Insights into the phytochemical composition and antioxidant potential of the Tunisian Ceratonia siliqua L.

Carob (Ceratonia siliqua L.) corresponds to an evergreen leguminous tree (Fabaceae family). The high phenolic content of numerous parts of carob has been deeply associated with several nutritional and functional benefits. The aim of this study was to investigate the physicochemical properties of ground carob pods and seeds, the effect of different extraction procedures as well a comprehensive phytochemical characterization of hydro-methanolic extracts (80/20 v/v) of pods and seeds by HPLC-DAD ESI-Q-TOF-MS/MS. Additionally, their antioxidant activity was evaluated using in vitro assays. The results showed thatthe dry matter (DM) values were 88.09% for pods and 89.10% for seeds, protein contents were 0.41 g/100 g DM for pods and 0.88 g/100 g DM for seedsand total sugars contents were 0.35 g/100 DM for pods and 26.70 g/100 g DM for seeds. Furthermore, the oil holding capacities (OHC) were 10.43 g/g for pods and 7.53 g/g for seeds, while the water holding capacities were 8.46 g/g for pods and 2.59 g/g for seeds.The hydro-methanolic extracts of both pods and seeds showed the presence of 53 secondary bioactive metabolites belonging to various classes(flavonoids, phenolic acids, tannins and non-phenolic compounds). The antioxidant activities were evidenced in DPPH (22.24 mg/ml for pods and 26.37 mg/ml for seeds), ABTS (198.50 mmol Eq Trolox/100 g for pods and 201.04 mmol Eq Trolox/100 g for seeds) and FRAP (0.39 mmol Eq Trolox/100 g for pods and 0.53 mmol Eq Trolox/100 g for seeds).Moreover,high significant (p ≤ 0.01) correlation coefficients were found between the antioxidant activity estimated by the DPPH method and total phenols (r = 0.943), orthodiphenols (r = 0.996), flavonoids (r = 0.880) and flavonols (r = 0.982). Nevertheless, lower correlations were detected with ABTS and FRAP methods.These results demonstrated that carob parts displayed an interesting potential that can be of interest for further valorizations as a natural antioxidant with multiple applications, namely functional food ingredients or prevention of many health problems.

Untargeted Phytochemical Profiling, Antioxidant, and Antimicrobial Activities of a Tunisian Capsicum annuum Cultivar.

Peppers are among the spices possessing a wide plethora of biological properties due to their excellent supply of health-related metabolites. Capsicum annuum L. (Solanaceae) is cultivated throughout Tunisia, and there is a shortage of information on the identification of the secondary metabolites in the seeds of this species as well as on their biological activities. In the present work, we intended to undertake a chemical characterization of the bioactive compounds from the hydro-methanolic seed extract of C. annuum as well as an evaluation of its broad spectrum of antimicrobial and antioxidant activities. The chemical profile was evaluated by RP-HPLC-DAD-QTOF-MS/MS, whereas the total phenol and flavonoid content, antioxidant, and antimicrobial activities were determined in in vitro assays. In this work, 45 compounds belonging to various phytochemical classes, such as organic acids (2), phenolic compounds (4 phenolic acids and 5 flavonoids), capsaicinoids (3), capsianosides (5), fatty acids (13), amino acids (1), sphingolipids (10), and steroids (2) were identified in the hydro-methanolic seed extract of C. annuum. The phenolic and flavonoid content (193.7 mg GAE/g DW and 25.1 mg QE/g DW, respectively) of the C. annuum extract correlated with the high antiradical activity (IC50 = 45.0 µg/mL), reducing power (EC50 = 61.3 µg/mL) and chelating power (IC50 = 79.0 µg/mL) activities. The hydro-methanolic seed extract showed an important antimicrobial activity against seven bacterial and four fungal strains. In fact, the inhibition zones (IZs) for bacteria ranged from 9.00 ± 1.00 mm to 12.00 ± 0.00 mm; for fungi, the IZs ranged from 12.66 ± 0.57 mm to 13.66 ± 0.57 mm. The minimal inhibition concentration and minimal bactericidal concentration values showed that the extract was more effective against fungi than bacteria.

