Evaluating birch leaf tea as a functional herbal beverage: Beneficial impact on the urinary tract, and metabolism in human organism.

Herbal teas are considered as a potential constituent of novel functional beverages consumed daily. One of the commonly used herbal teas is silver birch (Betula pendula Roth) leaf infusion, traditionally used in urinary tract diseases. In this study, the potential of birch leaf infusion as a functional beverage, emphasizing its active ingredients' bioavailability, anti-inflammatory, and antiadhesive properties concerning urinary tract health, was investigated. A complex approach was proposed, which included phytochemical screening, bioavailability, gut microbiota biotransformation, and an in vivo test for urine metabolomics assessment. The bioassays confirmed significant anti-inflammatory (interleukins IL-6 and IL-8 secretion) and anti-adhesive (Uropathogenic Escherichia coli and T24 bladder cells) activities. The high-resolution mass spectrometry metabolomics studies linked gut microbiota metabolites and the metabolites present in the urine. Several metabolites connected with phenolics' consumption were detected in the urine, e.g., glucuronides and sulfates of caffeic acid and dihydroxyphenyl-γ-valerolactones. Based on the presented results, the birch leaf should be considered useful in designing functional beverages, especially targeted to the groups at high risk of urinary diseases.

Heterogeneity of transport and metabolism of Tormentillae rhizoma constituents across human intestinal epithelium cellular model.

Tormentilla erecta (L.) Raeusch is a widespread plant in Europe and Western Asia. Its rhizomes (Tormentilae rhizoma) are the main ingredient of herbal alcoholic beverages and can be used as a natural preservative in beer production. Apart from its unique taste qualities, therapeutic properties in gastrointestinal tract ailments are attributed to the tincture obtained from Tormentillae rhizoma. The presented research aimed to determine the mutual relationship between the components of Tormentillae tincture, present in popular alcoholic beverages, and intestinal epithelium (Caco-2 cell monolayers). A comprehensive qualitative and quantitative analysis of the tincture was performed, including the determination of condensed and hydrolyzable tannins as well as triterpenoids (UHPLC-DAD-MS/MS). Incubation of the tincture with Caco-2 monolayers has shown that only triterpenes pass through the monolayer, while condensed tannins are mainly bound to the monolayer surface. Ellagic acid derivatives were the only components of the Tormentillae tinctura being metabolized by cell monolayers to the compounds not previously described in the literature, which may be crucial in the treatment of intestinal diseases with inflammatory background.

Antagonistic properties against Fusarium sporotrichioides and glycosylation of HT-2 and T-2 toxins by selected Trichoderma strains.

The present study assessed the ability of Trichoderma to combat F. sporotrichioides, focusing on their antagonistic properties. Tests showed that Trichoderma effectively inhibited F. sporotrichioides mycelial growth, particularly with T. atroviride strains. In co-cultures on rice grains, Trichoderma almost completely reduced the biosynthesis of T-2 and HT-2 toxins by Fusarium. T-2 toxin-α-glucoside (T-2-3α-G), HT-2 toxin-α-glucoside (HT-2-3α-G), and HT-2 toxin-ß-glucoside (HT-2-3ß-G) were observed in the common culture medium, while these substances were not present in the control medium. The study also revealed unique metabolites and varying metabolomic profiles in joint cultures of Trichoderma and Fusarium, suggesting complex interactions. This research offers insights into the processes of biocontrol by Trichoderma, highlighting its potential as a sustainable solution for managing cereal plant pathogens and ensuring food safety.

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.

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.

Effect of in vitro cultivation on human gut microbiota composition using 16S rDNA amplicon sequencing and metabolomics approach.

