Liquid Chromatography-electrospray Ionization-tandem Mass Spectrometry Research Articles

Development of a conventional immunochemical detection system for determination of Nδ-(5-hydro-5-methyl-4-imidazolone-2-yl)-ornithine in methylglyoxal-modified proteins.

Methylglyoxal (MGO) produced during glycolysis is known to react with arginine residues on proteins to generate advanced glycation end products, such as Nδ-(5-hydro-5-methyl-4-imidazolone-2-yl)-ornithine (MG-H1). Since the production of MGO is increased during hyperglycemia or metabolic disorders in vivo, it is considered that the measurement of MG-H1 is useful for evaluating abnormalities in carbohydrate metabolism. Thus, we prepared a monoclonal antibody against MG-H1 to develop a conventional measurement system for MG-H1. Reactivity and specificity of the antibody to MGO-modified protein were confirmed by enzyme-linked immunosorbent assay and western blotting, respectively. The measurement of MG-H1 content by the antibody was positively correlated with that by electrospray ionization-liquid chromatography-tandem mass spectrometry and the ratio of modified arginine residues by amino acid analysis. Our results demonstrated that immunochemical methods could be useful for the estimation of MG-H1 content in modified proteins.

Metabolic profiling and gene expression analysis provides insights into flavonoid and anthocyanin metabolism in poplar.

Poplar, a woody perennial model, is a common and widespread tree genus. We cultivated two red leaf poplar varieties from bud mutation of Populus sp. Linn. '2025' (also known as Zhonglin 2025, L2025 for shot): Populus deltoides varieties with bright red leaves (LHY) and completely red leaves (QHY). After measuring total contents of flavonoid, anthocyanin, chlorophyll and carotenoid metabolites, a liquid chromatography-electrospray ionization-tandem mass spectrometry system was used for the relative quantification of widely targeted metabolites in leaves of three poplar varieties. A total of 210 flavonoid metabolites (89 flavones, 40 flavonols, 25 flavanones, 18 anthocyanins, 16 isoflavones, 7 dihydroflavonols, 7 chalcones, 5 proanthocyanidins and 3 other flavonoid metabolites) were identified. Compared with L2025, 48 and 8 flavonoids were more and less abundant, respectively, in LHY, whereas 51 and 9 flavonoids were more and less abundant in QHY, respectively. On the basis of a comprehensive analysis of the metabolic network, gene expression levels were analyzed by deep sequencing to screen for potential reference genes for the red leaves. Most phenylpropanoid biosynthesis pathway-involved genes were differentially expressed among the examined varieties. Gene expression analysis also revealed several potential anthocyanin biosynthesis regulators including three MYB genes. The study results provide new insights into poplar flavonoid metabolites and represent the theoretical basis for future studies on leaf coloration in this model tree species.

Untargeted-metabolomics differentiation between poultry samples slaughtered with and without detaching spinal cord

Chicken meat is among the common and relatively inexpensive source of protein consumed worldwide from the poultry industry. Many communities show concern regarding the procedure of slaughtering animals for meat consumption due to ethical, religious, or cultural reasons. Liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) based untargeted metabolomics of 40 chicken meat samples were evaluated to differentiate meat samples based on slaughtering methods. Samples were grouped into, Zabiha (cutting neck without detaching spinal cord) and Non-Zabiha (completely detaching neck). A volcano plot reveals at least 150 features found significantly different between the two groups having ≥ 2-fold changes in intensities with p-values ≤ 0.05. Among them 05 identified and 25 unidentified metabolites have clear differences in peak intensities. The identified features can be employed to differentiate meat obtained from different slaughtering methods. A characteristic pattern based on principal component analysis (PCA) and orthogonal partial least square-discriminant analysis (OPLS-DA) was observed among the groups. The results will benefit Halal certification, food safety, and security agencies to curb food fraud.

