Bothrojaracin (apparent mol. wt 27,000) is a potent inhibitor of thrombin previously isolated from the venom of Bothrops jararaca. Several molecular variants (isoforms) have been identified in a pool of venom collected from a large number of animals. In order to determine whether an individual snake produces a single type of bothrojaracin or multiple isoforms, we analyzed the bothrojaracin content of venoms collected individually from six adult B. jararaca snakes. Bothrojaracin was found in all venoms, but its activity was especially high in three of them. After purification on an alpha-thrombin affinity column, followed by gel filtration on Superose 12 HR, proteins from these three venoms migrated on sodium dodecyl sulfate-polyacrylamide gel electrophoresis as single bands of 27,000 and their amino-terminal sequences (residues 1-28) revealed extensive homology with bothrojaracin. In contrast, the material purified from the three venoms with low bothrojaracin activity consisted of bothrojaracin together with inactive proteins. Differences in the sequences obtained for bothrojaracin isolated from individual venoms indicated the existence of more than one isoform in the venom of an individual snake.