Gut microbiota (GM) plays many key functions and helps maintain the host's health. Consequently, the development of GM cultivation under in vitro stimulating physiological conditions has gained extreme interest in different fields. In this study, we evaluated the impact of four culture media: Gut Microbiota Medium (GMM), Schaedler Broth (SM), Fermentation Medium (FM), and Carbohydrate Free Basal Medium (CFBM) on preserving the biodiversity and metabolic activity of human GM in batch in vitro cultures using PMA treatment coupled with 16S rDNA sequencing (PMA-seq) and LC-HR-MS/MS untargeted metabolomics supplemented with GC-MS SCFA profiling. Before the experiments, we determined the possibility of using the pooled faecal samples (MIX) from healthy donors (n = 15) as inoculum to reduce the number of variables and ensure the reproducibility of in vitro cultivation tests. Results showed the suitability of pooling faecal samples for in vitro cultivation study. Non-cultured MIX inoculum was characterized by higher α-diversity (Shannon effective count, and Effective microbial richness) compared to inocula from individual donors. After 24 h of cultivation, a significant effect of culture media composition on GM taxonomic and metabolomic profiles was observed. The SM and GMM had the highest α-diversity (Shannon effective count). The highest number of core ASVs (125) shared with non-cultured MIX inoculum and total SCFAs production was observed in the SM. These results might contribute to the development of standardized protocols for human GM in vitro cultivation by preventing methodological bias in the data.

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.

Gut Microbiota of Pigs Metabolizes Extracts of Filipendula ulmaria and Orthosiphon aristatus-Herbal Remedies Used in Urinary Tract Disorders.

Urinary tract infections influence the mortality rate in pigs and are linked to extensive antibiotic usage in the farm industry. Filipendula ulmaria (L.) Maxim. and Orthosiphon aristatus (Blume) Miq. are widespread medicinal plants traditionally used to treat urinary tract disorders. As their preparations are orally administered, the metabolism of their constituents by gut microbiota before absorption should be considered. Until now, no experiments had been performed to describe the biotransformation of tthose plants' extracts by animal gut microbiota. The study evaluates the influence of pig intestinal microbiota on the structure of active compounds in flowers of F. ulmaria and leaves of O. aristatus. The incubations of the extracts with piglet gut microbiota were performed in anaerobic conditions, and the samples of the batch culture were collected for 24 h. In F. ulmaria, the main metabolites were quercetin and kaempferol, which were products of the deglycosylation of flavonoids. After 24 h incubation of O. aristatus extract with the piglet gut microbiota, 2 main metabolites were observed. One, tentatively identified as 3-(3-dihydroxyphenyl)propionic acid, is likely the primary metabolite of the most abundant depsides and phenolic acids. The results confirm the formation of the compounds with anti-inflammatory and diuretic activity in the microbiota cultures, which might suggest F. ulmaria and O. aristatus for treating urinary tract disorders in piglets. Based on the similarities of human and pig gut microbiota, the pig model can help estimate the metabolic pathways of natural products in humans.

Antiadhesive activity of hydroethanolic extract from bean pods of Phaseolus vulgaris (common bean) against uropathogenic E. coli and permeability of its constituents through Caco-2 cells monolayer.

ETHNOPHARMACOLOGICAL RELEVANCE: Phaseaoli pericarpium (bean pods) is a pharmacopeial plant material traditionally used as a diuretic and antidiabetic agents. Diuretic activity of pod extracts was reported first in 1608. Since then Phaseoli pericarpium tea figures in many textbooks as medicinal plant material used by patients. AIM OF THE STUDY: Despite the traditional use of extracts from Phaseolium vulgaris pericarp, limited information is available on bioactivity, chemical composition, and bioavailability of such preparations. The following study aimed to investigate the phytochemical composition, the in vitro permeability of selected extract's constituents over the Caco-2 permeation system, and potential antivirulence activity against uropathogenic Escherichia coli of a hydroalcoholic Phaseoli pericarpium extract (PPX) in vitro to support its traditional use as a remedy used in urinary tract infections. MATERIAL AND METHODS: The chemical composition of the extract PPX [ethanol:water 7:3 (v/v)] investigated by using UHPLC-DAD-MSn and subsequent dereplication. The permeability of compounds present in PPX was evaluated using the Caco-2 monolayer permeation system. The influence of PPX on uropathogenic E. coli (UPEC) strain NU14 proliferation and against the bacterial adhesion to T24 epithelial cells was determined by turbidimetric assay and flow cytometry, respectively. The influence of the extract on the mitochondrial activity of T24 host cells was monitored by MTT assay. RESULTS: LC-MSn investigation and dereplication, indicated PPX extract to be dominated by a variety of flavonoids, with rutin as a major compound, and soyasaponin derivatives. Rutin, selected soyasaponins and fatty acids were shown to permeate the Caco-2 monolayer system, indicating potential bioavailability following oral intake. The extract did not influence the viability of T24 cells after 1.5h incubation at 2 mg/mL and UPEC. PPX significantly reduced the bacterial adhesion of UPEC to human bladder cells in a concentration-dependent manner (0.5-2 mg/mL). Detailed investigations by different incubation protocols indicated that PPX seems to interact with T24 cells, which subsequently leads to reduced recognition and adhesion of UPEC to the host cell membrane. CONCLUSIONS: PPX is characterised by the presence of flavonoids (e.g. rutin) and saponins, from which selected compounds might be bioavailable after oral application, as indicated by the Caco-2 permeation experiments. Rutin and some saponins can be considered as potentially bioavailable after the oral intake. The concentration-dependent inhibition of bacterial adhesion of UPEC to T24 cells justifies the traditional use of Phaseoli pericarpium in the prevention and treatment of urinary tract infections.

