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The metalloporphyrin heme acts as the oxygen-complexing prosthetic group of hemoglobin in blood. Heme has been noted to survive for many millions of years in fossils. Here, we investigate its stability and degradation under various conditions expected to occur during fossilization. Oxidative, reductive, aerobic, and anaerobic conditions were studied at neutral and alkaline pH values. Elevated temperatures were applied to accelerate degradation. High-performance liquid chromatography coupled to tandem mass spectrometry (HPLC-MS/MS) identified four main degradation products. The vinyl residues are oxidized to formyl and further to carboxylate groups. In the presence of air or H2O2, cleavage of the tetrapyrrole ring occurs, and hematinic acid is formed. The highest stability of heme was observed under anaerobic reductive conditions (half-life 9.5 days), while the lowest stability was found in the presence of H2O2 (half-life 1 min). We confirmed that the iron cation plays a crucial role in degradation, since protoporphyrin IX, lacking iron, remained significantly more stable. Under anaerobic, reductive conditions, the above-mentioned degradation products were not observed, suggesting a different degradation pathway. To our knowledge, this is the first molecular taphonomy study on heme, which will be useful for understanding its fate during fossilization.

The separation of cannabinoids from hemp materials is nowadays one of the most promising industrial applications of liquid-liquid chromatography (LLC). Despite various experimental research efforts to purify cannabinoids, there are currently few works on process modeling. Thus, this study aimed to explore a straightforward approach to model the LLC separation of cannabinoids from two hemp extracts with different compositions. The feed materials were simplified to mixtures of preselected key components (i.e., cannabidiol, tetrahydrocannabinol, cannabigerol, and cannabinol). The elution profiles of cannabinoids were simulated using the equilibrium-cell model with an empirical nonlinear correlation. The model parameters were derived from the elution profiles of single-solute pulse injections. For the validation of the proposed approach, LLC separations with the two hemp extracts were performed in descending mode with the solvent system composed of hexane/methanol/water 10/8/2 (v/v/v). The injected sample concentrations were gradually increased from 5 to 100 mg/mL. The results showed that the approach could describe reasonably well the elution behavior of the cannabinoids, with deviations of only 1-2 min between simulated and experimental elution times. However, to improve the prediction accuracy, the model parameters can be refitted to the elution profiles of 3-4 systematically selected pulse injections with specific hemp extracts., (© 2024 The Author(s). Journal of Separation Science published by Wiley‐VCH GmbH.)

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., Competing Interests: Declaration of competing interest The authors declare that they have no conflict of interest., (Copyright © 2023. Published by Elsevier B.V.)

Hemp ( Cannabis sativa L.) has been used for millennia as a rich source of food and fibers, whereas hemp flowers have only recently gained an increased market interest due to the presence of cannabinoids and volatile terpenes. Currently, the hemp flower processing industry predominantly focuses on either cannabinoid or terpene extraction. In an attempt to maximize the valorization of hemp flowers, the current study aimed to evaluate the phytochemical composition and antimicrobial properties of several extracts obtained from post-distillation by-products (e.g., spent material, residual distillation water) in comparison to the essential oil and total extract obtained from unprocessed hemp flowers. A terpene analysis of the essential oil revealed 14 monoterpenes and 35 sesquiterpenes. The cannabinoid profiling of extracts showed seven acidic precursors and 14 neutral derivatives, with cannabidiol (CBD) reaching the highest concentration (up to 16 wt.%) in the spent material extract. The antimicrobial assessment of hemp EO, cannabinoid-containing extracts, and single compounds (i.e., CBD, cannabigerol, cannabinol, and cannabichromene) against a panel of 20 microbial strains demonstrated significant inhibitory activities against Gram-positive bacteria, Helicobacter pylori , and Trichophyton species. In conclusion, this work suggests promising opportunities to use cannabinoid-rich materials from hemp flower processing in functional foods, cosmetics, and pharmaceuticals with antimicrobial properties.

Liquid-liquid chromatography (LLC) is a separation technique that utilizes a biphasic solvent system as the mobile and stationary phases. The components are separated solely due to their different distributions between the two liquid phases. Gradient change in the mobile phase composition during the chromatographic process is a powerful method for improving the resolution of separation or shortening the process time. Gradient elution readily applies to LLC with biphasic solvent systems in which the stationary phase composition remains nearly constant when the mobile phase composition changes. This work proposes a model-based approach to optimize gradients in LLC and circumvent tedious trial-and-error experiments. The solutes' distribution constant depends on the mobile phase composition. Thus, the distribution constants were described as a function of the content of one of the solvents (= modifier) in the mobile phase. The dispersive and mass-transfer effects in the tubing and the column are modeled with a stage model. Only a few experiments are required to determine the model parameters. After the validation of the model and its parameters, the model can be used for LLC gradient optimization. The proposed approach was demonstrated for a gradient LLC separation of a mixture of four cannabinoids. Two different gradient shapes, one-step and linear gradient, were considered. For a pre-selected minimal purity requirement, the gradient was optimized for maximum process efficiency, defined as the product of productivity and yield. An experiment conducted with the optimized gradient conditions was in good agreement with the simulation, showing the potential of the proposed method., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier B.V.)

