Search Results (148)

Thyroid hormones (THs) are essential regulators of human brain development, and disrupted TH availability during pregnancy or early life is linked to adverse neurodevelopmental outcomes. Concerns that environmental chemicals interfere with TH signalling have increased the need for human-relevant in vitro systems to identify thyroid hormone system-disrupting chemicals (THSDCs) for risk assessment. Here, we compared two human-induced pluripotent stem cell (hiPSC)-derived brain organoid models for THSDC assessment: (i) human cortical organoids (COs) generated by unguided differentiation, offering higher architectural complexity but lower throughput; and (ii) neural stem cell-derived organoids (NSCOs), designed for scalability with reduced cellular diversity. Both models expressed key TH handling components, including the transporter SLC16A2 (MCT8) and the inactivating enzyme DIO3. Using LC–MS/MS, we show that exogenous T3 is depleted from culture media and metabolized to 3,3′-T2 and 3′-T1 in both models, alongside upregulation of T3-responsive genes (HR, KLF9, DIO3, SEMA3C). Pulse and chronic co-exposures to reference disruptors iopanoic acid (IA, deiodinase inhibitor) and silychristin (SC, MCT8 inhibitor) altered T3 metabolism and modulated T3-responsive transcriptional endpoints. In NSCOs, high-content imaging revealed treatment-associated changes in cell composition, with chronic T3 reducing the SOX2-positive progenitor pool and THSDCs blocking this effect. Together, these findings provide a framework for organoid qualification—linking TH handling, transcriptomic responsiveness, and scalable phenotypic readouts—as a necessary step toward model validation and implementation of brain organoids in THSDC risk assessment pipelines. Full article

Objective: Cerebral ischemia–reperfusion (I/R) injury constitutes a pivotal pathological driver in cerebrovascular disorders such as stroke, yet effective therapeutic interventions remain scarce. This study explored whether hydrogen sulfide (H₂S) mitigates endothelial cell damage in the cerebral vasculature during I/R by modulating nicotinamide phosphoribosyltransferase (NAMPT) activity and its S-sulfhydration status, consequently restoring mitochondrial integrity and energetic homeostasis. Methods: Primary cerebrovascular endothelial cells (ECs) were subjected to hypoxia/reoxygenation (H/R) conditions in vitro, while rats experienced middle cerebral artery occlusion/reperfusion (MCAO/R) in vivo. The H₂S donor sodium hydrosulfide (NaHS) was administered, and outcomes were evaluated through Western blot analysis, S-sulfhydration assays, mitochondrial functional tests, autophagy profiling, and neurobehavioral assessments. The contributions of NAMPT and S-sulfhydration were validated using FK866 and dithiothreitol (DTT), respectively. LC-MS/MS was employed to identify candidate S-sulfhydration sites on NAMPT triggered by H₂S. Results: In cellular models, NaHS substantially boosted NAMPT enzymatic activity, elevated NAD⁺ and ATP levels, and enhanced cell survival. These protective benefits were nullified upon NAMPT inhibition with FK866 or reversal of S-sulfhydration via DTT. In animal studies, NaHS treatment significantly diminished infarct volume and ameliorated neurological deficits in MCAO/R rats; however, pretreatment with FK866 or DTT attenuated these benefits. Mechanistic investigations revealed that NaHS promoted S-sulfhydration of NAMPT, thereby activating autophagy of dysfunctional mitochondria. LC-MS/MS analysis confirmed enhanced S-sulfhydration at Cys39 and Cys397 residues of NAMPT following H₂S exposure. Conclusions: H₂S exerts neuroprotection against cerebral I/R injury in rats through S-sulfhydration-mediated activation of NAMPT, which improves mitochondrial performance and stimulates autophagy in cerebrovascular ECs. Full article

