Daptomycin was previously shown to reduce gentamicin renal toxicity and this toxicity was not delayed by the concomitant injection of daptomycin (Thibault N., L. Grenier, M. Simard, M. G. Bergeron, and D. Beauchamp, Antimicrob. Agents Chemother., 38 1027-1035 (1994)). The protective effect of daptomycin against gentamicin toxicity was evaluated in 96 female Sprague-Dawley rats. Normal and nephrectomized rats were treated with saline (NaCl, 0.9%), gentamicin (30 mg/kg/12 hrs, i.p.), daptomycin (10 mg/kg/12 hrs, s.c.) or with a combination of daptomycin plus gentamicin during 4 and 10 days. On day 4, gentamicin and daptomycin cortical levels were higher in nephrectomized gentamicin-daptomycin-treated rats (p < 0.05) as compared to all other groups. The accumulation of gentamicin or daptomycin in nephrectomized gentamicin-daptomycin-treated or gentamicin-saline-treated rats was higher on day 4 (p < 0.01) than on day 10. Other parameters such as the sphingomyelinase activity in the renal cortex, the serum creatinine, and the histopathology showed significantly fewer changes in daptomycin-gentamicin-treated rats as compared to animals given gentamicin alone. On the other hand, the protection of daptomycin was less extensive in nephrectomized rats as compared to normal rats. Daptomycin and gentamicin were localized in the lysosomes of proximal tubular cells of animals treated with daptomycin and gentamicin given alone or in combination. These results suggest that daptomycin protects against gentamicin toxicity in nephrectomized rats but to a lesser extent than in normal rats.