PubMed

Metabolic changes associated with methionine stress sensitivity in MDA-MB-468 breast cancer cells. Cancer Metab. 2016;4:9 Authors: Borrego SL, Fahrmann J, Datta R, Stringari C, Grapov D, Zeller M, Chen Y, Wang P, Baldi P, Gratton E, Fiehn O, Kaiser P Abstract BACKGROUND: The majority of cancer cells have a unique metabolic requirement for methionine that is not observed in normal, non-tumorigenic cells. This phenotype is described as "methionine dependence" or "methionine stress sensitivity" in which cancer cells are unable to proliferate when methionine has been replaced with its metabolic precursor, homocysteine, in cell culture growth media. We focus on the metabolic response to methionine stress in the triple negative breast cancer cell line MDA-MB-468 and its methionine insensitive derivative cell line MDA-MB-468res-R8. RESULTS: Using a variety of techniques including fluorescence lifetime imaging microscopy (FLIM) and extracellular flux assays, we identified a metabolic down-regulation of oxidative phosphorylation in both MDA-MB-468 and MDA-MB-468res-R8 cell types when cultured in homocysteine media. Untargeted metabolomics was performed by way of gas chromatography/time-of-flight mass spectrometry on both cell types cultured in homocysteine media over a period of 2 to 24 h. We determined unique metabolic responses between the two cell lines in specific pathways including methionine salvage, purine/pyrimidine synthesis, and the tricarboxylic acid cycle. Stable isotope tracer studies using deuterium-labeled homocysteine indicated a redirection of homocysteine metabolism toward the transsulfuration pathway and glutathione synthesis. This data corroborates with increased glutathione levels concomitant with increased levels of oxidized glutathione. Redirection of homocysteine flux resulted in reduced generation of methionine from homocysteine particularly in MDA-MB-468 cells. Consequently, synthesis of the important one-carbon donor S-adenosylmethionine (SAM) was decreased, perturbing the SAM to S-adenosylhomocysteine ratio in MDA-MB-468 cells, which is an indicator of the cellular methylation potential. CONCLUSION: This study indicates a differential metabolic response between the methionine sensitive MDA-MB-468 cells and the methionine insensitive derivative cell line MDA-MB-468res-R8. Both cell lines appear to experience oxidative stress when methionine was replaced with its metabolic precursor homocysteine, forcing cells to redirect homocysteine metabolism toward the transsulfuration pathway to increase glutathione synthesis. The methionine stress resistant MDA-MB-468res-R8 cells responded to this cellular stress earlier than the methionine stress sensitive MDA-MB468 cells and coped better with metabolic demands. Additionally, it is evident that S-adenosylmethionine metabolism is dependent on methionine availability in cancer cells, which cannot be sufficiently supplied by homocysteine metabolism under these conditions. PMID: 27141305 [PubMed - as supplied by publisher]

Low-level environmental phthalate exposure associates with urine metabolome alteration in a Chinese male cohort. Environ Sci Technol. 2016 May 3; Authors: Zhang J, Liu L, Wang X, Huang Q, Tian M, Shen H Abstract The general population is exposed to phthalates through various sources and routes. Integration of omics data and epidemiological data is a key step towards directly linking phthalate bio-monitoring data with biological response. Urine metabolomics is a powerful tool to identify exposure biomarkers and delineate the modes of action of environmental stressors. The objectives of this study are to investigate the association between low-level environmental phthalate exposure and urine metabolome alteration in male population, and to unveil the metabolic pathways involved in the mechanisms of phthalate toxicity. In this retrospective cross-sectional study, we studied the urine metabolomic profiles of 364 male subjects exposed to low-level environmental phthalates. Di(2-ethylhexyl) phthalate (DEHP) and dibutyl phthalate (DBP) are the most widely used phthalates. ΣDEHP and MBP (the major metabolite of DBP) were associated with significant alteration of global urine metabolome in the male population. We observed significant increase in the levels of acetylneuraminic acid, carnitine C8:1, carnitine C18:0, cystine, phenylglycine, phenylpyruvic acid and glutamylphenylalanine; and meanwhile, decrease in the levels of carnitine C16:2, diacetylspermine, alanine, taurine, tryptophan, ornithine, methylglutaconic acid, hydroxyl-PEG2 and keto-PGE2 in high exposure group. The observations indicated that low-level environmental phthalate exposure associated with increased oxidative stress and fatty acid oxidation and decreased prostaglandin metabolism. Urea cycle, tryptophan and phenylalanine metabolism disruption was also observed. The urine metabolome disruption effects associated with ΣDEHP and MEP were similar, but not identical. The multi-biomarker models presented AUC values of 0.845 and 0.834 for ΣDEHP and MEP, respectively. The predictive accuracy rates of established models were 81% for ΣDEHP and 73% for MEP. Our results suggest that low-level environmental phthalate exposure associates with urine metabolome disruption in male population, providing new insight into the early molecular events of phthalate exposure. PMID: 27138838 [PubMed - as supplied by publisher]

