High-performance Liquid Chromatography Coupled With Mass Spectrometry Research Articles

Cobalt-catalysed bicarbonate-activated peroxide as a promising system for the advanced oxidation of epirubicin in wastewaters

Epirubicin (EPI) is a common cytostatic drug often found as a pollutant in hospital wastewater. While many studies have focused on the degradation kinetics of its epimer, doxorubicin (DOX), none have reported the kinetics of EPI degradation. Herein, we report a new cost-efficient, environmentally friendly system for the advanced degradation of EPI consisting of cobalt-catalysed bicarbonate-activated peroxide. We propose a new kinetic model for EPI degradation that shows the hydroperoxyl radical is the most important reactive oxygen species (ROS) involved in the degradation process. Seven degradation products are identified using high-performance liquid chromatography coupled with mass spectrometry (HPLC-MS). The effect of surfactants, commonly found in wastewaters, is also investigated and the presence of the cationic surfactant hexadecylpyridinium chloride (CPC) in the reaction medium leads to a 6.1-fold increase in the degradation rate. Modelling the reaction kinetics using the Piszkiewicz model unveils the rate constant for the micellar degradation of EPI to be 4.66×10‐3s‐1. The ecotoxicity study of the degradation products shows a dramatic decrease in both acute and chronic toxicity. Although the level of mineralisation of the products is less than 10%, this method is still suitable as a pre-treatment of wastewaters. The same system is successfully used for the degradation of DOX, mitoxantrone, and quinizarin. In the case of DOX, a rate constant of 3.74×10‐2s‐1 is obtained, the highest reported so far.

Assessing Matrix Solid-Phase Dispersion Extraction Strategies for Determining Bisphenols and Phthalates in Gilthead Sea Bream Samples

Microplastics (MPs) and nanoplastics (NPs) are widely spread in the environment, generating significant concern due to their potential impact on environmental health. Marine species usually ingest plastic fragments, mistaking them for food. Many toxic compounds, such as plastic additives that are not chemically bound to the plastic matrix, can be released from MPs and NPs and reach humans via the food chain. This paper highlights the development and validation of a straightforward solid–liquid extraction clean-up procedure in combination with a matrix solid-phase dispersion method using high-performance liquid chromatography coupled with mass spectrometry (HPLC-MS) detection, enabling facile, precise, and reliable identification and quantitation of a total of six bisphenols and phthalates in gilthead sea breams. Under the optimized conditions, the developed method showed good linearity (R2 > 0.993) for all target compounds. The recoveries obtained were between 70 and 92%. The relative standard deviations (RSDs) for reproducibility (inter-day) and repeatability (intra-day) were less than 9% and 10%, respectively. The limit of detection (LOD) and limit of quantification (LOQ) for the target compounds ranged from 0.11 to 0.68 µg/kg and from 0.37 to 2.28 µg/kg, respectively. A new, efficient extraction methodology for the determination of BPA, BPS, BPF, DBP, DEP, and DHEP in gilthead seabream has been optimized and validated.

TRACING THE COORDINATED CONTROL OF NAD+ HOMEOSTASIS USING C. ELEGANS

Abstract Nicotinamide adenine dinucleotide (NAD+) is a vital coenzyme involved in redox reactions and cellular signaling. Its levels decline with age in humans, mice, and worms, leading to metabolic disturbances associated with age-related diseases. Interestingly, our studies in aged mice indicate that the rate of NAD+ synthesis remains unchanged, ruling out a lack of precursors as the cause of NAD+ decline. Instead, we found that tissues with lower NAD+ concentrations in aged mice exhibit higher fractional turnover, suggesting increased consumption as the primary driver of NAD+ decline during aging. This led us to hypothesize that the loss of NAD+ metabolic robustness and resiliency with age is linked to the hyper-activation and competition of NAD+-consuming enzymes. To investigate the physiological impact of NAD+ consuming enzymes on metabolic robustness, we used genetic tools and mass spectrometry in C. elegans. Surprisingly, we identified novel reproductive and aging phenotypes in mutant animals lacking specific NAD+ consuming enzymes, including Poly ADP-Ribose Polymerase (parp-1 and parp-2), TANKyrase (tank-1), and Toll and Interleukin 1 Receptor (tir-1). These mutants displayed significant reductions in fecundity and extensions in longevity. Additionally, employing High-Performance Liquid Chromatography coupled with Mass Spectrometry (HP-LC-MS), we examined the NAD+ metabolome across different developmental stages in our mutant strains, revealing notable metabolic disruptions associated with the loss of NAD+ consumers. In summary, our findings establish a novel connection between the hyper-activation of NAD+ degradation and the onset of reproductive aging. This research provides valuable insights into the role of NAD+ metabolism and its impact on aging-related processes.

