Many herbal products have a long history of use, but there are increasing concerns over product efficacy, safety and quality in the wake of recent cases exposing discrepancies between labeling and constituents. When it comes to St. John’s wort (Hypericum perforatum L.) herbal products, there is limited oversight, frequent off-label use and insufficient monitoring of adverse drug reactions. In this study, we use amplicon metabarcoding (AMB) to authenticate 78 H. perforatum herbal products and evaluate its ability to detect substitution compared to standard methods using thin-layer chromatography (TLC) and high performance liquid chromatography coupled with mass spectrometry (HPLC-MS). Hypericum perforatum was detected in 68% of the products using AMB. Furthermore, AMB detected incongruence between constituent species and those listed on the label in all products. Neither TLC nor HPLC-MS could be used to unambiguously identify H. perforatum. They are accurate methods for authenticating presence of the target compounds, but have limited efficiency in detecting infrageneric substitution and do not yield any information on other plant ingredients in the products. Random post-marketing AMB of herbal products by regulatory agencies could raise awareness among consumers of substitution and would provide an incentive to manufacturers to increase quality control from raw ingredients to commercialized products.

Antennaria dioica (L.) Gaertn. and Helichrysum arenarium (L.) Moench. are two species of the Asteraceae family, known in Romanian traditional medicine for their diuretic, choleretic, and anti-inflammatory properties. The aim of the present study was to evaluate the phenolic and sterolic composition of flowers from the two species and to assess their antioxidant, antibacterial and antifungal properties. LC-MS analyses were performed on methanolic, ethanolic and 70% v/v ethanolic extracts, before and after acid hydrolysis, and revealed high amounts of polyphenols. Chlorogenic acid was found as the main compound for the flowers of A. dioica (502.70 ± 25.11 mg/100 g d.w.), while quercitrin was dominant in H. arenarium (424.28 ± 21.21 mg/100 g d.w.) in 70% v/v ethanolic extracts before hydrolysis. Antioxidant capacity assays showed an important antioxidant potential, which can be correlated with the determined polyphenolic compounds, showing the 70% v/v ethanolic extracts of the two species as being the most effective antioxidant samples for the DPPH assay. Antibacterial and antifungal assays confirm a modest biological potential for the same extract of both species. Results obtained in the present study bring important data and offer scientific evidence on the chemical composition and on the biological activities of the flowers belonging to the two species.

Even though Salvia genus is one of the most known and studied taxa of Lamiaceae family, the knowledge regarding the chemical composition and health-related benefits of some locally used Salvia species (mostly endemic) is still scarce. In this regard, the present work aims to evaluate the chemical profile and potential bioactivities of 70% (v/v) ethanolic extracts obtained from the less-studied S. transsylvanica and S. glutinosa in comparison with S. officinalis. HPLC-PDA analysis revealed the presence of rutin and catechin as the main compounds in the extracts of the three studied species (using the employed HPLC method), whereas the presence of naringenin was highlighted only in S. glutinosa extract. Chlorogenic acid, rutin and quercetin were identified and quantified for the first time in S. transsylvanica extracts. The in vitro antioxidant capacity of each extract was tested through complementary methods (phosphomolybdenum assay, DPPH, ABTS, CUPRAC and FRAP assays), and correlated with the presence of phenolics (especially flavonoids) in high amounts. The neuroprotective and antidiabetic abilities of S. officinalis (the most active as AChE, BChE and α-glucosidase inhibitor), S. glutinosa (the most active as α-amylase inhibitor) and S. transsylvanica were also studied. For each extract it was determined the antimicrobial, antifungal and cytotoxic effects using in vitro assays. The obtained results confirm the potential of S. transsylvanica and S. glutinosa as promising sources of bioactive compounds and as a starting point for further analyses.

Romanian traditional medicine describes the use of aerial parts of Fumaria species to treat hepatobiliary diseases as well as diuretic agents. The present study aims to investigate the chemical composition, antioxidant properties, and diuretic effects of several Fumaria species. LC/MS analysis revealed that Fumaria species contain phenolic acids and high amounts of flavonoids with rutin and isoquercitrin as main compounds. Concerning antioxidant capacity, the most significant results were obtained for F. capreolata and F. vailantii. Both species showed a good correlation between the antioxidant capacity and a high amount of flavonoids. Furthermore, the extracts of F. officinalis and F. schleicheri produced a strong increase in urinary volumetric excretion of saline-loaded rats, 24 h after the oral administration of a single dose of 250 mg/kg bw. Moreover, both extracts of F. officinalis and F. schleicheri increased the urinary excretion of Na+ and K+. Results from the present study offer a new perspective concerning the chemical composition and bioactivities of traditionally used fumitory species.

