The present work focuses on in vitro cultures of Ruta montana L. in temporary immersion Plantform^TM bioreactors. The main aim of the study was to evaluate the effects of cultivation time (5 and 6 weeks) and different concentrations (0.1–1.0 mg/L) of plant growth and development regulators (NAA and BAP) on the increase in biomass and the accumulation of secondary metabolites. Consequently, the antioxidant, antibacterial, and antibiofilm potentials of methanol extracts obtained from the in vitro-cultured biomass of R. montana were evaluated. High-performance liquid chromatography analysis was performed to characterize furanocoumarins, furoquinoline alkaloids, phenolic acids, and catechins. The major secondary metabolites in R. montana cultures were coumarins (maximum total content of 1824.3 mg/100 g DM), and the dominant compounds among them were xanthotoxin and bergapten. The maximum content of alkaloids was 561.7 mg/100 g DM. Concerning the antioxidant activity, the extract obtained from the biomass grown on the 0.1/0.1 LS medium variant, with an IC₅₀ 0.90 ± 0.03 mg/mL, showed the best chelating ability among the extracts, while the 0.1/0.1 and 0.5/1.0 LS media variants showed the best antibacterial (MIC range 125–500 µg/mL) and antibiofilm activity against resistant Staphylococcus aureus strains.

Recently, our research team has started a study on Brassica fruticulosa subsp. fruticulosa, an edible plant traditionally used to treat various ailments, little investigated to date. Good in vitro antioxidant properties were highlighted for the leaf hydroalcoholic extract, with the secondary higher than the primary ones. In continuation of the ongoing research, this work was designed to elucidate the antioxidant properties of the phenolic compounds contained in the extract. For this purpose, a phenolic-rich ethyl acetate fraction (Bff-EAF) was obtained from the crude extract by liquid–liquid extraction. The phenolic composition was characterized by HPLC-PDA/ESI-MS analysis and the antioxidant potential was investigated by different in vitro methods. Furthermore, the cytotoxic properties were evaluated by MTT, LDH and ROS determinations on human colorectal epithelial adenocarcinoma cells (CaCo-2) and human normal fibroblasts (HFF-1). Twenty phenolic compounds (flavonoid and phenolic acid derivatives) were identified in Bff-EAF. The fraction exhibited good radical scavenging activity in the DPPH test (IC₅₀ = 0.81 ± 0.02 mg/mL), and moderate reducing power (ASE/mL = 13.10 ± 0.94) and chelating properties (IC₅₀ = 2.27 ± 0.18 mg/mL), contrary to what previously observed for the crude extract. Bff-EAF reduced in a dose-dependent manner CaCo-2 cell proliferation after 72 h of treatment. This effect was accompanied by the destabilization of the cellular redox state due to the antioxidant and pro-oxidant activities displayed by the fraction at lower and higher concentrations. No cytotoxic effect was observed on HFF-1 fibroblasts, used as control cell line.

In continuation of research conducted on species of the spontaneous flora of Sicily (Italy) belonging to the Brassicaceae family, Brassica fruticulosa subsp. fruticulosa was selected. It is an edible species utilized in Sicilian traditional medicine. In this study, for the first time, the phenolic and the volatile compounds and the antioxidant properties of the hydroalcoholic extract obtained from the leaves of B. fruticulosa subsp. fruticulosa were characterized. Through HPLC-PDA/ESI-MS analysis, a total of 22 polyphenolic compounds (20 flavonoids and 2 phenolic acids) were identified, with 3-hydroxiferuloylsophoroside-7-O-glucoside (1.30 mg/g ± 0.01) and kaempferol-3-O-feruloylsophoroside-7-O-glucoside (1.28 mg/g ± 0.01) as the most abundant compounds. Through SPME-GC/MS several volatiles belonging to different chemical classes were characterized, with nitriles and aldehydes accounting for more than 54% of the whole volatile fraction. The extract of B. fruticulosa subsp. fruticulosa showed moderate activity in the DPPH assay (IC₅₀ = 1.65 ± 0.08 mg/mL), weak reducing power (17.47 ± 0.65 ASE/mL), and good chelating properties (IC₅₀ = 0.38 ± 0.02 mg/mL), reaching approximately 90% activity at the highest tested concentration. Lastly, the extract was non-toxic against Artemia salina, indicating its potential safety. According to the findings, it can be stated that B. fruticulosa subsp. fruticulosa represents a new valuable source of bioactive compounds.

Brassica incana subsp. raimondoi is an endemic taxon present in a restricted area located on steep limestone cliffs at an altitude of about 500 m a.s.l. in eastern Sicily. In this research, for the first time, studies on the phytochemical profile, the antioxidant properties in cell-free and cell-based systems, the cytotoxicity on normal and cancer cells by 3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide (MTT) assay, and on Artemia salina Leach, were performed. The total phenolic, flavonoid, and condensed tannin contents of the leaf hydroalcoholic extract were spectrophotometrically determined. Ultra-performance liquid chromatography—tandem mass spectrometer (UPLC-MS/MS) analysis highlighted the presence of several phenolic acids, flavonoids, and carotenoids, while High-Performance Liquid Chromatography with Diode-Array Detection (HPLC-DAD) identified various kaempferol and isorhamnetin derivatives. The extract exhibited different antioxidant properties according to the five in vitro methods used. Cytotoxicity by MTT assay evidenced no impact on normal human fibroblasts (HFF-1) and prostate cancer cells (DU145), and cytotoxicity accompanied by necrotic cell death for colon cancer cells (CaCo-2) and hepatoma cells (HepG2), starting from 100 μg/mL and 500 μg/mL, respectively. No cytotoxic effects were detected by the A. salina lethality bioassay. In the H₂O₂-induced oxidative stress cell model, the extract counteracted cellular reactive oxygen species (ROS) production and preserved non-protein thiol groups (RSH) affected by H₂O₂ exposure in HepG2 cells. Results suggest the potential of B. incana subsp. raimondoi as a source of bioactive molecules.

Ferula L., belonging to the Apiaceae family, is represented by about 170 species predominantly present in areas with a mild–warm–arid climate, including the Mediterranean region, North Africa and Central Asia. Numerous beneficial activities have been reported for this plant in traditional medicine, including antidiabetic, antimicrobial, antiproliferative, anti-dysentery, stomachache with diarrhea and cramps remedies. FER-E was obtained from the plant F. communis, and precisely from the root, collected in Sardinia, Italy. A total of 25 g of root was mixed with 125 g of acetone (ratio 1:5, room temperature). The solution was filtered, and the liquid fraction was subjected to high pressure liquid chromatographic separation (HPLC). In particular, 10 mg of dry root extract powder, from F. communis, was dissolved in 10.0 mL of methanol, filtered with a 0.2 µm PTFE filter and subjected to HPLC analysis. The net dry powder yield obtained was 2.2 g. In addition, to reduce the toxicity of FER-E, the component ferulenol was removed. High concentrations of FER-E have demonstrated a toxic effect against breast cancer, with a mechanism independent of the oxidative potential, which is absent in this extract. In fact, some in vitro tests were used and showed little or no oxidizing activity by the extract. In addition, we appreciated less damage on the respective healthy cell lines (breast), assuming that this extract could be used for its potential role against uncontrolled cancer growth. The results of this research have also shown that F. communis extract could be used together with tamoxifen, increasing its effectiveness, and reducing side effects. However, further confirmatory experiments should be carried out.