This study aims to evaluate the toxicity, antibacterial activity, and chemical profiling of Gracilaria salicornia extract. The research stages included the extraction, bioactivity test, GC-MS, and docking analysis. The antibacterial activity showed better sensitivity against Escherichia coli than Staphylococcus aureus. The inhibition zone against E. coli ranged from 7.73-17.30 mm, while S. aureus ranged from 6.91-13.44 mm, and all of the extracts were toxic to Artemia salina Leach. The chloroform extract showed the highest activity with the clear zone diameter was 17.3±1.65 mm and 13.44 ± 0.04 mm to E. coli and S. aureus respectively. The presence of steroids, terpenoids, fatty acids, and alkane based on GC-MS and docking analysis were responsible for these activities. Molecular docking analysis showed that Cholest-5-en-3-ol and Cholesta-4,6-dien-3-ol showed better interactions than control with binding energy -9.99 and -9.67 kcal.mol-1 against 5BMM, -8.00 and -7.93 kcal.mol-1 against 3LPS respectively. Based on this data, it can be stated that G. salicornia extract is highly potential as the source of antibacterial compounds.