Modified Biejia Jianwan decoction restrains PD-L1-mediated immune evasion through the HIF-1α/STAT3/NF-κB signaling pathway

Ethnopharmacological relevanceModified Biejia Jianwan (M-BJJW), a Traditional Chinese Medicine (TCM) decoction, has exhibited great potential in treating hepatocellular carcinoma (HCC). However, its underlying functional mechanism still remains unknown. Aim of the studyThe study aimed to explore the anti-hepatocarcinogenic effects of M-BJJW, specifically its influence on PD-L1-mediated immune evasion in hypoxic conditions, and elucidate the related molecular mechanisms in HCC. Materials and methodsTo investigate the therapeutic efficacy and mechanisms underlying M-BJJW's effects on HCC, we employed a diethylnitrosamine (DEN)-induced rat model maintained for 120 days. Following model establishment, flow cytometry was utilized to assess the distribution of immune cell populations in peripheral blood, spleens, and tumor tissues after M-BJJW administration. Simultaneously, enzyme-linked immunosorbent assays (ELISA) were conducted to analyze cytokine profiles in serum samples. Immunohistochemistry was employed to determine the expression levels of crucial proteins within tumor tissues. Furthermore, HCC cells exposed to CoCl2 underwent Western blot analysis to validate the expression levels of HIF-1α, PD-L1, STAT3, and nuclear factor kappa B (NF-κB) p65. The modulatory effects of STAT3 and NF-κB p65 were investigated using specific inhibitors and activators in wild-type cell lines. High-performance liquid chromatography coupled with mass spectrometry (HPLC/MS) was utilized to identify the chemical constituents present in M-BJJW-medicated serum. The immunomodulatory properties and the anti-tumor activities of M-BJJW were evaluated by co-culturing with peripheral blood mononuclear cells (PBMC) and the CCK-8 assay. Additionally, we assessed M-BJJW's impact on hypoxia-induced alterations in HCC cell lines using immunofluorescence and Western blot assessments. ResultsM-BJJW exhibited substantial therapeutic advantages by effectively alleviating pathological deterioration within the HCC microenvironment. In the DEN-induced rat model, M-BJJW administration notably reduced tumor growth. Flow cytometry analyses revealed an increased proportion of Cytotoxic T lymphocytes (CTLs) accompanied by a simultaneous decrease in regulatory T cells (Tregs). ELISA data supported a marked decrease in pro-inflammatory cytokines, including interleukin-6 (IL-6), interleukin-10 (IL-10), and tumor necrosis factor α (TNF-α). Immunohistochemistry confirmed the suppressive effect of M-BJJW on the expression of HIF-1α and PD-L1. Notably, western blotting unveiled the role of HIF-1α in regulating PD-L1 expression via the STAT3 and NF-κB signaling pathways in HCC cell lines, which was validated using activators and inhibitors of STAT3 and NF-κB. The CCK-8 assay and co-culture techniques demonstrated the anti-tumor activity of M-BJJW. Immunofluorescence and western blotting further confirmed that M-BJJW-containing serum dose-dependently inhibited HIF-1α, PD-L1, p-STAT3, and p-p65 in hypoxic HCC cell lines. ConclusionsM-BJJW demonstrates significant therapeutic potential against HCC by influencing the hypoxic microenvironment, thereby regulating the immunosuppressive milieu. Specifically, M-BJJW modulates the HIF-1α/STAT3/NF-κB signaling pathway, leading to reduced PD-L1 expression and an elevated ratio of cytotoxic T lymphocytes (CTLs), while concurrently decreasing T regulatory cells (Tregs) and immunosuppressive factors. These synergistic effects aid in countering PD-L1-mediated immune evasion, presenting compelling pharmacological evidence supporting the clinical application of M-BJJW as a therapeutic approach for HCC.