Non-targeted metabolomics identified a common metabolic signature of lethal ventricular tachyarrhythmia (LVTA) in two rat models. Mol Biosyst. 2016 May 3; Authors: Wang X, Wang D, Yu X, Zhang G, Wu J, Zhu G, Su R, Lv J Abstract Lethal ventricular tachyarrhythmia (LVTA) is the predominant underlying mechanism of sudden cardiac death (SCD). The aim of this study is to characterize the metabolic features of myocardia following LVTA, and identify potential biomarkers to diagnose LVTA. We developed two LVTA rat models induced by aconitine injection or coronary artery ligation, which represent cardiac ion channel disease-related and cardiac ischemia-related SCD, respectively. The myocardial metabolic profile was investigated by gas chromatography-mass spectrometry and proton nuclear magnetic resonance-based metabolomics. Twenty-three aconitine-injected and 14 coronary artery ligation-treated rats developed LVTA SCD. A total of 38 differential metabolites of myocardia were identified in aconitine-induced LVTA rats, of which 31 metabolites showed a similar change in coronary artery ligation-related LVTA rats. Fatty acids (stearic, palmitic, linoleic, elaidic, and myristic) and branched-chain amino acids (valine, leucine, and isoleucine) were the most down-regulated metabolites. Furthermore, elevated ADP, phosphate, lactate, glutamate, aspartate, threonine, choline and arginine were also observed. Major pathways regarding these dysregulated metabolites post LVTA are energy excessive consumption and deficit, ionic imbalance, oxidative stress, cardiac cytotoxicity and membrane injury. Valine, stearic acid and leucine collectively enable a precision of 92.9% to distinguish LVTA from its control, and are correlated with several arrhythmia indices. Our results uncovered a common metabolic feature of LVTA in myocardia in two rat models, which represent cardiac ion channel disease and cardiac ischemia, respectively. l-Valine, l-leucine and stearic acid jointly confer good potential for distinguishing LVTA, which might be potential biomarkers of LVTA-related SCD. PMID: 27138062 [PubMed - as supplied by publisher]

SWAPDT: A method for Short-time Withering Assessment of Probability for Drought Tolerance in Camellia sinensis validated by targeted metabolomics. J Plant Physiol. 2016 Apr 19;198:39-48 Authors: Nyarukowa C, Koech R, Loots T, Apostolides Z Abstract Climate change is causing droughts affecting crop production on a global scale. Classical breeding and selection strategies for drought-tolerant cultivars will help prevent crop losses. Plant breeders, for all crops, need a simple and reliable method to identify drought-tolerant cultivars, but such a method is missing. Plant metabolism is often disrupted by abiotic stress conditions. To survive drought, plants reconfigure their metabolic pathways. Studies have documented the importance of metabolic regulation, i.e. osmolyte accumulation such as polyols and sugars (mannitol, sorbitol); amino acids (proline) during drought. This study identified and quantified metabolites in drought tolerant and drought susceptible Camellia sinensis cultivars under wet and drought stress conditions. For analyses, GC-MS and LC-MS were employed for metabolomics analysis.%RWC results show how the two drought tolerant and two drought susceptible cultivars differed significantly (p≤0.05) from one another; the drought susceptible exhibited rapid water loss compared to the drought tolerant. There was a significant variation (p<0.05) in metabolite content (amino acid, sugars) between drought tolerant and drought susceptible tea cultivars after short-time withering conditions. These metabolite changes were similar to those seen in other plant species under drought conditions, thus validating this method. The Short-time Withering Assessment of Probability for Drought Tolerance (SWAPDT) method presented here provides an easy method to identify drought tolerant tea cultivars that will mitigate the effects of drought due to climate change on crop losses. PMID: 27137993 [PubMed - as supplied by publisher]

