One of the most common complications in patients with incontinence is incontinence-associated dermatitis. This study aimed to examine the influences of lipidolytic enzymes and/or proteases on skin barrier and tissue structure on the development of incontinence-associated dermatitis. Two animal experiments, ex vivo and in vivo, were performed using rats to examine the influences of 3 factors (maceration, proteases, and lipidolytic enzymes) alone or in various combinations on the barrier function and histology of the skin. As a result, skin treatments, including both of the skin maceration and proteases application, caused erythrocyte leakage from the blood vessels in the dermis. The erythrocyte leakage was observed in a larger area in the skin treated with proteases and lipidolytic enzymes with maceration than in the skin treated with proteases with maceration, that is, the addition of lipidolytic enzymes to skin maceration with proteases enhanced erythrocyte leakage. Lipidolytic enzymes in macerated skin are factors that accelerate tissue damage via skin barrier impairment, and proteases are the factors that trigger the development of incontinence-associated dermatitis via tissue damage. Advanced nursing care of perineal skin in patients with faecal incontinence is required because of the deleterious influence of lipidolytic enzymes and proteases.