Bioguided fractionation of procyanidin B2 as potent anti coxsackie virus B and Herpes simplex from cypress (Cupressus sempervirens L.)

The methanolic extracts of cypress (Cupressus sempervirens L.) collected at three phenological stages were evaluated for their cytotoxicity on Vero cells by MTT test as well as on Herpes simplex (HSV-2) and coxsackie (CVB-3) viruses by plaque reduction assay. The methanolic extract exhibited the highest cytotoxicity against HSV-2 (IC50 = 20.40 mu g/mL) and CVB-3 (IC50 = 47.50 mu g/mL) at the flowering stage. This extract also exhibited a virucidal action both during the entry of viruses and the release of newly formed virions. The methanolic extract bioguided purification showed that the ethyl-acetate fraction was responsible for virucidal activity. This fraction was endowed with more important selectivity index of 8.15 for HSV-2 and 4.40 for CVB-3. The ethyl acetate fraction was subjected to thin layer chromatography fractionation and identification by HPLC-DAD-ESI-MSn. Results showed that the condensed tannin procyanidin B2 was identified for the first time responsible of the antiviral activity of cypress.