Bridging the Gaps: the Promise of Omics Studies in Pediatric Exercise Research. Pediatr Exerc Sci. 2016 May;28(2):194-201 Authors: Radom-Aizik S, Cooper DM Abstract In this review, we highlight promising new discoveries that may generate useful and clinically relevant insights into the mechanisms that link exercise with growth during critical periods of development. Growth in childhood and adolescence is unique among mammals and is a dynamic process regulated by an evolution of hormonal and inflammatory mediators, age-dependent progression of gene expression, and environmentally modulated epigenetic mechanisms. Many of these same processes likely affect molecular transducers of physical activity. How the molecular signaling associated with growth is synchronized with signaling associated with exercise is poorly understood. Recent advances in "omics"-namely genomics and epigenetics, metabolomics, and proteomics-now provide exciting approaches and tools that can be used for the first time to address this gap. A biologic definition of "healthy" exercise that links the metabolic transducers of physical activity with parallel processes that regulate growth will transform health policy and guidelines that promote optimal use of physical activity. PMID: 27137166 [PubMed - as supplied by publisher]

Related Articles Profiling the Oxylipin and Endocannabinoid Metabolome by UPLC-ESI-MS/MS in Human Plasma to Monitor Postprandial Inflammation. PLoS One. 2015;10(7):e0132042 Authors: Gouveia-Figueira S, Späth J, Zivkovic AM, Nording ML Abstract Bioactive lipids, including oxylipins, endocannabinoids, and related compounds may function as specific biochemical markers of certain aspects of inflammation. However, the postprandial responsiveness of these compounds is largely unknown; therefore, changes in the circulating oxylipin and endocannabinoid metabolome in response to a challenge meal were investigated at six occasions in a subject who freely modified her usual diet. The dietary change, and especially the challenge meal itself, represented a modification of precursor fatty acid status, with expectedly subtle effects on bioactive lipid levels. To detect even the slightest alteration, highly sensitive ultra-performance liquid chromatography (UPLC) coupled to electrospray ionization (ESI) tandem mass spectrometry (MS/MS) methods for bioactive lipid profiling was employed. A previously validated UPLC-ESI-MS/MS method for profiling the endocannabinoid metabolome was used, while validation of an UPLC-ESI-MS/MS method for oxylipin analysis was performed with acceptable outcomes for a majority of the parameters according to the US Food and Drug Administration guidelines for linearity (0.9938 < R2 < 0.9996), limit of detection (0.0005-2.1 pg on column), limit of quantification (0.0005-4.2 pg on column), inter- and intraday accuracy (85-115%) and precision (< 5%), recovery (40-109%) and stability (40-105%). Forty-seven of fifty-two bioactive lipids were detected in plasma samples at fasting and in the postprandial state (0.5, 1, and 3 hours after the meal). Multivariate analysis showed a significant shift of bioactive lipid profiles in the postprandial state due to inclusion of dairy products in the diet, which was in line with univariate analysis revealing seven compounds (NAGly, 9-HODE, 13-oxo-ODE, 9(10)-EpOME, 12(13)-EpOME, 20-HETE, and 11,12-DHET) that were significantly different between background diets in the postprandial state (but not at fasting). The only change in baseline levels at fasting was displayed by TXB2. Furthermore, postprandial responsiveness was detected for seven compounds (POEA, SEA, 9(10)-DiHOME, 12(13)-DiHOME, 13-oxo-ODE, 9-HODE, and 13-HODE). Hence, the data confirm that the UPLC-ESI-MS/MS method performance was sufficient to detect i) a shift, in the current case most notably in the postprandial bioactive lipid metabolome, caused by changes in diet and ii) responsiveness to a challenge meal for a subset of the oxylipin and endocannabinoid metabolome. To summarize, we have shown proof-of-concept of our UPLC-ESI-MS/MS bioactive lipid protocols for the purpose of monitoring subtle shifts, and thereby useful to address lipid-mediated postprandial inflammation. PMID: 26186333 [PubMed - indexed for MEDLINE]

