Identification of hydroxyquinazoline alkaloids from Justicia adhatoda L. leaves, a traditional natural remedy with NF-κB and AP-1-mediated anti-inflammatory properties and antioxidant activity.

ETHNOPHARMACOLOGICAL RELEVANCE: Justicia adhatoda L. is used as traditional medicine in Nepal to treat cough, asthma, and inflammatory disorders, and is indicated as "Asuro". Leaves are used worldwide as herbal medicine due to cardiotonic, expectorant, anti-asthmatic, and bronchodilatory properties. The aim of this work was to study the phytochemical composition of leaves of Nepalese J. adhatoda and assess their anti-inflammatory and antioxidant properties in vitro. MATERIALS AND METHODS: Secondary metabolites were extracted from dried leaves using methanol (JAME: J. adhatoda methanol extract). They were analysed by means of liquid chromatography coupled with multiple-stage mass spectrometry (LC-MSn). Anti-inflammatory potential was determined by the NF-κB and AP-1 inhibition assay, and DPPH, ABTS, and ß-carotene bleaching assays were performed to assess its antioxidant properties. RESULTS: JAME is a rich source of secondary metabolites, especially quinazoline alkaloids such as vasicine, vasicinone, vasicoline, and adhatodine. 7-Hydroxy derivatives of peganidine, vasicolinone, and adhatodine were also identified by means of MSn data and are here reported in J. adhatoda for the first time. JAME inhibited NF-κB and AP-1 expression in THP-1 cells to a greater extent than the positive control prednisolone. A moderate radical-quenching property was observed in DPPH and ABTS assays, but the anti-carotene bleaching activity was significantly higher than the reference BHT. CONCLUSIONS: To the best of our knowledge, this is the first insight into the phytochemical composition of Asuro leaves from Nepal and their bioactivity. Our results will contribute to the valorisation of this medicinal species still widely used in the traditional and complementary medicine.

Gene Silencing of Angiopoietin-like 3 (ANGPTL3) Induced De Novo Lipogenesis and Lipid Accumulation in Huh7 Cell Line.

Angiopoietin-like 3 (ANGPTL3) is a hepatokine acting as a negative regulator of lipoprotein lipase (LPL). Vupanorsen, an ANGPTL3 directed antisense oligonucleotide, showed an unexpected increase in liver fat content in humans. Here, we investigated the molecular mechanism linking ANGPTL3 silencing to hepatocyte fat accumulation. Human hepatocarcinoma Huh7 cells were treated with small interfering RNA (siRNA) directed to ANGPTL3, human recombinant ANGPTL3 (recANGPTL3), or their combination. Using Western blot, Oil Red-O, biochemical assays, and ELISA, we analyzed the expression of genes and proteins involved in lipid metabolism. Oil Red-O staining demonstrated that lipid content increased after 48 h of ANGPTL3 silencing (5.89 ± 0.33 fold), incubation with recANGPTL3 (4.08 ± 0.35 fold), or their combination (8.56 ± 0.18 fold), compared to untreated cells. This effect was also confirmed in Huh7-LX2 spheroids. A total of 48 h of ANGPTL3 silencing induced the expression of genes involved in the de novo lipogenesis, such as fatty acid synthase, stearoyl-CoA desaturase, ATP citrate lyase, and Acetyl-Coenzyme A Carboxylase 1 together with the proprotein convertase subtilisin/kexin 9 (PCSK9). Time-course experiments revealed that 6 h post transfection with ANGPTL3-siRNA, the cholesterol esterification by Acyl-coenzyme A cholesterol acyltransferase (ACAT) was reduced, as well as total cholesterol content, while an opposite effect was observed at 48 h. Under the same experimental conditions, no differences in secreted apoB and PCSK9 were observed. Since PCSK9 was altered by the treatment, we tested a possible co-regulation between the two genes. The effect of ANGPTL3-siRNA on the expression of genes involved in the de novo lipogenesis was not counteracted by gene silencing of PCSK9. In conclusion, our in vitro study suggests that ANGPTL3 silencing determines lipid accumulation in Huh7 cells by inducing the de novo lipogenesis independently from PCSK9.

Dibenzocyclooctadiene Lignans from Schisandra chinensis with Anti-Inflammatory Effects.

