The crust shows large variations in Cu isotopic composition (δ 65 Cu relative to NIST 976) resulting from redox reactions and other supergene processes. The significant range of δ 65 Cu in mantle peridotites thus could be ascribed to recycled crustal materials and/or oxidative mantle metasomatism. However, the influence of normal magmatic fractionation processes on δ 65 Cu variations remains poorly understood, and it is unclear whether the magmatic processes also lead to large δ 65 Cu variations in mantle rocks. To address the issue, we present bulk-rock δ 65 Cu of fresh mantle pyroxenites from the Balmuccia peridotite massif, Northern Italy. These pyroxenites formed by melt-peridotite reaction and mineral accumulation from MORB-to OIB-like, sulfide-saturated basic magmas. Mass balance calculations show that sulfide phases host >98 wt % of the Cu budget of bulk rocks (87−484 μg/g). The pyroxenites show significant variations of δ 65 Cu from −0.66‰ to 0.66‰, which cover the range known for peridotites, including metasomatized ones. Three subsamples from the same pyroxenite layer display >0.3‰ decrease in δ 65 Cu with progressive differentiation. Nevertheless, δ 65 Cu does not display correlations with indicators of magmatic differentiation (e.g., Mg# and Cu/Pd) for all pyroxenites in this study. This might be attributed to variable extents of magmatic sulfide segregation for different samples and/or varying δ 65 Cu of the pyroxenite parent magmas, which were also affected by reactive migration through peridotites. The combined processes can change δ 65 Cu of evolving magmas and reacted peridotites and lead to Cu isotopic heterogeneity in the mantle, without involvement of oxidative metasomatism or recycled crustal materials.