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Together with proteins and fats, carbohydrates are one of the macronutrients in the human diet. Digestible carbohydrates, such as starch, starch-based products, sucrose, lactose, glucose and some sugar alcohols and unusual (and fairly rare) α-linked glucans, directly provide us with energy while other carbohydrates including high molecular weight polysaccharides, mainly from plant cell walls, provide us with dietary fibre. Carbohydrates which are efficiently digested in the small intestine are not available in appreciable quantities to act as substrates for gut bacteria. Some oligo- and polysaccharides, many of which are also dietary fibres, are resistant to digestion in the small intestines and enter the colon where they provide substrates for the complex bacterial ecosystem that resides there. This review will focus on these non-digestible carbohydrates (NDC) and examine their impact on the gut microbiota and their physiological impact. Of particular focus will be the potential of non-digestible carbohydrates to act as prebiotics, but the review will also evaluate direct effects of NDC on human cells and systems.

Although phthalate esters (PAEs) are plasticizers widely utilized nowadays to enhance the flexibility and pro-cessability of polymeric materials, their presence and detection in the food environment have become an important concern. In this study, phthalate esters (butyl benzyl phthalate (BBP), dibutyl phthalate (DBP), di-(2-ethylhexyl) phthalate (DEHP), and diisononyl phthalate (DINP)) were detected in 40 coffee samples in single-use or large plastic-based packages of different brands collected from the Turkish market. PAEs exposure and non -carcinogenic and carcinogenic health risks were calculated for females and males aged 15-64, depending on their coffee consumption within 24 h. DBP and DINP exhibited the highest level of migration to coffee samples (10-4) reported by the US EPA. Our research findings showed that almost all age groups over 15 were exposed to PAEs orally from the coffee they frequently consumed, which might lead to an excessive risk of cancer in individuals. Therefore, it was concluded that more comprehensive studies should be conducted based on this study, especially regarding cancer risk., Scientific Research Project Centre of Agri I?brahim Cecen University; [SYO.21.002], Funding This research was financed by the Scientific Research Project Centre of Agri I?brahim Cecen University (Project No: SYO.21.002) .

Post-translational modifications (PTMs) to the tails of the core histone proteins are critically involved in epigenetic regulation. Hypoxia affects histone modifications by altering the activities of histone-modifying enzymes and the levels of hypoxia-inducible factor (HIF) isoforms. Synthetic hypoxia mimetics promote a similar response, but how accurately the hypoxia mimetics replicate the effects of limited oxygen availability on the levels of histone PTMs is uncertain. Here we report studies on the profiling of the global changes to PTMs on intact histones in response to hypoxia/hypoxia-related stresses using liquid chromatography-mass spectrometry (LC-MS). We demonstrate that intact protein LC-MS profiling is a relatively simple and robust method for investigating potential effects of drugs on histone modifications. The results provide insights into the profiles of PTMs associated with hypoxia and inform on the extent to which hypoxia and hypoxia mimetics cause similar changes to histones. These findings imply chemically-induced hypoxia does not completely replicate the substantial effects of physiological hypoxia on histone PTMs, highlighting that caution should be used in interpreting data from their use.

Background Weaning is known to result in barrier dysfunction and villus atrophy in the immediate post-weaning phase, and the magnitude of these responses is hypothesized to correlate with changes in the glutathione (GSH) redox system. Therefore, these parameters were simultaneously measured throughout the weaning phase, in piglets differing in birth weight category and weaning age, as these pre-weaning factors are important determinants for the weaning transition. Low birth weight (LBW) and normal birth weight (NBW) littermates were assigned to one of three weaning treatments; i.e. weaning at 3 weeks of age (3w), weaning at 4 weeks of age (4w) and removal from the sow at 3 d of age and fed a milk replacer until weaning at 3 weeks of age (3d3w). For each of these treatments, six LBW and six NBW piglets were euthanized at 0, 2, 5, 12 or 28 d post-weaning piglets, adding up 180 piglets. Results Weaning increased the glutathione peroxidase activity on d 5 post-weaning in plasma, and duodenal and jejunal mucosa. Small intestinal glutathione-S-transferase activity gradually increased until d 12 post-weaning, and this was combined with a progressive rise of mucosal GSH up till d 12 post-weaning. Oxidation of the GSH redox status (GSH/GSSG Eh) was only observed in the small intestinal mucosa of 3d3w weaned piglets at d 5 post-weaning. These piglets also demonstrated increased fluorescein isothiocyanate dextran (FD4) and horseradish peroxidase fluxes in the duodenum and distal jejunum during the experiment, and specifically demonstrated increased FD4 fluxes at d 2 to d 5 post-weaning. On the other hand, profound villus atrophy was observed during the weaning transition for all weaning treatments. Finally, LBW and NBW piglets did not demonstrate notable differences in GSH redox status, small intestinal barrier function and histo-morphology throughout the experiment. Conclusion Although moderate changes in the GSH redox system were observed upon weaning, the GSH redox status remained at a steady state level in 3w and 4w weaned piglets and was therefore not associated with weaning induced villus atrophy. Conversely, 3d3w weaned piglets demonstrated GSH redox imbalance in the small intestinal mucosa, and this co-occurred with a temporal malfunction of their intestinal barrier function.

Oxidative stress is associated with a myriad of diseases including pregnancy pathologies with long-term cardiovascular repercussions for both the mother and baby. Aberrant redox signalling coupled with deficient antioxidant defence leads to chronic molecular impairment. Abnormal placentation has been considered the primary source for reactive species; however, placental dysfunction has been deemed secondary to maternal cardiovascular maladaptation in pregnancy. While various therapeutic interventions, aimed at combating deregulated oxidative stress during pregnancy have shown promise in experimental models, they often result as inconclusive or detrimental in clinical trials, warranting the need for further research to identify candidates. The strengths and limitations of current experimental methods in redox research are discussed. Assessment of redox status and oxidative stress in experimental models and in clinical practice remains challenging; the state-of-the-art of computational models in this field is presented in this review, comparing static and dynamic models which provide functional information such as protein-protein interactions, as well as the impact of changes in molecular species on the redox-status of the system, respectively. Enhanced knowledge of redox biology in during pregnancy through computational modelling such as generation of Systems Biology Markup Language model which integrates existing models to a larger network in the context of placenta physiology.

Drug-metabolizing enzymes (DMEs), a diverse group of enzymes responsible for the metabolic elimination of drugs and other xenobiotics, have been recognized as the critical determinants to drug safety and efficacy. Deciphering and understanding the key roles of individual DMEs in drug metabolism and toxicity, as well as characterizing the interactions of central DMEs with xenobiotics require reliable, practical and highly specific tools for sensing the activities of these enzymes in biological systems. In the last few decades, the scientists have developed a variety of optical substrates for sensing human DMEs, parts of them have been successfully used for studying target enzyme(s) in tissue preparations and living systems. Herein, molecular design principals and recent advances in the development and applications of optical substrates for human DMEs have been reviewed systematically. Furthermore, the challenges and future perspectives in this field are also highlighted. The presented information offers a group of practical approaches and imaging tools for sensing DMEs activities in complex biological systems, which strongly facilitates high-throughput screening the modulators of target DMEs and studies on drug/herb-drug interactions, as well as promotes the fundamental researches for exploring the relevance of DMEs to human diseases and drug treatment outcomes. (C) 2022 Chinese Pharmaceutical Association and Institute of Materia Medica, Chinese Academy of Medical Sciences. Production and hosting by Elsevier B.V.