Schisandra chinensis (Schisandraceae) is a medicinal plant widely used in traditional Chinese medicine. Under the name Wu Wei Zi, it is used to treat many diseases, especially as a stimulant, adaptogen, and hepatoprotective. Dibenzocyclooctadiene lignans are the main compounds responsible for the effect of S. chinensis. As a part of ongoing studies to identify and evaluate anti-inflammatory natural compounds, we isolated a series of dibenzocyclooctadiene lignans and evaluated their biological activity. Furthermore, we isolated new sesquiterpene 7,7-dimethyl-11-methylidenespiro[5.5]undec-2-ene-3-carboxylic acid. Selected dibenzocyclooctadiene lignans were tested to assess their anti-inflammatory potential in LPS-stimulated monocytes by monitoring their anti-NF-κB activity, antioxidant activity in CAA assay, and their effect on gap junction intercellular communication in WB-ras cells. Some S. chinensis lignans showed antioxidant activity in CAA mode and affected the gap junction intercellular communication. The anti-inflammatory activity was proven for (-)-gomisin N, (+)-γ-schisandrin, rubrisandrin A, and (-)-gomisin J.

A Bronze Age lip-paint from southeastern Iran.

A small chlorite vial, discovered among numerous artifacts looted and recovered in the Jiroft region of Kerman province, southeastern Iran, contains a deep red cosmetic preparation that is likely a lip-coloring paint or paste. Through analytical research involving XRD (X-ray diffraction), SEM-EDS (scanning electron microscopy-energy-dispersive spectroscopy), and HPLC-MS (high-performance liquid chromatography-mass spectrometry) analyses, the mineral components of the reddish substance were identified as hematite, darkened with manganite and braunite, and traces of galena and anglesite, mixed with vegetal waxes and other organic substances. The mixture, thus observed, bears a striking resemblance to the recipes of contemporary lipsticks. We also report the first radiocarbon date ever obtained from a Bronze age cosmetic in the ancient Near East: results place the pigment in the early 2nd millennium BCE, a date compatible with several mentions of the powerful eastern-iranian civilization of Marhasi in coeval cuneiform texts of Mesopotamia, as well as with its currently emerging archaeological picture.

Advances in sample preparation methods for pesticide residue analysis in medicinal plants: A focus on Nepal.

Medicinal plant safety is a rising challenge worldwide due to the continued overuse of pesticides to their maximum residue limits. Due to the high demand for medicinal plants, their production is being increased and sometimes protected by pesticide use. The analysis of these residues requires robust analytical methods to ensure the safety and quality of medicinal plants. Developing effective sample preparation for detecting pesticides is challenging, due to their diverse natures, classes, and physico-chemical characteristics. Hence, existing techniques and strategies are needed to improve the reliability of the results. The review discusses the current state of sample preparation techniques, analytical methods, and instrumental technologies employed in pesticide residue analysis in medicinal plants. It highlights the challenges, limitations, and advancements in the field, providing insights into the analytical strategies used to detect and quantify pesticide residues. Reliable, accessible, affordable, and high-resolution analytical procedures are essential to ensure that pesticide levels in medicinal plants are effectively regulated. By understanding the complexities of pesticide residue analysis in medicinal plants, this review article aims to support the conservation of medicinal plant resources, promote public health, and contribute to the development of sustainable strategies for ensuring the safety and quality of medicinal plants in Nepal. The findings of this review will benefit researchers, policymakers, and stakeholders involved in the conservation of medicinal plant resources and the promotion of public health.

3D human foreskin model for testing topical formulations of sildenafil citrate.