Background: Symphytum (comfrey) genus, particularly Symphytum officinale, has been empirically used in folk medicine mainly for its potent anti-inflammatory properties. In an attempt to shed light on the valorization of less known taxa, the current study evaluated the metabolite profile and antioxidant and enzyme inhibitory effects of nine Symphytum species., Results: Phenolic acids, flavonoids and pyrrolizidine alkaloids were the most representative compounds in all comfrey samples. Hierarchical cluster analysis revealed that, within the roots, S. grandiflorum was slightly different from S. ibericum, S. caucasicum and the remaining species. Within the aerial parts, S. caucasicum and S. asperum differed from the other samples. All Symphytum species showed good antioxidant and enzyme inhibitory activities, as evaluated in DPPH (up to 50.17 mg Trolox equivalents (TE) g -1 ), ABTS (up to 49.92 mg TE g -1 ), cupric reducing antioxidant capacity (CUPRAC, up to 92.93 mg TE g -1 ), ferric reducing antioxidant power (FRAP, up to 53.63 mg TE g -1 ), acetylcholinesterase (AChE, up to 0.52 mg galanthamine equivalents (GALAE) g -1 ), butyrylcholinesterase (BChE, up to 0.96 mg GALAE g -1 ), tyrosinase (up to 13.58 mg kojic acid equivalents g -1 ) and glucosidase (up to 0.28 mmol acarbose equivalents g -1 ) tests. Pearson correlation analysis revealed potential links between danshensu and ABTS/FRAP/CUPRAC, quercetin-O-hexoside and DPPH/CUPRAC, or rabdosiin and anti-BChE activity., Conclusions: By assessing for the first time in a comparative manner the phytochemical-biological profile of a considerably high number of Symphytum samples, this study unveils the potential use of less common comfrey species as novel phytopharmaceutical or agricultural raw materials. © 2024 The Authors. Journal of The Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry., (© 2024 The Authors. Journal of The Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.)

In liquid-liquid chromatography (LLC), mixture components are separated due to their different distribution between the phases of a biphasic liquid system composed of three or four solvents. LLC separations are typically modeled assuming that only the solutes distribute between the two liquid phases and their distribution can be described with a concentration-independent distribution constant. With increasing solute concentration, the physicochemical properties of the biphasic system change, and the distribution of the solutes becomes a function of their concentration. However, the experimental determination of liquid-liquid equilibria in multicomponent systems is time-intensive, and its prediction using thermodynamic models is often not sufficiently accurate for process design purposes. Thus, in this work, we propose a simple approach to model and simulate LLC separations in the nonlinear (concentration-dependent) range of the solutes' distribution equilibria, namely cannabidiol (CBD) and cannabigerol (CBG). Using the inverse method, the distribution equilibrium equation parameters were estimated from pulse injection experiments of single solutes at concentrations ranging from 1 to 100 mg/mL and 1-50 mg/mL for CBD and CBG, respectively. The obtained parameters were then successfully used to predict the elution profiles of binary mixtures of different compositions at 40 mg/mL total cannabinoid concentration. The approach was demonstrated and validated for CBD and CBG as model compounds and n-hexane/methanol/water 10/7.5/2.5 (v/v/v) as the biphasic solvent system. It should be noted that the applicability of the proposed approach is system-dependent, and hence, it should be evaluated for each separation task individually., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier B.V.)