A fast, accurate and reliable ultra-high performance liquid chromatography–tandem mass spectrometry (UHPLC-MS/MS) method was developed for simultaneous quantification of ivermectin (IVER), doramectin (DORA), and moxidectin (MOXI) in bovine plasma. A priority for sample preparation was the eradication of possible infectious diseases to avoid travel restrictions. The sample preparation was based on protein precipitation using 1% formic acid in acetonitrile, followed by Ostro® 96-well plate pass-through sample clean-up. The simple and straightforward procedure, along with the short analysis time, makes the current method unique and suitable for a large set of sample analyses per day for PK studies. Chromatographic separation was performed using an Acquity UPLC HSS-T3 column, with 0.01% acetic acid in water and methanol, on an Acquity H-Class ultra-high performance liquid chromatograph (UHPLC) system. The MS/MS instrument was a Xevo TQ-S® mass spectrometer, operating in the positive electrospray ionization mode and two multiple reaction monitoring (MRM) transitions were monitored per component. The MRM transitions of m/z 897.50 > 753.4 for IVER, m/z 921.70 > 777.40 for DORA and m/z 640.40 > 123.10 for MOXI were used for quantification. The method validation was performed using matrix-matched calibration curves in a concentration range of 1 to 500 ng/mL. Calibration curves fitted a quadratic regression model with 1/x2 weighting (r ≥ 0.998 and GoF ≤ 4.85%). Limits of quantification (LOQ) values of 1 ng/mL were obtained for all the analytes, while the limits of detection (LOD) were 0.02 ng/mL for IVER, 0.03 ng/mL for DORA, and 0.58 ng/mL for MOXI. The results of within-day (RSD < 6.50%) and between-day (RSD < 8.10%) precision and accuracies fell within acceptance ranges. No carry-over and no peak were detected in the UHPLC-MS/MS chromatogram of blank samples showing good specificity of the method. The applicability of the developed method was proved by an analysis of the field PK samples.

Resumen del póster presentado al International Scientific Association for Probiotics and Prebiotics Annual Meeting (ISAPP), celebrado en Sitges. Barcelona (España) del 15 al 17 de junio de 2022., Carbohydrates entering the complex microbial ecosystem resident in the human gut can be fermented to different degrees by the bacterial species present. Whilst insoluble carbohydrates may be fermented to a low degree, they may still be classed dietary fibres. More extensively fermented carbohydrates frequently display a selectivity towards particular microbial groups; if the end result of this process is an improvement in host health, then we would consider those carbohydrates to be prebiotics. It is a complex task to identify the structural basis of the fermentation selectivity underlying a prebiotic effect. Individuals vary in terms of the diversity of their gut microbiome and in the response of this microbiome to any given carbohydrate. The task is further complicated by the use of varying microbiology techniques across studies. Some generic understanding can be identified: It is clear that low molecular weight carbohydrates of a wide range of structure are very readily metabolised by bifidobacteria and are frequently seen to select for this genus in in vitro studies.. This selectivity is explained by the widespread presence of exo-glycosidase and sugar transport system genes in bifidobacterial genomes. Metabolism of higher molecular weight carbohydrates is dependant of organisms possessing the relevant polysaccharide utilisation genes like the bacteroides, rendering them highly adept at polysaccharide metabolism. To understand the fate of any given carbohydrate in the gut microbiome, however, the presence of cross-feeding networks must be taken into account. We know of some such cross feeding networks likely to operate in the gut but there is much to learn. ILSI Europe has commissioned an expert group to produce an authoritative view of the state of the art in terms of our understanding of structure function relationships in fermented carbohydrates. This presentation will highlight our current understanding of structure function relationships and point to the gaps in knowledge and the questions that arise in this area.

Background: vegetative diversity is based on different climate and geographical origins. In terms of beekeeping, herbal diversity is strongly correlated to the production of a wide variety of honey. Therefore, based on the existing plant diversity in each country, multiple honey varieties are produced with different health characteristics. While beekeeping potential and consumption preferences are reflected in products' variety, this leads to an increase in the region's economy and extensive export. In the last years, monofloral honey has gained interest from consumers and especially in the medicinal field due to the presence of phytochemicals which are directly linked to health benefits, wound healing, antioxidant, anticancer and anti-inflammatory activities. Scope and approach: this review aims to highlight the physicochemical properties, mineral profiles and antioxidant activities of selected monofloral honeys based on their botanical and geographical origin. Moreover, this review focuses on the intercorrelation between monofloral honey's antioxidant compounds and in vitro and in vivo activities, focusing on the apoptosis and cell proliferation inhibition in various cell lines, with a final usage of honey as a potential therapeutic product in the fight towards reducing tumor growth. Key findings and conclusions: multiple studies have demonstrated that monofloral honeys have different physicochemical structures and bioactive compounds. Useful chemical markers to distinguish between monofloral honeys were evidenced, such as: 2-methoxybenzoic acid and trimethoxybenzoic acid are distinctive to Manuka honey while 4-methoxyphenylacetic acid is characteristic to Kanuka honey. Furthermore, resveratrol, epigallocatechin and pinostrobin are markers distinct to Sage honey, whereas carvacrol and thymol are found in Ziziphus honey. Due to their polyphenolic profile, monofloral honeys have significant antioxidant activity, as well as antidiabetic, antimicrobial and anticancer activities. It was demonstrated that Pine honey decreased the MDA and TBARS levels in liver, kidney, heart and brain tissues, whereas Malicia honey reduced the low-density lipoprotein level. Consumption of Clover, Acacia and Gelam honeys reduced the weight and adiposity, as well as trygliceride levels. Furthermore, the antiproliferative effect of chrysin, a natural flavone in Acacia honey, was demonstrated in human (A375) and murine (B16-F1) melanoma cell lines, whereas caffeic acid, a phenolic compound found in Kelulut honey, proves to be significant candidate in the chemoprevention of colon cancer. Based on these features, the use of hiney in the medicinal field (apitherapy), and the widespread usage of natural product consumption, is gaining interest by each year., [PN-III-P1-1.2-PCCDI-2017-0056-2PCCDI], The publication was funded by PN-III-P1-1.2-PCCDI-2017-0056-2PCCDI.

Graphical abstract, The recent approval of messenger RNA (mRNA)-based vaccines to combat the SARS-CoV-2 pandemic highlights the potential of both conventional mRNA and self-amplifying mRNA (saRNA) as a flexible immunotherapy platform to treat infectious diseases. Besides the antigen it encodes, mRNA itself has an immune-stimulating activity that can contribute to vaccine efficacy. This self-adjuvant effect, however, will interfere with mRNA translation and may influence the desired therapeutic outcome. To further exploit its potential as a versatile therapeutic platform, it will be crucial to control mRNA’s innate immune-stimulating properties. In this regard, we describe the mechanisms behind the innate immune recognition of mRNA and provide an extensive overview of strategies to control its innate immune-stimulating activity. These strategies range from modifications to the mRNA backbone itself, optimization of production and purification processes to the combination with innate immune inhibitors. Furthermore, we discuss the delicate balance of the self-adjuvant effect in mRNA vaccination strategies, which can be both beneficial and detrimental to the therapeutic outcome.

