Sea cucumber has antiviral activities against various viruses including herpes simplex virus type 1 (HSV-1). The purpose of the current study was to determine the chemical profile and inhibitory effects of tegument ethanolic extract of Holothuria parva on HSV-1 infection and to elucidate the mechanism of antiviral action of this marine invertebrate. Cytotoxic activity of the extract on Vero cell line was determined using the methyl thiazolyl tetrazolium (MTT) method. The different components in H. parva were determined by GC-MS analysis. To assess the antiviral activity of the extract, MTT and 50% tissue culture infective dose (TCID50) were applied. Finally, computational molecular docking was performed to screen the potential binding ability of extract contents with HSV-1 surface glycoproteins and host cell surface receptors. Using MTT assay, the non-cytotoxic concentration of the extract was measured 46.5 μg/mL. Octadecanoic acid 2-hydroxy-1-(hydroxymethyl) ethyl ester and 2′,6′-acetoxylidide were two major constituents in the H. parva extract. Pre-treatment of HSV-1 with the ethanolic extract of H. parva led to a 2.1 log10 TCID50 reduction in virus titers when compared to the control group (P = 0.002). The log10 TCID50 reductions relative to the control group for co-penetration and post-penetration assays were 1.5 (P = 0.009) and 0.7 (P = 0.09), respectively. The tegument ethanolic extract of H. parva has significant antiviral properties against HSV-1. Docking analysis demonstrated that compounds of the extract [lidocaine and 2-hydroxy-1-(hydroxymethyl) ethyl ester octadecanoic acid] may cover similarly both virus and host cells binding domains leading to interference in virus attachment to cell receptors.