In Vitro and In Vivo Pharmacological Profiles of LENART01, a Dermorphin-Ranatensin Hybrid Peptide.

Diverse chemical and pharmacological strategies are currently being explored to minimize the unwanted side effects of currently used opioid analgesics while achieving effective pain relief. The use of multitarget ligands with activity at more than one receptor represents a promising therapeutic approach. We recently reported a bifunctional peptide-based hybrid LENART01 combining dermorphin and ranatensin pharmacophores, which displays activity to the mu-opioid receptor (MOR) and dopamine D2 receptor (D2R) in rat brains and spinal cords. In this study, we investigated the in vitro binding and functional activities to the human MOR and the in vivo pharmacology of LENART01 in mice after subcutaneous administration. In vitro binding assays showed LENART01 to bind and be selective to the human MOR over the other opioid receptor subtypes and delta, kappa and nociceptin receptors. In the [35S]GTPγS binding assay, LENART01 acted as a potent and full agonist to the human MOR. In mice, LENART01 produced dose-dependent antinociceptive effects in formalin-induced inflammatory pain, with increased potency than morphine. Antinociceptive effects were reversed by naloxone, indicating MOR activation in vivo. Behavioral studies also demonstrated LENART01's properties to induce less adverse effects without locomotor dysfunction and withdrawal syndrome compared to conventional opioid analgesics, such as morphine. LENART01 is the first peptide-based MOR-D2R ligand known to date and the first dual MOR-dopamine D2R ligand for which in vivo pharmacology is reported with antinociceptive efficacy and reduced opioid-related side effects. Our current findings may pave the way to new pain therapeutics with limited side effects in acute and chronic use.

Urolithin A conjugation with NSAIDs inhibits its glucuronidation and maintains improvement of Caco-2 monolayers' barrier function.

Urolithin A (UA) is an ellagitannin-derived postbiotic metabolite which emerged as a promising health-boosting agent, promoting mitophagy, improving skeletal muscle function, and suppressing the inflammatory response. However, phase II intestinal metabolism severely limits its biopotency, leading to the formation of nonactive glucuronides. To address this constraint, a set of new UA derivatives (UADs), conjugated with nonsteroidal anti-inflammatory drugs (NSAIDs), was synthesized. The bioavailability and inhibitory activity of UADs against UA-glucuronidation were evaluated using differentiated Caco-2 cell monolayers. Parallelly, after the administration of tested substances, the transepithelial electrical resistance (TEER) of the cell monolayers was continuously monitored using the CellZscope device. Though investigated UADs did not penetrate Caco-2 monolayers, all of them significantly suppressed the glucuronidation rate of UA, while conjugates with diclofenac increased the concentration of free molecule on the basolateral side. Moreover, esters of UA with diclofenac (DicloUA) and aspirin (AspUA) positively influenced cell membrane integrity. Western blot analysis revealed that some UADs, including DicloUA, increased the expression of pore-sealing tight junction proteins and decreased the level of pore-forming claudin-2, which may contribute to their beneficial activity towards the barrier function. To provide comprehensive insight into the mechanism of action of DicloUA, Caco-2 cells were subjected to transcriptomic analysis. Next-generation sequencing (NGS) uncovered substantial changes in the expression of genes involved, for instance, in multivesicular body organization and zinc ion homeostasis. The results presented in this study offer new perspectives on the beneficial effects of modifying UA's structure on its intestinal metabolism and bioactivity in vitro.

Chemical Composition and Insulin-Resistance Activity of Arginine-Loaded American Cranberry (Vaccinium macrocarpon Aiton, Ericaceae) Leaf Extracts.