Following a request from the European Commission, the EFSA Panel on Nutrition, Novel Foods and Food Allergens (NDA) was asked to deliver an opinion on the relationship between alpha-lipoic acid (ALA) and the risk of insulin autoimmune syndrome (IAS). The Panel was also asked to advise on the dose below which ALA added to foods is not expected to cause IAS. A review of all possible adverse effects associated with consumption of ALA was not requested. This mandate refers to the procedure under Article 8(2) of Regulation (EC) No 1925/2006 on addition of vitamins, minerals and certain other substances to foods. No pre-established rule exists for the evaluation of the safety of foods when classical toxicity tests cannot be used, e.g. for autoimmune diseases. Published scientific evidence was retrieved through comprehensive literature searches, particularly 49 case reports in which IAS developed following ALA consumption. In all cases, IAS resolved after a few weeks to months when ALA was discontinued. No publication linking the intake of ALA naturally occurring in foods to IAS was identified. The Panel concludes that the consumption of ALA added to foods, including food supplements, is likely to increase the risk of developing IAS in individuals with certain genetic polymorphisms, who cannot be readily identified without genetic testing. The plausible mechanism of such an effect has not yet been fully elucidated. The incidence of IAS in Europe is low and likely lower than in Japan where it has been estimated to be 0.017 per 100,000 inhabitants in 2017-2018. Considering the limited data available, the risk associated with the development of IAS following ALA consumption cannot be quantified precisely. An ALA dose below which IAS is not expected to occur is likely to vary between individuals and cannot be determined from the available data. (c) 2021 European Food Safety Authority. EFSA Journal published by John Wiley and Sons Ltd on behalf of European Food Safety Authority.

Oxidatively generated damage to DNA has been implicated in the pathogenesis of a wide variety of diseases. Increasingly, interest is also focusing upon the effects of damage to the other nucleic acids, RNA and the (2′-deoxy-)ribonucleotide pools, and evidence is growing that these too may have an important role in disease. LC-MS/MS has the ability to provide absolute quantification of specific biomarkers, such as 8-oxo-7,8-dihydro-2′-deoxyGuo (8-oxodG), in both nuclear and mitochondrial DNA, and 8-oxoGuo in RNA. However, significant quantities of tissue are needed, limiting its use in human biomonitoring studies. In contrast, the comet assay requires much less material, and as little as 5 μL of blood may be used, offering a minimally invasive means of assessing oxidative stress in vivo, but this is restricted to nuclear DNA damage only. Urine is an ideal matrix in which to non-invasively study nucleic acid-derived biomarkers of oxidative stress, and considerable progress has been made towards robustly validating these measurements, not least through the efforts of the European Standards Committee on Urinary (DNA) Lesion Analysis. For urine, LC-MS/MS is considered the gold standard approach, and although there have been improvements to the ELISA methodology, this is largely limited to 8-oxodG. Emerging DNA adductomics approaches, which either comprehensively assess the totality of adducts in DNA, or map DNA damage across the nuclear and mitochondrial genomes, offer the potential to considerably advance our understanding of the mechanistic role of oxidatively damaged nucleic acids in disease., Graphical abstract Image 1, Highlights • Oxidatively damaged nucleic acids are implicated in the pathogenesis of disease. • LC-MS/MS, comet assay and ELISA are often used to study oxidatively damaged DNA. • Urinary oxidatively damaged nucleic acids non-invasively reflect oxidative stress. • DNA adductomics will aid understanding the role of ROS damaged DNA in disease.

The present article examines recently published literature on lipids, mainly focusing on research involving glycero-, glycerophospho- and sphingo-lipids. The primary aim is identification of distinct profiles in biologic lipidomic systems by ultra-high-performance liquid chromatography (UHPLC) coupled with mass spectrometry (MS, tandem MS) with multivariate data analysis. This review specifically targets lipid biomarkers and disease pathway mechanisms in humans and artificial targets. Different specimen matrices such as primary blood derivatives (plasma, serum, erythrocytes, and blood platelets), faecal matter, urine, as well as biologic tissues (liver, lung and kidney) are highlighted.

Amikacin, gentamicin and tobramycin are aminoglycoside antibiotics mostly used for the treatment of a wide range of aerobic Gram negative bacteria. Lack of a UV chromophore as well as their polar and non-volatile nature make direct determination of these aminoglycosides by conventional LC and GC very challenging. Existing analytical methodology is either expensive, complicated or time consuming. In this work, a thin layer chromatography – flame ionization method was developed for fast analysis of these aminoglycosides in pharmaceutical formulations. Paromomycin was used as internal standard for quantitation. Development of the rod with one single solvent mixture was sufficient to achieve the desired separation of target components. Good determination coefficients (>0.997) were achieved for all target analytes. Repeatability, recovery and detection limits were determined and reported. A total of 5 samples in duplicate can be analyzed simultaneously within 25 s. This method offers the advantage of being able to analyze these aminoglycosides without any need for derivatization or laborious coloration. The developed method was applied to the analysis of some pharmaceutical commercial samples.

Vincristine (VCR) is frequently used in pediatric oncology and can be administered intravenously through push injections or 1 h infusions. The effects of administration duration on population pharmacokinetics (PK) are unknown. We described PK differences related to administration duration and the relation between PK and VCR-induced peripheral neuropathy (VIPN). PK was assessed in 1&ndash, 5 occasions (1&ndash, 8 samples in 24 h per occasion). Samples were analyzed using high-performance liquid chromatography/tandem mass spectrometry. Population PK of VCR and its relationship with administration duration was determined using a non-linear mixed effect. We estimated individual post-hoc parameters: area under the concentration time curve (AUC) and maximum concentration (Cmax) in the plasma and peripheral compartment. VIPN was assessed using Common Terminology Criteria for Adverse Events (CTCAE) and the pediatric-modified total neuropathy score (ped-mTNS). Overall, 70 PK assessments in 35 children were evaluated. The population estimated that the intercompartmental clearance (IC-Cl), volume of the peripheral compartment (V2), and Cmax were significantly higher in the push group. Furthermore, higher IC-Cl was significantly correlated with VIPN development. Administration of VCR by push led to increased IC-Cl, V2, and Cmax, but were similar to AUC, compared to 1 h infusions. Administration of VCR by 1 h infusions led to similar or higher exposure of VCR without increasing VIPN.

Vitamin K and its essential role in coagulation (vitamin K [Koagulation]) have been well established and accepted the world over. Many countries have a Recommended Daily Intake (RDI) for vitamin K based on early research, and its necessary role in the activation of vitamin K-dependent coagulation proteins is known. In the past few decades, the role of vitamin K-dependent proteins in processes beyond coagulation has been discovered. Various isoforms of vitamin K have been identified, and vitamin K2 specifically has been highlighted for its long half-life and extrahepatic activity, whereas the dietary form vitamin K1 has a shorter half-life. In this review, we highlight the specific activity of vitamin K2 based upon proposed frameworks necessary for a bioactive substance to be recommended for an RDI. Vitamin K2 meets all these criteria and should be considered for a specific dietary recommendation intake.

Nutrients 12(6), 1852 (2020). doi:10.3390/nu12061852 special issue: "Special Issue "Physiology and Pathology of Vitamin K Intake" / Special Issue Editors: Dr. Maria Fusaro, Prof. Dr. Maurizio Gallieni", Published by MDPI, Basel

Micronutrients are essential to health at every life stage and their deficiencies are associated with increased incidence of various pathophysiological states and poor quality of life. Efficient methods are therefore needed to monitor micronutrient status of individuals and to improve evidenced-based recommendations for populations. This review (i) reports current approaches to assess the vitamin and mineral status in humans, (ii) summarizes current analytical advantages and shortcomings and (iii) provides practical information for both nutrition research and nutritional status diagnostics. Future analytical perspectives are discussed in relation to micronutrient profiling, analytical sensitivity, and miniaturized technologies. (C) 2018 Elsevier B.V. All rights reserved.

