This study compared the respective intramuscular (IM) safety margins of two preparations of botulinum toxin (BTX) serotype A and one preparation each of BTX serotypes B and F in mice. Mice received an IM injection (0-200 U kg(-1) body weight) of BTX-A (BOTOX or DYSPORT), an experimental preparation of BTX-B (WAKO Chemicals, Inc.), or an experimental preparation of BTX-F (WAKO). An observer who was masked to treatment scored muscle weakness using the Digit Abduction Scoring (DAS) assay. Peak DAS responses were plotted and IM ED(50) values calculated. The safety margin for each BTX preparation was calculated as a ratio of the IM median lethal dose after hind limb injection to the median effective dose in the DAS assay (IM LD(50)/IM ED(50)). Experiments were repeated 4-6-times for each preparation (10 mice/dose). Mean safety margin values were highest for BTX-F (WAKO; 16.7+/-3.9) and one of the BTX-A preparations (BOTOX; 13.9+/-1.7). Mean safety margins values for the other BTX-A preparation (DYSPORT) and BTX-B (WAKO) were significantly lower (7.6+/-0.9 and 4.8+/-1.1, respectively). Thus, the BTX preparations exhibited different safety margins in mice. These results support the hypothesis that the preparations are unique therapeutics and are not interchangeable based on a simple dose ratio.