Three-dimensional (3D) copper oxide (CuO) nanostructures were synthesized in a regenerated Bombyx mori silk fibroin aqueous solution at room temperature. In the synthesis process, silk fibroin served as the template and helped to form the hierarchical CuO nanostructures by self-assembly. Cu(OH)(2) nanowires were formed initially, and then they transformed into almond-like CuO nanostructures with branched edges and a compact middle. The size of the final CuO nanostructures can be tuned by varying the concentration of silk fibroin in the reaction system. A possible mechanism has been proposed based on various characterization techniques, such as scanning and transmission electron microscopy, X-ray diffraction, and thermogravimetric analysis. The synthesized CuO nanostructured material has been evaluated as an anode material for lithium ion batteries, and the result showed that they had a good electrochemical performance. The straightforward energy-saving method developed in this research may provide a useful preparation strategy for other functional inorganic materials through an environmentally friendly process.