The aim of the present study was to determine whether matrix metalloproteinase (MMP) activities are associated with the clinicopathological features of oral squamous cell carcinomas (OSCCs). Forty-four samples of primary OSCCs were microdissected, incubated in serum-free cell culture medium for 16 h, and the activity of secreted MMPs analyzed by gelatin zymography and densitometry. Two major enzymes with gelatinolytic activity were produced by all tumor samples and characterized as MMP-2 and -9 by their molecular weight, immunoprecipitation using monoclonal antibodies, and specific inhibition by 1,10-phenanthroline or EDTA. Patients with tumors showing elevated activity of MMP-2 and -9 had shorter disease-free survival after treatment than patients with tumors exhibiting low MMP activities. These results suggest that the zymographic measurement of the MMP-2 and -9 activities in OSCC specimens may be useful to predict the disease-free survival period of patients affected by these tumors.