Petasites hybridus L. (butterbur, Asteraceae) is a well-known medicinal plant traditionally used as a remedy for neurological, respiratory, cardiovascular, and gastrointestinal disorders. Eremophilane-type sesquiterpenes (petasins) are considered to be the major bioactive constituents of butterbur. However, efficient methods to isolate high-purity petasins in sufficient amounts for further analytical and biological testing are lacking. In this study, various sesquiterpenes were separated from a methanol rootstock extract of P. hybridus with liquid-liquid chromatography (LLC). The appropriate biphasic solvent system was selected using the predictive thermodynamic model COSMO-RS and shake-flask experiments. After the selection of the feed (extract) concentration and operating flow rate, a batch LLC experiment was performed with n-hexane/ethyl acetate/methanol/water 5/1/5/1 (v/v/v/v). For those LLC fractions containing petasin derivatives with purities < 95%, a preparative high-performance liquid chromatography purification step followed. All isolated compounds were identified by state-of-the-art spectroscopic methods, i.e., liquid chromatography coupled with high-resolution tandem mass spectrometry and nuclear magnetic resonance techniques. As a result, six compounds were obtained, namely 8β-hydroxyeremophil-7(11)-en-12,8-olide, 2-[(angeloyl)oxy]eremophil-7(11)-en-12,8-olide, 8α/β-H-eremophil-7(11)-en-12,8-olide, neopetasin, petasin, and isopetasin. The isolated petasins can be further used as reference materials for standardization and pharmacological evaluation., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Bioactivity-guided isolation of natural products from plant matrices is widely used in drug discovery. Here, this strategy was applied to identify trypanocidal coumarins effective against the parasite Trypanosoma cruzi, the etiologic agent of Chagas disease (American trypanosomiasis). Previously, phylogenetic relationships of trypanocidal activity revealed a coumarin-associated antichagasic hotspot in the Apiaceae. In continuation, a total of 35 ethyl acetate extracts of different Apiaceae species were profiled for selective cytotoxicity against T. cruzi epimastigotes over host CHO-K1 and RAW264.7 cells at 10 μg/mL. A flow cytometry-based T. cruzi trypomastigote cellular infection assay was employed to measure toxicity against the intracellular amastigote stage. Among the tested extracts, Seseli andronakii aerial parts, Portenschlagiella ramosissima and Angelica archangelica subsp. litoralis roots exhibited selective trypanocidal activity and were subjected to bioactivity-guided fractionation and isolation by countercurrent chromatography. The khellactone ester isosamidin isolated from the aerial parts of S. andronakii emerged as a selective trypanocidal molecule (selectivity index ∼9) and inhibited amastigote replication in CHO-K1 cells, though it was significantly less potent than benznidazole. The khellactone ester praeruptorin B and the linear dihydropyranochromones 3'-O-acetylhamaudol and ledebouriellol isolated from the roots of P. ramosissima were more potent and efficiently inhibited the intracellular amastigote replication at < 10 μM. The furanocoumarins imperatorin, isoimperatorin and phellopterin from A. archangelica inhibited T. cruzi replication in host cells only in combination, indicative of superadditive effects, while alloimperatorin was more active in fractions. Our study reports preliminary structure-activity relationships of trypanocidal coumarins and shows that pyranocoumarins and dihydropyranochromones are potential chemical scaffolds for antichagasic drug discovery., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Prof. Jürg Gertsch ic co-guest editor of this Special Issue., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

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 (IC 50 = 45.0 µg/mL), reducing power (EC 50 = 61.3 µg/mL) and chelating power (IC 50 = 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.

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 (CE 50 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., Competing Interests: The authors declare no conflicts of interest.

Anxiety is one of the most common central nervous system disorders, affecting at least one-quarter of the worldwide population. The medications routinely used for the treatment of anxiety (mainly benzodiazepines) are a cause of addiction and are characterized by many undesirable side effects. Thus, there is an important and urgent need for screening and finding novel drug candidates that can be used in the prevention or treatment of anxiety. Simple coumarins usually do not show side effects, or these effects are much lower than in the case of synthetic drugs acting on the central nervous system (CNS). This study aimed to evaluate the anxiolytic activity of three simple coumarins from Peucedanum luxurians Tamamsch, namely officinalin, stenocarpin isobutyrate, and officinalin isobutyrate, in a 5 dpf larval zebrafish model. Moreover, the influence of the tested coumarins on the expression of genes involved in the neural activity ( c-fos , bdnf ) or dopaminergic ( th1 ), serotoninergic ( htr1Aa , htr1b , htr2b ), GABA-ergic ( gabarapa , gabarapb ), enkephalinergic ( penka , penkb ), and galaninergic ( galn ) neurotransmission was assessed by quantitative PCR. All tested coumarins showed significant anxiolytic activity, with officinalin as the most potent compound. The presence of a free hydroxyl group at position C-7 and the lack of methoxy moiety at position C-8 might be key structural features responsible for the observed effects. In addition, officinalin and its isobutyrate upregulated the expression of genes involved in neurotransmission and decreased the expression of genes connected with neural activity. Therefore, the coumarins from P. luxurians might be considered as promising drug candidates for the therapy of anxiety and related disorders.

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 (IC 50 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.

The essential oil of Myristica fragrans Hutt. (nutmeg) is an important commodity used as a flavoring agent in the food, pharmaceutical, and cosmetic fields. Hydrodistillation is chiefly employed at the industrial scale for nutmeg essential oil isolation, but such a technique generates large quantities of post-distillation by-products (e.g., spent plant material and residual distillation water). Therefore, our work aimed to propose a novel strategy for the valorization of nutmeg wastes, with beneficial economic and ecological advantages. Thus, the current study assessed the phytochemical (GC-MS, LC-HRMS/MS) and biological (antioxidant, enzyme inhibitory, antimicrobial) profile of nutmeg crude materials (essential oil and total extract) and post-distillation by-products (residual water and spent material extract). Identified in these were 43 volatile compounds, with sabinene (21.71%), α -pinene (15.81%), myristicin (13.39%), and β -pinene (12.70%) as the main constituents. LC-HRMS/MS analysis of the nutmeg extracts noted fifteen metabolites (e.g., organic acids, flavonoids, phenolic acids, lignans, and diarylnonanoids). Among the investigated nutmeg samples, the spent material extract was highlighted as an important source of bioactive compounds, with a total phenolic and flavonoid content of 63.31 ± 0.72 mg GAE/g and 8.31 ± 0.06 mg RE/g, respectively. Moreover, it showed prominent radical-scavenging and metal-reducing properties and significantly inhibited butyrylcholinesterase (4.78 ± 0.03 mg GALAE/g). Further, the spent material extract displayed strong antimicrobial effects against Streptococcus pneumoniae , Micrococcus luteus , and Bacillus cereus (minimum inhibitory concentrations of 62.5 mg/L). Overall, our study brings evidence on the health-promoting (antioxidant, anti-enzymatic, antimicrobial) potential of nutmeg post-distillation by-products with future reference to their valorization in the pharmaceutical, cosmeceutical, and food industries.