One of the key pathogenetic links in type 2 diabetes mellitus (T2DM) is the formation of insulin resistance (IR). Besides a wide selection of synthetic antidiabetic drugs, various plant-origin extracts are also available to support the treatment of T2DM. This study aimed to investigate and gain knowledge of the chemical composition and potential IR correction effect of American cranberry (Vaccinium macrocarpon Aiton) leaf extracts and formulate novel 3D-printed oral dosage forms for such extracts. The bioactivity and IR of L-arginine-loaded cranberry leaf extracts were studied in vivo in rats. The cranberry leaf extracts consisted of quinic, 3-caffeoylquinic (chlorogenic), p-coumaroylquinic acids, quercetin 3-O-galactoside, quercetin-3-O-glucoside, quercetin-3-xyloside, quercetin-3-O-arabino pyranoside, quercetin-3-O-arabinofuranoside, quercetin 3-O-rhamnoside, and quercetin-O-p-coumaroyl hexoside-2 identified by HPLC. In vivo studies with rats showed that the oral administration of the cranberry leaf extracts had a positive effect on insulin sensitivity coefficients under the insulin tolerance test and affected homeostasis model assessment IR levels and liver lipid content with experimental IR. A novel 3D-printed immediate-release dosage form was developed for the oral administration of cranberry leaf extracts using polyethylene oxide as a carrier gel in semi-solid extrusion 3D printing. In conclusion, American cranberry leaf extracts loaded with L-arginine could find uses in preventing health issues associated with IR.

Therapeutic Effects of Oral Application of Menthol and Extracts from Tormentil (Potentilla erecta), Raspberry Leaves (Rubus idaeus), and Loosestrife (Lythrum salicaria) during Acute Murine Campylobacteriosis.

Human food-borne infections with the enteropathogen Campylobacter jejuni are becoming increasingly prevalent worldwide. Since antibiotics are usually not indicated in campylobacteriosis, alternative treatment regimens are important. We here investigated potential disease-alleviating effects of menthol and of extracts from tormentil, raspberry leaves, and loosestrife in acute murine campylobacteriosis. Therefore, C. jejuni-infected microbiota-depleted IL-10-/- mice were orally treated with the compounds alone or all in combination from day 2 until day 6 post-infection. Whereas neither treatment regimen affected gastrointestinal pathogen loads, the combination of compounds alleviated C. jejuni-induced diarrheal symptoms in diseased mice on day 6 post-infection. Furthermore, the therapeutic application of tormentil and menthol alone and the combination of the four compounds resulted in lower colonic T cell numbers in infected mice when compared to placebo counterparts. Notably, pro-inflammatory cytokines measured in mesenteric lymph nodes taken from C. jejuni-infected mice following tormentil, menthol, and combination treatment did not differ from basal concentrations. However, neither treatment regimen could dampen extra-intestinal immune responses, including systemic pro-inflammatory cytokine secretion on day 6 post-infection. In conclusion, the combination of menthol and of extracts from tormentil, raspberry leaves, and loosestrife constitutes an antibiotic-independent approach to alleviate campylobacteriosis symptoms.

Synthesis and anthelmintic activity of novel thiosemicarbazide and 1,2,4-triazole derivatives: In vitro, in vivo, and in silico study.

INTRODUCTION: Intestinal parasitic infections are neglected diseases and, due to the increasing resistance of parasites to available drugs, they pose an increasing therapeutic challenge. Therefore, there is a great need for finding new compounds with antiparasitic activity. OBJECTIVES: In this work, new thiosemicarbazide and 1,2,4-triazole derivatives were synthesized and tested for their anthelmintic activity. METHODS: The synthesis was carried out by classical methods of organic chemistry. Anthelmintic activity tests were carried out in vitro (Rhabditis sp., Haemonchus contortus, Strongylidae sp.) in vitro (Heligmosomoides polygyrus/bakeri), and in silico analysis was performed. RESULTS: Quinoline-6-carboxylic acid derivative compounds were designed and synthesized. The highest activity in the screening tests in the Rhabditis model was demonstrated by compound II-1 with a methoxyphenyl substituent LC50 = 0.3 mg/mL. In the next stage of the research, compound II-1 was analyzed in the H. contortus model. The results showed that compound II-1 was active and had ovicidal (percentage of dead eggs > 45 %) and larvicidal (percentage of dead larvae > 75 %) properties. Studies in the Strongylidae sp. model confirmed the ovicidal activity of compound II-1 (percentage of dead eggs ≥ 55 %). In vivo studies conducted in the H. polygyrus/bakeri nematode model showed that the number of nematodes decreased by an average of 30 % under the influence of compound II-1. In silico studies have shown two possible modes of action of compound II-1, i.e. inhibition of tubulin polymerization and SDH. The test compound did not show any systemic toxic effects. Its influence on drug metabolism related to the activity of cytochrome CYP450 enzymes was also investigated. CONCLUSION: The results obtained in the in vitro, in vivo, and in silico studies indicate that the test compound can be described as a HIT, which in the future may be used in the treatment of parasitic diseases in humans and animals.

