction of MORB melts with mantle peridotites, and that the Fuchuan harzburgites present the dual features of MORB and supra-subduction zone peridotites (SSZP). Trace and rare earth element (REE) analyses of olivines and orthopyroxenes from the Fuchuan harzburgites hint at the possibility of mantle metasomatism influenced by SSZ-subducted fluids. Finally, integrating with previous study, the Fuchuan ophiolite and chromitites might have been formed in a back-arc spreading ridge between the Yangtze and Cathaysia blocks during the Neoproterozoic.