Black pepper (P. nigrum L.) is considered one of the most valuable spices and a promising candidate in natural product research. In this study, the influence of different combinations of pressures (100-300 bar) and temperatures (40-60 °C) on the supercritical CO 2 (SC-CO 2 ) recovery of several key compounds from black pepper was evaluated systematically. The extraction curves showed that terpenes were recovered in a short time under all studied conditions. In contrast, higher pressure values were required to extract piperamides efficiently. Furthermore, the differences in the extraction kinetics of piperine, piperettine, pellitorine, guineensine, and N-isobutyl-2,4,14-eicosatrienamide were linked with several structural features, such as the nature of the amine group or the terminal part of the fatty acid. The data from the isocratic experiments represented the starting point for designing a two-step pressure gradient SC-CO 2 process in which one terpene-rich and one piperamide-rich product were successively obtained., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

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, AgNO 3 concentration, ratio of bark extract and AgNO 3 , 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 (IC 50 = 2.40 ± 0.21 and 6.02 ± 0.61 μg/mL, respectively). The phytosynthesized AgNPs also showed antioxidant and antibacterial effects.

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-1β, 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-1β, 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., Competing Interests: Declaration of competing interest The authors declare no conflict of interest., (Copyright © 2022. Published by Elsevier B.V.)

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., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Epilepsy is a neurological disease that burdens over 50 million people worldwide. Despite the considerable number of available antiseizure medications, it is estimated that around 30% of patients still do not respond to available treatment. Herbal medicines represent a promising source of new antiseizure drugs. This study aimed to identify new drug lead candidates with antiseizure activity from endemic plants of New Caledonia. The crude methanolic leaf extract of Halfordia kendack Guillaumin (Rutaceae) significantly decreased (75 μg/mL and 100 μg/mL) seizure-like behaviour compared to sodium valproate in a zebrafish pentylenetetrazole (PTZ)-induced acute seizure model. The main coumarin compound, halfordin, was subsequently isolated by liquid-liquid chromatography and subjected to locomotor, local field potential (LFP), and gene expression assays. Halfordin (20 μM) significantly decreased convulsive-like behaviour in the locomotor and LFP analysis (by 41.4% and 60%, respectively) and significantly modulated galn , and penka gene expression.

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 CO 2 (SC-CO 2 ) 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-CO 2 extracts, solvent extracts) and by-products/waste materials (post-distillation residual water, spent plant material extracts, and post-supercritical CO 2 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-CO 2 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.

There is currently no use for the vast quantities of post-distillation by-products, such as spent plant materials and residual waters, produced by the essential oil (EO) industry of aromatic herbs. In this study, the EOs of three Lamiaceae species (thyme, oregano, and basil) and their total, spent, and residual water extracts were phytochemically characterized and biologically assessed. The collected information was put through a series of analyses, including principal component analysis, heatmap analysis, and Pearson correlation analysis. Concerning the EOs, 58 volatile compounds were present in thyme (e.g., p -cymene, thymol), 44 compounds in oregano (e.g., thymol, carvacrol), and 67 compounds in basil (e.g., eucalyptol, linalool, estragole, ( E )-methyl cinnamate). The LC-HRMS/MS analysis of the total, spent, and residual water extracts showed the presence of 31 compounds in thyme (e.g., quercetin- O -hexoside, pebrellin, eriodictyol), 31 compounds in oregano (e.g., rosmarinic acid, apigenin, kaempferol, salvianolic acids I, B, and E), and 25 compounds in basil (e.g., fertaric acid, cichoric acid, caftaric acid, salvianolic acid A). The EOs of the three Lamiaceae species showed the highest metal-reducing properties (up to 1792.32 mg TE/g in the CUPRAC assay), whereas the spent extracts of oregano and basil displayed very high radical-scavenging properties (up to 266.59 mg TE/g in DPPH assay). All extracts exhibited anti-acetylcholinesterase (up to 3.29 mg GALAE/g), anti-tyrosinase (up to 70.00 mg KAE/g), anti-amylase (up to 0.66 mmol ACAE/g), and anti-glucosidase (up to 1.22 mmol ACAE/g) effects. Thus, the present research demonstrated that both the raw extracts (EOs and total extracts) and the post-distillation by-products (spent material and residual water extracts) are rich in bioactive metabolites with antioxidant and enzyme inhibitory properties.

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 IC 50 32.23±2.91 μM, while the reference (galantamine) had IC 50 =1.85±0.12 μM. Compounds 9 (IC 50 75.14±1.82 μM), 13 (IC 50 =16.14±0.43 μM), were determined to be stronger BChE inhibitors than the reference galantamine (IC 50 =93.53±2.23 μM). The IC 50 value of compound 16 for BChE inhibition (IC 50 =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., (© 2022 Wiley-VHCA AG, Zurich, Switzerland.)

