Gouania lupuloides (L.) Urb. (Rhamnaceae), a medicinal plant commonly used in the Caribbean for dental health, is known as chew stick (chewstick) or bejuco de Indio. Few studies have assessed its chemistry and biological activity. The overall aim of this research is to understand the phytochemistry and antibacterial activity of Gouania lupuloides, as well as its continued cultural use. Authenticated Gouania lupuloides samples collected in Jamaica were extracted, subjected to solvent-solvent partitioning, and evaluated for inhibitory activity against pathogenic oral bacteria. Commercial Caribbean samples of bejuco de Indio available in the United States were compared chemically to known Gouania lupuloides samples through targeted and untargeted UPLC-QTOF-MS analysis. Ethnobotanical fieldwork in rural and urban Jamaica demonstrated the ongoing cultural use of Gouania lupuloides as a natural toothbrush, and in the preservation of traditional fermented beverages. Antibacterial assays demonstrated that triterpenes from Gouania lupuloides, including ceanothic acid, had activity against several of the tested oral pathogens, such as Actinomyces viscosus, Prevotella intermedia, and Peptostreptococcus micros. Principal component analysis indicates that at least one of the chew stick samples sold as bejuco de Indio was chemically distinct from Gouania lupuloides type specimens. Presence of ceanothic acid was used to characterize a chemical fingerprint typical of Gouania lupuloides to aid in the chemotaxonomic identification of unknown chew sticks. Additionally, antibacterial ceanothic acid was present in similar relative abundance in all samples identified as Gouania lupuloides, further supporting its traditional use for oral health and hygiene.