Phosphorus is an indispensable key element in life systems and materials science. Here in this work, several cyanoterphenyl-based phosphinic acid-bridged liquid crystal (LC) dimers of 2(CTOn)P (n = 6, 11) and their methyl esterification derivatives of 2(CTOn)P1E have been synthesized through hydrophosphination reaction followed by Suzuki coupling. The cyanoterphenyl LC dimers of 2(CTOn)P and methyl esterified 2(CTOn)P1E exhibit rich enantiotropic LC mesophases such as nematic (N), smectic A (SmA) and highly ordered smectic E (SmE), rather than the monotropic N or twist bend nematic (N_TB) displayed by the analogous phosphinic acid-bridged cyanobiphenyl LC dimers of 2(CBOn)P as reported previously. The phase transition temperatures of the cyanoterphenyl LC dimers 2(CTOn)P are also significantly higher than those of the cyanobiphenyl series, which is attributed to the larger π-conjugated system of cyanoterphenyl as compared with cyanobiphenyl, resulting in much enhanced π-π stacking interactions. However, the significantly enhanced interactions also make them extremely insoluble; thus, a different two-step synthesis pathway combining hydrophosphination with Suzuki coupling reactions was adopted. It is worth pointing out that by combining multiple characterization techniques, including DEPT 135°, ¹³C NMR, and HR-MS spectra, the definite molecular composition and structure of a byproduct with a third pro-mesogen attached via a branching alkyl spacer has been unambiguously demonstrated, which evidently deepens our understanding of the free radical-mediated hydrophosphination reaction mechanism, thereby providing valuable guidance for diminishing side reactions and achieving well-preparation of the high-purity phosphorus-containing LC dimers. Such phosphinic acid functionalized LC materials are envisioned to bear some unique application prospects. Full article

Inflammation is an immune response to foreign substances, pathogens, and cellular damage, characterized by redness, pain, and swelling. The use of synthetic anti-inflammatory drugs may cause adverse side effects, prompting the need for natural alternatives. This study aimed to evaluate the total flavonoid content and anti-inflammatory activity of single and combined extracts of sembung (Blumea balsamifera) [S], binahong (Anredera cordifolia) [B], and meniran (Phyllanthus niruri) [M] leaves. Extraction was performed using a 70% ethanol and water solution. Total flavonoid content was determined spectrophotometrically using quercetin as the standard. LC-HR-MS/MS results showed the presence of flavonoids, terpenoids, phenolics, and amino acids in each single extract (S, B, and M). Anti-inflammatory activity was assessed through the inhibition of albumin denaturation and the inhibition of cyclooxygenase-2 (COX-2). The highest flavonoid content (35.27 mg QE/g DW) in single extracts was found in B of the ethanolic extract, while the highest flavonoid content (40.24 mg QE/g DW) in all formulations was discovered in a combination of the ethanolic extract S:B:M (1:0:1). Moreover, the ethanolic extract S:B:M (1:0:1) gave the strongest inhibition (76.47%) of the albumin denaturation at 300 µg/mL and the strongest inhibition (98.5%) of the COX-2 inhibition at 100 µg/mL. Sembung extract was highest for reducing the expression of pro-inflammatory cytokines and TNF-α was 97.66% at an extract concentration of 15.625 ppm. As for S, S:B:M (1:0:1) and S:B:M (1:1:1) can reduce the expression of pro-inflammatory cytokines, as IL-6 was 100% in 0 pg/mL at an extract concentration of 15.625 ppm. Full article

The rising demand for sustainable and circular approaches in the agro-industrial sector has generated interest in repurposing herbal tea residues as sources of high-value bioactive compounds. This work focusses on recovering phytochemicals from Rosa canina L. peel and seed dust (by-products of processing of herbal tea in filter tea bags) using green extraction techniques. Two environmentally friendly technologies were used: ultrasound-assisted extraction (UAE) with a sonotrode and subcritical fluid extraction (SBFE). The extracts were qualitatively profiled using (HR) LC-ESI-QToF-MS/MS and quantified using HPLC-PDA. Both by-products contained phenolic substances, including gallic acid derivatives, ellagic acid, and flavonoids such as quercetin and quercetin-3-O-glucoside (only in the peel). Additionally, Folin–Ciocalteu’s assay was used to determine Total Phenolic content (TP). The extraction efficiency was considered in terms of phenolic compound recovery and total phenolic content obtained under the respective experimental conditions. The maximum TP for SBFE was reported in samples extracted with ethanol–water (48:52) at 180 °C, producing 3876.67 GAE mg/L for peel and 1648.57 GAE mg/L for seeds. In the UAE, extraction with ethanol–water (48:52) for 10 min yielded the maximum TP of 2773.81 GAE mg/L for peel and 957.86 GAE mg/L for seeds. These findings highlight the potential of R. canina infusion by-products as long-term sources of bioactive compounds for use in nutraceutical, cosmetic, and pharmaceutical industries. Full article