The application of in-source fragmentation in ultra-high performance liquid chromatography-electrospray ionization -tandem mass spectrometry for pesticide residue analysis

In this study, in-source fragmentation in ultra-high performance liquid chromatography-electrospray ionization-tandem mass spectrometry (UHPLC-ESI-MS/MS) was investigated and applied for pesticide residue analysis. Over 400 pesticides were tested, among which 26 pesticides were found to be sensitive to in-source fragmentation, producing 33 in-source fragments. The fragment pathways were studied and severe in-source fragmentation was observed due to the cleavage of C-O bond of carbamate, phosphate, ester, and ether groups, especially for isocarbophos and methoprene, leading to false negative results. High source temperatures could significantly increase the extent of in-source fragmentation. A multiple reaction monitoring (MRM) based multiresidue method was established for the pesticide residue analysis in vegetables and fruits, applying both pesticides and in-source fragments as precursors. The quantification ability of the method was validated and compared in terms of recovery, linearity, and the limit of quantification. In-source fragmentswere found to be more suitable than their parent pesticides as precursor ions for MRM analysis.

Valorization of fruit processing by-products: free, esterified, and insoluble bound phytochemical extraction from cherry (Prunus avium) tissues and their biological activities

Fruit processing by-products having limited utilization and no commercial value are generated in high amounts. However, they are rich in phytochemicals with high nutritional value. Therefore, different techniques and applications including extractions, purifications, and fermentations have been evolved for the recovery of these phytochemicals into high value-added products. Similarly, free (F), esterified (E) and insoluble-bound (B) phenolic fractions of sweet cherry processing by-products including stalk, pulp, seed, and leaf were extracted and the potential of these extracts as a source of natural biological compounds were investigated in this study. Total contents of phenolic (TPC), flavonoid (TFC), and hydrolysable tannin compounds (HTC) were analyzed. Individual phenolics were identified and quantified using liquid chromatography–electrospray ionization–tandem mass spectrometry (LC–ESI–MS/MS). The biological activity of these phytochemicals was determined in terms of antioxidant (DPPH, ABTS, FRAP, and CUPRAC), antidiabetic, and antimicrobial activity. TPC, TFC, and HTC were highly found in F phenolic fractions. Nine phenolic compounds were quantified, most notably in the F phenolic fractions. The enzyme inhibition activity against α-glucosidase (5.88 mg/mL) and α-amylase (0.46 mg/mL) of the E phenolic fractions of seed were significantly higher (p < 0.05) than those of F and B extracts. F fractions of stalk had remarkable antimicrobial activity against selected microorganisms except for Escherichia coli. Two uncorrelated principal components explaining 84.51% of the total variance helped to classify sweet cherry tissues. The results clearly showed that F, E, and B phenolics with their biological properties could be suitable food additives or supplements for enrichment agents in the different formulations.

Simultaneous quantification of the lipids phosphatidylcholine, 3-sn-phosphatidylethanolamine, sphingomyelin, and L-α-lysophosphatidylcholine extracted from the tissues of the invasive golden apple snail (Pomacea canaliculata) using UHPLC-ESI-MS/MS

Lipids such as phosphatidylcholine (PC), 3-sn-phosphatidylethanolamine (PE), sphingomyelin (SM) and L-α-lysophosphatidylcholine (LPC) are the major components of biological membranes and play important roles in physiological functions. Here, PC, PE, SM, and LPC were extracted from golden apple snails (GAS, Pomacea canaliculata) and GAS flesh (GASF) using an ethanol/hexane sequential scheme and quantified simultaneously using ultra-high performance liquid chromatography-electrospray ionization-tandem mass spectrometry (UHPLC-ESI-MS/MS) to evaluate whether the GAS could be the source of the four lipids. Our results suggest that ethanol extracts contained the most crude lipids, and the yield of dry (evaporated) lipids were 3.45 g per 100 g fresh GASF and 1.82 g per 100 g of fresh GAS. Quantification of the lipids using UHPLC-ESI-MS/MS suggested that GAS contained PE, PC, SM and LPC, with SM being the most abundant lipid (after purification: 1.71 and 1.42 mg g−1 dry weight from 100 g of GASF and GAS, respectively). The method we used is cost-effective, and the recovery rates of ethanol and hexane ranged from 80−91% and 87−91% respectively. Overall, GAS and GASF are potential raw materials for lipids such as SM and PC extraction using the ethanol/hexane method. Comparatively, lipids extraction from the GAS is more effective and timesaving. Our finding would provide a way to utilize GAS and potentially control its invasion.

