BACKGROUND: Periprosthetic joint infection is the most common cause of readmissions after total joint arthroplasty (TJA). Intrawound vancomycin powder (VP) has reduced infection rates in spine surgery; however, there are no data regarding VP in primary TJA. METHODS: Thirty-four TJA patients received 2 g of VP intraoperatively to investigate VP's pharmacokinetics. Serum and wound concentrations were measured at multiple intervals over 24 hours after closure. RESULTS: All serum concentrations were subtherapeutic (<15mug/mL) and peaked 12 hours after closure (4.7mug/mL; standard deviation [SD], 3.2). Wound concentrations were 922 mug/mL (SD, 523) 3 hours after closure and 207 mug/mL (SD, 317) at 24 hours. VP had a half-life of 7.2 hours (95% confidence interval, 7.0-9.3) in TJA wounds. CONCLUSIONS: VP produced highly therapeutic intrawound concentrations while yielding low systemic levels in TJA. VP may serve as a safe adjunct in the prevention of periprosthetic joint infection.