Background: Therapeutic drug monitoring is useful in the treatment of tuberculosis to assure adequate exposure, minimize antibiotic resistance, and reduce toxicity. Salivary therapeutic drug monitoring could reduce the risks, burden, and costs of blood-based therapeutic drug monitoring. This systematic review compared human pharmacokinetics of antituberculosis drugs in saliva and blood to determine if salivary therapeutic drug monitoring could be a promising alternative.Methods: On December 2, 2016, PubMed and the Institute for Scientific Information Web of Knowledge were searched for pharmacokinetic studies reporting human salivary and blood concentrations of antituberculosis drugs. Data on study population, study design, analytical method, salivary Cmax, salivary area under the time-concentration curve, plasma/serum Cmax, plasma/serum area under the time-concentration curve, and saliva-plasma or saliva-serum ratio were extracted. All included articles were assessed for risk of bias.Results: In total, 42 studies were included in this systematic review. For the majority of antituberculosis drugs, including the first-line drugs ethambutol and pyrazinamide, no pharmacokinetic studies in saliva were found. For amikacin, pharmacokinetic studies without saliva-plasma or saliva-serum ratios were found.Conclusions: For gatifloxacin and linezolid, salivary therapeutic drug monitoring is likely possible due to a narrow range of saliva-plasma and saliva-serum ratios. For isoniazid, rifampicin, moxifloxacin, ofloxacin, and clarithromycin, salivary therapeutic drug monitoring might be possible; however, a large variability in saliva-plasma and saliva-serum ratios was observed. Unfortunately, sali-vary therapeutic drug monitoring is probably not possible for doripenem and amoxicillin/clavulanate, as a result of very low salivary drug concentrations.

Carminic acid, a glucosylated anthraquinone found in scale insects like Dactylopius coccus, has since ancient times been used as a red colorant in various applications. Here we show that a membrane-bound C-glucosyltransferase, isolated from D. coccus and designated DcUGT2, catalyzes the glucosylation of flavokermesic acid and kermesic acid into their respective C-glucosides dcII and carminic acid. DcUGT2 is predicted to be a type I integral endoplasmic reticulum (ER) membrane protein, containing a cleavable N-terminal signal peptide and a C-terminal transmembrane helix that anchors the protein to the ER, followed by a short cytoplasmic tail. DcUGT2 is found to be heavily glycosylated. Truncated DcUGT2 proteins synthesized in yeast indicate the presence of an internal ER-targeting signal. The cleavable N-terminal signal peptide is shown to be essential for the activity of DcUGT2, whereas the transmembrane helix/cytoplasmic domains, although important, are not crucial for its catalytic function., Carminic acid is a widely applied red colorant that is still harvested from insects because its biosynthesis is not fully understood. Here, the authors identify and characterize a membrane-bound C-glucosyltransferase catalyzing the final step during carminic acid biosynthesis.

Membrane lipid composition varies greatly within submembrane compartments, different organelle membranes, and also between cells of different cell stage, cell and tissue types, and organisms. Environmental factors (such as diet) also influence membrane composition. The membrane lipid composition is tightly regulated by the cell, maintaining a homeostasis that, if disrupted, can impair cell function and lead to disease. This is especially pronounced in the brain, where defects in lipid regulation are linked to various neurological diseases. The tightly regulated diversity raises questions on how complex changes in composition affect overall bilayer properties, dynamics, and lipid organization of cellular membranes. Here, we utilize recent advances in computational power and molecular dynamics force fields to develop and test a realistically complex human brain plasma membrane (PM) lipid model and extend previous work on an idealized, "average" mammalian PM. The PMs showed both striking similarities, despite significantly different lipid composition, and interesting differences. The main differences in composition (higher cholesterol concentration and increased tail unsaturation in brain PM) appear to have opposite, yet complementary, influences on many bilayer properties. Both mixtures exhibit a range of dynamic lipid lateral inhomogeneities ("domains"). The domains can be small and transient or larger and more persistent and can correlate between the leaflets depending on lipid mixture, Brain or Average, as well as on the extent of bilayer undulations.

Purpose: To explore factors associated with pentosidine accumulation in the human vitreous.Methods: Vitreous samples were obtained during trans pars plana vitrectomy for macular hole or rhegmatogenous retinal detachment. Patient characteristics included age, gender, and diabetes mellitus. Ocular characteristics included pseudophakia, posterior vitreous detachment, and presence of intraocular fibrosis (epiretinal membrane, proliferative vitreoretinopathy, or both). Pentosidine concentration as a measure of accumulation of advanced glycation end products was determined by high performance liquid chromatography.Results: Pentosidine concentrations were measured in 222 vitrectomy samples (118 female and 104 male patients [median age 66 years], treated for macular hole [n = 105] or rhegmatogenous retinal detachment [n = 117]). Pentosidine was found to accumulate significantly with age (P Conclusion: This study confirmed an age-related pentosidine accumulation in the vitreous and found new factors relating to pentosidine levels. Findings support the hypothesis of enzyme-induced vitreous liquefaction and the hypothesis of pentosidine as a pro-fibrotic factor.

Human milk oligosaccharides have been recognized as an important, functional biomolecule in mothers’ milk. Moreover, these oligosaccharides have been recognized as the third most abundant component of human milk, ranging from 10−15 g/L in mature milk and up to and over 20 g/L reported in colostrum. Initially, health benefits of human milk oligosaccharides were assigned via observational studies on the differences between breastfed and bottle fed infants. Later, pools of milk oligosaccharides were isolated and used in functional studies and in recent years more specific studies into structure−function relationships have identified some advanced roles for milk oligosaccharides in the healthy development of infants. In other research, the levels, diversity, and complexity of human milk oligosaccharides have been studied, showing a wide variation in results. This review gives a critical overview of challenges in the analysis of human milk oligosaccharides. In view of the myriad functions that can be assigned, often to specific structures or classes of structures, it is very relevant to assess the levels of these structures in the human milk correctly, as well as in other biological sample materials. Ultimately, the review makes a case for a comparative, inter-laboratory study on quantitative human milk oligosaccharide analysis in all relevant biological samples.

Quercetin has been shown to alleviate mucosal damage and modulate the glutathione (GSH) redox system in the colon of rodents. In the current study, we assessed whether quercetin was able to mitigate small intestinal dysfunction in weaned pigs. Here, 224 weaned piglets were fed a diet containing quercetin at either 0, 100, 300, or 900 mg/kg diet until d14 post-weaning, followed by a common basal diet until d42. Eight animals per treatment were sampled at d5 and d14 post-weaning. In these animals, the small intestinal histomorphology, barrier function, and protein abundance of occludin, caspase-3, and proliferating cell nuclear antigen were assessed. None of these parameters were affected, and neither did quercetin improve performance up to d42 post-weaning. The GSH redox system was evaluated in blood, small intestinal mucosa, and liver. Quercetin did not affect the glutathione peroxidase, glutathione reductase, and glutamate&ndash, cysteine ligase activity in these tissues. In contrast, the hepatic glutathione transferase (GST) activity was significantly increased by quercetin supplementation at d5 post-weaning of 100, 300, and 900 mg/kg. Importantly, d5 was characterized by a more oxidized GSH redox status. To conclude, dietary quercetin had little effect on the small intestine, but did upregulate hepatic GST in the occurrence of redox disturbance.

