Mitigation and control of bacterial corrosion must have been prioritized for failure anticipation by corrosion associated with microorganisms. Using synthetic inhibitor for material protection from corrosion have problems because it is not eco-friendly. This work conducted preliminary studies on T. cordifolia stem extracts at various methanol ratios against P. aeruginosa biofilm as a new eco-friendly inhibitor. The T. cordifolia stem was extracted by the maceration method with a different ratio of methanol solvent (100, 75 and 50 %). The bacterial activity was assessed using the dilution method (MTT assay) to determine the minimum inhibitory concentration (MIC). The total phenolic and flavonoid contents were determined using Follin-Ciocalteu and aluminum chloride (AlCl3) colorimetric, respectively. Meanwhile, the structure of the active compounds in the extract was identified by using the liquid chromatography-mass spectrometry/mass spectrometry (LC-MS/MS). The yields of T. cordifolia extracts are 9.45, 7.56 and 8.40 % at 100, 75 and 50 % of methanol ratios, respectively. The total phenolic content (TPC) in crude methanol extract of T. cordifolia is 11.20, 9.46 and 6.56 % for 100, 75 and 50 % of methanol ratios. Meanwhile, the flavonoid content (TFC) was obtained is about 5.25, 0.64 and 0.33 % for 100, 75 and 50 % of methanol concentrations, respectively. The T. cordifolia extract has antibacterial activity against P. aeruginosa in all ratios of methanol concentrations. The minimum inhibitory concentration (MIC) of T. cordifolia methanol extract was found at 4096 µg/mL. Methanol solvent concentration with the most active antibacterial activity of the extract was 50 %, followed by 75 and 100 %. The structures of the active compound in the methanolic extract of T. cordifolia are a phenolic group, and it is in the alkaloid derivatives (Calopiptin, d-Lirioferine (Lirioferine), Moupinamide, Piperanine, and Yuanhunine). © 2023, Walailak University. All rights reserved.