Microwave-assisted extraction of β-Sitosterol: A by-product from Agave angustifolia Haw bagasse

β-sitosterol (BSS) and β-sitosterol glucoside (BSSG) were extracted from Agave angustifolia Haw bagasse using microwave-assisted extraction (MAE). The quantification and characterization of BSS and BSSG were performed through high-performance thin-layer chromatography (HPTLC), Fourier-transform infrared spectroscopy (FT-IR), high-performance liquid chromatography coupled with mass spectrometry (HPLC-MS), and gas chromatography-mass spectrometry (GC-MS). With an extraction time of 9 seconds using MAE, a higher amount of BSS (103.6 mg of β-sitosterol per gram of dry weight of extract) was obtained compared to BSSG (61.6 mg of β-sitosterol glucoside per gram of dry weight of extract). MAE emerges as a promising method for the efficient recovery of β-sitosterol (BSS) and β-sitosterol glucoside (BSSG) from A. angustifolia bagasse. This enriches scientific knowledge regarding phytochemical extraction and holds great potential for various industries, such as pharmaceuticals and nutraceuticals. Additionally, by valorizing the by-products of the agave industry, this research contributes to the sustainable utilization of resources, aligning with the principles of green and circular economy practices.

Acetylcholine-Binding Protein Affinity Profiling of Neurotoxins in Snake Venoms with Parallel Toxin Identification

Snakebite is considered a concerning issue and a neglected tropical disease. Three-finger toxins (3FTxs) in snake venoms primarily cause neurotoxic effects since they have high affinity for nicotinic acetylcholine receptors (nAChRs). Their small molecular size makes 3FTxs weakly immunogenic and therefore not appropriately targeted by current antivenoms. This study aims at presenting and applying an analytical method for investigating the therapeutic potential of the acetylcholine-binding protein (AChBP), an efficient nAChR mimic that can capture 3FTxs, for alternative treatment of elapid snakebites. In this analytical methodology, snake venom toxins were separated and characterised using high-performance liquid chromatography coupled with mass spectrometry (HPLC-MS) and high-throughput venomics. By subsequent nanofractionation analytics, binding profiling of toxins to the AChBP was achieved with a post-column plate reader-based fluorescence-enhancement ligand displacement bioassay. The integrated method was established and applied to profiling venoms of six elapid snakes (Naja mossambica, Ophiophagus hannah, Dendroaspis polylepis, Naja kaouthia, Naja haje and Bungarus multicinctus). The methodology demonstrated that the AChBP is able to effectively bind long-chain 3FTxs with relatively high affinity, but has low or no binding affinity towards short-chain 3FTxs, and as such provides an efficient analytical platform to investigate binding affinity of 3FTxs to the AChBP and mutants thereof and to rapidly identify bound toxins.

Preparation of nickel-doped iron-based bimetallic organic framework carbonized derivatives for heterogeneous catalytic hydrogen peroxide degradation of 4-NP

A nickel-doped iron-bimetallic organic framework carbon-derived material (Fe-Ni-MOF@C) was successfully constructed by microwave-pyrolysis and used as a new non-homogeneous Fenton-like catalyst for the catalytic degradation of p-nitrophenol (4-NP) by hydrogen peroxide (H2O2). Several characterization results such as SEM and XRD show that Fe-Ni-MOF@C is successfully synthesized and has porous, strong stability and magnetic properties. Fe-Ni-MOF@C materials exhibit excellent catalytic performance for the degradation over a wide temperature (15–45 °C) and pH (3.00–9.00). Complete removal of 4-NP (30 mg/L) was achieved in 30 min at 25 °C and pH = 3. The material properties can be restored by calcination. According to the radical burst experiments, electron paramagnetic resonance spectroscopy (EPR) combined with the characterization results, the radicals acting in the Fe-Ni-MOF@C/H2O2 system are mainly •OH and, to a lesser extent, •O2−. The synergistic effect between Fe and Ni species accelerates electron transfer and radical generation, which is the reason for the material's high catalytic performance. High-performance liquid chromatography coupled with mass spectrometry (HPLC-MS) was used to analyze the main degradation products and pathways. This study demonstrates that bimetallic organic framework carbon-derived materials with synergistic effects have great application prospects as non-homogeneous Fenton-like catalysts in the field of H2O2 oxidation of organic pollutants.

The Comparison of Serum Exosome Protein Profile in Diagnosis of NSCLC Patients.

