Polyprenylated Acylphloroglucinols from Hypericum rochelii and Hypericum olympicum—Cytotoxic Effects on Non-Tumorigenic Cell Lines and Antibacterial Potential

<b>Objectives</b>: Research on the antimicrobial effect of <i>Hypericum</i> plant constituents is very rarely accompanied by studies of the cytotoxic effect on cell lines. In the current study, besides microbiological tests, an investigation of the cytotoxicity of <i>Hypericum</i> active ingredients on five non-tumorigenic cell lines, as well as research into the effect on other factors of host homeostasis, was performed. <b>Methods</b>: The main methods applied included an MTT assay, the broth microdilution method (BMD), real-time PCR, live cell imaging with Hoechst dye, Western blot, an enzyme-linked immunosorbent assay (ELISA), and skin irritation test on rabbits. <b>Results</b>: The mean inhibitory concentrations (IC₅₀) of six selected agents-previously phytochemically characterized extracts and compounds-ranged from 0.63 to 48 µg/mL. Due to their strong antimicrobial effect and favorable cytotoxic profile, the extract RochC from <i>Hypericum rochelii</i> and the compound olympiforin B from <i>Hypericum olympicum</i> were selected for subsequent studies at their previously determined minimum inhibitory concentrations (MICs) against <i>Staphylococcus aureus</i>-0.625 and 1 µg/mL, respectively. These doses were lower than their IC₅₀ values and the maximum tolerated concentrations (MTCs), according to ISO 10993-5, Annex C, for fibroblast cells, including a human gingival line. The MIC values of RochC and Olympiforin B against the cariogenic <i>Streptococcus mutans</i> were 6 and 3 µg/mL, respectively, values lower than the IC₅₀ values of the gingival cells. Olympiforin B inhibited the gene expression of the staphylococcal biofilm-related genes <i>ica</i>A and <i>ica</i>D, while RochC induced <i>ica</i>A and had a versatile effect on <i>ica</i>D. The MIC values for lactobacilli strains were higher than for <i>S. aureus</i>. The phytoconstituents did not cause cytopathic effects or apoptosis in CCL-1 fibroblasts at 2 × MIC. However, the agents at 1 × MIC significantly induced Atg5 and Atg7, proteins related to autophagy. Cytochrome P450 was not induced in liver cells, with the exception of a dose of 2 × MIC of RochC. The agents did not irritate rabbit skin in vivo at a dose of even 10 × MIC. <b>Conclusions</b>: The extract and compound have potential for further pharmacological development.