<b>Background and Objective:</b> Biofilms as a bacterial defense are relatively more difficult to eradicate with antibiotics, thus pathogenic bacteria in their biofilm form can cause serious problems for human health. Lakum <i>(Causonis trifolia</i> L.) is an herbaceous plant with many biological activities, one of which is an antimicrobial compound containing flavonoids, squalene, nimbidin, saponins, anthocyanins, tannins, myricetin, others. This study aimed to determine the antibiofilm activity of Lakum leaf extract against<i> Staphylococcus aureus </i>bacteria and the active compounds that play a role in inhibiting monomicrobial biofilms. <b>Materials and Methods:</b> This research method was carried out with an <i>in vitro</i> experimental study design using observations of phytochemical screening test results and the effectiveness of Lakum leaf antibiofilm on<i> Staphylococcus aureus</i> through microplate reader readings that measure optical density values. <b>Results:</b> This study showed that Lakum leaves contain alkaloids, flavonoids, phenolics, polyphenols, tannins and saponins. In addition, Lakum leaves gave biofilm inhibitory activity in the middle and maturation phase with the highest concentration in 1% extract of 76.95±0.0007 and 72.85± 0.0003%, respectively. Meanwhile, the lowest concentration was 0.125% extract of 65.65±0.0001% in the middle phase and 59.71±0.0003% in the maturation phase. <b>Conclusion:</b> That Lakum leaves have biofilm inhibitory activity on <i>Staphylococcus aureus</i> with flavonoid compounds, tannins and polyphenols that work as active substances in inhibiting the biofilm formation.