A thorough study of the exosomal proteomic cargo may enable the identification of proteins that play an important role in cancer development. The aim of this study was to compare the protein profiles of the serum exosomes derived from non-small lung cancer (NSCLC) patients and healthy volunteers (control) using the high-performance liquid chromatography coupled to mass spectrometry (HPLC-MS) method to identify potentially new diagnostic and/or prognostic protein biomarkers. Proteins exclusively identified in NSCLC and control groups were analyzed using several bioinformatic tools and platforms (FunRich, Vesiclepedia, STRING, and TIMER2.0) to find key protein hubs involved in NSCLC progression and the acquisition of metastatic potential. This analysis revealed 150 NSCLC proteins, which are significantly involved in osmoregulation, cell-cell adhesion, cell motility, and differentiation. Among them, 3 proteins: Interleukin-34 (IL-34), HLA class II histocompatibility antigen, DM alpha chain (HLA-DMA), and HLA class II histocompatibility antigen, DO beta chain (HLA-DOB) were shown to be significantly involved in the cancer-associated fibroblasts (CAFs) and tumor-associated macrophages (TAMs) infiltration processes. Additionally, detected proteins were analyzed according to the presence of lymph node metastasis, showing that differences in frequency of detection of protein FAM166B, killer cell immunoglobulin-like receptor 2DL1, and olfactory receptor 52R1 correlate with the N feature according to the TNM Classification of Malignant Tumors. These results prove their involvement in NSCLC lymph node spread and metastasis. However, this study requires further investigation.

Knotwood and Branchwood Polyphenolic Extractives of Silver Fir, Spruce and Douglas Fir and Their Antioxidant, Antifungal and Antibacterial Properties.

The extractive contents of three softwood species largely used in the wood industry, namely Abies alba (Silver fir), Picea abies (spruce) and Pseudotsuga menziesii (Douglas fir), have been determined quantitatively for knots and at different points chosen along their branches, before analysis using high-performance liquid chromatography coupled with Mass Spectrometry (HPLC-MS). The results indicated that branchwood samples located in close proximity to the stem present high contents of extractives similar to those recorded for the knots. HPLC analysis showed quite similar chemical compositions, indicating that first cm of the branches could be considered as an additional source of knotwood. The antibacterial, antifungal and antioxidant activities of knot's extractives have been investigated with the dual objective of better understanding the role of high levels of secondary metabolites present in the knot and evaluating their potential for biorefinery applications. The antioxidant activity study showed that crude extracts of Douglas fir knotwood presented higher radical scavenging activity levels than the extracts of Silver fir and spruce, which presented more or less the same activities. Silver fir and spruce knotwood extracts presented higher antibacterial activity levels than the Douglas fir knotwood extracts did, while Douglas fir knotwood extracts presented more fungal growth inhibition than the spruce and fir knotwood extracts did. The structure-activity relationships indicate that radical scavenging and antifungal activities are associated with a higher relative quantity of flavonoids in the crude extracts, while higher relative quantities of lignans are associated with antibacterial activity.

HPLC-Mass spectrometry analysis of phenolics comparing traditional bilberry and blueberry liqueurs

The purpose of this study was to investigate the differences between the phenolics compositions of bilberry and blueberry liqueurs, to determine whether these are comparable. Both bilberry and blueberry liqueurs have the same name on the market, however they differ in phenolic composition and content. A total of 43 compounds were identified, 36 in the bilberry liqueur, and 21 in the blueberry liqueur. High performance liquid chromatography coupled with mass spectrometry (HPLC-MS) was used to identify and quantify these compounds, where they were fragmented to the MS4 fragments. Anthocyanins were the major phenolics group detected in the bilberry liqueurs, but only the second most abundant phenolics group in the blueberry liqueurs. The most abundant phenolics group in the blueberry liqueurs was hydroxycinnamic acids. The lowest content of individual phenolic compounds was for blueberry liqueur made from whole fruit. This study will provide valuable data to consumers in their choice of a more expensive bilberry liqueur or a cheaper blueberry liqueur.

LC-MS guided isolation of N,β-glucopyranosyl vincosamide and other compounds from the curare ingredient Strychnos peckii

Strychnos peckii (Loganiaceae) is an important active ingredient in curare poisons in the Amazon rainforest. Although previous studies have identified this species as a promising source of monoterpene indole alkaloids (MIAs), knowledge about other natural products is still scarce. Thus, to detect and guide the isolation of unprecedented bioactive compounds from the leaves of S. peckii, an untargeted high-performance liquid chromatography coupled with mass spectrometry (HPLC-MS) analysis was performed with the leaf aqueous extract. The HPLC-MS analysis allowed the detection of eleven compounds, including the alkaloids harman-3-carboxylic acid (5) and N,β-glucopyranosyl vincosamide (6), and the flavonoids quercetin 3-O-rhamnopyranoside (9) and kaempferol 3-O-rhamnopyranoside (10), all not previously reported in the Loganiaceae family. These compounds, along with strictosidine (3), 5-carboxystrictosidine (7), and desoxycordifoline (8) were isolated through modern chromatographic techniques and determined by using NMR spectroscopy in combination with MS. Overall, the untargeted HPLC-MS analysis proved to be a simple and effective approach to guide the isolation of substances not yet identified from S. peckii.