Background/Objectives: Propolis is a complex bee product with a composition that varies according to local vegetation, environmental conditions, and bee foraging behaviours. Recently, gas chromatography–mass spectrometry (GC–MS) has been employed in Uganda to analyse its volatile components. This study examined Ugandan propolis non-volatile metabolites to determine chemotypes and identify antibacterial compounds. Methods: Ethanolic extracts were analysed using liquid chromatography–high-resolution quadrupole time-of-flight mass spectrometry (LC-HR-QTOF-MS) in an untargeted MS/MS mode. Data processing was carried out using MZmine, then annotated with Global Natural Products Social Molecular Networking (GNPS) and SIRIUS. Chemometric methods assisted in identifying regional chemical signatures. Metabolites highlighted by the heatmap were evaluated for antibacterial activity using molecular docking against bacterial targets, followed by ADMET (absorption, distribution, metabolism, excretion, and toxicity) assessments. Results: Out of 3252 features, 234 and 52 putative compounds were annotated in GNPS and SIRIUS, respectively, as indicated by molecular networking, suggesting high chemical complexity. The chemical space mainly comprises flavonoids (including glycosides, aglycones, methylated, and prenylated derivatives), phenolic acids, amides, hydroxycinnamate derivatives, lignans, megastigmanes, and various diterpenoid skeletons. Multivariate analyses clearly distinguish geographical chemotypes, separating flavonoid-rich regions from diterpenoid-rich regions. Docking studies revealed flavonoids, diterpenoids, and lignans with strong predicted antibacterial activities and favourable ADMET profiles. Conclusions: This study provides the first LC–MS characterisation of the non-volatile metabolome of Ugandan propolis, thereby expanding its chemical diversity. Metabolomics and computational approaches lay a foundation for future ecological, chemotaxonomic, and pharmacological research. Full article

(−)-Epicatechin (EC), taxifolin (Tax), and quercetin (Q) are flavonoids with different C-ring structures. We compared their physicochemical properties and biological activities. The comparison of their stability and redox properties was conducted at mildly acidic or neutral conditions mimicking the plant vacuole or gut, and their sympathetic hyperactivation ability was examined using catecholamine (CA) excretion and blood flow. At pH 5, flavonoids were stable, but at pH 7, their retention rates decreased in the order EC > Q > Tax. LC-MS analysis detected brown oxidized oligomers in EC, while Tax and Q primarily yielded degradation products. All flavonoids exhibited O₂^•− scavenging activity after incubation at pH 5. At pH 7 after 45-min, EC and Tax promoted O₂^•−, while Q only scavenged O₂^•− after 24-hr incubation, EC’s properties decreased but Tax’s properties enhanced. Computational chemistry analysis indicated EC has higher reactivity compared to Tax and Q. EC caused a significant increase in CA excretion and blood flow, which were not observed with Tax or Q or 24-h cultured EC. These results suggest that the C-ring structure of flavonoids plays a crucial role in their stability and redox properties. Furthermore, reactive oxygen species generated by flavonoids in the gut may exert beneficial effects through sympathetic activation. Full article

Background: Globally, breast cancer is the most frequently diagnosed neoplasm among women, with an estimated 2.3 million new cases reported in 2022. Treatment for hormone receptor-positive (HR+) advanced breast cancer includes aromatase inhibitors and CDK4/6 inhibitors such as palbociclib. Objective: This study evaluated the bioequivalence and tolerability of two palbociclib capsule formulations to support the regulatory approval of a branded generic product in Latin America countries. Methods: Healthy participants were enrolled in an open-label, randomized, single-dose study using a two-treatment, two-sequence, two-period crossover design. Study participants received a single-dose test product, a palbociclib 125 mg capsule (Laboratório LKM S.A., Argentina), and a reference product, an Ibrance^® 125 mg capsule (Pfizer Manufacturing Deutschland GmbH), under fed conditions separated by a 14-day washout period. Blood samples were obtained at scheduled intervals over a 72 h period following administration, and the palbociclib plasma concentrations were determined using a validated LC-MS/MS method. Pharmacokinetic parameters were computed via non-compartmental analysis methods. A total of 52 healthy subjects were enrolled, and 50 subjects completed the study. Results: The geometric mean ratios (90% confidence intervals) for C_max and AUC_0–72 were 107.07% (101.98–112.42) and 109.77% (106.51–113.13), respectively. Conclusions: Both formulations were well-tolerated in healthy subjects. In accordance with regulatory standards, bioequivalence between the test formulation and the reference product was successfully demonstrated. Full article