Sildenafil citrate is an approved drug used for the treatment of erectile dysfunction and premature ejaculation. Despite a widespread application, sildenafil citrate shows numerous adverse cardiovascular effects in high-risk patients. Local transdermal drug delivery of this drug is therefore being explored as an interesting and noninvasive alternative administration method that avoids adverse effects arised from peak plasma drug concentrations. Although human and animal skin represents the most reliable models to perform penetration studies, they involve a series of ethical issues and restrictions. For these reasons new in vitro approaches based on artificially reconstructed human skin or "human skin equivalents" are being developed as possible alternatives for transdermal testing. There is little information, however, on the efficiency of such new in vitro methods on cutaneous penetration of active ingredients. The objective of the current study was to investigate the sildenafil citrate loaded in three commercial transdermal vehicles using 3D full-thickness skin equivalent and compare the results with the permeability experiments using porcine skin. Our results demonstrated that, while the formulation plays an imperative role in an appropriate dermal uptake of sildenafil citrate, the D coefficient results obtained by using the 3D skin equivalent are comparable to those obtained by using the porcine skin when a simple drug suspension is applied (1.17 × 10-10 ± 0.92 × 10-10 cm2/s vs 3.5 × 102 ± 3.3 × 102 cm2/s), suggesting that in such case, this 3D skin model can be a valid alternative for ex-vivo skin absorption experiments.

Bioactive agents from Parkia biglobosa (Jacq.) R.Br. ex G. Don bark extracts for health promotion and nutraceutical uses.

BACKGROUND: Parkia biglobosa stem bark extracts were prepared using methanol, methanol 80%, water and ethyl acetate to investigate their phytochemical contents, as well as antioxidant and enzyme inhibitory properties. RESULTS: Liquid chromatography (LC) quadrupole time-of-flight mass spectrometry (MS) and LC-MSn revealed the presence of flavonoids, hydroxycinnamic acid derivatives and gallotannins. Particularly, the water extract contained rutin (480 µg per 100 mg) and 3-caffeoylquinic acid (1109 µg per 100 mg) in higher amounts, whereas the 80% methanol extract contains methoxyluteolin-7-O-rutinoside and catechin derivatives as major compounds. Total phenolic and flavonoid contents of the extracts were yielded in the range of 32.26-119.88 mg gallic acid equivalents g-1 and 0.60-2.39 mg rutin equivalents g-1 , respectively. Total antioxidant capacity was also displayed in the range of 0.53-6.34 mmol Trolox equivalents (TE) g-1 . Both the methanolic extracts showed higher total antioxidant capacity that could be related to the total phenolic contents. Radical scavenging capacity in DPPH (2,2-diphenyl-2-picryl-hydrazyl) (37.21-508.30 mg TE g-1 ) and ABTS [2,2-azinobis(3-ethylbenzothiazoline- 6-sulfonic acid)] (60.95-1068.06 mg TE g-1 ) assays, reducing power in cupric ion reducing antioxidant capacity (54.23-1002.78 mg TE g-1 ) and ferric ion reducing antioxidant power (33.18-558.68 mg TE g-1 ) assays, as well as metal chelating activity (2.45-11.28 mg EDTA equivalents g-1 ), were exhibited by all extracts. All extracts were found to inhibit acetylcholinesterase [0.23-2.47 mg galanthamine equivalents (GALAE) g-1 ], tyrosinase [27.20-83.33 mg kojic acid equivalents g-1 ], amylase [mmol acarbose equivalents (ACAE) g-1 ]. On the other hand, all extracts, except the water extract, inhibited butyrylcholinesterase (5.38-6.56 mg GALAE g-1 ), whereas only the water and ethyl acetate extract showed glucosidase inhibitory potential (1.96 and 1.82 mmol ACAE g-1 ). In general, the water extract was found to be a weaker enzyme inhibitor suggesting that water is not the preferrable extraction solvent to obtain active products. CONCLUSION: The present study demonstrated that the stem bark extracts of P. biglobosa contains good amount of phytochemical and extracts present significant antioxidant, as well as reasonable enzyme inhibitory effects. Hence, these findings suggest that further studies can be performed on more specific biological targets and models of bioactivity to determine their safe usage as a nutraceutical or for the preparation functional foods. © 2023 The Authors. Journal of The Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Karanjin, A Promising Bioactive Compound Possessing Anti-cancer Activity against Experimental Model of Non-small Cell Lung Cancer Cells.

