Background: Hand sanitizer use increased dramatically during the COVID-19 pandemic, yet knowledge about its safe use and potential health impacts remains limited, particularly across different occupational groups. There is a need to better understand the frequency and health effects of hand sanitizer use, especially when comparing healthcare professionals and non-healthcare professionals. Methods: This cross-sectional study included 305 adults from a central district of Türkiye, with 48.5% being healthcare professionals. Data were collected on participants’ knowledge of sanitizer contents, frequency of use, and any health problems experienced. Regression analysis was performed to identify factors associated with sanitizer-related health problems. Results: Only 28.4% of healthcare professionals and 12.1% of non-healthcare professionals reported full knowledge of the contents of the sanitizers they used. Daily use of hand sanitizer 10 or more times was reported by 53.4% of healthcare professionals and 33.8% of non-healthcare professionals. Regression analysis indicated that female gender and being a healthcare professional were significantly associated with sanitizer-related health problems (p < .01). Despite frequent use, both groups demonstrated insufficient knowledge regarding safe sanitizer use, which may contribute to increased health risks such as skin irritation or allergic reactions. Conclusions: These findings highlight the importance of implementing targeted, evidence-based educational interventions and developing clear guidelines on the safe use of hand sanitizers for both healthcare and non-healthcare professionals. Future studies should focus on the effectiveness of these interventions in reducing healthcare-associated infections and improving hygiene practices. Application to Practice: Occupational health and public health professionals can use these results to design workplace and community-based training programs, promote safer hygiene practices, and minimize sanitizer-related health risks across diverse populations.