Related Articles Gene Transcriptional and Metabolic Profile Changes in Mimetic Aging Mice Induced by D-Galactose. PLoS One. 2015;10(7):e0132088 Authors: Zhou YY, Ji XF, Fu JP, Zhu XJ, Li RH, Mu CK, Wang CL, Song WW Abstract D-galactose injection has been shown to induce many changes in mice that represent accelerated aging. This mouse model has been widely used for pharmacological studies of anti-aging agents. The underlying mechanism of D-galactose induced aging remains unclear, however, it appears to relate to glucose and 1ipid metabolic disorders. Currently, there has yet to be a study that focuses on investigating gene expression changes in D-galactose aging mice. In this study, integrated analysis of gas chromatography/mass spectrometry-based metabonomics and gene expression profiles was used to investigate the changes in transcriptional and metabolic profiles in mimetic aging mice injected with D-galactose. Our findings demonstrated that 48 mRNAs were differentially expressed between control and D-galactose mice, and 51 potential biomarkers were identified at the metabolic level. The effects of D-galactose on aging could be attributed to glucose and 1ipid metabolic disorders, oxidative damage, accumulation of advanced glycation end products (AGEs), reduction in abnormal substance elimination, cell apoptosis, and insulin resistance. PMID: 26176541 [PubMed - indexed for MEDLINE]

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Related Articles EIder: A compound identification tool for gas chromatography mass spectrometry data. J Chromatogr A. 2016 Apr 23; Authors: Koo I, Kim S, Shi B, Lorkiewicz P, Song M, McClain C, Zhang X Abstract We report software entitled EIder (EI mass spectrum identifier) that provides users with eight literature reported spectrum matching algorithms for compound identification from gas chromatography mass spectrometry (GC-MS) data. EIder calculates retention index according to experimental conditions categorized by column class, column type and data type, where 9 empirical distribution functions of the absolute retention index deviation to its mean value were constructed using the National Institute of Standards and Technology (NIST) 2011 retention index database to improve the accuracy of compound identification. EIder filters compound candidates based on elementary composition and derivatization reagent, and automatically adds the molecular information of the native compound to each derivatized compound using a manually created database. When multiple samples are analyzed together, EIder performs cross-sample alignment and provides an option of using an average mass spectrum for compound identification. Furthermore, a suite of graphical user interfaces are implemented in EIder to allow users to both manually and automatically modify the identification results using experimental information at various analysis stages. Analysis of three types of GC-MS datasets indicates that the developed EIder software can improve the accuracy of compound identification. PMID: 27131963 [PubMed - as supplied by publisher]

Related Articles Effect of organochlorine pesticides exposure on the maize root metabolome assessed using high-resolution magic-angle spinning (1)H NMR spectroscopy. Environ Pollut. 2016 Apr 28;214:539-548 Authors: Blondel C, Khelalfa F, Reynaud S, Fauvelle F, Raveton M Abstract (1)H-HRMAS NMR-based metabolomics was used to better understand the toxic effects on maize root tips of organochlorine pesticides (OCPs), namely lindane (γHCH) and chlordecone (CLD). Maize seedlings were exposed to 2.5 μM γHCH (mimicking basic environmental contaminations) for 7 days and compared to 2.5 μM CLD and 25 μM γHCH for 7 days (mimicking hot spot contaminations). The (1)H-HRMAS NMR-based metabolomic profiles provided details of the changes in carbohydrates, amino acids, tricarboxylic acid (TCA) cycle intermediates and fatty acids with a significant separation between the control and OCP-exposed root tips. First of all, alterations in the balance between glycolysis/gluconeogenesis were observed with sucrose depletion and with dose-dependent fluctuations in glucose content. Secondly, observations indicated that OCPs might inactivate the TCA cycle, with sizeable succinate and fumarate depletion. Thirdly, disturbances in the amino acid composition (GABA, glutamine/glutamate, asparagine, isoleucine) reflected a new distribution of internal nitrogen compounds under OCP stress. Finally, OCP exposure caused an increase in fatty acid content, concomitant with a marked rise in oxidized fatty acids which could indicate failures in cell integrity and vitality. Moreover, the accumulation of asparagine and oxidized fatty acids with the induction of LOX3 transcription levels under OCP exposure highlighted an induction of protein and lipid catabolism. The overall data indicated that the effect of OCPs on primary metabolism could have broader physiological consequences on root development. Therefore, (1)H-HRMAS NMR metabolomics is a sensitive tool for understanding molecular disturbances under OCP exposure and can be used to perform a rapid assessment of phytotoxicity. PMID: 27131813 [PubMed - as supplied by publisher]