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.

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., (Copyright © 2022 The Author(s). Published by Elsevier B.V. All rights reserved.)

The Artemisia L. genus comprises over 500 species with important medicinal and economic attributes. Our study aimed at providing a comprehensive metabolite profiling and bioactivity assessment of five Artemisia species collected from northeastern Romania ( A. absinthium L., A. annua L., A. austriaca Jacq., A. pontica L. and A. vulgaris L.). Liquid chromatography-tandem high-resolution mass spectrometry (LC-HRMS/MS) analysis of methanol and chloroform extracts obtained from the roots and aerial parts of the plants led to the identification of 15 phenolic acids (mostly hydroxycinnamic acid derivatives), 26 flavonoids (poly-hydroxylated/poly-methoxylated flavone derivatives, present only in the aerial parts), 14 sesquiterpene lactones, 3 coumarins, 1 lignan and 7 fatty acids. Clustered image map (CIM) analysis of the phytochemical profiles revealed that A. annua was similar to A. absinthium and that A. pontica was similar to A. austriaca, whereas A. vulgaris represented a cluster of its own. Correlated with their total phenolic contents, the methanol extracts from both parts of the plants showed the highest antioxidant effects, as assessed by the DPPH and ABTS radical scavenging, CUPRAC, FRAP and total antioxidant capacity methods. Artemisia extracts proved to be promising sources of enzyme inhibitory agents, with the methanol aerial part extracts being the most active samples against acetylcholinesterase and glucosidase. All Artemisia samples displayed good antibacterial effects against Mycobacterium tuberculosis H37Ra, with MIC values of 64-256 mg/L. In conclusion, the investigated Artemisia species proved to be rich sources of bioactives endowed with antioxidant, enzyme inhibitory and anti-mycobacterial properties.

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., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

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.

The essential oil industry of aromatic herbs and spices is currently producing a significant amount of by-products, such as the spent plant materials remaining after steam or hydrodistillation, that are simply discarded. The aim of this study was to comparatively investigate the phytochemical composition, antioxidant and multi-enzymatic inhibitory potential of the essential oils and spent plant material extractives obtained from cinnamon, cumin, clove, laurel, and black pepper. The essential oils were characterized by the presence of several phytochemical markers (cinnamaldehyde, cuminaldehyde, eugenol, eucalyptol, α -terpinene, limonene, β -caryophyllene or β -pinene). On the other hand, the LC-HRMS/MS profiling of the spent material extracts allowed the annotation of species specific and non-specific metabolites, such as organic acids, phenolic acids, flavonoids, proanthocyanidins, hydrolysable tannins, fatty acids, or piperamides. All samples exhibited very strong antioxidant effects, with the clove essential oil displaying the strongest radical scavenging (525.78 and 936.44 mg TE/g in DPPH and ABTS assays), reducing (2848.28 and 1927.98 mg TE/g in CUPRAC and FRAP), and total antioxidant capacity (68.19 mmol TE/g). With respect to the anti-acetylcholinesterase (0.73-2.95 mg GALAE/g), anti-butyrylcholinesterase (0-3.41 mg GALAE/g), anti-tyrosinase (0-76.86 mg KAE/g), anti-amylase and anti-glucosidase (both 0-1.00 mmol ACAE/g) assays, the spice samples showed a modest activity. Overall, our study reports that, not only the volatile fractions of common spices, but also their spent plant materials remaining after hydrodistillation can be regarded as rich sources of bioactive molecules with antioxidant and multi-enzymatic inhibitory effects.

Aside from Δ 9 -tetrahydrocannabinol (THC) and cannabidiol (CBD), other less common cannabinoids have recently gained an increasing popularity, mostly due to their promising biological potential. However, time-saving and cost-effective methods for their preparative purification are missing. In this study, trapping multiple dual mode (MDM), a flow-reversal liquid-liquid chromatography (LLC) operating mode, was used for the separation of different minor cannabinoids from a hemp extract. Separation task specific biphasic solvent systems were selected for the purification of the target constituents, as follows: n-hexane/methanol/water 10/6.5/3.5 for cannabielsoin (CBE); n-hexane/methanol/water 10/7/3 for cannabidivarin (CBDV) and cannabigerol (CBG); n-hexane/methanol/water 10/8/2 for cannabinol (CBN) and n-hexane/methanol/water 10/9/1 for cannabichromene (CBC) and cannabicylol (CBL). For each separation task, the concentration of the hemp extract in the feed stream and mobile phase flow rate were selected by shake-flask and stationary phase retention experiments, respectively. For the determination of the trapping MDM operating parameters, the short-cut method was implemented and followed by equilibrium-cell model-based simulations. The trapping MDM allowed the separation of the targeted cannabinoids with purities of 93-99%, yields of 73-95%, solvent consumption 2-4-fold lower and productivities almost double than those obtained using batch separation., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021. Published by Elsevier B.V.)

