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Sample records for detecting tamiflu metabolite

  1. Mixture toxicity of the antiviral drug Tamiflu((R)) (oseltamivir ethylester) and its active metabolite oseltamivir acid.

    PubMed

    Escher, Beate I; Bramaz, Nadine; Lienert, Judit; Neuwoehner, Judith; Straub, Jürg Oliver

    2010-02-18

    Tamiflu (oseltamivir ethylester) is an antiviral agent for the treatment of influenza A and B. The pro-drug Tamiflu is converted in the human body to the pharmacologically active metabolite, oseltamivir acid, with a yield of 75%. Oseltamivir acid is indirectly photodegradable and slowly biodegradable in sewage works and sediment/water systems. A previous environmental risk assessment has concluded that there is no bioaccumulation potential of either of the compounds. However, little was known about the ecotoxicity of the metabolite. Ester hydrolysis typically reduces the hydrophobicity and thus the toxicity of a compound. In this case, a zwitterionic, but overall neutral species is formed from the charged parent compound. If the speciation and predicted partitioning into biological membranes is considered, the metabolite may have a relevant contribution to the overall toxicity. These theoretical considerations triggered a study to investigate the toxicity of oseltamivir acid (OA), alone and in binary mixtures with its parent compound oseltamivir ethylester (OE). OE and OA were found to be baseline toxicants in the bioluminescence inhibition test with Vibrio fischeri. Their mixture effect lay between predictions for concentration addition and independent action for the mixture ratio excreted in urine and nine additional mixture ratios of OE and OA. In contrast, OE was an order of magnitude more toxic than OA towards algae, with a more pronounced effect when the direct inhibition of photosystem II was used as toxicity endpoint opposed to the 24h growth rate endpoint. The binary mixtures in this assay yielded experimental mixture effects that agreed with predictions for independent action. This is consistent with the finding that OE exhibits slightly enhanced toxicity, while OA acts as baseline toxicant. Therefore, with respect to mixture classification, the two compounds can be considered as acting according to different modes of toxic action, although there are

  2. The Tamiflu fiasco and lessons learnt.

    PubMed

    Gupta, Yogendra Kumar; Meenu, Meenakshi; Mohan, Prafull

    2015-01-01

    Oseltamivir (Tamiflu), a neuraminidase inhibitor, was approved for seasonal flu by US Food and Drug Administration in 1999. A number of randomized controlled trials, systematic reviews, and meta-analysis emphasized a favorable efficacy and safety profile. Majority of them were funded by Roche, which also first marketed and promoted this drug. In 2005 and 2009, the looming fear of pandemic flu led to recommendation by prominent regulatory bodies such as World Health Organization (WHO), Centers for Disease Control and Prevention, European Medicines Agency and others for its use in treatment and prophylaxis of influenza, and it's stockpiling as a measure to tide over the crisis. Serious Adverse Events, especially neuropsychiatric events associated with Tamiflu started getting reported leading to a cascade of questions on clinical utility of this drug. A recent Cochrane review and related articles have questioned the risk-benefit ratio of the drug, besides raising doubts about the regulatory decision of approving it. The recommendations for stockpiling the said drug as given by various international organizations viz WHO have also been put to scrutiny. Although many reviewers have labeled the Tamiflu saga as a "costly mistake," the episode leaves us with some important lessons. This article takes a comprehensive relook on the subject, and we proceed to suggest some ways and means to avoid a similar situation in the future. PMID:25821304

  3. The Tamiflu fiasco and lessons learnt

    PubMed Central

    Gupta, Yogendra Kumar; Meenu, Meenakshi; Mohan, Prafull

    2015-01-01

    Oseltamivir (Tamiflu), a neuraminidase inhibitor, was approved for seasonal flu by US Food and Drug Administration in 1999. A number of randomized controlled trials, systematic reviews, and meta-analysis emphasized a favorable efficacy and safety profile. Majority of them were funded by Roche, which also first marketed and promoted this drug. In 2005 and 2009, the looming fear of pandemic flu led to recommendation by prominent regulatory bodies such as World Health Organization (WHO), Centers for Disease Control and Prevention, European Medicines Agency and others for its use in treatment and prophylaxis of influenza, and it's stockpiling as a measure to tide over the crisis. Serious Adverse Events, especially neuropsychiatric events associated with Tamiflu started getting reported leading to a cascade of questions on clinical utility of this drug. A recent Cochrane review and related articles have questioned the risk-benefit ratio of the drug, besides raising doubts about the regulatory decision of approving it. The recommendations for stockpiling the said drug as given by various international organizations viz WHO have also been put to scrutiny. Although many reviewers have labeled the Tamiflu saga as a “costly mistake,” the episode leaves us with some important lessons. This article takes a comprehensive relook on the subject, and we proceed to suggest some ways and means to avoid a similar situation in the future. PMID:25821304

  4. Occurrence and fate of oseltamivir carboxylate (Tamiflu) and amantadine in sewage treatment plants.

    PubMed

    Ghosh, Gopal Chandra; Nakada, Norihide; Yamashita, Naoyuki; Tanaka, Hiroaki

    2010-09-01

    We investigated the occurrence and fate of the two antiviral drugs oseltamivir carboxylate (OC)-the active metabolite of Tamiflu-and amantadine (AMT) at three sewage treatment plants (STPs) during the 2008-2009 and 2009-2010 influenza seasons in Japan. Both compounds were detected in all samples analyzed. The concentrations in raw influents at the STPs ranged from 140 to 460 ng L(-1) OC and from 184 to 538 ng L(-1) AMT. Primary treatment gave no substantial removal of the drugs (OC, 2-9%; AMT, 7-17%). Biological nutrient-removal-based secondary treatment (anoxic-oxic-anoxic-oxic and anaerobic-anoxic-oxic) removed 20-37% of OC, whereas extended-aeration-based conventional activated sludge treatment removed <20%. STPs using primary plus biological secondary treatment removed <50% of the drugs. The incorporation of tertiary treatment by ozonation removed >90%. Ozonation after secondary treatment in STPs will be necessary during an influenza pandemic to reduce the risks associated with the widespread use of antiviral drugs. PMID:20692015

  5. Detection and characterization of clostebol sulfate metabolites in Caucasian population.

    PubMed

    Balcells, Georgina; Pozo, Oscar J; Garrostas, Lorena; Esquivel, Argitxu; Matabosch, Xavier; Kotronoulas, Aristotelis; Joglar, Jesús; Ventura, Rosa

    2016-06-01

    Anabolic androgenic steroids (AAS) are synthetic testosterone derivatives which undergo extensive metabolism in man. Differences in the excretion of phase II metabolites are strongly associated with inter-individual and inter-ethnic variations. Sulfate metabolites have been described as long-term metabolites for some AAS. Clostebol is the 4-chloro derivative of testosterone and the aim of the present study was the evaluation of clostebol sulfate metabolites in Caucasian population by LC-MS/MS technology. Clostebol was orally administered to four healthy Caucasian male volunteers, and excretion study urines were collected up to 31 days. Several analytical strategies (neutral loss scan, precursor ion scan and selected reaction monitoring acquisitions modes) were applied to detect sulfate metabolites in post-administration samples. Sixteen sulfate metabolites were detected, five of them having detectability times above 10 days (S1a, S2a, S3b, S3g and S4b). Interestingly, metabolite S1a could be detected up to the last collected sample of all excretion studies and it was characterized by LC-MS/MS and GC-MS as 4ξ-chloro-5α-androst-3β-ol-17-one 3β-sulfate. Thus, monitoring of S1a improves the detection time of clostebol misuse with respect to the commonly monitored metabolites, excreted in the glucuronide fraction. Importantly, this new metabolite can be incorporated into recently developed LC-MS/MS screening methods base on the direct detection of phase II metabolites. PMID:27085012

  6. Development and validation of a rapid HPLC method for the determination of oseltamivir phosphate in Tamiflu and generic versions.

    PubMed

    Joseph-Charles, J; Geneste, C; Laborde-Kummer, E; Gheyouche, R; Boudis, H; Dubost, J-P

    2007-08-15

    Oseltamivir phosphate (OP) is an antiviral drug that is used in the treatment and prophylaxis of both influenza A and influenza B. It is effective against all known influenza viruses than can infect humans, including pandemic influenza viruses and may be the most appropriate antiviral option against avian influenza caused by H5N1 virus. Tamiflu, the registered trademark used under exclusive license by Roche laboratories with OP as active pharmaceutical ingredient, is considered the best treatment for the bird flu disease. A simple, selective, linear, accurate and precise HPLC method was developed and validated for rapid assay of OP aimed to the quality control of Tamiflu capsules and generic versions. Isocratic elution at a flow rate of 1.2 mL/min was employed on a Zorbax CN column (150 mm x 4.6mm; 5 microm) at ambient temperature. The mobile phase consisted of methanol and 0.04 M formic acid pH 3.0 (50:50, v/v). The UV detection wavelength was 226 nm and 20 microL of sample was injected. Sotalol hydrochloride was used as the internal standard (IS). The retention times for OP and IS were 3.40 and 2.25 min, respectively. The method was successfully applied to commercial pharmaceuticals, Tamiflu and generic versions. The proposed method could be applicable for routine analysis of OP and monitoring of the quality of marketed drugs as possibly counterfeit Tamiflu. PMID:17512688

  7. Efficacy of Influenza Vaccination and Tamiflu® Treatment – Comparative Studies with Eurasian Swine Influenza Viruses in Pigs

    PubMed Central

    Duerrwald, Ralf; Schlegel, Michael; Bauer, Katja; Vissiennon, Théophile; Wutzler, Peter; Schmidtke, Michaela

    2013-01-01

    Recent epidemiological developments demonstrated that gene segments of swine influenza A viruses can account for antigenic changes as well as reduced drug susceptibility of pandemic influenza A viruses. This raises questions about the efficacy of preventive measures against swine influenza A viruses. Here, the protective effect of vaccination was compared with that of prophylactic Tamiflu® treatment against two Eurasian swine influenza A viruses. 11-week-old pigs were infected by aerosol nebulisation with high doses of influenza virus A/swine/Potsdam/15/1981 (H1N1/1981, heterologous challenge to H1N1 vaccine strain) and A/swine/Bakum/1832/2000 (H1N2/2000, homologous challenge to H1N2 vaccine strain) in two independent trials. In each trial (i) 10 pigs were vaccinated twice with a trivalent vaccine (RESPIPORC® FLU3; 28 and 7 days before infection), (ii) another 10 pigs received 150 mg/day of Tamiflu® for 5 days starting 12 h before infection, and (iii) 12 virus-infected pigs were left unvaccinated and untreated and served as controls. Both viruses replicated efficiently in porcine respiratory organs causing influenza with fever, dyspnoea, and pneumonia. Tamiflu® treatment as well as vaccination prevented clinical signs and significantly reduced virus shedding. Whereas after homologous challenge with H1N2/2000 no infectious virus in lung and hardly any lung inflammation were detected, the virus titre was not and the lung pathology was only partially reduced in H1N1/1981, heterologous challenged pigs. Tamiflu® application did not affect these study parameters. In conclusion, all tested preventive measures provided protection against disease. Vaccination additionally prevented virus replication and histopathological changes in the lung of homologous challenged pigs. PMID:23630601

  8. Detection and quantification of boscalid and its metabolites in honeybees.

    PubMed

    Jabot, Claire; Daniele, Gaëlle; Giroud, Barbara; Tchamitchian, Sylvie; Belzunces, Luc P; Casabianca, Hervé; Vulliet, Emmanuelle

    2016-08-01

    Boscalid is a new-generation fungicide that has been detected in several bee matrices. The objective of this work was to characterize boscalid metabolites in honeybees based on in vivo experimentation, and next to verify the presence of theses metabolites into honeybees from colonies presenting troubles. A methodology based on complementary mass spectrometric tools, namely ultra-high performance liquid chromatography coupled to high-resolution mass spectrometry (UHPLC-QToF) or triple quadrupole mass spectrometry (UHPLC-QqQ) was implemented. Honeybees were sprayed with boscalid, at field rate (to induce the metabolization process) and the parent compound with its generated metabolites were then extracted using modified EU-QuEChERS method. The mass characteristics including exact mass, isotopic profile and mass fragments allowed assuming the structure of several metabolites. Some of them were unambiguously identified by comparison with synthesized analytical standards. The metabolites were resulted from hydroxylation and dechlorination of the parent compound as well as the substitution of a chlorine atom with an hydroxyl group. The metabolites were then quantified in bee samples collected from various beehives located in France. Boscalid and three of its metabolites were present in some samples at a level ranged between 0.2 and 36.3 ng/g. PMID:27179242

  9. Metabolite

    MedlinePlus

    A metabolite is any substance produced during metabolism (digestion or other bodily chemical processes). The term metabolite may also refer to the product that remains after a drug is broken down (metabolized) by the body.

  10. Optimizing Urine Processing Protocols for Protein and Metabolite Detection

    PubMed Central

    Siddiqui, Nazema Y; DuBois, Laura G; St John-Williams, Lisa; Will, Thompson J; Grenier, Carole; Burke, Emily; Fraser, Matthew O; Amundsen, Cindy L; Murphy, Susan K

    2016-01-01

    Background In urine, factors such as timing of voids, and duration at room temperature (RT) may affect the quality of recovered protein and metabolite data. Additives may aid with detection, but can add more complexity in sample collection or analysis. We aimed to identify the optimal urine processing protocol for clinically-obtained urine samples that allows for the highest protein and metabolite yields with minimal degradation. Methods Healthy women provided multiple urine samples during the same day. Women collected their first morning (1st AM) void and another “random void”. Random voids were aliquotted with: 1) no additive; 2) boric acid (BA); 3) protease inhibitor (PI); or 4) both BA + PI. Of these aliquots, some were immediately stored at 4°C, and some were left at RT for 4 hours. Proteins and individual metabolites were quantified, normalized to creatinine concentrations, and compared across processing conditions. Sample pools corresponding to each processing condition were analyzed using mass spectrometry to assess protein degradation. Results Ten Caucasian women between 35-65 years of age provided paired 1st morning and random voided urine samples. Normalized protein concentrations were slightly higher in 1st AM compared to random “spot” voids. The addition of BA did not significantly change proteins, while PI significantly improved normalized protein concentrations, regardless of whether samples were immediately cooled or left at RT for 4 hours. In pooled samples, there were minimal differences in protein degradation under the various conditions we tested. In metabolite analyses, there were significant differences in individual amino acids based on the timing of the void. Conclusions For comparative translational research using urine, information about void timing should be collected and standardized. For urine samples processed in the same day, BA does not appear to be necessary while the addition of PI enhances protein yields, regardless of 4

  11. In situ detection of anaerobic alkane metabolites in subsurface environments

    PubMed Central

    Agrawal, Akhil; Gieg, Lisa M.

    2013-01-01

    Alkanes comprise a substantial fraction of crude oil and refined fuels. As such, they are prevalent within deep subsurface fossil fuel deposits and in shallow subsurface environments such as aquifers that are contaminated with hydrocarbons. These environments are typically anaerobic, and host diverse microbial communities that can potentially use alkanes as substrates. Anaerobic alkane biodegradation has been reported to occur under nitrate-reducing, sulfate-reducing, and methanogenic conditions. Elucidating the pathways of anaerobic alkane metabolism has been of interest in order to understand how microbes can be used to remediate contaminated sites. Alkane activation primarily occurs by addition to fumarate, yielding alkylsuccinates, unique anaerobic metabolites that can be used to indicate in situ anaerobic alkane metabolism. These metabolites have been detected in hydrocarbon-contaminated shallow aquifers, offering strong evidence for intrinsic anaerobic bioremediation. Recently, studies have also revealed that alkylsuccinates are present in oil and coal seam production waters, indicating that anaerobic microbial communities can utilize alkanes in these deeper subsurface environments. In many crude oil reservoirs, the in situ anaerobic metabolism of hydrocarbons such as alkanes may be contributing to modern-day detrimental effects such as oilfield souring, or may lead to more beneficial technologies such as enhanced energy recovery from mature oilfields. In this review, we briefly describe the key metabolic pathways for anaerobic alkane (including n-alkanes, isoalkanes, and cyclic alkanes) metabolism and highlight several field reports wherein alkylsuccinates have provided evidence for anaerobic in situ alkane metabolism in shallow and deep subsurface environments. PMID:23761789

  12. Detection of enrofloxacin and its metabolite ciprofloxacin in equine hair.

    PubMed

    Dunnett, Mark; Richardson, Dean W; Lees, Peter

    2004-10-01

    Hair analysis to detect drug administration has not been studied extensively in horses. This study aimed to (a) develop an analytical method for enrofloxacin and its metabolite ciprofloxacin in mane and tail hair, (b) relate measured values to doses, routes of administration, hair colour, and (c) demonstrate long-term detectability. Samples were extracted in trifluoroacetic acid at 70 degrees C. Extracts were cleaned-up by solid-phase extraction and analysed by high-performance liquid chromatography with UV-diode array detection. Analyte recoveries were > 87%. Horses were sampled after therapeutic enrofloxacin administration either orally at 7.5 mg/kg daily for 3-13 days or twice daily for 10-14 days (Group 1, n=7) or intravenously at 5.0 mg/kg daily for 12 and 15 days (Group 2, n=2). Enrofloxacin and ciprofloxacin were detected at concentrations up to 452 and 19 ng/mg, respectively, up to 10 months post-treatment. In vitro, enrofloxacin and ciprofloxacin were extensively bound to melanin (> 96%) and in vivo, their uptake was 40-fold greater in black than white hair. Enrofloxacin and ciprofloxacin concentrations correlated to enrofloxacin dose (r2=0.777 and r2=0.769). Enrofloxacin:ciprofloxacin ratios were 21:1 and 13:1 following intravenous and oral administration, respectively. Longitudinal analyte distributions correlated to treatment-sampling interval. PMID:15196904

  13. Acute intoxication with aniline: detection of acetaminophen as aniline metabolite.

    PubMed

    Iwersen-Bergmann, S; Schmoldt, A

    2000-01-01

    A 47-year-old woman unwittingly ingested an unknown substance together with her breakfast coffee. She suffered effects such as strong headache, generalized cyanosis, and a burning sensation of the lips and collapsed some minutes later. After admission into hospital a methemoglobin level of 35% was determined in the blood. Treatment by administration of tolonium chloride (toluidine blue) resulted in complete recovery of the patient. The toxic agent was identified as aniline by GC with mass selective detection after organic solvent extraction and 11 h after ingestion the plasma aniline level was 0.13 mg/l. Acetanilide (0.79 mg/ml) and acetaminophen (2.3 mg/ml) were identified in plasma as metabolites of aniline. It was assumed that a high metabolic capacity for acetylation protected the victim from more severe reactions. Her husband confessed later that he had tried to poison her. PMID:10876991

  14. Simultaneous determination of tricarboxylic acid cycle metabolites by high-performance liquid chromatography with ultraviolet detection.

    PubMed

    Shurubor, Yevgeniya I; Cooper, Arthur J L; Isakova, Elena P; Deryabina, Yulia I; Beal, M Flint; Krasnikov, Boris F

    2016-06-15

    Here we describe a simple high-performance liquid chromatography (HPLC) procedure for the simultaneous detection and quantitation in standard solutions of 13 important metabolites of cellular energy metabolism, including 9 tricarboxylic acid (TCA) cycle components and 4 additional metabolites. The metabolites are detected by their absorbance at 210 nm. The procedure does not require prior derivatization, and an analysis can be carried out at ambient temperature within 15 min. The significance of the current work is that the current HPLC procedure should motivate the development of simplified TCA cycle enzyme assays, isotopomer analysis, and determination of selected TCA metabolite levels in plasma/tissues. PMID:27001310

  15. Detection of Nitrobenzodiazepines and Their 7-Amino Metabolites in Oral Fluid.

    PubMed

    Vindenes, Vigdis; Strand, Dag Helge; Koksæter, Paul; Gjerde, Hallvard

    2016-05-01

    Clonazepam, nitrazepam and flunitrazepam are frequently used benzodiazepines, both as prescribed medication and as drugs of abuse. Little is, however, known about how these drugs are excreted in oral fluid. It has been claimed that the parent drugs are more likely to be detected in oral fluid than the 7-amino metabolites. The aim of this study was to investigate whether the parent drugs or the 7-amino metabolites of the nitrobenzodiazepines were most frequently detected in authentic oral fluid samples. Oral fluid samples were collected from patients undergoing opioid maintenance treatment. Cases where clonazepam, nitrazepam, flunitrazepam and/or their metabolites were detected were included. The samples were collected using the Intercept(®)Oral Specimen Collection Device. A cutoff concentration of 1 nM (∼0.3 ng/mL) in oral fluid-buffer mixture was applied for all the substances. A total of 1,001 oral fluid samples were positive for clonazepam and/or 7-aminoclonazepam; both substances were detected in 707 samples, only the parent drug in 64 cases and only the metabolite in 230 cases. For nitrazepam, both substances were detected in 139 samples; only the parent drug in 16 cases and only the metabolite in 56 cases. Flunitrazepam only was not detected in any sample; both substances were detected in one of these cases, and only the metabolite in three cases. This study revealed that 7-amino metabolites were more likely to be detected in oral fluid than the parent drugs. PMID:27013620

  16. Detection of Volatile Metabolites of Garlic in Human Breast Milk

    PubMed Central

    Scheffler, Laura; Sauermann, Yvonne; Zeh, Gina; Hauf, Katharina; Heinlein, Anja; Sharapa, Constanze; Buettner, Andrea

    2016-01-01

    The odor of human breast milk after ingestion of raw garlic at food-relevant concentrations by breastfeeding mothers was investigated for the first time chemo-analytically using gas chromatography−mass spectrometry/olfactometry (GC-MS/O), as well as sensorially using a trained human sensory panel. Sensory evaluation revealed a clear garlic/cabbage-like odor that appeared in breast milk about 2.5 h after consumption of garlic. GC-MS/O analyses confirmed the occurrence of garlic-derived metabolites in breast milk, namely allyl methyl sulfide (AMS), allyl methyl sulfoxide (AMSO) and allyl methyl sulfone (AMSO2). Of these, only AMS had a garlic-like odor whereas the other two metabolites were odorless. This demonstrates that the odor change in human milk is not related to a direct transfer of garlic odorants, as is currently believed, but rather derives from a single metabolite. The formation of these metabolites is not fully understood, but AMSO and AMSO2 are most likely formed by the oxidation of AMS in the human body. The excretion rates of these metabolites into breast milk were strongly time-dependent with large inter-individual differences. PMID:27275838

  17. Detection of Volatile Metabolites of Garlic in Human Breast Milk.

    PubMed

    Scheffler, Laura; Sauermann, Yvonne; Zeh, Gina; Hauf, Katharina; Heinlein, Anja; Sharapa, Constanze; Buettner, Andrea

    2016-01-01

    The odor of human breast milk after ingestion of raw garlic at food-relevant concentrations by breastfeeding mothers was investigated for the first time chemo-analytically using gas chromatography-mass spectrometry/olfactometry (GC-MS/O), as well as sensorially using a trained human sensory panel. Sensory evaluation revealed a clear garlic/cabbage-like odor that appeared in breast milk about 2.5 h after consumption of garlic. GC-MS/O analyses confirmed the occurrence of garlic-derived metabolites in breast milk, namely allyl methyl sulfide (AMS), allyl methyl sulfoxide (AMSO) and allyl methyl sulfone (AMSO₂). Of these, only AMS had a garlic-like odor whereas the other two metabolites were odorless. This demonstrates that the odor change in human milk is not related to a direct transfer of garlic odorants, as is currently believed, but rather derives from a single metabolite. The formation of these metabolites is not fully understood, but AMSO and AMSO₂ are most likely formed by the oxidation of AMS in the human body. The excretion rates of these metabolites into breast milk were strongly time-dependent with large inter-individual differences. PMID:27275838

  18. [Comparison of four immunoassay screening devices for detection of benzodiazepine and its metabolites in urine: mainly detection of etizolam, thienodiazepine and its metabolites].

    PubMed

    Namera, Akira; Makita, Ryosuke; Nagao, Masataka

    2011-03-01

    The immunoassay screening of benzodiazepines in urine is one of the most important methods of drug analysis in clinical and forensic laboratories. We experienced an unusual case of poisoning wherein the result of Triage DOA immunoassay screening was negative, although Depas (etizolam) was detected in the blood of the victim who had been suspected to prescribe Depas by gas chromatography-mass spectrometry. Depas has been widely used for the treatment of anxiety in Japan. Three immunoassay screening devices (AccuSign BZO, Monitect-3, and Fastect II) were evaluated for their specificity for etizolam, its 2 major metabolites M-III and M-VI, and other metabolites of benzodiazepines in urine. With AccuSign BZO, etizolam, M-III, and M-VI could be detected at concentrations of 1,000 ng/mL in urine; however, they could not be detected even at concentrations of 25,000 ng/mL with the other kits. In the case of etizolam poisoning, the result of AccuSign BZO was positive; however, Triage DOA, which is mainly used for the detection of drugs in urine at intensive care units (ICUs) or forensic laboratories, showed negative result for benzodiazepines. The concentrations of etizolam and its metabolites in urine were measured by the established high-performance liquid chromatographic method. The concentrations of M-III and M-V were 700 and 1,600 ng/mL, respectively. AccuSign BZO demonstrated higher specificity-than the other screening kits for the detection of etizolam and its metabolites in urine. Therefore, the types of drugs detected would be increased by combining Triage DOA with AccuSign BZO in ICUs or forensic laboratories. PMID:21485120

  19. Detection of driver metabolites in the human liver metabolic network using structural controllability analysis

    PubMed Central

    2014-01-01

    Background Abnormal states in human liver metabolism are major causes of human liver diseases ranging from hepatitis to hepatic tumor. The accumulation in relevant data makes it feasible to derive a large-scale human liver metabolic network (HLMN) and to discover important biological principles or drug-targets based on network analysis. Some studies have shown that interesting biological phenomenon and drug-targets could be discovered by applying structural controllability analysis (which is a newly prevailed concept in networks) to biological networks. The exploration on the connections between structural controllability theory and the HLMN could be used to uncover valuable information on the human liver metabolism from a fresh perspective. Results We applied structural controllability analysis to the HLMN and detected driver metabolites. The driver metabolites tend to have strong ability to influence the states of other metabolites and weak susceptibility to be influenced by the states of others. In addition, the metabolites were classified into three classes: critical, high-frequency and low-frequency driver metabolites. Among the identified 36 critical driver metabolites, 27 metabolites were found to be essential; the high-frequency driver metabolites tend to participate in different metabolic pathways, which are important in regulating the whole metabolic systems. Moreover, we explored some other possible connections between the structural controllability theory and the HLMN, and find that transport reactions and the environment play important roles in the human liver metabolism. Conclusion There are interesting connections between the structural controllability theory and the human liver metabolism: driver metabolites have essential biological functions; the crucial role of extracellular metabolites and transport reactions in controlling the HLMN highlights the importance of the environment in the health of human liver metabolism. PMID:24885538

  20. Detection and characterization of betamethasone metabolites in human urine by LC-MS/MS.

    PubMed

    Matabosch, Xavier; Pozo, Oscar J; Monfort, Núria; Pérez-Mañá, Clara; Farré, Magí; Segura, Jordi; Ventura, Rosa

    2015-08-01

    Glucocorticosteroids are prohibited in sports when administered by systemic routes and allowed using other administrations for therapeutic reasons. Therefore, markers to distinguish between routes of administration through the analysis of urine samples are needed in anti-doping control. As a first step to achieve that goal, the metabolism of betamethasone (BET) was investigated in the present work. Urine samples obtained after BET intramuscular injection were hydrolyzed with β-glucuronidase and subjected to liquid-liquid extraction with ethyl acetate in alkaline conditions. The extracts were analyzed by liquid chromatography coupled to tandem mass spectrometry. Common open screening methods for fluorine containing corticosteroids (precursor ion scan method of m/z 121, 147, 171, and neutral loss (NL) scan methods of 20 and 38 Da in positive ionization, and 46 and 76 Da in negative ionization) were applied to detect BET metabolites. Moreover, an NL method was applied to detect A-ring reduced metabolites of BET, which are ionized as [M+NH4 ](+) (NL of 55, 73, and 91 Da, corresponding to the consecutive losses of NH3 , HF and one, two and three water molecules, respectively). BET and 24 metabolites were detected. Six metabolites were identified by comparison with standards, and for ten, feasible structures were proposed based on mass spectrometric data. Eleven of the characterized metabolites had not been previously reported. Metabolites resulting from 11-oxidation, 6-hydroxylation, C20 or 4-ene-3-one reduction and combination of some of them were detected. Moreover one metabolite resulting from cleavage of the side chain with subsequent oxidation of carbon at C17 was also detected. PMID:25545348

  1. Detection, synthesis and characterization of metabolites of steroid hormones conjugated with cysteine.

    PubMed

    Fabregat, Andreu; Kotronoulas, Aristotelis; Marcos, Josep; Joglar, Jesús; Alfonso, Ignacio; Segura, Jordi; Ventura, Rosa; Pozo, Oscar J

    2013-03-01

    The occurrence of several polyunsaturated testosterone related compounds (including 4,6-androstadien-3,17-dione and 4,6-androstadien-17β-ol-3-one) in urine after alkaline treatment of the sample has been recently reported. Although several experiments seem to indicate that they are testosterone metabolites, their origin is still unknown. In this study, it is demonstrated that these metabolites are produced from the degradation of cysteine conjugates. Several testosterone metabolites conjugated with cysteine have been synthesized and characterized by NMR techniques. Their detection in human urine has been performed by LC-MS/MS. The acquisition of several transitions in the SRM mode and the comparison between ion ratios and retention times allowed for the unequivocal confirmation of the presence of cysteine conjugates in urine. The analysis of urine samples collected after testosterone administration confirmed that synthesized cysteine conjugates are testosterone metabolites. The fact that these conjugates result in polyunsaturated compounds in urine after alkaline treatment was demonstrated by fraction collection and alkaline treatment of each fraction. Besides, the presence of these metabolites was also confirmed in human plasma. The formation of these metabolites implies an unreported metabolic biotransformation: 6,7-dehydrogenation as phase I metabolism followed by conjugation with glutathione and subsequent transformation to cysteine conjugates. Finally, the existence of similar metabolites for cortisol and progesterone was also confirmed by LC-MS/MS indicating that the presented metabolic pathway is not exclusively active in androgens, but common to progestagens and glucocorticoids. PMID:23261958

  2. Lateral flow biosensor for multiplex detection of nitrofuran metabolites based on functionalized magnetic beads.

    PubMed

    Lu, Xuewen; Liang, Xiaoling; Dong, Jianghong; Fang, Zhiyuan; Zeng, Lingwen

    2016-09-01

    The use of potential mutagenic nitrofuran antibiotic in food animal production has been banned world-wide. Common methods for nitrofuran detection involve complex extraction procedures. In the present study, magnetic beads functionalized with antibody against nitrofuran derivative were used as both the extraction and color developing media in lateral flow biosensor. Derivatization reagent carboxybenzaldehyde is firstly modified with ractopamine. After reaction with nitrofuran metabolites, the resultant molecule has two functional groups: the metabolite moiety and the ractopamine moiety. Metabolite moiety is captured by the antibody that is coated on magnetic beads. This duplex is then loaded onto biosensor and ractopamine moiety is further captured by the antibody immobilized on the test zone of nitrocellulose membrane. Without tedious organic reagent-based extraction procedure, this biosensor was capable of visually detecting four metabolites simultaneously with a detection limit of 0.1 μg/L. No cross-reactivity was observed in the presence of 50 μg/L interferential components. Graphical abstract Derivatization of nitrofuran metabolites (AHD, AOZ, SEM, or AMOZ) and LFA detection of the derivative products. PMID:27438720

  3. Nanomaterial based electrochemical sensors for in vitro detection of small molecule metabolites.

    PubMed

    Xiao, Fei; Wang, Lu; Duan, Hongwei

    2016-01-01

    Small molecule metabolites secreted by pathological processes can act as molecular biomarkers for clinical diagnosis. In vitro detection of the metabolites such as glucose and reactive oxygen species is of great significance for precise screening, monitoring and prognosis of metabolic disorders and relevant diseases such as cancer, and has been under intense research and development in clinical chemistry and molecular diagnostics. In this review, we summarize recent developments in nanomaterial based electrochemical (bio)sensors for in vitro detection of glucose and reactive oxygen species and the progress in utilizing lightweight and flexible electrodes and micro/nanoscale electrodes for flexible and miniaturized sensors. PMID:26845060

  4. Development of A Concise Synthesis of (−)-Oseltamivir (Tamiflu®)

    PubMed Central

    Trost, Barry M.; Zhang, Ting

    2011-01-01

    We report a full account of our work towards the development of an eight-step synthesis of anti-influenza drug (−)-oseltamivir (Tamiflu®) from commercially available starting material. The final synthetic route proceeds with an overall yield of 30 %. Key transformations include a novel palladium-catalyzed asymmetric allylic alkylation reaction (Pd-AAA) as well as a rhodium-catalyzed chemo-, regio-, and stereoselective aziridination reaction. PMID:21365707

  5. Detection of hepatotoxicity potential with metabolite profiling (metabolomics) of rat plasma.

    PubMed

    Mattes, W; Davis, K; Fabian, E; Greenhaw, J; Herold, M; Looser, R; Mellert, W; Groeters, S; Marxfeld, H; Moeller, N; Montoya-Parra, G; Prokoudine, A; van Ravenzwaay, B; Strauss, V; Walk, T; Kamp, H

    2014-11-01

    While conventional parameters used to detect hepatotoxicity in drug safety assessment studies are generally informative, the need remains for parameters that can detect the potential for hepatotoxicity at lower doses and/or at earlier time points. Previous work has shown that metabolite profiling (metabonomics/metabolomics) can detect signals of potential hepatotoxicity in rats treated with doxorubicin at doses that do not elicit hepatotoxicity as monitored with conventional parameters. The current study extended this observation to the question of whether such signals could be detected in rats treated with compounds that can elicit hepatotoxicity in humans (i.e., drug-induced liver injury, DILI) but have not been reported to do so in rats. Nine compounds were selected on the basis of their known DILI potential, with six other compounds chosen as negative for DILI potential. A database of rat plasma metabolite profiles, MetaMap(®)Tox (developed by metanomics GmbH and BASF SE) was used for both metabolite profiles and mode of action (MoA) metabolite signatures for a number of known toxicities. Eight of the nine compounds with DILI potential elicited metabolite profiles that matched with MoA patterns of various rat liver toxicities, including cholestasis, oxidative stress, acetaminophen-type toxicity and peroxisome proliferation. By contrast, only one of the six non-DILI compounds showed a weak match with rat liver toxicity. These results suggest that metabolite profiling may indeed have promise to detect signals of hepatotoxicity in rats treated with compounds having DILI potential. PMID:25086301

  6. Metabolism of boldione in humans by mass spectrometric techniques: detection of pseudoendogenous metabolites.

    PubMed

    de la Torre, Xavier; Curcio, Davide; Colamonici, Cristiana; Molaioni, Francesco; Botrè, Francesco

    2013-01-01

    Boldione is an anabolic androgenic steroid (AAS) related to boldenone, androstenedione, and testosterone bearing two double bonds in C1 and C4 positions. Boldione is rapidly transformed to the well-known AAS boldenone, being both compounds included in the list of prohibited substances and methods published yearly by the World Anti-Doping Agency (WADA). After the administration of boldione to a male volunteer, the already described urinary metabolites of boldenone produced after reduction in C4, oxydoreduction in C3 and C17, and hydroxylation have been detected. In addition, minor new metabolites have been detected and their structure postulated after mass spectrometric analyses. Finally, the reduction of the double bound in C1 produces metabolites identical to the endogenously produced ones. A method based on gas chromatography coupled to isotope ratio mass spectrometry (GC/C/IRMS) after a urine sample purification by high performance liquid chromatography (HPLC) permitted to confirm the main synthetic like boldione/boldenone metabolite (17β-hydroxy-5β-androst-1-en-3-one) and boldenone at trace levels (< 5 ng/mL) and then to establish its synthetic or endogenous origin, and to determine the exogenous origin of metabolites with the same chemical structure of the endogenous ones. The detection of pseudoendogenous androgens of synthetic origin partially overlapped boldenone and its main metabolite detection, being an additional proof of synthetic steroids misuse. By the use of IRMS, the correct evaluation of the modifications of the steroid profile after the administration of synthetic AAS that could be converted into endogenous like ones is possible. PMID:24259377

  7. Chemoselective detection and discrimination of carbonyl-containing compounds in metabolite mixtures by 1H-detected 15N NMR

    PubMed Central

    Lane, Andrew N.; Arumugam, Sengodagounder; Lorkiewicz, Pawel K.; Higashi, Richard M.; Laulhé, Sébastien; Nantz, Michael H.; Moseley, Hunter N.B.; Fan, Teresa W.-M.

    2015-01-01

    NMR spectra of mixtures of metabolites extracted from cells or tissues are extremely complex, reflecting the large number of compounds that are present over a wide range of concentrations. Although multidimensional NMR can greatly improve resolution as well as improve reliability of compound assignments, lower abundance metabolites often remain hidden. We have developed a carbonyl selective aminooxy probe that specifically reacts with free keto and aldehyde functions, but not carboxylates. By incorporating 15N in the aminooxy functional group, 15N-edited NMR was used to select exclusively those metabolites that contain a free carbonyl function while all other metabolites are rejected. Here we demonstrate that the chemical shifts of the aminooxy adducts of ketones and aldehydes are very different, which can be used to discriminate between aldoses and ketoses for example. Utilizing the 2 or 3 bond 15N-1H couplings, the 15N-edited NMR analysis was optimized first with authentic standards and then applied to an extract of the lung adenocarcinoma cell line A549. More than 30 carbonyl containing compounds at NMR detectable levels, 6 of which we have assigned by reference to our database. As the aminooxy probe contains a permanently charged quaternary ammonium group, the adducts are also optimized for detection by mass spectrometry. Thus, this sample preparation technique provides a better link between the two structural determination tools, thereby paving the way to faster and more reliable identification of both known and unknown metabolites directly in crude biological extracts. PMID:25616249

  8. [Detection of fungal metabolites showing toxic activity through Artemia salina bioassay].

    PubMed

    González, Ana María; Presa, Maximiliano; Latorre, María Gabriela; Lurá, María Cristina

    2007-03-01

    The aim of this study was to detect toxic metabolites from fungi contaminating food and medicinal herbs by applying the toxicity assay to Artemia salina. According to toxicity percentages, the extracts were classified as nontoxic (NT), slightly toxic (ST), toxic (T) and highly toxic (HT). Those classified as T and HT were assayed for mycotoxins. Only 6 out of 71 strains were found to be T (8.5%) for A. salina. Penicillium brevicompactum Dierckx, isolated from sausages, was found to be HT, mainly due to the presence of ochratoxin A and two other unidentified metabolites. PMID:17592895

  9. Graphene nano-ink biosensor arrays on a microfluidic paper for multiplexed detection of metabolites.

    PubMed

    Labroo, Pratima; Cui, Yue

    2014-02-27

    The development of a miniaturized and low-cost platform for the highly sensitive, selective and rapid detection of multiplexed metabolites is of great interest for healthcare, pharmaceuticals, food science, and environmental monitoring. Graphene is a delicate single-layer, two-dimensional network of carbon atoms with extraordinary electrical sensing capability. Microfluidic paper with printing technique is a low cost matrix. Here, we demonstrated the development of graphene-ink based biosensor arrays on a microfluidic paper for the multiplexed detection of different metabolites, such as glucose, lactate, xanthine and cholesterol. Our results show that the graphene biosensor arrays can detect multiple metabolites on a microfluidic paper sensitively, rapidly and simultaneously. The device exhibits a fast measuring time of less than 2 min, a low detection limit of 0.3 μM, and a dynamic detection range of 0.3-15 μM. The process is simple and inexpensive to operate and requires a low consumption of sample volume. We anticipate that these results could open exciting opportunities for a variety of applications. PMID:24528665

  10. CEST-MRI detects metabolite levels altered by breast cancer cell aggressiveness and chemotherapy response.

    PubMed

    Chan, Kannie W Y; Jiang, Lu; Cheng, Menglin; Wijnen, Jannie P; Liu, Guanshu; Huang, Peng; van Zijl, Peter C M; McMahon, Michael T; Glunde, Kristine

    2016-06-01

    Chemical exchange saturation transfer (CEST) is an MRI contrast mechanism that detects the exchange of protons from distinct hydroxyl, amine, and amide groups to tissue water through the transfer of signal loss, with repeated exchange enhancing their effective signal. We applied CEST to detect systematically 15 common cellular metabolites in a panel of differentially aggressive human breast cancer cell lines. The highest CEST contrast was generated by creatine, myo-inositol, glutamate, and glycerophosphocholine, whose cellular concentrations decreased with increasing breast cancer aggressiveness. These decreased metabolite concentrations resulted in turn in a decreased CEST profile with increasing breast cancer aggressiveness in water-soluble extracts of breast cell lines. Treatment of both breast cancer cell lines with the chemotherapy drug doxorubicin resulted in increased metabolic CEST profiles, which correlated with significant increases in creatine, phosphocreatine, and glycerophosphocholine. CEST can detect breast cancer aggressiveness and response to chemotherapy in water-soluble extracts of breast cell lines. The presented results help shed light on possible contributions from CEST-active metabolites to the CEST contrast produced by breast cancers. The metabolic CEST profile may improve detection sensitivity over conventional MRS, and may have the potential to assess breast cancer aggressiveness and response to chemotherapy non-invasively using MRI if specialized metabolic CEST profile detection can be realized in vivo. Copyright © 2016 John Wiley & Sons, Ltd. PMID:27100284

  11. Separation and detection of amino acid metabolites of Escherichia coli in microbial fuel cell with CE.

    PubMed

    Wang, Wei; Ma, Lihong; Lin, Ping; Xu, Kaixuan

    2016-07-01

    In this work, CE-LIF was employed to investigate the amino acid metabolites produced by Escherichia coli (E. coli) in microbial fuel cell (MFC). Two peptides, l-carnosine and l-alanyl-glycine, together with six amino acids, cystine, alanine, lysine, methionine, tyrosine, arginine were separated and detected in advance by a CE-LIF system coupled with a homemade spontaneous injection device. The injection device was devised to alleviate the effect of electrical discrimination for analytes during sample injection. All analytes could be completely separated within 8 min with detection limits of 20-300 nmol/L. Then this method was applied to analyze the substrate solution containing amino acid metabolites produced by E. coli. l-carnosine, l-alanyl-glycine, and cystine were used as the carbon, nitrogen, and sulfur source for the E. coli culture in the MFC to investigate the amino acid metabolites during metabolism. Two MFCs were used to compare the activity of metabolism of the bacteria. In the sample collected at the running time 200 h of MFC, the amino acid methionine was discovered as the metabolite with the concentrations 23.3 μg/L. PMID:27121957

  12. Detection and identification of plasma progesterone metabolites in the female Florida manatee (Trichechus manatus latirostris) using GC/MS/MS.