Related Articles Plant secondary metabolism linked glycosyltransferases: An update on expanding knowledge and scopes. Biotechnol Adv. 2016 Apr 27; Authors: Tiwari P, Sangwan RS, Sangwan NS Abstract The multigene family of enzymes known as glycosyltransferases or popularly known as GTs catalyze the addition of carbohydrate moiety to a variety of synthetic as well as natural compounds. Glycosylation of plant secondary metabolites is an emerging area of research in drug designing and development. The unsurpassing complexity and diversity among natural products arising due to glycosylation type of alterations including glycodiversification and glycorandomization are emerging as the promising approaches in pharmacological studies. While, some GTs with broad spectrum of substrate specificity are promising candidates for glycoengineering while others with stringent specificity pose limitations in accepting molecules and performing catalysis. With the rising trends in diseases and the efficacy/potential of natural products in their treatment, glycosylation of plant secondary metabolites constitutes a key mechanism in biogeneration of their glycoconjugates possessing medicinal properties. The present review highlights the role of glycosyltransferases in plant secondary metabolism with an overview of their identification strategies, catalytic mechanism and structural studies on plant GTs. Furthermore, the article discusses the biotechnological and biomedical application of GTs ranging from detoxification of xenobiotics and hormone homeostasis to the synthesis of glycoconjugates and crop engineering. The future directions in glycosyltransferase research should focus on the synthesis of bioactive glycoconjugates via metabolic engineering and manipulation of enzyme's active site leading to improved/desirable catalytic properties. The multiple advantages of glycosylation in plant secondary metabolomics highlight the increasing significance of the GTs, and in near future, the enzyme superfamily may serve as promising path for progress in expanding drug targets for pharmacophore discovery and development. PMID: 27131396 [PubMed - as supplied by publisher]

Related Articles Cyanate as an energy source for nitrifiers. Nature. 2015 Aug 6;524(7563):105-8 Authors: Palatinszky M, Herbold C, Jehmlich N, Pogoda M, Han P, von Bergen M, Lagkouvardos I, Karst SM, Galushko A, Koch H, Berry D, Daims H, Wagner M Abstract Ammonia- and nitrite-oxidizing microorganisms are collectively responsible for the aerobic oxidation of ammonia via nitrite to nitrate and have essential roles in the global biogeochemical nitrogen cycle. The physiology of nitrifiers has been intensively studied, and urea and ammonia are the only recognized energy sources that promote the aerobic growth of ammonia-oxidizing bacteria and archaea. Here we report the aerobic growth of a pure culture of the ammonia-oxidizing thaumarchaeote Nitrososphaera gargensis using cyanate as the sole source of energy and reductant; to our knowledge, the first organism known to do so. Cyanate, a potentially important source of reduced nitrogen in aquatic and terrestrial ecosystems, is converted to ammonium and carbon dioxide in Nitrososphaera gargensis by a cyanase enzyme that is induced upon addition of this compound. Within the cyanase gene family, this cyanase is a member of a distinct clade also containing cyanases of nitrite-oxidizing bacteria of the genus Nitrospira. We demonstrate by co-culture experiments that these nitrite oxidizers supply cyanase-lacking ammonia oxidizers with ammonium from cyanate, which is fully nitrified by this microbial consortium through reciprocal feeding. By screening a comprehensive set of more than 3,000 publically available metagenomes from environmental samples, we reveal that cyanase-encoding genes clustering with the cyanases of these nitrifiers are widespread in the environment. Our results demonstrate an unexpected metabolic versatility of nitrifying microorganisms, and suggest a previously unrecognized importance of cyanate in cycling of nitrogen compounds in the environment. PMID: 26222031 [PubMed - indexed for MEDLINE]