Liquid chromatography (LC) hyphenated to mass spectrometry is currently the most widely used means of determining metabolic phenotypes via both untargeted and targeted analysis. At present a range of analytical separations, including reversed-phase, hydrophilic interaction and ion-pair LC are employed to maximise metabolome coverage with ultra (high) performance liquid chromatography (UHPLC) increasingly displacing conventional high performance liquid chromatography because of the need for short analysis times and high peak capacity in such applications. However, it is widely recognized that these methodologies do not entirely solve the problems facing researchers trying to perform comprehensive metabolic phenotyping and in addition to these "routine" approaches there are continuing investigations of alternative separation methods including 2-dimensional/multi column approaches. These involve either new stationary phases or multidimensional combinations of the more conventional materials currently used, as well as application of miniaturization or "new" approaches such as supercritical HP and UHP- chromatographic separations. There is also a considerable amount of interest in the combination of chromatographic and ion mobility separations, with the latter providing both an increase in resolution and the potential to provide additional structural information via the determination of molecular collision cross section data. However, key problems remain to be solved including ensuring quality, comparability across different laboratories and the ever present difficulty of identifying unknowns.

Fat is an important component of human milk and infant formula (IF), delivering half of the energy a baby needs. Nowadays, mostly vegetable fats are used in IFs; however, the use of bovine milk fat in formulas is currently increasing. Bovine milk fat contains a composition of fatty acids and lipid components different from those of vegetable fats. We have compared the lipid profile of human and bovine milk with infant formulas with different fat sources. Furthermore, current knowledge of how infant digestion, absorption, metabolic responses, gut immunity, microbiota and/or cognition is affected by dietary fat is reviewed. The possible opportunities and drawbacks of the application of bovine milk fat in infant nutrition are described. Future perspectives for the development of IF containing bovine milk fat and future research directions are highlighted.

Due to its multifunctional and complex role in the honey bee colony functioning and health (construction material allowing food storage, brood rearing, thermoregulation, mediation in chemical and mechanical communication, substrate for pathogens, toxins and waste), Apis mellifera beeswax has been widely studied over the last five decades. This is supported by a comprehensive set of scientific reports covering different aspects of beeswax research. In this article, we present an overview of the methods for studying chemical, biological, constructional, and quality aspects of beeswax. We provide a detailed description of the methods for investigating wax scales, comb construction and growth pattern, cell properties, chemical composition of beeswax using different analytical tools, as well as the analytical procedures for provenancing beeswax and beeswax-derived compounds based on the hydrogen isotope ratio (IRMS). Along with classical physico-chemical and sensory analysis, we describe more precise and accurate methods for detection of adulterants in beeswax (GC-MS and FTIR-ATR). Moreover, we present methods for studying the influence of beeswax (comb foundation) adulteration on comb construction. Analytical protocols for determining the pesticide residues using different chromatographic and spectroscopic techniques are also described. As beeswax is an agent of high risk for the transmission of bee diseases, we present methods for detection of pathogens in beeswax. To ensure the reproducibility of experiments and results, we present best practice approaches and detailed protocols for all methods described, as well as their advantages and disadvantages. COST ActionEuropean Cooperation in Science and Technology (COST) [FA0803]; Ricola Foundation - Nature Culture We would like to acknowledge the Ricola Foundation - Nature & Culture, and COST Action FA0803 for funding the COLOSS (Prevention of honey bee COlony LOSSes) network and making the BEEBOOK possible. We would also like to express our gratitude to Editor Vincent Dietemann for his kind support all the way, and for providing helpful and constructive suggestions.

Introduction As large scale metabolic phenotyping is increasingly employed in preclinical studies and in the investigation of human health and disease the current LC–MS/MS profiling methodologies adopted for large sample sets can result in lengthy analysis times, putting strain on available resources. As a result of these pressures rapid methods of untargeted analysis may have value where large numbers of samples require screening. Objectives To develop, characterise and evaluate a rapid UHP-HILIC-MS-based method for the analysis of polar metabolites in rat urine and then extend the capabilities of this approach by the addition of IMS to the system. Methods A rapid untargeted HILIC LC–MS/MS profiling method for the analysis of small polar molecules has been developed. The 3.3 min separation used a Waters BEH amide (1 mm ID) analytical column on a Waters Synapt G2-Si Q-Tof enabled with ion mobility spectrometry (IMS). The methodology, was applied to the metabolic profiling of a series of rodent urine samples from vehicle-treated control rats and animals administered tienilic acid. The same separation was subsequently linked to IMS and MS to evaluate the benefits that IMS might provide for metabolome characterisation. Results The rapid HILIC–MS method was successfully applied to rapid analysis of rat urine and found, based on the data generated from the data acquired for the pooled quality control samples analysed at regular intervals throughout the analysis, to be robust. Peak area and retention times for the compounds detected in these samples showed good reproducibility across the batch. When used to profile the urine samples obtained from vehicle-dosed control and those administered tienilic acid the HILIC-MS method detected 3007 mass/retention time features. Analysis of the same samples using HILIC–IMS–MS enabled the detection of 6711 features. Provisional metabolite identification for a number of compounds was performed using the high collision energy MS/MS information compared against the Metlin MS/MS database and, in addition, both calculated and measured CCS values from an experimentally derived CCS database. Conclusion A rapid metabolic profiling method for the analysis of polar metabolites has been developed. The method has the advantages of speed and both reducing sample and solvent consumption compared to conventional profiling methods. The addition of IMS added an additional dimension for feature detection and the identification of metabolites. Electronic supplementary material The online version of this article (10.1007/s11306-019-1474-9) contains supplementary material, which is available to authorized users.

Effluents from wastewater treatment plants (WWTPs) are a major source of estrogenic compounds to the aquatic environment. In the present work, estrogenic activities of effluents from eight municipal WWTPs in Finland were studied. The main objectives of the study were to quantify the concentrations of selected estrogenic compounds, to evaluate their contribution to estrogenic potency and to test the feasibility of the commercial bioassays for wastewater analysis. The effluent samples were analyzed by two in vitro tests, i.e. ERα-CALUX(®) and ELISA-E2, and by liquid chromatography mass spectrometry for six estrogenic compounds: estrone (E1), 17β-estradiol (E2), estriol (E3), 17α-ethinylestradiol (EE2), 17α-estradiol and bisphenol A (BPA). Estrogenic effects were found in all of the effluent samples with both of the bioassays. The concentrations measured with ELISA-E2 (8.6-61.6 ng/L) were clearly higher but exhibited a similar pattern than those with chemical analysis (E2

International audience; The numbers of potential neurotoxicants in the environment are raising and pose a great risk for humans and the environment. Currently neurotoxicity assessment is mostly performed to predict and prevent harm to human populations. Despite all the efforts invested in the last years in developing novel in vitro or in silico test systems, in vivo tests with rodents are still the only accepted test for neurotoxicity risk assessment in Europe. Despite an increasing number of reports of species showing altered behaviour, neurotoxicity assessment for species in the environment is not required and therefore mostly not performed. Considering the increasing numbers of environmental contaminants with potential neurotoxic potential, eco-neurotoxicity should be also considered in risk assessment. In order to do so novel test systems are needed that can cope with species differences within ecosystems. In the field, online-biomonitoring systems using behavioural information could be used to detect neurotoxic effects and effect-directed analyses could be applied to identify the neurotoxicants causing the effect. Additionally, toxic pressure calculations in combination with mixture modelling could use environmental chemical monitoring data to predict adverse effects and prioritize pollutants for laboratory testing. Cheminformatics based on computational toxicological data from in vitro and in vivo studies could help to identify potential neurotoxicants. An array of in vitro assays covering different modes of action could be applied to screen compounds for neurotoxicity. The selection of in vitro assays could be guided by AOPs relevant for eco-neurotoxicity. In order to be able to perform risk assessment for eco-neurotoxicity, methods need to focus on the most sensitive species in an ecosystem. A test battery using species from different trophic levels might be the best approach. To implement eco-neurotoxicity assessment into European risk assessment, cheminformatics and in vitro screening tests could be used as first approach to identify eco-neurotoxic pollutants. In a second step, a small species test battery could be applied to assess the risks of ecosystems.

