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Citrus fruits exert various beneficial health effects due to the large amount of polyphenols they contain. Citrus peels, often considered food waste, contain several health-promoting polyphenols. Among these, flavonoids have long been quantified through colorimetric assays which, if not adequately applied, can lead to conflicting results. Flavonoids possess strong antioxidant properties and can decrease circulating free radicals, thereby reducing oxidative stress phenomena. Quantifying flavonoids and properly estimating their antioxidant capacity allows us to predict plausible beneficial effects of citrus fruits on human health. The aim of this research was to analyze the advantageous phenolic compounds found in the peels of citrus fruits commonly found in the Mediterranean region. The objective was to measure their antioxidant capacity and ability to neutralize free radicals. To achieve this purpose, UV-visible spectrophotometric analyses, liquid chromatography (LC) and Electron Paramagnetic Spectroscopy (EPR) were utilized and compared, finally suggesting an innovative approach for assessing the overall flavonoid content by the nitrite-aluminum assay. HPLC data demonstrated that hesperidin was the most abundant flavonoid in all peel extracts except for orange peels, in which naringin was the predominant flavonoid. The total flavonoid content was greater than 1.3 mg/mL in all extracts, with tangerine and orange yielding the best results. Citrus peel polyphenols exerted strong antioxidant and free radical scavenging effects, inhibiting up to 75% of the free radicals used as reference in the EPR analyses.

Salvia rosmarinus Spenn. is a native Mediterranean shrub belonging to the Lamiaceae family and is well-known as a flavoring and spicing agent. In addition to its classical use, it has drawn attention because its biological activities, due particularly to the presence of polyphenols, including carnosic acid and rosmarinic acid, and phenolic diterpenes as carnosol. In this study, the aerial part of rosemary was extracted with a hydroalcoholic solution through maceration, followed by ultrasound sonication, to obtain a terpenoids-rich Salvia rosmarinus extract (TRSrE) and a polyphenols-rich Salvia rosmarinus extract (PRSrE). After phytochemical characterization, both extracts were investigated for their antioxidant activity through a classical assay and with electron paramagnetic resonance (EPR) for their DPPH and hydroxyl radicals scavenging. Finally, their potential beneficial effects to reduce lipid accumulation in an in vitro model of NAFLD were evaluated.

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.

Olea europaea L. is a plant belonging to the Oleaceae family, widely grown around the Mediterranean Basin and its leaves are a source of phenolic compounds with antioxidant and anti-inflammatory capacity. Among these, oleuropein and luteolin-7-O-glucoside represent two major polyphenolic compounds in olive-leaf extract. Herein, a polystyrene resin was used to recover the polyphenolic fraction from the acetone-water leaf extract from Nocellara del Belice cultivar, which showed the higher level of analysed bioactive compounds, compared to Carolea cultivar. The antioxidant activity of the extract concentrated in phenolic compounds (OLECp) was evaluated through a classical assay and electron paramagnetic resonance (EPR) for DPPH and hydroxyl radicals scavenging. Thus, the anti-inflammatory activity and the potential beneficial effects in reducing lipid accumulation in an in vitro model of NAFLD using McA-RH7777 cells exposed to oleic acid (OA) were evaluated. Nile Red and Oil Red O have been used to stain the lipid accumulation, while the inflammatory status was assessed by Cytokines Bioplex Assay. OLECp (TPC: 92.93 ± 9.35 mg GAE/g, TFC: 728.12 ± 16.04 mg RE/g; 1 g of extract contains 315.250 mg of oleuropein and 17.44 mg of luteolin-7-O-glucoside) exerted a good radical scavenging capability (IC50: 2.30 ± 0.18 mg/mL) with a neutralizing power against DPPH and hydroxyl radicals, as confirmed by the decreased signal area of the EPR spectra. Moreover, OLECp at concentration of 25, 50 and 100 μg/mL counteracted the intracellular inflammatory status, as result of decreased intracellular lipid content. Our results highlighted the multiple properties and applications of an O. europaea extract concentrated in polyphenols, and the possibility to formulate novel nutraceuticals with antioxidant properties, destined to ameliorate human health.

Ferula communis L. (F. communis) belongs to the Apiaceae family and is a herbaceous plant with various pharmaceutical properties, due to the different contents of bioactive compounds extracted mainly from its roots, as well as its leaves and rhizome. To date, this plant extract has demonstrated estrogenic, anti-inflammatory, antiproliferative, cytotoxic, antimicrobial and anti-neoplastic properties. Its estrogenic activity is justified by the presence of ferutinin, an ester of a sesquiterpenic alcohol that acts as an agonist for estrogen receptors, with a chemical formula equal to C22H3O4. The component present in F. communis responsible for the toxicity of the plant is ferulenol, a prenylated coumarin with the chemical formula C24H30O3. This compound is capable of inducing mortality via its strong anti-coagulant properties, leading to a lethal hemorrhagic syndrome, ferulosis, in animals that feed on a chemotype of F. communis containing a high amount of ferulenol. The removal of the component ferulenol makes extracts of Ferula non-toxic. In fact, the remaining prenylated coumarins are not present in concentrations sufficient to induce toxicity. The intake of high concentrations of the extract of this plant leads a double dose-dependent effect that is typical of sesquiterpenes such as ferutinin. Here, we assessed the cytotoxicity and the estrogenic properties of the F. communis phytocomplex obtained through extraction using a mixture of acetone and water. Among the active constituents of F. communis, the identification of ferutinin and ferulenol was performed using HPLC. The effects of the extract were evaluated, following the removal of ferulenol, on three cell lines: human breast cancer MCF-7, human cervical cancer HeLa and human osteoblastic sarcoma Saos-2. The choice of these cell lines was justified by the need to mimic certain processes which may occur in vivo and which are estrogen-dependent. The obtained results demonstrated that F. communis extract, in addition to possessing an estrogenic-like property, showed a dose-dependent effect. Low concentrations (0.1–0.8 μM) demonstrated a hyperproliferative effect, whereas higher concentrations (1.6–50 μM) were toxic. Therefore, this extract could be an excellent candidate to make up for a reduction or lack of estrogen.