Chemical Profile and Bioactivity Evaluation of Salvia Species from Eastern Europe.

The Salvia genus comprises about 1000 species endowed with medicinal, aromatic, cosmetic, and ornamental applications. Even though the genus is one of the most-studied taxa of the Lamiaceae family, data on the chemical composition and biological properties of certain locally used Salvia species are still scarce. The present work aimed to evaluate the phytochemical profile and antimicrobial, antioxidant, and cytotoxic potential of ten Salvia species that grow in Eastern Europe (e.g., the Republic of Moldova). LC-HRMS/MS metabolite profiling allowed for the annotation of 15 phenolic and organic acids, 18 flavonoids, 19 diterpenes, 5 sesterpenes, and 2 triterpenes. Multivariate analysis (e.g., principal component analysis, hierarchical cluster analysis) revealed that S. austriaca, S. nutans, and S. officinalis formed individual clusters, whereas the remaining species had a similar composition. S. officinalis showed the highest activity against Staphylococcus aureus and Streptococcus pneumoniae (MIC = 0.625 mg/mL). As evaluated in DPPH, ABTS, and FRAP assays, S. officinalis was one of the most potent radical scavenging and metal-reducing agents (CE50 values of 25.33, 8.13, and 21.01 µg/mL, respectively), followed by S. verticillata, S. sclarea, S. kopetdaghensis, S. aethiopis, and S. tesquicola. Pearson correlation analysis revealed strong correlations with rosmarinic acid, luteolin-O-glucuronide, and hydroxybenzoic acid. When the cytotoxic activity was evaluated in human breast carcinoma MCF-7 and MDA-MB-231 cells, no significant reduction in cell viability was observed over the concentrations ranging from 25 and 100 µg/mL. The results confirm the potential use of understudied Salvia species as promising sources of antioxidant compounds for developing novel pharmaceutical, nutraceutical, or cosmeceutical products.

Cannabidiol and Minor Phytocannabinoids: A Preliminary Study to Assess Their Anti-Melanoma, Anti-Melanogenic, and Anti-Tyrosinase Properties.

Currently, there is an increased interest from both scientists and consumers in the application of cannabis/hemp/phytocannabinoids in skin-related disorders. However, most previous investigations assessed the pharmacological properties of hemp extracts, cannabidiol (CBD), or tetrahydrocannabinol (THC), with very few studies focusing on minor phytocannabinoids from hemp. In this context, the current work explored the in vitro anti-melanoma, anti-melanogenic, and anti-tyrosinase effects of cannabidiol (CBD) and three minor phytocannabinoids, namely cannabigerol (CBG), cannabinol (CBN), and cannabichromene (CBC). Among the tested human malignant melanoma cells (A375, SH4, and G361), only A375 cells were highly susceptible to the 48 h treatment with the four phytocannabinoids (IC50 values between 12.02 and 25.13 µg/mL). When melanogenesis was induced in murine melanoma B16F10 cells by α-melanocyte stimulating hormone (αMSH), CBD, CBG, and CBN significantly decreased the extracellular (29.76-45.14% of αMSH+ cells) and intracellular (60.59-67.87% of αMSH+ cells) melanin content at 5 µg/mL. Lastly, CBN (50-200 µg/mL) inhibited both mushroom and murine tyrosinase, whereas CBG (50-200 µg/mL) and CBC (100-200 µg/mL) down-regulated only the mushroom tyrosinase activity; in contrast, CBD was practically inactive. The current data show that tyrosinase inhibition might not be responsible for reducing the melanin biosynthesis in α-MSH-treated B16F10 cells. By evaluating for the first time the preliminary anti-melanoma, anti-melanogenic, and anti-tyrosinase properties of CBN and CBC and confirming similar effects for CBD and CBG, this study can expand the utilization of CBD and, in particular, of minor phytocannabinoids to novel cosmeceutical products for skin care.

Silver Nanoparticles Synthesized from Abies alba and Pinus sylvestris Bark Extracts: Characterization, Antioxidant, Cytotoxic, and Antibacterial Effects.