International audience; 3-Carboxy-4-methyl-5-propyl-2-furanpropionate (CMPF) is a metabolite of furan fatty acid and a marker of fish oil intake. CMPF is described as a protein-bound uremic toxin and interacts with free oxygen radicals, which can induce cell damages. However, the clinical consequences of CMPF accumulation in haemodialysis patients remain poorly documented. The aims of this study are to investigate potential association between CMPF levels and (i) biochemical and nutritional parameters; (ii) cardiovascular events and (iii) mortality. Two hundred and fifty-two patients undergoing maintenance haemodialysis were included. Routine clinical biochemistry tests and assay for CMPF by HPLC technique were performed at the inclusion. Body composition parameters were measured using a bioimpedance spectroscopy method. The enrolled patients were prospectively monitored for cardiovascular events and mortality. CMPF level was positively correlated with nutritional parameters and lean mass and is significantly higher in patients without protein-energy wasting. However, the multivariate linear regression analysis indicated that CMPF level was not independently associated with albumin, prealbumin, creatinemia and body mass index. Elevated serum CMPF was not associated with mortality and cardiovascular morbidity. Our results indicate that CMPF is not a relevant uremic toxin in haemodialysis and in contrast could be a marker of healthy diet and omega 3 intakes.

Vitamin K₂ (menaquinone) concentrations were measured in a wide range of cheeses and the effects of fat content, ripening and origin of the cheeses were investigated. Moreover, the menaquinone content of cheese was compared with that of other foods known to contain vitamin K₂. It was found that cheese and curd are the most important sources of long-chain menaquinones in the Western diet and, in general, hard cheeses are richer in menaquinones than soft cheeses. However, the actual menaquinone content varies substantially and is dependent on the type of cheese, the time of ripening, the fat content and the geographic area where the cheeses are produced. Given the fact that poor vitamin K status has been mentioned as a risk factor for cardiovascular disease and mortality, while there is no clear evidence for adverse cardiovascular effects of dairy fats, cheese should be considered as a recommendable component in a heart-healthy diet.

Salsolinol (1-methyl-6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline) is a tetrahydroisoquinoline derivative whose presence in humans was first detected in the urine of Parkinsonian patients on l-DOPA (l-dihydroxyphenylalanine) medication. Thus far, multiple hypotheses regarding its physiological/pathophysiological roles have been proposed, especially related to Parkinson’s disease or alcohol addiction. The aim of this review was to outline studies related to salsolinol, with special focus on in vivo and in vitro experimental models. To begin with, the chemical structure of salsolinol together with its biochemical implications and the role in neurotransmission are discussed. Numerous experimental studies are summarized in tables and the most relevant ones are stressed. Finally, the ability of salsolinol to cross the blood–brain barrier and its possible double-faced neurobiological potential are reviewed.

Five different quorum sensing peptides (QSP) were iodinated using different iodination techniques. These iodinated peptides were analyzed using a C-18 reversed phase HPLC system, applying a linear gradient of water and acetonitrile containing 0.1% (m/v) formic acid as mobile phase. Electrospray ionization (ESI) ion trap mass spectrometry was used for the identification of the modified peptides, while semi-quantification was performed using total ion current (TIC) spectra. Non-iodinated peptides and mono- and di-iodinated peptides (NIP, MIP and DIP respectively) were well separated and eluted in that order. Depending on the used iodination method, iodination yields varied from low (2%) to high (57%).

A sensitive and specific method for the quantitative determination of Fumonisin B1 (FB1), its partially hydrolysed metabolites pHFB1a+b and hydrolysed metabolite HFB1, and Fumonisin B2 (FB2) in broiler chicken plasma using ultra-performance liquid chromatography combined with tandem mass spectrometry (UPLC-MS/MS) was developed. The sample preparation was rapid, straightforward and consisted of a deproteinization and phospholipid removal step using an Oasis((R)) Ostro(TM) 96-well plate. Chromatography was performed on an Acquity HSS-T3 column, using 0.3% formic acid and 10 mM ammonium formate in water, and acetonitrile as mobile phases. The MS/MS instrument was operated in the positive electrospray ionization mode and the two multiple reaction monitoring transitions were monitored for each component for quantification and identification, respectively. The method was validated in-house: matrix-matched calibration graphs were prepared and good linearity (r 0.99) was achieved over the concentration ranges tested (1-500 ng/mL for FB1 and FB2; 0.86-860 ng/mL for pHFB1a; 0.72-1430 ng/mL for pHFB1b and 2.5-2500 ng/mL for HFB1). Limits of quantification (LOQ) and detection (LOD) in plasma ranged between 0.72 to 2.5 ng/mL and 0.03 to 0.17 ng/mL, respectively. The results for the within-day and between-day precision and accuracy fell within the specified ranges. Moreover, the method was transferred to an UPLC high-resolution mass spectrometry (HR-MS) instrument in order to determine potential metabolites of HFB1, such as N-acyl-HFB1s and phase II metabolites. The method has been successfully applied to investigate the toxicokinetics and biotransformation of HFB1 in broiler chickens.

In this study, an ID-LC-MS method was developed and validated for the determination of 14 sulfonamides in milk samples. Recovery results were found to be in the range of 91%-114% for milk samples. The relative measurement uncertainty was between 7.5%-12.7%. Validated method was performed on milk samples obtained from market and street vendors. The amount of sulfonamides in the analyzed samples was found to be below the legal limits (Sulfamethazine: 6.46 +/- 0.76 ng/g and sulfisoxazole: 7.3 +/- 0.71 ng/g).

In agriforensics, time of administration is often debated when illegal drug residues, such as clenbuterol, are found in frequently traded cattle. In this proof-of-concept work, the feasibility of obtaining retrospective timeline information from segmented calf tail hair analyses has been studied. First, an ultraperformance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) hair analysis method was adapted to accommodate smaller sample sizes and in-house validated. Then, longitudinal 1 cm segments of calf tail hair were analyzed to obtain clenbuterol concentration profiles. The profiles found were in good agreement with calculated, theoretical positions of the clenbuterol residues along the hair. Following assessment of the average growth rate of calf tail hair, time of clenbuterol administration could be retrospectively determined from segmented hair analysis data. The data from the initial animal treatment study (n = 2) suggest that time of treatment can be retrospectively estimated with an error of 3-17 days.

Lichens and free-living terrestrial algae are widespread across many habitats and develop successfully in ecosystems where a cold winter limits survival. With the goal of comparing photoprotective responses in free-living and lichenized algae, the physiological responses to chilling and photochilling conditions were studied in three lichens and their isolated algal photobionts together as well as in a fourth free-living algal species. We specifically addressed the following questions: (i) Are there general patterns of acclimation in green algae under chilling and photochilling stresses? (ii) Do free-living algae exhibit a similar pattern of responses as their lichenized counterparts? (iii) Are these responses influenced by the selection pressure of environmental conditions or by the phylogenetic position of each species? To answer these questions, photosynthetic fluorescence measurements as well as pigment and low molecular weight carbohydrate pool analyses were performed under controlled laboratory conditions. In general, photochemical efficiency in all free-living algae decreased with increasing duration of the stress, while the majority of lichens maintained an unchanged photochemical activity. Nevertheless, these patterns cannot be generalized because the alga Trebouxia arboricola and the lichen Ramalina pollinaria (associated with Trebouxia photobionts) both showed a similar decrease in photochemical efficiency. In contrast, in the couple Elliptochloris bilobata-Baeomyces rufus, only the algal partner exhibited a broad physiological performance under stress. This study also highlights the importance of the xanthophyll cycle in response to the studied lichens and algae to photochilling stress, while the accumulation of sugars was not related to cold acclimation, except in the alga E. bilobata. The differences in response patterns detected among species can be mainly explained by their geographic origin, although the phylogenetic position should also be considered, especially in some species. FM received a fellowship for PhD students (2012-2015) from the Government of the Basque Country as well as a mobility grant which allowed the collaboration between University of Basque Country (UPV/EHU) and University of Rostock. LG thanks the Deutsche Forschungsgemeinschaft for financial support (DFG GU 1278/1-1). The work was also supported by the project BFU 2010-15021 and CTM2014-53902-C2-2-P from the MINECO and the European Regional Development Fund ERDF (FEDER), and by the Basque Government (UPV/EHU-GV IT-624-13 and UPV/EHU-GV IT-1018-16).

