The purpose of the present research was to determine if metalloproteinase levels were elevated in human chronic wound fluid. Samples of blood and wound fluid from acute (mastectomy) and chronic (leg ulcer) wounds were collected, and metalloproteinase profiles of the samples were determined by gelatin zymography. Compared to serum, acute wound fluid (mastectomy fluid) contained markedly increased levels (five- to tenfold) of several metalloproteinases including 72- kDa and 94-kDa gelatinases (MMP-2 and MMP-9). In chronic wound fluid, not only were these enzyme levels increased another five- to tenfold over mastectomy fluid, but also both activated enzyme and proenzyme species appeared to be present. Our results suggest that non-healing ulcers develop an environment containing high levels of activated metalloproteinases, which may result in chronic tissue turnover and failed wound closure.