Foodomics as part of the host-microbiota-exposome interplay. J Proteomics. 2016 Apr 26; Authors: Putignani L, Dallapiccola B Abstract The functional complexity of human gut microbiota and its relationship with host physiology and environmental modulating factors, offers the opportunity to investigate (i) the host and microbiota role in organism-environment relationship; (ii) the individual functional diversity and response to environmental stimuli (exposome); (iii) the host' genome and microbiota metagenomes' modifications by diet-mediated epigenomic controls (nutriepigenomics); and (iv) the genotype-phenotype "trajectories" under physiological and disease constraints. Systems biology-based approaches aim at integrating biological data at cellular, tissue and organ organization levels, using computational modeling to interpret diseases' physiopathological mechanisms (i.e., onset and progression). Proteomics improves the existing gene models by profiling molecular phenotypes at protein abundance level, by analyzing post-translational modifications and protein-protein interactions and providing specific pathway information, hence contributing to functional molecular networks. Transcriptomics and metabolomics may determine host ad microbiota changes induced by food ingredients at molecular level, complementing functional genomics and proteomics data. Since foodomics is an -omic wide methodology may feed back all integrative data to foster the omics-based systems medicine field. Hence, coupled to ecological genomics of gut microbial communities, foodomics may highlight health benefits from nutrients, dissecting diet-induced gut microbiota eubiosis mechanisms and significantly contributing to understand and prevent complex disease phenotypes. BIOLOGICAL SIGNIFICANCE: Besides transcriptomics and proteomics there is a growing interest in applying metabolic profiling to food science for the development of functional foods. Indeed, one of the biggest challenges of modern nutrition is to propose a healthy diet to populations worldwide, intrinsically respecting the high inter-individual variability, driven by complex host/nutrients/microbiota/environment interactions. Therefore, metabolic profiling can assist at various levels for the development of functional foods, starting from screening for food composition to identification of new biomarkers to trace food intake. This current approach can support diet intervention strategies, epidemiological studies, and controlling of metabolic disorders worldwide spreading, hence ensuring healthy aging. With high-throughput molecular technologies driving foodomics, studying bidirectional interactions of host-microbial co-metabolism, innate immune development, dysfunctional nutrient absorption and processing, complex signaling pathways involved in nutritional metabolism, is now likely. In all cases, as microbiome pipeline efforts continue, it is possible that enhanced standardized protocols can be developed, which may lead to new testable biological and clinical hypotheses. This Review provides a comprehensive update on the current state-of-the-art of the integrated -omics route in food, microbiota and host co-metabolism studies, which may revolutionize the design of new dietary intervention strategies. PMID: 27130534 [PubMed - as supplied by publisher]

Multi-omic data integration and analysis using systems genomics approaches: methods and applications in animal production, health and welfare. Genet Sel Evol. 2016;48(1):38 Authors: Suravajhala P, Kogelman LJ, Kadarmideen HN Abstract In the past years, there has been a remarkable development of high-throughput omics (HTO) technologies such as genomics, epigenomics, transcriptomics, proteomics and metabolomics across all facets of biology. This has spearheaded the progress of the systems biology era, including applications on animal production and health traits. However, notwithstanding these new HTO technologies, there remains an emerging challenge in data analysis. On the one hand, different HTO technologies judged on their own merit are appropriate for the identification of disease-causing genes, biomarkers for prevention and drug targets for the treatment of diseases and for individualized genomic predictions of performance or disease risks. On the other hand, integration of multi-omic data and joint modelling and analyses are very powerful and accurate to understand the systems biology of healthy and sustainable production of animals. We present an overview of current and emerging HTO technologies each with a focus on their applications in animal and veterinary sciences before introducing an integrative systems genomics framework for analysing and integrating multi-omic data towards improved animal production, health and welfare. We conclude that there are big challenges in multi-omic data integration, modelling and systems-level analyses, particularly with the fast emerging HTO technologies. We highlight existing and emerging systems genomics approaches and discuss how they contribute to our understanding of the biology of complex traits or diseases and holistic improvement of production performance, disease resistance and welfare. PMID: 27130220 [PubMed - in process]

