Opportunistic pathogenic bacteria, such as Pseudomonas aeruginosa, pose a serious risk to patients suffering from a compromised immune system and those patients with cystic fibrosis. Confirming their presence often requires culturing the bacteria, which can take days. Herein is proposed a rapid electrochemical detection method based on P. aeruginosa-generated small molecules employed for chemical communication, referred to as quorum sensing (QS) molecules, within the biofilm formed by the bacteria; specifically, 4-hydroxy-2-heptylquinoline (HHQ) and 2-heptyl-3,4-dihydroxyquinoline (pseudomonas quinolone signal, PQS). This method does not depend on the redox activity of the QS molecules. Instead, as a proof-of-concept, electrochemical monitoring was achieved through aqueous alkali metal ion and proton interfacial complexation with organic-solubilized HHQ and PQS at an interface between two immiscible electrolytic solutions, specifically, between water and 1,2-dichloroethane. The proton/HHQ and proton/PQS binding stoichiometries were discovered to be 1:3 and 1:2, respectively, which is likely due to the relatively high concentrations of QS molecules employed. Owing to the biphasic nature of the methodology, experimental complications due to the poor solubility of the hydrophobic QS molecules in aqueous media were avoided.