Aims: This study aimed to evaluate the antibacterial effect of two new cationic surfactants based on phenylalanine-arginine (LPAM) and tryptophan-arginine (LTAM). Materials & methods: Antibacterial activity, mechanism of action and interactions with Staphylococcus aureus enzymes were measured through microbiological, flow cytometry and molecular docking assays, respectively. Results & conclusion: These compounds showed antibacterial activity in the range of 4.06-16.24 μg/ml against planktonic cells and no activity against mature biofilms, since they caused a loss of membrane integrity and increased DNA damage, as revealed by flow cytometry analysis. In silico assays revealed the existence of molecular bonds such as hydrogen bonds, mainly with DNA. Therefore, these compounds have promising pharmacological activity against MRSA strains.