    PubMed

    Tripp, K M; Dubois, M; Delahaut, P; Verstegen, J P

    2009-08-01

    Florida manatees (Trichechus manatus latirostris) have relatively low peripheral concentrations of progesterone (P4). The objective of this study was to determine if these relatively low P4 concentrations are associated with a high ratio of progestin metabolites and to document metabolite concentrations from individual blood samples obtained from manatees during diestrus or pregnancy. Metabolites known to exist in elephants-terrestrial manatee relatives-were targeted. These included 5alpha-reduced progestins (5alpha-pregnane-3,20-dione [5alpha-DHP] and 3alpha-hydroxy-5alpha-pregnan-20-one [5alpha-P3-OH]) and 17alpha-hydroxyprogesterone (17alpha-OHP), which occurs in Asian elephants. An additional, inactive metabolite, 20alpha-hydroxyprogesterone (20alpha-OHP), indicative of P4 overproduction, was also targeted. Progesterone itself was the predominant progestin detected in pregnant and nonpregnant manatee plasma (n = 10) using gas chromatography-mass spectrometry with tandem quadrupole detectors (GC/MS/MS). Progesterone concentrations in pregnant females varied from early (moderate to high) through mid and late (low) pregnancy. Progesterone concentrations ranged from low to high in nonpregnant, nonlactating females. The most commonly detected metabolite was 5alpha-P3-OH (n = 7), which occurred in pregnant (lower limit of detection [LLOD] to high) and nonpregnant (trace to high) females. The 5alpha-DHP metabolite was also detected in pregnant (LLOD to moderate) and nonpregnant (low) females. The 17alpha-OHP metabolite was not detected in any tested female. The 20alpha-OHP metabolite was detected in one nonpregnant, nonlactating, captive female (LLOD). Metabolites were most prevalent during early pregnancy, concurrent with maximum P4 concentrations. Based on their concentrations in peripheral circulation, we inferred that these metabolites may have, opposite to elephants, a limited physiologic role during luteal, pregnant, and nonpregnant phases in the manatee. PMID

  13. SECONDARY METABOLITE PRODUCTION BY THE FUNGAL PATHOGEN EUTYPA LATA: ANALYSIS OF EXTRACTS FROM GRAPEVINE CULTURES AND DETECTION OF THOSE METABOLITES IN PLANTA

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Eutypa dieback of grapevines is caused by the fungal pathogen Eutypa lata and reduces vineyard longevity worldwide. Early detection could reduce losses due to this disease, so our aim was to identify acetylenic phenol metabolites of E. lata that could prove suitable as chemical markers in an early d...

  14. Determining zolpidem compliance: urinary metabolite detection and prevalence in chronic pain patients.

    PubMed

    Schwope, David M; DePriest, Anne; Black, David L; Caplan, Yale H; Cone, Edward J; Heltsley, Rebecca

    2014-10-01

    Zolpidem (Ambien(®)) is the most prescribed insomnia treatment in the USA; however, little is known about zolpidem metabolite excretion in chronic pain patients. As zolpidem is extensively metabolized in vivo to zolpidem 4-phenyl carboxylic acid (ZCA), metabolite detection may provide improved accuracy for compliance determinations, thereby improving clinical decisions. Zolpidem and ZCA were extracted from 1 mL human urine by mixed-mode solid-phase extraction. Samples were analyzed by LC-MS-MS using positive electrospray ionization with multiple reaction monitoring mode employed for detection and quantification. Gradient chromatographic separation was achieved with a reversed-phase column in a rapid 1.8 min analysis. The assay was linear from 4 to 1,000 µg/L for zolpidem and 4 to 10,000 µg/L for ZCA. Interday recovery (bias) and imprecision (n = 20) were 100-107% of target and 2.4-3.7% relative standard deviation, respectively. Extraction efficiencies were 78-90%. Pain compliance samples (n = 3,142) were de-identified and analyzed for zolpidem and ZCA. Zolpidem was detected greater than limit of quantification in 720 specimens (22.9%), while ZCA was detected in 1,579 specimens (50.3%). Only five specimens contained zolpidem alone. ZCA was observed without parent zolpidem in 864 specimens, thereby increasing population detection rates by 27.5%. Addition of a zolpidem metabolite to compliance determinations substantially improved detection for zolpidem intake and also should prove useful in clinical and forensic settings. PMID:25217539

  15. A comparative study of the interaction of Tamiflu and Oseltamivir carboxylate with bovine serum albumin.

    PubMed

    Vishkaee, Tahereh Sadigh; Mohajerani, Niloufar; Nafisi, Shohreh

    2013-02-01

    Oseltamivir phosphate (Tamiflu) is a pro-drug that is metabolized to its active form (Oseltamivir carboxylate), after oral administration. OC inhibits influenza A and B neuraminidases in vitro and OP inhibits influenza virus infection and replication in vitro. Serum albumin is the most abundant of the proteins in the circulatory system of a wide variety of organisms and plays an important role in the transport and deposition of many drugs. The aim of this study was to examine the interaction of BSA with Tamiflu and Oseltamivir carboxylate in aqueous solution at physiological conditions, using a constant protein concentration and various drug contents. FTIR, UV-Vis spectroscopic methods were used to determine the drugs binding mode, the binding constant and the effects of drug complexation on protein secondary structure. Structural analysis showed that OP and OC bind BSA with overall binding constants of K(OP-BSA)=1.88 (±0.16)×10(4)M(-1) and K(OC-BSA)=5.7 (±0.09)×10(2)M(-1). Drug complexation alters protein conformation by major reduction of α-helix and random coil and increase of β-sheet and turn structures that indicate a partial protein destabilization. The results suggest that BSA might act as carrier proteins for OP in delivering it to target molecules. PMID:23353784

  16. Detection of rare species of volatile organic selenium metabolites in male golden hamster urine.

    PubMed

    Kwak, Jae; Ohrnberger, Sarah A; Valencak, Teresa G

    2016-07-01

    Selenium has been considered as an essential trace element in mammals and its intake comes mainly from food. Mammals can metabolize both inorganic and organic species, and urinary excretion is the primary elimination route of selenium. Selenosugars and trimethylselenonium ion have been identified as major urinary metabolites. Other metabolites have been reported, but they were detected in some studies and not in others. Still, a large portion of the ingested selenium eliminated from the body is unknown. Volatile selenium species may account for a certain portion of the unknown species since they can easily be lost during sample analyses. While we analyzed male golden hamster urine in search of potential volatile pheromone(s), four volatile selenium compounds were detected. They were dimethyl selenenylsulfide, dimethyl diselenide, dimethyl bis(thio)selenide, and dimethyl selenodisulfide. When the urine samples were aged and dried for 48 h, dimethyl selenodisulfide tended to increase, while others decreased. The increase might be due to the formation of dimethyl selenodisulfide via reaction of dimethyl diselenide and dimethyl trisulfide whose concentration increased as urine aged. To our knowledge, dimethyl bis(thio)selenide and dimethyl selenodisulfide have never been demonstrated in urine. It remains to be determined whether these species are common metabolites in other animals or hamster-specific. PMID:27129975

  17. Effects of oseltamivir phosphate (Tamiflu) in human sera on results of microneutralization and hemagglutinin-inhibition tests for H5N2 avian influenza virus.

    PubMed

    Yamazaki, Yoshinao; Doy, M; Yamato, S; Kawada, Y; Ogata, T

    2008-01-01

    To determine the influence of oseltamivir phosphate (Tamiflu) on the results of microneutralization and hemagglutinin-inhibition (HI) tests in human sera with H5N2 influenza virus, ten volunteers were administered Tamiflu and blood samples were collected. In the microneutralization test, no consistent effects were observed. However, in the HI test, specimens from all volunteers taken at 4 and 7 h after drug administration showed a higher titer as compared to 0 and 24 h after administration when mammalian cells (horse, guinea pig, and human) were used. These results suggest that the administration of Tamiflu may affect the results of HI tests for H5N2 virus. PMID:18227965

  18. Detection of Pesticides and Pesticide Metabolites Using the Cross Reactivity of Enzyme Immunoassays

    USGS Publications Warehouse

    Thurman, E.M.; Aga, D.S.

    2001-01-01

    Enzyme immunoassay is an important environmental analysis method that may be used to identify many pesticide analytes in water samples. Because of similarities in chemical structure between various members of a pesticide class, there often may be an unwanted response that is characterized by a percentage of cross reactivity. Also, there may be cross reactivity caused by degradation products of the target analyte that may be present in the sample. In this paper, the concept of cross reactivity caused by degradation products or by nontarget analytes is explored as a tool for identification of metabolites or structurally similar compounds not previously known to be present in water samples. Two examples are examined in this paper from various water quality studies. They are alachlor and its metabolite, alachlor ethane sulfonic acid, and atrazine and its class members, prometryn and propazine. A method for using cross reactivity for the detection of these compounds is explained in this paper.

  19. Aflatoxin metabolism in humans: detection of metabolites and nucleic acid adducts in urine by affinity chromatography

    SciTech Connect

    Groopman, J.D.; Donahue, P.R.; Zhu, J.Q.; Chen, J.S.; Wogan, G.N.

    1985-10-01

    A high-affinity IgM monoclonal antibody specific for aflatoxins was covalently bound to Sepharose 4B and used as a preparative column to isolate aflatoxin derivatives from the urine of people and experimental animals who had been exposed to the carcinogen environmentally or under laboratory conditions. Aflatoxin levels were quantified by radioimmunoassay and high-performance liquid chromatography after elution from the affinity column. In studies on rats injected with ( UC)aflatoxin B1, the authors identified the major aflatoxin-DNA adduct, 2,3-dihydro-2-(N7-guanyl)-3-hydroxy-aflatoxin B1 (AFB1-N7-Gua), and the oxidative metabolites M1 and P1 as the major aflatoxin species present in the urine. When this methodology was applied to human urine samples obtained from people from the Guangxi Province of China exposed to aflatoxin B1 through dietary contamination, the aflatoxin metabolites detected were also AFB1-N7-Gua and aflatoxins M1 and P1. Therefore, affinity chromatography using a monoclonal antibody represents a useful and rapid technique with which to isolate this carcinogen and its metabolites in biochemical epidemiology and for subsequent quantitative measurements, providing exposure information that can be used for risk assessment.

  20. In vivo detection of the new psychoactive substance AM-694 and its metabolites.

    PubMed

    Bertol, Elisabetta; Vaiano, Fabio; Di Milia, Maria Grazia; Mari, Francesco

    2015-11-01

    AM-694 or 1-(5-fluoropentyl)-3-(2-iodobenzoyl)indole is a synthetic cannabinoid that acts as a selective and a powerful agonist for CB1 receptor, inducing cannabinoid-like effects (euphoria, sedation, hallucinations and anxiety). Its spread, like for other synthetic cannabinoids, has increased in recent years and many web sources freely supply these kinds of new drugs. It can be taken by smoking or through oral consumption. A 25-years-old man was hospitalized at the local hospital following a major trauma after ingestion of alcohol and an unknown pill. Urine and blood samples were sent to our Forensic Toxicology Division to investigate on possible substance abuse. A general unknown screening of biological samples, extracted by liquid-liquid extraction (ethylacetate and dichloromethane) in basic, acidic and neutral conditions, was achieved to verify the presence of drugs of abuse and/or their metabolites, both in gas chromatography-mass spectrometry (GC-MS) and liquid chromatography-tandem mass spectrometry (LC-MS/MS). For the quantification of AM-694, urine was extracted by solid phase extraction (SPE) on a Bond Elut Certify cartridge; an acidic hydrolysis (HCl 30%, 95°C, 60min) was necessary before liquid-liquid extraction of metabolites. For the detection of benzodiazepines and their metabolites, an enzymatic hydrolysis was applied (β-glucuronidase, pH 4.5, 50°C, 18h). Quantification of AM-694 (internal standard AM-2201), midazolam and α-hydroxymidazolam (internal standard halazepam) were performed by LC-MS/MS analysis in multiple reaction monitoring ([M+H](+): m/z 436→190, 272, AM-694; m/z 360→155, 127, AM-2201; m/z 326→291, 223, midazolam; m/z 342→168, 203, α-hydroxymidazolam; m/z 353→241, 222, halazepam). The general unknown screening revealed the presence of AM-694 (urine sample) and benzodiazepines (urine and blood). The concentration of AM-694, obtained by LC-MS/MS, was 0.084μg/L. Midazolam and α-hydroxymidazolam were detected in urine (0

  1. Proton observed phosphorus editing (POPE) for in vivo detection of phospholipid metabolites.

    PubMed

    Wijnen, Jannie P; Klomp, Dennis W J; Nabuurs, Christine I H C; de Graaf, Robin A; van Kalleveen, Irene M L; van der Kemp, Wybe J M; Luijten, Peter R; Kruit, Mark C; Webb, Andrew; Kan, Hermien E; Boer, Vincent O

    2016-09-01

    The purpose of this article was to compare the sensitivity of proton observed phosphorus editing (POPE) with direct (31) P MRS with Ernst angle excitation for (1) H-(31) P coupled metabolites at 7 T. POPE sequences were developed for detecting phosphocholine (PC), phosphoethanolamine (PE), glycerophosphocholine (GPC), and glycerophosphoethanolamine (GPE) on the (1) H channel, thereby using the enhanced sensitivity of the (1) H nuclei over (31) P detection. Five healthy volunteers were examined with POPE and (31) P-MRS. POPE editing showed a more than doubled sensitivity in an ideal phantom experiment as compared with direct (31) P MRS with Ernst angle excitation. In vivo, despite increased relaxation losses, significant gains in signal-to-noise ratio (SNR) of 30-40% were shown for PE and GPE + PC levels in the human brain. The SNR of GPC was lower in the POPE measurement compared with the (31) P-MRS measurement. Furthermore, selective narrowband editing on the (31) P channel showed the ability to separate the overlapping GPE and PE signals in the (1) H spectrum. POPE can be used for enhanced detection of (1) H-(31) P coupled metabolites in vivo. Copyright © 2015 John Wiley & Sons, Ltd. PMID:26601921

  2. Removal of stigmasterol from Kraft mill effluent by aerobic biological treatment with steroidal metabolite detection.

    PubMed

    Chamorro, Soledad; Vergara, Juan P; Jarpa, Mayra; Hernandez, Victor; Becerra, Jose; Vidal, Gladys

    2016-10-14

    Stigmasterol is a phytosterol contained in Kraft mill effluent that is able to increase over 100% after aerobic biological treatment. This compound can act as an endocrine disrupter as its structure is similar to that of cholesterol. The aim of this study was to evaluate the removal of stigmasterol from Kraft mill effluents treated by a moving bed biofilm reactor (MBBR) with steroidal metabolite detection. The MBBR was operated for 145 days, with a hydraulic retention time of 2 days. Stigmasterol and steroidal metabolites were detected by gas chromatography with a flame ionization detector during MBBR operation. The results show that the MBBR removed 87.4% of biological oxygen demand (BOD5), 61.5% of chemical oxygen demand (COD), 24.5% of phenol and 31.5% of lignin, expressed in average values. The MBBR system successfully removed 100% of the stigmasterol contained in the influent (33 µg L(-1)) after 5 weeks of operation. In that case, the organic load rate was 0.343 kg COD m(-3) d(-1). Furthermore, different steroidal compounds (e.g., testosterone propionate, stigmast-4-en-3-one, 5α-pregnan-12-one-20α-hydroxy, 5α-pregnane-3,11,20-trione and 3α-hydroxy-5α-androstane-11,17-dione were detected in the Kraft mill effluent as potential products of phytosterol biotransformation. PMID:27399163

  3. Meat, the metabolites: an integrated metabolite profiling and lipidomics approach for the detection of the adulteration of beef with pork

    PubMed Central

    Trivedi, Drupad K.; Hollywood, Katherine A.; Rattray, Nicholas J. W.; Ward, Holli; Trivedi, Dakshat K.; Greenwood, Joseph; Ellis, David I.

    2016-01-01

    Adulteration of high quality food products with sub-standard and cheaper grades is a world-wide problem taxing the global economy. Currently, many traditional tests suffer from poor specificity, highly complex outputs and a lack of high-throughput processing. Metabolomics has been successfully used as an accurate discriminatory technique in a number of applications including microbiology, cancer research and environmental studies and certain types of food fraud. In this study, we have developed metabolomics as a technique to assess the adulteration of meat as an improvement on current methods. Different grades of beef mince and pork mince, purchased from a national retail outlet were combined in a number of percentage ratios and analysed using GC-MS and UHPLC-MS. These techniques were chosen because GC-MS enables investigations of metabolites involved in primary metabolism whilst UHPLC-MS using reversed phase chromatography provides information on lipophilic species. With the application of chemometrics and statistical analyses, a panel of differential metabolites were found for identification of each of the two meat types. Additionally, correlation was observed between metabolite content and percentage of fat declared on meat products’ labelling. PMID:26911805

  4. Meat, the metabolites: an integrated metabolite profiling and lipidomics approach for the detection of the adulteration of beef with pork.

    PubMed

    Trivedi, Drupad K; Hollywood, Katherine A; Rattray, Nicholas J W; Ward, Holli; Trivedi, Dakshat K; Greenwood, Joseph; Ellis, David I; Goodacre, Royston

    2016-04-01

    Adulteration of high quality food products with sub-standard and cheaper grades is a world-wide problem taxing the global economy. Currently, many traditional tests suffer from poor specificity, highly complex outputs and a lack of high-throughput processing. Metabolomics has been successfully used as an accurate discriminatory technique in a number of applications including microbiology, cancer research and environmental studies and certain types of food fraud. In this study, we have developed metabolomics as a technique to assess the adulteration of meat as an improvement on current methods. Different grades of beef mince and pork mince, purchased from a national retail outlet were combined in a number of percentage ratios and analysed using GC-MS and UHPLC-MS. These techniques were chosen because GC-MS enables investigations of metabolites involved in primary metabolism whilst UHPLC-MS using reversed phase chromatography provides information on lipophilic species. With the application of chemometrics and statistical analyses, a panel of differential metabolites were found for identification of each of the two meat types. Additionally, correlation was observed between metabolite content and percentage of fat declared on meat products' labelling. PMID:26911805

  5. Detection of Phenolic Metabolites of Styrene in Mouse Liver and Lung Microsomal IncubationsS⃞

    PubMed Central

    Shen, Shuijie; Zhang, Fan; Gao, Lingbo; Zeng, Su

    2010-01-01

    Metabolic activation is considered to be a critical step for styrene-induced pulmonary toxicity. Styrene-7,8-oxide is a primary oxidative metabolite generated by vinyl epoxidation of styrene. In addition, urinary 4-vinylphenol (4-VP), a phenolic metabolite formed by aromatic hydroxylation, has been detected in workers and experimental animals after exposure to styrene. In the present study, new oxidative metabolites of styrene, including 2-vinylphenol (2-VP), 3-vinylphenol (3-VP), vinyl-1,4-hydroquinone, and 2-hydroxystyrene glycol were detected in mouse liver microsomal incubations. The production rates of 2-VP, 3-VP, 4-VP, and styrene glycol were 0.0527 ± 0.0045, 0.0019 ± 0.0006, 0.0053 ± 0.0002, and 4.42 ± 0.33 nmol/(min · mg protein) in mouse liver microsomes, respectively. Both disulfiram (100 μM) and 5-phenyl-1-pentyne (5 μM) significantly inhibited the formation of the VPs and styrene glycol. 2-VP, 3-VP, and 4-VP were metabolized in mouse liver microsomes at rates of 2.50 ± 0.30, 2.63 ± 0.13, and 3.45 ± 0.11 nmol/(min · mg protein), respectively. The three VPs were further metabolized to vinylcatechols and/or vinyl-1,4-hydroquinone and the corresponding glycols. Pulmonary toxicity of 2-VP, 3-VP, and 4-VP was evaluated in CD-1 mice, and 4-VP was found to be more toxic than 2-VP and 3-VP. PMID:20724499

  6. Development and in silico evaluation of large-scale metabolite identification methods using functional group detection for metabolomics

    PubMed Central

    Mitchell, Joshua M.; Fan, Teresa W.-M.; Lane, Andrew N.; Moseley, Hunter N. B.

    2014-01-01

    Large-scale identification of metabolites is key to elucidating and modeling metabolism at the systems level. Advances in metabolomics technologies, particularly ultra-high resolution mass spectrometry (MS) enable comprehensive and rapid analysis of metabolites. However, a significant barrier to meaningful data interpretation is the identification of a wide range of metabolites including unknowns and the determination of their role(s) in various metabolic networks. Chemoselective (CS) probes to tag metabolite functional groups combined with high mass accuracy provide additional structural constraints for metabolite identification and quantification. We have developed a novel algorithm, Chemically Aware Substructure Search (CASS) that efficiently detects functional groups within existing metabolite databases, allowing for combined molecular formula and functional group (from CS tagging) queries to aid in metabolite identification without a priori knowledge. Analysis of the isomeric compounds in both Human Metabolome Database (HMDB) and KEGG Ligand demonstrated a high percentage of isomeric molecular formulae (43 and 28%, respectively), indicating the necessity for techniques such as CS-tagging. Furthermore, these two databases have only moderate overlap in molecular formulae. Thus, it is prudent to use multiple databases in metabolite assignment, since each major metabolite database represents different portions of metabolism within the biosphere. In silico analysis of various CS-tagging strategies under different conditions for adduct formation demonstrate that combined FT-MS derived molecular formulae and CS-tagging can uniquely identify up to 71% of KEGG and 37% of the combined KEGG/HMDB database vs. 41 and 17%, respectively without adduct formation. This difference between database isomer disambiguation highlights the strength of CS-tagging for non-lipid metabolite identification. However, unique identification of complex lipids still needs additional

  7. Detection of "bath salt" synthetic cathinones and metabolites in urine via DART-MS and solid phase microextraction.

    PubMed

    LaPointe, Joseph; Musselman, Brian; O'Neill, Teresa; Shepard, Jason R E

    2015-01-01

    A rapid and sensitive method, direct analysis in real time mass spectrometry (DART-MS) was applied to the characterization and semiquantitative analysis of synthetic cathinones and their metabolites in urine. DART-MS was capable of detecting three different cathinones and three metabolites down to sub-clinical levels directly without any sample preparations. The process produced a spectrum within seconds because no extraction or derivatization was required for analysis and the high mass accuracy of the instrumentation allowed analysis without the need for lengthy chromatographic separations. The use of solid phase microextration demonstrated a relative increase in the detectability of both drugs and metabolites, improving the detection signal on average more than an order of magnitude over direct detection, while providing cleaner spectra devoid of the major peaks associated with urine that oftentimes dominate such samples. PMID:25315459

  8. Detection of "Bath Salt" Synthetic Cathinones and Metabolites in Urine via DART-MS and Solid Phase Microextraction

    NASA Astrophysics Data System (ADS)

    LaPointe, Joseph; Musselman, Brian; O'Neill, Teresa; Shepard, Jason R. E.

    2015-01-01

    A rapid and sensitive method, direct analysis in real time mass spectrometry (DART-MS) was applied to the characterization and semiquantitative analysis of synthetic cathinones and their metabolites in urine. DART-MS was capable of detecting three different cathinones and three metabolites down to sub-clinical levels directly without any sample preparations. The process produced a spectrum within seconds because no extraction or derivatization was required for analysis and the high mass accuracy of the instrumentation allowed analysis without the need for lengthy chromatographic separations. The use of solid phase microextration demonstrated a relative increase in the detectability of both drugs and metabolites, improving the detection signal on average more than an order of magnitude over direct detection, while providing cleaner spectra devoid of the major peaks associated with urine that oftentimes dominate such samples.

  9. Simultaneous detection of lysine metabolites by a single LC-MS/MS method: monitoring lysine degradation in mouse plasma.

    PubMed

    Pena, Izabella A; Marques, Lygia A; Laranjeira, Angelo B A; Yunes, José A; Eberlin, Marcos N; Arruda, Paulo

    2016-01-01

    Detection and quantification of lysine degradation metabolites in plasma is necessary for the diagnosis and follow-up of diseases such as pyridoxine-dependent epilepsy. The principal metabolites involved in the disease are related to the first steps of lysine oxidation, either through the saccharopine or the pipecolate pathways. Currently, there are three different analytical methods used to assess the content of these metabolites in urine and plasma, but they require different sample preparations and analytical equipment. Here, we describe a protocol that calls for a simple sample preparation and uses liquid chromatography tandem mass spectrometry (LC-MS/MS) that allows simultaneous detection and quantification of underivatized l-saccharopine, l-aminoadipic acid, l-pipecolic acid, piperideine-6-carboxylate, l-glutamic acid, and pyridoxal-5-phosphate in plasma samples. To validate the method we analyzed the time course degradation after intraperitoneal injection of l-lysine in C57BL/6/J mice. We observed that the degradation of lysine through the saccharopine pathway reached a maximum within the first 2 h. At this time point there was an increase in the levels of the metabolites saccharopine, aminoadipic acid, and pipecolic acid by 3-, 24- and 3.4-fold, respectively, compared to time zero levels. These metabolites returned to basal levels after 4-6 h. In conclusion, we have developed a LC-MS/MS approach, which allows simultaneous analysis of lysine degradation metabolites without the need for derivatization. PMID:27026869

  10. The production and characterisation of an antibody to detect the coccidiostat toltrazuril and its metabolite ponazuril.

    PubMed

    Connolly, Lisa; Fodey, Terence L; Crooks, Steven R H; Elliott, Christopher T

    2003-05-01

    The production of an antibody to detect toltrazuril or its metabolite ponazuril is complicated due to structural constraints of conjugating these coccidiostats to a carrier protein. Therefore a search was carried out for a compound that shared a common substructure to use as an antigen mimic. The chosen compound, trifluoraminoether, was conjugated to two carrier proteins (HSA and BTG) and used in the immunisation of six rabbits. Two immunogen doses (1 mg and 0.1 mg) were also used. All six rabbits produced an immunological response to the hapten regardless of the carrier protein or immunogen dose used. The most sensitive polyclonal antibody produced, designated R609, was subsequently characterised. This antiserum exhibited an IC50 of 18 ng ml(-1) using a competitive ELISA format. Cross reactivity studies show that this serum is specific for toltrazuril and its metabolites (toltrazuril sulfoxide and toltrazuril sulfone) but does not cross-react with other coccidiostats such as halofuginone, nitroimidazoles or nicarbazin. This is the first reported production of an antibody capable of specifically binding toltrazuril and ponazuril. PMID:12790197

  11. The Detection of Metabolite-Mediated Gene Module Co-Expression Using Multivariate Linear Models

    PubMed Central

    Padayachee, Trishanta; Khamiakova, Tatsiana; Shkedy, Ziv; Perola, Markus; Salo, Perttu; Burzykowski, Tomasz

    2016-01-01

    Investigating whether metabolites regulate the co-expression of a predefined gene module is one of the relevant questions posed in the integrative analysis of metabolomic and transcriptomic data. This article concerns the integrative analysis of the two high-dimensional datasets by means of multivariate models and statistical tests for the dependence between metabolites and the co-expression of a gene module. The general linear model (GLM) for correlated data that we propose models the dependence between adjusted gene expression values through a block-diagonal variance-covariance structure formed by metabolic-subset specific general variance-covariance blocks. Performance of statistical tests for the inference of conditional co-expression are evaluated through a simulation study. The proposed methodology is applied to the gene expression data of the previously characterized lipid-leukocyte module. Our results show that the GLM approach improves on a previous approach by being less prone to the detection of spurious conditional co-expression. PMID:26918614

  12. Analysis of the cyanolichen Lichina pygmaea metabolites using in situ DART-MS: from detection to thermochemistry of mycosporine serinol.

    PubMed

    Le Pogam, Pierre; Legouin, Béatrice; Le Lamer, Anne-Cécile; Boustie, Joël; Rondeau, David

    2015-03-01

    Direct Analysis in Real Time DART-HRMS is here first applied to the detection of molecules from a lichen, Lichina pygmaea. The aim was to propose an innovative method of in situ detection of lichen secondary metabolites using the possibilities of elemental composition determination available when a DART source is interfaced with a TOF analyzer. Three kinds of samples have been submitted to DART ionization, i.e. an intact thallus, a powder obtained from the crushed lichen and an aqueous extract. In situ analysis of crushed lichen, yields an extensive chemical profile, comparable to what is obtained from the aqueous extract, comprising both major polar metabolites described in literature along with some other signals that could correspond to potentially unknown metabolites. One of the detected secondary metabolites, mycosporine serinol, underwent a dehydration reaction prior to its transfer in the gas-phase by DART ionization. The consideration of the thermal transfers involved in the DART ionization process and the possibility to record time-dependent mass spectra through the use of the TOF analyzer allowed establishing Arrhenius plots of this water molecule loss to obtain associated thermodynamic quantities. The low values of corresponding activation enthalpy (Δr‡Hm° of the order of 25 kJ mol(-1)) enabled formulating some assumption regarding a possible role of such metabolites in the lichen. PMID:25800181

  13. Direct Standard-Free Quantitation of Tamiflu® and Other Pharmaceutical Tablets using Clustering Agents with Electrospray Ionization Mass Spectrometry

    PubMed Central

    Flick, Tawnya G.; Leib, Ryan D.; Williams, Evan R.

    2010-01-01

    Accurate and rapid quantitation is advantageous to identify counterfeit and substandard pharmaceutical drugs. A standard-free electrospray ionization mass spectrometry method is used to directly determine the dosage in the prescription and over-the-counter drugs, Tamiflu®, Sudafed®, and Dramamine®. A tablet of each drug was dissolved in aqueous solution, filtered, and introduced into solutions containing a known concentration of either L-tryptophan, L-phenylalanine or prednisone as clustering agents. The active ingredient(s) incorporates statistically into large clusters of the clustering agent where effects of differential ionization/detection are substantially reduced. From the abundances of large clusters, the dosages of the active ingredients in each of the tablets were determined to typically better than 20% accuracy even when the ionization/detection efficiency of the individual components differed by over 100×. Although this unorthodox method for quantitation is not as accurate as using conventional standards, it has the advantages that it is fast, it can be applied to mixtures where the identities of the analytes are unknown, and it can be used when suitable standards may not be readily available, such as schedule I or II controlled substances or new designer drugs that have not previously been identified. PMID:20092258

  14. Insights into the Anaerobic Biodegradation Pathway of n-Alkanes in Oil Reservoirs by Detection of Signature Metabolites.

    PubMed

    Bian, Xin-Yu; Mbadinga, Serge Maurice; Liu, Yi-Fan; Yang, Shi-Zhong; Liu, Jin-Feng; Ye, Ru-Qiang; Gu, Ji-Dong; Mu, Bo-Zhong

    2015-01-01

    Anaerobic degradation of alkanes in hydrocarbon-rich environments has been documented and different degradation strategies proposed, of which the most encountered one is fumarate addition mechanism, generating alkylsuccinates as specific biomarkers. However, little is known about the mechanisms of anaerobic degradation of alkanes in oil reservoirs, due to low concentrations of signature metabolites and lack of mass spectral characteristics to allow identification. In this work, we used a multidisciplinary approach combining metabolite profiling and selective gene assays to establish the biodegradation mechanism of alkanes in oil reservoirs. A total of twelve production fluids from three different oil reservoirs were collected and treated with alkali; organic acids were extracted, derivatized with ethanol to form ethyl esters and determined using GC-MS analysis. Collectively, signature metabolite alkylsuccinates of parent compounds from C1 to C8 together with their (putative) downstream metabolites were detected from these samples. Additionally, metabolites indicative of the anaerobic degradation of mono- and poly-aromatic hydrocarbons (2-benzylsuccinate, naphthoate, 5,6,7,8-tetrahydro-naphthoate) were also observed. The detection of alkylsuccinates and genes encoding for alkylsuccinate synthase shows that anaerobic degradation of alkanes via fumarate addition occurs in oil reservoirs. This work provides strong evidence on the in situ anaerobic biodegradation mechanisms of hydrocarbons by fumarate addition. PMID:25966798

  15. Insights into the Anaerobic Biodegradation Pathway of n-Alkanes in Oil Reservoirs by Detection of Signature Metabolites

    PubMed Central

    Bian, Xin-Yu; Maurice Mbadinga, Serge; Liu, Yi-Fan; Yang, Shi-Zhong; Liu, Jin-Feng; Ye, Ru-Qiang; Gu, Ji-Dong; Mu, Bo-Zhong

    2015-01-01

    Anaerobic degradation of alkanes in hydrocarbon-rich environments has been documented and different degradation strategies proposed, of which the most encountered one is fumarate addition mechanism, generating alkylsuccinates as specific biomarkers. However, little is known about the mechanisms of anaerobic degradation of alkanes in oil reservoirs, due to low concentrations of signature metabolites and lack of mass spectral characteristics to allow identification. In this work, we used a multidisciplinary approach combining metabolite profiling and selective gene assays to establish the biodegradation mechanism of alkanes in oil reservoirs. A total of twelve production fluids from three different oil reservoirs were collected and treated with alkali; organic acids were extracted, derivatized with ethanol to form ethyl esters and determined using GC-MS analysis. Collectively, signature metabolite alkylsuccinates of parent compounds from C1 to C8 together with their (putative) downstream metabolites were detected from these samples. Additionally, metabolites indicative of the anaerobic degradation of mono- and poly-aromatic hydrocarbons (2-benzylsuccinate, naphthoate, 5,6,7,8-tetrahydro-naphthoate) were also observed. The detection of alkylsuccinates and genes encoding for alkylsuccinate synthase shows that anaerobic degradation of alkanes via fumarate addition occurs in oil reservoirs. This work provides strong evidence on the in situ anaerobic biodegradation mechanisms of hydrocarbons by fumarate addition. PMID:25966798

  16. Detection of metabolites of trapped humans using ion mobility spectrometry coupled with gas chromatography.

    PubMed

    Vautz, Wolfgang; Slodzynski, Rafael; Hariharan, Chandrasekhara; Seifert, Luzia; Nolte, Jürgen; Fobbe, Rita; Sielemann, Stefanie; Lao, Bolan C; Huo, Ran; Thomas, C L Paul; Hildebrand, Lars

    2013-02-19

    For the first time, ion mobility spectrometry coupled with rapid gas chromatography, using multicapillary columns, was applied for the development of a pattern of signs of life for the localization of entrapped victims after disaster events (e.g., earthquake, terroristic attack). During a simulation experiment with entrapped volunteers, 12 human metabolites could be detected in the air of the void with sufficient sensitivity to enable a valid decision on the presence of a living person. Using a basic normalized summation of the measured concentrations, all volunteers involved in the particular experiments could be recognized only few minutes after they entered the simulation void and after less than 3 min of analysis time. An additional independent validation experiment enabled the recognition of a person in a room of ∼25 m(3) after ∼30 min with sufficiently high sensitivity to detect even a person briefly leaving the room. Undoubtedly, additional work must be done on analysis time and weight of the equipment, as well as on validation during real disaster events. However, the enormous potential of the method as a significantly helpful tool for search-and-rescue operations, in addition to trained canines, could be demonstrated. PMID:23249433

  17. Detection of methamphetamine and its main metabolite in fingermarks by liquid chromatography-mass spectrometry.

    PubMed

    Zhang, Ting; Chen, Xueguo; Yang, Ruiqin; Xu, Yingjian

    2015-03-01

    A sensitive and efficient method applying liquid chromatography-mass spectrometry for the analysis of methamphetamine and its main metabolite in fingermark deposits was described. Using this method, good linear relationship of methamphetamine was obtained in the range of 0.005μg to 0.5μg per cotton swab, the limit of detection was 1.5ng per cotton swab, the limit of quantitation was 5.0ng per cotton swab and the average values of recovery ratios were above 70.1%. Moreover, the influence factors for the detection of methamphetamine in fingermarks, such as kinds of substrates, development methods and extraction methods, were all discussed in details. The results showed that good recovery ratios could be obtained on painted wood and smooth substrates surfaces. Development methods in commercial powder could not influence the quality of examination of exogenous drug in latent fingermark. Furthermore, the results indicated that the method mentioned here could be applied in the analysis of forensic trace evidences and samples obtained in clinically addicted cases. PMID:25576675

  18. Detection of main metabolites of XLR-11 and its thermal degradation product in human hepatoma HepaRG cells and human urine.

    PubMed

    Kanamori, Tatsuyuki; Kanda, Koji; Yamamuro, Tadashi; Kuwayama, Kenji; Tsujikawa, Kenji; Iwata, Yuko Togawa; Inoue, Hiroyuki

    2015-04-01

    The metabolism of (1-(5-fluoropentyl)-1H-indol-3-yl)(2,2,3,3-tetramethylcyclopropyl)methanone (XLR-11), a novel synthetic cannabinoid, was studied using a HepaRG cell culture. The HepaRG cells were incubated with the drug for 48 hours and the metabolites were extracted from the culture medium by liquid-liquid extraction. The extract was analyzed by liquid chromatography/mass spectrometry to detect the metabolites. N-(5-Hydroxypentyl) metabolite and N-pentanoic acid metabolite were identified in the culture medium of XLR-11, and several other metabolites, presumably formed by oxidation of the first two metabolites and XLR-11, were detected. The extract of an XLR-11 user's urine was also analyzed; however, the metabolites detected in the urine were different from XLR-11 metabolites in the medium. A metabolic experiment with the thermal degradation product of XLR-11, XLR-11 degradant, using HepaRG cells revealed that the urinary metabolites were almost identical to the XLR-11 degradant metabolites. These findings suggest that most of the XLR-11 was degraded by heating when the user smoked the herbal product containing XLR-11. PMID:25600155

  19. Novel Metabolite Biomarkers of Huntington's Disease As Detected by High-Resolution Mass Spectrometry.

    PubMed

    Graham, Stewart F; Kumar, Praveen; Bahado-Singh, Ray O; Robinson, Andrew; Mann, David; Green, Brian D

    2016-05-01

    Huntington's disease (HD) is a fatal autosomal-dominant neurodegenerative disorder that affects approximately 3-10 people per 100 000 in the Western world. The median age of onset is 40 years, with death typically following 15-20 years later. In this study, we biochemically profiled post-mortem frontal lobe and striatum from HD sufferers (n = 14) and compared their profiles with controls (n = 14). LC-LTQ-Orbitrap-MS detected a total of 5579 and 5880 features for frontal lobe and striatum, respectively. An ROC curve combining two spectral features from frontal lobe had an AUC value of 0.916 (0.794 to 1.000) and following statistical cross-validation had an 83% predictive accuracy for HD. Similarly, two striatum biomarkers gave an ROC AUC of 0.935 (0.806 to 1.000) and after statistical cross-validation predicted HD with 91.8% accuracy. A range of metabolite disturbances were evident including but-2-enoic acid and uric acid, which were altered in both frontal lobe and striatum. A total of seven biochemical pathways (three in frontal lobe and four in striatum) were significantly altered as a result of HD. This study highlights the utility of high-resolution metabolomics for the study of HD. Further characterization of the brain metabolome could lead to the identification of new biomarkers and novel treatment strategies for HD. PMID:27018767

  20. Oxidative stress/reactive metabolite gene expression signature in rat liver detects idiosyncratic hepatotoxicants.

    PubMed

    Leone, Angelique; Nie, Alex; Brandon Parker, J; Sawant, Sharmilee; Piechta, Leigh-Anne; Kelley, Michael F; Mark Kao, L; Jim Proctor, S; Verheyen, Geert; Johnson, Mark D; Lord, Peter G; McMillian, Michael K

    2014-03-15

    Previously we reported a gene expression signature in rat liver for detecting a specific type of oxidative stress (OS) related to reactive metabolites (RM). High doses of the drugs disulfiram, ethinyl estradiol and nimesulide were used with another dozen paradigm OS/RM compounds, and three other drugs flutamide, phenacetin and sulindac were identified by this signature. In a second study, antiepileptic drugs were compared for covalent binding and their effects on OS/RM; felbamate, carbamazepine, and phenobarbital produced robust OS/RM gene expression. In the present study, liver RNA samples from drug-treated rats from more recent experiments were examined for statistical fit to the OS/RM signature. Of all 97 drugs examined, in addition to the nine drugs noted above, 19 more were identified as OS/RM-producing compounds-chlorpromazine, clozapine, cyproterone acetate, dantrolene, dipyridamole, glibenclamide, isoniazid, ketoconazole, methapyrilene, naltrexone, nifedipine, sulfamethoxazole, tamoxifen, coumarin, ritonavir, amitriptyline, valproic acid, enalapril, and chloramphenicol. Importantly, all of the OS/RM drugs listed above have been linked to idiosyncratic hepatotoxicity, excepting chloramphenicol, which does not have a package label for hepatotoxicity, but does have a black box warning for idiosyncratic bone marrow suppression. Most of these drugs are not acutely toxic in the rat. The OS/RM signature should be useful to avoid idiosyncratic hepatotoxicity of drug candidates. PMID:24486436

  1. Detection of urinary metabolites common to structurally related 17alpha-alkyl anabolic steroids in horses and application to doping tests in racehorses: methandienone, methandriol, and oxymetholone.

