Two protein bound polysaccharides, a neutral protein bound polysaccharide (NPBP) and an acidic protein bound polysaccharide (APBP), were isolated from water soluble substances of Ganoderma lucidum by EtOH precipitation and DEAE-cellulose column chromatography. Their antiviral activities against herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2) were then investigated by plaque reduction assay. APBP exhibited more potent HSV-1 and HSV-2 antiviral activity than NPBP with 50% effective concentration (EC(50)) of 300-520 microg/ml. In order to examine the possible mode of the antiviral activity of APBP its virucidal effect, antiviral activity in preincubation, attachment and penetration assay were tested with HSV-1 and HSV-2. APBP was found to have a direct virucidal effect on HSV-1 and HSV-2. APBP did not induce IFN or IFN-like materials in vitro and is not expected to induce a change from a normal state to an antiviral state. APBP in concentrations of 100 and 90 microg/ml inhibited up to 50% of the attachment of HSV-1 and HSV-2 to Vero cells and was also found to prevent penetration of both types of HSV into Vero cells. These results show that the antiherpetic activity of APBP seems to be related to its binding with HSV-specific glycoproteins responsible for the attachment and penetration, and APBP impedes the complex interactions of viruses with cell plasma membranes.