Male fertility issues account for approximately 50% of all infertility cases in western society. With infertility affecting 1 in 6 couples in Australia, there is a pressing need to understand the underlying cause. Growing evidence suggests that the overproduction of reactive oxygen species (ROS) reduces the fertilisation potential of spermatozoa. The specific mechanism of this functional decline appears to be tied to a lipoxygenase-lipid peroxidation pathway. This occurs through the overproduction of ROS within the cell, liberating poly-unsaturated fatty acids in the plasma membrane, which are subsequently broken down by lipoxygenase proteins. We have recently confirmed that arachidonate 15-lipoxygenase (ALOX15) catalyses lipid peroxidation in human spermatozoa, yielding the cytotoxic aldehyde product 4-hydroxynonenal (4HNE). The involvement of ALOX15 within the lipid peroxidation pathway of human spermatozoa was assessed using the ALOX15 inhibitor, PD146176. PD146176 treatment, alongside an oxidative insult (H2O2), resulted in significant reductions to both cellular ROS (p < 0.01) and lipid peroxidation levels (p < 0.01) compared to our H2O2 treated controls. Excitingly, our functional data indicate that the inhibition of ALOX15 provides protection against oxidative stress with significant improvements in sperm motility (p < 0.01), acrosome reaction rates (p < 0.05) and the competence of these cells to interact with homologous human oocytes (p < 0.05). In addition to ALOX15, we have now established the presence of two additional lipoxygenase proteins, ALOX5 and ALOX12, in human spermatozoa that may contribute to cellular decline. To date, the use of a selective ALOX5 inhibitor (BW-B70C) and an ALOX5, 12 and 15 inhibitor (2-TEDC) have both shown significant reductions in lipid peroxidation levels compared to a treated control (p < 0.05 and p < 0.005 respectively). This work presents the possibility of targeting lipoxygenase proteins as a novel therapeutic strategy to lower oxidative stress in the male germline.