Pharyngitis is a leading cause of outpatient antibiotic use, despite its typically viral or self-limiting nature. Such unnecessary antibiotic therapies are not only the cause of increasing antibiotic resistance, but also significant changes in the human microbiota in the intestines and other locations, which translate into immune disorders and an increased risk of developing several chronic diseases. Orally administered octenidine-containing lozenges provide a topical alternative; however, their effects on the host microbiota of the oral cavity, throat, and intestine remain unclear. In this study, we evaluated the antimicrobial and antibiofilm in vitro activity of octenidine lozenges against 106 microbial strains, including pathogens and commensals from the oral cavity, pharynx, and large intestine. Minimal biocidal concentrations (MBCs) and minimal biofilm eradication concentrations (MBECs) were determined under physiologically relevant exposure times: 23 min for oral contact and 24 h for intestinal transit. ADME in silico analysis confirmed the lack of absorption of octenidine through the blood–brain barrier and the gastric intestinal mucosa. At concentrations achievable in saliva and the intestinal lumen, octenidine effectively eradicated in vitro all oropharyngeal pathogens while leaving intestinal commensals unaffected. Its impact on oral commensals resembled that of routine mechanical cleaning. These in vitro findings are of high translative value because they support the use of octenidine lozenges as a safe topical treatment for pharyngeal infections, “sore throat”, without adverse effects on the gut microbiota.