Introduction. Immobilized subtilisins are currently used as a fibrinolytic agent in antithrombotic drugs. The effect of immobilized subtilisins (ISs) on the contractile function of the heart has not been previously studied. Aim. To study the effect of coronary perfusion with immobilized subtilisins (ISs) on the contractile function of an isolated rat heart in an experiment. Materials and methods. The study used a Langendorff model of coronary retrograde perfusion of an isolated rat heart with solutions of ISs at different concentrations. The experiment included 50 male rats. The animals were divided into 5 groups: group 1 – control (hearts perfused only with Krebs-Henseleit solution) and 4 experimental groups – hearts perfused with a solution of ISs at concentrations of 170, 340, 510 and 1020 IU/l, respectively. The contractile function (inotropic effect) of the isolated rat heart was assessed in terms of the pressure developed by the left ventricle (LVP). Results. Perfusion of an isolated rat heart with a solution of ISs for 40 min causes a positive inotropic effect in all experimental groups. At a concentration of 170 IU/l, the effect is observed from the 10th minute, at higher concentrations – from the 5th minute. The duration of the effect’s rising when using ISs in the dose range of 170–510 U/l manifests itself up to the 20th minute, and when using a dose of 1020 U/l – up to the 30th minute. Then there is a weakening of the inotropic effect. In groups of animals with the introduction of immobilized subtilisins at a concentration of 340–1020 U/l, there is a decrease of LVP by the 40th minute, but the values of this indicator remain higher compared to the control group. Conclusion. Perfusion of the isolated heart with a solution of ISs increases the contractile activity of the isolated heart. When the heart is perfused with ISs, there is an increase of LVP. The inotropic effect is dose-dependent at various time points.