Eugenia sonderiana O. Berg leaves: Phytochemical characterization, evaluation of in vitro and in vivo antidiabetic effects, and structure-activity correlation

Several medicinal plants have drawn the attention of researchers by its phytochemical composition regarding their potential for treating chronic complications of diabetes mellitus. In this context, plants of the Myrtaceae family popularly used in Brazil for the treatment of diabetes mellitus, including Eugenia sonderiana, have shown beneficial effects due to the presence of phenolic compounds and saponins in their chemical constitution. Thus, the present work aimed to perform the phytochemical characterization of the hydroethanolic extract of E. sonderiana leaves using high-performance liquid chromatography coupled with mass spectrometry (HPLC-MS), along with in vitro and in vivo studies of antidiabetic activity. The chemical characterization revealed the presence of phenolic compounds, flavonoids, neolignans, tannins, and saponins. In addition, the extract exhibited minimum inhibitory concentrations of alpha-amylase and alpha-glycosidase higher than the acarbose in the in vitro tests. Also, the in vivo tests revealed a slight increase in body mass in diabetic rats, as well as a significant decrease in water and feed consumption provided by the extract. Regarding serum biochemical parameters, the extract showed significant activity in decreasing the levels of glucose, hepatic enzymes, and triglycerides, in addition to maintaining HDL cholesterol levels within normal ranges, protecting the cell membranes against oxidative damage. Thus, the extract of E. sonderiana leaves was considered promising pharmaceutical ingredient in the production of a phytotherapy medication.

Cannabis sativa L. roots from Northeast Brazil reduce abdominal contortions in a mouse model of primary dysmenorrhea

Ethnopharmacological relevanceAlthough the root of Cannabis sativa L. has been mentioned in some regions, such as the Vale do São Francisco, for its potential traditional medicinal use as an anti-inflammatory, anti-asthmatic, and against gastrointestinal diseases, it has received little exploration and discussion. Aim of the studyThis study aimed to perform a chemical analysis of an aqueous extract of Cannabis sativa roots (AqECsR) and evaluate its pharmacological effects against uterine disorders, in vivo and ex vivo, in rodents. Materials and methodsThe roots were provided by the Brazilian Federal Police, and the freeze-dried extract was used for the chemical analysis of the AqECsR by high performance liquid chromatography coupled with mass spectrometry (HPLC-MS). The sample was subsequently used in three doses for pharmacological assays (12.5, 25, and 50 mg/kg), which included the spasmolytic activity test and the primary dysmenorrhea test. The primary dysmenorrhea test aimed to verify the effect of AqECsR on induced abdominal contortions in female mice in vivo and to perform a morphometric analysis of the organs. Association tests at subtherapeutic doses of AqECsR with antidysmenorrheic drugs were also performed. ResultsThe data obtained by HPLC-MS suggested the presence of four substances: cannabisativine, anhydrocannabisativine, feruloyltyramine, and p-coumaroyltyramine. In the pharmacological assays, the AqECsR showed no spasmolytic effect. However, in the antidysmenorrheal activity test, AqECsR demonstrated a significant in vivo effect of reducing oxytocin-induced abdominal contortions. Morphometric analysis of the uterus showed no significant organ enlargement effect, and the association of AqECsR with subtherapeutic doses of three drugs used in antidysmenorrheal therapy (mefenamic acid, scopolamine, and nifedipine) showed an effect in reducing abdominal contortions. ConclusionsIn conclusion, AqECsR contains four chemical compounds and exhibits an antidysmenorrheic effect both alone and in association with drugs, reducing abdominal contortions in female mice without generating organ enlargement in the animals. Further studies are needed to prove the mechanism of action by which AqECsR promotes its effect on primary dysmenorrhea and to explore its associations.

Estrogenic and anti-amnesic potential of Millettia griffoniana Baill. (Fabaceae) ethanolic extract on scopolamine-induced memory impairment in ovariectomized Wistar rats.