Chemical composition of extracts from leaves, stems and roots of wasabi (Eutrema japonicum) and their anti-cancer, anti-inflammatory and anti-microbial activities.

The purpose of our study was to evaluate the composition of the extracts obtained from the roots and leaves of Eutrema japonicum cultivated in Poland. For this purpose, LC-DAD-IT-MS and LC-Q-TOF-MS analyses were used. The results revealed the presence of forty-two constituents comprising glycosinolates, phenylpropanoid glycosides, flavone glycosides, hydroxycinnamic acids, and other compounds. Then, the resultant extracts were subjected to an assessment of the potential cytotoxic effect on human colon adenocarcinoma cells, the effect on the growth of probiotic and intestinal pathogenic strains, as well as their anti-inflammatory activity. It was demonstrated that 60% ethanol extract from the biennial roots (WR2) had the strongest anti-inflammatory, antibacterial, and cytotoxic activities compared to the other samples. Our results suggest that extracts from E. japonicum may be considered as a promising compound for the production of health-promoting supplements.

HPLC-DAD-MS3 fingerprints of phenolics of selected Polygonum taxa and their chemometric analysis.

Many Polygonaceae taxa such as Bistorta officinalis, Persicaria amphibia, Persicaria hydropiper, Persicaria lapathifolia, Persicaria maculosa, Persicaria mitis, Polygonum aviculare occur naturally in the entire territory of Poland and are also common in other European countries. Many of these species are also utilised as medicinal plants. In this manuscript we establish the phytochemical profiles of selected taxa from the Polygonaceae focusing on phenolics. Additionally, we try to find chemophenetic markers for the species investigated. Compounds were detected and characterised based on HPLC-DAD-MS data, quantified, and furtherly analysed using multivariate analyses. Chemophenetic markers were identified also considering previous literature.

Phytotherapy of mood disorders in the light of microbiota-gut-brain axis.

BACKGROUND: Clinical research in natural product-based psychopharmacology has revealed a variety of promising herbal medicines that may provide benefit in the treatment of mild mood disorders, however failed to unambiguously indicate pharmacologically active constituents. The emerging role of the microbiota-gut-brain axis opens new possibilities in the search for effective methods of treatment and prevention of mood disorders. PURPOSE: Considering the clinically proven effectiveness juxtaposed with inconsistencies regarding the indication of active principles for many medicinal plants applied in the treatment of anxiety and depression, the aim of the review is to look at their therapeutic properties from the perspective of the microbiota-gut-brain axis. METHOD: A literature-based survey was performed using Scopus, Pubmed, and Google Scholar databases. The current state of knowledge regarding Hypericum perforatum, Valeriana officinalis, Piper methysticum, Passiflora incarnata, Humulus lupulus, Melissa officinalis, Lavandula officinalis, and Rhodiola rosea in terms of their antimicrobial activity, bioavailability, clinical effectiveness in depression/anxiety and gut microbiota - natural products interaction was summarized and analyzed. RESULTS: Recent studies have provided direct and indirect evidence that herbal extracts and isolated compounds are potent modulators of gut microbiota structure. Additionally, some of the formed postbiotic metabolites exert positive effects and ameliorate depression-related behaviors in animal models of mood disorders. The review underlines the gap in research on natural products - gut microbiota interaction in the context of mood disorders. CONCLUSION: Modification of microbiota-gut-brain axis by natural products is a plausible explanation of their therapeutic properties. Future studies evaluating the effectiveness of herbal medicine and isolated compounds in treating mild mood disorders should consider the bidirectional interplay between phytoconstituents and the gut microbiota community.

In vitro cytotoxic activity of Cuphea ingrata Cham. & Schltdl. extracts related to the oenothein B content.

Cuphea ingrata is a traditional medicinal plant species of the Lythraceae family. This work reports on the cytotoxic activity of the methanolic extract from the aerial parts of C. ingrata and the n-butanol and ethyl acetate fractions against human skin and prostate cancer cells. The selectivity of action was tested in normal skin keratinocytes HaCaT and prostate epithelial cells PNT2. The ethyl acetate fraction showed the highest activity in all three human skin cancer cell lines: A375, HTB-140, WM793, with IC50 = 15.90; 3.40; 18.75 µg/mL, respectively. To obtain comparative information on the chemical composition, a quantitative analysis of oenothein B was performed using the UHPLC-PDA method. An analysis of its cytotoxic activity was also carried out.