Cherry stems (CS) represent a by-product intensively used in Eastern European countries as a traditional remedy for urinary tract disorders. Ethnopharmacological evidences sustain the use of CS as aqueous preparations (infusion and decoction), but few data were previously reported about phytochemical profile and pharmacological potential of CS hydroalcoholic extracts. In this regard, we aimed to evaluate the phenolic profile, in vitro antioxidant and tyrosinase inhibitory potential, and in vivo diuretic activity of 70% hydroethanolic cherry stems extract and cherry stems decoction (CSD). LC-DAD-ESI/MSn analysis revealed the presence of flavonoid-type compounds as main constituents for both preparations, especially flavanones (naringenin glycosides). Antioxidant activity evaluated through DPPH, ABTS, and FRAP methods was superior for cherry stems extract, probably due to the presence of phenolic-derived compounds in higher amounts than CSD. On the other hand, tyrosinase inhibitory potential and diuretic effect exerted by CSD were stronger, highlighting that other types of hydrophilic secondary metabolites are responsible for this bioactivity. Overall, our findings indicate that CS preparations could be used as promising mild diuretic agents and encourage further investigations regarding the correlation between their chemical composition and bioactive potential.

Objectives The aim of the present study consisted in the isolation of flavonoids from the leaves of Bryonia alba L. and evaluation of their antioxidant activity and inhibition on peroxidase‐catalysed reactions. Methods Flavonoids were isolated by preparative HPLC‐DAD and their structures were elucidated by MS and NMR. Inhibitory effect was tested by the horseradish peroxidase and the myeloperoxidase assays. Cellular antioxidant assays consisted in testing the inhibitory activity on the reactive oxygen species released upon activation of neutrophils freshly isolated ex vivo from equine blood and of human monocytes‐derived macrophages in vitro. Whole organism toxicity was assessed on zebrafish larvae. Key findings Four flavonoids (lutonarin, saponarin, isoorientin and isovitexin) were isolated. The performed assays showed significant antioxidant activity and inhibition for the peroxidase‐catalysed reactions. Absence of cellular and zebrafish toxicity was confirmed. Conclusions Bryonia alba L. leaves are particularly interesting for their flavonoids content and showed significant inhibitory effect on peroxidase‐catalysed oxidation of substrates (Amplex Red and L012), as well as antioxidant/antiradical activity, proving that this species has a medicinal potential. Moreover, the present study highlights the absence of the toxicity of these leaves and offers though a novel perspective on the species, previously known as being toxic.

Bryonia alba L. is the only Bryonia species found in Romanian flora, being known as a remedy for inflammatory pathologies or for its hepatoprotective and adaptogen activities. The present investigation studied the flavonoid composition and antioxidant activities of the aerial parts of this species. Flavonoid profile was evaluated by HPLC coupled with Diode Array Detection (DAD), while antioxidant capacity was assessed by various methods, testing different antioxidant mechanisms: DPPH (2,2-diphenyl-1-picrylhydrazyl), CUPRAC (cupric reducing antioxidant capacity), FRAP (ferric reducing ability of plasma), TEAC (Trolox equivalent antioxidant capacity), EPR (electron paramagnetic resonance method) and SNPAC (silver nanoparticles antioxidant capacity). Cytotoxicity was tested on human cancerous and healthy cell lines. Anti-plasmodial tests were performed on two strains of Plasmodium falciparum. Whole organism toxicity was assessed on zebrafish larvae. The HPLC-DAD analysis proved the presence of lutonarin, saponarin, isoorientin, and isovitexin as the major flavonoids in the composition of tested samples. Significant results were obtained for all antioxidant capacity assays. The cytotoxicity tests proved the absence of cellular and parasitic toxicity and these results were confirmed by the lack of toxicity on the zebrafish larvae model. This study proves a promising potential of the aerial parts of Bryonia alba L. as antioxidant agents.

& A new rapid, sensitive UPLC method coupled with ESI-MS=MS detection was developed and validated for the simultaneous quantification of six phytoestrogens (daidzin, genistin, genistein, daidzein, formononetin, and coumestrol) from Genista tinctoria L. extracts. An adequate chromatographic separation was achieved with gradient elution and a mobile phase consisting of a mixture of 0.1% acetic acid=methanol (70=30, v=v). The total run time was of 5.5 min. The detection of target compounds was performed in multiple reaction monitoring mode. The method was validated and proved to be linear, accurate, and precise over the range of 10-800 ng=mL. The LLOQ values estimated based on the signal to noise ratio were of 5 ng=mL for each analyte, except for daidzein (LLOQ ¼ 10.78 ng=mL). The elaborated assay shows shorter run time, lower solvent consumption, and higher sensitivity than most of the existing LC methods described in the literature. The method was used to quantify the selected phytoestrogens in Genista tinctoria herba or flos extracts. The major compounds found were genistein and its glycosidic form genistin. The method proved to be adequate for the characterization of plant extracts from the point of view of their phytoestrogen content, a group of active compounds with great therapeutic potential.