Wolffia globosa-Mankai Plant-Based Protein Contains Bioactive Vitamin B12 and Is Well Absorbed in Humans.

Background: Rare plants that contain corrinoid compounds mostly comprise cobalamin analogues, which may compete with cobalamin (vitamin B12 (B12)) metabolism. We examined the presence of B12 in a cultivated strain of an aquatic plant: Wolffia globosa (Mankai), and predicted functional pathways using gut-bioreactor, and the effects of long-term Mankai consumption as a partial meat substitute, on serum B12 concentrations. Methods: We used microbiological assay, liquid-chromatography/electrospray-ionization-tandem-mass-spectrometry (LC-MS/MS), and anoxic bioreactors for the B12 experiments. We explored the effect of a green Mediterranean/low-meat diet, containing 100 g of frozen Mankai shake/day, on serum B12 levels during the 18-month DIRECT-PLUS (ID:NCT03020186) weight-loss trial, compared with control and Mediterranean diet groups. Results: The B12 content of Mankai was consistent at different seasons (p = 0.76). Several cobalamin congeners (Hydroxocobalamin(OH-B12); 5-deoxyadenosylcobalamin(Ado-B12); methylcobalamin(Me-B12); cyanocobalamin(CN-B12)) were identified in Mankai extracts, whereas no pseudo B12 was detected. A higher abundance of 16S-rRNA gene amplicon sequences associated with a genome containing a KEGG ortholog involved in microbial B12 metabolism were observed, compared with control bioreactors that lacked Mankai. Following the DIRECT-PLUS intervention (n = 294 participants; retention-rate = 89%; baseline B12 = 420.5 ± 187.8 pg/mL), serum B12 increased by 5.2% in control, 9.9% in Mediterranean, and 15.4% in Mankai-containing green Mediterranean/low-meat diets (p = 0.025 between extreme groups). Conclusions: Mankai plant contains bioactive B12 compounds and could serve as a B12 plant-based food source.

Metabolomics and gene expression analysis reveal the accumulation patterns of phenylpropanoids and flavonoids in different colored-grain wheats (Triticum aestivum L.)

Colored-grain wheats have received increasing attention owing to their high nutritional values. In this study, we compared the metabolomes of four pigmented wheat cultivars with conventional yellow wheat using an ultra-performance liquid chromatography-electrospray ionization-tandem mass spectrometry (UPLC-ESI-MS/MS)-based metabolomics approach. A total of 711 metabolites were identified, and considerable differences were observed in the flavonoid and phenylpropanoid metabolites among five samples by orthogonal signal correction and partial least squares-discriminant analysis (OPLS-DA) analysis. These differential metabolites were significantly enriched in the “anthocyanin biosynthesis”, “flavones and flavonols biosynthesis”, and “flavonoids biosynthesis” pathways. Furthermore, the expression of 9 structural genes and 2 regulatory genes involved in flavonoid biosynthesis pathway were investigated by quantitative real-time PCR (qRT-PCR). Results suggested that blue, red, purple, and black wheat cultivars showed higher transcription levels of structural and regulatory genes in the flavonoid pathway than that of conventional yellow wheat, possibly accounting for the abundant anthocyanin accumulation in the grains of these four cultivars. This study laid a foundation for understanding the accumulation of flavonoids and coloration mechanisms in colored-grain wheats, and provides a theoretical basis for their sufficient utilization.

Analysis of hair and plasma samples for methotrexate (MTX) and metabolite using high-performance liquid chromatography triple quadrupole mass spectrometry (LC-MS/MS) detection.

Methotrexate (MTX), a folate antagonist, is the anchor drug used to treat several diseases. Therapeutic effects are attributed to intracellular levels of various methotrexate conjugates that are present in the cell as polyglutamates (MTX-Glu). The present study was conducted to develop a new liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS)-based assay to separately quantitate the MTX-Glu in hair cells, red blood cells, and serum using internal standards. Sample preparation consisted of extraction with an organic solution followed by solid-phase extraction. The presented methodology was applied for the analysis of methotrexate and its polyglutamates in hair cells, red blood cells, and serum obtained from clinical patients. The developed LC-ESI-MS/MS method for the quantitative measurement of MTX-Glu was both sensitive and precise within the clinically relevant range. This method is possibly be superior with respect to sensitivity, selectivity, and speed than all previously described approaches and can be easily applied in routine clinical tests owing to the combination of a simple pretreatment process with robust LC-MS/MS.