    PubMed

    Yamada, Masayuki; Aramaki, Sugako; Kurosawa, Masahiko; Saito, Koichi; Nakazawa, Hiroyuki

    2008-06-01

    Methandienone, methandriol, and oxymetholone, which are anabolic steroids possessing 17alpha-methyl and 17beta-hydroxy groups, were developed as oral formulations for therapeutic purposes. However, they have been used in racehorses to enhance racing performance. In humans, it has been reported that structurally related anabolic steroids having the 17alpha-methyl and 17beta-hydroxy groups, including 17alpha-methyltestosterone, mestanolone, methandienone, methandriol, and oxymetholone, have metabolites in common. In this study, we found that metabolites common to those of 17alpha-methyltestosterone and mestanolone were detected in horse urine after the administration of oxymetholone, methandienone, and methandriol. Based on analytical data, we confirmed these to be the common metabolites of five structurally related steroids, 17alpha-methyltestosterone, mestanolone, oxymetholone, methandienone, and methandriol. Furthermore, we detected hitherto unknown urinary metabolites of methandriol and oxymetholone in horses. The parent steroid itself was detected in horse urine after the administration of methandriol, other than metabolites common to 17alpha-methyltestosterone and mestanolone. On the other hand, the major metabolite of oxymetholone was mestanolone, aside from metabolites presumed to be the stereoisomers of 2-hydroxymethyl-17alpha-methyl-5alpha-androstan-3,17beta-diol and 2,17alpha-di(hydroxymethyl)-5alpha-androstan-3,17beta-diol. The simultaneous detection of common metabolites and other main metabolites would help us narrow down the candidate-administered steroid for the doping tests in racehorses. PMID:18544225

  2. Detection and quantification of residues and metabolites of medicinal products in environmental compartments, food commodities and workplaces. A review.

    PubMed

    Bottoni, Paola; Caroli, Sergio

    2015-03-15

    The toxicological assessment of medicinal products (MPs) and their residues and metabolites in the environment have become a challenging task worldwide. The contamination of environmental compartments, biota, workplace, foodstuff and feedstuff by residues and metabolites of these substances poses a risk to human health which is still far from being fully understood. On the other hand, existing analytical methods not always possess sufficient detection power to quantify residues of MPs at very low concentrations. This review sets forth some of the most significant contributions made in this field over the past decade with a special focus on novel fit-for-purpose analytical approaches for the detection, identification and quantification of these pollutants and the assessment of their noxious potential for human beings and the environment. PMID:25591909

  3. Measurement of lumefantrine and its metabolite in plasma by high performance liquid chromatography with ultraviolet detection.

    PubMed

    Khalil, Insaf F; Abildrup, Ulla; Alifrangis, Lene H; Maiga, Deogratius; Alifrangis, Michael; Hoegberg, Lotte; Vestergaard, Lasse S; Persson, Ola Per-Eric; Nyagonde, Nyagonde; Lemnge, Martha M; Theander, Thor G; Bygbjerg, Ib C

    2011-01-01

    Artemether-lumefantrine (ARM-LUM) has in recent years become the first-line treatment for uncomplicated malaria in many Sub-Saharan African countries. Vigorous monitoring of the therapeutic efficacy of this treatment is needed. This requires high-quality studies following standard protocols; ideally, such studies should incorporate measurement of drug levels in the study patients to exclude the possibility that insufficient drug levels explain an observed treatment failure. Several methods for measuring lumefantrine (LUM) in plasma by HPLC are available; however, several of these methods have some limitations in terms of high costs and limited feasibility arising from large required sample volumes and demanding sample preparation. Therefore, we set out to develop a simpler reversed phase high performance liquid chromatography (RP-HPLC) method based on UV detection for simultaneous measurement of LUM and its major metabolite the desbutyl LUM (DL) in plasma. Halofantrine was used as an internal standard. Liquid-liquid extraction of samples was carried out using hexane-ethyl acetate (70:30, v/v). Chromatographic separation was carried out on a Synergi Polar-RP column (250 mm × 300 mm, particle size 4 μm). The mobile phase consisted of acetonitrile-0.1M ammonium acetate buffer adjusted to pH 4.9 (85:15%, v/v). Absorbance of the compounds was monitored at 335 nm using a reference wavelength of 360 nm. Absolute extraction recovery for LUM and DL were 88% and 90%, respectively. Inter- and intraday coefficients of variation for LUM and DL were ≤ 10%. The lower limits of quantification for LUM and DL were 12.5 and 6.5 ng/ml, respectively. After validation, the methodology was transferred to a local laboratory in Tanga Tanzania and samples from a small subset of malaria patients were analysed for LUM. The method appears to be applicable in settings with limited facilities. PMID:20832961

  4. Oxidative stress/reactive metabolite gene expression signature in rat liver detects idiosyncratic hepatotoxicants

    SciTech Connect

    Leone, Angelique; Nie, Alex; Brandon Parker, J.; Sawant, Sharmilee; Piechta, Leigh-Anne; Kelley, Michael F. Mark Kao, L.; Jim Proctor, S.; Verheyen, Geert; Johnson, Mark D.; Lord, Peter G.; McMillian, Michael K.

    2014-03-15

    Previously we reported a gene expression signature in rat liver for detecting a specific type of oxidative stress (OS) related to reactive metabolites (RM). High doses of the drugs disulfiram, ethinyl estradiol and nimesulide were used with another dozen paradigm OS/RM compounds, and three other drugs flutamide, phenacetin and sulindac were identified by this signature. In a second study, antiepileptic drugs were compared for covalent binding and their effects on OS/RM; felbamate, carbamazepine, and phenobarbital produced robust OS/RM gene expression. In the present study, liver RNA samples from drug-treated rats from more recent experiments were examined for statistical fit to the OS/RM signature. Of all 97 drugs examined, in addition to the nine drugs noted above, 19 more were identified as OS/RM-producing compounds—chlorpromazine, clozapine, cyproterone acetate, dantrolene, dipyridamole, glibenclamide, isoniazid, ketoconazole, methapyrilene, naltrexone, nifedipine, sulfamethoxazole, tamoxifen, coumarin, ritonavir, amitriptyline, valproic acid, enalapril, and chloramphenicol. Importantly, all of the OS/RM drugs listed above have been linked to idiosyncratic hepatotoxicity, excepting chloramphenicol, which does not have a package label for hepatotoxicity, but does have a black box warning for idiosyncratic bone marrow suppression. Most of these drugs are not acutely toxic in the rat. The OS/RM signature should be useful to avoid idiosyncratic hepatotoxicity of drug candidates. - Highlights: • 28 of 97 drugs gave a positive OS/RM gene expression signature in rat liver. • The specificity of the signature for human idiosyncratic hepatotoxicants was 98%. • The sensitivity of the signature for human idiosyncratic hepatotoxicants was 75%. • The signature can help eliminate hepatotoxicants from drug development.

  5. High-performance liquid chromatography with fluorescence detection for the determination of nitrofuran metabolites in pork muscle.

    PubMed

    Sheng, Liang-Quan; Chen, Ming-Ming; Chen, Shui-Sheng; Du, Na-Na; Liu, Zhao-Di; Song, Chong-Fu; Qiao, Rui

    2013-01-01

    A simple and sensitive HPLC method with fluorescence detection (HPLC-FLD) is reported for the simultaneous determination of metabolites of four nitrofuran drugs (furazolidone, furaltadone, nitrofurantoin and nitrofurazone) in pork muscle. The method involves acid hydrolysis of the protein-bound drug metabolites and the conjugation of the released side-chains with a novel fluorescence agent 2-hydroxy-1-naphthaldehyde. After liquid-liquid extraction and effective separation of the derivatives on a YMC-Pack Polymer C18 column at 40°C under alkaline conditions, the high fluorescence intensity of these derivatives at emission wavelength λem = 463 nm enables their simultaneous determination in pork muscle at concentrations as low as 1 µg kg⁻¹. The method was validated using blank pork muscle fortified with all four metabolites at 0.5, 1.0 and 2.0 µg kg⁻¹. Recoveries were > 92.3% with RSDs < 8.5% for all four metabolites. The results obtained with HPLC-FLD and LC-MS/MS methods showed very good agreement for pork muscle samples. PMID:24283965

  6. Simultaneous detection of 93 conventional and emerging drugs of abuse and their metabolites in urine by UHPLC-MS/MS.

    PubMed

    Tang, Magdalene H Y; Ching, C K; Lee, Caroline Y W; Lam, Ying-Hoo; Mak, Tony W L

    2014-10-15

    Novel psychoactive substances (NPS) are becoming increasingly popular worldwide in recent years, some of which have been reported to cause considerable harm and even fatalities. Currently, simultaneous screening for a comprehensive panel of conventional and novel drugs of abuse is not widely available in most clinical laboratories. The aim of this study was to establish a chromatography/mass spectrometry-based analytical system for the simultaneous detection of conventional drugs of abuse and NPS in urine. Sample preparation entails enzyme digestion and solid phase extraction; analytes were then detected by liquid-chromatography tandem mass spectrometry (LC-MS/MS) with multiple reaction monitoring. Forty-seven conventional drugs (28 parent drugs, 19 metabolites) and 46 NPS analytes (44 parent drugs, two metabolites) are covered by the established method, which has been validated according to international guidelines. The method was then applied to 964 urine samples collected from drug abusers and the results revealed the presence of two NPS - TFMPP and methcathinone - as well as conventional drugs of abuse. To conclude, an LC-MS/MS method has been established that allows the simultaneous detection of over 90 conventional as well as novel psychoactive substances and metabolites in urine samples. The method was successfully applied to authentic specimens revealing the presence of conventional as well as novel drugs of abuse in the local population. PMID:25203724

  7. Abortion after deliberate Arthrotec® addition to food. Mass spectrometric detection of diclofenac, misoprostol acid, and their urinary metabolites.

    PubMed

    Watzer, Bernhard; Lusthof, Klaas J; Schweer, Horst

    2015-07-01

    Arthrotec(®) (AT) is a combination of diclofenac, a nonsteroidal anti-inflammatory drug (NSAID), and misoprostol (MP), a synthetic analogue of prostaglandin E1 (PGE1). MP is a lipophilic methyl ester prodrug. It is readily metabolized to the biologically active misoprostol acid (MPA). During the last few years, medical studies exhibited MP to be an excellent abortive. In this paper, we describe a rare criminal case of MP abortion, initiated by the expectant father. After the abortion, samples of vomit and urine were collected. Systemic exposure to MP is difficult to prove, because both MP and the active metabolite MPA are hardly excreted in urine. Therefore, in addition to routine toxicological analysis, we used slightly modified, well-established liquid and gas chromatographic/tandem mass spectrometric (LC/MS/MS and GC/MS/MS) methods, for the direct and the indirect detection of MPA and its metabolites. In this case, we were able to demonstrate the presence of the major MP metabolites 2,3-dinor-MPA and 2,3,4,5-tetranor-MPA in the urine of the victim. We also detected paracetamol, 3-methoxyparacetamol and diclofenac-glucuronide in the urine. In the vomit of the victim, we detected diclofenac and MPA. These results, combined with the criminal investigations, showed that the accused had mixed MP into the food of his pregnant girlfriend. Finally, these investigations contributed to a confession of the accused. PMID:25524762

  8. Integrated circuit-based electrochemical sensor for spatially resolved detection of redox-active metabolites in biofilms

    PubMed Central

    Bellin, Daniel L.; Sakhtah, Hassan; Rosenstein, Jacob K.; Levine, Peter M.; Thimot, Jordan; Emmett, Kevin; Dietrich, Lars E. P.; Shepard, Kenneth L.

    2014-01-01

    Despite advances in monitoring spatiotemporal expression patterns of genes and proteins with fluorescent probes, direct detection of metabolites and small molecules remains challenging. A technique for spatially resolved detection of small molecules would benefit the study of redox-active metabolites produced by microbial biofilms, which can drastically affect colony development. Here we present an integrated circuit-based electrochemical sensing platform featuring an array of working electrodes and parallel potentiostat channels. “Images” over a 3.25 × 0.9 mm area can be captured with a diffusion-limited spatial resolution of 750 μm. We demonstrate that square wave voltammetry can be used to detect, identify, and quantify (for concentrations as low as 2.6 μM) four distinct redox-active metabolites called phenazines. We characterize phenazine production in both wild-type and mutant Pseudomonas aeruginosa PA14 colony biofilms, and find correlations with fluorescent reporter imaging of phenazine biosynthetic gene expression. PMID:24510163

  9. A Comparison of Self-Reported Analgesic Use and Detection of Urinary Ibuprofen and Acetaminophen Metabolites by Means of Metabonomics

    PubMed Central

    Loo, Ruey Leng; Chan, Queenie; Brown, Ian J.; Robertson, Claire E.; Stamler, Jeremiah; Nicholson, Jeremy K.; Holmes, Elaine; Elliott, Paul

    2012-01-01

    Information on dietary supplements, medications, and other xenobiotics in epidemiologic surveys is usually obtained from questionnaires and is subject to recall and reporting biases. The authors used metabolite data obtained from hydrogen-1 (or proton) nuclear magnetic resonance (1H NMR) analysis of human urine specimens from the International Study of Macro-/Micro-Nutrients and Blood Pressure (INTERMAP Study) to validate self-reported analgesic use. Metabolic profiling of two 24-hour urine specimens per individual was carried out for 4,630 participants aged 40–59 years from 17 population samples in Japan, China, the United Kingdom, and the United States (data collection, 1996–1999). 1H NMR-detected acetaminophen and ibuprofen use was low (∼4%) among East Asian population samples and higher (>16%) in Western population samples. In a comparison of self-reported acetaminophen and ibuprofen use with 1H NMR-detected acetaminophen and ibuprofen metabolites among 496 participants from Chicago, Illinois, and Belfast, Northern Ireland, the overall rate of concordance was 81%–84%; the rate of underreporting was 15%–17%; and the rate of underdetection was approximately 1%. Comparison of self-reported unspecified analgesic use with 1H NMR-detected acetaminophen and ibuprofen metabolites among 2,660 Western INTERMAP participants revealed similar levels of concordance and underreporting. Screening for urinary metabolites of acetaminophen and ibuprofen improved the accuracy of exposure information. This approach has the potential to reduce recall bias and other biases in epidemiologic studies for a range of substances, including pharmaceuticals, dietary supplements, and foods. PMID:22223708

  10. An integrated approach for profiling oxidative metabolites and glutathione adducts using liquid chromatography coupled with ultraviolet detection and triple quadrupole-linear ion trap mass spectrometry.

    PubMed

    Chen, Guiying; Cheng, Zhongzhe; Zhang, Kerong; Jiang, Hongliang; Zhu, Mingshe

    2016-09-10

    The use of liquid chromatography (LC) coupled with triple quadrupole linear ion trap (Qtrap) mass spectrometry (MS) for both quantitative and qualitative analysis in drug metabolism and pharmacokinetic studies is of great interest. Here, a new Qtrap-based analytical methodology for simultaneous detection, structural characterization and semi-quantitation of in vitro oxidative metabolites and glutathione trapped reactive metabolites was reported. In the current study, combined multiple ion monitoring and multiple reaction monitoring were served as surveying scans to trigger product ion spectral acquisition of oxidative metabolites and glutathione adduct, respectively. Then, detection of metabolites and recovery of their MS/MS spectra were accomplished using multiple data mining approaches. Additionally, on-line ultraviolet (UV) detection was employed to determine relative concentrations of major metabolites. Analyses of metabolites of clozapine and nomifensine in rat liver microsomes not only revealed multiple oxidative metabolites and glutathione adducts, but also identified their major oxidative metabolism and bioactivation pathways. The results demonstrated that the LC/UV/MS method enabled Qtrap to perform the comprehensive profiling of oxidative metabolites and glutathione adducts in vitro. PMID:27497649

  11. Detoxification of Atrazine by Endophytic Streptomyces sp. Isolated from Sugarcane and Detection of Nontoxic Metabolite.

    PubMed

    Mesquini, Josiane A; Sawaya, Alexandra C H F; López, Begonã G C; Oliveira, Valéria M; Miyasaka, Natalia R S

    2015-12-01

    Atrazine is still one of the most used agricultural pesticides worldwide and it has been recognized as a major contaminant of surface and ground water. The aims of this research were to isolate an endophytic microorganism from leaves of sugarcane, evaluate its ability to degrade atrazine, and investigate the formation of metabolites. By sequencing of the 16S rRNA gene, the endophytic isolate atz2 was identified as Streptomyces sp. The reduction in atrazine concentration by Streptomyces sp. atz2 was 98 % and UHPLC-MS/MS analyses showed the appearance of an unknown metabolite observed as m/z 311. Ecotoxicity tests with an aquatic organism, Daphnia similis, confirmed that this metabolite was nontoxic. This mechanism of detoxification of atrazine is different from the ones of other free-living microorganisms that inhabit the soil or rhizosphere. The results show new aspects of atrazine detoxification, highlighting a new role of endophytic bacteria in plants. PMID:26467569

  12. Isoflavone metabolism in domestic cats (Felis catus): comparison of plasma metabolites detected after ingestion of two different dietary forms of genistein and daidzein.

    PubMed

    Whitehouse-Tedd, K M; Cave, N J; Ugarte, C E; Waldron, L A; Prasain, J K; Arabshahi, A; Barnes, S; Hendriks, W H; Thomas, D G

    2013-03-01

    Some felid diets contain isoflavones but the metabolic capacity of cats toward isoflavones is relatively unknown, despite the understanding that isoflavones have divergent biological potential according to their metabolite end products. The objective of this study was to determine the plasma metabolites detectable in domestic cats after exposure to 2 different dietary forms of isoflavones, either as a soy extract tablet (n = 6) or as part of a dietary matrix (n = 4). Serial blood samples were collected after isoflavone exposure to identify the plasma metabolites of each cat. Genistein was detected in its unconjugated form or as a monosulfate. Daidzein was detected as both a mono- and disulfate as well as in its unconjugated form. Other daidzein metabolites detected included equol mono- and disulfate, dihydrodaidzein, and O-desmethylangolensin. No β-glucuronide metabolites of either isoflavone were detected. Equol was produced in markedly fewer cats after ingestion of a soy extract tablet as a single oral bolus compared with cats consuming an isoflavone-containing diet. The detectable metabolites of the isoflavones, genistein and daidzein, in domestic cat plasma after dietary ingestion has been described in the present study for the first time. The metabolic capacity for isoflavones by domestic cats appears to be efficient, with only minimal proportions of the ingested amount detected in their unconjugated forms. This has implications for the potential of isoflavones to exert physiological activity in the domestic cat when consumed at concentrations representative of typical dietary intake. PMID:23307849

  13. Capillary electrophoresis with UV detection and mass spectrometry in method development for profiling metabolites of steroid hormone metabolism.

    PubMed

    Sirén, Heli; Seppänen-Laakso, Tuulikki; Oresic, Matej

    2008-08-15

    The aim of this study was to develop a method for comprehensive profiling of metabolites involved in mammalian steroid metabolism. The study was performed using the partial filling micellar electrokinetic chromatography (PF-MEKC) technique for determination of endogenous low-hydrophilic steroids. The detection techniques in capillary electrophoresis were UV absorption and electrospray mass spectrometry (ESI-MS). Thirteen steroids were included in the method development, and the selected were metabolites involved in major pathways of steroid biosynthesis. Although only eight of them could be separated and detected with UV, they could be identified by ESI-MS using selected ion monitoring (SIM) technique. Tandem MS spectra were also collected. UV detection was more sensitive than MS due to better separation of compounds and the selective signal sensitivity. The lowest limits of detection were 10-100 ng/mL for cortisone, corticosterone, hydrocortisone and testosterone. The other steroids could be detected at 500-1000 ng/mL. The identification of cortisone, corticosterone, hydrocortisone, estrogen and testosterone were made in patient urine samples and their concentrations were 1-40 microg/L. PMID:18585986

  14. Gas chromatographic-mass spectrometry method for the detection of busulphan and its metabolites in plasma and urine.

    PubMed

    El-Serafi, Ibrahim; Terelius, Ylva; Twelkmeyer, Brigitte; Hagbjörk, Ann-Louise; Hassan, Zuzana; Hassan, Moustapha

    2013-01-15

    Busulphan is an alkylating agent used as conditioning regimen prior to stem cell transplantation. Busulphan is metabolized in the liver and four major metabolites have been identified. The first metabolite is tetrahydrothiophene which is oxidized to tetrahydrothiophene 1-oxide, then sulfolane and finally 3-hydroxy sulfolane. Despite the low molecular weight and wide polarity range of busulphan and its four metabolites, the use of a fused silica non-polar column significantly enhanced the automated gas chromatography-mass spectrometry of their detection in one simple method. The limit of quantification was 0.5μM for busulphan and all its metabolites except 3-OH sulfolane, which was 1.25μM. This method was validated for all the compounds in both human plasma and urine. Lower limits of quantifications (LLOQs) were run in pentaplicate per compound and all results were within 20% of the nominal values. The recovery was determined by comparing the peak area of two quality control (QC) samples, before and after extraction in plasma and urine, in triplicate. Acceptable precision and accuracy have been obtained; at least 3 standard curves have been run for each compound using three different QCs covering the calibration curve in triplicate. The QC values were within 15% (SD) of the nominal values. Selectivity and sensitivity of all compounds have been measured. Compounds were stable up to 50 days after extraction in -20°C and 48h at RT. Moreover, the compounds were stable for three cycles of freezing and thawing. The method was applied in a clinical case where the patient received high dose busulphan; all the compounds have been detected, identified and quantified both in plasma and urine. PMID:23286981

  15. Enzyme-DNA biocolloids for DNA adduct and reactive metabolite detection by chromatography-mass spectrometry.

    PubMed

    Bajrami, Besnik; Hvastkovs, Eli G; Jensen, Gary C; Schenkman, John B; Rusling, James F

    2008-02-15

    Silica microbead bioreactors (0.5 microm diameter) coated with DNA and enzymes were fabricated to measure reactive metabolite and DNA-adduct formation rates relevant to genotoxicity screening. Cytochrome (cyt) P450 2E1, cyt P450(cam), and myoglobin (Mb) were incorporated into thin films with DNA using the electrostatic layer-by-layer (LbL) method. The utility of these biocolloids was demonstrated by oxidation of guaiacol, styrene, and (4-methylnitrosoamino)-1-(3-pyridyl)-1-butanone (NNK). Enzyme turnover rates for formation of reactive metabolites were monitored using gas chromatography/mass spectrometry (GC/MS) and liquid chromatography-mass spectrometry (LC-MS). Capillary LC-MS/MS was employed to determine DNA nucleobase adducts after catalyzing the reactive metabolite formation with DNA-enzyme biocolloids and then using neutral thermal hydrolysis on the biocolloids. Dramatic improvements in surface area to volume ratio over similar films on macroscopic surfaces opens new avenues for genotoxicity screening and enabled the first use of pure cyt P450 enzymes in enzyme-DNA films to produce DNA adducts. The method makes possible identification and formation rate measurements of major and minor DNA adducts as well as the metabolites themselves in <5 min of reaction time using relevant human liver enzymes. PMID:18217727

  16. Showing Its Colors. Thin-Layer Chromatographic Detection of Cannabinoid Metabolites.

    ERIC Educational Resources Information Center

    Bonicamp, Judith M.

    1986-01-01

    Describes a chemistry laboratory experiment in which thin-layer chromatography (TLC) is used to analyze urine specimens containing metabolites of the drug tetrahydro-cannabinol, which comes from the marijuana plant. The materials needed to conduct the experiment are listed, and the procedure and expected results are outlined. (TW)

  17. Mutation-induced loop opening and energetics for binding of tamiflu to influenza N8 neuraminidase.

    PubMed

    Kar, Parimal; Knecht, Volker

    2012-05-31

    Tamiflu, also known as oseltamivir (OTV), binds to influenza A neuraminidase (H5N1) with very high affinity (0.32 nM). However, this inhibitor binds to other neuraminidases as well. In the present work, a systematic computational study is performed to investigate the mechanism underlying the binding of oseltamivir to N8 neuraminidase (NA) in "open" and "closed" conformations of the 150-loop through molecular dynamics simulations and the popular and well established molecular mechanics Poisson-Boltzmann (MM-PBSA) free energy calculation method. Whereas the closed conformation is stable for wild type N8, it transforms into the open conformation for the mutants Y252H, H274Y, and R292K, indicating that bound to oseltamivir these mutants are preferentially in the open conformation. Our calculations show that the binding of wild type oseltamivir to the closed conformation of N8 neuraminidase is energetically favored compared to the binding to the open conformation. We observe water mediated binding of oseltamivir to the N8 neuraminidase in both conformations which is not seen in the case of binding of the same drug to the H5N1 neuraminidase. The decomposition of the binding free energy reveals the mechanisms underlying the binding and changes in affinity due to mutations. Considering the mutant N8 variants in the open conformation adopted during the simulations, we observe a significant loss in the size of the total binding free energy for the N8(Y252H)-OTV, N8(H274Y)-OTV, and N8(R292K)-OTV complexes compared to N8(WT)-OTV, mainly due to the decrease in the size of the intermolecular electrostatic energy. For R292K, an unfavorable shift in the van der Waals interactions also contributes to the drug resistance. The mutations cause a significant expansion in the active site cavity, increasing its solvent accessible surface compared to the crystal structures of both the open and closed conformations. Our study underscores the need to consider dynamics in rationalizing the

  18. Advances in Electronic-Nose Technologies for the Detection of Volatile Biomarker Metabolites in the Human Breath

    PubMed Central

    Wilson, Alphus D.

    2015-01-01

    Recent advancements in the use of electronic-nose (e-nose) devices to analyze human breath profiles for the presence of specific volatile metabolites, known as biomarkers or chemical bio-indicators of specific human diseases, metabolic disorders and the overall health status of individuals, are providing the potential for new noninvasive tools and techniques useful to point-of-care clinical disease diagnoses. This exciting new area of electronic disease detection and diagnosis promises to yield much faster and earlier detection of human diseases and disorders, allowing earlier, more effective treatments, resulting in more rapid patient recovery from various afflictions. E-nose devices are particularly suited for the field of disease diagnostics, because they are sensitive to a wide range of volatile organic compounds (VOCs) and can effectively distinguish between different complex gaseous mixtures via analysis of electronic aroma sensor-array output profiles of volatile metabolites present in the human breath. This review provides a summary of some recent developments of electronic-nose technologies, particularly involving breath analysis, with the potential for providing many new diagnostic applications for the detection of specific human diseases associated with different organs in the body, detectable from e-nose analyses of aberrant disease-associated VOCs present in air expired from the lungs. PMID:25738426

  19. Detection and mapping of illicit drugs and their metabolites in fingermarks by MALDI MS and compatibility with forensic techniques.

    PubMed

    Groeneveld, G; de Puit, M; Bleay, S; Bradshaw, R; Francese, S

    2015-01-01

    Despite the proven capabilities of Matrix Assisted Laser Desorption Ionisation Mass Spectrometry (MALDI MS) in laboratory settings, research is still needed to integrate this technique into current forensic fingerprinting practice. Optimised protocols enabling the compatible application of MALDI to developed fingermarks will allow additional intelligence to be gathered around a suspect's lifestyle and activities prior to the deposition of their fingermarks while committing a crime. The detection and mapping of illicit drugs and metabolites in latent fingermarks would provide intelligence that is beneficial for both police investigations and court cases. This study investigated MALDI MS detection and mapping capabilities for a large range of drugs of abuse and their metabolites in fingermarks; the detection and mapping of a mixture of these drugs in marks, with and without prior development with cyanoacrylate fuming or Vacuum Metal Deposition, was also examined. Our findings indicate the versatility of MALDI technology and its ability to retrieve chemical intelligence either by detecting the compounds investigated or by using their ion signals to reconstruct 2D maps of fingermark ridge details. PMID:26118853

  20. Detection and mapping of illicit drugs and their metabolites in fingermarks by MALDI MS and compatibility with forensic techniques

    PubMed Central

    Groeneveld, G.; de Puit, M.; Bleay, S.; Bradshaw, R.; Francese, S.

    2015-01-01

    Despite the proven capabilities of Matrix Assisted Laser Desorption Ionisation Mass Spectrometry (MALDI MS) in laboratory settings, research is still needed to integrate this technique into current forensic fingerprinting practice. Optimised protocols enabling the compatible application of MALDI to developed fingermarks will allow additional intelligence to be gathered around a suspect’s lifestyle and activities prior to the deposition of their fingermarks while committing a crime. The detection and mapping of illicit drugs and metabolites in latent fingermarks would provide intelligence that is beneficial for both police investigations and court cases. This study investigated MALDI MS detection and mapping capabilities for a large range of drugs of abuse and their metabolites in fingermarks; the detection and mapping of a mixture of these drugs in marks, with and without prior development with cyanoacrylate fuming or Vacuum Metal Deposition, was also examined. Our findings indicate the versatility of MALDI technology and its ability to retrieve chemical intelligence either by detecting the compounds investigated or by using their ion signals to reconstruct 2D maps of fingermark ridge details. PMID:26118853

  1. Detection and mapping of illicit drugs and their metabolites in fingermarks by MALDI MS and compatibility with forensic techniques

    NASA Astrophysics Data System (ADS)

    Groeneveld, G.; de Puit, M.; Bleay, S.; Bradshaw, R.; Francese, S.

    2015-06-01

    Despite the proven capabilities of Matrix Assisted Laser Desorption Ionisation Mass Spectrometry (MALDI MS) in laboratory settings, research is still needed to integrate this technique into current forensic fingerprinting practice. Optimised protocols enabling the compatible application of MALDI to developed fingermarks will allow additional intelligence to be gathered around a suspect’s lifestyle and activities prior to the deposition of their fingermarks while committing a crime. The detection and mapping of illicit drugs and metabolites in latent fingermarks would provide intelligence that is beneficial for both police investigations and court cases. This study investigated MALDI MS detection and mapping capabilities for a large range of drugs of abuse and their metabolites in fingermarks; the detection and mapping of a mixture of these drugs in marks, with and without prior development with cyanoacrylate fuming or Vacuum Metal Deposition, was also examined. Our findings indicate the versatility of MALDI technology and its ability to retrieve chemical intelligence either by detecting the compounds investigated or by using their ion signals to reconstruct 2D maps of fingermark ridge details.

  2. Efficient approach for the detection and identification of new androgenic metabolites by applying SRM GC-CI-MS/MS: a methandienone case study.

    PubMed

    Polet, Michael; Van Gansbeke, Wim; Van Eenoo, Peter; Deventer, Koen

    2016-07-01

    Identification of anabolic androgenic steroids (AAS) is a vital issue in doping control and toxicology, and searching for metabolites with longer detection times remains an important task. Recently, a gas chromatography chemical ionization triple quadrupole mass spectrometry (GC-CI-MS/MS) method was introduced, and CI, in comparison with electron ionization (EI), proved to be capable of increasing the sensitivity significantly. In addition, correlations between AAS structure and fragmentation behavior could be revealed. This enables the search for previously unknown but expected metabolites by selection of their predicted transitions. The combination of both factors allows the setup of an efficient approach to search for new metabolites. The approach uses selected reaction monitoring which is inherently more sensitive than full scan or precursor ion scan. Additionally, structural information obtained from the structure specific CI fragmentation pattern facilitates metabolite identification. The procedure was demonstrated by a methandienone case study. Its metabolites have been studied extensively in the past, and this allowed an adequate evaluation of the efficiency of the approach. Thirty three metabolites were detected, including all relevant previously discovered metabolites. In our study, the previously reported long-term metabolite (18-nor-17β-hydroxymethyl,17α-methyl-androst-1,4,13-trien-3-one) could be detected up to 26 days by using GC-CI-MS/MS. The study proves the validity of the approach to search for metabolites of new synthetic AAS and new long-term metabolites of less studied AAS and illustrates the increase in sensitivity by using CI. Copyright © 2016 John Wiley & Sons, Ltd. PMID:27434811

  3. Production and quantification of sesquiterpenes in Saccharomyces cerevisiae, including extraction, detection and quantification of terpene products and key related metabolites.

    PubMed

    Rodriguez, Sarah; Kirby, James; Denby, Charles M; Keasling, Jay D

    2014-08-01

    The procedures described here are designed for engineering Saccharomyces cerevisiae to produce sesquiterpenes with an aim to either increase product titers or to simply generate a quantity of product sufficient for identification and/or downstream experimentation. Engineering high-level sesquiterpene production in S. cerevisiae often requires iterations of strain modifications and metabolite analysis. To address the latter, the methods described here were tailored for robust measurement of metabolites that we have found to be fundamental indicators of pathway flux, using only gas chromatography and mass spectrometry (GC-MS) instrumentation. Thus, by focusing on heterologous production of sesquiterpenes via the mevalonate (MEV) pathway in S. cerevisiae, we detail procedures for extraction and detection of the key pathway metabolites MEV, squalene and ergosterol, as well as the farnesyl pyrophosphate (FPP)-derived side products farnesol and nerolidol. Analysis of these compounds is important for quality control, because they are possible indicators of pathway imbalance. As many of the sesquiterpene synthase (STS) genes encountered in nature are of plant origin and often not optimal for expression in yeast, we provide guidelines for designing gene expression cassettes to enable expression in S. cerevisiae. As a case study for these protocols, we have selected the sesquiterpene amorphadiene, native to Artemisia annua and related plants. The analytical steps can be completed within 1-2 working days, and a typical experiment might take 1 week. PMID:25058645

  4. Determination of spinosad and its metabolites in food and environmental matrices. 1. High-performance liquid chromatography with ultraviolet detection.

    PubMed

    West, S D; Yeh, L T; Turner, L G; Schwedler, D A; Thomas, A D; Duebelbeis, D O

    2000-11-01

    Spinosad is an insect control agent that is derived from a naturally occurring soil bacterium and is effective on several classes of insects, especially Lepidoptera larvae. Spinosad is registered in many countries for use on a variety of crops, including cotton, corn, soybeans, fruits, and vegetables. Residue methods utilizing high-performance liquid chromatography (HPLC) with ultraviolet (UV) detection have been described for determining spinosad and its metabolites in environmental and food matrices. These residue methods typically involve an extraction with organic solvents, followed by purification using liquid-liquid partitioning and/or solid phase extraction prior to measurement by HPLC-UV. The residue methods determine the active ingredients (spinosyns A and D) and up to three minor metabolites (spinosyn B, spinosyn K, and N-demethylspinosyn D). The methods have validated limits of quantitation ranging from 0.010 to 0.040 microgram/g. This paper briefly reviews the residue methodology for spinosad and metabolites in food and environmental matrices and provides a summary of method validation results for 61 different sample types, including newly published results for 37 additional crop matrices and processed commodities. PMID:11087448

  5. Analysis of mitragynine and metabolites in human urine for detecting the use of the psychoactive plant kratom.

    PubMed

    Le, David; Goggin, Melissa M; Janis, Gregory C

    2012-01-01

    The leaves of the South Asian plant kratom are described as having stimulating effects at low doses, and opiate-like analgesic and euphoric effects at high doses. A long history of use and abuse has led to the classification of kratom as a controlled substance in its native Thailand and other South Asian countries. However, kratom is not controlled in the United States, and the ready availability of kratom has led to its emergence as an herbal drug of abuse. With the growing popularity of kratom, efficient procedures are needed to detect kratom use. In the current study, both ultra-high-performance liquid chromatography and high-performance liquid chromatography-tandem mass spectrometry methods have been developed and validated for monitoring the major alkaloids and metabolites found in urine following kratom use. The primary unique alkaloid mitragynine is quantified in human urine from 1.00-500.00 ng/mL using mitraphylline as an internal standard. In addition, two metabolites (5-desmethylmitragynine and 17-desmethyldihydromitragynine) and the related active, alkaloid 7-hydroxy-mitragynine, are simultaneously qualitatively monitored. The presence of analytes are confirmed by an information-dependent acquisition-enhanced product ion procedure generating full fragmentation data used to positively identify detected analytes. The validated method has been utilized for clinical and forensic analyses of urine for the detection of kratom use. PMID:23024321

  6. [Detection and identification of major metabolites of clorprenaline in swine urine using ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry].

    PubMed

    Bi, Yanfeng; Wang, Yilin; Ye, Ni; Sun, Lei; Wang, Hejia; Xu, Shixin; Xiao, Xilong

    2015-07-01

    This study was conducted to detect and identify the metabolites of clorprenaline in swine urine using ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC/Q-TOF MS), and the major metabolic pathways of clorprenaline were proposed. The swines were administered a single dose each of 10 mg/kg b. w. clorprenaline by oral gavage. The urine samples were collected before and after administration. After a simple preparation, the urine samples were analyzed using UPLC/Q-TOF MS. Combined with data processing techniques including extracted ion chromatography (EIC) and mass defect filtering (MDF), two phase I and seven phase II metabolites were detected in the urine samples collected 0-24 h after administration. The structures of detected metabolites were elucidated by comparing their characteristic product ions with those of the parent clorprenaline. Based on the identified metabolites, the metabolic pathways of clorprenaline included hydroxylation, glucuronidation and sulphate conjugates. Among those detected metabolites, hydroxylated-clorprenaline and its conjugates were responsible for over 60% of the total MS responses, much greater than those of clorprenaline, and were proposed as the primary metabolites in swine urine. This study can provide scientific basis for determining appropriate marker residues of clorprenaline, and facilitate to effectively control clorprenaline residues in animals. PMID:26672198

  7. Application of stable isotope labeled glutathione and rapid scanning mass spectrometers in detecting and characterizing reactive metabolites.

    PubMed

    Mutlib, Abdul; Lam, Wing; Atherton, Jim; Chen, Hao; Galatsis, Paul; Stolle, Wayne

    2005-01-01

    The formation of reactive metabolites from a number of compounds was studied in vitro using a mixture of non-labeled and stable isotope labeled glutathione (GSH) as a trapping agent. GSH was labeled by incorporating [1,2-(13)C(2),(15)N]glycine into the tripeptide to give an overall increase of 3 Da over the naturally occurring substance. Detection and characterization of reactive metabolites was greatly facilitated by using the data-dependent scanning features of the linear ion trap mass spectrometers to give complimentary and confirmatory data in a single analytical run. A comparison was made by analyzing the samples simultaneously on a triple-stage quadrupole mass spectrometer operated in the constant neutral loss mode. The compounds studied included 2-acetamidophenol, 3-acetamidophenol, 4-acetamidophenol (acetaminophen), and flufenamic acid. GSH adducts for each of these compounds produced a characteristic pattern of 'twin ions' separated by 3 Da in the mass spectral data. This greatly facilitated the detection and characterization of any GSH-related adducts present in the microsomal extracts. Furthermore, characterization of these adducts was greatly facilitated by the rapid scanning capability of linear ion trap instruments that provided full-scan, MS/MS and MS(3) data in one single analysis. This method of detecting and characterizing reactive metabolites generated in vitro was found to be far superior to any of the existing methods previously employed in this laboratory. The combination of two techniques, stable isotope labeled glutathione and linear ion traps, provided a very sensitive and specific method of identifying compounds capable of producing reactive metabolites in a discovery setting. The complimentary set of mass spectral data (including full-scan, MS/MS and MS(3) mass spectra), obtained rapidly in a single analysis with the linear ion trap instruments, greatly accelerated identification of metabolically bioactivated soft spots on the molecules

  8. [Cross-reactivity of Instant-View M-1 for detection of benzodiazepine-related drugs and their metabolites in urine].

    PubMed

    Torikoshi, Aiko; Namera, Akira; Arima, Yousuke; Toubou, Hirokazu; Tajima, Takashi; Shiraishi, Hiroaki; Nagao, Masataka

    2014-03-01

    Immunoassays are useful methods for the determination of regulated drugs in clinical and forensic laboratories. Although the Instant-View M-1 (IV M-1) immunoassay kit is frequently used to screen drugs in laboratories in Japan, basic information about the IV M-1 such as its specificity and reactivity is not available. In this study, we determined the specificity and cross-reactivity of IV M-1 for the detection of benzodiazepine-related drugs and their metabolites in urine. The IV M-1 could detect triazolobenzodiazepines such as triazolam in urine at concentrations > or = 300 ng/mL. However, thienodiazepines such as etizolam could not be detected because of lack of cross reactivity. A correlation was observed between the structure of the metabolites and the reactivity of the kit; 4-hydroxy metabolites of alprazolam and triazolam were detectable, whereas a-hydroxy metabolites were not. Furthermore, 7-amino metabolites such as nitrazepam could not be detected at any concentration, including high concentrations. The specificity and reactivity of various kits used for detection of drugs in urine are different. Therefore, it is necessary to consider the basic features of the kit used while assessing the results obtained. PMID:24724359

  9. Determination of spinosad and its metabolites in citrus crops and orange processed commodities by HPLC with UV detection.

    PubMed

    West, S D; Turner, L G

    2000-02-01

    Spinosad is an insect control agent that is derived from a naturally occurring organism and is effective on a wide variety of crops, including citrus crops. A method is described for the determination of spinosad and its metabolites in citrus crops and orange processed commodities. The method determines residues of the active ingredients (spinosyns A and D) and three minor metabolites (spinosyn B, spinosyn K, and N-demethylspinosyn D). For dried orange pulp and orange oil, the method has a limit of quantitation (LOQ) of 0.02 microg/g and a limit of detection (LOD) of 0.006 microg/g. For all other sample matrices (whole fruit, edible fruit, juice, and peel), the method has an LOQ of 0.01 microg/g and an LOD of 0.003 microg/g. The analytes are extracted from the various sample types using appropriate solvents, and the extracts are purified by liquid-liquid partitioning and/or solid-phase extraction. All five analytes are determined simultaneously in the purified extracts by reversed-phase high-performance liquid chromatography with ultraviolet detection at 250 nm. PMID:10691641

  10. The detection of androstenedione abuse in sport: a mass spectrometry strategy to identify the 4-hydroxyandrostenedione metabolite.

    PubMed

    Cawley, Adam T; Trout, Graham J; Kazlauskas, Rymantas; George, Adrian V

    2008-12-01

    Studies have shown that the administration of androstenedione (ADIONE) significantly increases the urinary ratio of testosterone glucuronide to epitestosterone glucuronide (T/E) - measured by gas chromatography/mass spectrometry (GC/MS) - in subjects with a normal ( approximately 1) or naturally high (>1) initial values. However, the urinary T/E ratio has been shown not to increase in subjects with naturally low (<1) initial values. Such cases then rely on the detection of C(6)-hydroxylated metabolites shown to be indicative of ADIONE administration. While these markers may be measured in the routine GC/MS steroid profile, their relatively low urinary excretion limits the use of gas chromatography/combustion/isotope ratio mass spectrometry (GC/C/IRMS) to specifically confirm ADIONE administration based on depleted (13)C content. A mass spectrometry strategy was used in this study to identify metabolites of ADIONE with the potential to provide compound-specific detection. C(4)-hydroxylation was subsequently shown to be a major metabolic pathway following ADIONE administration, thereby resulting in urinary excretion of 4-hydroxyandrostenedione (4OH-ADIONE). Complementary analysis of 4OH-ADIONE by GC/MS and GC/C/IRMS was used to confirm ADIONE administration. PMID:19025884

  11. A sensitive GC-EIMS method for simultaneous detection and quantification of JWH-018 and JWH-073 carboxylic acid and hydroxy metabolites in urine.

