Background <p>Testicular ischemia-reperfusion injury (tIRI) has been shown to cause testicular damage and lower sperm quality through the initiation of oxidative stress and inflammation. Conversely, vitamins C and E have been independently established to confer cellular protection by suppressing oxidative stress. However, the effect of vitamins C and E, when administered singly or in combination, on tIRI is yet to be explored.</p> Methods <p>Therefore, this study investigated the impact of vitamins C and E, when administered singly or in combination, on tIRI and the associated mechanism. Fifty adult male Wistar albino rats were randomly divided into sham-operated, tIRI, Vitamin C+ tIRI, Vitamin E + tIRI, and Vitamin C + E+ tIRI groups.</p> Results <p>tIRI led to a distortion of testicular histo-architecture (<i>p</i> = 0.0166) and a reduction in sperm count (<i>p</i> = 0.0200), and viability (<i>p</i> = 0.0067), as well as an increased number of sperm with abnormal morphology (<i>p</i> &lt; 0.0001). tIRI suppressed circulating FSH (<i>p</i> = 0.0372), LH (<i>p</i> = 0.0043), and testosterone (<i>p</i> = 0.0029), significantly increased testicular MDA levels (<i>p</i> &lt; 0.0001) and reduced GSH rats (<i>p</i> = 0.0318) and protein thiol levels (<i>p</i> = 0.0136), and SOD (<i>p</i> = 0.0164) and catalase activities (<i>p</i> = 0.0223). tIRI also led to an increased NO level (<i>p</i> = 0.0068), myeloperoxidase activity (<i>p</i> = 0.0049), and NF-kB expression (<i>p</i> &lt; 0.0001). tIRI-induced histological and biochemical alterations were attenuated by vitamins C and E, administered singly and when combined. However, co-administration of vitamins C and E did not exert a superior effect compared to the administration of either vitamin C or vitamin E.</p> Conclusion <p>In conclusion, vitamins C and E attenuated tIRI by suppressing NF-kB-dependent inflammation and oxidative stress.</p>