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The aim of this work was to determine the effect of standardized aqueous extracts from Epilobium angustifolium L., E. parviflorum Schreb. and E. hirsutum L. herbs on the apoptosis of hormone-dependent prostate cancer cells (LNCaP).The extracts were characterized using high-performance liquid chromatography−diode array detector coupled with mass spectrometry (HPLC-DAD-MS/MS). Apoptosis in the cells was analysed using Annexin V–fluorescein isothiocyanate, and mitochondrial potential, Δψm, using JC-1 by flow cytometry. Caspase-3 activity was determined by enzyme-linked immunosorbent assay.Using the HPLC-DAD-MS/MS method, 38 constituents were characterized. Extracts contained significant amounts of oenothein B as well as flavonoids and phenolic acids. Exposure of LNCaP cells to the extracts (20, 50 and 70 μg/ml) resulted in a significant increase in the level apoptotic cells, from 2.86 ± 0.5% (for untreated cells) up to 86.6 ± 1.0%. All extracts significantly decreased the mitochondrial potential, Δψm, resulting in an increase in the activity of caspase-3 from 0.3 ± 0.07 ng/mg of protein (for untreated cells) up to 1.26 ± 0.32 ng/mg of protein.This study demonstrated that Epilobium extracts are active against LNCaP prostate cancer cells and that their apoptotic activity is related to activation of the mitochondrial pathway. The high oenothein B content may influence the biological activity of these plant materials.