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Diabetes mellitus, a metabolic disorder alters gonadal development and spermatogenesis, reactive oxygen species production, DNA damage, and apoptosis, which subsequently lead to male subfertility. Eugenol is an antioxidant, traditionally used as medication for digestive disorders and antioxidant therapy, decrease transport of glucose from GIT to systemic circulation. This experiment was aimed to decipher cellular and molecular insights of eugenol in protecting diabetic germ cells in rats. Rats were assigned randomly into five groups: control, eugenol control (Eugenol 400; EUG), diabetic (DIA), diabetic + eugenol 100 (DIA + EUG 100), and diabetic + eugenol 400 (DIA + EUG 400). EUG 400 and DIA + EUG 400 groups received 400 mg/kg eugenol orally. DIA + EUG 100 group received 100 mg/kg eugenol. Treatment was conducted for 4 weeks. Type 1 diabetes was induced by injecting a single i.p. dose of streptozotocin (55 mg/kg). Morphometric, biochemical, sperm parameters, oxidative stress, hormonal levels, histopathology, and fibrosis in the testis and epididymis, were evaluated. DNA damage was evaluated using halo and comet assays; DNA fragmentation and apoptosis using TUNEL assay. Eugenol treatment significantly normalized biochemical parameters, reduced MDA while increased albumin and GSH levels in diabetes. Eugenol significantly increased sperm numbers, motility and attenuated abnormal sperm head morphology in diabetes. Moreover, eugenol significantly reversed diabetes‐induced cellular damages, altered spermatogenesis, and collagen deposition in testis and epididymis. It also significantly attenuated diabetes‐associated DNA breaks and apoptosis. These findings suggest that 4 weeks treatment with 400 mg/kg of eugenol could be beneficial for diabetic patients to prevent subfertility. Highlights: Protective role of eugenol mediated by cellular and molecular mechanisms in diabetes‐induced testicular toxicity. Eugenol treatment significantly ameliorated oxidative stress in diabetic rat testes.Eugenol treatment prominently normalized spermatogenesis and sperm parameters in diabetes.Eugenol significantly protected diabetes‐induced DNA damage and apoptosis of testes and sperm. [ABSTRACT FROM AUTHOR]