Abstract
Surface-enhanced Raman spectroscopy (SERS) in plasmonic nanocavities enables single-molecule detection through dramatic enhancement of the local electromagnetic field. However, single-molecule SERS (SM-SERS) signals exhibit pronounced fluctuations in both absolute and relative band intensities, as well as abrupt signal dropouts, which complicate reliable analyte detection and identification. A key contributor to this temporal variability is the translational and rotational mobility of molecules within the plasmonic cavity. In this work, we investigated how confining thionine (Th) molecules within the macrocycle cucurbit[7]-uril (CB[7]) suppresses molecular motion and improves spectroscopic stability. We employed two high-field-enhancement geometries nanoparticle-on-mirror and spherical gold oligomers. The spectral analyses were supported with density functional theory (DFT) calculations and simulations. Our results demonstrate that CB[7] encapsulation improves SM-SERS detection reliability by reducing amplitude fluctuations. Although the average SERS intensity decreases by several tens of percent, signal decay during initial illumination accelerates. Under electronic-resonant excitation of the analyte, detection probability increases owing to the CB[7]-enforced optimal alignment of Th's transition dipole moment with the nanocavity's electromagnetic field. Limiting analyte mobility through encapsulation diminishes amplitude fluctuations, while spectral diffusion remains unaffected. These complementary results disentangle two fluctuation mechanisms: molecular motion suppressed by CB[7] and substrate/adatom dynamics unchanged by encapsulation. These findings provide fundamental insights into molecule-nanocavity interactions and establish new strategies for enhancing the reliability of single-molecule detection. The approach opens promising avenues for probing the dynamics of biologically and catalytically relevant species with improved temporal stability and reduced measurement uncertainty.