    PubMed

    Paul, Buddha D; Bosy, Thomas

    2015-04-01

    Synthetic cannabinoids, including JWH-018 and JWH-073, belong to a class of aminoalkylindoles (AAIs) that are smoked to produce an effect similar to tetrahydrocannabinol. Compounds in this class are often collectively known as 'Spice'. After ingestion, these compounds are extensively metabolized to their hydroxy and carboxylic acid metabolites. During forensic analysis, detection of these metabolites in urine is an indication of past exposure to the parent compounds. The analytical process involved hydrolysis of conjugated metabolites by glucuronidase, solvent extraction, derivatization by trifluoroacetic anhydride and hexafluoroisopropanol and GC-EIMS detection. Identification of the unknown was based on the criteria of GC retention time within ±2% and mass spectral ion ratio within ±20% of that of a standard. Deuterated internal standards of the carboxylic acid metabolites were used for quantification. The acid (JWH-018-COOH, JWH-073-COOH) and hydroxy (JWH-018-OH, JWH-073-OH) metabolites were linear over the concentration range of 0.1-10 and 0.2-10 ng/mL, respectively, with a correlation coefficient-square, R(2) > 0.999 (N = 5). Extraction recoveries of the metabolites were within 79 and 87%. The method was applied to 17 urine specimens collected as part of a military law enforcement investigation. Nine of the specimens tested positive for one or more of the metabolites. When the procedure was extended to screen other AAI compounds, two of the specimens were found to contain JWH-210, JWH-250 (JWH-302 or JWH-201) and JWH-250 (C4 isomers). The GC-EIMS method presented here was found to be suitable for detecting JWH-018 and JWH-073 metabolites and other AAI compounds in urine. PMID:25691387

  12. Detection and structural characterization of glutathione-trapped reactive metabolites using liquid chromatography-high-resolution mass spectrometry and mass defect filtering.

    PubMed

    Zhu, Mingshe; Ma, Li; Zhang, Haiying; Humphreys, W Griffith

    2007-11-01

    The present study was designed to apply the mass defect filter (MDF) approach to the screening and identification of reactive metabolites using high-resolution mass spectrometry. Glutathione (GSH)-trapped reactive metabolites of acetaminophen, diclofenac, carbamazepine, clozapine, p-cresol, 4-ethylphenol, and 3-methylindole in human liver microsomes (HLM) were analyzed by HPLC coupled with Orbitrap or Fourier transform ion cyclotron resonance mass spectrometry. Through the selective removal of all ions that fall outside of the GSH adduct MDF template windows, the processed full scan MS chromatograms displayed GSH adducts as major components with no or a few interference peaks. The accurate mass LC-MS data sets were also utilized for the elimination of false positive peaks, detection of stable oxidative metabolites with other MDF templates, and determination of metabolite molecular formulas. Compared to the neutral loss scan by a triple quadrupole instrument, the MDF approach was more sensitive and selective in screening for GSH-trapped reactive metabolites in HLM and rat bile and far more effective in detecting GSH adducts that do not afford the neutral loss of 129 Da as a significant fragmentation pathway. The GSH adduct screening capability of the MDF approach, together with the utility of accurate mass MS/MS information in structural elucidation, makes high-resolution LC-MS a useful tool for analyzing reactive metabolites. PMID:17918967

  13. Ibuprofen exposure in Lemna gibba L.: Evaluation of growth and phytotoxic indicators, detection of ibuprofen and identification of its metabolites in plant and in the medium.

    PubMed

    Pietrini, F; Di Baccio, D; Aceña, J; Pérez, S; Barceló, D; Zacchini, M

    2015-12-30

    Ibuprofen (IBU) is detected worldwide in water bodies due to the incomplete removal by wastewater treatments. Contrasting results have been reported on the toxicity of IBU on aquatic biomonitor plants such as duckweed, and no data about IBU detection and metabolism in plants has been reported. In this work, the effects of 1 mg L(-1) IBU on Lemna gibba L. were monitored in an 8-day laboratory test. In particular, an increase in frond number (+12%) and multiplication rate (+10%) while no variations in photosynthetic pigment content were observed. Moreover, UPLC-HRMS analysis of the presence of IBU and its metabolites in plants and in the growth medium was performed. The results showed that, besides IBU, 11 IBU metabolites were detected in plants. Among the IBU metabolites, hydroxyl- and dihydroxyl-IBU were found, whereas carboxyl-IBU was undetectable. Interestingly, some IBU metabolites were detected in the plant growth solution at the end of the IBU treatment, while no IBU products were found in the IBU solution without plants, suggesting a role for L. gibba in IBU metabolism. The findings of this work represent an important step for a better evaluation of the effects of IBU and its metabolites in duckweed, with notable implications for the eco-toxicological assessment of IBU in the aquatic ecosystem. PMID:26184801

  14. Liquid Microjunction Surface Sampling Probe Electrospray Mass Spectrometry for Detection of Drugs and Metabolites in Thin Tissue Sections

    SciTech Connect

    Van Berkel, Gary J; Kertesz, Vilmos; Koeplinger, Kenneth A.; Vavek, Marissa; Kong, Ah-Ng Tony

    2008-01-01

    A self-aspirating, liquid micro-junction surface sampling probe/electrospray emitter mass spectrometry system was demonstrated for use in the direct analysis of spotted and dosed drugs and their metabolites in thin tissue sections. Proof-of-principle sampling and analysis directly from tissue without the need for sample preparation was demonstrated first by raster scanning a region on a section of rat liver onto which reserpine was spotted. The mass spectral signal from selected reaction monitoring was used to develop a chemical image of the spotted drug on the tissue. The probe was also used to selectively spot sample areas of sagittal whole mouse body tissue sections that had been dosed orally (90 mg/kg) with R,S-sulforaphane 3 hrs prior to sacrifice. Sulforaphane and its glutathione and N-acetyl cysteine conjugates were monitored with selected reaction monitoring and detected in the stomach and various other tissues from the dosed mouse. No signal for these species was observed in the tissue from a control mouse. The same dosed tissue section was used to illustrate the possibility of obtaining a line scan across the whole body section. In total these results illustrate the potential for rapid screening of the distribution of drugs and metabolites in tissue sections with the micro-liquid junction surface sampling probe/electrospray mass spectrometry approach.

  15. Direct Analysis in Real Time-Mass Spectrometry for the Rapid Detection of Metabolites of Aconite Alkaloids in Intestinal Bacteria

    NASA Astrophysics Data System (ADS)

    Li, Xue; Hou, Guangyue; Xing, Junpeng; Song, Fengrui; Liu, Zhiqiang; Liu, Shuying

    2014-12-01

    In the present work, direct analysis of real time ionization combined with multi-stage tandem mass spectrometry (DART-MSn) was used to investigate the metabolic profile of aconite alkaloids in rat intestinal bacteria. A total of 36 metabolites from three aconite alkaloids were identified by using DART-MSn, and the feasibility of quantitative analysis of these analytes was examined. Key parameters of the DART ion source, such as helium gas temperature and pressure, the source-to-MS distance, and the speed of the autosampler, were optimized to achieve high sensitivity, enhance reproducibility, and reduce the occurrence of fragmentation. The instrument analysis time for one sample can be less than 10 s for this method. Compared with ESI-MS and UPLC-MS, the DART-MS is more efficient for directly detecting metabolic samples, and has the advantage of being a simple, high-speed, high-throughput method.

  16. Detection of benzimidazole carbamates and amino metabolites in liver by surface plasmon resonance-biosensor

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Two surface plasmon resonance (SPR) biosensor screening assays were developed and validated to detect 11 benzimidazole carbamate (BZT) and four amino-benzimidazole veterinary drug residues in liver tissue. The assays used polyclonal antibodies, raised in sheep, to detect BZTs and amino-benzimidazole...

  17. Old Meets New: Using Interspecies Interactions to Detect Secondary Metabolite Production in Actinomycetes

    PubMed Central

    Clardy, Jon; Kolter, Roberto

    2014-01-01

    Actinomycetes, a group of filamentous, Gram-positive bacteria, have long been a remarkable source of useful therapeutics. Recent genome sequencing and transcriptomic studies have shown that these bacteria, responsible for half of the clinically used antibiotics, also harbor a large reservoir of gene clusters, which have the potential to produce novel secreted small molecules. Yet, many of these clusters are not expressed under common culture conditions. One reason why these clusters have not been linked to a secreted small molecule lies in the way that actinomycetes have typically been studied: as pure cultures in nutrient-rich media that do not mimic the complex environments in which these bacteria evolved. New methods based on multispecies culture conditions provide an alternative approach to investigating the products of these gene clusters. We have recently implemented binary interspecies interaction assays to mine for new secondary metabolites and to study the underlying biology in inter-actinomycete interactions. Here we describe the detailed biological and chemical methods comprising these studies. PMID:23084935

  18. Determination of mesotrione residues and metabolites in crops, soil, and water by liquid chromatography with fluorescence detection.

    PubMed

    Alferness, Philip; Wiebe, Lawrence

    2002-07-01

    A method for the determination of residues of mesotrione and two metabolites in a variety of environmental matrixes has been developed. Mesotrione, a new selective herbicide for use in corn, is 2-(4-methylsulfonyl-2-nitrobenzoyl)-1,3-cyclohexanedione. The metabolite 4-methylsulfonyl-2-nitrobenzoic acid (MNBA) is determined with the parent compound in crops, whereas two metabolites, 2-amino-4-methylsulfonyl-benzoic acid (AMBA) and MNBA are determined with parent in soil and water. Crop samples are macerated with an acetonitrile/water mixture, and an aliquot is evaporated and acidified then centrifuged. Soil is shaken with an ammonium hydroxide solution, and an aliquot is acidified then centrifuged. For water analysis, an aliquot is acidified. Crop and soil extracts, and water, are cleaned up using reverse-phase high-performance liquid chromatography (RPHPLC) with mesotrione and MNBA isolated using a fraction collector. During this clean up, AMBA is determined in soil and water samples using fluorescence detection. The collected mesotrione and MNBA fractions are converted into AMBA via oxidation followed by reduction in the case of mesotrione, or by reduction alone in the case of MNBA. Both fractions are analyzed by RPHPLC with fluorescence detection using an AMBA external reference standard. The method was tested on corn grain, fodder, and forage, as well as on sugar cane. The limits of quantitation (LOQ) for each analyte are 0.01 mg/kg for crops, 0.005 mg/kg for soil, and 0.10 microg/L for water. Method fortification recoveries from all crop commodities averaged 79% (CV = 7%, n = 37 and 82% (CV = 5%, n = 37) for mesotrione and MNBA, respectively. Soil was fortified at 0.005 and 0.05 mg/kg. Recoveries were 79% (CV = 4%, n = 12), 96% (CV = 2%, n = 12), and 89% (CV = 2%, n = 12) for mesotrione, MNBA, and AMBA, respectively. Groundwater, drinking water, seawater, and river water were fortified at 0.1 and 1.0 microg/L. Recoveries for all waters were 80% (CV = 7%, n = 51

  19. Detection of volatile metabolites of moulds isolated from a contaminated library.

    PubMed

    Micheluz, Anna; Manente, Sabrina; Rovea, Manuela; Slanzi, Debora; Varese, Giovanna Cristina; Ravagnan, Giampietro; Formenton, Gianmaria

    2016-09-01

    The principal fungal species isolated from a contaminated library environment were tested for their microbial volatile organic compound (MVOC) production ability. Aspergillus creber, A. penicillioides, Cladosporium cladosporioides, Eurotium chevalieri, E. halophilicum, Penicillium brevicompactum and P. chrysogenum were cultivated on suitable culture media inside sample bottles specifically designed and created for direct MVOC injection to a GC-MS instrument. The fungal emissions were monitored over several weeks to detect changes with the aging of the colonies, monitored also by respirometric tests. A total of 55 different MVOCs were detected and isopropyl alcohol, 3-methyl-1-butanol and 2-butanone were the principal compounds in common between the selected fungal species. Moreover, 2,4-dimethylheptane, 1,4-pentadiene, styrene, ethanol, 2-methyl-1-butanol, acetone, furan and 2-methylfuran were the most detected compounds. For the first time, the MVOC production for particular fungal species was detected. The species A. creber, which belongs to the recently revised group Aspergillus section Versicolores, was characterized by the production of ethanol, furan and 1,4-pentadiene. For the xerophilic fungus E. halophilicum, specific production of acetone, 2-butanone and 1,4-pentadiene was detected, supported also by respirometric data. The results demonstrated the potential use of this method for the detection of fungal contamination phenomena inside Cultural Heritage's preservation environments. PMID:27392938

  20. Prototype Systems Containing Human Cytochrome P450 for High-Throughput Real-Time Detection of DNA Damage by Compounds That Form DNA-Reactive Metabolites.

    PubMed

    Brito Palma, Bernardo; Fisher, Charles W; Rueff, José; Kranendonk, Michel

    2016-05-16

    The formation of reactive metabolites through biotransformation is the suspected cause of many adverse drug reactions. Testing for the propensity of a drug to form reactive metabolites has increasingly become an integral part of lead-optimization strategy in drug discovery. DNA reactivity is one undesirable facet of a drug or its metabolites and can lead to increased risk of cancer and reproductive toxicity. Many drugs are metabolized by cytochromes P450 in the liver and other tissues, and these reactions can generate hard electrophiles. These hard electrophilic reactive metabolites may react with DNA and may be detected in standard in vitro genotoxicity assays; however, the majority of these assays fall short due to the use of animal-derived organ extracts that inadequately represent human metabolism. The current study describes the development of bacterial systems that efficiently detect DNA-damaging electrophilic reactive metabolites generated by human P450 biotransformation. These assays use a GFP reporter system that detects DNA damage through induction of the SOS response and a GFP reporter to control for cytotoxicity. Two human CYP1A2-competent prototypes presented here have appropriate characteristics for the detection of DNA-damaging reactive metabolites in a high-throughput manner. The advantages of this approach include a short assay time (120-180 min) with real-time measurement, sensitivity to small amounts of compound, and adaptability to a microplate format. These systems are suitable for high-throughput assays and can serve as prototypes for the development of future enhanced versions. PMID:27031942

  1. Detection of the metabolites of human plasma and follicular fluid in IVF-ET with microextraction and LC-TOF-MS.

    PubMed

    Wang, Gang; Yao, Shun; Liang, Xin; Zuo, Tao; Zhu, Minghui

    2015-01-01

    The metabolites from human plasma and follicular fluid in IVF-ET are related with the metabolic activity of follicular cells and further with oocyte quality. With liquid chromatography-time-of-flight-mass spectrometry (LC-TOF-MS) and organic extraction from plasma and follicular fluid, a fast and efficient method was established for detecting the metabolites of human plasma and follicular fluid. Certain metabolites were closely related with human physiological activities, such as hormone, amino acid, estrogen and fatty acid, and estrogen fatty acyllipid lipid were successfully detected in human plasma and follicular fluid. The metabolites in follicular fluid and plasma on the day of oocyte retrieval were found to be similar. But those in the latter and basal plasma were partially different. Methyl (3α ,5β ,7α ,12α)-3,7,12-trihydroxycholan-24-oate, androst-4-ene-3,17-dione and tryptophan were identified and might be potential biomarkers on oocyte quality. It demonstrates that LC-TOF-MS combined with organic extraction could be an efficient tool to investigate the metabolites of human plasma and follicular fluid. Based on this study, it is expected to provide the basis for the further steps to explore possible influential factors on oocyte quality. PMID:26410324

  2. Standardized high-performance liquid chromatography of 182 mycotoxins and other fungal metabolites based on alkylphenone retention indices and UV-VIS spectra (diode array detection).

    PubMed

    Frisvad, J C; Thrane, U

    1987-08-28

    A general standardized method for the analysis of mycotoxins and other fungal secondary metabolites has been developed, based on high-performance liquid chromatography (HPLC) with an alkylphenone retention index and photodiode-array detection combined with thin-layer chromatography (TLC) in two different eluents. Each fungal secondary metabolite is characterized by its bracketed alkylphenone retention time index, its UV-VIS absorption maxima and its retardation factors relative to griseofulvin in two TLC eluents. This system is effective for the comparison of chemotaxonomic data in different laboratories and for a precise identification of fungi based on organic solvent extracts of fungal cultures. All important groups of mycotoxins and other fungal secondary metabolites could be detected in the HPLC system described and data are listed for 182 metabolites. The fungal secondary metabolites separated and characterized include aflatoxin B1, B2, G1 and G2, ochratoxin A, citrinin, penicillin acid, viomellein, penitrem A, patulin, sterigmatocystin, alternariol, tenuazonic acid, trichothecenes, roquefortines, fusarin C, zearalenone, PR-toxin, citreoviridin, viridicatumtoxin, verruculogen, rugulosin, cyclopiazonic acid, penicillin G and many other alkaloids, polyketides and terpenes. PMID:3680432

  3. In Vivo Detection of Perinatal Brain Metabolite Changes in a Rabbit Model of Intrauterine Growth Restriction (IUGR)

    PubMed Central

    Simões, Rui V.; Muñoz-Moreno, Emma; Carbajo, Rodrigo J.; González-Tendero, Anna; Illa, Miriam; Sanz-Cortés, Magdalena; Pineda-Lucena, Antonio; Gratacós, Eduard

    2015-01-01

    Background Intrauterine growth restriction (IUGR) is a risk factor for abnormal neurodevelopment. We studied a rabbit model of IUGR by magnetic resonance imaging (MRI) and spectroscopy (MRS), to assess in vivo brain structural and metabolic consequences, and identify potential metabolic biomarkers for clinical translation. Methods IUGR was induced in 3 pregnant rabbits at gestational day 25, by 40–50% uteroplacental vessel ligation in one horn; the contralateral horn was used as control. Fetuses were delivered at day 30 and weighted. A total of 6 controls and 5 IUGR pups underwent T2-w MRI and localized proton MRS within the first 8 hours of life, at 7T. Changes in brain tissue volumes and respective contributions to each MRS voxel were estimated by semi-automated registration of MRI images with a digital atlas of the rabbit brain. MRS data were used for: (i) absolute metabolite quantifications, using linear fitting; (ii) local temperature estimations, based on the water chemical shift; and (iii) classification, using spectral pattern analysis. Results Lower birth weight was associated with (i) smaller brain sizes, (ii) slightly lower brain temperatures, and (iii) differential metabolite profile changes in specific regions of the brain parenchyma. Specifically, we found estimated lower levels of aspartate and N-acetylaspartate (NAA) in the cerebral cortex and hippocampus (suggesting neuronal impairment), and higher glycine levels in the striatum (possible marker of brain injury). Our results also suggest that the metabolic changes in cortical regions are more prevalent than those detected in hippocampus and striatum. Conclusions IUGR was associated with brain metabolic changes in vivo, which correlate well with the neurostructural changes and neurodevelopment problems described in IUGR. Metabolic parameters could constitute non invasive biomarkers for the diagnosis and abnormal neurodevelopment of perinatal origin. PMID:26208165

  4. Photocatalytic degradation of the antiviral drug Tamiflu by UV-A/TiO2: Kinetics and mechanisms.

    PubMed

    Wang, Wen-Long; Wu, Qian-Yuan; Wang, Zheng-Ming; Hu, Hong-Ying; Negishi, Nobuaki; Torimura, Masaki

    2015-07-01

    The photocatalytic degradation of the antiviral drug Tamiflu (oseltamivir phosphate, OP) by TiO2 - P25, ST-01 and ATO was investigated in aqueous solution under ultraviolet (UV-A) irradiation. The photocatalysis of OP is well described by pseudo-first-order kinetics with r2>98.0% for all cases. The kinetic constant of P25 with 80% anatase and 20% rutile (0.040 min(-1)) is 4 and 10 times higher than that of ATO and ST-01 with 100% purity of anatase, respectively. We examined the effects of the catalyst loading and initial OP concentration on the photodegradation of OP, and used potassium iodine, isopropanol, and calcium fluorine as radical quenchers to evaluate the contributions of the hydroxyl radical (OH) and photo hole (h+) in the photodegradation. Results confirmed that 80% of the contribution came from the OH species. Although more than 95% of the OP (21 μM) was removed after 80 min of UV-A irradiation with 20 and 100 mg L(-1) P25, the removal efficiencies of total organic carbon (TOC) were only 45.6% and 67.0%, respectively, after 360 min UV-A irradiation. Based on an intermediate analysis by HPLC coupled with a triple quadrupole spectrometer and an ion trap mass spectrometer, typical intermediate species such as hydration derivatives, hydroxyl substitutes and keto-derivatives were identified and possible degradation pathways of OP by P25 were proposed. PMID:25765262

  5. Removal of oseltamivir (Tamiflu) and other selected pharmaceuticals from wastewater using a granular bioplastic formulation entrapping propagules of Phanerochaete chrysosporium.

    PubMed

    Accinelli, Cesare; Saccà, Maria Ludovica; Batisson, Isabelle; Fick, Jerker; Mencarelli, Mariangela; Grabic, Roman

    2010-09-01

    The capacity of the ligninolytic fungus Phanerochaete chrysosporium to degrade a wide variety of environmentally persistent xenobiotics has been largely reported in the literature. Beside other factors, one barrier to a wider use of this bioremediation fungus is the availability of effective formulations that ensure easy preparation, handling and application. In this series of laboratory experiments, we evaluated the efficiency of a granular bioplastic formulation entrapping propagules of P. chrysosporium for removal of four selected pharmaceuticals from wastewater samples. Addition of inoculated granules to samples of the wastewater treatment plant of Bologna significantly increased the removal of the antiviral drug oseltamivir (Tamiflu), and the antibiotics, erythromycin, sulfamethoxazol, and ciprofloxacin. Similar effects were also observed in effluent water. Oseltamivir was the most persistent of the four active substances. After 30d of incubation, approximately two times more oseltamivir was removed in bioremediated wastewater than controls. The highest removal efficiency of the bioplastic formulation was observed with the antibiotic ciprofloxacin. Microbiological DNA-based analysis showed that the bioplastic matrix supported the growth of P. chrysosporium, thus facilitating its adaptation to unusual environment such as wastewater. PMID:20673959

  6. Spectroscopic imaging and spatial localization using adiabatic pulses and applications to detect transmural metabolite distribution in the canine heart.

    PubMed

    Robitaille, P M; Merkle, H; Sublett, E; Hendrich, K; Lew, B; Path, G; From, A H; Bache, R J; Garwood, M; Uğurbil, K

    1989-04-01

    Adiabatic pulses have been employed in spectroscopic imaging and relaxation rate measurements at 4.7 T to demonstrate the feasibility of obtaining spectroscopic data from the complete sensitive volume of a surface coil using the surface coil as a transmitter and receiver. With conventional B1 sensitive pulses, spectroscopic localization or imaging techniques, such as chemical-shift imaging, yield resonance intensities that are distorted severely as a function of space, and maximal signal is detected from a small region within the complete sensitive volume of the coil. With adiabatic pulses, however, this problem is eliminated completely. In addition, a new method of spatial localization is introduced. This method, referred to as FLAX-ISIS, is a derivative of longitudinally modulated Fourier series window and ISIS approaches and utilizes adiabatic inversion and excitation pulses. The method allows construction of localized spectra for multiple regions along the surface coil axis by postacquisition data manipulation of a single set of free induction decays. These techniques were applied to the study of the myocardium using an implanted surface coil in an instrumented closed-chest canine model and in an open-chest preparation. The results demonstrate that one-dimensional techniques are adequate for transmural detection of metabolites provided signal origin is restricted to a column perpendicular to the left ventricle wall. PMID:2755331

  7. Real-time bilinear rotation decoupling in absorptive mode J-spectroscopy: Detecting low-intensity metabolite peak close to high-intensity metabolite peak with convenience.

    PubMed

    Verma, Ajay; Baishya, Bikash

    2016-05-01

    "Pure shift" NMR spectra display singlet peak per chemical site. Thus, high resolution is offered at the cost of valuable J-coupling information. In the present work, real-time BIRD (BIlinear Rotation Decoupling) is applied to the absorptive-mode 2D J-spectroscopy to provide pure shift spectrum in the direct dimension and J-coupling information in the indirect dimension. Quite often in metabolomics, proton NMR spectra from complex bio-fluids display tremendous signal overlap. Although conventional J-spectroscopy in principle overcomes this problem by separating the multiplet information from chemical shift information, however, only magnitude mode of the experiment is practical, sacrificing much of the potential high resolution that could be achieved. Few J-spectroscopy methods have been reported so far that produce high-resolution pure shift spectrum along with J-coupling information for crowded spectral regions. In the present work, high-quality J-resolved spectrum from important metabolomic mixture such as tissue extract from rat cortex is demonstrated. Many low-intensity metabolite peaks which are obscured by the broad dispersive tails from high-intensity metabolite peaks in regular magnitude mode J-spectrum can be clearly identified in real-time BIRD J-resolved spectrum. The general practice of removing such spectral overlap is tedious and time-consuming as it involves repeated sample preparation to change the pH of the tissue extract sample and subsequent spectra recording. PMID:27026651

  8. Real-time bilinear rotation decoupling in absorptive mode J-spectroscopy: Detecting low-intensity metabolite peak close to high-intensity metabolite peak with convenience

    NASA Astrophysics Data System (ADS)

    Verma, Ajay; Baishya, Bikash

    2016-05-01

    "Pure shift" NMR spectra display singlet peak per chemical site. Thus, high resolution is offered at the cost of valuable J-coupling information. In the present work, real-time BIRD (BIlinear Rotation Decoupling) is applied to the absorptive-mode 2D J-spectroscopy to provide pure shift spectrum in the direct dimension and J-coupling information in the indirect dimension. Quite often in metabolomics, proton NMR spectra from complex bio-fluids display tremendous signal overlap. Although conventional J-spectroscopy in principle overcomes this problem by separating the multiplet information from chemical shift information, however, only magnitude mode of the experiment is practical, sacrificing much of the potential high resolution that could be achieved. Few J-spectroscopy methods have been reported so far that produce high-resolution pure shift spectrum along with J-coupling information for crowded spectral regions. In the present work, high-quality J-resolved spectrum from important metabolomic mixture such as tissue extract from rat cortex is demonstrated. Many low-intensity metabolite peaks which are obscured by the broad dispersive tails from high-intensity metabolite peaks in regular magnitude mode J-spectrum can be clearly identified in real-time BIRD J-resolved spectrum. The general practice of removing such spectral overlap is tedious and time-consuming as it involves repeated sample preparation to change the pH of the tissue extract sample and subsequent spectra recording.

  9. Screening for anabolic steroids in sports: analytical strategy based on the detection of phase I and phase II intact urinary metabolites by liquid chromatography tandem mass spectrometry.

    PubMed

    Balcells, Georgina; Pozo, Oscar J; Esquivel, Argitxu; Kotronoulas, Aristotelis; Joglar, Jesús; Segura, Jordi; Ventura, Rosa

    2015-04-10

    In order to improve the detection capabilities of anabolic androgenic steroids (AAS) in sports, a liquid chromatography-tandem mass spectrometry (LC-MS/MS) screening method for the simultaneous detection of AAS phase I and phase II intact urinary metabolites (glucuronides and sulfates) was developed. A total of 36 metabolites (7 unconjugated; 19 glucuronides and 10 sulfates) corresponding to 15 of the most reported AAS were included. Analytes were extracted from urine using C18 cartridges. LC and MS conditions were studied in-depth to determine the most sensitive and selective conditions for each analyte. A selected reaction monitoring method was set up. The optimization of the experimental parameters for 13 metabolites not available as standards was performed using excretion study urines. Extraction recoveries were above 77% for all 23 validated analytes. Intra-day precision was lower than 21%, and LODs were in the range 0.25-4ng/mL for 18 of the 23 analytes. Matrix effect was evaluated using post column infusion and ranged from 92 to 147%. The method was successfully applied to excretion study urines of different exogenous AAS. The suitability of the strategy was demonstrated with methyltestosterone and stanozolol excretion study urines by achieving detection times of 22 and 21 days, respectively. The method is compliant with the World Antidoping Agency requirements for most of the studied compounds. It represents a cost-effective approach that improves the detection capabilities of AAS by increasing the sensitivity for some metabolites and by including recently described phase II long-term metabolites not detectable using the current screening strategy. PMID:25746760

  10. The detection of the urinary metabolites of 3-[(adamantan-1-yl)carbonyl]-1-pentylindole (AB-001), a novel cannabimimetic, by gas chromatography-mass spectrometry.

    PubMed

    Grigoryev, Andrej; Kavanagh, Pierce; Melnik, Aleksandra

    2012-06-01

    3-[(Adamantan-1-yl)carbonyl]-1-pentylindole (AB-001), a synthetic cannabimimetic, was identified in head shop products in Ireland in 2010. German authorities also reported it to the European Monitoring Centre for Drugs and Drug Addiction (EMCDDA) via the Early Warning System (EWS) in 2011. As indole-derived cannabimimetics, such as JWH-018, JWH-073, and JWH-250, undergo extensive metabolism, it was expected that AB-001 would behave similarly. To include it in our toxicological screening protocols, we have identified its urinary metabolites in humans following oral administration. The major metabolites were found to be adamantane mono-hydroxylated and adamantane mono-hydroxylated/N-dealkylated products. No parent compound was found in urine, and metabolites were detectable for up to 160 h following administration. PMID:22102533

  11. Pesticide Exposures to Migrant Farmworkers in Eastern NC: Detection of metabolites in farmworker urine associated with housing violations and camp characteristics

    PubMed Central

    Raymer, J. H.; Studabaker, W.; Gardner, M.; Talton, J.; Quandt, S.A.; Chen, H.; Michael, L. C.; McCombs, M.; Arcury, T.A.

    2014-01-01

    Background The purpose of this paper is to present and evaluate descriptively bivariate associations between urinary metabolites of pesticides and herbicides and migrant camp conditions, violations and personal worker behaviors at home for farmworkers who do not apply pesticides. Methods We studied 183 migrant farmworker camps in eastern North Carolina in 2010. Data and urine samples were collected from 371 men. Predictor measures included violations in six domains of housing regulations and nonviolation characteristics and personal behaviors that might impact urinary metabolites. Results Cockroaches and bathroom violations were predictive of increased exposure to pyrethroids and cyfluthrin/chlorpyrifos, respectively. Changing and storing clothing and shoes in sleeping rooms increased the number of detects for the diazinon metabolite. Conclusions Farmworkers had exposures to multiple chemicals. No single housing domain was identified as critical to mitigating housing-related exposure;specific attention should be paid to changing and storing soiled clothing in sleeping rooms, and insect infestations. PMID:24273087

  12. Determination of chloroacetanilide herbicide metabolites in water using high-performance liquid chromatography-diode array detection and high-performance liquid chromatography/mass spectrometry

    USGS Publications Warehouse

    Hostetler, K.A.; Thurman, E.M.

    2000-01-01

    Analytical methods using high-performance liquid chromatography-diode array detection (HPLC-DAD) and high-performance liquid chromatography/mass spectrometry (HPLC/MS) were developed for the analysis of the following chloroacetanilide herbicide metabolites in water: alachlor ethanesulfonic acid (ESA); alachlor oxanilic acid; acetochlor ESA; acetochlor oxanilic acid; metolachlor ESA; and metolachlor oxanilic acid. Good precision and accuracy were demonstrated for both the HPLC-DAD and HPLC/MS methods in reagent water, surface water, and ground water. The average HPLC-DAD recoveries of the chloroacetanilide herbicide metabolites from water samples spiked at 0.25, 0.5 and 2.0 ??g/l ranged from 84 to 112%, with relative standard deviations of 18% or less. The average HPLC/MS recoveries of the metabolites from water samples spiked at 0.05, 0.2 and 2.0 ??g/l ranged from 81 to 118%, with relative standard deviations of 20% or less. The limit of quantitation (LOQ) for all metabolites using the HPLC-DAD method was 0.20 ??g/l, whereas the LOQ using the HPLC/MS method was at 0.05 ??g/l. These metabolite-determination methods are valuable for acquiring information about water quality and the fate and transport of the parent chloroacetanilide herbicides in water. Copyright (C) 2000 Elsevier Science B.V.

  13. Competitive immunoassays for simultaneous detection of metabolites and proteins using micromosaic patterning.

    PubMed

    Murphy, Brian M; He, Xinya; Dandy, David; Henry, Charles S

    2008-01-15

    New high-throughput immunoassay methods for rapid point-of-care diagnostic applications represent an unmet need and current focus of numerous innovative methods. We report a new micromosaic competitive immunoassay developed for the analysis of the thyroid hormone thyroxine (T4), inflammation biomarker C-reactive protein (CRP), and the oxidative damage marker 3-nitrotyrosine (BSA-3NT) on a silicon nitride substrate. To demonstrate the versatility of the method, both direct and indirect format competitive immunoassays were developed and could be applied simultaneously for single samples. Signals from standard solutions were fit to a logistic equation, allowing simultaneous detection of T4 (7.7-257.2 nM), CRP (0.3-4.2 microg/mL), and BSA-3NT (0.03-22.3 microg/mL). Total assay time including sample introduction, washing, and fluorescence measurement was less than 45 min. Dissociation constants for affinity pairs in the system have been estimated using regression. This proof-of-concept experiment shows that both small and macromolecular biomarkers can be quantified from a single sample using the method and suggests that groups of clinically related analytes may be analyzed by competitive micromosaic immunoassay techniques. PMID:18092765

  14. An immunoassay for the detection of triclosan-O-glucuronide, a primary human urinary metabolite of triclosan.

    PubMed

    Ranganathan, Anupama; Gee, Shirley J; Hammock, Bruce D

    2015-09-01

    Triclosan-O-glucuronide (TCSG) is one of the primary urinary metabolites of the antibacterial compound triclosan or TCS that is found in many personal care products and consumer goods. We have developed a competitive, indirect heterologous ELISA for the detection of the target TCSG in urine. Such an ELISA for TCSG could be developed as a useful tool to measure this important biomarker of human exposure to TCS. Immunogens were prepared by conjugating TCSG to thyroglobulin, via heterobifunctional cross-linkers AEDP or 3-[(2-aminoethyl)dithio] propionic acid•hydrochloride and TFCS or N-[ε-trifluoroacetylcaproyloxy]succinimide ester. The coating antigen was prepared by the direct conjugation of TCSG to bovine serum albumin. Antibodies raised in rabbits 2619, 2621 (immunogen TCSG-AEDP-Thy), and 2623 (immunogen TCSG-TFCS-Thy), and the coating antigen were screened and characterized to determine their optimal concentrations. The optimized ELISA, developed with antibody 2621, gave an IC50 value of 2.85 ng/mL, with the linear range (IC20-IC80) determined to be 2.6-24.8 ng/mL. Selectivity of the assay was assessed by measuring cross-reactivity of antibody 2621 to related congeners such as the aglycone TCS, triclosan-O-sulfate, triclocarban, a polybrominated diphenyl ether derivative, and 3-phenoxybenzyl alcohol glucuronide. There was virtually no recognition by antibody 2621 to any of these cross-reactants. Graphical Abstract Urinary biomarker analysis of triclosan glucuronide. PMID:26255293

  15. Detection of the marijuana metabolite 11-nor-Delta9-tetrahydrocannabinol-9-carboxylic acid in oral fluid specimens and its contribution to positive results in screening assays.

    PubMed

    Moore, Christine; Ross, Wayne; Coulter, Cynthia; Adams, Laura; Rana, Sumandeep; Vincent, Michael; Soares, James

    2006-09-01

    The detection of the marijuana metabolite 11-nor-Delta(9)-tetrahydrocannabinol-9-carboxylic acid (THC-COOH) in oral fluid specimens is described, and its contribution to an immunoassay for the detection of cannabinoids is investigated. Oral fluid specimens, screened using an enzyme-linked immunosorbent immunoassay (ELISA), were carried forward to confirmation for both tetrahydrocannabinol (THC) and THC-COOH using gas chromatography-mass spectrometry (GC-MS). One hundred and fifty-three specimens were analyzed, of which 143 screened positive for cannabinoids. Ninety-five (66.4%) of these specimens were positive for both THC and THC-COOH; 14 (9.7%) were positive for THC-COOH only, and 27 (18.8%) were positive for THC only. The GC-MS assay for the detection of THC-COOH in oral fluid was linear to 160 pg/mL with a limit of quantitation of 2 pg/mL. The detection of the marijuana metabolite, THC-COOH, in 76.2% of oral fluid specimens screening positive for cannabinoids is reported. As a potential defense against passive exposure claims, proposed SAMHSA regulations may require the simultaneous collection of a urine sample when oral fluid samples are used. The detection of the metabolite, THC-COOH, is a significant alternative to this approach because its presence in oral fluid minimizes the argument for passive exposure to marijuana in drug testing cases. PMID:16959132

  16. Sensitive and selective detection of urinary 1-nitropyrene metabolites following administration of a single intragastric dose of diesel exhaust particles (SRM 2975) to rats.

    PubMed

    van Bekkum, Y M; van den Broek, P H; Scheepers, P T; Bos, R P

    1998-11-01

    1-Nitropyrene (1-NP) has been proposed as a marker for exposure to diesel exhaust particles (DEP). Since the extent of the actual intake of 1-NP adsorbed on DEP will be relatively low, sensitive and selective methods are needed regarding human exposure assessment. Two analytical methods are presented for the assessment of 1-NP metabolites in urine of male Sprague-Dawley rats administered a single intragastric dose of native DEP (SRM 2975, 20 mg, 35.7 microgram of 1-NP/g). Enzymatically hydrolyzed urine was extracted using Blue Rayon. The extracts were analyzed directly, using HPLC with postcolumn on-line reduction and fluorescence detection (HPLC-Flu), or were processed further for GC/MS/MS analysis. Although sensitive to several metabolites, the HPLC-Flu method lacked selectivity for quantitation of some important metabolites in rat urinary extracts, and therefore seems suitable for screening purposes only. With regard to GC/MS/MS analysis, derivatization with heptafluorobutyrylimidazole (HFBI) yielded low limits of determination for hydroxy-1-aminopyrenes, hydroxy-N-acetyl-1-aminopyrenes (converted to derivatized hydroxy-1-aminopyrenes by the reagent), and 1-aminopyrene (1.8-9.2 fmol on the column). Derivatization of hydroxy-1-nitropyrenes yielded relatively high limits of determination, and therefore, hydroxy-1-nitropyrenes were reduced to hydroxy-1-aminopyrenes prior to derivatization with HFBI. Intragastric administration of DEP to rats resulted in urinary excretion of 6-hydroxy-N-acetyl-1-aminopyrene, 8-hydroxy-N-acetyl-1-aminopyrene, 6-hydroxy-1-nitropyrene, 8-hydroxy-1-nitropyrene, and 3-hydroxy-1-nitropyrene (7, 1.2, 1.6, 0.3, and 0.5% of the dose within 12 h, respectively). 1-Nitropyrene, N-acetyl-1-aminopyrene, and 3-, 6-, and 8-hydroxy-1-aminopyrene were not observed as urinary metabolites following administration of a single dose of DEP. The observed excretion pattern and urinary metabolite concentrations suggest that 1-NP present on unmodified DEP

  17. Development and Validation of an HPLC Method for Determination of Amifostine and/or Its Metabolite (WR-1065) In Human Plasma Using OPA Derivatization and UV Detection.

    PubMed

    Samiei, Nasim; Foroutan, Seyed Mohsen; Shafaati, Alireza; Zarghi, Afshin

    2015-01-01

    A rapid, sensitive and reproducible HPLC method was developed and validated for the analysis of amifostine (AMF) and/or its metabolite, WR-1065 in human plasma. The method involves the alkylation of free sulfydryl group with iodoacetic acid followed by derivatization of the drug and its metabolite with o-phthaldialdehyde (OPA) and UVdetection at 340 nm. The derivatized AMF and WR-1065 were eluted in less than 11 min, and in the case of the metabolite with no interferences from the endogenous plasma peaks. Cystein was used as the internal standard. Analysis was carried out on a Eurosphere Performance (RP-18e, 100 × 4.6 mm) analytical column. The mobile phase was a mixture of methanol and phosphate buffer 0.03 M pH = 2.7 at a ratio of 40: 60v/v, respectively, with a flow rate of 1.5 mLmin(-1). Limit of detection was 0.5 µgmL(-1). The method involved a simple extraction procedure for AMF and/or its metabolite and analytical recovery was 90 ± 0.9%.The calibration curve was linear over the concentration range of 1-200 µgmL(-1). The coefficients of variation for intra-day and inter-day assays were less than 10%. PMID:26664371

  18. Development and Validation of an HPLC Method for Determination of Amifostine and/or Its Metabolite (WR-1065) In Human Plasma Using OPA Derivatization and UV Detection

    PubMed Central

    Samiei, Nasim; Foroutan, Seyed Mohsen; Shafaati, Alireza; Zarghi, Afshin

    2015-01-01

    A rapid, sensitive and reproducible HPLC method was developed and validated for the analysis of amifostine (AMF) and/or its metabolite, WR-1065 in human plasma. The method involves the alkylation of free sulfydryl group with iodoacetic acid followed by derivatization of the drug and its metabolite with o-phthaldialdehyde (OPA) and UVdetection at 340 nm. The derivatized AMF and WR-1065 were eluted in less than 11 min, and in the case of the metabolite with no interferences from the endogenous plasma peaks. Cystein was used as the internal standard. Analysis was carried out on a Eurosphere Performance (RP-18e, 100 × 4.6 mm) analytical column. The mobile phase was a mixture of methanol and phosphate buffer 0.03 M pH = 2.7 at a ratio of 40: 60v/v, respectively, with a flow rate of 1.5 mLmin-1. Limit of detection was 0.5 µgmL-1. The method involved a simple extraction procedure for AMF and/or its metabolite and analytical recovery was 90 ± 0.9%.The calibration curve was linear over the concentration range of 1-200 µgmL-1. The coefficients of variation for intra-day and inter-day assays were less than 10%. PMID:26664371

  19. Enantioselective determination of metoprolol and its metabolites in human urine high-performance liquid chromatography with fluorescence detection (HPLC-FLD) and tandem mass spectrometry (MS/MS).