Exercise-induced recovery of plasma lipids perturbed by ageing with nanoflow UHPLC-ESI-MS/MS.

Daily physical exercise is an essential part of life and is required for remaining healthy; it enhances therapeutic efficacy in the elderly and prevents age-related diseases associated with lipid profile alterations, such as cardiovascular disease, diabetes mellitus, and dementia. To more efficiently analyse the lipid profiles and unveil the effect of exercise in aged mice, we optimized our study by examining the effects of using ionization modifiers in the mobile phase and in-source fragmentation of lysophospholipids on the simultaneous analysis of fatty acids (FAs) including hydroxyl fatty acids, glycerophospholipids, sphingolipids, and glycerolipids using nanoflow ultrahigh performance liquid chromatography-electrospray ionization-tandem mass spectrometry. We applied the optimization to investigate the lipidomic plasma alterations in young (7weeks old) and aged (84weeks old) mice (C57BL/6) subjected to treadmill exercise. Of the 390 identified lipid species, 159 were quantified to investigate ageing-related lipid species responsive to physical exercise. In particular, circulating lysophosphatidylcholine and lysophosphatidylethanolamine levels showed a significant decrease, and lysophosphatidic acid showed a simultaneous increase with ageing. The saturated FA (16:0 and 18:0) increased with ageing while the unsaturated FA 22:6 decreased. Dihydroxy fatty acid (18:1_2OH) showed an exercise-induced recovery against ageing. It is notable that the levels of five triacylglycerol species significantly increased by as much as threefold with ageing, but their levels largely recovered to those observed in the young mice after exercise. These findings can help understand the influence of ageing on lipid perturbation and the role of physical exercise on lipidomic recovery in response to ageing-associated loss of physical status. Graphical abstract.

Analysis of Flavonoid Metabolites in Chaenomeles Petals Using UPLC-ESI-MS/MS.

Chaenomeles species are used for both ornamental decoration and medicinal purposes. In order to have a better understanding of the flavonoid profile of Chaenomeles, the petals of four Chaenomeles species, including Chaenomeles japonica (RB), Chaenomeles speciose (ZP), Chaenomeles sinensis (GP), and Chaenomeles cathayensis (MY), were selected as experimental material. The total flavonoid content of GP was found to be the highest, followed by MY, ZP, and RB. In total, 179 flavonoid metabolites (including 49 flavonols, 46 flavonoids, 19 flavone C-glycosides, 17 procyanidins, 15 anthocyanins, 10 flavanols, 10 dihydroflavonoids, 6 isoflavones, 5 dihydroflavonols, and 2 chalcones) were identified by Ultra-Performance Liquid Chromatography-Electrospray Ionization-Tandem Mass Spectrometry. Screening of differential flavonoid metabolites showed that GP had higher levels of metabolites when compared with the other three Chaenomeles species. Annotation and enrichment analysis of flavonoid metabolites revealed that cyanidin 3,5-diglucoside and pelargonidin-3,5-diglucoside anthocyanins are likely responsible for the color differences of the four Chaenomeles petals. Additionally, a large number of flavonoids, flavonols, and isoflavones were enriched in the petals of GP. This study provides new insights into the development and utilization of Chaenomeles petals and provides a basis for future investigations into their utilization.

