Usnic Acid and Usnea barbata (L.) F.H. Wigg. Dry Extracts Promote Apoptosis and DNA Damage in Human Blood Cells through Enhancing ROS Levels.

Nowadays, numerous biomedical studies performed on natural compounds and plant extracts aim to obtain highly selective pharmacological activities without unwanted toxic effects. In the big world of medicinal plants, Usnea barbata (L) F.H. Wigg (U. barbata) and usnic acid (UA) are well-known for their therapeutical properties. One of the most studied properties is their cytotoxicity on various tumor cells. This work aims to evaluate their cytotoxic potential on normal blood cells. Three dry U. barbata extracts in various solvents: ethyl acetate (UBEA), acetone (UBA), and ethanol (UBE) were prepared. From UBEA we isolated usnic acid with high purity by semipreparative chromatography. Then, UA, UBA, and UBE dissolved in 1% dimethyl sulfoxide (DMSO) and diluted in four concentrations were tested for their toxicity on human blood cells. The blood samples were collected from a healthy non-smoker donor; the obtained blood cell cultures were treated with the tested samples. After 24 h, the cytotoxic effect was analyzed through the mechanisms that can cause cell death: early and late apoptosis, caspase 3/7 activity, nuclear apoptosis, autophagy, reactive oxygen species (ROS) level and DNA damage. Generally, the cytotoxic effect was directly proportional to the increase of concentrations, usnic acid inducing the most significant response. At high concentrations, usnic acid and U. barbata extracts induced apoptosis and DNA damage in human blood cells, increasing ROS levels. Our study reveals the importance of prior natural products toxicity evaluation on normal cells to anticipate their limits and benefits as potential anticancer drugs.

In Vitro Anticancer Activity and Oxidative Stress Biomarkers Status Determined by Usnea barbata (L.) F.H. Wigg. Dry Extracts.

Lichens represent an important resource for common traditional medicines due to their numerous metabolites that can exert diverse pharmacological activities including anticancer effects. To find new anticancer compounds with fewer side effects and low tumor resistance, a bioprospective study of Usnea barbata (L.) F.H. Wigg. (U. barbata), a lichen from the Calimani Mountains (Suceava county, Romania) was performed. The aim of this research was to investigate the anticancer potential, morphologic changes, wound healing property, clonogenesis, and oxidative stress biomarker status of four extracts of U. barbata in different solvents (methanol, ethanol, acetone, and ethyl acetate), and also of usnic acid (UA) as a positive control on the CAL-27 (ATCC® CRL-2095™) oral squamous carcinoma (OSCC) cell line and V79 (ATCC® CCL-93™) lung fibroblasts as normal cells. Using the MTT assay and according to IC50 values, it was found that the most potent anticancer property was displayed by acetone and ethyl acetate extracts. All U. barbata extracts determined morphological modifications (losing adhesion capacity, membrane shrinkage, formation of abnormal cellular wrinkles, and vacuolization) with higher intensity in tumor cells than in normal ones. The most intense anti-migration effect was established in the acetone extract treatment. The clonogenic assay showed that some U. barbata extracts decreased the ability of cancer cells to form colonies compared to untreated cells, suggesting a potential anti-tumorigenic property of the tested extracts. Therefore, all the U. barbata extracts manifest anticancer activity of different intensity, based, at least partially, on an imbalance in antioxidant defense mechanisms, causing oxidative stress.

Antioxidant and Cytotoxic Activities of Usnea barbata (L.) F.H. Wigg. Dry Extracts in Different Solvents.

Lichens represent a significant source of antioxidants due to numerous metabolites that can reduce free radicals. Usnea barbata (L.) F.H. Wigg. has been recognized and used since ancient times for its therapeutic effects, some of which are based on its antioxidant properties. The present study aims to analyze the phytochemical profile and to evaluate the antioxidant and cytotoxic potential of this lichen species. Five dry extracts of U. barbata (UBDE) in different solvents (acetone, ethyl acetate, ethanol, methanol, water) were prepared by refluxing at Soxhlet to achieve these proposed objectives and to identify which solvent is the most effective for the extraction. The usnic acid content (UAC) was quantified by ultra-high performance liquid chromatography (UHPLC). The total polyphenols content (TPC) and tannins content (TC) were evaluated by spectrophotometry, and the total polysaccharides (PSC) were extracted by a gravimetric method. The 2,2-diphenyl-1-picryl-hydrazyl-hydrate (DPPH) free radical method was used to assess the antioxidant activity (AA) and the Brine Shrimp Lethality (BSL) assay was the biotest for cytotoxic activity evaluation. The ethyl acetate extract had the highest usnic acid content, and acetone extract had the highest content of total polyphenols and tannins. The most significant antioxidant effect was reported to methanol extract, and all the extracts proved high cytotoxicity. The water extract has the lowest cytotoxicity because usnic acid is slightly soluble in this solvent, and it was not found at UHPLC analysis. All extracts recorded a moderate correlation between the content of usnic acid, polyphenols, tannins, and AA; furthermore, it has been observed that the cytotoxicity varies inversely with the antioxidant effect.

Exploring New Antioxidant and Mineral Compounds from Nymphaea alba Wild-Grown in Danube Delta Biosphere.

Nymphaea alba is an aquatic flowering plant from the Nymphaeaceae family that has been used for hundreds of years in traditional herbal medicine. The plant is characterized by different phytochemicals, depending on the geographical location. Herein, we have carried out, for the first time, the separation and HPLC-MS/MS identification of some antioxidant compounds, such as polyphenols and flavonoids from N. alba extracts from the Danube Delta Biosphere, and investigated their possible antiradical properties. An ultrasonic method has been exhaustively used for the extraction of the antioxidant compounds from the different anatomic parts of N. alba (fruit, flower, leaf, stem, and root). The extracts that were obtained using ultrasound irradiation showed a large polyphenol (19.42 mg EqGA/100 mg extract) and flavonoid (0.97 mg EqQ/100 mg extract) content. The fruit and flower extracts showed the highest antioxidant activity index (AAI). Among the 27 phytochemical compounds identified in all of the N. alba extracts, rutin and p-coumaric acid were found as the major components. The content of macroelements and microelements in N. alba extracts were compared, and it was found that their concentrations depend on the different anatomic parts of the plant. This research contributes to the study of Nymphaeaceae family, being the first exhaustive phytochemical study of N. alba from a wild population in Romania.