Purpose The aim of this prospective study was to compare in non-Asian subjects the pharmacokinetics (PK), pharmacodynamics (PD), safety, and tolerability of two short-acting cardioselective beta 1-adrenergic antagonists, landiolol and esmolol, after administration of three different bolus dosages. Materials and methods We conducted a single-center, prospective, double-blinded, randomized study in three cross-over periods with 12 healthy subjects (7 women and 5 men, mean age of 24.5 +/- 6.9 years) each receiving three doses of landiolol (0.1, 0.2, and 0.3 mg/kg BW) either in a newly developed concentrate i.v. formulation (RapiblocA (R) 20 mg/2 mL concentrate) or a lyophilized formulation, or three doses of esmolol (0.5, 1, and 1.5 mg/kg BW) in an i.v. formulation (BreviblocA (R) 100 mg/10 mL). PK and PD parameters, safety, and tolerability were assessed. Findings Results of the two landiolol formulations were reported previously and were similar. For the landiolol concentrate formulation and esmolol, maximum blood concentrations were rapidly reached (mean t(max) ranged between 1.8 and 3.0 min for landiolol and 1.8 to 2.4 min for esmolol). The parent drugs disappeared very fast from the blood stream, with a t(1/2) of 3.2 +/- 1.2 (SD) minutes and 3.7 +/- 2.1 (SD) minutes for the low doses of landiolol and esmolol, respectively. Despite comparable injection rates (0.1 or 0.5 mg/kg/15 s for landiolol and esmolol, respectively), the onset of significant heart rate reduction occurred earlier in response to landiolol (1 min) than in response to esmolol (2 min). In addition, significantly lower heart rate values were obtained at every dose level of landiolol, in comparison to esmolol (p < 0.05). Both compounds reduced the systolic blood pressure to a comparable degree. Especially at the highest dose, the duration of blood pressure reduction was longer under esmolol compared to landiolol. Seven mild to moderate adverse events occurred after administration of landiolol, and five occurred after administration of esmolol. No serious adverse events were reported in this study.