Pancreatic lipase and α-amylase inhibitory activity of extracts from selected plant materials after gastrointestinal digestion in vitro.

A screening of inhibitory activity of α-amylase, as well as pancreatic lipase (PL), under the influence of aqueous and ethanolic preparations from 12 plant materials was performed. The most active aqueous extracts from the fruits of Chaenomeles japonica (CJ) and Hippophaë rhamnoides (HR) were selected for artificial gastrointestinal digestion (GID). The aim of this study was to evaluate the inhibitory effect of the fractions obtained after GID on PL and α-amylase activities using a fluorescence assay. The changes in the composition of crude extracts in GID aliquots were followed by analysis with HPLC-DAD-MSn method in order to indicate active constituents. The main constituents of CJ and HR extracts were procyanidins and isorhamnetin derivatives, respectively. The most abundant compounds of extracts were found in all compartments of the digestion model correlated with relevant lipase/α-amylase inhibitory activity. What is more, the gastric and intestinal fractions inhibited enzymatic activity by at least 40%.

Gut microbiota metabolism and the permeability of natural products contained in infusions from herb of European goldenrod Solidago virgaurea L.

ETHNOPHARMACOLOGICAL RELEVANCE: Solidago virgaurea L. (also known as European goldenrod) is a pharmacopoeial plant material popularly used by patients in the form of an infusion. It was traditionally used in Europe and North America for the treatment of urinary tract conditions. It is also reported as a topical agent for skin disorders. AIM OF THE STUDY: Gut microbiota metabolism plays a crucial role in the bioavailability of natural products contained in plant extracts taken orally. The aim of the current study was to establish the biotransformation of compounds contained in an infusion from goldenrod using human and piglet fecal microbiota in vitro. The permeability of unmetabolized natural products and gut microbiota metabolites was evaluated using a Caco-2 cell model. Preliminary anti-inflammatory assays of raw extract using human neutrophils were also established. MATERIAL AND METHODS: An infusion was prepared from Solidaginis virgaureae herba commercially available on the market. The characterization of the raw extract was performed by UHPLC-DAD-MS method. The infusion was incubated with human or swine fecal samples in anaerobic conditions. Metabolism products were analyzed and identified by UHPLC-DAD-MS technique. The permeability of the natural products contained in the raw infusion and after metabolism was checked by UHPLC method. The influence of raw extracts on proinflammatory functions of human neutrophils after LPS stimulation was established by flow cytometry and ELISA. RESULTS: The experiments showed that goldenrod infusion contains mainly caffeoylquinic acid derivatives, flavonoids, and some phenylpropanoids. Natural products present in the extract were transformed by human and swine microbiota to smaller molecules mainly phenylpropanoid acid derivatives. The permeability assays showed that most of the parental compound present in the infusion cannot cross the gut epithelial barrier. In contrast, metabolites were able to cross the Caco-2 monolayer. Depending on the structure, different possible mechanisms of transport were observed. The infusion did not significantly influence the proinflammatory functions of human neutrophils. CONCLUSIONS: Following oral administration of goldenrod infusion, phytochemicals are prone to undergoing metabolism by gut microbiota to smaller phenylpropionic acid derivatives that can be bioavailable after crossing the gut epithelial barrier to be further metabolized and distributed. Detected metabolites should be considered as potentially active compounds responsible for the bioactivity of the raw plant material in vivo.

