β-Cyclodextrin (β-CD)-based polymers have shown high adsorption capacities for removing per- and polyfluoroalkyl substances (PFAS) from drinking water. PFAS capture by these materials involves many physical and chemical processes, such as adsorption and inclusion complexation. Quantifying the underlying host–guest binding between CDs and PFAS nevertheless remains essential because it governs the primary inclusion step. Here, we investigated native and linker-modified CD–PFAS inclusion complexes in aqueous solution using isothermal titration calorimetry (ITC) and molecular dynamics (MD) simulations with the attach–pull–release (APR) method. We computed the Gibbs free energies of binding for α-, β-, and γ-CDs with seven linear PFAS, including perfluorocarboxylic acids and perfluorosulfonic acids, and found reasonable agreement with our experimental measurements and previously reported data. Comparison between implicit and explicit solvent calculations suggests that the apparently better agreement of the implicit solvent model for some native monomer complexes is likely due to error cancellation rather than a more transferable physical description, whereas explicit solvent is required to capture solvent-related salt and linker effects. Overall, β-CD exhibited the strongest binding affinities, whereas α-CD showed negligible affinities and γ-CD bound PFAS more weakly than β-CD. Hydrogen bonding, interaction-energy decomposition, and solvent-accessible surface area showed that host–guest hydrogen bonding cannot uniquely predict affinities, and that hydrophobic dehydration plays a dominant role in binding. We further examined how background ion concentration affects β-CD–PFAS binding and found that explicit solvent simulations capture a clear salt dependence, with Li/Merz ion parameters describing the high-salinity trend more reasonably than the Joung–Cheatham model. To mimic the local microenvironment surrounding CD units in polymers, we also examined three linker-modified β-CD models containing phenyl groups. Bind3P water correctly reproduced the experimentally reported enhancement of PFAS adsorption with increasing linker number, and energy decomposition showed that linker groups strengthen PFAS binding by enhancing local hydrophobic confinement and specific linker–guest interactions. Overall, this combined experimental and computational study provides molecular-level insight into the building blocks of cyclodextrin polymers and lays the groundwork for future in silico construction of CD polymer models for PFAS adsorption under diverse water-matrix conditions. ... Read more