This study investigated the influence of selenium nanoparticle (SeNP) allotropy on bioavailability, tissue distribution, antioxidant activity, and retention rates in animal models, with a focus on adult male Japanese quails. Red amorphous and grey crystalline selenium nanoparticles were synthesized and characterized using scanning electron microscopy with energy-dispersive X-ray spectroscopy (SEM–EDS), X-ray diffraction (XRD), Raman spectroscopy, and fluorescence analysis to confirm their structural differences at the nanoscale. Two in vivo experiments were conducted. In the first trial, 20 quails were assigned to five dietary treatments: control (no SeNPs), red SeNPs (0.05 and 0.5 mg/kg), and grey SeNPs (0.05 and 0.5 mg/kg). Growth parameters (Body weight and feed intake), organ indices, and tissue selenium concentrations were evaluated after 28 days. In the second experiment, 60 quails received control, red SeNPs (0.5 and 5 mg/kg), or grey SeNPs (0.5 and 5 mg/kg) for 28 days, followed by a 7-day selenium withdrawal period. Selenium distribution, antioxidant biomarkers (glutathione peroxidase, superoxide dismutase, and total antioxidant capacity), and selenium retention/depletion were assessed. Physicochemical characterization confirmed distinct allotropes: red SeNPs were amorphous, spherical particles (~218 nm), while grey SeNPs exhibited crystalline, needle-like morphology (Length=575±202 nm; Width=33.9±11nm). Fluorescence analysis revealed stronger optical activity for grey SeNPs, indicating structural-dependent electronic properties. Final Body weights and feed intakes were not significantly affected by SeNP supplementation in either experiment (P > 0.05), confirming the physiological safety of the applied doses. However, selenium metabolism showed clear form- and dose-dependent differences. Red SeNPs resulted in higher selenium accumulation in metabolically active tissues, including liver, red blood cells, and breast muscle, indicating greater bioavailability and rapid systemic distribution. In contrast, grey SeNPs demonstrated more selective accumulation, particularly in spleen and testis, suggesting organ-specific targeting. Antioxidant responses were significantly influenced by SeNP supplementation. High-dose red SeNPs (5 mg/kg) produced the greatest increase in hepatic glutathione peroxidase activity and serum total antioxidant capacity (P < 0.05), while grey SeNPs showed a moderate but consistent effect. Serum superoxide dismutase activity decreased in supplemented groups, indicating reduced oxidative stress. Selenium retention during the withdrawal phase was highest at moderate supplementation levels (0.5 mg/kg), reaching approximately 91% for red SeNPs and 88% for grey SeNPs. In contrast, high-dose treatments showed reduced retention and increased depletion rates, reflecting homeostatic regulation and enhanced excretion of excess selenium. In conclusion, selenium nanoparticle allotropy plays a critical role in determining selenium bioavailability and biological function. Red amorphous SeNPs act as a rapidly available selenium source with strong antioxidant effects, whereas grey crystalline SeNPs provide slower, more controlled release with organ-specific deposition. Moderate supplementation levels optimize selenium retention and biological efficiency. These findings provide a mechanistic basis for the targeted use of nano-selenium in animal nutrition to improve antioxidant status, tissue enrichment, and overall physiological function.