Vicia faba L. is a widely cultivated legume known to contain numerous specialised metabolites. In this study, the seed coats and cotyledons of two ancient V. faba L. varieties, historically consumed in southern Italy and distinguished by black and purple seed coats, were extracted using 80% methanol and 80% ethanol. Extracts were analysed for total polyphenol, flavonoid and proanthocyanidin contents, and antioxidant activity using DPPH, ABTS, and FRAP assays. The purple seed coats exhibited the highest levels of phenolics and antioxidant capacity, exceeding those of black seed coats. Next, liquid chromatography coupled with high-resolution mass spectrometry (LC-ESI-HR-MS) was used to characterise the bioactive metabolites in both seed coats and cotyledons. The purple variety showed a higher phytochemical content, with a greater level of flavonoids and proanthocyanidins in methanolic extract. Furthermore, the purple seed coat exhibited in vitro anti-inflammatory activity by inhibiting soluble epoxide hydrolase (sEH), a key enzyme in the arachidonic acid cascade, with an IC₅₀ of 31.51 ± 1.16 µg/µL. Elemental analysis was performed for both varieties to assess their nutritional value. Specifically, the purple seed coats were found to represent a valuable source of bioactive compounds and micronutrients, highlighting their potential applications in nutraceutical, cosmetic, and food supplement sectors. Full article

Background/Objectives: Cytochrome P450 (CYP) enzymes are crucial for drug metabolism, yet inter-individual variability in their activity remains a significant clinical challenge. Current phenotyping methods are often impractical or even impossible, particularly in forensic toxicology and vulnerable populations. This proof-of-concept study investigated the feasibility of using in vitro assays with human liver microsomes (HLM) and recombinant CYP enzymes (isoenzymes), combined with untargeted metabolomics, to identify potential endogenous biomarker candidates indicative of CYP phenotype. Methods: This study uses in vitro incubations of HLM and isoenzymes in tandem with targeted and untargeted LC-(HR)MS and metabolomics techniques as well as statistical processing. Results: We demonstrate that HLM and isoenzymes maintain activity in the presence of complex biological matrices (blood/plasma), enabling metabolomic profiling. Untargeted analysis of assays in plasma revealed numerous potential biomarkers, with several showing significant correlations to enzyme activity. Conclusions: While identification remains the major challenge, this approach offers a promising avenue for developing accessible and efficient methods for indirect CYP phenotyping, potentially facilitating investigations in scenarios where traditional approaches are limited. This work provides a foundation for future studies focused on further developing in vitro assays and validating the proposed biomarkers, as well as establishing their utility in clinical and forensic settings. Full article

Agarwood from Aquilaria agallocha, known as chim-hyuang in Korea, is widely distributed throughout Southeast Asia and has traditionally been used to treat asthma, pain, and gastrointestinal disorders. As part of our ongoing efforts to identify bioactive metabolites from natural sources, a phytochemical investigation of the EtOAc fraction of A. agallocha extract led to the isolation and identification of four compounds, N-trans-feruloyltyramine (1), (3R,5R)-octahydrocurcumin (2), 1,7-bis(4-hydroxyphenyl)heptane (3), and trans-caffeoyltyramine (4), via HPLC purification and LC/MS-based analysis. Structural elucidation of the isolated compounds was achieved using NMR spectroscopy, LC/MS, and high-resolution electrospray ionization mass spectrometry (HR-ESIMS). The absolute configuration of compound 2 was further confirmed by optical rotation and electronic circular dichroism (ECD) analyses. All isolated compounds (1–4) were evaluated for their inhibitory activity against tau protein aggregation. Notably, compound 2 exhibited a 43.7% reduction in tau aggregation at 20 μM, without cytotoxicity at the same concentration. These findings indicate that phytochemicals from A. agallocha, particularly the diarylheptanoid compound 2, hold promise as natural lead candidates for the development of therapeutic agents targeting tau protein aggregation. Full article

