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Competitiveness is an essential feature of human social interactions. Despite an extensive body of research on the underlying psychological and cultural factors regulating competitive behavior, the role of biological factors remains poorly understood. Extant research has focused primarily on sex hormones, with equivocal findings. Here, we examined if intranasal administration of the neuropeptide oxytocin (OT) – a key regulator of human social behavior and cognition – interacts with changes in endogenous testosterone (T) levels in regulating the willingness to engage in competition. In a double-blind placebo-control design, 204 subjects (102 females) self-administrated OT or placebo and were assessed for their willingness to compete via an extensively-validated economic laboratory competition paradigm, in which, before completing a set of incentivized arithmetic tasks, subjects are asked to decide what percentage of their payoffs will be based on tournament paying-scheme. Salivary T concentrations (n = 197) were measured throughout the task to assess endogenous reactivity. Under both OT and placebo, T-reactivity during competition was not associated with competitiveness in females. However, in males, the association between T-reactivity and competitiveness was OT-dependent. That is, males under placebo demonstrated a positive correlation between T-reactivity and the willingness to engage in competition, while no association was observed in males receiving OT. The interaction between OT, T-reactivity, and sex on competitive preferences remained significant even after controlling for potential mediators such as performance, self-confidence, and risk-aversion, suggesting that this three-way interaction effect was specific to competitive motivation rather than to other generalized processes. These findings deepen our understanding of the biological processes underlying human preferences for competition and extend the evidence base for the interplay between hormones in affecting human social behavior.