A metabolomics approach to identify and quantify the phytochemicals in watermelons by quantitative (1)HNMR. Talanta. 2016 Jun 1;153:268-77 Authors: Jayaprakasha GK, Patil BS Abstract Watermelon (Citrullus vulgaris) contains many health-promoting compounds, such as ascorbic acid, carotenoids, phenolic acids and amino acids including l-citrulline, arginine, and glutathione. Reported HPLC method for quantification of l-citrulline and sugars in watermelon involves, time-consuming sample preparation, post-column color development and detection with fluorescence and refractive index detectors. The present study describes development of a method to identify and quantify amino acids and sugars simultaneously from watermelon samples using quantitative proton NMR. Lyophilized watermelon samples (30-50mg) were extracted with deuterium oxide (D2O) by sonication and the centrifuged extract was directly used for quantification and identification with (1)HNMR. An external coaxial insert containing a 65µL of 0.012% 3-(trimethylsilyl) propionic-(2,2,3,3-d4) acid sodium salt (TSP-d4) in D2O was used as a quantitative reference. The levels of l-citrulline and sugars were measured in less than 6min. This rapid quantitation method was validated for specificity, linearity, accuracy, precision, reproducibility, and robustness. The limit of detection for l-citrulline was 38µg/mL and the limit of quantification was 71µg/mL; for sugars, the limits were 59-94µg/mL and 120µg/mL, respectively. This method can be used widely for confirmation and rapid quantitation of multiple compounds in large number of biological or breeding samples for routine analysis. PMID: 27130118 [PubMed - in process]

Simultaneous Determination of Thirteen Kinds of Amino Acid and Eight Kinds of Acylcarnitine in Human Serum by LC-MS/MS and Its Application to Measure the Serum Concentration of Lung Cancer Patients. Biomed Chromatogr. 2016 Apr 30; Authors: Ni J, Xu L, Li W, Wu L Abstract Easy-to-use early cancer detection methods based on metabolomics using serum samples have been developed recently. Among metabolites, amino acids and acylcarnitine are two of the most suitable candidates for diagnosing lung cancer. The purpose of the present study was to develop a novel, sensitive and specific liquid chromatography-tandem mass spectrometry (LC-MS/MS) method to simultaneously determine thirteen amino acids and eight acylcarnitines in lung cancer patients in serum. After derivatization, the 21 analytes were separated by a C18 column with gradient elution program in 14 minutes, obtaining recovery within 90.4%-113.8% and precision within 0.3%-14.8%. The method was successfully applied in concentration determination of lung cancer patients and healthy controls. The results showed that the serum concentration of lung cancer patients were significant from those of healthy controls. PMID: 27129889 [PubMed - as supplied by publisher]

Sexual Dimorphism in the Response of Mercurialis annua to Stress. Metabolites. 2016;6(2) Authors: Orlofsky EM, Kozhoridze G, Lyubenova L, Ostrozhenkova E, Winkler JB, Schröder P, Bacher A, Eisenreich W, Guy M, Golan-Goldhirsh A Abstract The research presented stemmed from the observations that female plants of the annual dioecious Mercurialis annua outlive male plants. This led to the hypothesis that female plants of M. annua would be more tolerant to stress than male plants. This hypothesis was addressed in a comprehensive way, by comparing morphological, biochemical and metabolomics changes in female and male plants during their development and under salinity. There were practically no differences between the genders in vegetative development and physiological parameters. However, under salinity conditions, female plants produced significantly more new reproductive nodes. Gender-linked differences in peroxidase (POD) and glutathione transferases (GSTs) were involved in anti-oxidation, detoxification and developmental processes in M. annua. ¹H NMR metabolite profiling of female and male M. annua plants showed that under salinity the activity of the TCA cycle increased. There was also an increase in betaine in both genders, which may be explainable by its osmo-compatible function under salinity. The concentration of ten metabolites changed in both genders, while 'Female-only-response' to salinity was detected for five metabolites. In conclusion, dimorphic responses of M. annua plant genders to stress may be attributed to female plants' capacity to survive and complete the reproductive life cycle. PMID: 27128954 [PubMed - as supplied by publisher]

Advantages and Pitfalls of Mass Spectrometry Based Metabolome Profiling in Systems Biology. Int J Mol Sci. 2016;17(5) Authors: Aretz I, Meierhofer D Abstract Mass spectrometry-based metabolome profiling became the method of choice in systems biology approaches and aims to enhance biological understanding of complex biological systems. Genomics, transcriptomics, and proteomics are well established technologies and are commonly used by many scientists. In comparison, metabolomics is an emerging field and has not reached such high-throughput, routine and coverage than other omics technologies. Nevertheless, substantial improvements were achieved during the last years. Integrated data derived from multi-omics approaches will provide a deeper understanding of entire biological systems. Metabolome profiling is mainly hampered by its diversity, variation of metabolite concentration by several orders of magnitude and biological data interpretation. Thus, multiple approaches are required to cover most of the metabolites. No software tool is capable of comprehensively translating all the data into a biologically meaningful context yet. In this review, we discuss the advantages of metabolome profiling and main obstacles limiting progress in systems biology. PMID: 27128910 [PubMed - as supplied by publisher]