AIMS: The aim of this study is to isolate the Millettia pinnata (Karanj) leaf extract for pure compound with anticancer properties and to study the molecular target of the isolates in non-small cell lung cancer cell lines. BACKGROUND: In our earlier research Millettia pinnata leaf extract has demonstrated potential anticancer activities. Thus, in pursuit of the bioactive compounds, the most potential active extract from our previous study was purified. Furthermore, the anticancer properties of the isolated compound karanjin was studied and aimed for apoptosis and restraining growth. METHODS: A novel method was developed through column chromatography for isolation and purification of the compound karanjin from leaf chloroform extract. The purified component was then characterised using FTIR, mass spectrometry, and NMR. An MTT-based cytotoxicity assay was used to analyse cell cytotoxicity, whereas fluorescence staining was used for apoptosis and reactive oxygen species inhibition quantification. Furthermore, the real-time PCR assay was used to determine the molecular mechanism of action in cells causing cytotoxicity induced by karanjin dosing. RESULTS: The anticancer activity of karanjin in A549 cell line exhibited prominent activity revealing IC50 value of 4.85 µM. Conferring the predicted molecular pathway study, karanjin restrains the proliferation of cancer cells through apoptosis, which is controlled by extrinsic pathway proteins FAS/FADD/Caspases 8/3/9. Downregulation of KRAS and dependent gene expression also stopped cell proliferation. CONCLUSION: Karanjin has been identified as a compound with potential effect in non-small cell lung cancer cells. Molecular mechanism for apoptosis and inhibition of reactive oxygen species induced through H2O2 were observed, concluding karanjin have medicinal and antioxidant properties.

Uncovering chemical profiles, biological potentials, and protection effect against ECM destruction in H2 O2 -treated HDF cells of the extracts of Stachys tundjeliensis.

The genus Stachys L., one of the largest genera of the Lamiaceae family, is highly represented in Turkey. This study was conducted to determine the bio-pharmaceutical potential and phenolic contents of six different extracts from aerial parts of Stachys tundjeliensis. The obtained results showed that the ethanol extract exhibited the highest antioxidant activity in the antioxidant assays. Meanwhile, the ethanol extract displayed strong inhibitory activity against α-tyrosinase, the dichloromethane extract exhibited potent inhibition against butyrylcholinesterase, and the n-hexane extract against α-amylase. Based on ultra-high performance liquid chromatography coupled to high-resolution mass spectrometry analysis, more than 90 secondary metabolites, including hydroxybenzoic acid, hydroxycinnamic acid, and their glycosides, acylquinic acids, phenylethanoid glycosides, and various flavonoids were identified or tentatively annotated in the studied S. tundjeliensis extracts. It was observed that the application of S. tundjeliensis eliminated H2 O2 -induced oxidative stress. It was determined that protein levels of phospho-nuclear factor kappa B (NF-κB), receptor for advanced glycation endproducts, and activator protein-1, which are activated in the nucleus, decreased, and the synthesis of matrix metalloproteinase (MMP)-2 and MMP-9 also decreased to basal levels. Overall, these findings suggest that S. tundjeliensis contains diverse bioactive compounds for the development of nutraceuticals or functional foods with potent biological properties.

Phytochemical analysis of Clinopodium candidissimum (Munby) Kuntze growing in Algeria by an integrated HS-SPME-GC-MS, NMR and HPLC-DAD-MSn approach: valorisation of an endemic natural source of bioactive compounds.

Clinopodium candidissimum (Munby) Kuntze (Lamiaceae) is used in traditional medicine and as a food condiment in Algeria, where it is known as Zaater cheleuh and Nabta elbida. Here, we report the comprehensive characterisation of non-volatile polar constituents extracted from C. candidissimum aerial parts (a mixture of inflorescences, stems and leaves), and their aroma profile. Qualitative 1H-NMR and quali-quantitative HPLC-MSn analyses of fractions obtained with solvents at different polarity revealed the presence of aglyconic and glycosylated flavonoids (3.1%), phenylpropanoids (3.6%), gallic acid derivatives (0.76%), and triterpenoids (0.62%), among the others. On the other hand, HS-SPME-GC-MS allowed to identify 38 volatile constituents, among which the oxygenated monoterpenes pulegone (44.8%), piperitenone (6.6%), isopulegone (5.8%) and neo-menthol (3.8%), and the sesquiterpene hydrocarbons germacrene D (16.2%) and bicyclogermacrene (3.0%) were the most abundant. Overall, results indicate that C. candidissimum represents an endemic natural source of antioxidants and bioactive compounds, and they will be useful for further studies on this species.

