Multiplex Nanopore Detection of Structurally Diverse Per- and Polyfluoroalkyl Substances in Surface Water

Perfluoroalkyl and polyfluoroalkyl substances (PFASs) make up a large class of emerging chemical pollutants that have caused extensive contamination of global water sources. As the toxicity of PFAS becomes increasingly recognized, there is a growing demand for low-cost and rapid sensors capable of screening water samples for multiple PFAS species. However, most electrochemical and optical sensors can detect only one or two PFASs, despite the high structural diversity of these compounds in aquatic environments. Here, we report a single-molecule nanopore sensor that enables simultaneous detection of nine PFASs in a single measurement. By incorporating a β-cyclodextrin (β-CD) adapter into a mutant α-hemolysin (α-HL) nanopore, translocating PFAS molecules produce distinct current blockades that allow clear discrimination based on carbon chain lengths, hydrogen substitutions, and terminal functional groups. With assistance from a machine learning classifier, an overall identification accuracy of 95.83% is achieved. This strategy allows direct, label-free, and rapid discrimination of multiple PFASs in surface water samples at environmentally relevant concentrations as low as the microgram per liter level, without chemical labeling, separation, or enrichment. The successful demonstration of nanopore sensing in complex real-world matrices highlights its strong potential for practical, field-deployable environmental analysis.