Identification of a Binding Site for Unsaturated Fatty Acids in the Orphan Nuclear Receptor Nurr1. ACS Chem Biol. 2016 Apr 29; Authors: de Vera IM, Giri PK, Munoz-Tello P, Brust R, Fuhrmann J, Matta-Camacho E, Shang J, Campbell S, Wilson HD, Granados J, Gardner WJ, Creamer TP, Solt LA, Kojetin DJ Abstract Nurr1/NR4A2 is an orphan nuclear receptor, and currently there are no known natural ligands that bind Nurr1. A recent metabolomics study identified unsaturated fatty acids, including arachidonic acid and docosahexaenoic acid (DHA), that interact with the ligand-binding domain (LBD) of a related orphan receptor, Nur77/NR4A1. However, the binding location and whether these ligands bind other NR4A receptors were not defined. Here, we show that unsaturated fatty acids also interact with the Nurr1 LBD, and solution NMR spectroscopy reveals the binding epitope of DHA at its putative ligand-binding pocket. Biochemical assays reveal that DHA-bound Nurr1 interacts with high affinity with a peptide derived from PIASγ, a protein that interacts with Nurr1 in cellular extracts, and DHA also affects cellular Nurr1 transactivation. This work is the first structural report of a natural ligand binding to a canonical NR4A ligand-binding pocket and indicates a natural ligand can bind and affect Nurr1 function. PMID: 27128111 [PubMed - as supplied by publisher]

Mitochondrial Reactive Oxygen Species Mediate Lysophosphatidylcholine-Induced Endothelial Cell Activation. Arterioscler Thromb Vasc Biol. 2016 Apr 28; Authors: Li X, Fang P, Li Y, Kuo YM, Andrews AJ, Nanayakkara G, Johnson C, Fu H, Shan H, Du F, Hoffman NE, Yu D, Eguchi S, Madesh M, Koch WJ, Sun J, Jiang X, Wang H, Yang X Abstract OBJECTIVE: Hyperlipidemia-induced endothelial cell (EC) activation is considered as an initial event responsible for monocyte recruitment in atherogenesis. However, it remains poorly defined what is the mechanism underlying hyperlipidemia-induced EC activation. Here, we tested a novel hypothesis that mitochondrial reactive oxygen species (mtROS) serves as signaling mediators for EC activation in early atherosclerosis. APPROACH AND RESULTS: Metabolomics and transcriptomics analyses revealed that several lysophosphatidylcholine (LPC) species, such as 16:0, 18:0, and 18:1, and their processing enzymes, including Pla2g7 and Pla2g4c, were significantly induced in the aortas of apolipoprotein E knockout mice during early atherosclerosis. Using electron spin resonance and flow cytometry, we found that LPC 16:0, 18:0, and 18:1 induced mtROS in primary human aortic ECs, independently of the activities of nicotinamide adenine dinucleotide phosphate oxidase. Mechanistically, using confocal microscopy and Seahorse XF mitochondrial analyzer, we showed that LPC induced mtROS via unique calcium entry-mediated increase of proton leak and mitochondrial O2 reduction. In addition, we found that mtROS contributed to LPC-induced EC activation by regulating nuclear binding of activator protein-1 and inducing intercellular adhesion molecule-1 gene expression in vitro. Furthermore, we showed that mtROS inhibitor MitoTEMPO suppressed EC activation and aortic monocyte recruitment in apolipoprotein E knockout mice using intravital microscopy and flow cytometry methods. CONCLUSIONS: ATP synthesis-uncoupled, but proton leak-coupled, mtROS increase mediates LPC-induced EC activation during early atherosclerosis. These results indicate that mitochondrial antioxidants are promising therapies for vascular inflammation and cardiovascular diseases. PMID: 27127201 [PubMed - as supplied by publisher]