The kynurenine pathway in treatment-resistant schizophrenia at the crossroads between pathophysiology and pharmacotherapy.

Two cardinal elements in the complex and multifaceted pathophysiology of schizophrenia (SCZ) are neuroinflammation and dysregulation of glutamatergic neurotransmission, with the latter being especially involved in treatment-resistant schizophrenia (TRS). Interestingly, the Kynurenine (KYN) pathway (KP) is at the crossroad between them, constituting a potential causal link and a therapeutic target. Although there is preclinical and clinical evidence indicating a dysregulation of KP associated with the clinical phenotype of SCZ, clinical studies investigating the possible relationship between changes in biomarkers of the KP and response to pharmacotherapy are still limited. Therefore, we have studied possible differences in the circulating levels of biomarkers of the metabolism of tryptophan along the KP in 43 responders to first-line treatments (FLR) and 32 TRS patients treated with clozapine, and their possible associations with psychopathology in the two subgroups. Plasma levels of KYN were significantly higher in TRS patients than in FLR patients, indicating a greater activation of KP. Furthermore, the levels of quinolinic (NMDA receptor agonist) and kynurenic acid (NMDA negative allosteric modulator) showed a negative and a positive correlation with several dimensions and the overall symptomatology in the whole sample and in FLR, but not in TRS, suggesting a putative modulating effect of clozapine elicited through the NMDA receptors. Despite the cross-sectional design of the study that prevents us from demonstrating causation, these findings show a significant relationship among circulating KP biomarkers, psychopathology, and response to pharmacotherapy in SCZ. Therefore, plasma KP biomarkers should be further investigated for developing personalized medicine approaches in SCZ.

Melatonin MT1 receptors as a target for the psychopharmacology of bipolar disorder: A translational study.

The treatment of bipolar disorder (BD) still remains a challenge. Melatonin (MLT), acting through its two receptors MT1 and MT2, plays a key role in regulating circadian rhythms which are dysfunctional in BD. Using a translational approach, we examined the implication and potential of MT1 receptors in the pathophysiology and psychopharmacology of BD. We employed a murine model of the manic phase of BD (Clock mutant (ClockΔ19) mice) to study the activation of MT1 receptors by UCM871, a selective partial agonist, in behavioral pharmacology tests and in-vivo electrophysiology. We then performed a high-resolution Nuclear Magnetic Resonance study on isolated membranes to characterize the molecular mechanism of interaction of UCM871. Finally, in a cohort of BD patients, we investigated the link between clinical measures of BD and genetic variants located in the MT1 receptor and CLOCK genes. We demonstrated that: 1) UCM871 can revert behavioral and electrophysiological abnormalities of ClockΔ19 mice; 2) UCM871 promotes the activation state of MT1 receptors; 3) there is a significant association between the number of severe manic episodes and MLT levels, depending on the genetic configuration of the MT1 rs2165666 variant. Overall, this work lends support to the potentiality of MT1 receptors as target for the treatment of BD.

Chemical Characterization of Corydalis chaerophylla D.C. Extracts and Preliminary Evaluation of Their in Vitro and in Vivo Biological Properties.

Genus Corydalis is a rich source of isoquinoline alkaloids reported to having potential bioactivities. Corydalis chaerophylla collected from Nepal at an altitude of 2400-4800 m was extracted using hexane, methanol and chloroform as solvents. The resulting hexane, methanol and chloroform extracts were subjected to LC-DAD-MSn analysis to yield fifteen different alkaloids. To assess any potential pharmacological properties, antimicrobial activity against two Gram-positive, two Gram-negative bacterial strains and one fungal strain was assessed, revealing significant inhibitive action of the methanol and chloroform extracts. Of the extracts obtained using chloroform contained the highest content of phenolic compounds at 113 mg GAE/g, while the highest total flavonoid content was found for the hexane extract with a value of 46.45 mg QE/g. The chloroform extract also exhibited a considerable antioxidant activity at IC50 value, 261.5±3 µg/mL, for the DPPH assay. Conversely, the methanol extract exhibited the highest LC50 value for Brine Shrimp cytotoxicity at 196±3 µg/mL being least potential for the test. The methanol extract was found to be the most active against α-amylase inhibition with an IC50 of 51.52±2 µg/mL. In an in vivo acute oral toxicity study against mice, methanol and chloroform extracts presented harmful effects with 1000.36 mg/kg BW and 515 mg/kg BW for LD50 , respectively. By analyzing all the results of the solvents used, the chloroform extract was found to be the most active, a feature that will be used in future isolation procedures and other pharmacological tests.