In recent years, phytofunctionalized AgNPs have attracted great interest due to their remarkable biological activities. In the present study, AgNPs were synthesized using Abies alba and Pinus sylvestris bark extracts. The chemical profile of these bark extracts was analyzed by LC-HRMS/MS. As a first step, the synthesis parameters (pH, AgNO3 concentration, ratio of bark extract and AgNO3, temperature, and reaction time) were optimized. The synthesized AgNPs were characterized by ATR-FTIR spectroscopy, DLS, SEM, EDX, and TEM. Their antioxidant, cytotoxic, and antibacterial properties were evaluated by the DPPH, ABTS, MTT, and broth microdilution assays, respectively. Abies alba and Pinus sylvestris bark extract-derived AgNPs were well-dispersed, spherical, small (average particle size of 9.92 and 24.49 nm, respectively), stable (zeta potential values of -10.9 and -10.8 mV, respectively), and cytotoxic to A-375 human malignant melanoma cells (IC50 = 2.40 ± 0.21 and 6.02 ± 0.61 µg/mL, respectively). The phytosynthesized AgNPs also showed antioxidant and antibacterial effects.

Value-Added Compounds with Antimicrobial, Antioxidant, and Enzyme-Inhibitory Effects from Post-Distillation and Post-Supercritical CO2 Extraction By-Products of Rosemary.

Hydrodistillation is the main technique to obtain essential oils from rosemary for the aroma industry. However, this technique is wasteful, producing numerous by-products (residual water, spent materials) that are usually discarded in the environment. Supercritical CO2 (SC-CO2) extraction is considered an alternative greener technology for producing aroma compounds. However, there have been no discussions about the spent plant material leftover. Therefore, this work investigated the chemical profile (GC-MS, LC-HRMS/MS) and multi-biological activity (antimicrobial, antioxidant, enzyme inhibitory) of several raw rosemary materials (essential oil, SC-CO2 extracts, solvent extracts) and by-products/waste materials (post-distillation residual water, spent plant material extracts, and post-supercritical CO2 spent plant material extracts). More than 55 volatile organic compounds (e.g., pinene, eucalyptol, borneol, camphor, caryophyllene, etc.) were identified in the rosemary essential oil and SC-CO2 extracts. The LC-HRMS/MS profiling of the solvent extracts revealed around 25 specialized metabolites (e.g., caffeic acid, rosmarinic acid, salvianolic acids, luteolin derivatives, rosmanol derivatives, carnosol derivatives, etc.). Minimum inhibitory concentrations of 15.6-62.5 mg/L were obtained for some rosemary extracts against Micrococcus luteus, Bacilus cereus, or Staphylococcus aureus MRSA. Evaluated in six different in vitro tests, the antioxidant potential revealed strong activity for the polyphenol-containing extracts. In contrast, the terpene-rich extracts were more potent in inhibiting various key enzymes (e.g., acetylcholinesterase, butyrylcholinesterase, tyrosinase, amylase, and glucosidase). The current work brings new insightful contributions to the continuously developing body of knowledge about the valorization of rosemary by-products as a low-cost source of high-added-value constituents in the food, pharmaceutical, and cosmeceutical industries.

Nonlinear liquid-liquid chromatography: Beyond a constant distribution coefficient.

Liquid-liquid chromatography (LLC) is a technique in which the separation of mixture components is achieved due to their different distribution between the two phases of a pre-equilibrated biphasic solvent system. In this work, the LLC operation in the nonlinear range of the distribution isotherm was systematically examined for the first time. The influence of the feed concentration on the elution profiles of a model component (cannabidiol, CBD) was studied in three LLC units of different types and sizes ranging from ∼20 mL to ∼2 L. A series of pulse injections with CBD concentrations varying from 1 to 300 mg/mL was performed with n-hexane/methanol/water 5/4/1 (v/v/v) in descending mode (lower phase as the mobile phase). The elution profiles were simulated using the equilibrium-cell model and an anti-Langmuir-like equation for describing the CBD distribution equilibria. The distribution equilibria equation parameters were fitted to the CBD elution profiles using the peak fitting method. The model was validated and provided good predictions of the CBD elution profiles in the entire concentration range for all three LLC units.

Influence of pyrrolizidine alkaloids depletion upon the biological activity of Symphytum officinale L. extracts.

