This preclinical study compared the muscle weakening efficacy, duration, and safety margin of the recently approved botulinum toxin type B (BTX-B; Myobloc/Neurobloc) to botulinum toxin type A (BTX-A; BOTOX((R))). Mice received a single hind limb intramuscular injection of BTX-B (1-150U/kg) or BTX-A (1-120U/kg). An observer who was masked to treatment assessed the magnitude and duration of muscle weakening efficacy on a 0-4 scale using the digit abduction scoring assay. Safety margins were determined as the ratio of the IM median lethal dose to the IM dose that produced half-maximal muscle weakness in the DAS. BTX-A produced muscle weakness at lower doses than BTX-B (IM ED(50): 6.2+/-0.6 vs. 20.8+/-1.4U/kg, respectively) (p<0.0001). BTX-A at 29U/kg and BTX-B at 67U/kg produced comparable peak DAS scores of approximately 4 indicating maximal muscle weakness. At these doses, the duration of BTX-A was longer, with a return to baseline by day 36 compared to a return to baseline by day 14 with BTX-B. The mean dose that was lethal in 50% of mice was lower for BTX-A than BTX-B (81.4+/-3.5 vs. 104.6+/-1.9U/kg, respectively) (p<0.001) and the safety margin was higher (13.9+/-1.7 vs. 5.4+/-0.3, respectively (p<0.001). These results indicate that the BTX-A:BTX-B dose ratio for muscle weakening efficacy is different from the ratio for systemic effects following IM injections and suggest that no single dose ratio is adequate to compare these preparations. The in vivo differences found are consistent with the different clinical profiles reported for these two products.