Nutraceutical formulation for immune system modulation: Active constituents, in vitro antibacterial and immunomodulatory activity, and metabolomics analysis.

There is a large demand for nutraceuticals in the market and studies related to their action are needed. In this paper, the antimicrobial activity and the immunomodulatory effect of a nutraceutical formulation containing 14.39% of ascorbic acid, 7.17% of coenzyme Q10, 1.33% of Echinacea polyphenols, 0.99% of pine flavan-3-ols, 0.69% of resveratrol and 0.023% of Echinacea alkylamides were studied using in vitro assays and cell-based metabolomics. Chromatographic analysis allowed us to study the nutraceutical composition. The antibacterial activity was evaluated on S. aureus, K. pneumoniae, P. aeruginosa, E. coli, H. influenzae, S. pyogenes, S. pneumoniae and M. catarrhalis. The immunomodulatory activity was assessed on human macrophages and dendritic cells. The production of IL-1ß, IL-12p70, IL-10 and IL-8 was evaluated on culture medium by ELISA and the activation/maturation of dendritic cells with cytofluorimetric analysis. Treated and untreated macrophages and dendritic cell lysates were analysed by liquid chromatography coupled with high-resolution mass spectrometry, and results were compared using multivariate data analysis to identify biological markers related to the treatment with the food supplement. The food supplement decreased K. pneumoniae, P. aeruginosa, E. coli, Methicillin-resistant Staphylococcus aureus (MRSA) and M. catharralis growth, reduced the inflammatory response in macrophages exposed to lipopolysaccharide (LPS) and modulated the activation and maturation of the dendritic cells. Oxidized phospholipids were identified as the main biological markers of treated cell lysates, compared with controls.

Effect of different atmospheric and subatmospheric cooking techniques on physical and chemical qualitative properties of pork loin.

In this paper, the effects of four cooking procedures were evaluated, two occurring in atmospheric (in ventilated and steam oven) and two in subatmospheric (vacuum and sous vide cooking) conditions on pork Longissimus lumborum. The main objective of the study was to compare and evaluate the physical and chemical characteristics. Samples were cooked in four independent trials namely Oven (O), Steaming (ST), Vacuum Cooking (VC) and Sous Vide (SV). The analyses included temperature, cooking effect, percentage weight loss, texture (cutting and double compression tests), colour (superficially and inside the sample), microstructure (optical microscopy) and fibres shortening analysis. To assess cooking effects on significant nutritional constituents, the fatty acid composition and the content of B vitamins were analysed. Volatile profiles of samples were also compared using solid-phase microextraction. SV cooking resulted in the less favourable meat texture, presenting the highest hardness and chewiness. Moreover, high hardness values measured on SV samples is also related to the high weight loss. The technique of oven cooking (O) demonstrated superior results in terms of mechanical properties, which are closely associated with the cooking values. Specifically, the cook value C0 was significantly higher in the case of oven cooking compared to SV, VC, and ST. Mild temperature conditions and cooking times of the four considered cooking techniques did not induce significant variations in the fatty acid composition and volatile profile. Conversely, SV and VC allowed the highest amount of vitamin B retention in cooked meat. This work suggests that some differences emerged on the effects due to sub-atmospheric and atmospheric cooking compared to traditional ones.

Legacy perfluoro-alkyl substances impair LDL-cholesterol uptake independently from PCSK9-function.