    PubMed

    Baranowska, Irena; Adolf, Weronika; Magiera, Sylwia

    2015-11-01

    A sensitive, stereoselective assay using solid phase extraction and high-performance liquid chromatography (HPLC) with fluorescence detection (FLD) was developed and validated for the analysis of enantiomers of metoprolol and its metabolites (α-hydroxymetoprolol, O-desmethylmetoprolol). Chiral separation was achieved using a CHIRALCEL OD-RH column, packed with cellulose tris-(3,5-dimethylphenyl-carbamate) stationary phase, employing a mobile phase composed by a mixture of 0.2% diethylamine in water and acetonitrile in gradient elution mode. Linear calibration curves were obtained over the range of 0.025-2.0μg/mL (R(2)>0.994) in urine for both enantiomers of metoprolol and its metabolites with quantitation limit of 0.025μg/mL. Intra and inter-day precision and accuracy were below 15% for both metoprolol and metabolites enantiomers. The recovery of enantiomer of metoprolol and its metabolite was greater than 68.0%, utilizing a SPE procedure. The method was tested with urine quality control samples and human urine fractions after administration of 50mg rac-metoprolol. PMID:26451466

  20. Rapid determination of letrozole, citalopram and their metabolites by high performance liquid chromatography-fluorescence detection in urine: Method validation and application to real samples.

    PubMed

    Rodríguez, J; Castañeda, G; Muñoz, L

    2013-01-15

    This work reports the validation of a high precision and accuracy method for the simultaneous determination of letrozole, citalopram and their metabolites in urine by high performance liquid chromatography with fluorescence detection. Dilution (urine:mobile phase, 1:2, v/v) was the only sample preparation step. The separation was carried out in a Kromasil C(18) (150mm×4.6mm) column, and the mobile phase was phosphate buffer 80mM (pH 3.0) and acetonitrile (65:35, v/v) at a flow rate of 1.0mL/min. The analytes were detected at 295nm after excitation at 230nm. Linearity was observed in the range of 1.0-1000ng/mL for letrozole and its metabolite and 2.5-1000ng/mL for citalopram and their metabolites, with limits of detection and quantification between 0.09-1.0 and 0.27-1.65ng/mL, respectively. The precisions were satisfactory with RSDs between 0.17 and 5.71%. The accuracy was studied by spiking three urines from healthy female volunteers, and the recoveries were from 85 to 103%. The method was applied to urine samples from women under treatment for breast cancer and depression diseases. PMID:23262245

  1. Monitoring in Situ Anaerobic Alkylbenzene Biodegradation Based on Mass Spectrometric Detection of Unique Metabolites or Real-Time PCR Detection of a Catabolic Gene

    NASA Astrophysics Data System (ADS)

    Beller, H. R.; Kane, S. R.

    2002-12-01

    Monitored natural attenuation (MNA) can be a cost-effective and viable approach for remediation of hydrocarbon-contaminated groundwater. However, regulatory acceptance of the approach is often contingent on monitoring that can convincingly demonstrate the role of microbial degradation. Recent advances in anaerobic hydrocarbon biochemistry, analytical chemistry, and molecular biology have fostered the development of powerful new techniques that can be applied to MNA of BTEX (benzene, toluene, ethylbenzene, and xylenes). Here we report two independent methods that have been developed to monitor in situ, anaerobic biodegradation of toluene and xylenes. A method has been developed for rapid, sensitive, and highly selective detection of distinctive indicators of anaerobic alkylbenzene metabolism. The target metabolites, benzylsuccinic acid (BS) and methylbenzylsuccinic acid (MeBS) isomers, have no known sources other than anaerobic toluene or xylene degradation; thus, their mere presence in groundwater provides definitive evidence of in situ metabolism. The method, which involves small sample size (<1 mL) and no extraction/concentration steps, relies on isotope dilution liquid chromatography/tandem mass spectrometry (LC/MS/MS) with selected reaction monitoring. Detection limits for benzylsuccinates were determined to be ca. 0.3 μg/L and accuracy and precision were favorable in a groundwater matrix. The LC/MS/MS method was used to characterize geographic and temporal distributions of benzylsuccinates in an anaerobic, hydrocarbon-contaminated aquifer. BS was never detected and MeBS isomers were detected in the three wells with the highest concentrations of BTEX; MeBS concentrations ranged from <0.3 to 205 μg/L. A strong linear correlation was found between concentrations of total MeBS isomers and their parent compounds, xylenes. A monitoring method based on real-time Polymerase Chain Reaction (PCR) analysis has been developed to specifically quantify populations of

  2. Extraction of nitrofurantoin and its toxic metabolite from urine by supercritical fluids. Quantitation by high performance liquid chromatography with UV detection.

    PubMed

    Arancibia, Verónica; Valderrama, Mauricio; Madariaga, Andrea; Zúñiga, Marı́a Carolina; Segura, Rodrigo

    2003-11-01

    A procedure is described for the determination of nitrofurantoin and its toxic metabolite in urine from patients with urinary infection using supercritical fluid extraction (SFE) and liquid chromatography. The standard solution of toxic metabolite (radical anion) was obtained by electrochemical reduction of nitrofurantoin in an aprotic medium and chemical reoxidation with oxygen. In our initial SFE studies to find the adequate extraction parameters, drug solutions were impregnated onto filter paper. Quantitative extractions were achieved when the experiments were carried out under 2500 psi of pressure at a temperature of 80 degrees C (oven and restrictor) after 20 min of static extraction and 5 min of dynamic extraction. The modifier used was acetonitrile (2.0 ml in a 10 ml extraction column). Nitrofurantoin and its toxic metabolite were detected in urine samples. Both compounds were quantified in the extracts by high performance liquid chromatography (HPLC) with detection at 310 nm. The calibration graph of these compounds in acetonitrile was linear between 10.9 and 378.0 muM (R=0.9995) for nitrofurantoin and between 3.0x10(-3) and 21.0 muM (R=0.9992) for the metabolite. The detection limits (LOD) were 12.1 and 0.9 muM, respectively. The drug was administered to two patients during 7 days, and all the urine eliminated between 1 day before and 2 days after administration was analyzed. One patient consumed the drug in the form of microcrystals and the other as macrocrystals. PMID:18969197

  3. 'Dilute-and-shoot' RSLC-MS-MS method for fast detection of nerve and vesicant chemical warfare agent metabolites in urine.

    PubMed

    Rodin, Igor; Braun, Arcady; Stavrianidi, Andrey; Baygildiev, Timur; Shpigun, Oleg; Oreshkin, Dmitry; Rybalchenko, Igor

    2015-01-01

    A sensitive screening method based on fast liquid chromatography tandem mass-spectrometry (RSLC-MS-MS) has shown the feasibility of separation and detection of low concentration β-lyase metabolites of sulfur mustard and of nerve agent phosphonic acids in urine. The analysis of these compounds is of interest because they are specific metabolites of the chemical warfare agents (CWAs), sulfur mustard (HD), sarin (GB), soman (GD), VX and Russian VX (RVX). The 'dilute-and-shoot' RSLC-MS-MS method provides a sensitive and direct approach for determining CWA exposure in non-extracted non-derivatized samples from urine. Chromatographic separation of the metabolites was achieved using a reverse phase column with gradient mobile phases consisting of 0.5% formic acid in water and acetonitrile. Identification and quantification of species were achieved using electrospray ionization-tandem mass-spectrometry monitoring two precursor-to-product ion transitions for each compound. The method demonstrates linearity over at least two orders of magnitude and had detection limits of 0.5 ng/mL in urine. PMID:25326204

  4. Comparison of Two Automated Solid Phase Extractions for the Detection of Ten Fentanyl Analogs and Metabolites in Human Urine Using Liquid Chromatography Tandem Mass Spectrometry

    PubMed Central

    Shaner, Rebecca L.; Kaplan, Pearl; Hamelin, Elizabeth I.; Bragg, William; Johnson, Rudolph C.

    2016-01-01

    Two types of automated solid phase extraction (SPE) were assessed for the determination of human exposure to fentanyls in urine. High sensitivity is required to detect these compounds following exposure because of the low dose required for therapeutic effect and the rapid clearance from the body for these compounds. To achieve this sensitivity, two acceptable methods for the detection of human exposure to seven fentanyl analogs and three metabolites were developed using either off-line 96-well plate SPE or on-line SPE. Each system offers different advantages: off-line 96-well plate SPE allows for high throughput analysis of many samples, which is needed for large sample numbers, while on-line SPE removes almost all analyst manipulation of the samples, minimizing the analyst time needed for sample preparation. Both sample preparations were coupled with reversed phase liquid chromatography and isotope dilution tandem mass spectrometry (LC-MS/MS) for analyte detection. For both methods, the resulting precision was within 15%, the accuracy within 25%, and the sensitivity was comparable with the limits of detection ranging from 0.002-0.041ng/mL. Additionally, matrix effects were substantially decreased from previous reports for both extraction protocols. The results of this comparison showed that both methods were acceptable for the detection of exposures to fentanyl analogs and metabolites in urine. PMID:24893271

  5. Combined derivatization and high-performance liquid chromatography with fluorescence and ultraviolet detection for simultaneous analysis of octreotide and gabexate mesylate metabolite in human pancreatic juice samples.

    PubMed

    Carlucci, Giuseppe; Selvaggi, Federico; Sulpizio, Sara; Bassi, Claudio; Carlucci, Maura; Cotellese, Roberto; Ferrone, Vincenzo; Innocenti, Paolo; Locatelli, Marcello

    2015-06-01

    A simple and sensitive method based on the combination of derivatization and high-performance liquid chromatography with ultraviolet and fluorimetric detection was developed for the simultaneous determination of octreotide and gabexate mesylate metabolite in human pancreatic juice samples. Parameters of the derivatization procedure affecting extraction efficiency were optimized. The developed method was validated according to the International Conference on Harmonization guidelines. The calibration curves were linear over a range of 0.1-15 µg/mL for octreotide and 0.20-15 µg/mL for gabexate mesylate metabolite. Derivatized products of octreotide and gabexate mesylate metabolite were separated on a Luna C18 column (4.6 × 250 mm; 5 µm particle size) using a gradient with a run time of 36 min, without further purification. The limits of detection were 0.025 and 0.05, respectively, for octreotide and gabexate mesylate metabolite. This paper reports the validation of a quantitative high performance liquid chromatography-photodiode array-fluorescence (HPLC-PDA-FL) method for the simultaneous analysis of octreotide and gabexate mesylate metabolite in pancreatic juice by protein precipitation using zinc sulfate-methanol-acetonitrile containing the derivatizing reagent, 4-fluoro-7-nitro-[2,1,3]-benzoxadiazole (NBD-F). Derivatized products of octreotide and gabexate mesylate metabolite were separated on a Luna C18 column (4.6 × 250 mm; 5 µm particle size) using a gradient with a run time of 36 min, without further purification. The method was validated over the concentration ranges 0.1-15 and 0.2-15 µg/mL for octreotide and gabexate mesylate metabolite, respectively, in human pancreatic juice. Biphalin and methyl-p-hydroxybenzoate were used as the internal standards. This method was successfully utilized to support clinical studies in humans. The results from assay validations show that the method is selective, sensitive and robust. The limit

  6. A high-resolution 2D J-resolved NMR detection technique for metabolite analyses of biological samples

    PubMed Central

    Huang, Yuqing; Zhang, Zhiyong; Chen, Hao; Feng, Jianghua; Cai, Shuhui; Chen, Zhong

    2015-01-01

    NMR spectroscopy is a commonly used technique for metabolite analyses. Due to the observed macroscopic magnetic susceptibility in biological tissues, current NMR acquisitions in measurements of biological tissues are generally performed on tissue extracts using liquid NMR or on tissues using magic-angle spinning techniques. In this study, we propose an NMR method to achieve high-resolution J-resolved information for metabolite analyses directly from intact biological samples. A dramatic improvement in spectral resolution is evident in our contrastive demonstrations on a sample of pig brain tissue. Metabolite analyses for a postmortem fish from fresh to decayed statuses are presented to further reveal the capability of the proposed method. This method is a previously-unreported high-resolution 2D J-resolved spectroscopy for biological applications without specialised hardware requirements or complicated sample pretreatments. It provides a significant contribution to metabolite analyses of biological samples, and may be potentially applicable to in vivo samples. Furthermore, this method also can be applied to measurements of semisolid and viscous samples. PMID:25670027

  7. A high-resolution 2D J-resolved NMR detection technique for metabolite analyses of biological samples.

    PubMed

    Huang, Yuqing; Zhang, Zhiyong; Chen, Hao; Feng, Jianghua; Cai, Shuhui; Chen, Zhong

    2015-01-01

    NMR spectroscopy is a commonly used technique for metabolite analyses. Due to the observed macroscopic magnetic susceptibility in biological tissues, current NMR acquisitions in measurements of biological tissues are generally performed on tissue extracts using liquid NMR or on tissues using magic-angle spinning techniques. In this study, we propose an NMR method to achieve high-resolution J-resolved information for metabolite analyses directly from intact biological samples. A dramatic improvement in spectral resolution is evident in our contrastive demonstrations on a sample of pig brain tissue. Metabolite analyses for a postmortem fish from fresh to decayed statuses are presented to further reveal the capability of the proposed method. This method is a previously-unreported high-resolution 2D J-resolved spectroscopy for biological applications without specialised hardware requirements or complicated sample pretreatments. It provides a significant contribution to metabolite analyses of biological samples, and may be potentially applicable to in vivo samples. Furthermore, this method also can be applied to measurements of semisolid and viscous samples. PMID:25670027

  8. The Tamiflu saga continues: will our conduct change after the publication of the latest systematic review on benefits and harms of oseltamivir?

    PubMed

    Bachelet, Vivienne C

    2014-01-01

    In 2013, we wrote about the harm, waste and deception stemming from conducts adopted by the pharmaceutical industry, by concealing raw data and Clinical Study Reports (CSRs) from the regulator’s view when requesting the marketing patent. We described the case of Tamiflu (Roche), a drug that has been widely used in our population and profusely prescribed by physicians. Health authorities, entailing a great cost for the countries in the region, have also purchased it. In this editorial, we will show how the idea of using antivirals for prophylaxis and treatment of influenza took hold, starting from the first enthusiastic recommendations up to the systematic review published last month in the BMJ. PMID:25383799

  9. Determination of a metabolite of the herbicide pyridate in drinking and groundwater using high-performance liquid chromatography with amperometric detection.

    PubMed

    Pachinger, A; Eisner, E; Begutter, H; Klus, H

    1991-10-11

    The main metabolite of the herbicide pyridate is 3-phenyl-4-hydroxy-6-chlorpyridazine (CL9673). A high-performance liquid chromatographic method with amperometric detection is described for determining CL9673 at residue levels in water samples. Sample preconcentration is performed by passage through a C18 extraction cartridge. A recovery study using tap water samples spiked with CL9673 at a concentration of 0.1 micrograms/l showed a recovery of 84.8% (coefficient of variation 6.2%). The method is suitable for the determination of CL9673 in drinking and groundwater. PMID:1797812

  10. Therapeutic targeting of Neu1 sialidase with oseltamivir phosphate (Tamiflu®) disables cancer cell survival in human pancreatic cancer with acquired chemoresistance

    PubMed Central

    O’Shea, Leah K; Abdulkhalek, Samar; Allison, Stephanie; Neufeld, Ronald J; Szewczuk, Myron R

    2014-01-01

    Background Resistance to drug therapy, along with high rates of metastasis, contributes to the low survival rate in patients diagnosed with pancreatic cancer. An alternate treatment for human pancreatic cancer involving targeting of Neu1 sialidase with oseltamivir phosphate (Tamiflu®) was investigated in human pancreatic cancer (PANC1) cells with acquired resistance to cisplatin and gemcitabine. Its efficacy in overcoming the intrinsic resistance of the cell to chemotherapeutics and metastasis was evaluated. Methods Microscopic imaging, immunocytochemistry, immunohistochemistry, and WST-1 cell viability assays were used to evaluate cell survival, morphologic changes, and expression levels of E-cadherin, N-cadherin, and VE-cadherin before and after treatment with oseltamivir phosphate in PANC1 cells with established resistance to cisplatin, gemcitabine, or a combination of the two agents, and in archived paraffin-embedded PANC1 tumors grown in RAGxCγ double mutant mice. Results Oseltamivir phosphate overcame the chemoresistance of PANC1 to cisplatin and gemcitabine alone or in combination in a dose-dependent manner, and disabled the cancer cell survival mechanism(s). Oseltamivir phosphate also reversed the epithelial-mesenchymal transition characteristic of the phenotypic E-cadherin to N-cadherin changes associated with resistance to drug therapy. Low-dose oseltamivir phosphate alone or in combination with gemcitabine in heterotopic xenografts of PANC1 tumors growing in RAGxCγ double mutant mice did not prevent metastatic spread to the liver and lung. Conclusion Therapeutic targeting of Neu1 sialidase with oseltamivir phosphate at the growth factor receptor level disables the intrinsic signaling platform for cancer cell survival in human pancreatic cancer with acquired chemoresistance. These findings provide evidence for oseltamivir phosphate (Tamiflu) as a potential therapeutic agent for pancreatic cancer resistant to drug therapy. PMID:24470763

  11. Rapid and sensitive detection of fipronil and its metabolites in edible oils by solid-phase extraction based on humic acid bonded silica combined with gas chromatography with electron capture detection.

    PubMed

    Peng, Xi-Tian; Li, Yu-Nan; Xia, Hong; Peng, Li-Jun; Feng, Yu-Qi

    2016-06-01

    Solid-phase extraction based on humic acid bonded silica followed by gas chromatography with electron capture detection was developed to determine fipronil and its metabolites in edible oil. To achieve the best extraction performance, we systematically investigated a series of solid-phase extraction parameters. Under the optimized conditions, the method was validated according to linearity, recovery, and precision. Good linearities were obtained with R(2) more than 0.9996 for all analytes. The limits of detection were between 0.3 and 0.5 ng/g, and the recoveries ranged from 83.1 to 104.0% at three spiked concentrations with intra- and interday relative standard deviation values less than 8.7%. Finally, the proposed method was applied to determine fipronil and its metabolites in 11 edible oil samples taken from Wuhan markets. Fipronil was detectable in four samples with concentrations ranging from 3.0 to 5.2 ng/g. In China, the maximum residue limits of fipronil in some vegetables and maize are 20 and 100 ng/g (GB/T 2763-2014), respectively. The residues of fipronil and its metabolites in commercial edible oils might exhibit some potential threat to human health as a result of high consumption of edible oil as part of daily intake. PMID:27280701

  12. Multi-residue enantiomeric analysis of human and veterinary pharmaceuticals and their metabolites in environmental samples by chiral liquid chromatography coupled with tandem mass spectrometry detection.

    PubMed

    Camacho-Muñoz, Dolores; Kasprzyk-Hordern, Barbara

    2015-12-01

    Enantiomeric profiling of chiral pharmacologically active compounds (PACs) in the environment has hardly been investigated. This manuscript describes, for the first time, a multi-residue enantioselective method for the analysis of human and veterinary chiral PACs and their main metabolites from different therapeutic groups in complex environmental samples such as wastewater and river water. Several analytes targeted in this paper have not been analysed in the environment at enantiomeric level before. These are aminorex, carboxyibuprofen, carprofen, cephalexin, 3-N-dechloroethylifosfamide, 10,11-dihydro-10-hydroxycarbamazepine, dihydroketoprofen, fenoprofen, fexofenadine, flurbiprofen, 2-hydroxyibuprofen, ifosfamide, indoprofen, mandelic acid, 2-phenylpropionic acid, praziquantel and tetramisole. The method is based on chiral liquid chromatography utilising a chiral α1-acid glycoprotein column and tandem mass spectrometry detection. Excellent chromatographic separation of enantiomers (Rs≥1.0) was achieved for chloramphenicol, fexofenadine, ifosfamide, naproxen, tetramisole, ibuprofen and their metabolites: aminorex and dihydroketoprofen (three of four enantiomers), and partial separation (Rs = 0.7-1.0) was achieved for ketoprofen, praziquantel and the following metabolites: 3-N-dechloroethylifosfamide and 10,11-dihydro-10-hydroxycarbamazepine. The overall performance of the method was satisfactory for most of the compounds targeted. Method detection limits were at low nanogram per litre for surface water and effluent wastewater. Method intra-day precision was on average under 20% and sample pre-concentration using solid phase extraction yielded recoveries >70% for most of the analytes. This novel, selective and sensitive method has been applied for the quantification of chiral PACs in surface water and effluent wastewater providing excellent enantioresolution of multicomponent mixtures in complex environmental samples. It will help with better understanding

  13. Identification of metabolites from liquid chromatography-coulometric array detection profiling: gas chromatography-mass spectrometry and refractionation provide essential information orthogonal to LC-MS/microNMR.

    PubMed

    Gathungu, Rose M; Bird, Susan S; Sheldon, Diane P; Kautz, Roger; Vouros, Paul; Matson, Wayne R; Kristal, Bruce S

    2014-06-01

    Liquid chromatography-coulometric array detection (LC-EC) is a sensitive, quantitative, and robust metabolomics profiling tool that complements the commonly used mass spectrometry (MS) and nuclear magnetic resonance (NMR)-based approaches. However, LC-EC provides little structural information. We recently demonstrated a workflow for the structural characterization of metabolites detected by LC-EC profiling combined with LC-electrospray ionization (ESI)-MS and microNMR. This methodology is now extended to include (i) gas chromatography (GC)-electron ionization (EI)-MS analysis to fill structural gaps left by LC-ESI-MS and NMR and (ii) secondary fractionation of LC-collected fractions containing multiple coeluting analytes. GC-EI-MS spectra have more informative fragment ions that are reproducible for database searches. Secondary fractionation provides enhanced metabolite characterization by reducing spectral overlap in NMR and ion suppression in LC-ESI-MS. The need for these additional methods in the analysis of the broad chemical classes and concentration ranges found in plasma is illustrated with discussion of four specific examples: (i) characterization of compounds for which one or more of the detectors is insensitive (e.g., positional isomers in LC-MS, the direct detection of carboxylic groups and sulfonic groups in (1)H NMR, or nonvolatile species in GC-MS), (ii) detection of labile compounds, (iii) resolution of closely eluting and/or coeluting compounds, and (iv) the capability to harness structural similarities common in many biologically related, LC-EC-detectable compounds. PMID:24657819

  14. Simultaneous analysis of acetaminophen, p-aminophenol and aspirin metabolites by hydrophilic interaction and strong anion exchange capillary liquid chromatography coupled to amperometric detection.

    PubMed

    Zheng, Minmin; Wu, Yimin; Lu, Lanxiang; Ding, Kang; Tang, Fengxiang; Lin, Zian; Wu, Xiaoping

    2011-08-01

    A simple and sensitive method has been developed for the simultaneous determination of polar nonsteroidal pharmaceuticals and metabolites, including acetaminophen, p-aminophenol and several aspirin metabolites (salicylic acid, gentisic acid, salicyluric acid and 2,3-dihydroxybenzoic acid), by capillary liquid chromatography with amperometric detection. Using a capillary monolithic column with mixed mode stationary phases and a mobile phase composed of acetonitrile and Tris buffer, rapid separation of six polar analytes was achieved within 8 min, and a hydrophilic interaction and strong anion exchange separation mechanism were exhibited. Method detection limits of six analytes ranged from 10 to 50 ng/mL. In terms of precision, the intra- and interday relative standard deviation values in all analytes never exceeded 3.1% for migration time and 8.9% for peak areas, respectively. This method provided a simple, rapid and cost-effective approach for the analysis of polar pharmaceuticals. The applicability of the method in pharmacokinetics was verified by spiking human serum samples with the compounds and analyzing the recoveries. PMID:21644253

  15. Determination of herbicides and its metabolite in soil and water samples by capillary electrophoresis-laser induced fluorescence detection using microwave-assisted derivatization.

    PubMed

    Cao, Liwei; Deng, Tao; Liang, Siliu; Tan, Xiaofang; Meng, Jianxin

    2014-01-01

    Methods were developed to determine glufosinate (GLUF), glyphosate (GLYP) and its metabolite, aminomethylphosphonic acid (AMPA) by capillary electrophoresis-laser induced fluorescence detection using 5-(4,6-dichlorotriazinylamino) fluorescein (DTAF) and fluorescein isothiocyanate (FITC) as the derivatizing reagents. To accelerate the labeling speed, a microwave-assisted derivatization method was adopted. The derivatizing reaction time was reduced to 180 and 150 s for DTAF and FITC, whose reaction time for conventional labeling was 50 min and 5 h, respectively. The optimum separation conditions for derivatives were as follows: a back ground electrolyte (BGE) of 30 mmol L(-1) sodium tetraborate containing 15 mmol L(-1) brij-35, hydrodynamic injection 15 s and a 10 kV separation voltage. Under these conditions, the LODs (S/N = 3) for DTAF derivatives were 0.32, 0.19 and 0.15 nmol L(-1) for GLUF, GLYP, and AMPA, respectively. The LODs (S/N = 3) for FITC derivatives were 2.60, 3.88 and 2.42 nmol L(-1) for GLUF, GLYP, and AMPA, respectively. The applicability of the developed method was demonstrated by the detection of the above herbicides and metabolite in water and soil samples. PMID:25007936

  16. Analytical Methodologies for Detection of Gamma-Valerolactone, Delta-Valerolactone, Acephate and Azinphos Methyl and Their Associated Metabolites in Complex Biological Matrices

    SciTech Connect

    Zink, E.; Clark, R.; Grant, K.; Campbell, J.; Hoppe, E.

    2005-01-01

    Non-invasive biomonitoring for chemicals of interest in law enforcement and similar monitoring of pesticides, together with their metabolites, can not only save money but can lead to faster medical attention for individuals exposed to these chemicals. This study describes methods developed for the analysis of gamma-valerolactone (GVL), delta-valerolactone (DVL), acephate, and azinphos methyl in saliva and serum. Liquid chromatography/mass spectrometry (LC/MS) operated in the negative and positive ion mode and gas chromatography/mass spectrometry (GC/MS) were used to analyze GVL and DVL. Although both analytical techniques worked well, lower detection limits were obtained with GC/MS. The lactones and their corresponding sodium salts were spiked into both saliva and serum. The lactones were isolated from saliva or serum using newly developed extraction techniques and then subsequently analyzed using GC/MS. The sodium salts of the lactones are nonvolatile and require derivatization prior to analysis by this method. N-methyl-N-(t-butyldimethylsilyl)-trifluoroacetamide (MTBSTFA) was ultimately selected as the reagent for derivatization because the acidic conditions required for reactions with diazomethane caused the salts to undergo intramolecular cyclization to the corresponding lactones. In vitro studies were conducted using rat liver microsomes to determine other metabolites associated with these compounds. Azinphos methyl and acephate are classified as organophosphate pesticides, and are known to be cholinesterase inhibitors in humans and insects, causing neurotoxicity. For this reason they have both exposure and environmental impact implications. These compounds were spiked into serum and saliva and prepared for analysis by GC/MS. Continuation of this research would include analysis by GC/MS under positive ion mode to determine the parent ions of the unknown metabolites. Further research is planned through an in vivo analysis of the lactones and pesticides. These

  17. Analytical Methodologies for Detection of Gamma-valerolactone, Delta-valerolactone, Acephate, and Azinphos Methyl and their Associated Metabolites in Complex Biological Matrices

    SciTech Connect

    Zink, Erika M.; Clark, Ryan J.; Grant, Karen E.; Campbell, James A.; Hoppe, Eric W.

    2005-01-01

    Non-invasive biomonitoring for chemicals of interest in law enforcement and similar monitoring of pesticides together with their metabolites can not only save money but can lead to faster medical attention for individuals exposed to these chemicals. This study describes methods developed for the analysis of gamma-valerolactone (GVL), delta-valerolactone (DVL), acephate, and azinphos methyl in saliva and serum. Liquid chromatography/mass spectrometry (LC/MS) operated in the negative ion mode and in the positive ion mode and gas chromatography/mass spectrometry (GC/MS) were used to analyze GVL and DVL. Although both analytical techniques worked well, lower detection limits were obtained with GC/MS. The lactones and their corresponding sodium salts were spiked into both saliva and serum. The lactones were isolated from saliva or serum using newly developed extraction techniques and then subsequently analyzed using GC/MS. The sodium salts of the lactones are nonvolatile and require derivatization prior to analysis by this method. N-methyl-N-(t-butyldimethylsilyl)-trifluoroacetamide (MTBSTFA) was ultimately selected as the reagent for derivatization because the acidic conditions required for reactions with diazomethane caused the salts to undergo intramolecular cyclization to the corresponding lactones. In vitro studies were conducted using rat liver microsomes to determine other metabolites associated with these compounds. Azinphos methyl and acephate are classified as organophosphate pesticides, and are known to be cholinesterase inhibitors in humans and insects, causing neurotoxicity. For this reason they have both exposure and environmental impact implications. These compounds were spiked into serum and saliva and prepared for analysis by GC/MS. Continuation of this research would include analysis by GC/MS under positive ion mode to determine the parent ions of the unknown metabolites. Further research is planned through an in vivo analysis of the lactones and

  18. A sensitive and rapid ultra HPLC-MS/MS method for the simultaneous detection of clopidogrel and its derivatized active thiol metabolite in human plasma.

    PubMed

    Peer, Cody J; Spencer, Shawn D; VanDenBerg, Dustin A H; Pacanowski, Michael A; Horenstein, Richard B; Figg, William D

    2012-01-01

    A sensitive, selective, and rapid ultra-high performance liquid chromatography-tandem mass spectrometry (uHPLC-MS/MS) was developed for the simultaneous quantification of clopidogrel (Plavix(®)) and its derivatized active metabolite (CAMD) in human plasma. Derivatization of the active metabolite in blood with 2-bromo-3'-methoxy acetophenone (MPB) immediately after collection ensured metabolite stability during sample handling and storage. Following addition of ticlopidine as an internal standard and simple protein precipitation, the analytes were separated on a Waters Acquity UPLC™ sub-2 μm-C(18) column via gradient elution before detection on a triple-quadrupole MS with multiple-reaction-monitoring via electrospray ionization. The method was validated across the clinically relevant concentration range of 0.01-50 ng/mL for parent clopidogrel and 0.1-150 ng/mL (r(2)=0.99) for CAMD, with a fast run time of 1.5 min to support pharmacokinetic studies using 75, 150, or 300 mg oral doses of clopidogrel. The analytical method measured concentrations of clopidogrel and CAMD with accuracy (%DEV) <±12% and precision (%CV) of <±6%. The method was successfully applied to measure the plasma concentrations of clopidogrel and CAMD in three subjects administered single oral doses of 75, 150, and 300 mg clopidogrel. It was further demonstrated that the derivatizing agent (MPB) does not affect clopidogrel levels, thus from one aliquot of blood drawn clinically, this method can simultaneously quantify both clopidogrel and CAMD with sensitivity in the picogram per mL range. PMID:22169056

  19. MIDDAS-M: Motif-Independent De Novo Detection of Secondary Metabolite Gene Clusters through the Integration of Genome Sequencing and Transcriptome Data

    PubMed Central

    Umemura, Myco; Koike, Hideaki; Nagano, Nozomi; Ishii, Tomoko; Kawano, Jin; Yamane, Noriko; Kozone, Ikuko; Horimoto, Katsuhisa; Shin-ya, Kazuo; Asai, Kiyoshi; Yu, Jiujiang; Bennett, Joan W.; Machida, Masayuki

    2013-01-01

    Many bioactive natural products are produced as “secondary metabolites” by plants, bacteria, and fungi. During the middle of the 20th century, several secondary metabolites from fungi revolutionized the pharmaceutical industry, for example, penicillin, lovastatin, and cyclosporine. They are generally biosynthesized by enzymes encoded by clusters of coordinately regulated genes, and several motif-based methods have been developed to detect secondary metabolite biosynthetic (SMB) gene clusters using the sequence information of typical SMB core genes such as polyketide synthases (PKS) and non-ribosomal peptide synthetases (NRPS). However, no detection method exists for SMB gene clusters that are functional and do not include core SMB genes at present. To advance the exploration of SMB gene clusters, especially those without known core genes, we developed MIDDAS-M, a motif-independent de novo detection algorithm for SMB gene clusters. We integrated virtual gene cluster generation in an annotated genome sequence with highly sensitive scoring of the cooperative transcriptional regulation of cluster member genes. MIDDAS-M accurately predicted 38 SMB gene clusters that have been experimentally confirmed and/or predicted by other motif-based methods in 3 fungal strains. MIDDAS-M further identified a new SMB gene cluster for ustiloxin B, which was experimentally validated. Sequence analysis of the cluster genes indicated a novel mechanism for peptide biosynthesis independent of NRPS. Because it is fully computational and independent of empirical knowledge about SMB core genes, MIDDAS-M allows a large-scale, comprehensive analysis of SMB gene clusters, including those with novel biosynthetic mechanisms that do not contain any functionally characterized genes. PMID:24391870

  20. Detection of pharmaceuticals and phytochemicals together with their metabolites in hospital effluents in Japan, and their contribution to sewage treatment plant influents.

    PubMed

    Azuma, Takashi; Arima, Natsumi; Tsukada, Ai; Hirami, Satoru; Matsuoka, Rie; Moriwake, Ryogo; Ishiuchi, Hirotaka; Inoyama, Tomomi; Teranishi, Yusuke; Yamaoka, Misato; Mino, Yoshiki; Hayashi, Tetsuya; Fujita, Yoshikazu; Masada, Mikio

    2016-04-01

    The occurrence of 41 pharmaceuticals and phytochemicals (PPs) including their metabolites was surveyed in hospital effluent in an urban area of Japan. A detailed survey of sewage treatment plant (STP) influent and effluent, and river water was also conducted. Finally, mass balances with mass fluxes of the target PPs through the water flow were evaluated and the degree of contribution of hospital effluent to the environmental discharge was estimated. The results indicate that 38 compounds were detectable in hospital effluent over a wide concentration range from ng/L to μg/L, with a maximum of 92μg/L. The contributions of PPs in the hospital effluent to STP influent varied widely from <0.1% to 14.8%. Although almost all of the remaining components could be removed below 1.0ng/L at STPs by the addition of ozone treatment, a number of PPs still remained above 10ng/L in STP effluent. These findings suggest the importance of applying highly developed treatments to hospital effluents and at STPs in the future to reduce the environmental risks posed by PPs. To our knowledge, this is the first demonstration of the presence of two conjugated metabolites of acetaminophen, acetaminophen glucuronide and acetaminophen sulfate, as well as of loxoprofen and loxoprofen alcohol, in hospital effluent, STP, and river waters. PMID:26802347

  1. Selective observation of biologically important 15N-labeled metabolites in isolated rat brain and liver by 1H-detected multiple-quantum-coherence spectroscopy

    NASA Astrophysics Data System (ADS)

    Kanamori, Keiko; Ross, Brian D.; Parivar, Farhad

    Four cerebral metabolites of importance in neurotransmission, serotonin, L-tryptophan, L-glutamine, and N-acetyl- L-aspartate, and two hepatic urea-cycle intermediates, citrulline and urea, were found to be observable by 1H- 15N heteronuclear multiple-quantum-coherence (HMQC) spectroscopy in aqueous solution at physiological pH and temperature, through the protons spin-coupled to their indole, amide, or ureido nitrogen. Their 1H chemical shifts were well dispersed over a 5-10 ppm region while the 1J 15N- 1H values were 87-99 Hz. For [γ- 15N]glutamine, a 50- to 100-fold increase in sensitivity over direct 15N detection was achieved, in contrast to a 2-fold increase by the polarization-transfer method. In the isolated brain of portacaval-shunted rats, the amide protons of biologically 15N-enriched [γ- 15N]glutamine were observed in 2 min of acquisition, with suppression of proton signals from all other cerebral metabolites. In isolated liver of 15N-enriched control rats, [ 15NIurea protons were observed in 16 min. The HMQC method is likely to be effective for the in vivo study of cerebral and hepatic nitrogen metabolism.

  2. Cross validation of gas chromatography-flame photometric detection and gas chromatography-mass spectrometry methods for measuring dialkylphosphate metabolites of organophosphate pesticides in human urine.

    PubMed

    Prapamontol, Tippawan; Sutan, Kunrunya; Laoyang, Sompong; Hongsibsong, Surat; Lee, Grace; Yano, Yukiko; Hunter, Ronald Elton; Ryan, P Barry; Barr, Dana Boyd; Panuwet, Parinya

    2014-01-01

    We report two analytical methods for the measurement of dialkylphosphate (DAP) metabolites of organophosphate pesticides in human urine. These methods were independently developed/modified and implemented in two separate laboratories and cross validated. The aim was to develop simple, cost effective, and reliable methods that could use available resources and sample matrices in Thailand and the United States. While several methods already exist, we found that direct application of these methods required modification of sample preparation and chromatographic conditions to render accurate, reliable data. The problems encountered with existing methods were attributable to urinary matrix interferences, and differences in the pH of urine samples and reagents used during the extraction and derivatization processes. Thus, we provide information on key parameters that require attention during method modification and execution that affect the ruggedness of the methods. The methods presented here employ gas chromatography (GC) coupled with either flame photometric detection (FPD) or electron impact ionization-mass spectrometry (EI-MS) with isotopic dilution quantification. The limits of detection were reported from 0.10ng/mL urine to 2.5ng/mL urine (for GC-FPD), while the limits of quantification were reported from 0.25ng/mL urine to 2.5ng/mL urine (for GC-MS), for all six common DAP metabolites (i.e., dimethylphosphate, dimethylthiophosphate, dimethyldithiophosphate, diethylphosphate, diethylthiophosphate, and diethyldithiophosphate). Each method showed a relative recovery range of 94-119% (for GC-FPD) and 92-103% (for GC-MS), and relative standard deviations (RSD) of less than 20%. Cross-validation was performed on the same set of urine samples (n=46) collected from pregnant women residing in the agricultural areas of northern Thailand. The results from split sample analysis from both laboratories agreed well for each metabolite, suggesting that each method can produce

  3. High-performance liquid chromatographic method with amperometric detection employing boron-doped diamond electrode for the determination of sildenafil, vardenafil and their main metabolites in plasma.

    PubMed

    Bartošová, Zdenka; Jirovský, David; Horna, Aleš

    2011-11-01

    A simple, fast and sensitive HPLC method with electrochemical detection employing boron-doped diamond electrode (BDD) for the determination of sildenafil (Viagra™), vardenafil (Levitra™) and their main metabolites, N-desmethyl sildenafil and N-desethyl vardenafil in human plasma is presented. The assay involved drug extraction by tert-butyl methyl ether and isocratic reversed-phase liquid chromatography with amperometric detection. Complete separation of all analytes was achieved within 12 min. The mobile phase consisted of 20mM sodium dihydrogen phosphate with 40 mM sodium perchlorate/acetonitrile (70:30, v/v), pH 3.5. The electrode working potential was +1520 mV (vs. Pd/H(2)). Calibration curves were linear over the concentration range of 10-400 ng mL(-1). Phloretin was used as an internal standard. The limits of detection (LOD) and quantification (LOQ) for the studied analytes were within the range of 2-4 ng mL(-1) and 7.0-13.4 ng mL(-1), respectively. The developed method was applied to human plasma samples spiked with analytes at therapeutic concentrations. The study confirms the method's suitability for both pharmacokinetic studies and therapeutic monitoring. PMID:21943935

  4. Fast detection of choline-containing metabolites in liver using 2D 1H- 14N three-bond correlation (HN3BC) spectroscopy

    NASA Astrophysics Data System (ADS)

    Mao, Xi-an; Li, Ning; Mao, Jiezhen; Li, Qiurong; Xiao, Nan; Jiang, Bin; Jiang, Ling; Wang, Xu-xia; Liu, Maili

    2012-01-01

    Detection and quantification of total choline-containing metabolites (CCMs) in tissues by magnetic resonance spectroscopy (MRS) has received considerable attention as a biomarker of cancer. Tissue CCMs are mainly choline (Cho), phosphocholine (PCho), and glycerophosphocholine (GPCho). Because the methyl 1H resonances of tissue CCMs exhibit small chemical shift differences and overlap significantly in 1D 1H MRS, quantification of individual components is precluded. Development of a MRS method capably of resolving individual components of tissue CCMs would be a significant advance. Herein, a modification of the 2D 1H- 14N HSQC technique is targeted on the two methylene 1H in the CH 2O group ( 3J1H14N = 2.7 Hz) and applied to ex vivo mouse and human liver samples at physiological temperature (37 °C). Specifically, the 1H- 14N HSQC technique is modified into a 2D 1H- 14N three-bond correlation (HN3BC) experiment, which selectively detects the 1H of CH 2O coupled to 14N in CCMs. Separate signals from Cho, PCho, and GPCho components are resolved with high detection sensitivity. A 2D HN3BC spectrum can be recorded from mouse liver in only 1.5 min and from human carcinoma liver tissue in less than 3 min with effective sample volume of 0.2 ml at 14.1 T.

  5. [High throuput analysis of organophosphorus pesticide residues and their metabolites in animal original foods by dual gas chromatography-dual pulse flame photometric detection].

    PubMed

    Yang, Lixin; Li, Heli; Miao, Hong; Zeng, Fangang; Li, Ruifeng; Chen, Huijing; Zhao, Yunfeng; Wu, Yongning

    2011-10-01

    A method was established for the quantitative determination of 54 organophosphorus pesticide residues and their metabolites in foods of animal origin by dual gas chromatography-dual pulse flame photometric detection. Homogenized samples were extracted with acetone and methylene chloride, and cleaned-up by gel permeation chromatography (GPC). The response of each analyte showed a good linearity with a correlation coefficient not less than 0. 99. The recovery experiments were performed by a blank sample spiked at low, medium and high fortification levels. The recoveries for beef, mutton, pork, chicken were in the range of 50. 5% -128. 1% with the relative standard deviations (n = 6) of 1. 1% -25. 5%, which demonstrated the good precision and accuracy of the present method. The limits of detection for the analytes were in the range of 0. 001 -0. 170 mg/kg, and the limits of quantification were in the range of 0. 002 -0. 455 mg/kg. Animal food samples collected from markets such as meat, liver and kidney were analyzed, and the residues of dichlorovos and disulfoton-sulfoxide were found in the some samples. The established method is sensitive and selective enough to detect organophosphorus pesticide residues in animal foods. PMID:22268359

  6. An environmental risk assessment for oseltamivir (Tamiflu) for sewage works and surface waters under seasonal-influenza- and pandemic-use conditions.

