•Extracts and essential oils of four different hops varieties from the territory of Slovenia were obtained.•Monoterpnenes and sesquiterpene hydrocarbons were dominant components in isolated essential oils.•All obtained extracts and oils were characterized by strong antimicrobial activity.•All extracts and essential oils were selective towards normal cells. Hydrophilic, lipophilic extracts and essential oil of four hops varieties from Slovenia were examined in this study. Lipophilic extracts were obtained by supercritical extraction (SFE), while for hydrophilic extracts ultrasound and microwave extraction were employed. Essential oils were isolated by the hydrodistillation process. The lipophilic composition of essential oils and lipophilic extracts was determined by GC–MS analysis. Monoterpenes and sesquiterpene hydrocarbons were the most abundant class of compounds in oils (62.27–79.65 %), with myrcene being the most abundant constituent. Limonene and trans-caryophyllene were two terpenes determined in all essential oils while only trans-caryophyllene was detected in SFE samples. Antioxidant, antimicrobial, and cytotoxic activity, determined by applying in vitro assays, was more influenced by extraction technique than by varieties. Molecular docking was carried out to gain insight into the potential cancer protein targets BCL-2 and MMP9, whereby humulene epoxide II displayed good binding configuration within the cavities of the two proteins.