Toxin components and toxicological importance of Galerina marginata from Turkey

Amatoxins, most of which are hepatotoxic, can cause fatal intoxication. While mushrooms in the amatoxin-containing Galerina genus are rare, they can poison humans and animals worldwide. Few studies have profiled the toxicity of Galerina marginata. In addition, many studies indicate that macrofungi can have different characteristics in different regions. In this study, the quantities of toxins present in G. marginata from different provinces in Turkey were analysed using reversed-phase high-performance liquid chromatography with ultraviolet detection (RP-HPLC-UV) and liquid chromatography–electrospray ionization–tandem mass spectrometry (LC–ESI–MS/MS). G. marginata samples were collected from three different regions of Turkey. The taxonomic categorization of mushrooms was based on their micro- and macroscopic characteristics. The presence of toxins α-amanitin (AA), β-amanitin (BA), γ-amanitin (GA), phalloidin (PHD) and phallacidin (PHC) quantities were measured using RP-HPLC-UV and then were confirmed using LC-ESI-MS/MS. BA levels were higher than AA levels in G. marginata mushrooms collected from all three regions. Moreover, the levels of GA were below the detection limit and no phallotoxins were detected. This is the first study to identify and test the toxicity of G. marginata collected from three different regions of Turkey using RP-HPLC-UV. This is also the first study to confirm the UV absorption of amatoxins in G. marginata using LC–ESI–MS/MS, which is a far more sensitive process. More studies evaluating the toxicity of G. marginata in other geographic regions of the world are needed.

Enhanced Sensitivity for the Analysis of Perfluoroethercarboxylic Acids Using LC-ESI-MS/MS: Effects of Probe Position, Mobile Phase Additive, and Capillary Voltage.

Perfluoroethercarboxylic acids (PFECAs) have recently emerged as replacements for toxic per- and polyfluorinated alkyl substances (PFAS) including perfluorooctanoic acid (PFOA) and perfluorooctanesulfonic acid (PFOS). Compared with other PFAS, many PFECAs including hexafluoropropylene oxide dimer acid (HFPO-DA, trade name GenX) exhibit poor sensitivity during analysis using liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) and are therefore often difficult to quantify. This study examined changes in ESI probe position, mobile phase additive, and capillary voltage with the goal of enhancing PFECA sensitivity. In addition, the relative contributions of existing mechanistic theories for PFAS ionization during ESI are discussed. Results indicated that the LC-ESI-MS/MS sensitivity for 9 PFECAs can be improved significantly by altering the ESI probe position. At the optimal probe position, lowering the capillary voltage from 2.0 to 0.5 kV universally enhanced the LC-ESI-MS/MS sensitivity for PFAS analysis. For most analytes, the use of ammonium bicarbonate rather than ammonium acetate as a mobile phase additive also enhanced the analytical response. These effects have not been previously reported and suggest that many laboratories may be conducting analyses of PFECAs under suboptimal conditions. Using the strategies outlined in this study, PFECAs can be more easily incorporated into comprehensive methods for PFAS analysis. Here, we describe analytical parameters that enhance the sensitivity for some PFECAs by up to 36-fold while maintaining high sensitivity for legacy PFAS. This work not only highlights solutions to mitigate inadequate PFECA sensitivity but also provides insight into the mechanisms underlying PFAS ionization efficiency during LC-ESI-MS/MS.

Phytochemical Composition and Antimicrobial Activity of Corydalis solida and Pseudofumaria lutea.

Corydalis and Pseudofumaria are two closely related genera from the Papaveraceae subfamily Fumarioideae with Corydalis solida (C. solida) and Pseudofumaria lutea (P. lutea) as two representative species. Phytochemical analysis revealed significant differences in the quality and quantity of isoquinoline alkaloids, phenolic compounds and non-phenolic carboxylic acids between aerial and underground parts of both species. Using the Liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) technique, 21 compounds were identified: five protoberberine derivatives, three protopine derivatives, four phenanthridine derivatives, as well as three carboxylic acids, two hydroxycinnamic acids, one chlorogenic acid, one phenolic aldehyde, and two flavonoids. Moroever, significant differences in the content of individual compounds were observed between the two studied species. The phytochemical profile of C. solida showed a higher variety of compounds that were present in lower amounts, whereas P. lutea extracts contained fewer compounds but in larger quantities. Protopine was one of the most abundant constituents in C. solida (440–1125 µg/g d.w.) and in P. lutea (1036–1934 µg/g d.w.). Moreover, considerable amounts of coptisine (1526 µg/g) and quercetin (3247 µg/g) were detected in the aerial parts of P. lutea. Extracts from aerial and underground parts of both species were also examined for the antimicrobial potential against S. aureus, P. aeruginosa and C. albicans. P. lutea herb extract was the most effective (MIC at 0.39 mg/L) against all three pathogens.

Isoform 1 of Fibrinogen Alpha Chain Precursor is a Potential Biomarker for Steroid-Induced Osteonecrosis of the Femoral Head.