    PubMed

    Straub, Jürg Oliver

    2009-09-01

    In the event of an influenza pandemic, anti-viral medications such as oseltamivir (Tamiflu) are expected to be used in high amounts over a duration of several weeks. Oseltamivir has been predicted to reach high concentrations in surface waters and sewage works. New oseltamivir environmental fate and toxicity studies permit an environmental risk assessment (ERA) under seasonal- and pandemic-use scenarios. The environmental fate data for sewage works (no removal), surface waters (no significant degradation), and water/sediment systems (>50% primary degradation in 100 days) were used for the derivation of new predicted environmental concentrations (PECs) for western Europe and the River Lee catchment in the UK. Existing worst-case PECs for western Europe, the River Lee catchment in the UK and the Lower Colorado basin in the USA under pandemic conditions (< or =98.1 microg/L for surface waters, < or =348 microg/L for sewage works) were also considered for the ERA. PECs were compared with predicted no-effect concentrations (PNECs) based on new chronic ecotoxicity data (no observed effect concentration for algae, daphnia, and fish > or =1 mg/L). Based on all PEC/PNEC risk ratios, no significant risk is evident to surface waters or sewage works during both regular seasonal-use and high pandemic-use of oseltamivir. PMID:19560203

  7. Effects of Water Models on Binding Affinity: Evidence from All-Atom Simulation of Binding of Tamiflu to A/H5N1 Neuraminidase

    PubMed Central

    Nguyen, Trang Truc; Viet, Man Hoang

    2014-01-01

    The influence of water models SPC, SPC/E, TIP3P, and TIP4P on ligand binding affinity is examined by calculating the binding free energy ΔGbind of oseltamivir carboxylate (Tamiflu) to the wild type of glycoprotein neuraminidase from the pandemic A/H5N1 virus. ΔGbind is estimated by the Molecular Mechanic-Poisson Boltzmann Surface Area method and all-atom simulations with different combinations of these aqueous models and four force fields AMBER99SB, CHARMM27, GROMOS96 43a1, and OPLS-AA/L. It is shown that there is no correlation between the binding free energy and the water density in the binding pocket in CHARMM. However, for three remaining force fields ΔGbind decays with increase of water density. SPC/E provides the lowest binding free energy for any force field, while the water effect is the most pronounced in CHARMM. In agreement with the popular GROMACS recommendation, the binding score obtained by combinations of AMBER-TIP3P, OPLS-TIP4P, and GROMOS-SPC is the most relevant to the experiments. For wild-type neuraminidase we have found that SPC is more suitable for CHARMM than TIP3P recommended by GROMACS for studying ligand binding. However, our study for three of its mutants reveals that TIP3P is presumably the best choice for CHARMM. PMID:24672329

  8. Volatile Metabolites

    PubMed Central

    Rowan, Daryl D.

    2011-01-01

    Volatile organic compounds (volatiles) comprise a chemically diverse class of low molecular weight organic compounds having an appreciable vapor pressure under ambient conditions. Volatiles produced by plants attract pollinators and seed dispersers, and provide defense against pests and pathogens. For insects, volatiles may act as pheromones directing social behavior or as cues for finding hosts or prey. For humans, volatiles are important as flavorants and as possible disease biomarkers. The marine environment is also a major source of halogenated and sulfur-containing volatiles which participate in the global cycling of these elements. While volatile analysis commonly measures a rather restricted set of analytes, the diverse and extreme physical properties of volatiles provide unique analytical challenges. Volatiles constitute only a small proportion of the total number of metabolites produced by living organisms, however, because of their roles as signaling molecules (semiochemicals) both within and between organisms, accurately measuring and determining the roles of these compounds is crucial to an integrated understanding of living systems. This review summarizes recent developments in volatile research from a metabolomics perspective with a focus on the role of recent technical innovation in developing new areas of volatile research and expanding the range of ecological interactions which may be mediated by volatile organic metabolites. PMID:24957243

  9. Time-course measurements of caffeine and its metabolites extracted from fingertips after coffee intake: a preliminary study for the detection of drugs from fingerprints.

    PubMed

    Kuwayama, Kenji; Tsujikawa, Kenji; Miyaguchi, Hajime; Kanamori, Tatsuyuki; Iwata, Yuko T; Inoue, Hiroyuki

    2013-05-01

    The aim of this study was to determine whether an ingested drug and its metabolites could be detected in the subject's fingerprints. Caffeine (CF) was chosen as the model drug. Three healthy subjects were asked to consume a cup of coffee (ca. 100 mL) containing 80 micro micro mg CF as the total dose, which is the normal amount in one cup of coffee. After washing hands with water to remove external contaminants, each subject pressed the index fingertip to a collecting matrix just before consuming the test cup of coffee, and then again pressed the index fingertip to the collecting matrix after 1, 3, 5, and 7 h. The time curve of the amounts of CF and its metabolites-theobromine (TB), paraxanthine (PX), and theophylline (TP)-in fingerprints and blood was determined using liquid chromatography/tandem mass spectrometry (LC/MS). A filter paper wetted with water (50 μL) was an efficient collecting matrix for extracting the analytes from the fingertip. With optimized sample preparation and LC/MS conditions, the total operating time, from taking the fingerprints to obtaining the analytical result, was approximately 10 min. The lower limits of quantification for CF, TB, PX, and TP were 0.5, 5, 0.5, and 5 ng/fingerprint, respectively. The amount of CF or PX determined in fingerprints obtained over 7 h after coffee intake was significantly greater than the amount determined in fingerprints taken before drinking coffee. Fingerprints were a more efficient source for drug testing than other biological samples, such as blood and sweat, because the procedures for sampling and extracting the drugs were simpler and took less time. The method could be used to prove drug intake in criminal investigations. PMID:23187828

  10. FISH BILIARY POLYCYCLIC AROMATIC HYDROCARBON METABOLITES ESTIMATED BY FIXED-WAVELENGTH FLUORESCENCE: COMPARISON WITH HPLC-FLUORESCENT DETECTION

    EPA Science Inventory

    Fixed wavelength fluorescence (FF) was compared to high-performance liquid chromatography with fluorescence detection (HPLC-F) as an estimation of polycyclic aromatic hydrocarbon (PAH) exposure to fish. Two excitation/emission wavelength pairs were used to measure naphthalene- an...

  11. Measuring levels of biogenic amines and their metabolites in rat brain tissue using high-performance liquid chromatography with photodiode array detection.

    PubMed

    Gu, Min-Jung; Jeon, Ji-Hyun; Oh, Myung Sook; Hong, Seon-Pyo

    2016-01-01

    We developed a method to detect biogenic amines and their metabolites in rat brain tissue using simultaneous high-performance liquid chromatography and a photodiode array detection. Measurements were made using a Hypersil Gold C-18 column (250 × 2.1 mm, 5 µm). The mobile phase was 5 mM perchloric acid containing 5 % acetonitrile. The correlation coefficient was 0.9995-0.9999. LODs (S/N = 3) and LOQs (S/N = 10) were as follows: dopamine 0.4 and 1.3 pg, 3, 4-dihydroxyphenylacetic acid 8.4 and 28.0 pg, serotonin 0.4 and 1.3 pg, 5-hydroxyindolacetic acid 3.4 and 11.3 pg, and homovanillic acid 8.4 and 28.0 pg. This method does not require derivatization steps, and is more sensitive than the widely used HPLC-UV method. PMID:26463700

  12. Analytical method for the determination of atrazine and its dealkylated chlorotriazine metabolites in water using gas chromatography/mass selective detection.

    PubMed

    Yokley, R A; Cheung, M W

    2000-10-01

    A multiresidue method is reported for the determination of atrazine and its dealkylated chlorotriazine metabolites in water. Water samples are buffered to pH 10 and partitioned in ethyl acetate. Final analysis is accomplished using gas chromatography/mass selective detection (GC/MSD) in the selected ion monitoring (SIM) mode. The limit of detection (LOD) is 0.050 ng and the limit of quantification (LOQ) is 0.10 ppb for 2-chloro-4-(ethylamino)-6-(isopropylamino)-s-triazine (atrazine), 2-amino-4-chloro-6-(isopropylamino)-s-triazine (G-30033), 2-amino-4-chloro-6-(ethylamino)-s-triazine (G-28279), and 2, 4-diamino-6-chloro-s-triazine (G-28273). The mean procedural recoveries were 90, 92, 98, and 85% and the standard deviations were 12, 13, 16, and 20% for atrazine, G-30033, G-28279, and G-28273, respectively (n = 30). The study was conducted under U.S. EPA FIFRA Good Laboratory Practice Guidelines 40 CFR 160 for method validation. The reported procedure accounts for residues of G-28273 in water that are not included in EPA Method 507. PMID:11052690

  13. Determination of heroin metabolites in human urine using capillary zone electrophoresis with beta-cyclodextrin and UV detection.

    PubMed

    Alnajjar, Ahmed; McCord, Bruce

    2003-10-15

    A method has been developed for the detection of a mixture of morphine, codeine, 6-acetyl morphine (6-AM) and normorphine using capillary zone electrophoresis (CZE). The method utilized urinary 6-AM as a diagnostic indicator of heroin abuse because it is not a product of either morphine or codeine metabolism. The electrophoretic separation was achieved using an uncoated (50 microm I.D.) fused-silica capillary, 77 cm long, containing the detector window 10.0 cm from the outlet end. The running buffer (pH 6.0) contained 50 mM sodium phosphate and 0.015 M beta-cyclodextrins (beta-CD). The samples were first extracted using a mixed-mode solid-phase extraction procedure and then analyzed by CZE. The UV absorbance detection was monitored at 214 nm. It has been found that beta-CDs can improve separation efficiency due to their hydrophobic cavity. The effect of the concentration of beta-CD and pH was also evaluated. The application of electrokinetic injection with field amplified sample stacking results in low detection limits (40 ng/ml for each analyte) and the method has good reproducibility, precision, accuracy, and high recovery. PMID:14550865

  14. Simultaneous determination of Aprepitant and two metabolites in human plasma by high-performance liquid chromatography with tandem mass spectrometric detection.

    PubMed

    Chavez-Eng, C M; Constanzer, M L; Matuszewski, B K

    2004-09-01

    A method for the simultaneous determination of Aprepitant, I (5-[[2(R)-[1(R)-(3,5-bistrifluoromethylphenyl)ethoxy]-3(S)-(4-fluorophenyl) morpholin-4-yl]methyl]-2,4-dihydro-[1,2,4]triazol-3-one) and two active metabolites (II and III) in human plasma has been developed. The method was based on high-performance liquid chromatography (HPLC) with atmospheric pressure chemical ionization tandem mass spectrometric (APCI-MS-MS) detection in positive ionization mode using a heated nebulizer interface. The analytes and internal standard (IV) (Fig. 1) were isolated from basified plasma using liquid-liquid extraction. The organic extracts were dried, reconstituted in mobile phase and injected into the HPLC-MS/MS system. The analytes were chromatographed on a narrow bore (50 mm x 2.0 mm, 3 microm) Keystone Scientific's Prism R.P. analytical column, with mobile phase consisting of acetonitrile (ACN):water containing trifluoroacetic acid with pH adjusted to 3 (40:60, v/v) pumped at a flow rate of 0.5 ml/min. The MS-MS detection was performed on a Sciex API 3000 tandem mass spectrometer operated in selected reaction monitoring mode. The precursor-->product ion combinations of m/z 535-->277, 438-->180, 452-->223 and 503-->259 were used to quantify I, II, III, and IV, respectively, after chromatographic separation of the analytes. The assay was validated in the concentration range of 10-5000 ng/ml for I and II and 25-5000 ng/ml for III when 1 ml of plasma was processed. The precision of the assay (expressed as coefficient of variation, CV) was less than 10% at all concentrations within the standard curve range, with adequate assay accuracy. Matrix effect experiments were performed to demonstrate the absence of any significant change in ionization of the analytes when comparing neat standards to analytes in the presence of plasma matrix. This assay was utilized to support a clinical study where multiple oral doses of I were administered to healthy subjects to investigate the

  15. Methods of analysis by the U.S. Geological Survey Organic Geochemistry Research Group; determination of chloroacetanilide herbicide metabolites in water using high-performance liquid chromatography-diode array detection and high-performance liquid chromatography/mass spectrometry

    USGS Publications Warehouse

    Zimmerman, L.R.; Hostetler, K.A.; Thurman, E.M.

    2000-01-01

    Analytical methods using high-performance liquid chromatography-diode array detection (HPLC-DAD) and high-performance liquid chromatography/mass spectrometry (HPLC/MS) were developed for the analysis of the following chloroacetanilide herbicide metabolites in water: acetochlor ethanesulfonic acid (ESA), acetochlor oxanilic acid (OXA), alachlor ESA, alachlor OXA, metolachlor ESA, and metolachlor OXA. Good precision and accuracy were demonstrated for both the HPLC-DAD and HPLC/MS methods in reagent water, surface water, and ground water. The mean HPLC-DAD recoveries of the chloroacetanilide herbicide metabolites from water samples spiked at 0.25, 0.50, and 2.0 mg/L (micrograms per liter) ranged from 84 to 112 percent, with relative standard deviations of 18 percent or less. The mean HPLC/MS recoveries of the metabolites from water samples spiked at 0.05, 0.20, and 2.0 mg/L ranged from 81 to 125 percent, with relative standard deviations of 20 percent or less. The limit of quantitation (LOQ) for all metabolites using the HPLC-DAD method was 0.20 mg/L, whereas the LOQ using the HPLC/MS method was 0.05 mg/L. These metabolite-determination methods are valuable for acquiring information about water quality and the fate and transport of the parent chloroacetanilide herbicides in water.

  16. Detection of 3-methylmethcathinone and its metabolites 3-methylephedrine and 3-methylnorephedrine in pubic hair samples by liquid chromatography-high resolution/high accuracy Orbitrap mass spectrometry.

    PubMed

    Frison, Giampietro; Frasson, Samuela; Zancanaro, Flavio; Tedeschi, Gianpaola; Zamengo, Luca

    2016-08-01

    Hair testing is considered to be one of the most efficient tool to investigate drug-related histories, particularly when the period of use needs to be tested back to many days or even months before sampling. High-resolution mass spectrometry represents today one of the most specific and sensitive analytical techniques to detect psychoactive substances in hair samples following single or multiple drug exposures. In this study pubic hair testing, by means of liquid chromatography-high resolution/high accuracy Orbitrap mass spectrometry, was employed to document the potential intake of five new psychoactive substances by a drug dealer. Pubic hair samples were decontaminated and pulverized with a ball mill, and, after the addition of the internal standard 3,4-methylenedioxypropylamphetamine, extracted with methanol:trifluoroacetic acid 9:1 at 45°C for one night. The obtained extracts were analyzed on a Thermo Fisher Scientific Accela 1250 liquid chromatography system coupled to a Thermo Fisher Scientific single-stage Exactive HCD mass spectrometry system. 3-methylmethcathinone (3-MMC) was found to be present at a concentration of 25.8ng/mg in the pubic hair sample, whereas the other four designer drugs were found to be absent. 3-methylephedrines and 3-methylnorephedrines, metabolites of 3-MMC, were identified in the same sample, thereby proving the 3-MMC intake by the drug dealer. PMID:26901638

  17. Comparison of immunoassay screening tests and LC-MS-MS for urine detection of benzodiazepines and their metabolites: results of a national proficiency test.

    PubMed

    Bertol, Elisabetta; Vaiano, Fabio; Borsotti, Maurizio; Quercioli, Massimo; Mari, Francesco

    2013-01-01

    For most diverse purposes, different immunoassay (IA) screening methods are usually used to detect benzodiazepines and their metabolites in urine. In this study, we compared the main IAs used in forensic toxicology (Cloned Enzyme Donor Immunoassay, CEDIA®; Enzyme-Multiplied Immunoassay Technique, EMIT®; Fluorescent Polarization ImmunoAssay, FPIA®; Kinetic Interaction of Microparticles in Solution, KIMS® and Immunochromatographic Techniques, IMC) with liquid chromatography-tandem mass spectrometry (LC-MS-MS). Twelve urine specimens were analyzed by 178 laboratories in Italy that participated in a National Proficiency Test, providing both qualitative and semi-quantitative results. Each IA was evaluated by the parameters: true positive, true negative, false positive (FP), false negative (FN), sensitivity (SENS), specificity (SPEC), positive predictive value, negative predictive value (NPV) and accuracy. SPEC was affected by a high FP rate for all IAs. The lowest SENS and NPV were provided by FPIA due to a high number of FN cases. Comparing IA semi-quantitative data with LC-MS-MS results, an overestimation of benzodiazepine amount is noted. This paper draws attention to the problem of the careless use of IA tests for forensic purposes as they may provide FP and/or FN results that can lead to errors of great severity. PMID:23943436

  18. Gas chromatographic method using nitrogen-phosphorus detection for the measurement of tramadol and its O-desmethyl metabolite in plasma and brain tissue of mice and rats.

    PubMed

    Tao, Q; Stone, D J; Borenstein, M R; Jean-Bart, V; Codd, E E; Coogan, T P; Desai-Krieger, D; Liao, S; Raffa, R B

    2001-11-01

    A method that allows the measurement of plasma and brain levels of the centrally-acting analgesic tramadol and its major metabolite (O-desmethyl tramadol) in mice and rats was developed using gas chromatography equipped with nitrogen-phosphorus detection (GC-NPD). Plasma samples were extracted with methyl tert.-butyl ether (MTBE) and were injected directly into the GC system. Brain tissue homogenates were precipitated with methanol, the resulting supernatant was dried then acidified with hydrochloric acid. The aqueous solution was washed with MTBE twice, alkalinized, and extracted with MTBE. The MTBE layer was dried, reconstituted and injected into the GC system. The GC assay used a DB-1 capillary column with an oven temperature ramp (135 to 179 degrees C at 4 degrees C/min). Dextromethorphan was used as the internal standard. The calibration curves for tramadol and O-desmethyl tramadol in plasma and brain tissue were linear in the range of 10 to 10000 ng/ml (plasma) and ng/g (brain). Assay accuracy and precision of back calculated standards were within +/- 15%. PMID:11710575

  19. Detection of Pancreatic Cancer-Induced Cachexia Using a Fluorescent Myoblast Reporter System and Analysis of Metabolite Abundance.

    PubMed

    Winnard, Paul T; Bharti, Santosh K; Penet, Marie-France; Marik, Radharani; Mironchik, Yelena; Wildes, Flonne; Maitra, Anirban; Bhujwalla, Zaver M

    2016-03-15

    The dire effects of cancer-induced cachexia undermine treatment and contribute to decreased survival rates. Therapeutic options for this syndrome are limited, and therefore efforts to identify signs of precachexia in cancer patients are necessary for early intervention. The applications of molecular and functional imaging that would enable a whole-body "holistic" approach to this problem may lead to new insights and advances for diagnosis and treatment of this syndrome. Here we have developed a myoblast optical reporter system with the purpose of identifying early cachectic events. We generated a myoblast cell line expressing a dual tdTomato:GFP construct that was grafted onto the muscle of mice-bearing human pancreatic cancer xenografts to provide noninvasive live imaging of events associated with cancer-induced cachexia (i.e., weight loss). Real-time optical imaging detected a strong tdTomato fluorescent signal from skeletal muscle grafts in mice with weight losses of only 1.2% to 2.7% and tumor burdens of only approximately 79 to 170 mm(3). Weight loss in cachectic animals was also associated with a depletion of lipid, cholesterol, valine, and alanine levels, which may provide informative biomarkers of cachexia. Taken together, our findings demonstrate the utility of a reporter system that is capable of tracking tumor-induced weight loss, an early marker of cachexia. Future studies incorporating resected tissue from human pancreatic ductal adenocarcinoma into a reporter-carrying mouse may be able to provide a risk assessment of cachexia, with possible implications for therapeutic development. PMID:26719527

  20. Identification and determination of phase II nabumetone metabolites by high-performance liquid chromatography with photodiode array and mass spectrometric detection.

    PubMed

    Nobilis, M; Holcapek, M; Kolárová, L; Kopecký, J; Kunes, M; Svoboda, Z; Kvetina, J

    2004-03-26

    Chromatographic analyses play an important role in the identification and determination of phase I and phase II drug metabolites. While the chemical standards of phase I metabolites are usually available from commercial sources or by various synthetic, degradation or isolation methods, the phase II drug metabolites have usually more complicated structures, their standards are in general inaccessible and their identification and determination require a comprehensive analytical approach involving the use of xenobiochemical methods and the employment of hyphenated analytical techniques. In this work, various high-performance liquid chromatography (HPLC) methods were employed in the evaluation of xenobiochemical experiments leading to the identification and determination of phase II nabumetone metabolites. Optimal conditions for the quantitative enzymatic deconjugation of phase II metabolites were found for the samples of minipig bile, small intestine contents and urine. Comparative HPLC analyses of the samples of above-mentioned biomatrices and of the same biomatrices after their enzymatic treatment using beta-glucuronidase and arylsulfatase afforded the qualitative and quantitative information about phase II nabumetone metabolites. Hereby, three principal phase II nabumetone metabolites (ether glucuronides) were discovered in minipig's body fluids and their structures were confirmed using liquid chromatography (LC)-electrospray ionization mass spectrometric (MS) analyses. PMID:15058587

  1. Automated analysis of oxidative metabolites

    NASA Technical Reports Server (NTRS)

    Furner, R. L. (Inventor)

    1974-01-01

    An automated system for the study of drug metabolism is described. The system monitors the oxidative metabolites of aromatic amines and of compounds which produce formaldehyde on oxidative dealkylation. It includes color developing compositions suitable for detecting hyroxylated aromatic amines and formaldehyde.

  2. High-performance liquid chromatography-tandem mass spectrometry method for simultaneous detection of ochratoxin A and relative metabolites in Aspergillus species and dried vine fruits.

    PubMed

    Zhang, Xiaoxu; Li, Jingming; Cheng, Zhan; Zhou, Ziying; Ma, Liyan

    2016-08-01

    A simple, sensitive and reliable quantification and identification method was developed for simultaneous analysis of ochratoxin A (OTA) and its related metabolites ochratoxin alpha (OTα), ochratoxin B (OTB) and mellein. The method was assessed by spiking analytes into blank culture media and dried vine fruits. Performance was tested in terms of accuracy, selectivity and repeatability. The method involves an ultrasonic extraction step for culture samples using methanol aqueous solution (7:3, v/v); the mycotoxin is quantified by high-performance liquid chromatography coupled with electrospray ionisation and triple quadrupole mass spectrometry (LC-ESI-MS/MS). The recoveries were 74.5-108.8%, with relative standard deviations (RSDs) of 0.4-8.4% for fungal culture. The limits of detection (LODs) were in the range of 0.03-0.87 μg l(-)(1), and the limits of quantification (LOQs) ranged from 0.07 to 2.90 μg l(-)(1). In addition, the extraction solutions and clean-up columns were optimised specifically for dried vine fruit samples to improve the performance of the method. Methanol-1% sodium bicarbonate extraction solution (6:4, v/v) was determined to be the most efficient. Solid-phase extraction (SPE) was performed as a clean-up step prior to HPLC-MS/MS analysis to reduce matrix effects. Recoveries ranged from 80.1% to 110.8%. RSDs ranged from 0.1% to 3.6%. LODs and LOQs ranged from 0.06 to 0.40 μg kg(-)(1) and from 0.19 to 1.20 μg kg(-)(1), respectively. The analytical method was established and used to identify and quantify OTA and related compounds from Aspergillus carbonarius and Aspergillus ochraceus in cultures and dried vine fruits. The results showed that A. carbonarius produced OTα, OTB and OTA, whereas A. ochraceus produced OTB, OTA and mellein after 7 days of cultivation. Of 30 commercial samples analysed, 10 were contaminated with ochratoxins; OTB, OTα and mellein were also detected in different samples. PMID:27442910

  3. Metabolite profiling of licorice (Glycyrrhiza glabra) from different locations using comprehensive two-dimensional liquid chromatography coupled to diode array and tandem mass spectrometry detection.

    PubMed

    Montero, Lidia; Ibáñez, Elena; Russo, Mariateresa; di Sanzo, Rosa; Rastrelli, Luca; Piccinelli, Anna Lisa; Celano, Rita; Cifuentes, Alejandro; Herrero, Miguel

    2016-03-24

    Profiling of the main metabolites from several licorice (Glycyrrhiza glabra) samples collected at different locations is carried out in this work by using comprehensive two-dimensional liquid chromatography (LC × LC) coupled to diode array (DAD) and mass spectrometry (MS) detectors. The optimized method was based on the application of a HILIC-based separation in the first dimension combined with fast RP-based second dimension separation. This set-up was shown to possess powerful separation capabilities allowing separating as much as 89 different metabolites in a single sample. Identification and grouping of metabolites according to their chemical class were achieved using the DAD, MS and MS/MS data. Triterpene saponins were the most abundant metabolites followed by glycosylated flavanones and chalcones, whereas glycyrrhizic acid, as expected, was confirmed as the main component in all the studied samples. LC × LC-DAD-MS/MS was able to resolve these complex licorice samples providing with specific metabolite profiles to the different licorice samples depending on their geographical origin. Namely, from 19 to 50 specific compounds were exclusively determined in the 2D-chromatograms from the different licorice samples depending on their geographical origin, which can be used as a typical pattern that could potentially be related to their geographical location and authentication. PMID:26944999

  4. The metabolism of 4-trifluoromethoxyaniline and [13C]-4-trifluoromethoxyacetanilide in the rat: detection and identification of metabolites excreted in the urine by NMR and HPLC-NMR.

    PubMed

    Tugnait, M; Lenz, E M; Phillips, P; Hofmann, M; Spraul, M; Lindon, J C; Nicholson, J K; Wilson, I D

    2002-06-01

    A combination of 19F, 1H NMR and HPLC-NMR spectroscopic approaches have been used to quantify and identify the urinary-excreted metabolites of 4-trifluoromethoxyaniline (4-TFMeA) and its [13C]-labelled acetanilide following i.p. administration at 50 mg/kg to rats. The major metabolite excreted in the urine for both compounds was a sulphated ring-hydroxylated metabolite (either 2- or 3-trifluoromethyl-5-aminosulphate) which accounted for approximately 32.3% of the dose following the administration of 4-TFMeA and approximately 29.9% following dosing of the acetanilide. The trifluoromethoxy-substituent appeared to be metabolically stable, with no evidence of O-detrifluoromethylation. There was no evidence of the excretion of N-oxanilic acids in urine, of the type seen with 4-trifluoromethylaniline. PMID:12039629

  5. Detection of ketamine and its metabolites in human hair using an integrated nanoflow liquid chromatography column and electrospray emitter fritted with a single porous 10 μm bead.

    PubMed

    Parkin, Mark C; Longmoore, Alana M; Turfus, Sophie C; Braithwaite, Robin A; Cowan, David A; Elliott, Simon; Kicman, Andrew T

    2013-02-15

    Targeting metabolites incorporated into hair following drug administration is useful for evidential purposes as this approach can aid in differentiating between administration and passive exposure. Greater analytical sensitivity is required than for targeting the parent drug alone. A 20 μm i.d. fused silica capillary column with an integrated electrospray emitter fritted with a single porous 10 μm polymeric bead has been successfully fabricated to facilitate this purpose. The sensitivity gains through the use of these integrated single fritted columns coupled to a nanoelectrospray source (nanoflow-LC nanoESI) over conventional liquid chromatography-tandem mass spectrometry (LC-MS/MS) columns was explored by their application to the detection of ketamine and its phase I metabolites in human hair. Hair was collected from 4 volunteers following the administration of a small oral dose of ketamine (50 mg) and subsequently analysed by the capillary-LC nanoESI approach. The drug and its metabolites were extracted from hair using solid phase extraction following a methanolic wash, pulverisation with a ball mill and acid digestion. From a 50 μL extract, 1 μL was injected and the method provided a limit of detection estimated to be 5 fg on column for ketamine and norketamine and 10 fg for dehydronorketamine. The single porous frit minimises extra column band broadening and offers an alternative fritting approach which reduces the blocking of the electrospray emitter, in comparison with other approaches such as sintering and polymerisation. PMID:23332304

  6. An approach for quantitative analysis of vitamins D and B9 and their metabolites in human biofluids by on-line orthogonal sample preparation and sequential mass spectrometry detection.

    PubMed

    Ferreiro-Vera, Carlos; Priego-Capote, Feliciano; Luque de Castro, María Dolores

    2013-04-01

    An approach for quantitative analysis of two vitamins with different polarities (vitamins D and B9) and their metabolites is presented here. The approach is based on an experimental setup based on hyphenation of an automated workstation for preparation of liquid samples and an LC-MS/MS system with a triple quadrupole mass spectrometer. This configuration enabled development of an orthogonal protocol for sequential SPE retention of analytes with different polarities for subsequent elution and chromatographic separation prior to detection. The resulting method was validated by application to three human biofluids. Estimation of recovery factors in the SPE step led to values from 85.2 to 100% for vitamin D and metabolites and from 93.1 to 100% for vitamin B9 and metabolites (folic acid and folates). The influence of sample matrix variability by analysis of human serum, urine and breast milk was minimized with a complete optimization of the SPE step. The utility of the proposed configuration is shown by the sensitivity and precision of the method, expressed as limits of detection (between 0.2 and 0.30 ng mL(-1) or 4 and 60 pg on-column) and within-laboratory reproducibility (lower than 6.7%, as relative standard deviation). The present application represents an example of determination methods involving targeted analysis of compounds with different polarities using a single aliquot of the sample. PMID:23435066

  7. Secondary metabolite profiles of leaves of healthy and huanglongbing-infected orange (Citrus sinensis L.) seedlings measured by HPLC-fluorescence detection

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Leaves of greenhouse-grown ‘Hamlin’ and ‘Valencia’ orange (Citrus sinensis L.) seedlings were analyzed by high performance liquid chromatography in a study of the progression of changes in secondary metabolite profiles resulting from infection by Candidatus Liberibacter asiaticus (CLas), the Huanglo...

  8. Analysis and detection of the chemical constituents of Radix Polygalae and their metabolites in rats after oral administration by ultra high-performance liquid chromatography coupled with electrospray ionization quadrupole time-of-flight tandem mass spectrometry.

    PubMed

    Ling, Yun; Li, Zhixiong; Chen, Mingcang; Sun, Zhaolin; Fan, Mingsong; Huang, Chenggang

    2013-11-01

    Radix Polygalae (RP), the dried root of Polygala tenuifolia Willd., is a well-known traditional Chinese medicine to mediate sedative, antipsychotic, cognitive improving, neuroprotective, and anti-inflammatory therapeutic effects on the central nervous system. In this work, ultra high-performance liquid chromatography coupled with electrospray ionization quadrupole time-of-flight tandem mass spectrometry (UHPLC/ESI-Q-TOF-MS/MS) was established for the separation and characterization of the chemical constituents in Radix Polygalae and their metabolites in rat plasma and urine after oral administration. Samples were separated on an Agilent Zorbax Eclipse Plus-C18 column (100mm×2.1mm, 1.8μm) with 0.1% formic acid aqueous solution and acetonitrile as the mobile phase under gradient conditions. Overall, 50 compounds were characterized from the RP, 9 of which are to our knowledge reported for the first time. In vivo, 10 components and 2 metabolites were observed in rat plasma, and 27 components and 7 metabolites were detected in rat urine. The results from this work improve our understanding on the chemical constituents of RP and their metabolic profiling. PMID:23860503

  9. A simple HPLC method with fluorescence detection for simultaneous determination of 10-methoxycamptothecin and its metabolite 10-hydroxycamptothecin in rat liver tissue.

    PubMed

    Liu, Y; Shao, C; Fan, B; Li, S; Wang, Y; Zheng, J

    2015-03-01

    A simple HPLC method to determine the amount of 10-methoxycamptothecin (MCPT) and its major metabolite 10-hydroxycamptothecin (HCPT) in rat liver tissue was developed in the present study. Camptothecin (CPT) was used as internal standard (IS). A piecewise linear function was used over lower and higher concentrations, respectively. The calibration curves were linear (r (2) >0.99) over concentrations from 2.5 to 20 ng/mL and 20 to 320 ng/mL for both MCPT and HCPT. The method had an accuracy of 92.74% to 112.76%, and the intra- and inter-day precision (RSD%) were 11.85% or less for MCPT and HCPT. The stability data showed no significant degradation occurred under the experimental conditions. This method was successfully applied to the tissue distribution study of MCPT and its metabolite HCPT in liver tissue samples after intravenous administration. PMID:24782285

  10. Characterization of flavonoid metabolites in rat plasma, urine, and feces after oral administration of Semen Ziziphi Spinosae extract by HPLC-diode-array detection (DAD) and ion-trap mass spectrometry (MS(n)).

    PubMed

    Bao, Kang-De; Li, Ping; Qi, Lian-Wen; Li, Hui-Jun; Yi, Ling; Wang, Wei; Wang, Ya-Qiong

    2009-02-01

    A highly specific and sensitive method using high performance liquid chromatography coupled with diode-array detection and ion-trap mass spectrometry (HPLC-DAD-MS(n)) was developed for study of the constituents of flavonoid extract of jujube seeds and the metabolites in rat plasma, urine, and feces samples after oral administration of flavonoid extract of jujube seeds. Two major flavonoids (spinosin and 6'''-feruloylspinosin) with content >60% in the flavonoid extract of jujube seeds were detected and confirmed by comparison with the reference standards. Furthermore, five metabolic components in plasma, seven in urine, and four in feces were detected and elucidated. The scientific and plausible biotransformation pathways of the main components in flavonoid extract of jujube seeds were also proposed, together with presentation of clues for potential bioactive mechanisms. This convenient HPLC-DAD-MS(n) method could be used to identify the chemical components of flavonoid extract of jujube seeds as well as their metabolites, and to reveal their possible metabolic mechanism of action in vivo. PMID:19182403

  11. A wood preservative metabolite in river water.

    PubMed

    Khoroshko, Larisa O; Petrova, Varvara N; Viktorovskii, Igor V; Lahtiperä, Mirja; Sinkkonen, Seija; Paasivirta, Jaakko

    2005-01-01

    A previously unknown pollutant in river water was identified to be 2-mercaptobenzothiazole (2-MBT) by interpretation and simulation of its GC/LRMS spectrum. Further GC/HRMS measurement of the isotope composition of the molecular ion verified this structure. 2-MBT is a well-known agent for corrosion inhibition and a stable metabolite of several other benzothiazoles. The present 2-MBT trace was most probably a metabolite of the wood preservative TCMTB which leaked from an upstream sawmill. The metabolite had been detected earlier in urine of the sawmill workers, but now was identified in the recipient water environment for the first time. PMID:15768735

  12. Novel and sensitive reversed-phase high-pressure liquid chromatography method with electrochemical detection for the simultaneous and fast determination of eight biogenic amines and metabolites in human brain tissue.

    PubMed

    Van Dam, Debby; Vermeiren, Yannick; Aerts, Tony; De Deyn, Peter Paul

    2014-08-01

    A fast and simple RP-HPLC method with electrochemical detection (ECD) and ion pair chromatography was developed, optimized and validated in order to simultaneously determine eight different biogenic amines and metabolites in post-mortem human brain tissue in a single-run analytical approach. The compounds of interest are the indolamine serotonin (5-hydroxytryptamine, 5-HT), the catecholamines dopamine (DA) and (nor)epinephrine ((N)E), as well as their respective metabolites, i.e. 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA), 5-hydroxy-3-indoleacetic acid (5-HIAA) and 3-methoxy-4-hydroxyphenylglycol (MHPG). A two-level fractional factorial experimental design was applied to study the effect of five experimental factors (i.e. the ion-pair counter concentration, the level of organic modifier, the pH of the mobile phase, the temperature of the column, and the voltage setting of the detector) on the chromatographic behaviour. The cross effect between the five quantitative factors and the capacity and separation factors of the analytes were then analysed using a Standard Least Squares model. The optimized method was fully validated according to the requirements of SFSTP (Société Française des Sciences et Techniques Pharmaceutiques). Our human brain tissue sample preparation procedure is straightforward and relatively short, which allows samples to be loaded onto the HPLC system within approximately 4h. Additionally, a high sample throughput was achieved after optimization due to a total runtime of maximally 40min per sample. The conditions and settings of the HPLC system were found to be accurate with high intra and inter-assay repeatability, recovery and accuracy rates. The robust analytical method results in very low detection limits and good separation for all of the eight biogenic amines and metabolites in this complex mixture of biological analytes. PMID:24857034

  13. Urinary metabolites of diisodecyl phthalate in rats.

    PubMed

    Kato, Kayoko; Silva, Manori J; Wolf, Cynthia; Gray, L Earl; Needham, Larry L; Calafat, Antonia M

    2007-07-01

    Diisodecyl phthalate (DiDP) is an isomeric mixture of phthalates with predominantly 10-carbon branched-dialkyl chains, widely used as a plasticizer for polyvinyl chloride. The extent of human exposure to DiDP is unknown in part because adequate biomarkers of exposure to DiDP are not available. We identified several major metabolites of DiDP in urine of adult female Sprague-Dawley rats after a single oral administration of DiDP (300 mg/kg). These metabolites can potentially be used as biomarkers of exposure to DiDP. The metabolites extracted from urine were chromatographically resolved and identified by their chromatographic behavior and full scan negative ion electrospray ionization mass spectrum. The identity of metabolites with similar molecular weights was further examined in accurate mass mode. For some metabolites, unequivocal identification was done using authentic standards. Among these were the hydrolytic monoester of DiDP, monoisodecyl phthalate (MiDP), detected as a minor metabolite, and one omega oxidation product of MiDP, mono(carboxy-isononyl) phthalate (MCiNP), which was the most abundant urinary metabolite. We also tentatively identified other secondary metabolites of MiDP, mono(hydroxy-isodecyl) phthalate, mono(oxo-isodecyl) phthalate, mono(carboxy-isoheptyl) phthalate, mono(carboxy-isohexyl) phthalate, mono(carboxy-isopentyl) phthalate, mono(carboxy-isobutyl) phthalate, and mono(carboxy-ethyl) phthalate. Oxidative metabolites of diisoundecyl phthalate (DiUdP) and diisononyl phthalate (DiNP) were also detected suggesting the presence of DiUdP and DiNP in the DiDP formulation. The urinary concentrations of all these metabolites gradually decreased in the 4 days following the administration of DiDP. MCiNP and other DiDP secondary metabolites are more abundant in urine than MiDP, suggesting that these oxidative products are better biomarkers for DiDP exposure assessment than MiDP. Additional research on the toxicokinetics of these metabolites is needed

  14. Validation of a novel in vitro assay using ultra performance liquid chromatography-mass spectrometry (UPLC/MS) to detect and quantify hydroxylated metabolites of BDE-99 in rat liver microsomes.

    PubMed

    Erratico, Claudio A; Szeitz, András; Bandiera, Stelvio M

    2010-06-01

    The purpose of this study was to develop and validate an ultra performance liquid chromatography-mass spectrometry (UPLC/MS) method to investigate the hepatic oxidative metabolism of 2,2',4,4',5-pentabromodiphenyl ether (BDE-99), a widely used flame retardant and ubiquitous environmental contaminant. Hydroxylated metabolites were extracted using liquid-to-liquid extraction, resolved on a C18 column with gradient elution and detected by mass spectrometry in single ion recording mode using electrospray negative ionization. The assay was validated for linearity, accuracy, precision, limit of quantification, range and recovery. Calibration curves were linear (R2 > or = 0.98) over a concentration range of 0.010-1.0 microM for 4-OH-2,2',3,4',5-pentabromodiphenyl ether (4-OH-BDE-90), 5'-OH-2,2',4,4',5-pentabromodiphenyl ether (5'-OH-BDE-99) and 6'-OH-2,2',4,4',5-pentabromodiphenyl ether (6'-OH-BDE-99), and a concentration range of 0.0625-12.5 microM for 2,4,5-tribromophenol (2,4,5-TBP). Inter- and intra-day accuracy values ranged from -2.0% to 6.0% and from -7.7% to 7.3%, respectively, and inter- and intra-day precision values ranged from 2.0% to 8.5% and from 2.2% to 8.6% (n=6), respectively. The limits of quantification were 0.010 microM for 4-OH-BDE-90, 5'-OH-BDE-99 and 6'-OH-BDE-99, and 0.0625 microM for 2,4,5-TBP. Recovery values ranged between 85 and 100% for the four analytes. The validated analytical method was applied to identify and quantify hydroxy BDE-99 metabolites formed in vitro. Incubation of BDE-99 with rat liver microsomes yielded 4-OH-BDE-90 and 6'-OH-BDE-99 as major metabolites and 5'-OH-BDE-99 and 2,4,5-TBP as minor metabolites. To our knowledge, this is the first validated UPLC/MS method to quantify hydroxylated metabolites of PBDEs without the need of derivatization. PMID:20451473

  15. Identification of Epoxide-Derived Metabolite(s) of Benzbromarone.

    PubMed

    Wang, Kai; Wang, Hui; Peng, Ying; Zheng, Jiang

    2016-04-01

    Benzbromarone (BBR) is a benzofuran derivative that has been quite useful for the treatment of gout; however, it was withdrawn from European markets in 2003 because of reported serious incidents of drug-induced liver injury. BBR-induced hepatotoxicity has been suggested to be associated with the formation of a quinone intermediate. The present study reported epoxide-derived intermediate(s) of BBR. An N-acetylcysteine (NAC) conjugate derived from epoxide metabolite(s) was detected in both microsomal incubations of BBR and urine samples of mice treated with BBR. The NAC conjugate was identified as 6-NAC BBR. Ketoconazole suppressed the bioactivation of BBR to the epoxide intermediate(s), and the CYP3A subfamily was the primary enzyme responsible for the formation of the epoxide(s). The present study provided new information on metabolic activation of BBR. PMID:26792818

  16. Rapid determination of dichlorodiphenyltrichloroethane and its main metabolites in aqueous samples by one-step microwave-assisted headspace controlled-temperature liquid-phase microextraction and gas chromatography with electron capture detection.

