Increased inflammation and heightened physiological stress reactivity have been associated with pathophysiology of depressive symptoms. The underlying biological mechanisms by which inflammation and stress can affect neurogenesis is through alterations of the kynurenine (KYN) pathway, which is activated under stress. Meanwhile, n-3 polyunsaturated fatty acids (n-3 PUFA) supplementation has anti-inflammatory properties and may enhance stress resilience. Whether n-3 PUFA alters stress responsivity KYN is unknown. This placebo-controlled trial examined the effect of n-3 PUFA on KYN metabolism, inflammation, depressive symptoms and mood; and stress-induced changes following a laboratory stressor. Using a placebo-controlled design, 47 healthy male adults were received 4g/d of Omega-3, or a placebo for 12 weeks. A significant group-by-time interaction was found for inflammation markers - gp130 (F= 7.07, p = 0.011), IL-6R alpha (F= 10.33, p = 0.003), and TNF_RI (F= 10.92, p = 0.002). No significant group-by-time interactions were found for mood or depressive symptoms, except for Hedonic Tone (F= 6.50, p = 0.014), neither for KYN metabolites and stress-induced changes to the KYN metabolites and mood following a laboratory stressor. Overall, elevating n-3 PUFAs levels in healthy males ameliorates inflammatory markers but does not ameliorate KYN metabolism, depressive symptoms, mood, or stress responsivity KYN metabolism and mood. The study was registered at ClinicalTrials.gov with identifier NCT05520437 (30/08/2022 first trial registration).