Early diagnosis is crucial to increase the chances of conservation treatment for patients with steroid-induced osteonecrosis of the femoral head (SIONFH). This study aimed to identify serum peptides as potential biomarkers to diagnose SIONFH. The serum proteome of 32 SIONFH patients and 24 healthy controls are analyzed using magnetic bead-based weak cation exchange (MB-WCX) and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF-MS). Next, candidate biomarkers are identified using liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS). Candidate biomarkers are then validated using ELISA and western blotting. 39 peaks are identified and the expression fold changes of seven peaks in the two groups are greater than 1.5. Three peaks (m/z: 1077.84Da; m/z: 1061.78Da; m/z: 1099.56Da) tend to be upregulated, while four peaks (m/z: 3973.92Da; m/z: 7766.53Da; m/z: 3957.31Da; m/z: 4212.02Da) tend to be down-regulated in SIONFH patients. The peak for a 1077.84Da peptide is identified as Isoform 1 of the Fibrinogen alpha chain precursor (FGA). ELISAs and western blot analyses reveal that the expression of FGA is significantly higher in SIONFH patients than healthy controls. FGA is overexpressed in SIONFH patients, and thus, is a novel potential biomarker for SIONFH.

Identification of cisplatin-resistant factor by integration of transcriptomic and proteomic data using head and neck carcinoma cell lines.

ABSTRACTCisplatin is an important drug for the treatment of head and neck squamous cell carcinoma (HNSCC). Determining chemoresistant factors prior to treatment will lead to great benefits for clinicians and patients. Here, we evaluated chemoresistant factors by integrating proteomic and transcriptomic data using HNSCC cell lines to identify a more precise chemoresistant factor in HNSCC. We used four HNSCC cell lines: cisplatin-sensitive, acquired cisplatin resistance, naturally cisplatin-resistant, and acquired 5-FU resistance. Proteomic analysis was performed using iTRAQ, tandem mass spectrometry, and liquid chromatography-electrospray ionization-tandem mass spectrometry. Transcriptomic analysis was performed using microarrays. By integrating these independent data, common factors were addressed and functional analysis was performed using small interfering RNAs (siRNAs) to change the chemosensitivity. Using iTRAQ analysis, 7 proteins were identified as specific for cisplatin chemoresistance factors. Transcriptomic analysis revealed hundreds of potential candidate factors. By combining and integrating these data, S100A2 was identified as a potential cisplatin-specific chemoresistance factor. Functional analysis with siRNA revealed that the expression of S100A2 was reduced and cisplatin sensitivity recovered in the acquired and naturally cisplatin-resistant cell lines, but not in the cisplatin-sensitive cell lines. S100A2 was identified as a cisplatin-specific chemoresistance factor by integrating the transcriptomic and proteomic results obtained using HNSCC cell lines. This is a novel technique that allows for a precise identification, also known as a comprehensive analysis. Our findings indicate that these proteins could be used as biomarkers of HNSCC treatments, providing physicians with new treatment strategies for patients with HNSCC, showing chemoresistance.

Carotenoid composition and antioxidant potential of Eucheuma denticulatum, Sargassum polycystum and Caulerpa lentillifera

Three species of Malaysian edible seaweed (Eucheuma denticulatum, Sargassum polycystum and Caulerpa lentillifera) were analyzed for their carotenoid composition using a combination of high-performance thin layer chromatography (HPTLC) and ultra-high-performance liquid chromatography-electrospray ionization-tandem mass spectrometry (UHPLC-ESI-MS/MS), while the antioxidant capacities were determined by 2,2-diphenyl-1-picrylhydrazyl (DPPH) and oxygen radical absorbance capacity (ORAC) assays. The HPTLC analysis exhibited a distinct carotenoid pattern among the three seaweed groups. The UHPLC-ESI-MS/MS analysis showed fucoxanthin as the major carotenoid present in S. polycystum while lutein and zeaxanthin in E. denticulatum. For C. lentillifera, β-carotene and canthaxanthin were the major carotenoids. Some of the carotenoids, such as rubixanthin, dinoxanthin, diatoxanthin and antheraxanthin, were also tentatively detected in E. denticulatum and S. polycystum. For antioxidant activity, S. polycystum (20 %) and E. denticulatum (1128 μmol TE/g) showed the highest activity in the DPPH and ORAC assays, respectively. The findings suggest the three edible varieties of seaweeds may provide a good dietary source with a potential to reduce antioxidative stress.