Dementias including Alzheimer disease (AD) are three times higher in menopausal women than in men. Phytoestrogens, a group of plant-derived compounds are known to alleviate menopausal complaints including dementia. Millettia griffoniana Baill is a phytoestrogen-rich plant used to treat menopausal complaints and dementia. Evaluating the estrogenic and neuroprotective potential of Millettia griffoniana on ovariectomized (OVX) rats. The in vitro safety of M. griffoniana ethanolic extract was assayed by MTT in human mammary epithelial (HMEC) and mouse neuronal (HT-22) cells and its lethal dose 50 (LD50) was estimated following OECD 423 guidelines. For estrogenicity, in vitro the well known E-screen assay on MCF-7cells was performed and in vivo four groups of OVX rats were treated either with 75, 150 and 300mg/kg M. griffoniana extract doses or estradiol (1mg/kg BW) for three days; and changes in uterine and vagina were analyzed. Then, for neuroprotective effect, Alzheimer-type dementia induction was achieved by scopolamine (1.5mg/kg B.W., i.p.) injection four days/week and M. griffoniana extract as well as piracetam (standard) were administered daily for 2 weeks to evaluate the extract's neuroprotective potential. The endpoints were the assessment of learning and working memory, oxidative stress state (SOD, CAT, and MDA) in brain, acetylcholine esterase (AChE) activity and the histopathological changes in hippocampus. No toxic effect was observed when incubating mammary (HMEC) and neuronal (HT-22) cells with M. griffoniana ethanol extract for 24h and its LD50 was found >2000mg/kg. The extract also exhibited both in vitro and in vivo estrogenic activities, displayed by a significant (p<0.01) increment in MCF-7cells population in vitro and an increase in the epithelium height of the vagina and the wet weight of the uterus mainly with the 150mg/kg BW extract dose compared to untreated OVX rats. The extract also reversed scopolamine-induced memory impairment in rat by improving learning, working and reference memory. This was associated with an increment in CAT and SOD expression, alongside a decrement in MDA content and AChE activity in hippocampus. Further, the extract reduced neuronal cell loss in hippocampal structures (CA1, CA3 and dentate gyrus). High Performance Liquid Chromatography coupled with Mass Spectrometry (HPLC-MS) spectra, revealed the presence of numerous phytoestrogens in M. griffoniana extract. M. griffoniana ethanolic extract has estrogenic, anticholinesterase and antioxidant activities that could account for its anti-amnesic effects. These findings therefore sheds light on why this plant is commonly used in the therapy of menopausal complaints and dementia.

In vitro Antioxidant Potential and In vivo Analgesic and Anti-Inflammatory Activities of Moroccan Equisetum ramosissimum

Background: Equisetum ramosissimum is a pteridophyte plant used in Moroccan traditional medicine for its diuretic and antidiabetic properties. The species is known to reduce blood cholesterol levels and is given in cases of gonorrhea. Objective: The present work aims to explore the potential use of the crude butanolic (BE) and methanolic (ME) extracts of E. ramosissimum in the management of pain, inflammation and oxidative stress, and their chemical characterization. Methods: The phytochemical investigation of E. ramosissimum was initially done by colorimetric methods, followed by High Performance Liquid Chromatography coupled with Mass Spectroscopy (HPLC-MS). Its antioxidant activity was evaluated using radical scavenging activity in 2,2- diphenyl-1-picrylhydrazyl radical (DPPH), reducing power and β-carotene/linoleic acid bleaching assays. The analgesic activity was evaluated by acetic acid in mice and hot-plate-test in rat models. Rate paw edema and ear edema were used as anti-inflammatory models. Results: The chemical quantification revealed appreciable levels of phenolic compounds. The results of HPLC-UV and HPLC-MS highlighted the presence of kaempferol3-O-sophoroside7-Oglucoside, kaempferol3,7-O-diglucoside, vanillin, ferulic acid and tannic acid. The maximum IC50 was obtained for BE by DPPH test (0.064±0.0004mg/mL). BE exhibited an interesting antinociceptive effect; inhibition of writhes, 3.83±0.48 at 600mg/kg, increased the latency period on the hotplate- test, i.e., 14.98S±0.57S at 400mg/kg after 120min. Furthermore, the results reported a significant inhibition of rate-paw edema and ear edema for both extracts at a dose of 400mg/kg. Conclusion: BE and ME of E. ramosissimum were found to contain a significant amount of flavonoids, especially derivatives of kaempferol, which can explain the interesting effectiveness properties. Possible applications in the food and pharmaceutical industries are suggested.