ETHNOPHARMACOLOGICAL RELEVANCE: Comfrey (Symphytum officinale L., Boraginaceae) root preparations are used as both traditional remedies and therapeutic agents in treating pain and inflammation associated with joint, bone, and muscle ailments. Even though numerous phytochemicals contribute to the beneficial effects of comfrey, the presence of toxic pyrrolizidine alkaloids (PAs) overshadows its uses. AIM OF THE STUDY: In this work, different PA-/mucilage-depleted/undepleted comfrey root extracts were subjected to detailed phytochemical characterization and biological evaluation. MATERIALS AND METHODS: The phytochemical profiling was performed by LC-HRMS/MS. The quantification of PAs and major phenolic compounds was carried out by LC-MS/MS and LC-DAD. Antioxidant and enzyme inhibitory activity was determined by in vitro free radical scavenging, ion reducing, metal chelating, cholinesterase, tyrosinase, amylase, and glucosidase assays. Using an ex vivo model of LPS-stimulated neutrophils, their viability (as measured by flow cytometry) and the release of IL-1ß, IL-8, and TNF-α were determined (ELISA assay). RESULTS: 12 phenolic acids, six PAs, three organic acids, two fatty acids, and two sugars were identified in the obtained comfrey extracts. The PA-depleted materials contained PAs levels below 2 ppm, whereas the removal of mucilage increased the content of rosmarinic acid, globoidnan A, globoidnan B, and rabdosiin. PA-depletion did not significantly affect the antioxidant potential. However, the radical scavenging and metal reducing properties were higher in the mucilage-depleted extracts. Neither PA-depletion nor mucilage-depletion had considerable effects on the in vitro inhibitory activity of cholinesterases, tyrosinase, amylase, and glucosidase or release of ex vivo pro-inflammatory cytokines (e.g., IL-1ß, IL-8, and TNF-α) in LPS-stimulated neutrophils. CONCLUSIONS: In light of their superior safety profiles, PA-depleted comfrey extracts can be utilized further in cosmetic and pharmaceutical products.

Exploring the Artemisia Genus: An Insight into the Phytochemical and Multi-Biological Potential of A. campestris subsp. lednicensis (Spreng.) Greuter & Raab-Straube.

The Artemisia L. genus includes over five hundred species with great economic and medicinal properties. Our study aimed to provide a comprehensive metabolite and bioactivity profile of Artemisia campestris subsp. lednicensis (Spreng.) Greuter & Raab-Straube collected from north-eastern Romania. Liquid chromatography with tandem high-resolution mass spectrometry (LC-HRMS/MS) analysis of different polarity extracts obtained from the aerial parts led to the identification of twelve flavonoids, three phenolic acids, two sesquiterpene lactones, two fatty acids, one coumarin, and one lignan. The antioxidant and enzyme inhibitory properties were shown in the DPPH (0.71−213.68 mg TE/g) and ABTS (20.57−356.35 mg TE/g) radical scavenging, CUPRAC (38.56−311.21 mg TE/g), FRAP (121.68−202.34 mg TE/g), chelating (12.88−22.25 mg EDTAE/g), phosphomolybdenum (0.92−2.11 mmol TE/g), anti-acetylcholinesterase (0.15−3.64 mg GALAE/g), anti-butyrylcholinesterase (0−3.18 mg GALAE/g), anti-amylase (0.05−0.38 mmol ACAE/g), anti-glucosidase (0.43−2.21 mmol ACAE/g), and anti-tyrosinase (18.62−48.60 mg KAE/g) assays. At 100 µg/mL, Artemisia extracts downregulated the secretion of tumor necrosis factor (TNF)-α in a lipopolysaccharide (LPS)-stimulated human neutrophil model (29.05−53.08% of LPS+ control). Finally, the Artemisia samples showed moderate to weak activity (minimum inhibitory concentration (MIC) > 625 mg/L) against the seventeen tested microbial strains (bacteria, yeasts, and dermatophytes). Overall, our study shows that A. campestris subsp. lednicensis is a promising source of bioactives with putative use as food, pharmaceutical and cosmetic ingredients.