Phytochemical Characterization and Effects on Cell Proliferation of Lentisk (Pistacia lentiscus) Berry Oil: a Revalued Source of Phenolics.

The ethno-pharmaceutical use of the edible fixed oil produced from lentisk (Pistacia lentiscus) berries covers a long tradition in several Mediterranean regions. Many of the health-promoting properties of lentisk berry oil (LBO) have been associated with the content of polar (poly)phenolic compounds. However, the polar fraction (methanol 80%, v/v) of LBO (LBO-pf) remains poorly and inadequately characterized. We assessed the phytochemical composition (fatty acids, phytosterols and polyphenols) of cold-pressed LBO produced in Cilento (Campania region, Italy) over four years of production (2015-2018). Main phenolic compounds present in LBO-pf were identified and semi-quantified combining ultra-high performance liquid chromatography-electrospray ionization-tandem mass spectrometry (UHPLC-ESI-MS/MS) and HPLC with diode array detection. Phenolic compounds, also responsible for oil stability and antioxidant properties, are relatively abundant in LBO, compared to other edible oils. LBO-pf induced clear dose-dependent effects on the growth of HT-29 cell line derived from human colorectal adenocarcinoma, as evidenced by the cell cycle arrest. Our data support the health-promoting properties of cold-pressed LBO, which is obtained with good yield from spontaneous plants growing in semiarid regions.

Quantitative determination of four nitrofurans and corresponding metabolites in the fish muscle by liquid chromatography-electrospray ionization-tandem mass spectrometry

A method validated based on the European Union (EU) regulations for determining the presence of furazolidone, furaltadone, nitrofurazone, nitrofurantoin and their corresponding metabolites AOZ, AMOZ, SC and AH in fish muscle was developed. Samples were acid-hydrolyzed, treated with 2-nitrobenzaldehyde and extracted with ethyl acetate. The extracts were dried in a N2 stream and redissolved in methanohwater (50:50, v/v). Analysis was performed by LC/ESI/MS/MS. This developed method carried limits of quantification lower than 10 μg/kg for the nitrofurans and 1.0 μg/kg for the metabolites. It was observed that the decision limit (CCα) ranged from 2.93 to 5.01 μg/kg for the nitrofurans and 0.19 to 0.43 μg/kg for the metabolites. The detection capability (CCβ) was between 3.62 and 6.20 μg/kg for the nitrofurans and between 0.23 and 0.54 μg/kg for the metabolites. The linear calibration curve parameters in the fortified fish muscle were between 1.0-100.0 μg/kg and 0.1-10.0 μg/kg for the nitrofurans and metabolites, respectively.

Defining in vivo dose-response curves for kidney DNA adduct formation of aristolochic acid I in rat, mouse and human by an in vitro and physiologically based kinetic modeling approach.

Aristolochic acid I (AAI) is a well‐known genotoxic kidney carcinogen. Metabolic conversion of AAI into the DNA‐reactive aristolactam‐nitrenium ion is involved in the mode of action of tumor formation. This study aims to predict in vivo AAI‐DNA adduct formation in the kidney of rat, mouse and human by translating the in vitro concentration‐response curves for AAI‐DNA adduct formation to the in vivo situation using physiologically based kinetic (PBK) modeling‐based reverse dosimetry. DNA adduct formation in kidney proximal tubular LLC‐PK1 cells exposed to AAI was quantified by liquid chromatography‐electrospray ionization‐tandem mass spectrometry. Subsequently, the in vitro concentration‐response curves were converted to predicted in vivo dose‐response curves in rat, mouse and human kidney using PBK models. Results obtained revealed a dose‐dependent increase in AAI‐DNA adduct formation in the rat, mouse and human kidney and the predicted DNA adduct levels were generally within an order of magnitude compared with values reported in the literature. It is concluded that the combined in vitro PBK modeling approach provides a novel way to define in vivo dose‐response curves for kidney DNA adduct formation in rat, mouse and human and contributes to the reduction, refinement and replacement of animal testing.