Phytochemical study, antioxidant and vasodilatation activities of leafy stem extracts of Flemingia faginea Guill. & Perr. (Barker), a medicinal plant used for the traditional treatment of arterial hypertension

BackgroundFlemingia faginea Guill. & Perr. is a medicinal plant used for the endogenous treatment of hypertension. This study aimed to establish the phytochemical profile, antioxidant power, and vasodilation effects of aqueous decoction (FAD), ethyl acetate (EAF) and aqueous residual fractions (ARF) of the leafy stem of Flemingia faginea (F. faginea). MethodsThe phytochemical study was performed using high-performance thin-layer chromatography technique and high-performance liquid chromatography coupled with mass spectrometry (HPLC-MS). Folin-Ciocalteu and aluminum trichloride colorimetric methods were used, respectively, to quantify total polyphenols and flavonoids. The antioxidant properties of the extracts were determined using 2,29-azinobis-3-ethylbenzothiazoline-6-sulfonic (ABTS•), 2,2-diphenyl-1-picrylhydrazyl (DPPH•) and ferric ion reduction (FRAP) methods. The vasodilatory effects of F. faginea extracts were performed using the DMT 620M myograph technique for an ex vivo study. ResultsThe phytochemical study revealed the presence of flavonoids, tannins, coumarins, sterols, and triterpenes, alkaloids, and saponins in the extracts studied. HPLC MS analysis identified as the main phenolic compounds, gallic acid-glucoside (16.56 ± 0.06 mg/g), myricetin rhamnoside (15.39 ± 0.04 mg/g), myricetin rutinoside (9.32 ± 0.00 mg/g), quercetin rutinoside (9.05 ± 0.00 mg/g), caffeoyl-glucoside (6.07 ± 0.02 mg/g) and 5-Caffeoylquinic acid (4.52 ± 0.00 mg/g). The present data also showed higher contents of phenolic compounds and flavonoids in fractions of ARF (280.49 ± 0.08 mgGAE/g) and EAF (33.31 ± 0.33 mgQE/g) fractions respectively. Furthermore, the ARF fraction had the best iron ion reduction capacity (16.96 ± 0.01µgAAE/100 mg), while, the EAF (ABTS: IC50=67.33 ± 0.04 µg/mL; DPPH: IC50=4.69 ± 0.20 µg/mL) was the best free radical scavenger. In endothelium-intact rings, study extracts showed a vasodilation effect in a concentration-dependent manner with half-maximum effective concentration (EC50) of 0.24 ± 0.03 mg/mL, 0.85 ± 0.07 mg/mL and 0.05±0.01 mg/mL respectively for FAD, ARF and EAF. In endothelium-denuded vessels, these values were 0.26 ± 0.09 mg/mL, 0.57 ± 0.05 mg/mL, and 0.85 ± 0.01 mg/mL for EAF, FAD, and ARF, respectively. The maximum effects were of 99.94 ± 0.15%, 86.89 ± 5.58%, and 62.69 ± 5.14% (endothelium-intact) and of 100 ± 0.06%, 96.55 ± 4.68%, and 74.16 ± 4.06% (endothelium-denuded), respectively, for EAF, FAD, and ARF with a significant difference between them. ConclusionThis study reports that the decoction and fractions of F. faginea are rich in various chemical compounds and possess antioxidant and vasodilatory properties. These preliminary data provide a scientific basis which contributed to justify that plant used in traditional medicine.

Estudio preliminar fitoquímico de fracciones hexánicas de hojas de Eugenia uniflora L. (Myrtaceae)

Fractions from the Hexane Extract (HE) of Eugenia unifloraL. leaves were subjected to various chromatographic systems. Germacrone sesquiterpene and bornyl acetate bicyclic ester were characterized by High Performance Liquid Chromatography coupled to Mass Spectrometry (HPLC-MS) with APCI Mass detector comparing with their homonymous spectrum provided by databases and characteristic fragmentation pathways were proposed. The monoterpene pulegone and the pentacyclic triterpene compound, ursolic acid, were found through High Performance Liquid Chromatography coupled to High Resolution Mass Spectrometry (HPLC -HRMS) by atmospheric pressure ionization (API) and the detector used was mass of Electronic Impact (IE). Both ursolic acid and bornyl acetate are present in other species of the same genus, but not in the species studied.

Using the Synergy between HPLC-MS and MALDI-MS Imaging to Explore the Lipidomics of Clear Cell Renal Cell Carcinoma.