Dermatophyte infections represent a significant public health concern, with an alarming negative impact caused by unsuccessful therapeutic regimens. Natural products have been highlighted as a promising alternative, due to their long-standing traditional use and increasing scientific recognition. In this study, honokiol and magnolol, the main bioactives from Magnolia spp. bark, were investigated for their antidermatophytic activity. The antifungal screening was performed using dermatophyte standard strains and clinical isolates. The minimal inhibitory concentration (MIC) and the minimal fungicidal concentration (MFC) were determined in accordance with EUCAST-AFST guidelines, with minor modifications. The effects on ergosterol biosynthesis were assessed in Trichophyton rubrum cells by HPLC-DAD. Putative interactions with terbinafine against T. rubrum were evaluated by the checkerboard method. Their impact on cells' viability and pro-inflammatory cytokines (IL-1β, IL-8 and TNF-α) was shown using an ex vivo human neutrophils model. Honokiol and magnolol were highly active against tested dermatophytes, with MIC and MFC values of 8 and 16 mg/L, respectively. The mechanism of action involved the inhibition of ergosterol biosynthesis, with accumulation of squalene in T. rubrum cells. Synergy was assessed for binary mixtures of magnolol with terbinafine (FICI = 0.50), while honokiol-terbinafine combinations displayed only additive effects (FICI = 0.56). In addition, magnolol displayed inhibitory effects towards IL-1β, IL-8 and TNF-α released from lipopolysaccharide (LPS)-stimulated human neutrophils, while honokiol only decreased IL-1β secretion, compared to the untreated control. Overall, honokiol and magnolol acted as fungicidal agents against dermatophytes, with impairment of ergosterol biosynthesis.

Dermatophyte infections represent an important public health concern, affecting up to 25% of the world's population. Trichophyton rubrum and T. mentagrophytes are the predominant dermatophytes in cutaneous infections, with a prevalence accounting for 70% of dermatophytoses. Although terbinafine represents the preferred treatment, its clinical use is hampered by side effects, drug-drug interactions, and the emergence of resistant clinical isolates. Combination therapy, associating terbinafine and essential oils (EOs), represents a promising strategy in the treatment of dermatophytosis. In this study, we screened the potential of selected Apiaceae EOs (ajowan, coriander, caraway, and anise) to improve the antifungal activity of terbinafine against T. rubrum ATCC 28188 and T. mentagrophytes ATCC 9533. The chemical profile of EOs was analyzed by gas chromatography. The minimal inhibitory concentration (MIC) and minimal fungicidal concentration (MFC) of EOs/main compounds were determined according to EUCAST-AFST guidelines, with minor modifications. The checkerboard microtiter method was used to identify putative synergistic combinations of EOs/main constituents with terbinafine. The influence of EOs on the viability and pro-inflammatory cytokine production (IL-1 β , IL-8 and TNF- α ) was determined using an ex vivo human neutrophils model. The binary associations of tested EOs with terbinafine were found to be synergistic against T. rubrum , with FICI values of 0.26-0.31. At the tested concentrations (6.25-25 mg/L), EOs did not exert cytotoxic effects towards human neutrophils. Anise EO was the most potent inhibitor of IL-1 β release (46.49% inhibition at 25 mg/L), while coriander EO displayed the highest inhibition towards IL-8 and TNF- α production (54.15% and 54.91%, respectively). In conclusion, the synergistic combinations of terbinafine and investigated Apiaceae EOs could be a starting point in the development of novel topical therapies against T. rubrum -related dermatophytosis.

Piper spices represent an inexhaustible reservoir of bioactive compounds that may act as drug leads in natural product research. The aim of this study was to investigate a series of methanolic fruit extracts obtained from P. nigrum (black, green, white and red), P. longum and P. retrofractum in comparative phytochemical and multi-directional biological (antimicrobial, antioxidant, anti-enzymatic and anti-melanogenic) assays. The metabolite profiling revealed the presence of 17 piperamides, with a total content of 247.75-591.42 mg piperine equivalents/g. Among the 22 tested microorganism strains, Piper spices were significantly active (MIC < 0.1 mg/mL) against the anaerobes Actinomyces israelii and Fusobacterium nucleatum . The antioxidant and anti-enzymatic activities were evidenced in DPPH (10.64-82.44 mg TE/g) and ABTS (14.20-77.60 mg TE/g) radical scavenging, CUPRAC (39.94-140.52 mg TE/g), FRAP (16.05-77.00 mg TE/g), chelating (0-34.80 mg EDTAE/g), anti-acetylcholinesterase (0-2.27 mg GALAE/g), anti-butyrylcholinesterase (0.60-3.11 mg GALAE/g), anti-amylase (0.62-1.11 mmol ACAE/g) and anti-glucosidase (0-1.22 mmol ACAE/g) assays. Several Piper extracts (10 μg/mL) inhibited both melanin synthesis (to 32.05-60.65% of αMSH+ cells) and release (38.06-45.78% of αMSH+ cells) in αMSH-stimulated B16F10 cells, partly explained by their tyrosinase inhibitory properties. Our study uncovers differences between Piper spices and sheds light on their potential use as nutraceuticals or cosmeceuticals for the management of different diseases linked to bacterial infections, Alzheimer's dementia, type 2 diabetes mellitus or hyperpigmentation.

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 (IC 50 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.