Evaluation of Anti-Alzheimer Activity of Synthetic Coumarins by Combination of in Vitro and in Silico Approaches.

Series of synthetic coumarin derivatives (1-16) were tested against acetylcholinesterase (AChE) and butyrylcholinesterase (BChE), two enzymes linked to the pathology of Alzheimer's disease (AD). Compound 16 was the most active AChE inhibitor with IC50 32.23±2.91 µM, while the reference (galantamine) had IC50 =1.85±0.12 µM. Compounds 9 (IC50 75.14±1.82 µM), 13 (IC50 =16.14±0.43 µM), were determined to be stronger BChE inhibitors than the reference galantamine (IC50 =93.53±2.23 µM). The IC50 value of compound 16 for BChE inhibition (IC50 =126.56±11.96 µM) was slightly higher than galantamine. The atomic interactions between the ligands and the key amino acids inside the binding cavities were simulated to determine their ligand-binding positions and free energies. The three inhibitory coumarins (9, 13, 16) were next tested for their effects on the genes associated with AD using human neuroblastoma (SH-SY5Y) cell lines. Our data indicate that they could be considered for further evaluation as new anti-Alzheimer drug candidates.

A review on the ethnobotany, phytochemistry, pharmacology and toxicology of butterbur species (Petasites L.).

ETHNOPHARMACOLOGICAL RELEVANCE: Petasites (butterbur, Asteraceae) species have been used since Ancient times in the traditional medicine of Asian and European countries to treat central nervous system (migraine), respiratory (asthma, allergic rhinitis, bronchitis, spastic cough), cardiovascular (hypertension), gastrointestinal (ulcers) and genitourinary (dysmenorrhea) disorders. AIM OF THE REVIEW: This study summarized and discussed the traditional uses, phytochemical, pharmacological and toxicological aspects of Petasites genus. MATERIALS AND METHODS: A systematic search of Petasites in online databases (Scopus, PubMed, ScienceDirect, Google Scholar) was performed, with the aim to find the phytochemical, toxicological and bioactivity studies. The Global Biodiversity Information Facility, Plants of the World Online, World Flora Online and The Plant List databases were used to describe the taxonomy and geographical distribution. RESULTS: The detailed phytochemistry of the potentially active compounds of Petasites genus (e.g. sesquiterpenes, pyrrolizidine alkaloids, polyphenols and essential oils components) was presented. The bioactivity studies (cell-free, cell-based, animal, and clinical) including the traditional uses of Petasites (e.g. anti-spasmolytic, hypotensive, anti-asthmatic activities) were addressed and followed by discussion of the main pharmacokinetical and toxicological issues related to the administration of butterbur-based formulations. CONCLUSIONS: This review provides a complete overview of the Petasites geographical distribution, traditional use, phytochemistry, bioactivity, and toxicity. More than 200 different sesquiterpenes (eremophilanes, furanoeremophilanes, bakkenolides), 50 phenolic compounds (phenolic acids, flavonoids, lignans) and volatile compounds (monoterpenes, sesquiterpenes) have been reported within the genus. Considering the phytochemical complexity and the polypharmacological potential, there is a growing research interest to extend the current therapeutical applications of Petasites preparations (anti-migraine, anti-allergic) to other human ailments, such as central nervous system, cardiovascular, malignant or microbial diseases. This research pathway is extremely important, especially in the recent context of the pandemic situation, when there is an imperious need for novel drug candidates.

Profiling the annual change of the neurobiological and antioxidant effects of five Origanum species in correlation with their phytochemical composition.

The ethanol extracts of five Origanum species: O. majorana L., O. onites L., O. syriacum L., O. vulgare subsp. hirtum (Link) Ietsw., and O. vulgare subsp. viride (Boiss.) Hayek, collected annually (each month), were investigated for their cholinesterase inhibition and antioxidant effects. The phytochemical composition of a total of 60 extracts was assessed by HPLC-DAD-ESI/HRMS, revealing the presence of a total of 73 compounds. Possible correlation between the bioactivity and metabolite profiles during 12 months was monitored. Acetylcholinesterase (AChE) inhibitory activity was found to be the highest between April and November (50.29-75.95%, 200 µg/mL), while the highest inhibition towards butyrylcholinesterase (BChE) was observed for the extracts between April and October (71.68-88.97%). Aromadendrin showed good correlation with anti-AChE, anti-BChE and reducing power activities. Furthermore, molecular docking data with aromadendrin, caffeoylarbutin and eriodictyol indicated that caffeoylarbutin had the lowest binding energy against both enzymes.

