Daptomycin is a lipopeptide antibiotic that exhibits bactericidal activity against Gram-positive bacteria. In pursuit of potential clinical dosing regimens for endocarditis, we evaluated two, once-daily daptomycin regimens against multiple, drug-resistant Gram-positive pathogens. Daptomycin susceptibility was determined in the absence and presence of physiologic concentrations of albumin. An in vitro pharmacodynamic model with simulated endocardial vegetations incorporating protein was used to simulate regimens of daptomycin at 6 and 8 mg/kg/day and vancomycin at 1 g every 12 h against methicillin-resistant S. aureus (MRSA-67 and 494) and S. epidermidis (MRSE-R227 and R617), glycopeptide-intermediate S. aureus and S. epidermidis (GISA-992 and GISE-12333), and vancomycin-resistant E. faecium (VREF-SF12047 and 12366). Bacterial quantification occurred over 72 h. Daptomycin MIC results for study isolates in the absence or presence of albumin ranged from 0.125 to 4 and 1 to 8, respectively. Both daptomycin regimens achieved greater than 99.9% kill by 8 h and demonstrated greater bacterial reduction than vancomycin against all tested isolates at 24, 48, and 72 h (p < 0.05). Undetectable limits of bacterial quantification was achieved and maintained by 8 mg/kg/day against MRSA-494 and 67, GISA-992, and VREF-590 for the study duration. Although slight regrowth was noted only for 6 mg/kg/day against MRSA-67, 99.9% kill was maintained throughout the study period without development of resistance. Pharmacodynamic profiles and drug exposure of daptomycin at 6 mg/kg/day corresponds to previously reported AUC/MIC requirements. These results suggest that 6 and 8 mg/kg/day of daptomycin represent potential regimens for further clinical evaluation in drug-resistant Gram-positive endocarditis.