This study was aimed at providing a comprehensive phytochemical characterization and multi-biological assessment of Symphytum officinale L., a medicinal plant with a noteworthy traditional use, and Anchusa ochroleuca M. Bieb., a Boraginaceae species from the Romanian flora. The dichloromethane, methanol and 65 % ethanol extracts obtained from the roots and aerial parts of both plants revealed the presence of numerous phenolic acids, oxygenated fatty acids, pyrrolizidine alkaloids (PAs) and flavonoids, as assessed by LC-HRMS/MS analysis. Consistent with their higher total phenolic content, the polar aerial part extracts of S. officinale and root extracts of A. ochroleuca showed the most significant antioxidant activities, as evaluated by DPPH (173.22-216.98 mg TE/g) and ABTS (219.41-311.97 mg TE/g) radical scavenging, CUPRAC (387.18-626.40 mg TE/g), FRAP (199.36-299.86 mg TE/g) and total antioxidant capacity (2.28-2.68 mmol TE/g). Furthermore, both plants exhibited good tyrosinase (19.11-43.89 mg KAE/g) and α-glucosidase (2.45-12.54 mmol ACAE/g) inhibitory effects. The orthogonal projections to latent structures discriminant analysis (OPLS-DA) allowed the objective differentiation between the roots and aerial parts of the two investigated species based on their phytochemical and biological profiles. The partial least square (PLS) analysis showed that several individual phenolic acids, such as danshensu, rabdosiin and rosmarinic acid, significantly contributed to the antioxidant potential of both Boraginaceae species, whilst the relative levels of sucrose were positively correlated with the anti-enzymatic properties. Overall, S. officinale and A. ochroleuca could be regarded as rich sources of bioactive phytochemicals that could further lead to developing novel phyto-pharmaceutical commodities., Competing Interests: Declaration of Competing Interest The authors declare that they have no actual or potential competing financial interests or personal relationships that could have appeared to influence the work reported in the present paper., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Comfrey ( Symphytum officinale L.) roots are well-known bioactive ingredients included in various cosmeceutical and pharmaceutical preparations. In this study, the influence of the post-harvest storage on the chemico-biological potential of roots collected from different European regions and stored for up to six months was investigated. Total phenolic content (TPC) and total phenolic acid content (TPAC) were spectrophotometrically estimated, whereas the levels of individual phenolic and pyrrolizidine alkaloidal markers were determined by HPLC-DAD and HPLC-MS/MS, respectively. The changes in the biological potential was tracked via antioxidant (DPPH, ABTS, CUPRAC, and FRAP) and anti-enzymatic (cholinesterase, tyrosinase, glucosidase, and amylase) assays. TPC and TPAC varied from 6.48-16.57 mg GAE/g d.w. root and from 2.67-9.03 mg CAE/g, respectively. The concentration of the four phenolics (rosmarinic acid, globoidnan A, globoidnan B, rabdosiin) and six pyrrolizidine alkaloids generally showed maximum values at 1-3 months, after which their levels significantly decreased. With respect to the bioassays, the samples showed a wide range of antioxidant and anti-enzymatic effects; however, a direct storage time-bioactivity relationship was not observed. Similar conclusions were also revealed by the multivariate and correlation analyses. Our study could improve the current knowledge of the shelf-life properties of comfrey-based products and enhance their industrial exploitation.

Introduction: Symphytum officinale L. (comfrey, Boraginaceae) is a cultivated or spontaneously growing medicinal plant that is traditionally used for the treatment of bone fractures, hematomas, muscle pains and joint pains. A wide range of topical preparations and dried roots for ex tempore applications are marketed in European drug stores or pharmacies., Objective: The aim of this study was to perform the qualitative and quantitative analysis of pyrrolizidine alkaloids (PAs) and phenolic compounds in the hydroethanolic extracts of 16 commercial comfrey root batches purchased from 12 different European countries., Methods: Liquid chromatography hyphenated with high-resolution tandem mass spectrometry (LC-HRMS/MS) was used for the profiling of PAs and phenolic compounds, whereas LC-MS/MS and liquid chromatography with diode array detection (LC-DAD) were used for their quantification., Results: 20 PAs (i.e. intermedine, lycopsamine, acetylintermedine, acetyllycopsamine, symphytine, symphytine-N-oxide), 17 phenolic compounds (i.e. caffeic and rosmarinic acids, rabdosiin, globoidnan A, globoidnan B) and 9 nonphenolic compounds (sugars, organic and fatty acids) were fully or partly annotated in the analysed samples. In addition, the quantitative analyses revealed that globoidnan B, rabdosiin and globoidnan A are new phenolic markers that can be used together with rosmarinic acid and PAs for the quality control of commercial comfrey root batches., Conclusions: This study brings new insights into the phytochemical complexity of S. officinale, revealing not only numerous toxic PAs, but also a significant number of valuable phenolic compounds that could contribute to the bioactivities of comfrey-based preparations., (© 2020 The Authors. Phytochemical Analysis published by John Wiley & Sons Ltd.)