Lipid imaging mass spectrometry (LIMS) has been tested in several pathological contexts, demonstrating its ability to segregate and isolate lipid signatures in complex tissues, thanks to the technique's spatial resolution. However, it cannot yet compete with the superior identification power of high-performance liquid chromatography coupled to mass spectrometry (HPLC-MS), and therefore, very often, the latter is used to refine the assignment of the species detected by LIMS. Also, it is not clear if the differences in sensitivity and spatial resolution between the two techniques lead to a similar panel of biomarkers for a given disease. Here, we explore the capabilities of LIMS and HPLC-MS to produce a panel of lipid biomarkers to screen nephrectomy samples from 40 clear cell renal cell carcinoma patients. The same set of samples was explored by both techniques, and despite the important differences between them in terms of the number of detected and identified species (148 by LIMS and 344 by HPLC-MS in negative-ion mode) and the presence/absence of image capabilities, similar conclusions were reached: using the lipid fingerprint, it is possible to set up classifiers that correctly identify the samples as either healthy or tumor samples. The spatial resolution of LIMS enables extraction of additional information, such as the existence of necrotic areas or the existence of different tumor cell populations, but such information does not seem determinant for the correct classification of the samples, or it may be somehow compensated by the higher analytical power of HPLC-MS. Similar conclusions were reached with two very different techniques, validating their use for the discovery of lipid biomarkers.

FREEZE-DRIED RASPBERRY POMACE: A RICH RESERVOIR OF POLYPHENOLS WITH ANTIOXIDANT AND PROTECTIVE EFFECTS AS A PROMISING FUNCTIONAL FOOD ADDITIVE

Background. Raspberries (Rubus idaeus L.) hold significant importance in the food industry. Rasp berry pomace, commonly regarded as a waste product during fruit processing, is a rich source of biologi cally active components. This study aimed to assess the detailed profile of polyphenols in fresh and freeze dried raspberry pomace. Furthermore, we examined its potential to protect normal human gastric epithelial cells against oxidative stress in an H2O2-induced stress model. A qualitative and quantitative analysis of phenolic compounds was performed using the high-performance liquid chromatography coupled with mass spectrometry (HPLC-MS) method. Cell viability was assessed based on the integrity of lysosomal membranes (neutral red NR assay) and mitochondrial dehydrogenase activity (MTT). Additionally, the reducing potential (FRAP), the ability to scavenge free radicals in the DPPH• test and studies on ABC HP021X cells based on the H2DCFDA test were evaluated. Results and conclusion. It was found that the extract from freeze-dried raspberry pomace is a rich source of various polyphenolic compounds, including ellagitannins, epicatechin and anthocyanins. The extract was characterized by high antioxidant potential and exhibited a protective effect on human gastric epithelial cells against H2O2-induced cytotoxicity. Cell incubation with the extracts prior to oxidative stress induction attenuated H2O2-induced cytotoxicity in a concentration-dependent manner, and at a con centration of 250 µg/cm3 , it abolished the detrimental effects of H2O2. Additionally, the extract at this concentration restored the redox status in cells with induced oxidative stress and reduced ROS levels to values similar to the control. Thus, raspberry pomace extracts may be considered a valuable food supplement.

Morus macroura Miq. Fruit extract protects against acetic acid-induced ulcerative colitis in rats: Novel mechanistic insights on its impact on miRNA-223 and on the TNFα/NFκB/NLRP3 inflammatory axis

Nod-like receptor pyrin domain-1 containing 3 (NLRP3) inflammasome/tumor necrosis factor alpha (TNFα)/nuclear factor kappa B (NFκB) inflammatory pathway is known to be involved in the pathogenesis of ulcerative colitis (UC). Inversely, miRNA-223 can exert counter-regulatory effect on NLRP3 expression. The mulberry tree (Morus macroura) fruit is attaining increased importance for its antioxidant and anti-inflammatory activity in addition to its high safety profile. Accordingly, we attempted to explore the possible protective effect of mulberry fruit extract (MFE) in acetic acid (AA)-induced UC rat model. Phytochemical constituents of MFE were characterized using high performance liquid chromatography coupled to mass spectrometry (HPLC-MS). In the in vivo study, three doses of MFE were orally given for seven days before intra-rectal induction of UC by AA on day eight. Screening study revealed that MFE (300 mg/kg) significantly reduced macroscopic and microscopic UC scores. Biochemically, MFE ameliorated oxidative stress, levels of TNFR1, NLRP3, p-NFκB p65, TNFα, IL-1β, and IL-18, caspase-1 activity, but enhanced miRNA-223 expression. In conclusion, our study provided a novel protective impact for MFE against UC, in which miRNA-223 and TNFα/NFκB/NLRP3 pathway are involved. These results provide a promising step that might encourage further investigations of MFE as a protective agent in UC patients.