Bioactive Potential: A Pharmacognostic Definition through the Screening of Four Hypericum Species from the Canary Islands.

In this work, we propose a general methodology to assess the bioactive potential (BP) of extracts in the quest of vegetable-based drugs. To exemplify the method, we studied the anticancer potential (AP) of four endemic species of genus Hypericum (Hypericum canariense L, Hypericum glandulosum Aiton, Hypericum grandifolium Choisy and Hypericum reflexum L.f) from the Canary Islands. Microextracts were obtained from the aerial parts of these species and were tested against six human tumor cell lines, A549 (non-small-cell lung), HBL-100 (breast), HeLa (cervix), SW1573 (non-small-cell lung), T-47D (breast) and WiDr (colon). The methanol-water microextracts were evaluated further for cell migration, autophagy and cell death. The most promising bioactive polar microextracts were analyzed by UHPLC-DAD-MS. The extraction yield, the bioactivity evaluation and the chemical profiling by LC-MS suggested that H. grandifolium was the species with the highest AP. Label-free live-cell imaging studies on HeLa cells exposed to the methanol-water microextract of H. grandifolium enabled observing cell death and several apoptotic hallmarks. Overall, this study allows us to select Hypericum grandifolium Choisy as a source of new chemical entities with a potential interest for cancer treatment.

Comparative Study of Chemical Stability of a PK20 Opioid-Neurotensin Hybrid Peptide and Its Analogue [Ile9]PK20-The Effect of Isomerism of a Single Amino Acid.

Chemical stability is one of the main problems during the discovery and development of potent drugs. When ignored, it may lead to unreliable biological and pharmacokinetics data, especially regarding the degradation of products' possible toxicity. Recently, two biologically active drug candidates were presented that combine both opioid and neurotensin pharmacophores in one entity, thus generating a hybrid compound. Importantly, these chimeras are structurally similar except for an amino acid change at position 9 of the peptide chain. In fact, isoleucine (C6H13NO2) was replaced with its isomer tert-leucine. These may further lead to various differences in hybrids' behavior under specific conditions (temperature, UV, oxidative, acid/base environment). Therefore, the purpose of the study is to assess and compare the chemical stability of two hybrid peptides that differ in nature by way of one amino acid (tert-leucine vs. isoleucine). The obtained results indicate that, opposite to biological activity, the substitution of tert-leucine into isoleucine did not substantially influence the compound's chemical stability. In fact, neither hydrolysis under alkaline and acidic conditions nor oxidative degradation resulted in spectacular differences between the two compounds-although the number of potential degradation products increased, particularly under acidic pH. However, such a modification significantly reduced the compound's half-life from 204.4 h (for PK20 exposed to 1M HCl) to 117.7 h for [Ile9]PK20.

Current Knowledge on Interactions of Plant Materials Traditionally Used in Skin Diseases in Poland and Ukraine with Human Skin Microbiota.

Skin disorders of different etiology, such as dermatitis, atopic dermatitis, eczema, psoriasis, wounds, burns, and others, are widely spread in the population. In severe cases, they require the topical application of drugs, such as antibiotics, steroids, and calcineurin inhibitors. With milder symptoms, which do not require acute pharmacological interventions, medications, dietary supplements, and cosmetic products of plant material origin are gaining greater popularity among professionals and patients. They are applied in various pharmaceutical forms, such as raw infusions, tinctures, creams, and ointments. Although plant-based formulations have been used by humankind since ancient times, it is often unclear what the mechanisms of the observed beneficial effects are. Recent advances in the contribution of the skin microbiota in maintaining skin homeostasis can shed new light on understanding the activity of topically applied plant-based products. Although the influence of various plants on skin-related ailments are well documented in vivo and in vitro, little is known about the interaction with the network of the skin microbial ecosystem. The review aims to summarize the hitherto scientific data on plant-based topical preparations used in Poland and Ukraine and indicate future directions of the studies respecting recent developments in understanding the etiology of skin diseases. The current knowledge on investigations of interactions of plant materials/extracts with skin microbiome was reviewed for the first time.

