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Stevioside is an important natural sweetener in food and beverages. There are concerns about its safety, but few methods for its determination. Silver nanoparticles (Ag NPs) functionalized with 4-mercaptophenylboronic acid (4-MPBA) were successfully used to determine stevioside in beverages based upon surface-enhanced Raman spectroscopy (SERS). The Ag NPs self-assemble with 4-MPBA through the covalent interaction of their sulfhydryl groups with the metal particles. Boronic acid groups bonded to 1,2-/1,3-diol groups form cyclic boronic esters with strong affinity and significant specificity for stevioside. The combination of the two leads to an alteration of the 4-MPBA vibrational mode. The change in the orientation of the moiety and the charge transfer resulted in a significant increase of the SERS signal. The substrates showed strong specificity and sensitivity to stevioside in the presence of other sweeteners. Also, the substrate modification conditions were optimized, including the time, ratio, and concentration. The limit of detection (LOD) for stevioside in beverages was 0.210 μmol/L, and the recovery was from 98.4% to 109.4%. In summary, the presented SERS method provides high sensitivity for stevioside. [ABSTRACT FROM AUTHOR]