    PubMed

    Vinoth Kumar, Ponnusamy; Jen, Jen-Fon

    2011-03-01

    A rapid and sensitive analytical method for the determination of dichlorodiphenyltrichloroethane (DDT) and its main metabolites in environmental aqueous samples has been developed using one-step microwave-assisted headspace controlled-temperature liquid-phase micro-extraction (MA-HS-CT-LPME) technique coupled with gas chromatography-electron-capture detection (GC-ECD). In this study, the one-step extraction of DDT and its main metabolites was achieved by using microwave heating to accelerate the evaporation of analytes into the controlled-temperature headspace to form a cloudy mist vapor zone for LPME sampling. Parameters influencing extraction efficiency were thoroughly optimized, and the best extraction for DDT and its main metabolites from 10-mL aqueous sample at pH 6.0 was achieved by using 1-octanol (4-μL) as the LPME solvent, sampling at 34°C for 6.5 min under 249W of microwave irradiation. Under optimum conditions, excellent linear relationship was obtained in the range of 0.05-1.0 μg/L for 1-dichloro-2,2-bis-(p'-chlorophenyl)ethylene (p,p'-DDE), 0.1-2.0 μg/L for o,p'-DDT, 0.15-3.0 μg/L for 1,1-dichloro-2,2-bis-(p'-chlorophenyl)ethane (p,p'-DDD) and p,p'-DDT, with detection limits of 20 ng/L for p,p'-DDE, and 30 ng/L for o,p'-DDT, p,p'-DDD and p,p'-DDT. Precision was in the range of 3.2-11.3% RSD. The proposed method was validated with environmental water samples. The spiked recovery was between 95.5% and 101.3% for agricultural-field water, between 94% and 99.7% for sea water and between 93.5% and 98% for river water. Thus the established method has been proved to be a simple, rapid, sensitive, inexpensive and eco-friendly procedure for the determination of DDT and its main metabolites in environmental water samples. PMID:21251695

  17. Reactive metabolites and agranulocytosis.

    PubMed

    Uetrecht, J P

    1996-01-01

    Central to most hypotheses of the mechanism of idiosyncratic drug-induced blood dyscrasias is the involvement of reactive metabolites. In view of the reactive nature of the majority of such metabolites, it is likely that they are formed by, or in close proximity to the blood cells affected. The major oxidative system of neutrophils generates hypochlorous acid. We have demonstrated that the drugs associated with the highest incidence of agranulocytosis are oxidized to reactive metabolites by hypochlorous acid and/or activated neutrophils. There are many mechanisms by which such reactive metabolites could induce agranulocytosis. In the case of aminopyrine-induced agranulocytosis, most cases appear to involve drug-dependent anti-neutrophil antibodies, and these are likely to be induced by cell membrane antigens modified by the reactive metabolite of aminopyrine. The target of agranulocytosis associated with many other drugs is usually neutrophil precursors and may involve cytotoxicity or a cell-mediated immune reaction induced by a reactive metabolite. In the case of aplastic anaemia, there is evidence in some cases for involvement of cytotoxic T cells, which could either be induced by metabolites generated by neutrophils, or more likely, by reactive metabolites generated by stem cells. PMID:8987247

  18. Advances in metabolite identification.

    PubMed

    Wishart, David S

    2011-08-01

    One of the central challenges to metabolomics is metabolite identification. Regardless of whether one uses so-called 'targeted' or 'untargeted' metabolomics, eventually all paths lead to the requirement of identifying (and quantifying) certain key metabolites. Indeed, without metabolite identification, the results of any metabolomic analysis are biologically and chemically uninterpretable. Given the chemical diversity of most metabolomes and the character of most metabolomic data, metabolite identification is intrinsically difficult. Consequently a great deal of effort in metabolomics over the past decade has been focused on making metabolite identification better, faster and cheaper. This review describes some of the newly emerging techniques or technologies in metabolomics that are making metabolite identification easier and more robust. In particular, it focuses on advances in metabolite identification that have occurred over the past 2 to 3 years concerning the technologies, methodologies and software as applied to NMR, MS and separation science. The strengths and limitations of some of these approaches are discussed along with some of the important trends in metabolite identification. PMID:21827274

  19. Blood styrene and urinary metabolites in styrene polymerisation.

    PubMed Central

    Wolff, M S; Lorimer, W V; Lilis, R; Selikoff, I J

    1978-01-01

    The results of the analysis of blood and urine samples for styrene and its metabolites in 491 workers in a styrene polymerisation plant in the United States are reported. The levels of exposure to styrene were estimated to be less than 10 ppm, but nevertheless styrene and metabolites were detectable in more than 50% of workers in polymerisation jobs, within 4 h of exposure. Workers involved in the manufacture and purification of styrene from ethyl benzene also had detectable blood styrene and urinary metabolites in 83% of recently exposed subjects. The relationship between styrene in blood and in subcutaneous fat and urinary metabolites as pharmacokinetic variables is discussed. PMID:737139

  20. Using multi-walled carbon nanotubes as solid phase extraction adsorbents to determine dichlorodiphenyltrichloroethane and its metabolites at trace level in water samples by high performance liquid chromatography with UV detection.

    PubMed

    Zhou, Qingxiang; Xiao, Junping; Wang, Weidong

    2006-09-01

    Carbon nanotubes (CNTs) are a kind of new carbon-based nano-materials which have drawn great attention in many application fields. The potential of multi-walled carbon nanotubes (MWNTs) as solid-phase extraction (SPE) adsorbents for the preconcentration of environmental pollutants has been investigated in recent years. The goal of this work was to investigate the feasibility of MWNTs used as SPE adsorbents to enrich dichlorodiphenyltrichloroethane (DDT) and its metabolites including 1,1-dichloro-2,2-bis-(4'-chlorophenyl)ethane (DDD) and 1,1-dichloro-2,2-bis-(4'-chlorophenyl)ethane (DDE) at trace level which are typical persistent organic pollutants in environment. Parameters that maybe influence the extraction efficiency such as the eluent volume, sample flow rate, sample pH and the sample volume were optimized in detail. The experimental results showed the excellent linear relationship between peak area and the concentration of DDT and its metabolites over the range of 0.2-60 microg L(-1), and the precisions (RSD) were 2.3-2.5% under the optimal conditions. The detection limits of proposed method could reach 4-13 ng L(-1) based on the ratio of chromatographic signal to base line noise (S/N = 3). Satisfied results were achieved when the proposed method was applied to determine the four target compounds in realworld water samples with spiked recoveries over the range of 89.7-115.5%. All these facts indicated that MWCNTs as SPE packing materials coupled to HPLC was an excellent alternative for the routine analysis of DDT and its metabolites at trace level in environment. PMID:16797570

  1. Familial resemblance for serum metabolite concentrations.

    PubMed

    Draisma, Harmen H M; Beekman, Marian; Pool, René; van Ommen, Gert-Jan B; Adamski, Jerzy; Prehn, Cornelia; Vaarhorst, Anika A M; de Craen, Anton J M; Willemsen, Gonneke; Slagboom, P Eline; Boomsma, Dorret I

    2013-10-01

    Metabolomics is the comprehensive study of metabolites, which are the substrates, intermediate, and end products of cellular metabolism. The heritability of the concentrations of circulating metabolites bears relevance for evaluating their suitability as biomarkers for disease. We report aspects of familial resemblance for the concentrations in human serum of more than 100 metabolites, measured using a targeted metabolomics platform. Age- and sex-corrected monozygotic twin correlations, midparent-offspring regression coefficients, and spouse correlations in subjects from two independent cohorts (Netherlands Twin Register and Leiden Longevity Study) were estimated for each metabolite. In the Netherlands Twin Register subjects, who were largely fasting, we found significant monozygotic twin correlations for 121 out of 123 metabolites. Heritability was confirmed by midparent-offspring regression. For most detected metabolites, the correlations between spouses were considerably lower than those between twins, indicating a contribution of genetic effects to familial resemblance. Remarkably high heritability was observed for free carnitine (monozygotic twin correlation 0.66), for the amino acids serine (monozygotic twin correlation 0.77) and threonine (monozygotic twin correlation 0.64), and for phosphatidylcholine acyl-alkyl C40:3 (monozygotic twin correlation 0.77). For octenoylcarnitine, a consistent point estimate of approximately 0.50 was found for the spouse correlations in the two cohorts as well as for the monozygotic twin correlation, suggesting that familiality for this metabolite is explained by shared environment. We conclude that for the majority of metabolites targeted by the used metabolomics platform, the familial resemblance of serum concentrations is largely genetic. Our results contribute to the knowledge of the heritability of fasting serum metabolite concentrations, which is relevant for biomarker research. PMID:23985338

  2. The profiling and identification of the metabolites of 8-prenylkaempferol and a study on their distribution in rats by high-performance liquid chromatography with diode array detection combined with electrospray ionization ion trap time-of-flight multistage mass spectrometry.

    PubMed

    Ma, Li-Man; Xu, Feng; Li, Feng-Chun; Wang, Jing-Zhe; Shang, Ming-Ying; Liu, Guang-Xue; Cai, Shao-Qing

    2016-02-01

    8-Prenylkaempferol is a prenylflavonoid that has various bioactivities and benefits for human health. A high-performance liquid chromatography with a diode array detector combined with electrospray ionization ion trap time-of-flight multistage mass spectrometry (HPLC-DAD-ESI-IT-TOF-MS(n) ) method was established to profile and identify the metabolites of 8-prenylkaempferol in rat in vivo and in vitro, and to study the distribution of these metabolites in rats for the first time. A total of 38 metabolites were detected and tentatively identified, 30 of which were identified as new compounds. The new in vivo metabolic reactions in rats of prenylflavonoids of isomerization, polymerization, sulfation, amino acid conjugation, vitamin C conjugation and other known metabolic reactions were found in the metabolism of 8-prenylkaempferol. The numbers of detected metabolites in feces, urine, plasma, small intestine, stomach, kidneys, liver, heart, lungs, spleen and hepatic S9 fraction were 31, 19, 1, 20, 13, 8, 7, 3, 3, 1 and 11, respectively. This indicated that small intestine and stomach were the major organs in which the 8-prenylkaempferol metabolites were distributed. Furthermore, 16 metabolites were determined to have bioactivities based on the literature and 'PharmMapper' analysis. These findings are useful for better comprehension of the effective forms, target organs and pharmacological actions of 8-prenylkaempferol. Moreover, they provide a reference for the study of the metabolism and distribution of prenylflavonoid aglycone compounds. PMID:26058713

  3. Enhanced metabolite generation

    DOEpatents

    Chidambaram, Devicharan

    2012-03-27

    The present invention relates to the enhanced production of metabolites by a process whereby a carbon source is oxidized with a fermentative microbe in a compartment having a portal. An electron acceptor is added to the compartment to assist the microbe in the removal of excess electrons. The electron acceptor accepts electrons from the microbe after oxidation of the carbon source. Other transfers of electrons can take place to enhance the production of the metabolite, such as acids, biofuels or brewed beverages.

  4. Transplacental transport of netobimin metabolites in ewes.

    PubMed

    Cristofol, C; Carretero, A; Fernandez, M; Navarro, M; Sautet, J; Ruberte, J; Arboix, M

    1995-01-01

    Neither netobimin (NTB) nor its metabolite albendazole (ABZ) were found in plasma after an oral administration of 20 mg/kg of NTB to pregnant ewes during the last third of gestation. ABZ metabolites, albendazole sulphoxide (ABZSO) and albendazole sulphone (ABZSO2) were found in plasma 30 min and 2 h, respectively, after administration. The maximal plasma concentration (Cmax) of ABZSO was detected at 11.6 +/- 1.0 h and for ABZSO2 at 16.5 +/- 2.3 h. The plasma levels of the latter remained constant for 36 h, and decreased as ABZSO was removed from the blood. Jugular plasma levels of both metabolites did not differ significantly from those observed in the ovarian vein, suggesting that there were no exchanges between foetal and placental tissues. Both metabolite concentrations were similar in the umbilical vein and artery and in the amniotic and allantoic fluids, their values were half the maternal plasma concentration, leading to the conclusion that there was transplacental movement of metabolites. Both metabolites reached the foetus and could be responsible for the teratogenicity of NTB in sheep. PMID:8751036

  5. Molecular modeling of swine influenza A/H1N1, Spanish H1N1, and avian H5N1 flu N1 neuraminidases bound to Tamiflu and Relenza

    PubMed Central

    Le, Ly; Lee, Eric; Schulten, Klaus; Truong, Thanh N.

    2011-01-01

    A molecular model of the swine influenza A/H1N1 ( also called H1N1pdm) type-I neuraminidase was built using the pathogenic avian H5N1 type-I neuraminidase as a basis, due to the higher sequence identity between A/H1N1 and H5N1 (91.47%) compared to Spanish H1N1 (88.37%) neuraminidase. All-atom molecular dynamics (MD) simulations of all three neuraminidases were performed, either as apo-structures or with commercial antiviral drugs Tamiflu or Relenza separately bound; the simulations allowed for the identification of both conserved and unique drug-protein interactions across all three proteins. Specifically, conserved networks of hydrogen bonds stabilizing the drugs in the sialic acid binding site of the simulated neuraminidases are analyzed, providing insight into how disruption due to mutations may lead to increased drug resistance. In addition, a possible mechanism through which the residue 294 mutation acquires drug resistance is proposed by mapping the mutation site onto an electrostatic pathway which may play a role in controlling drug access to the binding pocket of neuraminidase, establishing a starting point for further investigations of neuraminidase drug resistance. PMID:20029609

  6. Molecular modeling of swine influenza A/H1N1, Spanish H1N1, and avian H5N1 flu N1 neuraminidases bound to Tamiflu and Relenza.

    PubMed

    Le, Ly; Lee, Eric; Schulten, Klaus; Truong, Thanh N

    2009-01-01

    A molecular model of the swine influenza A/H1N1 ( also called H1N1pdm) type-I neuraminidase was built using the pathogenic avian H5N1 type-I neuraminidase as a basis, due to the higher sequence identity between A/H1N1 and H5N1 (91.47%) compared to Spanish H1N1 (88.37%) neuraminidase. All-atom molecular dynamics (MD) simulations of all three neuraminidases were performed, either as apo-structures or with commercial antiviral drugs Tamiflu or Relenza separately bound; the simulations allowed for the identification of both conserved and unique drug-protein interactions across all three proteins. Specifically, conserved networks of hydrogen bonds stabilizing the drugs in the sialic acid binding site of the simulated neuraminidases are analyzed, providing insight into how disruption due to mutations may lead to increased drug resistance. In addition, a possible mechanism through which the residue 294 mutation acquires drug resistance is proposed by mapping the mutation site onto an electrostatic pathway which may play a role in controlling drug access to the binding pocket of neuraminidase, establishing a starting point for further investigations of neuraminidase drug resistance. PMID:20029609

  7. Detection of reactive metabolites using isotope-labeled glutathione trapping and simultaneous neutral loss and precursor ion scanning with ultra-high-pressure liquid chromatography triple quadruple mass spectrometry.

    PubMed

    Huang, Ke; Huang, Lingyi; van Breemen, Richard B

    2015-04-01

    Metabolic activation of drugs to electrophilic species is responsible for over 60% of black box warnings and drug withdrawals from the market place in the United States. Reactive metabolite trapping using glutathione (GSH) and analysis using high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) or HPLC with high resolution mass spectrometry (mass defect filtering) have enabled screening for metabolic activation to become routine during drug development. However, current MS-based approaches cannot detect all GSH conjugates present in complex mixtures, especially those present in extracts of botanical dietary supplements. To overcome these limitations, a fast triple quadrupole mass spectrometer-based approach was developed that can detect positively and negatively charged GSH conjugates in a single analysis without the need for advanced knowledge of the elemental compositions of potential conjugates and while avoiding false positives. This approach utilized UHPLC instead of HPLC to shorten separation time and enhance sensitivity, incorporated stable-isotope labeled GSH to avoid false positives, and used fast polarity switching electrospray MS/MS to detect GSH conjugates that form positive and/or negative ions. The general new method was then used to test the licorice dietary supplement Glycyrrhiza glabra, which was found to form multiple GSH conjugates upon metabolic activation. Among the GSH conjugates found in the licorice assay were conjugates with isoliquiritigenin and glabridin, which is an irreversible inhibitor of cytochrome P450 enzymes. PMID:25774910

  8. Determination of 49 organophosphorus pesticide residues and their metabolites in fish, egg, and milk by dual gas chromatography-dual pulse flame photometric detection with gel permeation chromatography cleanup.

    PubMed

    Yang, Lixin; Li, Heli; Zeng, Fangang; Liu, Yinping; Li, Ruifeng; Chen, Huijing; Zhao, Yunfeng; Miao, Hong; Wu, Yongning

    2012-02-29

    A new method for the quantitative determination of 49 kinds of organophosphorus pesticide residues and their metabolites in fish, egg, and milk by dual gas chromatography-dual pulse flame photometric detection was developed. Homogenized samples were extracted with acetone and methylene chloride (1 + 1, v/v), and then the extracts were cleaned up by gel permeation chromatography (GPC). The response of each organophosphorus pesticide showed a good linearity with its concentration; the linearity correlation was not less than 0.99. The detection limits (S/N = 3) of pesticides were in the range of 0.001-0.025 mg kg⁻¹. The recovery experiments were performed by blank sample spiked at low, medium, and high fortification levels. The recoveries for fish, egg, and milk were 50.9-142.2, 53.3-137.2, and 50.3-139.4% with relative standard deviations (RSD, n = 6) of 2.3-24.9, 4.3-26.7, and 2.8-32.2%, respectively. The method was applied to detect organophosphorus pesticides in samples collected from the market, and satisfactory results were obtained. This quantitative method was highly sensitive and exact and could be applied to the accurate determination of organophosphorus contaminants in fish, egg, and milk. PMID:22300587

  9. Detection of Reactive Metabolites Using Isotope-Labeled Glutathione Trapping and Simultaneous Neutral Loss and Precursor Ion Scanning with Ultra-High-Pressure Liquid Chromatography Triple Quadrupole Mass Spectrometry

    PubMed Central

    Huang, Ke; Huang, Lingyi; van Breemen, Richard B.

    2015-01-01

    Metabolic activation of drugs to electrophilic species is responsible for over 60% of black box warnings and drug withdrawals from the market place in United States. Reactive metabolite trapping using glutathione (GSH) and analysis using high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) or HPLC with high resolution mass spectrometry (mass defect filtering) have enabled screening for metabolic activation to become routine during drug development. However, current MS-based approaches cannot detect all GSH conjugates present in complex mixtures, especially those present in extracts of botanical dietary supplements. To overcome these limitations, a fast triple quadrupole mass spectrometer-based approach was developed that can detect positively and negatively charged GSH conjugates in a single analysis without the need for advance knowledge of the elemental compositions of potential conjugates and while avoiding false positives. This approach utilized UHPLC instead of HPLC to shorten separation time and enhance sensitivity, incorporated stable-isotope labeled GSH to avoid false positives, and used fast polarity switching electrospray MS/MS to detect GSH conjugates that form positive and/or negative ions. The general new method was then used to test the licorice dietary supplement Glycyrrhiza glabra which was found to form multiple GSH conjugates upon metabolic activation. Among the GSH conjugates found in the licorice assay were conjugates with isoliquiritigenin and glabridin, which is an irreversible inhibitor of cytochrome P450 enzymes. PMID:25774910

  10. Quantification of free mycophenolic acid and its glucuronide metabolite in human plasma by liquid-chromatography using mass spectrometric and ultraviolet absorbance detection.

    PubMed

    Atcheson, Bronwyn; Taylor, Paul J; Mudge, David W; Johnson, David W; Pillans, Peter I; Tett, Susan E

    2004-01-01

    The immunosuppressant drug mycophenolic acid (MPA) and its major metabolite, mycophenolic acid glucuronide (MPAG), are highly bound to albumin. An HPLC-tandem-MS (HPLC/MS/MS) and an HPLC-UV assay were developed to measure free (unbound) concentrations of MPA and MPAG, respectively. Ultrafiltrate was prepared from plasma (500 microl) by ultrafiltration at 3000 x g for 20 min (20 degrees C). Both MPA and MPAG were isolated from ultrafiltrate (100 microl) by acidification and C18 solid-phase extraction. Free MPA was measured by electrospray tandem mass spectrometry using selected reactant monitoring (MPA: m/z 338.2--> 206.9) in positive ionisation mode. Chromatography was performed on a PFPP column (50 mm x 2 mm, 5 microm). Total analysis time was 7 min. The assay was linear over the range 1-200 microg/l with a limit of quantification of 1 microg/l. The inter-day accuracy and imprecision of quality controls (7.5, 40, 150 microg/l) were 94-99% and < 7%, respectively. Free MPAG was chromatographed on a C18 Nova-Pak column (150 mm x 3.9 mm, 5 microm) using a binary gradient over 20 min. The eluent was monitored at 254 nm. The assay was linear over the range 1-50 mg/l with the limit of quantification at 2.5 mg/l. The inter-day accuracy and imprecision of quality controls (5, 20, 45 mg/l) was 101-107% and < 8% (n = 4), respectively. For both methods no interfering substances were found in ultrafiltrate from patients not receiving MPA. The methods described have a suitable dynamic linear range to facilitate the investigation of free MPA and MPAG pharmacokinetics in transplant patients. Further, this is the first reported HPLC-UV method to determine free MPAG concentrations. PMID:14659448

  11. Maternal and Infant Urinary Phthalate Metabolite Concentrations: Are They Related?

    PubMed Central

    Sathyanarayana, S; Calafat, Antonia Maria; Liu, Fan; Swan, Shanna Helen

    2008-01-01

    Background Phthalates are synthetic chemicals that are ubiquitous in our society and may have adverse health effects in humans. Detectable concentrations of phthalate metabolites have been found in adults and children, but no studies have examined the relationship between maternal and infant phthalate metabolite concentrations. Objective We investigated the relationship between maternal and infant urinary phthalate metabolite concentrations. Methods We measured nine phthalate metabolites in urine samples from 210 mother/infant pairs collected on the same study visit day (1999–2005) and obtained demographic history from questionnaires. Using multivariate linear regression analyses, we examined the degree to which maternal urine phthalate metabolite concentration predicted infant phthalate metabolite concentration. All analyses were adjusted for infant age, creatinine concentration, and race. Results Correlation coefficients between phthalate metabolite concentrations in the urine of mothers and their infants were generally low but increased with decreasing age of infant. In multivariate analyses, mother’s phthalate metabolite concentrations were significantly associated with infants’ concentrations for six phthalate metabolites: monobenzyl phthalate, monoethyl phthalate, monoisobutyl phthalate, and three metabolites of di(2-ethylhexyl) phthalate: mono(2-ethylhexyl) phthalate, mono(2-ethyl-5-hydroxy-hexyl) phthalate and mono(2-ethyl-5-oxo-hexyl) phthalate (p-values for all coefficients <0.05). Discussion Mother’s urine phthalate metabolite concentration is significantly associated with infant urine phthalate metabolite concentration for six phthalate metabolites. It is plausible that shared exposures to phthalates in the immediate surrounding environment accounted for these relationships, but other unidentified sources may also contribute to infants’ phthalate exposures. This study indicates the importance of further identifying infant phthalate exposures

  12. Sensitive determination of three aconitum alkaloids and their metabolites in human plasma by matrix solid-phase dispersion with vortex-assisted dispersive liquid-liquid microextraction and HPLC with diode array detection.

    PubMed

    Wang, Xiaozhong; Li, Xuwen; Li, Lanjie; Li, Min; Liu, Ying; Wu, Qian; Li, Peng; Jin, Yongri

    2016-05-01

    A simple and sensitive method for determination of three aconitum alkaloids and their metabolites in human plasma was developed using matrix solid-phase dispersion combined with vortex-assisted dispersive liquid-liquid microextraction and high-performance liquid chromatography with diode array detection. The plasma sample was directly purified by matrix solid-phase dispersion and the eluate obtained was concentrated and further clarified by vortex-assisted dispersive liquid-liquid microextraction. Some important parameters affecting the extraction efficiency, such as type and amount of dispersing sorbent, type and volume of elution solvent, type and volume of extraction solvent, salt concentration as well as sample solution pH, were investigated in detail. Under optimal conditions, the proposed method has good repeatability and reproducibility with intraday and interday relative standard deviations lower than 5.44 and 5.75%, respectively. The recoveries of the aconitum alkaloids ranged from 73.81 to 101.82%, and the detection limits were achieved within the range of 1.6-2.1 ng/mL. The proposed method offered the advantages of good applicability, sensitivity, simplicity, and feasibility, which makes it suitable for the determination of trace amounts of aconitum alkaloids in human plasma samples. PMID:27005409

  13. Determination of two oxy-pyrimidine metabolites of diazinon in urine by gas chromatography/mass selective detection and liquid chromatography/electrospray ionization/mass spectrometry/mass spectrometry.

    PubMed

    Yokley, R A; Shen, N; Cheung, M W

    2000-01-01

    An analytical method was developed for the determination in urine of 2 metabolites of diazinon: 6-methyl-2-(1-methylethyl)-4(1H)-pyrimidinone (G-27550) and 2-(1-hydroxy-1-methylethyl)-6-methyl-4(1H)-pyrimidinone (GS-31144). Two of the urine sample preparation procedures presented rely on gas chromatography/mass selective detection (GC/MSD) in the selected ion monitoring mode for determination of G-27550. For fast sample preparation and a limit of quantitation (LOQ) of 1.0 ppb, urine samples were purified by using ENV+ solid-phase extraction (SPE) columns. For analyte confirmation at an LOQ of 0.50 ppb, classical liquid/liquid partitioning was used before further purification in a silica SPE column. An SPE sample preparation procedure and liquid chromatography/electrospray ionization/mass spectrometry/mass spectrometry (LC/ESI/MS/MS) were used for both G-27550 and GS-31144. The limit of detection was 0.01 ng for G-27550 with GC/MSD, and 0.016 ng when LC/ESI/MS/MS was used for both G-27550 and GS-31144. The LOQ was 0.50 ppb for G-27550 when GC/MSD and the partitioning/SPE sample preparation procedure were used, and 1.0 ppb for the SPE only sample preparation procedure. The LOQ was 1.0 ppb for both analytes when LC/ESI/MS/MS was used. PMID:11048865

  14. Characterizing Protein Modifications by Reactive Metabolites using Magnetic Bead Bioreactors and LC-MS/MS

    PubMed Central

    Li, Dandan; Fu, You-Jun; Rusling, James F.

    2015-01-01

    We report here label-free metabolite-protein adduct detection and identification employing magnetic beads coated with metabolic enzymes as bioreactors to generate metabolites and possible metabolite-protein adducts for analysis by liquid chromatography-tandem mass spectrometry. PMID:25693065

  15. Characterizing protein modifications by reactive metabolites using magnetic bead bioreactors and LC-MS/MS.

    PubMed

    Li, Dandan; Fu, You-Jun; Rusling, James F

    2015-03-18

    We report here label-free metabolite-protein adduct detection and identification employing magnetic beads coated with metabolic enzymes as bioreactors to generate metabolites and possible metabolite-protein adducts for analysis by liquid chromatography-tandem mass spectrometry. PMID:25693065

  16. Secondary metabolites from Ganoderma.

    PubMed

    Baby, Sabulal; Johnson, Anil John; Govindan, Balaji

    2015-06-01

    Ganoderma is a genus of medicinal mushrooms. This review deals with secondary metabolites isolated from Ganoderma and their biological significance. Phytochemical studies over the last 40years led to the isolation of 431 secondary metabolites from various Ganoderma species. The major secondary compounds isolated are (a) C30 lanostanes (ganoderic acids), (b) C30 lanostanes (aldehydes, alcohols, esters, glycosides, lactones, ketones), (c) C27 lanostanes (lucidenic acids), (d) C27 lanostanes (alcohols, lactones, esters), (e) C24, C25 lanostanes (f) C30 pentacyclic triterpenes, (g) meroterpenoids, (h) farnesyl hydroquinones (meroterpenoids), (i) C15 sesquiterpenoids, (j) steroids, (k) alkaloids, (l) prenyl hydroquinone (m) benzofurans, (n) benzopyran-4-one derivatives and (o) benzenoid derivatives. Ganoderma lucidum is the species extensively studied for its secondary metabolites and biological activities. Ganoderma applanatum, Ganoderma colossum, Ganoderma sinense, Ganoderma cochlear, Ganoderma tsugae, Ganoderma amboinense, Ganoderma orbiforme, Ganoderma resinaceum, Ganoderma hainanense, Ganoderma concinna, Ganoderma pfeifferi, Ganoderma neo-japonicum, Ganoderma tropicum, Ganoderma australe, Ganoderma carnosum, Ganoderma fornicatum, Ganoderma lipsiense (synonym G. applanatum), Ganoderma mastoporum, Ganoderma theaecolum, Ganoderma boninense, Ganoderma capense and Ganoderma annulare are the other Ganoderma species subjected to phytochemical studies. Further phytochemical studies on Ganoderma could lead to the discovery of hitherto unknown biologically active secondary metabolites. PMID:25975187

  17. Metabolite profiling of wheat (Triticum aestivum L.) phloem exudate

    PubMed Central

    2014-01-01

    Background Biofortification of staple crops with essential micronutrients relies on the efficient, long distance transport of nutrients to the developing seed. The main route of this transport in common wheat (Triticum aestivum) is via the phloem, but due to the reactive nature of some essential micronutrients (specifically Fe and Zn), they need to form ligands with metabolites for transport within the phloem. Current methods available in collecting phloem exudate allows for small volumes (μL or nL) to be collected which limits the breadth of metabolite analysis. We present a technical advance in the measurement of 79 metabolites in as little as 19.5 nL of phloem exudate. This was achieved by using mass spectrometry based, metabolomic techniques. Results Using gas chromatography–mass spectrometry (GC-MS), 79 metabolites were detected in wheat phloem. Of these, 53 were identified with respect to their chemistry and 26 were classified as unknowns. Using the ratio of ion area for each metabolite to the total ion area for all metabolites, 39 showed significant changes in metabolite profile with a change in wheat reproductive maturity, from 8–12 to 17–21 days after anthesis. Of these, 21 were shown to increase and 18 decreased as the plant matured. An amine group derivitisation method coupled with liquid chromatography MS (LC-MS) based metabolomics was able to quantify 26 metabolites and semi-quantitative data was available for a further 3 metabolites. Conclusions This study demonstrates that it is possible to determine metabolite profiles from extremely small volumes of phloem exudate and that this method can be used to determine variability within the metabolite profile of phloem that has occurred with changes in maturity. This is also believed to be the first report of the presence of the important metal complexing metabolite, nicotianamine in the phloem of wheat. PMID:25143779

  18. High fat diet leads to changes in metabolite patterns in pig plasma, fecal, and urine samples detected by a ultra-high performance liquid chromatography tandem with high resolution mass spectrometry metabolomic study

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Non-targeted metabolite profiling can identify robust biological markers of dietary exposure that can lead to a better understanding of causal interactions between diet and health. In this study, pigs were used as an animal model to develop an efficient procedure to discover metabolites in biolog...

  19. Understanding and Classifying Metabolite Space and Metabolite-Likeness

    PubMed Central

    Peironcely, Julio E.; Reijmers, Theo; Coulier, Leon; Bender, Andreas; Hankemeier, Thomas

    2011-01-01

    While the entirety of ‘Chemical Space’ is huge (and assumed to contain between 1063 and 10200 ‘small molecules’), distinct subsets of this space can nonetheless be defined according to certain structural parameters. An example of such a subspace is the chemical space spanned by endogenous metabolites, defined as ‘naturally occurring’ products of an organisms' metabolism. In order to understand this part of chemical space in more detail, we analyzed the chemical space populated by human metabolites in two ways. Firstly, in order to understand metabolite space better, we performed Principal Component Analysis (PCA), hierarchical clustering and scaffold analysis of metabolites and non-metabolites in order to analyze which chemical features are characteristic for both classes of compounds. Here we found that heteroatom (both oxygen and nitrogen) content, as well as the presence of particular ring systems was able to distinguish both groups of compounds. Secondly, we established which molecular descriptors and classifiers are capable of distinguishing metabolites from non-metabolites, by assigning a ‘metabolite-likeness’ score. It was found that the combination of MDL Public Keys and Random Forest exhibited best overall classification performance with an AUC value of 99.13%, a specificity of 99.84% and a selectivity of 88.79%. This performance is slightly better than previous classifiers; and interestingly we found that drugs occupy two distinct areas of metabolite-likeness, the one being more ‘synthetic’ and the other being more ‘metabolite-like’. Also, on a truly prospective dataset of 457 compounds, 95.84% correct classification was achieved. Overall, we are confident that we contributed to the tasks of classifying metabolites, as well as to understanding metabolite chemical space better. This knowledge can now be used in the development of new drugs that need to resemble metabolites, and in our work particularly for assessing the metabolite

  20. Analysis of selected herbicide metabolites in surface and ground water of the United States

    USGS Publications Warehouse

    Scribner, E.A.; Thurman, E.M.; Zimmerman, L.R.

    2000-01-01

    One of the primary goals of the US Geological Survey (USGS) Laboratory in Lawrence, Kansas, is to develop analytical methods for the analysis of herbicide metabolites in surface and ground water that are vital to the study of herbicide fate and degradation pathways in the environment. Methods to measure metabolite concentrations from three major classes of herbicides - triazine, chloroacetanilide and phenyl-urea - have been developed. Methods for triazine metabolite detection cover nine compounds: six compounds are detected by gas chromatography/mass spectrometry; one is detected by high-performance liquid chromatography with diode-array detection; and eight are detected by liquid chromatography/mass spectrometry. Two metabolites of the chloroacetanilide herbicides - ethane sulfonic acid and oxanilic acid - are detected by high-performance liquid chromatography with diode-array detection and liquid chromatography/mass spectrometry. Alachlor ethane sulfonic acid also has been detected by solid-phase extraction and enzyme-linked immunosorbent assay. Six phenylurea metabolites are all detected by liquid chromatography/mass spectrometry; four of the six metabolites also are detected by gas chromatography/mass spectrometry. Additionally, surveys of herbicides and their metabolites in surface water, ground water, lakes, reservoirs, and rainfall have been conducted through the USGS laboratory in Lawrence. These surveys have been useful in determining herbicide and metabolite occurrence and temporal distribution and have shown that metabolites may be useful in evaluation of non-point-source contamination. Copyright (C) 2000 Elsevier Science B.V.

  1. Detection of T-2 mycotoxin metabolites in urines of exposed rats. Comparison of a potentially fieldable kit with a laboratory assay. Interim report

    SciTech Connect

    Hewetson, J.F.; Wannemacher, R.W.; Hawley, R.J.

    1988-03-09

    Rapid methods to detect toxin exposure have been a concern of the Army since the reported use of T-2 mycotoxin as a biological warfare agent in Southeast Asia and Afghanistan. T-2 toxin was included in an exploratory development program of rapid identification systems for biological agents sponsored by the United States Army Medical Materiel Development Activity. Reported here is evidence of T-2W exposure in urines collected up to 2 weeks after rats were exposed to a sublethal dose of T-2 toxin. A laboratory radioimmunoassay (RIA) using polyclonal antibody was used to assay the urines for HT-2 or T-2 tetraol. The sensitivity of the RIA for HT-2 was 5 ng/ml and 50 ng/ml for T-2 tetraol. Some of the urines were assayed in parallel with a potentially fieldable enzyme-linked immunoassay (ELISA) developed for T-2 with a monoclonal antibody that cross reacts with HT-2.

  2. The quantitative spectrum of inositol phosphate metabolites in avian erythrocytes, analysed by proton n.m.r. and h.p.l.c. with direct isomer detection.

    PubMed Central

    Radenberg, T; Scholz, P; Bergmann, G; Mayr, G W

    1989-01-01

    The spectrum of inositol phosphate isomers present in avian erythrocytes was investigated in qualitative and quantitative terms. Inositol phosphates were isolated in micromolar quantities from turkey blood by anion-exchange chromatography on Q-Sepharose and subjected to proton n.m.r. and h.p.l.c. analysis. We employed a h.p.l.c. technique with a novel, recently described complexometric post-column detection system, called 'metal-dye detection' [Mayr (1988) Biochem. J. 254, 585-591], which enabled us to identify non-radioactively labelled inositol phosphate isomers and to determine their masses. The results indicate that avian erythrocytes contain the same inositol phosphate isomers as mammalian cells. Denoted by the 'lowest-locant rule' [NC-IUB Recommendations (1988) Biochem. J. 258, 1-2] irrespective of true enantiomerism, these are Ins(1,4)P2, Ins(1,6)P2, Ins(1,3,4)P3, Ins(1,4,5)P3, Ins(1,3,4,5)P4, Ins(1,3,4,6)P4, Ins(1,4,5,6)P4, Ins(1,3,4,5,6)P5, and InsP6. Furthermore, we identified two inositol trisphosphate isomers hitherto not described for mammalian cells, namely Ins(1,5,6)P3 and Ins(2,4,5)P3. The possible position of these two isomers in inositol phosphate metabolism and implications resulting from absolute abundances of inositol phosphates are discussed. PMID:2604720

  3. Sample preparation methods for LC-MS-based global aqueous metabolite profiling.

    PubMed

    Beltran, Antoni; Samino, Sara; Yanes, Oscar

    2014-01-01

    Metabolite extraction is a key step in metabolomic analyses, particularly for untargeted studies. The extraction determines the types of metabolites that will be detected and the analytical platform to be used. In this chapter we describe two protocols aimed at detecting polar metabolites from biological samples; the first is aimed at detecting reduced species by LC/MS, and the second satisfies the requirements for both NMR and LC/MS analysis simultaneously. PMID:25270923

  4. Microalgal metabolites: a new perspective.

    PubMed

    Shimizu, Y

    1996-01-01

    Occurrence of secondary metabolites in microalgae (protoctista) is discussed with respect to the phylogenic or taxonomic relationships of organisms. Biosynthetic mechanisms of certain metabolites such as paralytic shellfish poisoning toxins and polyether toxins are also discussed, and genetic aspects of the secondary metabolite production as well. PMID:8905087

  5. Metabolite Damage and Metabolite Damage Control in Plants.

    PubMed

    Hanson, Andrew D; Henry, Christopher S; Fiehn, Oliver; de Crécy-Lagard, Valérie

    2016-04-29

    It is increasingly clear that (a) many metabolites undergo spontaneous or enzyme-catalyzed side reactions in vivo, (b) the damaged metabolites formed by these reactions can be harmful, and (c) organisms have biochemical systems that limit the buildup of damaged metabolites. These damage-control systems either return a damaged molecule to its pristine state (metabolite repair) or convert harmful molecules to harmless ones (damage preemption). Because all organisms share a core set of metabolites that suffer the same chemical and enzymatic damage reactions, certain damage-control systems are widely conserved across the kingdoms of life. Relatively few damage reactions and damage-control systems are well known. Uncovering new damage reactions and identifying the corresponding damaged metabolites, damage-control genes, and enzymes demands a coordinated mix of chemistry, metabolomics, cheminformatics, biochemistry, and comparative genomics. This review illustrates the above points using examples from plants, which are at least as prone to metabolite damage as other organisms. PMID:26667673

  6. Ex vivo studies for the passive transdermal delivery of low-dose naltrexone from a cream; detection of naltrexone and its active metabolite, 6β-naltrexol, using a novel LC Q-ToF MS assay.

    PubMed

    Dodou, Kalliopi; Armstrong, Andrew; Kelly, Ivan; Wilkinson, Simon; Carr, Kevin; Shattock, Paul; Whiteley, Paul

    2015-01-01

    Naltrexone (NTX) is a long-acting opiate antagonist. Low-dose naltrexone (LDN) therapy has shown promising results in the treatment of several autoimmune disorders. Our aim was to formulate NTX into a cream for the delivery of LDN and develop an analytical technique for the quantification of NTX and its active metabolite 6-β-naltrexol (NTXol) during transdermal diffusion cell permeation studies. A 1% w/w NTX cream was formulated and drug permeation was examined over 24 h using static Franz diffusion cells mounted with pig skin. A Liquid Chromatography Quadrupole-Time of Flight Mass Spectrometry (LC-MS Q-ToF) method was developed for the detection of NTX and NTXol in the receptor solution, skin membrane and residual cream on the donor chamber after completion of the diffusion studies. The cream formulation exhibited steady state release of NTX over 24 h after an initial lag time of 2.74 h. The bioconversion of NTX to NTXol in the skin membrane was 1.1%. It was concluded that the cream may be an effective formulation for the sustained transdermal delivery of LDN. The novel LC Q-ToF MS method allowed the accurate measurement of NTX and NTXol levels across the diffusion cell assemblies and the quantification of NTX metabolism in the skin. PMID:24785567

  7. The syntheses of [(14) C]BMS-823778 for use in a human ADME clinical study and of [(13) CD3 (13) CD2 ]BMT-094817, a stable-isotope labeled standard of a newly detected human metabolite.

    PubMed

    Maxwell, Brad D; Tran, Scott B; Lago, Michael; Li, Jun; Bonacorsi, Samuel J

    2016-05-30

    Type 2 diabetes is a significant worldwide health problem. To support the development of BMS-823778 as an inhibitor of 11β-hydroxysteroid dehydrogenase type 1 for type 2 diabetes, the synthesis of carbon-14-labeled material was required for use in a human adsorption, distribution, metabolism, and excretion (ADME) study. The HCl salt form of [(14) C]BMS-823778 was synthesized in two steps from commercially available [2-(14) C]acetone. The radiochemical purity of the synthesized [(14) C]BMS-823778 after dilution with unlabeled clinical-grade BMS-823778 was 99.5% having a specific activity of 7.379 μCi/mg. One result of the human ADME study was the detection of a new human metabolite, BMT-094817. To support the quantification of BMT-094817 in clinical samples, it was necessary to synthesize [(13) CD3 (13) CD2 ]BMT-094817 for use as a liquid chromatography/mass spectrometry standard. [(13) CD3 (13) CD2 ]BMT-094817 was prepared in five labeled steps from [(13) CD3 ]iodomethane. PMID:27169762

  8. Development and validation of a multiresidue method for the simultaneous determination of organophosphorus insecticides and their toxic metabolites in sugarcane juice and refined sugar by gas chromatography with flame photometric detection.