Conjugates of urolithin A with NSAIDs, their stability, cytotoxicity, and anti-inflammatory potential.

Urolithin A (UA, 1), a gut microbiota postbiotic metabolite is attributed to express interesting biological activities indicated by in vitro, in vivo and clinical studies. Due to its strong anti-inflammatory properties it is considered as a promising lead molecule for further drug development, however, its strong phase II metabolism, severely limits its oral application. Therefore, monoesterified UA derivatives with selected NSAIDs: ibuprofen (Mix 3a/3b), mefenamic acid (Mix 4a/4b), diclofenac (Mix 5a/5b) and aspirin (Mix 6a/6b) were designed. Performed array of stability assays indicated Mix 4a/4b as a most suitable candidate for further studies due to its exceptional stability in human plasma. Thus, we evaluated effects of Mix 4a/4b on cell viability as well as the impact on cytokines secretion in THP-1 derived macrophages and compared it to UA. At high concentration (50 µM) Mix 4a/4b expressed a cytotoxic effect, however at concentration of 5 µM it significantly suppressed TNF-α secretion, and significantly increased ani-inflammatory IL-10 secretion at 10 µM without affecting cell viability. This work has led to selection of a novel UA derivatives, which are stable in solutions and in human plasma as well as posess anti-inflammatory activity towards THP-1 macrophages at non-cytotoxic concentrations.

Tiliae flos metabolites and their beneficial influence on human gut microbiota biodiversity ex vivo.

ETHNOPHARMACOLOGICAL RELEVANCE: The linden flower (Tiliae flos) has been used for centuries to treat and relieve symptoms of the common cold, throat irritation, and upper respiratory tract disturbances. Traditionally, this herb is administered orally, and thus it undergoes intestinal metabolism. Although it is pharmacopeial plant material, there are no reports about its interaction with human gut microbiota. AIM OF THE STUDY: The study aimed to determine the interaction between human gut microbiota and the linden flower extracts, resulting in the biotransformation of the extract's constituents and changes in the microbiota composition. MATERIAL AND METHODS: The linden flower metabolites were obtained by incubation of extract with human faecal slurries from 5 healthy donors. The UHPLC-DAD-MSn analysis determined the composition of raw extract and analysis of microbial metabolites. The intestinal microbiota isolation and sequencing were used to determine changes in microbiota composition. The anti-inflammatory activity was tested using the LPS-stimulated human neutrophils model and ELISA test. RESULTS: After incubation of linden flower extract with human gut microbiota, twenty metabolites were detected and characterized, and three among them were identified. The extract changed human gut microbiota composition but did not cause dysbiosis (change in the abundance of forty-three genera). Raw extract and their metabolites exhibit different levels of inhibition of cytokines production by LPS-stimulated neutrophils, but the reduction of TNF-α production was observed. CONCLUSIONS: The linden flower extract has a beneficial influence on human gut microbiota because it promotes increasing the abundance of bacteria responsible for SCFAs production. The anti-inflammatory effect might be linked to both microbiota composition changes and direct activity of bioavailable metabolites. Increased abundance of SCFAs producers may inhibit the production of pro-inflammatory cytokines. A low concentration of phenolic compounds in metabolized linden flower extract and responsible for anti-inflammatory properties, and the multitude of biological and chemical particles and their interactions may weaken these properties.

Polyphenols and Maillard Reaction Products in Dried Prunus spinosa Fruits: Quality Aspects and Contribution to Anti-Inflammatory and Antioxidant Activity in Human Immune Cells Ex Vivo.

Dried Prunus spinosa fruits (sloes) are folk phytotherapeutics applied to treat chronic inflammatory disorders. However, their pharmacological potential, activity vectors, and drying-related changes in bioactive components remain unexplored. Therefore, the present research aimed to evaluate the anti-inflammatory and antioxidant effects of dried sloes in ex vivo models of human neutrophils and peripheral blood mononuclear cells (PMBCs) and establish their main active components. It was revealed that the fruit extracts significantly and dose-dependently inhibited the respiratory burst, downregulated the production of elastase (ELA-2) and TNF-α, and upregulated the IL-10 secretion by immune cells under pro-inflammatory and pro-oxidant stimulation. The slightly reduced IL-6 and IL-8 secretion was also observed. The structural identification of active compounds, including 45 phenolics and three Maillard reaction products (MRPs) which were formed during drying, was performed by an integrated approach combining LC-MS/MS, preparative HPLC isolation, and NMR studies. The cellular tests of four isolated model compounds (chlorogenic acid, quercetin, procyanidin B2, and 5-hydroxymethylfurfural), supported by statistical correlation studies, revealed a significant polyphenolic contribution and a slight impact of MRPs on the extracts' effects. Moreover, a substantial synergy was observed for phenolic acids, flavonoids, condensed proanthocyanidins, and MPRs. These results might support the phytotherapeutic use of dried P. spinosa fruits to relieve inflammation and establish the quality control procedure for the extracts prepared thereof.

