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Abstract Objective To demonstrate that the intraosseous (IO) access is more effective compared with the intravenous (IV) access for prophylactic antibiotic administration in knee joint surgeries, using 36 pigs as live models. Materials and Methods Skin, subcutaneous tissue, cartilage, and bone samples were collected, analyzed and compared after the administration of IV or IO antibiotic in different groups. Results When comparing the IO and IV groups, the IO group showed a higher concentration of prophylactic antibiotic in the skin (p = 0.049), cartilage (p = 0.018), and bone (p = 0.002), in the analysis of the first 24 hours after 30 minutes of infusion. Conclusion Since complications regarding this practice are rare, the use of this pathway may be an alternative to reduce the risk of surgical site infection in orthopedic surgeries, leading to a decrease in morbidity and mortality and hospital expenses with readmission or prolonged hospitalization time. However, further research and further experimental studies in humans are required, as the effectiveness of the method in pigs has been proved.