The presence of abundant surface polysaccharide, or glycocalyx, on viridans streptococci has been associated with failure to eradicate the organism from experimental cardiac vegetations during penicillin treatment. The role of glycocalyx in retarding sterilization was tested by in vivo administration of dextranase, an endohydrolase that attacks internally situated alpha (1-6) linkages. Dextranase and penicillin, either singly or in combination, were used to treat experimental endocarditis. After two days of therapy, 100% of animals treated with penicillin or dextranase alone had infected vegetations, whereas only 25% treated with penicillin and dextranase had infected vegetations (P less than .01). After five days of therapy, 100% of the animals treated with penicillin had infected vegetations, versus none that were treated with penicillin and dextranase (P less than .01). We conclude that glycocalyx acts to retard antibiotic activity in vegetations and that partial enzymatic digestion of the glycocalyx facilitates penicillin sterilization of the infected valve.