Perfluoro-alkyl substances (PFAS) are pollutants, whose exposure was associated with altered levels of low-density lipoproteins (LDL) in humans. Here we investigated this clinical outcome in two groups of young male adults residing in areas of respectively low and high environmental exposure to perfluoro-octanoic-acid (PFOA). From the Regional Authority data on pollution areas, 38 not-exposed and 59 exposed age-matched participants were evaluated for serum levels of total cholesterol (Total-Chol), LDL-Chol, high-density lipoprotein cholesterol (HDL-Chol), triglycerides (Tgl) and chromatography quantified PFOA. Human hepato-carcinoma cell line HepG2 was exposed to PFOA or perfluoro-octane-sulfonate (PFOS), as legacy PFAAs, and C6O4 as new generation compound. Fluorimetry was used to evaluate the cell-uptake of labelled-LDL. Proprotein Convertase Subtilisin/Kexin 9 (PCSK9)-mediated LDL-receptor (LDL-R) degradation and sub-cellular localization of LDL-R were evaluated by western blot analysis. Serum levels of PFOA, were positively and significantly correlated with Total-Chol (ρ = 0.312, P = 0.002), LDL-Chol (ρ = 0.333, P = 0.001) and Tgl (ρ = 0.375, P < 0.001). Participants with high serum LDL-Chol and Tgl levels, according to the cardiovascular risk, were more prevalent in exposed compared to not-exposed subjects (respectively: 23.7% vs 5.3%, P = 0.023 and 18,6% vs 0%, P = 0.006). Exposure of HepG2 cells to PFOA or C6O4 100 ng/mL was associated with a significantly lower LDL uptake than controls but no major impact of any PFAAs on PCSK9-mediated LDL-R degradation was observed. Compared to controls, exposure to PFAS showed an unbalanced LDL-R partition between membrane and cytoplasm. Endocytosis inducer sphingosine restored LDL-R partition only in samples exposed to C6O4. These data suggest a novel endocytosis-based mechanism of altered lipid trafficking associated with the exposure to legacy PFAS.

LC-MS/MS and GC-MS profiling, antioxidant, enzyme inhibition, and antiproliferative activities of Thymus leucostomus H ausskn. & V elen. extracts.

The chemical composition as well as antioxidant, antiproliferative, and enzyme inhibition activities of extracts from aerial parts of Thymus leucostomus H ausskn. & V elen. obtained with hexane, methanol, and water were evaluated. Results showed that the methanol extract had significantly (p < 0.05) the highest total phenolic content (TPC; 107.80 mg GAE/g) and total flavonoids content (TFC; 25.21 mg RE/g) followed by the aqueous extract (102.72 mg GAE/g and 20.88 mg RE/g, respectively). LC-MS/MS-guided profiling of the three extracts revealed that rosmarinic acid (34.8%), hesperetin (42.9%), and linoleic acid (18%) were the dominant compounds in the methanol, aqueous and hexane extracts, respectively. GC-MS analysis of the hexane extract showed that É£-sitosterol (29.9%) was the major constituent. The methanol extract displayed significantly (p < 0.05) the highest Cu++ , Fe+++ , and Mo(VI) ions scavenging and reducing properties while the aqueous extract exerted significantly (p < 0.05) the highest metal chelating power (42.51 mg EDTAE/g). Both the hexane and methanol extracts effectively inhibited the acetylcholinesterase enzyme (2.63 and 2.65 mg GALAE/g, respectively) while the former extract exerted significantly (p < 0.05) the highest butyrylcholinesterase (2.32 mg GALAE/g), tyrosinase (19.73 mg KAE/g), and amylase (1.16 mmol ACAE/g) inhibition capacity. The aqueous extract exhibited the best glucosidase inhibition property (0.49 mmol ACAE/g). The methanol and hexane extracts exerted a higher cytotoxic effect on HT-29 (IC50 : 8.12 µg/mL) and HeLa (IC50 = 8.08 µg/mL) cells, respectively. In conclusion, these results provide valuable insight into the potential use of T. leucostomus bioactive extracts in different pharmaceutical applications.

Development and Characterization of Microparticles with Actinidia arguta Leaves Extract by Spray-Drying: A New Mind-Set Regarding Healthy Compounds for Oral Mucositis.