The Effect of Standardised Leaf Extracts of Gaultheria procumbens on Multiple Oxidants, Inflammation-Related Enzymes, and Pro-Oxidant and Pro-Inflammatory Functions of Human Neutrophils.

The leaves of Gaultheria procumbens are polyphenol-rich traditional medicines used to treat inflammation-related diseases. The present study aimed to optimise the solvent for the effective recovery of active leaf components through simple direct extraction and verify the biological effects of the selected extract in a model of human neutrophils ex vivo. The extracts were comprehensively standardised, and forty-one individual polyphenols, representing salicylates, catechins, procyanidins, phenolic acids, and flavonoids, were identified by UHPLC-PDA-ESI-MS3. The chosen methanol-water (75:25, v/v) extract (ME) was obtained with the highest extraction yield and total phenolic levels (397.9 mg/g extract's dw), including 98.9 mg/g salicylates and 299.0 mg/g non-salicylate polyphenols. In biological tests, ME revealed a significant and dose-dependent ability to modulate pro-oxidant and pro-inflammatory functions of human neutrophils: it strongly reduced the ROS level and downregulated the release of pro-inflammatory cytokines and tissue remodelling enzymes, especially IL-1ß and elastase 2, in cells stimulated by fMLP, LPS, or fMLP + cytochalasin B. The extracts were also potent direct scavengers of in vivo relevant oxidants (O2•-, •OH, and H2O2) and inhibitors of pro-inflammatory enzymes (cyclooxygenase-2, hyaluronidase, and lipoxygenase). The statistically significant correlations between the tested variables revealed the synergic contribution of individual polyphenols to the observed effects and indicated them as useful active markers for the standardisation of the extract/plant material. Moreover, the safety of ME was confirmed in cytotoxicity tests. The obtained results might partially explain the ethnomedicinal application of G. procumbens leaves and support the usage of the standardised leaf extract in the adjuvant treatment of oxidative stress and inflammation-related chronic diseases.

Polyphenol-Enriched Extracts of Prunus spinosa Fruits: Anti-Inflammatory and Antioxidant Effects in Human Immune Cells Ex Vivo in Relation to Phytochemical Profile.

The fresh fruits of Prunus spinosa L., a wild plum species, are traditionally used for dietary purposes and medicinal applications in disorders related to inflammation and oxidative stress. This study aimed to investigate the phytochemical composition of the fruits in the function of fractionated extraction and evaluate the biological potential of the extracts as functional products in two models of human immune cells ex vivo. Fifty-seven phenolic components were identified in the extracts by UHPLC-PDA-ESI-MS3, including twenty-eight new for the analysed fruits. Fractionation enabled the enrichment of polyphenols in the extracts up to 126.5 mg gallic acid equivalents/g dw total contents, 91.3 mg/g phenolic acids (caffeoyl-, coumaroyl-, and feruloylquinic acids), 41.1 mg/g flavonoids (mostly quercetin mono-, di- and triglycosides), 44.5 mg/g condensed proanthocyanidins, and 9.2 mg/g anthocyanins (cyanidin and peonidin glycosides). The hydroalcoholic extract and phenolic-enriched fractions of the fruits revealed significant ability to modulate pro-oxidant, pro-inflammatory, and anti-inflammatory functions of human neutrophils and peripheral blood mononuclear cells (PBMCs): they strongly downregulated the release of reactive oxygen species, TNF-α, and neutrophils elastase, upregulated the secretion of IL-10, and slightly inhibited the production of IL-8 and IL-6 in the cells stimulated by fMLP, fMLP+cytochalasin B, and LPS, depending on the test. Correlation studies and experiments on the pure compounds indicated a significant contribution of polyphenols to these effects. Moreover, cellular safety was confirmed for the extracts by flow cytometry in a wide range of concentrations. The results support the traditional use of fresh blackthorn fruits in inflammatory disorders and indicate extracts that are most promising for functional applications.