Liquid-Liquid Chromatography Separation of Guaiane-Type Sesquiterpene Lactones from Ferula penninervis Regel & Schmalh. and Evaluation of Their In Vitro Cytotoxic and Melanin Inhibitory Potential.

Ferula penninervis Regel & Schmalh. is a perennial plant used in Kazakh traditional folk medicine to treat epilepsy, neurosis, rheumatism, gastroduodenal ulcers, dyspepsia, wounds, abscesses or tumors. The aim of this work was to isolate series of sesquiterpene lactones from a crude methanolic root extract and investigate their in vitro cytotoxic potential against androgen-dependent prostate cancer LNCaP and epithelial prostate PNT2 cells, as well as to evaluate their melanin production inhibitory effects in murine melanoma B16F10 cells stimulated with α-melanocyte-stimulating hormone (αMSH). Two new (penninervin P and penninervin Q) and five known (olgin, laferin, olgoferin, oferin and daucoguainolactone F) guaiane-type sesquiterpene lactones were isolated with the use of a simple and fast liquid-liquid chromatography method. Olgin and laferin showed the most promising cytotoxic effects in LNCaP cells (IC50 of 31.03 and 23.26 µg/mL, respectively). Additionally, olgin, laferin, olgoferin, and oferin (10 µg/mL) potently impaired melanin release (40.67-65.48% of αMSH + cells) without influencing the viability of B16F10 cells. In summary, our findings might indicate that guaiane-type sesquiterpene lactones from F. penninervis could be regarded as promising candidates for further research in discovering new therapeutic agents with anti-prostate cancer and skin depigmentation properties.

Phyto-Functionalized Silver Nanoparticles Derived from Conifer Bark Extracts and Evaluation of Their Antimicrobial and Cytogenotoxic Effects.

Silver nanoparticles synthesized using plant extracts as reducing and capping agents showed various biological activities. In the present study, colloidal silver nanoparticle solutions were produced from the aqueous extracts of Picea abies and Pinus nigra bark. The phenolic profile of bark extracts was analyzed by liquid chromatography coupled to mass spectrometry. The synthesis of silver nanoparticles was monitored using UV-Vis spectroscopy by measuring the Surface Plasmon Resonance band. Silver nanoparticles were characterized by attenuated total reflection Fourier transform infrared spectroscopy, Raman spectroscopy, dynamic light scattering, scanning electron microscopy, energy dispersive X-ray and transmission electron microscopy analyses. The antimicrobial and cytogenotoxic effects of silver nanoparticles were evaluated by disk diffusion and Allium cepa assays, respectively. Picea abies and Pinus nigra bark extract derived silver nanoparticles were spherical (mean hydrodynamic diameters of 78.48 and 77.66 nm, respectively) and well dispersed, having a narrow particle size distribution (polydispersity index values of 0.334 and 0.224, respectively) and good stability (zeta potential values of -10.8 and -14.6 mV, respectively). Silver nanoparticles showed stronger antibacterial, antifungal, and antimitotic effects than the bark extracts used for their synthesis. Silver nanoparticles obtained in the present study are promising candidates for the development of novel formulations with various therapeutic applications.

Verbascum nigrum: Cytotoxicity Evaluation in A431 Epidermoid Carcinoma Cells and Untargeted LC-HR-MS/MS Metabolite Profiling.