International audience; RATIONALE: The first step in the diagnosis of oligosaccharidoses is to evidence abnormal oligosaccharides excreted in urine, usually performed by the poorly sensitive but efficient thin layer chromatography (TLC) method. Developing a tandem mass spectrometry (MS/MS) technique could be of great interest to replace TLC. METHODS: Abnormal underivatized oligosaccharides have been recently studied using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, allowing the unambiguous identification of oligosaccharidoses. Based on this previous work, we developed an advantageous and efficient liquid chromatography (LC)/MS/MS method using a more common triple quadrupole tandem mass spectrometer for oligosaccharides analysis. RESULTS: Oligosaccharidoses (n = 97) and control (n = 240) urine samples were analysed. A specific pattern was obtained for each oligosaccharidosis using this method. In urine, it allows not only the identification of all the oligosaccharidoses previously identified by TLC (fucosidosis, alphamannosidosis, aspartylglucosaminuria, GM1 gangliosidosis, sialidosis, galactosialidosis and Schindler disease), but also extends the field of diagnosis to mucolipidosis type II, Sandhoff disease, and beta-mannosidosis. The same technique was applied to 16 amniotic fluid supernatants from oligosaccharidosis-affected foetuses (n = 16) compared with 37 unaffected. All the affected foetuses could be clearly identified: sialidosis (n = 3), galactosialidosis (n = 4), aspartylglucosaminuria (n = 1), mucolipidosis type II (n = 4) or GM1 gangliosidosis (n = 4). This technique can be applied to early prenatal diagnosis as well as to the oligosaccharidosis screening in the case of non-immune hydrops fetalis. CONCLUSIONS: The method is quick and easy to run, with an LC analysis time of 13 min per sample. The quantitative validation could not be obtained in the absence of a specific standard and of a labelled internal standard for each compound. Even if this LC/MS/MS method is only qualitative, it is very specific and much more sensitive than TLC. It allows the urinary screening of oligosaccharidoses, even mild or late-onset forms, and the screening of antenatal forms in amniotic fluid. Copyright (C) 2017 John Wiley & Sons, Ltd.

A proportion of the plant's L-ascorbate (vitamin C) occurs in the apoplast, where it and its metabolitesmay act as pro-oxidants and anti-oxidants. One ascorbate metabolite is 2,3-dilcetogulonate (DKG), preparations of which can non-enzymically generate H2O2 and delay peroxidase action on aromatic substrates. As DKG itself generates several by-products, we characterised these and their ability to generate H2O2 and delay peroxidase action. DKG preparations rapidly produced a by-product, compound (1), with lambda(max) 271 and 251 nm at neutral and acidic pH respectively. On HPLC, (1) co-eluted with the major H2O2-generating and peroxidase-delaying principle. Compound (1) was slowly destroyed by ascorbate oxidase, and was less stable at pH 6 than at pH 1. Electrophoresis of an HPLC-enriched preparation of (1) suggested a strongly acidic (pK(a) approximate to 2.3) compound. Mass spectrometry suggested that un-ionised (1) has the formula C6H6O5, i.e. it is a reduction product of DKG (C6H8O7). In conclusion, compound (1) is the major H2O2-generating, peroxidase-delaying principle formed non-enzymically from DKG in the pathway ascorbate -> dehydroascorbic acid -> DKG -> (1). We hypothesise that (1) generates apoplastic H2O2 (and consequently hydroxyl radicals) and delays cell-wall crosslinking - both these effects favouring wall loosening, and possibly playing a role in pathogen defence. (C) 2017 The Authors. Published by Elsevier Inc.

The beneficial effects of honey on human health have long been recognized. Today, many of those positive effects have been studied to elucidate its mode of action. This review briefly summarizes the best studied features of honey, highlighting it as an appealing alternative medicine. In these reports, the health benefits of honey range from antioxidant, immunomodulatory, and anti-inflammatory activity to anticancer action, metabolic and cardiovascular benefits, prebiotic properties, human pathogen control, and antiviral activity. These studies also support that the honey's biological activity is mainly dependent on its floral or geographic origin. In addition, some promising synergies between honey and antibiotics have been found, as well as some antiviral properties that require further investigation. Altogether, these studies show that honey is effectively a nutraceutical foodstuff. info:eu-repo/semantics/publishedVersion

In this study, the stability constants of the copper(II) and aluminium(III) complexes of quercetin were determined potentiometrically by using Calvin-Bjerrum and Irwing Rossotti methods. The protonation constants of quercetin were found: logK(1) = 11.15 +/- 0.118, logK(2) = 10.42 +/- 0.144, logK(3) = 9.44 +/- 0.162, logK(4) = 8.28 +/- 0.151. For copper complex formation constant was found by Irwing-Rossotti method: logK(1) = 19.92 +/- 0.367 and for aluminium complex complex: logK(1) = 23.02 +/- 0.459. From the results, the components of quercetin / metal complexes are given as 1/1 both copper(II) and aluminium(III). A reversed phase high pressure liquid chromatographic method was developed for the metal complexes which were prepared according to potentiometric results with the ratio quercetin / metal = 1/1. Mobile phase was 0.01 M HClO4/8.33 x 10(-5) M quercetin in Methanol (40/60), column was XTerra RP18, 5 mu m, 4.6 x 150 mm, detector was Diode Array Detector at lambda = 373 ve 421 nm (band width 4nm).

International audience; Folates, also known as B9 vitamins, serve as donors and acceptors in one-carbon (C1) transfer reactions. The latter are involved in synthesis of many important biomolecules, such as amino acids, nucleic acids and vitamin B5. Folates also play a central role in the methyl cycle that provides one-carbon groups for methylation reactions. The important functions fulfilled by folates make them essential in all living organisms. Plants, being able to synthesize folates de novo, serve as an excellent dietary source of folates for animals that lack the respective biosynthetic pathway. Unfortunately, the most important staple crops such as rice, potato and maize are rather poor sources of folates. Insufficient folate consumption is known to cause severe developmental disorders in humans. Two approaches are employed to fight folate deficiency: pharmacological supplementation in the form of folate pills and biofortification of staple crops. As the former approach is considered rather costly for the major part of the world population, biofortification of staple crops is viewed as a decent alternative in the struggle against folate deficiency. Therefore, strategies, challenges and recent progress of folate enhancement in plants will be addressed in this review. Apart from the ever-growing need for the enhancement of nutritional quality of crops, the world population faces climate change catastrophes or environmental stresses, such as elevated temperatures, drought, salinity that severely affect growth and productivity of crops. Due to immense diversity of their biochemical functions, folates take part in virtually every aspect of plant physiology. Any disturbance to the plant folate metabolism leads to severe growth inhibition and, as a consequence, to a lower productivity. Whereas today's knowledge of folate biochemistry can be considered very profound, evidence on the physiological roles of folates in plants only starts to emerge. In the current review we will discuss the implication of folates in various aspects of plant physiology and development.