Determination of Flavonoids in Selected Scleranthus Species and Their Anti-Collagenase and Antioxidant Potential.

A new 5,7-dihydroxy-3'-methoxy-4'-acetoxyflavone-8-C-ß-d-arabinopyranoside-2″-O-(4‴-acetoxy)-glucoside (6) and three known flavone C-glycosides-5,7,3',4'-tetrahydroxyflavone-6-C-xyloside-8-C-ß-d-glucoside (lucenin-1) (7), 5,7,3'-trihydroxyflavone-6-C-glucoside-8-C-ß-d-glucoside (vicenin-2) (8), and 5,7,4'-trihydroxy-3'-methoxyflavone-6-C-ß-d-glucopyranoside-8-C-α-arabinopyranoside (chrysoeriol-6-C-ß-d-glucopyranoside-8-C-α-arabinopyranoside) (9)-were isolated from aerial parts of Scleranthus perennis L. (Caryophyllaceae). Their structures were determined through the use of comprehensive spectroscopic and spectrometric methods, and a method for the quantification of the major constituents of S. perennis and S. annuus L. was developed. Furthermore, the anti-collagenase and antioxidant activities of all isolated compounds obtained from extracts and fractions from both Scleranthus species were evaluated. The highest percentage of collagenase inhibition (at 400 µg/mL) was distinguished for methanolic extracts (22.06%, 32.04%) and ethyl acetate fractions (16.59%, 14.40%) from S. annuus and S. perennis. Compounds 6-9 displayed moderate inhibitory activity, with IC50 values ranging from 39.59-73.86 µM.

Sambucus nigra L. leaves inhibit TNF-α secretion by LPS-stimulated human neutrophils and strongly scavenge reactive oxygen species.

ETHNOPHARMACOLOGICAL RELEVANCE: Sambucus nigra (elderberry) leaves were used in folk medicine to treat skin inflammations, ulcers, burns or boils, as well as to treat wounds, including infected and chronic ones. For centuries, elderberry leaves have been used mainly in eastern and southern Europe, as well as in western Asia. AIM OF THE STUDY: The study aimed to investigate the anti-inflammatory and antioxidant activity of four different extracts, such as aqueous and ethanolic prepared at room temperature and the solvent's boiling point, from the leaves of elderberry. MATERIALS AND METHODS: The effect of extracts both on the secretion of cytokines (TNF-α, IL-1ß, and IL-8) and reactive oxygen species (ROS) by neutrophils stimulated with bacteria-derived products was investigated. The cytotoxicity of extracts was analyzed by staining with propidium iodide measured by flow cytometry. The anti-inflammatory activity of extracts was also investigated through their influence on lipoxygenase activity. The antioxidant properties, including scavenging superoxide anion, hydrogen peroxide, nitric oxide, and 2,2-diphenyl-1-picrylhydrazyl radical were investigated in cell-free systems. The total content of phenolic compounds was tested using the Folin-Ciocalteu reagent. The qualitative and quantitative determination of the content of individual phenolic acids and flavonoids was performed by HPLC-DAD-MSn and HPLC-DAD method, respectively. RESULTS: Elderberry leaves extracts turned out to affect the inflammatory response of neutrophils by inhibiting the secretion of TNF-α and ROS. The ethanolic and aqueous extracts at a concentration of 50 µg × mL-1 reduce the secretion of TNF-α by approximately 40% and 10%, respectively. ROS secretion was decreased by around 50% for all extracts at concentration of 5 µg × mL-1. All the extracts were able to inhibit the activity of lipoxygenase. The ethanolic extracts were characterized by a higher content of phenolic compounds and a higher antioxidant activity, especially against nitric oxide, compared to the aqueous extracts. CONCLUSIONS: Our research has confirmed that elderberry leaves are a plant material with anti-inflammatory activity, especially against reactive oxygen species, and a potentially rich source of antioxidants. Preliminary analyses performed in this study could be the first step in confirming the traditional use of elderberry leaves in relieving inflammation.