The crude methanolic extract obtained from Verbascum nigrum aerial parts (VNE) and its six fractions (VNF1-VNF6) were initially screened regarding their effects on the viability of immortalized HaCaT keratinocytes and A431 epidermoid carcinoma cells (MTT assay, 24 h). None of the tested samples affected the viability of HaCaT cells in a concentration range of 25-150 µg/mL. VNE and VNF4 exhibited significant cytotoxic effects in A431 cells, with IC50 values of 81.92 and 12.27 µg/mL, respectively; the selectivity index was higher than 10 for VNF4. The untargeted LC/HR-MS/MS metabolite profiling led to the tentative annotation of a total number of 23 compounds. Of these, VNE comprised mainly iridoid glycosides (harpagoside, laterioside, acylated aucubin derivatives), whereas VNF4 showed a high abundance of triterpene saponin glycosides (ilwensisaponins A and C, songarosaponins A and B), constituents known for their selective cytotoxic potential.

Symphytum officinale L.: Liquid-liquid chromatography isolation of caffeic acid oligomers and evaluation of their influence on pro-inflammatory cytokine release in LPS-stimulated neutrophils.

ETHNOPHARMACOLOGICAL RELEVANCE: Symphytum officinale L. (comfrey, Boraginaceae) has been traditionally used for millennia in joint distortions, myalgia, bone fractures and hematomas. However, key activity-determining constituents and molecular mechanisms underlying its use have not been completely elucidated. AIM OF THE STUDY: The objective of this study was to isolate and identify the major compounds from a hydroethanolic root extract of S. officinale and evaluate their antioxidant potential, alongside their effect on the cytokine production of ex vivo stimulated neutrophils, thus providing scientific support for the traditional use of comfrey root. MATERIAL AND METHODS: Four caffeic acid oligomers were isolated from comfrey roots by liquid-liquid chromatography, their structures being established by MS and NMR analyses. In vitro antioxidant evaluation was performed by DPPH and ABTS assays. The cytotoxicity of isolated compounds was established by flow cytometry. The effect on cytokine release, such as interleukin (IL)-1ß, IL-8 and tumor necrosis factor alpha (TNF-α), in lipopolysaccharide (LPS)-stimulated neutrophils was determined by enzyme-linked immunosorbent assay (ELISA). RESULTS: The main constituents found in comfrey root were represented by four caffeic acid oligomers, namely globoidnan B (1), rabdosiin (2), rosmarinic acid (3) and globoidnan A (4). Rabdosiin, globoidnans A and B were isolated for the first time from S. officinale. In the in vitro antioxidant tests, compound 2 was the most active, with EC50 values in DPPH and ABTS assays of 29.14 ± 0.43 and 11.13 ± 0.39, respectively. Neutrophils' viability over the tested concentration domain of 12.5-50 µM was not altered. At 50 µM, all compounds significantly inhibited IL-1ß release, with compound 3 (45.60% release vs. LPS stimulated neutrophils) being the most active, followed by compounds 1 (53.85%), 2 (69.89%) and 4 (60.68%). CONCLUSIONS: The four caffeic acid oligomers reported in S. officinale root may contribute to the overall anti-inflammatory activity for which comfrey preparations are used in traditional medicine.

Vasorelaxant effects of Crataegus pentagyna: Links with arginase inhibition and phenolic profile.

ETHNOPHARMACOLOGICAL RELEVANCE: Crataegus leaves, flowers and fruits have been traditionally used to improve blood circulation, numerous preclinical and clinical studies supporting the cardiovascular benefits of Crataegus preparations. In this respect, there is very limited data on Crataegus pentagyna; in addition, the chemical profile of this species is still incompletely elucidated. AIM OF THE STUDY: The objective of this study was to examine the cardiovascular benefits of Crataegus pentagyna Waldst. et Kit. ex Willd. (small-flowered black hawthorn, Rosaceae) extracts (leaf, flower and fruit ethyl acetate extracts) and the underlying mechanisms. We hypothesized that C. pentagyna extracts might exert vasodilatory effects and inhibit arginase activity due, in large part, to their polyphenolic constituents. MATERIALS AND METHODS: C. pentagyna extracts induced-relaxation and the mechanisms involved were studied ex vivo in isolated aortic rings from Sprague-Dawley rats. The inhibitory effects on bovine liver arginase I were assessed by an in vitro assay. Metabolite profiling of C. pentagyna extracts was performed and the most endothelium- and nitric oxide synthase-dependent; flower extract additionally reduced Ca2+ entry and, to a lesser extent, Ca2+ release from the sarcoplasmic reticulum. C. pentagyna proved to be an important source of arginase inhibitors with potential benefits in endothelial dysfunction that remains to be explored.

