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Glutaraldehyde is a high-efficiency disinfectant that has been included in the protocols of some hospitals for controlling the spread of SARS-CoV-2, together with sodium hypochlorite and quaternary ammonium disinfectants. However, exposure has been poorly studied in workplace settings, despite the association between glutaraldehyde and respiratory diseases and skin conditions in exposed workers. This study evaluated the magnitude of exposure associated with the use of glutaraldehyde in healthcare workers across various work areas of a first level of Hospital-Based Care in Colombia. Workers were classified into similar exposure groups (SEGs) based on work areas and tasks performed, and airborne concentrations of glutaraldehyde were measured in different work areas of each SEG through direct monitoring. The 95th percentile of glutaraldehyde concentrations in all SEGs studied exceeded the TLV-C immediately after use. Cleaning workers and nurses had the highest exposures to glutaraldehyde. Results indicate that workers were overexposed and highlight the need to implement controls to reduce exposure. The high-exposure levels also raise the need to consider glutaraldehyde substitution and adequate use of personal protective equipment (PPE)., (© The Author(s) 2023. Published by Oxford University Press on behalf of the British Occupational Hygiene Society.)