This study was originated in recognition that little data have been accumulated in the literature regarding concentrations of gentamicin in various infected tissues. The study was undertaken to determine if systemically administered gentamicin penetrates burn wound eschar and to determine if there is a relationship between gentamicin serum levels attained and concentration of gentamicin reaching the eschar. Five hospitalized adult patients requiring parenteral gentamicin for sepsis secondary to third-degree burns were studied prospectively. For the purpose of determining the amount of gentamicin reaching the eschar, biopsy of the burn wound eschar was performed on each patient. Venous blood samples were also obtained to examine the relationship of serial serum samples to eschar levels. Calculated pharmacokinetic parameters include elimination rate constants, apparent volume of distribution, β, and gentamicin half-life. The data indicate that systemically administered gentamicin does penetrate burn eschar in measurable quantities. Deeper layers of eschar tissue contained significantly greater quantities of gentamicin. Analysis of variance resulted in statistically significant (p < 0.05) rise in eschar concentration with time. A positive correlation between the peak serum gentamicin concentration and the concentration of gentamicin reaching the burn tissue was found. The quantity of gentamicin attained was sufficient to prevent proliferation of organisms cultured from the eschar of patients. Accumulation of gentamicin in the eschar occurred in every patient studied. This study suggests the eradication of organisms in eschar samples is due to the achievable concentration of gentamicin at this site.