6-O-(3″, 4″-di-O-trans-cinnamoyl)-α-l-rhamnopyranosylcatalpol and verbascoside: Cytotoxicity, cell cycle kinetics, apoptosis, and ROS production evaluation in tumor cells.

The aim of this study was to evaluate the impact that 6-O-(3″, 4″-di-O-trans-cinnamoyl)-α- l-rhamnopyranosylcatalpol (Dicinn) and verbascoside (Verb), two compounds simultaneously reported in Verbascum ovalifolium, have on tumor cell viability, apoptosis, cell cycle kinetics, and intracellular reactive oxygen species (ROS) level. At 100 µg/mL and 48 hours incubation time, Dicinn and Verb produced good cytotoxic effects in A549, HT-29, and MCF-7 cells. Dicinn induced cell-cycle arrest at the G0 /G1 phase and apoptosis, whereas Verb increased the population of subG1 cells and cell apoptosis rates. Furthermore, the two compounds exhibited time-dependent ROS generating effects in tumor cells (1-24 hours). Importantly, no cytotoxic effects were induced in nontumor MCF-10A cells by the two compounds up to 100 µg/mL. Overall, the effects exhibited by Verb in tumor cells were more potent, which can be correlated with its structural features, such as the presence of phenolic hydroxyl groups.

Bioactivity of dietary polyphenols: The role of metabolites.

A polyphenol-rich diet protects against chronic pathologies by modulating numerous physiological processes, such as cellular redox potential, enzymatic activity, cell proliferation and signaling transduction pathways. However, polyphenols have a low oral bioavailability mainly due to an extensive biotransformation mediated by phase I and phase II reactions in enterocytes and liver but also by gut microbiota. Despite low oral bioavailability, most polyphenols proved significant biological effects which brought into attention the low bioavailability/high bioactivity paradox. In recent years, polyphenol metabolites have attracted great interest as many of them showed similar or higher intrinsic biological effects in comparison to the parent compounds. There is a huge body of literature reporting on the biological functions of polyphenol metabolites generated by phase I and phase II metabolic reactions and gut microbiota-mediated biotransformation. In this respect, the review highlights the pharmacokinetic fate of the major dietary polyphenols (resveratrol, curcumin, quercetin, rutin, genistein, daidzein, ellagitannins, proanthocyanidins) in order to further address the efficacy of biometabolites as compared to parent molecules. The present work strongly supports the contribution of metabolites to the health benefits of polyphenols, thus offering a better perspective in understanding the role played by dietary polyphenols in human health.

Unravelling the potential of seaweeds from the Black Sea coast of Romania as bioactive compounds sources. Part I: Cystoseira barbata (Stackhouse) C. Agardh.

The Romanian coastlines of the Black Sea have abundant seaweed resources, but little effort has been done to investigate their biological potential. The aim of the present study was to assess the in vitro antioxidant and anti-proliferative effects of Cystoseira barbata (Stackhouse) C. Agardh (Sargassaceae), a brown alga inhabiting the Black Sea coast of Romania. The 70% acetone, methanol and water extracts of C. barbata were evaluated for their total phenolic content, antioxidant activity and anti-proliferative potential against human tumor cell lines (pulmonary A549, colon HT-29, mammary MCF-7) and the non-tumor mammary epithelial MCF-10A cell line. C. barbata 70% acetone extract (CBAE) displayed the highest antioxidant and cytotoxic activities. The mechanism of CBAE anti-proliferative activity involved initially increased intracellular ROS accumulation, followed by increased DNA content in the subG1 phase and DNA fragmentation leading to excessive apoptosis. Thus, our study provides a theoretical basis for the use of CBAE as a tumor preventive agent. Furthermore, UHPLC-DAD-QTOF-MS analysis of CBAE tentatively identified 18 phlorotannins as fucophlorethol and eckol derivatives, containing three up to seven phloroglucinol units. In conclusion, C. barbata represents a valuable source for the development of macroalgal-based products with putative use as nutraceuticals and pharmaceuticals.