We prepared 4.5% and 2.6% copper oxide/hydroxide nanoparticle highly dispersed in graphite-like carbon film (Cu–NDC) by RF co-sputtering copper and carbon together. The two film structures were investigated by X-ray photoelectron spectroscopy (XPS) and transmission electron microscope (TEM) techniques. The nanoparticles size is about 4–5 nm . The copper particles in the two different content films have different chemical state. The copper in 4.5% Cu–NDC film exists mainly as cupric hydroxide (Cu(OH)₂), while in 2.6% Cu–NDC film cuprous oxide (Cu₂O) is the predominant form. The electrochemical properties of the two kinds of Cu–NDC films were evaluated by detecting glucose in an alklaline medium at constant applied potentials in flow system. The 4.5% Cu–NDC film electrode has higher electrocatalytic ability than that of 2.6% Cu–NDC film electrode with regards to electrooxidation of glucose. Both of the two Cu–NDC film electrodes have good stability.