The plant genus Piper comprises extensively consumed spice taxa like black pepper (P. nigrum L.) or long pepper (P. longum L.). The chronic dietary use of different Piper spices has been associated with different health benefits, though the molecular mechanisms remain poorly understood. The aim of this work was to perform the liquid-chromatography-high-resolution tandem mass spectrometry (LC-HRMS/MS) profiling and LC-DAD quantification of piperamides in several Piper species and varieties and study their ability to modulate the endocannabinoid system (ECS). LC-HRMS/MS analysis revealed a number of 42 piperamides grouped into six structural classes, with 22 of them, notably piperine, retrofractamide B, guineensine, piperchabamide C, being also quantified by LC-DAD. The multivariate analysis showed that P. nigrum and P. longum are very similar with respect to their piperamide profile, while the other Piper spices (P. retrofractum, P. guineense, P. cubeba, P. borbonense) might have significantly different metabolite patterns. The results from the biological assays confirmed that guineensine and total piperamides are strongly correlated with anandamide (AEA) cellular uptake inhibition. While none of the Piper spice extracts showed binding activity at cannabinoid CB 1 receptors, some P. nigrum varieties exhibited moderate binding interactions with CB 2 receptors. Overall, the analytical profiling enabled global annotations of piperamides associated to cannabimimetic effects in Piper spices., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Different types of anxiety disorders have become the number one mental health issue in developed countries. The search for new, safer and effective drug-like molecules among naturally derived substances faces two difficulties: an efficient method of isolation compounds with a high-purity and high-throughput animal model for activity assay. Thus, the aim of the present study was to isolate by liquid-liquid chromatography high-purity rare coumarins from the fruits of Seseli devenyense Simonk. and evaluate their anxiolytic effect (defined as reversed thimotaxis) using a 5-days post-fertilization (dpf) Danio rerio larvae model. Liquid-liquid chromatography enabled the isolation of one simple hydroxycoumarin (devenyol) and four pyranocoumarins (cis-khellactone, d-laserpitin, isolaserpitin and octanoyllomatin). The anxiolytic effect was defined as a decrease in the time spent in the boundaries of the living space (also described as reversed thigmotaxis). Our results show that all isolated courmarins exerted a significant influence on the anxiety behavior (anxiolytic activity) in the zebrafish larvae model. According to our knowledge, this is the first report of anxiolytic activity of pyranocoumarins and devenyol.

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 IC 50 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., (© 2020 The Authors. Chemistry & Biodiversity published by Wiley-VHCA AG, Zurich, Switzerland.)

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)-1β, 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 EC 50 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-1β 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., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Naturally occurring coumarins are a group of compounds with many documented central nervous system (CNS) activities. However, dihydrofuranocoumarins have been infrequently investigated for their bioactivities at CNS level. Within the frame of this study, an efficient liquid-liquid chromatography method was developed to rapidly isolate rutamarin from Ruta graveolens L. (Rutaceae) dichloromethane extract (DCM). The crude DCM (9.78 mg/mL) and rutamarin (6.17 M) were found to be effective inhibitors of human monoamine oxidase B ( h MAO-B) with inhibition percentages of 89.98% and 95.26%, respectively. The inhibitory activity against human monoamine oxidase A ( h MAO-A) for the DCM extract was almost the same (88.22%). However, for rutamarin, it significantly dropped to 25.15%. To examine the molecular interaction of rutamarin with h MAO- B, an in silico evaluation was implemented. A docking study was performed for the two enantiomers ( R )-rutamarin and ( S )-rutamarin. The ( S )-rutamarin was found to bind stronger to the h MAO-B binging cavity.

The emergence of multidrug resistant pathogens is a great challenge to the medical field and a major global health threat requiring novel therapeutic strategies. Among plant products, essential oils have significant antimicrobial properties that make them promising agents in the fight against drug resistant human pathogens. The aim of the present review was to highlight the most important essential oil-based antimicrobial strategies as revealed by recent studies. Synergistic interactions between essential oils or their bioactive compounds in combination with known antibiotics are presented. Also, nanoformulation approaches to boost the antimicrobial activity of essential oils are reviewed in terms of bioefficiency, stability and design of the nanostructured delivery systems. The focus was mainly put on the antimicrobial activity against multi-drug resistant pathogens, also called "ESKAPE" organisms ( Enterococcus spp., Staphylococcus aureus , Klebsiella spp., Acinetobacter baumannii , Pseudomonas aeruginosa , and Enterobacter spp.). Thus, essential oils in combinatorial and nano-based strategies may cope with infections caused by drug-resistant bacteria and may offer possibilities for reducing antibiotic use. Research on the in vivo efficacy and safety of such strategies is required for further clinical antimicrobial chemotherapy. In this regard, the understanding of the interactions between essential oil-based strategies and biological interface is essential.

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 Ca 2+ entry and, to a lesser extent, Ca 2+ 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., Competing Interests: Declaration of competing interest The authors declare that they have no conflicts of interest., (Copyright © 2020 Elsevier B.V. All rights reserved.)

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 G 0 /G 1 phase and apoptosis, whereas Verb increased the population of subG 1 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., (© 2020 Wiley Periodicals, Inc.)

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.