High molecular pyrogens present in plant extracts interfere with examinations of their immunomodulatory properties in vitro.

The widely accepted strategy to justify the use of medicinal plant extracts in diseases with inflammatory background is their examination on in vitro models using immune cells. It is also a key initial step of research for active principles, which could be then isolated and tested on more advanced models, becoming new pharmacologically active lead molecules. The crucial aspect which has not been so far addressed in this context, is the presence of pyrogens in plant preparations. The aim of this study was the examination of pyrogens interference with in vitro evaluation of anti-inflammatory activity of plant extracts using human primary neutrophils model together with introduction of effective method of interfering factors elimination. The obtained results showed that chosen plant extracts contained pyrogens, which were responsible for concentration-dependent stimulation of pro-inflammatory cytokines production by human neutrophils in vitro in the same extent as LPS did. The ultrafiltration method was successfully applied for pyrogens elimination, which effectiveness was confirmed using LAL test. The determined interference of pyrogens implies the necessity of their consideration and removal when in vitro studies include direct addition of plant extracts to the cell culture, what can be obtained by ultrafiltration, which does not affect extract composition.

Lamalbid, Chlorogenic Acid, and Verbascoside as Tools for Standardization of Lamium album Flowers-Development and Validation of HPLC-DAD Method.

Preparations from the flowers or herb of the white dead nettle (Lamium album L.) are recommended for the treatment of upper respiratory tract disorders or as a topical medication for mild inflammation of the throat, mouth, and skin. Taking into consideration the significance of L. album in traditional medicines across Europe, as well as the lack of studies describing the quantities of their most abundant constituents, we aimed to design a high-performance liquid chromatography coupled with diode-array detection (HPLC-DAD) method for potential standardization procedures of extracts from flowers of L. album. The HPLC-DAD method was developed and validated for quantification of iridoids (lamalbid), phenolic acids/depsides (chlorogenic acid), phenylpropanoids (verbascoside), and flavonoids (rutin; quercetin malonylhexoside; tiliroside) in aqueous and ethanolic-aqueous extracts of Lamii albi flos. The method was specific, accurate, and precise. Lamalbid was the most abundant compound both in aqueous (39.09 ± 1.02 m/g dry weight) and ethanolic-aqueous (26.66 ± 0.64 m/g dry weight) extracts. The quantities of selected compounds, except for chlorogenic acid and tiliroside, were higher in the aqueous extract than in the ethanolic-aqueous one. In conclusion, the method developed allowed for quantitation of compounds from different classes. In particular, chlorogenic acid and verbascoside have been proposed as reference compounds for routine quantitative control of Lamii albi flos.

Ionic liquids as a key medium for efficient extraction of copper complexes from chia seeds (Salvia hispanica L.).

Due to insufficient information, the aim of study was to concern on the optimization of extraction procedure of selected metal complexes with flavonoids from chia seeds. Evaluation of the amount of elements in compound, not only their total concentration content, is highly important due to the fact, that only a part from total content of metal is absorbed by human body. At the beginning the total amount of elements in chia seeds was established as 14.51±0.42 µg g(-1) for copper, 57.44±1.23 µg g(-1) for manganese, 81.12±1.89 µg g(-1) for zinc and 0.35±0.13 µg g(-1) for cobalt. After the most suitable solvent was established, effects of several parameters on the efficiency of metal extraction were studied. Solvent concentration, solid-solvent ratio, extraction method, extraction time and temperature have been investigated as independent variables. The optimal extraction conditions included vortexing during 20 min in 50°C, using an ionic liquid (1-butyl-3-methylimidazolium bromide) as an extractant, with solid-solvent ratio of 1:20. The determination of total and extractable amount of metals in chia seeds was carried out by standalone ICP MS. In addition, a complementary analysis of extracted metal complexes was performed using SEC-ICP MS method. It was confirmed that the ionic liquid is able to extract different copper complexes in comparison with commonly used solvents. The study indicated that extraction by using an ionic liquid has been successfully applied for determination of metals and metal complexes in chia seeds.