Induced mutations have been used effectively for plant improvement. Physical and chemical mutagens induce a high frequency of genome variation. Recently, developed screening methods have allowed the detection of single nucleotide polymorphisms (SNPs) and the identification of traits that are difficult to identify at the molecular level by conventional breeding. With the assistance of reverse genetic techniques, sequence variation information can be linked to traits to investigate gene function. Targeting induced local lesions in genomes (TILLING) is a high-throughput technique to identify single nucleotide mutations in a specific region of a gene of interest with a powerful detection method resulted from chemical-induced mutagenesis. The main advantage of TILLING as a reverse genetics strategy is that it can be applied to any species, regardless of genome size and ploidy level. However, TILLING requires laborious and time-consuming steps, and a lack of complete genome sequence information for many crop species has slowed the development of suitable TILLING targets. Another method, high-resolution melting (HRM), which has assisted TILLING in mutation detection, is faster, simpler and less expensive with non-enzymatic screening system. Currently, the sequencing of crop genomes has completely changed our vision and interpretation of genome organization and evolution. Impressive progress in next-generation sequencing (NGS) technologies has paved the way for the detection and exploitation of genetic variation in a given DNA or RNA molecule. This review discusses the applications of TILLING in combination with HRM and NGS technologies for screening of induced mutations and discovering SNPs in mutation breeding programs.

A targeted reversed-phase gradient UPLC-MS/MS assay has been developed for the quantification/monitoring of amino acids and amino-containing compounds in human plasma and serum using pre-column derivatization with 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (AccQTag UltraTM). Derivatization of the target amino-containing compounds reagent required minimal sample preparation and resulted in analytes with excellent chromatographic and mass spectrometric properties. The resulting method, which requires only 10 µl of sample, provides the reproducible and robust separation of 66 analytes in 7.5 minutes, including baseline resolution of isomers such as e.g. leucine and isoleucine. The assay has been validated for the quantification of 33 amino compounds (predominantly amino acids) over a concentration range from 2-20 and 800µM. Intra- and inter-day accuracy of between 0.05-15.6 and 0.78 -13.7 % and precision between 0.91-16.9 % and 2.12-15.9 % were obtained. A further 33 biogenic amines can be monitored in samples for relative changes in concentration rather than quantification. Application of the assay to samples derived from healthy controls and patients suffering from acetaminophen (APAP, paracetamol) induced acute liver failure (ALF) showed significant differences in the amounts of aromatic and branched chain amino acids between the groups as well as a number of other analytes, including the novel observation of increased concentrations of sarcosine in ALF patients. The properties of the developed assay, including short analysis time, make it suitable for high throughput targeted UPLC-ESI-MS/MS metabonomic analysis in clinical and epidemiological environments.

This study has been aimed at providing a qualitative and quantitative evaluation of selected phytochemicals such as phenolic acids, flavonoids, oleuropein, fatty acids profile, and volatile oil compounds, present in wild olive leaves harvested in Portugal, as well as at determining their antioxidant and cytotoxic potential against human melanoma HTB-140 and WM793, prostate cancer DU-145 and PC-3, hepatocellular carcinoma Hep G2 cell lines, as well as normal human skin fibroblasts BJ and prostate epithelial cells PNT2. Gallic, protocatechuic, p-hydroxybenzoic, vanillic acids, apigenin 7-O-glucoside, luteolin 7-O-glucoside, and rutin were identified in olive leaves. The amount of oleuropein was equal to 22.64 g/kg dry weight. (E)-Anethole (32.35%), fenchone (11.89%), and (Z)-3-nonen-1-ol (8%) were found to be the main constituents of the oil volatile fraction, whereas palmitic, oleic, and alpha-linolenic acid were determined to be dominating fatty acids. Olive leaves methanol extract was observed to exerted a significant, selective cytotoxic effect on DU-145 and PC-3 cell lines. Except the essential oil composition, evaluated wild olive leaves, with regard to their quantitative and qualitative composition, do not substantially differ from the leaves of other cultivars grown for industrial purposes and they reveal considerable antioxidant and cytotoxic properties. Thus, the wild species may prove to be suitable for use in traditional medicine as cancer chemoprevention. Polish Ministry of Science and Higher Education [K/ZDS/004122, K/ZDS/006239, K/ZDS/006228] info:eu-repo/semantics/publishedVersion

Dehydroascorbate reductase (DHAR) catalyzes the glutathione (GSH)-dependent reduction of dehydroascorbate and plays a direct role in regenerating ascorbic acid, an essential plant antioxidant vital for defense against oxidative stress. DHAR enzymes bear close structural homology to the glutathione transferase (GST) superfamily of enzymes and contain the same active site motif, but most GSTs do not exhibit DHAR activity. The presence of a cysteine at the active site is essential for the catalytic functioning of DHAR, as mutation of this cysteine abolishes the activity. Here we present the crystal structure of DHAR2 from Arabidopsis thaliana with GSH bound to the catalytic cysteine. This structure reveals localized conformational differences around the active site which distinguishes the GSH-bound DHAR2 structure from that of DHAR1. We also unraveled the enzymatic step in which DHAR releases oxidized glutathione (GSSG). To consolidate our structural and kinetic findings, we investigated potential conformational flexibility in DHAR2 by normal mode analysis and found that subdomain mobility could be linked to GSH binding or GSSG release.

Haptoglobin (Hp) is an acute phase protein that has recently been linked to components of the metabolic syndrome (MetS). We aimed to evaluate Hp as marker of MetS, and to assess its association with long-term outcome in renal transplant recipients (RTR). We measured plasma Hp in a prospective cohort of 699 stable RTR and 149 healthy controls. Median plasma Hp concentration in RTR was 1.4 [interquartile range (IQR), 1.0–1.8] g/L, which was higher compared to 1.1 [0.9–1.4] g/L in controls (P

Based on the recent scientific opinion of the European Food Safety Authority (EFSA) Panel on Contaminants in the Food Chain on the risks to human and animal health related to the presence of T-2 and HT-2 toxins in food and feed that was published by EFSA in the EFSA Journal, this article provides an update on the determination of these Fusarium mycotoxins. After a brief introduction into the chemistry of these toxins, both chromatographic and immuno-analytical methods are discussed for the determination of these type A trichothecenes. During the last decade, liquid chromatography with (tandem) mass spectrometry has become the most frequently used method for the determination of T-2 and HT-2 toxins, often within a multi-analyte approach. However, complex matrices and the resulting signal suppression effects, as observed particularly in electrospray-mass spectrometry methods owing to matrix effects, may require careful optimisation of clean-up, usage of matrix matched standards, or e.g. the use of internal standards. For specific purposes where extremely low limits of quantification are needed, e.g. for the analysis of duplicate diets, a dedicated gas chromatography method with multistage mass spectrometry has become available. Other novel analytical approaches to determine T-2 and HT-2 toxins in food and feed include biosensor-based methods in surface plasmon resonance and electrochemical formats, as well as DNA microchip assays. For rapid screening, several immunochemical methods (mostly ELISAs) have become available and some are sold as commercial test kits. Whereas these methods work fast, cross-reactivities with other trichothecenes can have an undesired effect on their accuracy. While proficiency tests including T-2 and HT-2 toxins have been carried out, none of the chromatographic or immunochemical methods have been formally validated in interlaboratory validation studies. There are no certified reference materials available for T-2 and HT-2 toxins.