Actinidia arguta leaves have gained notoriety over the past years due to their rich bioactive composition with human pro-healthy effects, particularly in relation to antioxidants. Nevertheless, antioxidants are well known for their chemical instability, making it necessary to develop suitable delivery systems, such as microparticles, to provide protection and ensure a controlled release. The aim of this work was to produce polymeric particles of A. arguta leaves extract by spray-drying that may improve the oral mucositis condition. Microparticles were characterized by size, shape, antioxidant/antiradical activities, swelling capacity, moisture content, and effect on oral cells (TR146 and HSC-3) viability, with the aim to assess their potential application in this oral condition. The results attested the microparticles' spherical morphology and production yields of 41.43% and 36.40%, respectively, for empty and A. arguta leaves extract microparticles. The A. arguta leaves extract microparticles obtained the highest phenolic content (19.29 mg GAE/g) and antioxidant/antiradical activities (FRAP = 81.72 µmol FSE/g; DPPH = 4.90 mg TE/g), being perceived as an increase in moisture content and swelling capacity. No differences were observed between empty and loaded microparticles through FTIR analysis. Furthermore, the exposure to HSC-3 and TR146 did not lead to a viability decrease, attesting their safety for oral administration. Overall, these results highlight the significant potential of A. arguta leaves extract microparticles for applications in the pharmaceutical and nutraceutical industries.

Linking traditional medicine to modern phytotherapy: Chemical characterization and assessment of antioxidant and anticholinesterase effects in vitro of a natural Persian remedy for dementia.

Several natural remedies are used in the Traditional Persian Medicine (TPM) to prevent dementia, but their efficacy is debated. In this work, an improved "Safoof-e-Nesyan" formulation described in the "Qarabadin-e-Azam" pharmacopoeia was developed, and its chemical composition and antioxidant and anti-cholinesterase properties were assessed. The formulation contains a mixture (FM) of Cinnamomum cassia (CC), C. verum (CV), Pistacia lentiscus (PL), Rheum palmatum (RP), Syzygium aromaticum (SA), and Zingiber officinalis (ZO) powdered plants. Its total phenolic content is 110.45 mg GAE/g, while the total flavonoid content is 6.28 mg RE/g. 66 secondary metabolites (mainly tannins, flavonoids, anthraquinones, and gingerols) were identified by UPLC-QToF-MS analysis. FM exerts antioxidant effects by scavenging radicals, and by reducing and chelating metals such as Mb, Cu and Fe. The anticholinesterase activity of one gram of the FM equals that of 3.60 mg of the reference drug galantamine, on both acetyl- and butyryl-cholinesterase. Correlations between specific compounds and bioactivities were highlighted by multivariate analysis of data: lyoniresinol 9'-glucoside strongly correlates with antiradical activities on DPPH and ABTS and reducing activity on Cu, and with anti-AChE effects. Most of the identified flavonoids and the ellagic acid derivatives positively correlate with the reducing activity on Fe and Mb, and with anti-BChE effects. Finally, a tablet formulation of the FM was developed, and its physical properties were preliminarily assessed. Overall, our results indicate that the FM may be a useful natural remedy for dementia, although further safety and efficacy assessments in vivo are required.

Simulated Gastrointestinal Digestion of Chestnut (Castanea sativa Mill.) Shell Extract Prepared by Subcritical Water Extraction: Bioaccessibility, Bioactivity, and Intestinal Permeability by In Vitro Assays.

Chestnut shells (CSs) are an appealing source of bioactive molecules, and constitute a popular research topic. This study explores the effects of in vitro gastrointestinal digestion and intestinal permeability on the bioaccessibility and bioactivity of polyphenols from CS extract prepared by subcritical water extraction (SWE). The results unveiled higher phenolic concentrations retained after gastric and intestinal digestion. The bioaccessibility and antioxidant/antiradical properties were enhanced in the following order: oral < gastric ≤ intestinal digests, attaining 40% of the maximum bioaccessibility. Ellagic acid was the main polyphenol in the digested and undigested extract, while pyrogallol-protocatechuic acid derivative was only quantified in the digests. The CS extract revealed potential mild hypoglycemic (<25%) and neuroprotective (<75%) properties before and after in vitro digestion, along with upmodulating the antioxidant enzymes' activities and downregulating the lipid peroxidation. The intestinal permeation of ellagic acid achieved 22.89% after 240 min. This study highlighted the efficacy of the CS extract on the delivery of polyphenols, sustaining its promising use as nutraceutical ingredient.