6-Nitrodopamine is an endogenous catecholamine responsible for numerous biological activities. Here, an improved method for the synthesis of both 6-nitrohomovanillic acid (6-NHVA) and its regioisomer 5-nitrohomovanillic acid (5-NHVA) is reported. The developed one-step synthetic procedures ensured the efficient preparation of the target compounds in good yields. Comprehensive structural characterization was achieved through one- and two-dimensional NMR studies and by high-resolution mass spectrometry (HR-MS/MS). The presence of both substances was identified in human amniotic fluid by LC-MS/MS. Full article

Background: Glioblastoma is an aggressive brain tumor that invariably recurs despite treatment, partly due to metabolic adaptations, including altered lipid metabolism. This study investigates plasma lipidomic profiles in patients with glioblastoma to explore their potential as a liquid biopsy for disease monitoring. Methods: Plasma samples were collected from 36 patients with histopathologically confirmed IDH wild-type glioblastoma at four treatment stages: pre-surgery (n = 36), post-surgery (n = 32), pre-radiation (n = 28), and post-radiation (n = 17). Untargeted lipidomics analysis was performed using liquid chromatography-high resolution mass spectrometry (LC-HR-MS/MS). Results: Plasma lipidomic signatures differed significantly across treatment stages. Specifically, the lipidomic profile prior to surgery was statistically distinct from those at subsequent stages, demonstrating an increased compound abundance of numerous lipids that are decreased at subsequent stages, including linoleic acid (fold-change 2.58, p = 4.21 × 10⁻¹¹), behenic acid (fold-change 2.09, p = 9.3 × 10⁻¹⁰), and linolenic acid (fold-change 4.44, p = 5.83 × 10⁻⁶). Random forest modeling could predict pre-surgical samples with 85.7% accuracy. Conclusions: Plasma lipidomics shows promise as a potential liquid biopsy approach for monitoring glioblastoma treatment but future studies will need to examine these findings in larger and well-controlled cohorts. Full article

Vegetarians must rely on supplements to meet the recommended daily intake of vitamin B12. Therefore, it is essential to establish a rapid, inexpensive, and reliable method of determining B12 levels in order to accurately characterize and assess the quality of supplements. This study describes a methodology for quantifying vitamin B12 in the form of methylcobalamin and cyanocobalamin following 2 min of ultrasound-assisted extraction performed at pH 4. Vitamin B12 was determined using UV-Vis spectrophotometry and liquid chromatography-tandem mass spectrometry. Thus, LC-MS/MS validated the cost-effective UV-Vis method. The content and form of vitamin B12 in the tested supplements were investigated, and serious discrepancies were found in the content or form of vitamin B12 in three out of ten supplements. To verify the quality of the analyzed supplements, the presence of metal impurities (as Cd, Hg, and Pb) was also assessed using high-resolution continuum source electrothermal atomic absorption spectrometry. No risk associated with the presence of these metals has been noted. Nevertheless, our findings underscore the need for stricter quality control in supplement manufacturing. Full article

Olea europaea L. ‘Lavagnina’ is cultivated in the Eastern Ligurian coast (Italy), and during the pruning process a huge amount of pruning residues is produced. This by-product is generally disposed of by burning, despite still containing bioactive compounds. In particular, olive leaves are indeed rich in secondary metabolites, which can vary both in quality and quantity in relation to the cultivar considered and the area of cultivation. For this reason, we aimed to carry out a pharmacognostic study of the pruned leaves of the unexplored local cultivar ‘Lavagnina’, evaluating the possibility of reusing this by-product for new health applications. The micromorphological characterization was conducted by light and scanning electron microscopy. ‘Lavagnina’ leaf was micromorphologically similar to that of other olive cultivars; however, it differed in terms of midrib structure. Leaf extracts were obtained using solvents of increasing polarity (petroleum ether, chloroform, methanol) and the food-grade solvent, 70% ethanol. A high antioxidant activity was found only for the methanolic (ME) and hydroalcoholic (HAE) extracts, and, therefore, they were then characterized from a phytochemical point of view by LC-ESI-HR-MS. Such analysis allowed the identification of secondary metabolites belonging mainly to secoiridoids, flavonoids, and iridoids. Overall, the HAE had the highest antioxidant activity (17.3 ± 0.6 μg/mL), and it is, therefore, the best candidate for health applications related to a protective effect on a variety of inflammation-related diseases, also considering that inflammation may play a role in cancer progression. Full article