    PubMed

    Ramasubramanian, Thirumalaiandi; Paramasivam, Mariappan

    2016-06-01

    A multiresidue method has been developed and validated for the simultaneous determination of organophosphorus insecticides and their toxic metabolites in sugarcane juice and refined sugar by gas chromatography with flame photometric detection. Limits of quantification of the method varied between 0.007 and 0.01 μg/g. Ethyl acetate based extraction followed by dispersive solid-phase extraction cleanup with primary secondary amine yielded internationally acceptable recoveries of acephate, chlorpyrifos, dichlorvos, monocrotophos, malathion, malaoxon, phorate, phorate-sulfoxide, phorate-oxon, phorate-sulfone, and quinalphos from selected matrices. The recoveries of target analytes from cane juice were 75.55 ± 0.5-102.57 ± 4.2, 77.45 ± 4.7-103.33 ± 3.3, and 80.55 ± 6.6-105.82 ± 9.8% at 0.01, 0.02, and 0.1 μg/g levels of fortification, respectively. The recoveries from cane sugar were 73.24 ± 3.5-104.47 ± 1.9, 75.23 ± 1.5-116.10 ± 3.7, and 70.75 ± 5.7-110.15 ± 2.7%, respectively at 0.01, 0.02, and 0.1 μg/g levels of fortification. Matrix effect and measurement uncertainty were within the permissible limit (less than 20%) as prescribed for pesticide residue analysis. PMID:27061678

  9. Pressurized liquid extraction for the determination of cannabinoids and metabolites in hair: Detection of cut-off values by high performance liquid chromatography-high resolution tandem mass spectrometry.

    PubMed

    Montesano, Camilla; Simeoni, Maria Chiara; Vannutelli, Gabriele; Gregori, Adolfo; Ripani, Luigi; Sergi, Manuel; Compagnone, Dario; Curini, Roberta

    2015-08-01

    Hair analysis has become a routine procedure in most forensic laboratories since this alternative matrix presents clear advantages over classical matrices; particularly wider time window, non-invasive sampling and good stability of the analytes over time. There are, however, some major challenges for the analysis of cannabinoids in hair, mainly related to the low concentrations of 11-nor-9-carboxy-Δ(9)-tetrahydrocannabinol (THC-COOH), that is the major metabolite. In this study a fast, accurate and sensitive method for the determination of cannabinol, cannabidiol, THC and THC-COOH in hair has been developed. The extraction of analytes from hair (50mg) is based on an automated pressurized liquid extraction (PLE) using water modified with the surfactant sodium dodecyl sulphate as eluent phase. PLE extract is then cleaned up by SPE using polymeric reversed phase cartridges Strata XL before the injection in the HPLC-HRMS/MS system. Chromatographic conditions obtained with a fused-core column allowed a good separation of the analytes in less than 4min. The whole procedure has been validated according to SWGTOX guidelines. The LLOQs obtained for THC-COOH and the other analytes were respectively 0.1 and 2pg/mg. To the best of our knowledge, this is the first LC-MS/MS based method that allows the detection of THC-COOH in hair at values lower than the cut-off. PMID:26118805

  10. Detection of three herbicide, and one metabolite, residues in brown rice and rice straw using various versions of the QuEChERS method and liquid chromatography-tandem mass spectrometry.

    PubMed

    Lee, Young-Jun; Rahman, Md Musfiqur; Abd El-Aty, A M; Choi, Jeong-Heui; Chung, Hyung Suk; Kim, Sung-Woo; Abdel-Aty, Azza M; Shin, Ho-Chul; Shim, Jae-Han

    2016-11-01

    A single-run analytical method was developed to analyze the three herbicides azimsulfuron, bensulfuron-methyl, and mesotrione and its metabolite (4-methylsulfonyl-2-nitrobenzoic acid (MNBA)) in brown rice and rice straw using liquid chromatography-tandem mass spectrometry (LC/MS/MS). Samples extracted using various versions of Quick, Easy, Cheap, Effective, Rugged, and Safe "QuEChERS" (original unbuffered, acetate (AOAC), and citrate (EN) buffered) methods gave poor recoveries of all the tested analytes in both matrices. The extraction efficiency was improved when primary-secondary amine (PSA) sorbent was removed from the purification step, with the best recovery being achieved for EN-QuEChERS, which was subsequently used throughout the study. Overall, a determination coefficients (R(2))⩾0.995 was achieved at matrix-matched calibration curves at various concentration ranges. The recovery rates at three fortification levels (limit of quantification (LOQ), 1/2 maximum residue limit (1/2MRL), and MRL) ranged from 78 to 114.5, with relative standard deviations (RSDs)<18% for all the tested analytes in both matrices. The LOQs for all herbicides were lower than the MRL set by the Ministry of Food and Drug Safety (MFDS), Republic of Korea. Field trials with the recommended, or double the recommended dose, revealed that the herbicides can safely be applied to rice, as no residues were detected in the harvested samples at 110days. PMID:27211669

  11. Metabolic activation of 1,2-dibromo-3-chloropropane to mutagenic metabolites: detection and mechanism of formation of (Z)- and (E)-2-chloro-3-(bromomethyl)oxirane.

    PubMed

    Pearson, P G; Omichinski, J G; Myers, T G; Søderlund, E J; Dybing, E; Nelson, S D

    1990-01-01

    1,2-Dibromo-3-chloropropane (DBCP), a haloalkane nematocide and soil fumigant, is metabolically activated to chemically reactive species that are direct-acting mutagens in a Salmonella typhimurium TA 100 test system. Studies in vitro with rat liver microsomes indicated that oxidation at carbon 3 resulted in the formation of an unstable gem-chlorohydrin that rearranged with elimination of hydrogen bromide to form (Z)-2-chloro-3-(bromomethyl)oxirane [(Z)-CBPO] and (E)-2-chloro-3-(bromomethyl)oxirane [(E)-CBPO]. Gas chromatography-mass spectrometry (GC-MS) with positive ion chemical ionization (CI) was employed to identify (Z)-CBPO and (E)-CBPO by comparison of characteristic fragment ions in their CI mass spectra with those observed for authentic standards. Quantitative GC-MS methodology was exploited to quantitate the rate of formation of (Z)-CBPO and (E)-CBPO from DBCP and analogues of DBCP specifically deuterated at carbon 1 and carbon 3. The rate of formation of Z- and E-isomers of CBPO was 31 and 33 pmol/(min.mg of protein), respectively, from DBCP; substitution with deuterium at carbon 1 increased the rate of epoxide formation by 50%, whereas CBPO formation could not be detected from a substrate labeled with deuterium at carbon 3. Both epoxides were directly acting mutagens to S. typhimurium TA 100. (Z)-CBPO caused approximately twice as many his+ revertants/nmol compared to (E)-CBPO. Oxidation at carbon 2 of DBCP resulted in the formation of a bifunctional alkylating agent, 1-bromo-3-chloroacetone, presumably via the intermediacy of an unstable gem-bromohydrin.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:2133097

  12. Urinary pesticide metabolites in school students from northern Thailand.

    PubMed

    Panuwet, Parinya; Prapamontol, Tippawan; Chantara, Somporn; Barr, Dana B

    2009-05-01

    We evaluated exposure to pesticides among secondary school students aged 12-13 years old in Chiang Mai Province, Thailand. Pesticide-specific urinary metabolites were used as biomarkers of exposure for a variety of pesticides, including organophosphorus insecticides, synthetic pyrethroid insecticides and selected herbicides. We employed a simple solid-phase extraction with analysis using isotope dilution high-performance liquid chromatography tandem mass spectrometry (HPLC-MS/MS). A total of 207 urine samples from Thai students were analyzed for 18 specific pesticide metabolites. We found 14 metabolites in the urine samples tested; seven of them were detected with a frequency > or=17%. The most frequently detected metabolites were 2-[(dimethoxyphosphorothioyl) sulfanyl] succinic acid (malathion dicarboxylic acid), para-nitrophenol (PNP), 3,5,6-trichloro-2-pyridinol (TPCY; metabolite of chlorpyrifos), 2,4-dichlorophenoxyacetic acid (2,4-D), cis- and trans-3-(2,2-dichlorovinyl)-2,2-dimethylcyclopropane-1-carboxylic acids (c-DCCA and t-DCCA; metabolite of permethrin) and 3-phenoxybenzoic acid (3-PBA; metabolite of pyrethroids). The students were classified into 4 groups according to their parental occupations: farmers (N=60), merchants and traders (N=39), government and company employees (N=52), and laborers (N=56). Children of farmers had significantly higher urinary concentrations of pyrethroid insecticide metabolites than did other children (p<0.05). Similarly, children of agricultural families had significantly higher pyrethroid metabolite concentrations. Males had significantly higher values of PNP (Mann-Whitney test, p=0.009); however, no other sex-related differences were observed. Because parental occupation and agricultural activities seemed to have little influence on pesticide levels, dietary sources were the likely contributors to the metabolite levels observed. PMID:18760967

  13. Profiling Reactive Metabolites via Chemical Trapping and Targeted Mass Spectrometry.

    PubMed

    Chang, Jae Won; Lee, Gihoon; Coukos, John S; Moellering, Raymond E

    2016-07-01

    Metabolomic profiling studies aim to provide a comprehensive, quantitative, and dynamic portrait of the endogenous metabolites in a biological system. While contemporary technologies permit routine profiling of many metabolites, intrinsically labile metabolites are often improperly measured or omitted from studies due to unwanted chemical transformations that occur during sample preparation or mass spectrometric analysis. The primary glycolytic metabolite 1,3-bisphosphoglyceric acid (1,3-BPG) typifies this class of metabolites, and, despite its central position in metabolism, has largely eluded analysis in profiling studies. Here we take advantage of the reactive acylphosphate group in 1,3-BPG to chemically trap the metabolite with hydroxylamine during metabolite isolation, enabling quantitative analysis by targeted LC-MS/MS. This approach is compatible with complex cellular metabolome, permits specific detection of the reactive (1,3-) instead of nonreactive (2,3-) BPG isomer, and has enabled direct analysis of dynamic 1,3-BPG levels resulting from perturbations to glucose processing. These studies confirmed that standard metabolomic methods misrepresent cellular 1,3-BPG levels in response to altered glucose metabolism and underscore the potential for chemical trapping to be used for other classes of reactive metabolites. PMID:27314642

  14. NMR identification of endogenous metabolites interacting with fatted and non-fatted human serum albumin in blood plasma: Fatty acids influence the HSA-metabolite interaction

    NASA Astrophysics Data System (ADS)

    Jupin, Marc; Michiels, Paul J.; Girard, Frederic C.; Spraul, Manfred; Wijmenga, Sybren S.

    2013-03-01

    Metabolites and their concentrations are direct reporters on body biochemistry. Thanks to technical developments metabolic profiling of body fluids, such as blood plasma, by for instance NMR has in the past decade become increasingly accurate enabling successful clinical diagnostics. Human Serum Albumin (HSA) is the main plasma protein (˜60% of all plasma protein) and responsible for the transport of endogenous (e.g. fatty acids) and exogenous metabolites, which it achieves thanks to its multiple binding sites and its flexibility. HSA has been extensively studied with regard to its binding of drugs (exogenous metabolites), but only to a lesser extent with regard to its binding of endogenous (non-fatty acid) metabolites. To obtain correct NMR measured metabolic profiles of blood plasma and/or potentially extract information on HSA and fatty acids content, it is necessary to characterize these endogenous metabolite/plasma protein interactions. Here, we investigate these metabolite-HSA interactions in blood plasma and blood plasma mimics. The latter contain the roughly twenty metabolites routinely detected by NMR (also most abundant) in normal relative concentrations with fatted or non-fatted HSA added or not. First, we find that chemical shift changes are small and seen only for a few of the metabolites. In contrast, a significant number of the metabolites display reduced resonance integrals and reduced free concentrations in the presence of HSA or fatted HSA. For slow-exchange (or strong) interactions, NMR resonance integrals report the free metabolite concentration, while for fast exchange (weak binding) the chemical shift reports on the binding. Hence, these metabolites bind strongly to HSA and/or fatted HSA, but to a limited degree because for most metabolites their concentration is smaller than the HSA concentration. Most interestingly, fatty acids decrease the metabolite-HSA binding quite significantly for most of the interacting metabolites. We further find

  15. Variability of Non-Polar Secondary Metabolites in the Red Alga Portieria

    PubMed Central

    Payo, Dioli Ann; Colo, Joannamel; Calumpong, Hilconida; de Clerck, Olivier

    2011-01-01

    Possible sources of variation in non-polar secondary metabolites of Portieria hornemannii, sampled from two distinct regions in the Philippines (Batanes and Visayas), resulting from different life-history stages, presence of cryptic species, and/or spatiotemporal factors, were investigated. PCA analyses demonstrated secondary metabolite variation between, as well as within, five cryptic Batanes species. Intraspecific variation was even more pronounced in the three cryptic Visayas species, which included samples from six sites. Neither species groupings, nor spatial or temporal based patterns, were observed in the PCA analysis, however, intraspecific variation in secondary metabolites was detected between life-history stages. Male gametophytes (102 metabolites detected) were strongly discriminated from the two other stages, whilst female gametophyte (202 metabolites detected) and tetrasporophyte (106 metabolites detected) samples were partially discriminated. These results suggest that life-history driven variations, and possibly other microscale factors, may influence the variation within Portieria species. PMID:22163195

  16. Characterization and tissue distribution of conjugated metabolites of pyrene in the rat

    PubMed Central

    SAENGTIENCHAI, Aksorn; IKENAKA, Yoshinori; DARWISH, Wageh Sobhy; NAKAYAMA, Shouta M.M.; MIZUKAWA, Hazuki; ISHIZUKA, Mayumi

    2015-01-01

    Pyrene (PY) is a polycyclic aromatic hydrocarbon (PAH) that is often used as a biomarker for human and wildlife exposure to PAHs. As the metabolites of PAHs, similar to their parent compounds, pose public health risks, it is necessary to study their characteristics and tissue-specific distribution. The present study was performed to experimentally characterize PY metabolites and analyze the tissue-specific distribution of the conjugated metabolites after oral administration of PY to rats. PY metabolites, such as pyrenediol-disulfate (PYdiol-diS), pyrenediol-sulfate (PYdiol-S), pyrene-1-sufate (PYOS), pyrene-1-glucuronide (PYOG) and 1-hydroxypyrene (PYOH), were detected in rat urine. Although glucuronide conjugate was the predominant metabolite, the metabolite composition varied among tissues. Interestingly, the proportion of PYOH was high in the large intestine. Furthermore, PYOH was the only PY metabolite detected in feces. PMID:26028020

  17. Functional Genomics of Novel Secondary Metabolites from Diverse Cyanobacteria Using Untargeted Metabolomics

    PubMed Central

    Baran, Richard; Ivanova, Natalia N.; Jose, Nick; Garcia-Pichel, Ferran; Kyrpides, Nikos C.; Gugger, Muriel; Northen, Trent R.

    2013-01-01

    Mass spectrometry-based metabolomics has become a powerful tool for the detection of metabolites in complex biological systems and for the identification of novel metabolites. We previously identified a number of unexpected metabolites in the cyanobacterium Synechococcus sp. PCC 7002, such as histidine betaine, its derivatives and several unusual oligosaccharides. To test for the presence of these compounds and to assess the diversity of small polar metabolites in other cyanobacteria, we profiled cell extracts of nine strains representing much of the morphological and evolutionary diversification of this phylum. Spectral features in raw metabolite profiles obtained by normal phase liquid chromatography coupled to mass spectrometry (MS) were manually curated so that chemical formulae of metabolites could be assigned. For putative identification, retention times and MS/MS spectra were cross-referenced with those of standards or available sprectral library records. Overall, we detected 264 distinct metabolites. These included indeed different betaines, oligosaccharides as well as additional unidentified metabolites with chemical formulae not present in databases of metabolism. Some of these metabolites were detected only in a single strain, but some were present in more than one. Genomic interrogation of the strains revealed that generally, presence of a given metabolite corresponded well with the presence of its biosynthetic genes, if known. Our results show the potential of combining metabolite profiling and genomics for the identification of novel biosynthetic genes. PMID:24084783

  18. [Antiviral properties of basidiomycetes metabolites].

    PubMed

    Avtonomova, A V; Krasnopolskaya, L M

    2014-01-01

    The data on the antiviral action of the Ganoderma lucidum, Lentinus edodes, Grifola frondosa, Agaricus brasiliensis and other basidiomycetes metabolites are summurized. The metabolites of these species of basidiomycetes exhibit a direct antiviral effect on herpes simplex virus types I and II, human immunodeficiency virus (HIV), hepatitis B virus, vesicular stomatitis virus, influenza virus, Epstein-Barr virus, and others. Moreover, metabolites of basidiomycetes increased antiviral immunity. PMID:25975107

  19. Pryogalloloestrogens -- a new group of oestrogen metabolites.

    PubMed

    Stubenrauch, G; Gelbke, H P; Knuppen, R

    1976-01-01

    After incubation of radioactive catecholoestrogen monomethyl ethers with rat liver slices the following well known metabolic pathways were observed: 1) demethylation, 2) 16alpha-hydroxylation, 3) oxidoreduction at C-atom 17, and 4) conjugation with glutathione, sulphuric acid and glucuronic acid. In addition, for the first time a further aromatic ortho-hydroxylation, leading to pyrogalloloestrogen derivatives, was detected. Thus, the incubation of 2-methoxyoestrone yielded 2,4-dihydroxyoestrone 2-methyl ether as the main metabolite of the lipophile fraction. Under the same conditions, 4-methoxyoestrone was converted to 2,4-dihydroxyoestrone 4-methyl ether and 2,4-dihydroxyoestradiol-17beta 4-methyl ether; these compounds were the quantitatively most important metabolites not only in the lipophile but also in the sulphate and glucuronide fractions. The identity of these new metabolic products was established by chromatography, microchemical reactions and recrystallisation to constant specific radioactivity. PMID:1248801

  20. Sun, shade, and secondary metabolites

    Technology Transfer Automated Retrieval System (TEKTRAN)

    My research program focuses on understanding plant primary and secondary metabolites. Grape secondary metabolites, such as phenolics, have long been valuable for the organoleptic properties they impart to fruit and wine, and, more recently, for their possible health benefits. These compounds develop...

  1. Gas chromatography with pulsed flame photometric detection multiresidue method for organophosphate pesticide and metabolite residues at the parts-per-billion level in representatives commodities of fruits and vegetable crop groups.

    PubMed

    Podhorniak, L V; Negron, J F; Griffith, F D

    2001-01-01

    A gas chromatographic method with a pulsed flame photometric detector (P-FPD) is presented for the analysis of 28 parent organophosphate (OP) pesticides and their OP metabolites. A total of 57 organophosphates were analyzed in 10 representative fruit and vegetable crop groups. The method is based on a judicious selection of known procedures from FDA sources such as the Pesticide Analytical Manual and Laboratory Information Bulletins, combined in a manner to recover the OPs and their metabolite(s) at the part-per-billion (ppb) level. The method uses an acetone extraction with either miniaturized Hydromatrix column partitioning or alternately a miniaturized methylene dichloride liquid-liquid partitioning, followed by solid-phase extraction (SPE) cleanup with graphitized carbon black (GCB) and PSA cartridges. Determination of residues is by programmed temperature capillary column gas chromatography fitted with a P-FPD set in the phosphorus mode. The method is designed so that a set of samples can be prepared in 1 working day for overnight instrumental analysis. The recovery data indicates that a daily column-cutting procedure used in combination with the SPE extract cleanup effectively reduces matrix enhancement at the ppb level for many organophosphates. The OPs most susceptible to elevated recoveries around or greater than 150%, based on peak area calculations, were trichlorfon, phosmet, and the metabolites of dimethoate, fenamiphos, fenthion, and phorate. PMID:11417651

  2. Spatio-temporal distribution and natural variation of metabolites in citrus fruits.

    PubMed

    Wang, Shouchuang; Tu, Hong; Wan, Jian; Chen, Wei; Liu, Xianqing; Luo, Jie; Xu, Juan; Zhang, Hongyan

    2016-05-15

    To study the natural variation and spatio-temporal accumulation of citrus metabolites, liquid chromatography tandem mass spectrometry (LC-MS) based metabolome analysis was performed on four fruit tissues (flavedo, albedo, segment membrane and juice sacs) and different Citrus species (lemon, pummelo and grapefruit, sweet orange and mandarin). Using a non-targeted metabolomics approach, more than 2000 metabolite signals were detected, from which more than 54 metabolites, including amino acids, flavonoids and limonoids, were identified/annotated. Differential accumulation patterns of both primary metabolites and secondary metabolites in various tissues and species were revealed by our study. Further investigation indicated that flavedo accumulates more flavonoids while juice sacs contain more amino acids. Besides this, cluster analysis based on the levels of metabolites detected in 47 individual Citrus accessions clearly grouped them into four distinct clusters: pummelos and grapefruits, lemons, sweet oranges and mandarins, while the cluster of pummelos and grapefruits lay distinctly apart from the other three species. PMID:26775938

  3. Synthesis Of Labeled Metabolites

    DOEpatents

    Martinez, Rodolfo A.; Silks, III, Louis A.; Unkefer, Clifford J.; Atcher, Robert

    2004-03-23

    The present invention is directed to labeled compounds, for example, isotopically enriched mustard gas metabolites including: [1,1',2,2'-.sup.13 C.sub.4 ]ethane, 1,1'-sulfonylbis[2-(methylthio); [1,1',2,2'-.sup.13 C.sub.4 ]ethane, 1-[[2-(methylsulfinyl)ethyl]sulfonyl]-2-(methylthio); [1,1',2,2'-.sup.13 C.sub.4 ]ethane, 1,1'-sulfonylbis[2-(methylsulfinyl)]; and, 2,2'-sulfinylbis([1,2-.sup.13 C.sub.2 ]ethanol of the general formula ##STR1## where Q.sup.1 is selected from the group consisting of sulfide (--S--), sulfone (--S(O)--), sulfoxide (--S(O.sub.2)--) and oxide (--O--), at least one C* is .sup.13 C, X is selected from the group consisting of hydrogen and deuterium, and Z is selected from the group consisting of hydroxide (--OH), and --Q.sup.2 --R where Q.sup.2 is selected from the group consisting of sulfide (--S--), sulfone(--S(O)--), sulfoxide (--S(O.sub.2)--) and oxide (--O--), and R is selected from the group consisting of hydrogen, a C.sub.1 to C.sub.4 lower alkyl, and amino acid moieties, with the proviso that when Z is a hydroxide and Q.sup.1 is a sulfide, then at least one X is deuterium.

  4. Structural transformation of lignan compounds in rat gastrointestinal tract; II. Serum concentration of lignans and their metabolites.

    PubMed

    Nose, M; Fujimoto, T; Nishibe, S; Ogihara, Y

    1993-04-01

    Serum concentrations of arctiin, tracheloside, and their metabolites formed in the gastrointestinal tract were investigated in the rat. Arctiin or tracheloside was not detected in the serum after oral administration (200 mg/kg). In regard to their metabolites, each metabolite 1 (AM1, TM1), their genuine genins, appeared in the serum, and the serum concentration of arctiin metabolite 1 (AM1) reached its peak at 4 h and that of tracheloside metabolite 1 (TM1) reached its peak at 8 h. On the other hand, both metabolites 2 (AM2, TM2), which each possess a catechol moiety as reported previously, were not found in the serum. Now, we have studied the detection of their metabolites in the rat large intestinal contents after oral administration. It was revealed that all metabolites reported previously were certainly formed in rat gastrointestinal tract in vivo. Thus, we presumed a possibility that metabolite 2 was converted into metabolite 1 through C-3" methylation by catechol-O-methyltransferase (COMT) in rat liver. Each metabolite 2 was incubated with rat liver cytosol in the presence of S-adenosyl-L-methionine. It was proved that metabolite 2 was rapidly converted into metabolite 1 within 3 min. We suggest that arctiin or tracheloside was transformed to at least two metabolites in the gastrointestinal tract, and after absorption from the intestine, metabolite 2 was converted into metabolite 1 through methylation by COMT in the liver, and arctiin and tracheloside existed as metabolite 1, the genuine genin, in the blood stream. PMID:8387675

  5. The metabolite profiling of coastal coccolithophorid species Pleurochrysis carterae (Haptophyta)

    NASA Astrophysics Data System (ADS)

    Zhou, Chengxu; Luo, Jie; Ye, Yangfang; Yan, Xiaojun; Liu, Baoning; Wen, Xin

    2015-11-01

    Pleurochrysis carterae is a calcified coccolithophorid species that usually blooms in the coastal area and causes aquaculture losses. The cellular calcification, blooming and many other critical species specific eco-physiological processes are closely related to various metabolic pathways. The purpose of this study is to apply the unbiased and non-destructive method of nuclear magnetic resonance (NMR) to detect the unknown holistic metabolite of P. carterae. The results show that NMR spectroscopic method is practical in the analysis of metabolites of phytoplankton. The metabolome of P. carterae was dominated by 26 metabolites involved in a number of different primary and secondary metabolic pathways. Organic acids and their derivatives, amino acids, sugars, nucleic aides were mainly detected. The abundant metabolites are that closely related to the process of cellular osmotic adjustment, which possibly reflect the active ability of P. carterae to adapt to the versatile coastal niche. DMSP (dimethylsulphoniopropionate) was the most dominant metabolite in P. carterae, up to 2.065±0.278 mg/g lyophilized cells, followed by glutamate and lactose, the contents were 0.349±0.035 and 0.301±0.073 mg/g lyophilized cells respectively. Other metabolites that had the content ranged between 0.1-0.2 mg/g lyophilized cells were alanine, isethionate and arabinose. Amino acid (valine, phenylalanine, isoleucine, tyrosine), organic acid salts (lactate, succinate), scyllitol and uracil had content ranged from 0.01 to below 0.1 mg/g lyophilized cells. Trigonelline, fumarate and formate were detected in very low content (only thousandths of 1 mg per gram of lyophilized cells or below). Our results of the holistic metabolites of P. carterae are the basic references for the further studies when multiple problems will be addressed to this notorious blooming calcifying species.

  6. The metabolite profiling of coastal coccolithophorid species Pleurochrysis carterae (Haptophyta)

    NASA Astrophysics Data System (ADS)

    Zhou, Chengxu; Luo, Jie; Ye, Yangfang; Yan, Xiaojun; Liu, Baoning; Wen, Xin

    2016-07-01

    Pleurochrysis carterae is a calcified coccolithophorid species that usually blooms in the coastal area and causes aquaculture losses. The cellular calcification, blooming and many other critical species specific eco-physiological processes are closely related to various metabolic pathways. The purpose of this study is to apply the unbiased and non-destructive method of nuclear magnetic resonance (NMR) to detect the unknown holistic metabolite of P. carterae. The results show that NMR spectroscopic method is practical in the analysis of metabolites of phytoplankton. The metabolome of P. carterae was dominated by 26 metabolites involved in a number of different primary and secondary metabolic pathways. Organic acids and their derivatives, amino acids, sugars, nucleic aides were mainly detected. The abundant metabolites are that closely related to the process of cellular osmotic adjustment, which possibly reflect the active ability of P. carterae to adapt to the versatile coastal niche. DMSP (dimethylsulphoniopropionate) was the most dominant metabolite in P. carterae, up to 2.065±0.278 mg/g lyophilized cells, followed by glutamate and lactose, the contents were 0.349±0.035 and 0.301±0.073 mg/g lyophilized cells respectively. Other metabolites that had the content ranged between 0.1-0.2 mg/g lyophilized cells were alanine, isethionate and arabinose. Amino acid (valine, phenylalanine, isoleucine, tyrosine), organic acid salts (lactate, succinate), scyllitol and uracil had content ranged from 0.01 to below 0.1 mg/g lyophilized cells. Trigonelline, fumarate and formate were detected in very low content (only thousandths of 1 mg per gram of lyophilized cells or below). Our results of the holistic metabolites of P. carterae are the basic references for the further studies when multiple problems will be addressed to this notorious blooming calcifying species.

  7. Identification of novel metoclopramide metabolites in humans: in vitro and in vivo studies.

    PubMed

    Argikar, Upendra A; Gomez, Javier; Ung, Din; Parkman, Henry P; Nagar, Swati

    2010-08-01

    Metoclopramide (MCP) is frequently used to treat gastroparesis. Previous studies have documented MCP metabolism, but systematic structural identification of metabolites has not been performed. The aim of this study was to better understand MCP metabolism in humans. For examination of in vivo metabolism, a single oral 20-mg MCP dose was administered to eight healthy male volunteers, followed by complete urine collection over 24 h. In vitro incubations were performed in human liver microsomes (HLM) to characterize metabolism via cytochromes P450 and UDP-glucuronosyltransferases and in human liver cytosol for metabolism via sulfotransferases. Urine and subcellular incubations were analyzed for MCP metabolites on a mass spectrometer with accurate mass measurement capability. Five MCP metabolites were detected in vivo, and five additional metabolites were detected in vitro. The five metabolites of MCP identified both in vitro and in vivo were an N-O-glucuronide (M1), an N-sulfate (M2), a des-ethyl metabolite (M3), a hydroxylated metabolite (M4), and an oxidative deaminated metabolite (M5). To our knowledge, metabolites M1 and M4 have not been reported previously. M2 urinary levels varied 22-fold and M3 levels varied 16-fold among eight subjects. In vitro studies in HLM revealed the following additional metabolites: two ether glucuronides (M6 and M8), possibly on the phenyl ring after oxidation, an N-glucuronide (M7), a carbamic acid (M9), and a nitro metabolite (M10). Metabolites M6 to M10 have not been reported previously. In conclusion, this study describes the identification of MCP metabolites in vivo and in vitro in humans. PMID:20423954

  8. Exploring antagonistic metabolites of established biocontrol agent of marine origin.

    PubMed

    Rane, Makarand Ramesh; Sarode, Prashant Diwakar; Chaudhari, Bhushan Liladhar; Chincholkar, Sudhir Bhaskarrao

    2008-12-01

    Biocontrol ability of Pseudomonas aeruginosa ID 4365, a biocontrol agent of groundnut phytopathogens from marine origin, was previously attributed to the production of pyoverdin type of siderophores. However, pyoverdin-rich supernatants of this organism showed better antifungal activity compared to equivalent amount of purified pyoverdin indicating presence of undetected metabolite(s) in pyoverdin rich supernatants. On the basis of observation that antagonistic activity was iron-dependent and iron-independent, an attempt was made to detect the presence of additional metabolites. In addition to pyoverdin, strain produced additional siderophores, viz. pyochelin and salicylic acid. Two broad spectrum antifungal compounds, viz. pyocyanin and phenazine-1-carboxylic acid, were detected, characterized, and activity against phytopathogens was demonstrated. Iron- and phosphate-dependent co-production of siderophores and phenazines was confirmed. Strain showed additional features like production of hydrogen cyanide, indol-3-acetic acid, and phosphate solubilization. PMID:18626581

  9. Development of a new high-performance liquid chromatography method with diode array and electrospray ionization-mass spectrometry detection for the metabolite fingerprinting of bioactive compounds in Humulus lupulus L.

    PubMed

    Prencipe, Francesco Pio; Brighenti, Virginia; Rodolfi, Margherita; Mongelli, Andrea; dall'Asta, Chiara; Ganino, Tommaso; Bruni, Renato; Pellati, Federica

    2014-07-01

    The study was aimed at developing a new analytical method for the metabolite fingerprinting of bioactive compounds in Humulus lupulus L. (hop), together with a simple extraction procedure. Different extraction techniques, including maceration, heat reflux extraction (HRE), ultrasound-assisted extraction (UAE) and microwave-assisted extraction (MAE), were compared in order to obtain a high yield of the target analytes. Dynamic maceration for 30min with MeOH-HCOOH (99:1, v/v) as the extraction solvent provided the best result in terms of recovery of secondary metabolites. The analysis of hop constituents, including prenylflavonoids and prenylphloroglucinols (bitter acids), was carried out by means of HPLC-UV/DAD, HPLC-ESI-MS and MS(2), using an ion trap mass analyzer. An Ascentis Express C18 column (150mm×3.0mm I.D., 2.7μm) was used for the HPLC analysis, with a mobile phase composed of 0.25% formic acid in both water and acetonitrile, under gradient elution. The method validation was performed to show compliance with ICH guidelines. The validated technique was successfully applied to the phytochemical analysis of ten commercial cultivars and twenty-three wild Italian hop genotypes, thus demonstrating to be a reliable and useful tool for the comprehensive multi-component analysis of hop secondary metabolites. PMID:24857039

  10. Identification of flurochloridone metabolites in rat urine using liquid chromatography/high resolution mass spectrometry.

    PubMed

    Lu, Dasheng; Zhang, Suhui; Wang, Dongli; Feng, Chao; Liu, Shihong; Jin, Yu 'e; Xu, Qian; Lin, Yuanjie; Wu, Chunhua; Tang, Liming; She, Jianwen; Wang, Guoquan; Zhou, Zhijun

    2016-05-01

    It is of great interest to develop strategic methods to enable chemicals' metabolites to be accurately and rapidly screened and identified. To screen and identify a category of metabolites with distinct isotopic distribution, this study proposed a generic strategy using in silico metabolite prediction plus accurate-mass-based isotopic pattern recognition (AMBIPR) and library identification on the data acquired via the data dependent MS/MS scan of LC-Q Exactive Orbitrap mass spectrometry. The proposed method was evaluated by the analysis of flurochloridone (FLC) metabolites in rat urine sample collected from toxicity tests. Different from the traditional isotopic pattern recognition (IPR) approach, AMBIPR here was performed based on the potential metabolites predicted via in silico metabolite prediction tools. Thus, the AMBIPR treated FLC data was only associated with FLC metabolites, consequently not only avoiding great efforts made to remove FLC-unrelated information and reveal FLC metabolites, but also increasing the percent of positive hits. Among the FLC metabolite peaks screened using AMBIPR, 87% of them (corresponding 97 metabolites and 49 biotransformation) were successfully identified via multiple MS identification techniques packaged in an established FLC's metabolites library based on Mass Frontier. Noteworthy, 34 metabolites (89%) were identified without distinct naturally isotopic distribution. The universal strategic approach based on background subtraction (BS) and mass defect filtering (MDF) was used to evaluate the AMBIPR and no more false positive and negative metabolites were detected. Furthermore, our results revealed that AMBIPR is very effective, inherently sensitive and accurate, and is easily automated for the rapidly screening and profiling chemicals related metabolites. PMID:27063369

  11. Analysis of purine metabolites in maternal serum for evaluating the risk of gestosis.

    PubMed

    Senyavina, N V; Khaustova, S A; Grebennik, T K; Pavlovich, S V

    2013-09-01

    Metabolome analysis of the serum from pregnant patients aimed at detection of low-molecular-weight biomarkers of gestation process disorders indicated a relationship between the metabolic profile of maternal serum and risk of gestosis. In women with pre-eclampsia or preterm delivery, analysis of serum purine metabolites revealed changes in the metabolite concentrations, associated with pregnancy complications. PMID:24288739

  12. Investigating associations between milk metabolite profiles and milk traits of Holstein cows.

    PubMed

    Melzer, N; Wittenburg, D; Hartwig, S; Jakubowski, S; Kesting, U; Willmitzer, L; Lisec, J; Reinsch, N; Repsilber, D

    2013-03-01

    In the field of dairy cattle research, it is of great interest to improve the detection and prevention of diseases (e.g., mastitis and ketosis) and monitor specific traits related to the state of health and management. During the standard milk performance test, traditional milk traits are monitored, and quality and quantity are screened. In addition to the standard test, it is also now possible to analyze milk metabolites in a high-throughput manner and to consider them in connection with milk traits to identify functionally important metabolites that can also serve as biomarker candidates. We present a study in which 190 milk metabolites and 14 milk traits of 1,305 Holstein cows on 18 commercial farms were investigated to characterize interrelations of milk metabolites between each other, to milk traits from the milk standard performance test, and to influencing factors such as farm and sire effect (half-sib structure). The effect of influencing factors (e.g., farm) varied among metabolites and traditional milk traits. The investigations of associations between metabolites and milk traits revealed groups of metabolites that show, for example, positive correlations to protein and casein, and negative correlations to lactose and pH. On the other hand, groups of metabolites jointly associated with the investigated milk traits can be identified and functionally discussed. To enable a multivariate investigation, 2 machine learning methods were applied to detect important metabolites that are highly correlated with the investigated traditional milk traits. For somatic cell score, uracil, lactic acid, and 9 other important metabolites were detected. Lactic acid has already been proposed as a biomarker candidate for mastitis in the recent literature. In conclusion, we found sets of metabolites eligible to predict milk traits, enabling the analysis of milk traits from a metabolic perspective and discussion of the possible functional background for some of the detected

  13. Metabolite identification and quantitation in LC-MS/MS-based metabolomics

    PubMed Central

    Xiao, Jun Feng; Zhou, Bin; Ressom, Habtom W.

    2011-01-01

    Metabolomics aims at detection and quantitation of all metabolites in biological samples. The presence of metabolites with a wide variety of physicochemical properties and different levels of abundance challenges existing analytical platforms used for identification and quantitation of metabolites. Significant efforts have been made to improve analytical and computational methods for metabolomics studies. This review focuses on the use of liquid chromatography with tandem mass spectrometry (LC-MS/MS) for quantitative and qualitative metabolomics studies. It illustrates recent developments in computational methods for metabolite identification, including ion annotation, spectral interpretation and spectral matching. We also review selected reaction monitoring and high-resolution MS for metabolite quantitation. We discuss current challenges in metabolite identification and quantitation as well as potential solutions. PMID:22345829

  14. A Carbonyl Capture Approach for Profiling Oxidized Metabolites in Cell Extracts

    PubMed Central

    Mattingly, Stephanie J.; Xu, Tao; Nantz, Michael H.; Higashi, Richard M.; Fan, Teresa W.-M.

    2012-01-01

    Fourier-transform ion-cyclotron resonance mass spectrometry (FT-ICR-MS) detection of oxidized cellular metabolites is described using isotopologic, carbonyl-selective derivatizing agents that integrate aminooxy functionality for carbonyl capture, quaternary nitrogen for electrospray enhancement, and a hydrophobic domain for sample cleanup. These modular structural features enable rapid, sensitive analysis of complex mixtures of metabolite-derivatives by FT-ICR-MS via continuous nanoelectrospray infusion. Specifically, this approach can be used to globally assess levels of low abundance and labile aldehyde and ketone metabolites quantitatively and in high throughput manner. These metabolites are often key and unique indicators of various biochemical pathways and their perturbations. Analysis of lung adenocarcinoma A549 cells established a profile of carbonyl metabolites spanning multiple structural classes. We also demonstrate a procedure for metabolite quantification using pyruvate as a model analyte. PMID:23175637

  15. Nuclear Magnetic Resonance Identification of New Sulfonic Acid Metabolites of Chloroacetanilide Herbicides

    USGS Publications Warehouse

    Morton, M.D.; Walters, F.H.; Aga, D.S.; Thurman, E.M.; Larive, C.K.

    1997-01-01

    The detection of the sulfonic acid metabolites of the chloroacetanilide herbicides acetochlor, alachlor, butachlor, propachlor, and, more recently, metolachlor in surface and ground water suggests that a common mechanism for dechlorination exists via the glutathione conjugation pathway. The identification of these herbicides and their metabolites is important due to growing public awareness and concern about pesticide levels in drinking water. Although these herbicides are regulated, little is known about the fate of their metabolites in soil. The sulfonic acid metabolites were synthesized by reaction of the parent compounds with an excess of sodium sulfite. Acetochlor, alachlor, butachlor, metolachlor, and propachlor and their sulfonic acid metabolites were studied by nuclear magnetic resonance spectroscopy and fast atom bombardment mass spectrometry. This paper provides a direct method for the preparation and characterization of these compounds that will be useful in the analysis and study of chloracetanilide herbicides and their metabolites.

  16. Determination of XLR-11 and its metabolites in hair by liquid chromatography-tandem mass spectrometry.

    PubMed

    Park, Meejung; Yeon, Seonghoon; Lee, Jaesin; In, Sangwhan

    2015-10-10

    Analysis of drugs in hair is often used as a routine method to obtain detailed information about drug ingestion. However, few studies have been conducted on disposition of synthetic cannabinoids including cyclopropylindoles (UR-144 and XLR-11) and their metabolites in hair. XLR-11 has been widely abused in South Korea recently. Identification of metabolites in hair can be an important proof of synthetic cannabinoids use because it can exclude the possibility of passive smoke exposure. In this study, we described a quantitative analytical method of XLR-11 and its metabolites (UR-144, UR-144 N-5-hydroxypentyl metabolite, UR-144 N-4-hydroxypentyl metabolite, UR-144 N-pentanoic acid metabolite and XLR-11 N-4-hydroxypentyl metabolite) in hair by liquid chromatography with ESI-MS/MS. The target analytes were extracted with methanol from washed and cut hair samples and the extracts were evaporated, filtered and analyzed by LC-MS/MS with electrospray ion source in positive-ionization mode. JWH-018-d9 and JWH-018 N-5-hydroxypentyl metabolite-d5 were used as internal standards. Chromatographic separation was completed within 15 min. No interferences were detected in 10 blank hair samples. In intra- and inter-assay precision and accuracy study, CV (%) and bias (%) were below 12. The limit of detection (LOD) was 0.1∼2 pg/mg and the limit of quantification (LOQ) was 0.2-2 pg/mg, respectively. The validation results proved that the method was selective, accurate and precise with acceptable linearity within calibration range. No significant variation was observed by different sources of matrices. This method was applied to hair samples from 14 individual suspects of XLR-11 use. In this result, XLR-11, UR-144, UR-144 N-5-hydroxypentyl metabolite and UR-144 N-pentanoic acid metabolite, XLR-11 N-4-hydroxypentyl metabolite were detected. The concentration of XLR-11 as a parent drug was much higher than other metabolites. UR-144 N-5-hydroxy metabolite and UR-144 N-pentanoic acid

  17. High-resolution mass spectrometry elucidates metabonate (false metabolite) formation from alkylamine drugs during in vitro metabolite profiling.

    PubMed

    Barbara, Joanna E; Kazmi, Faraz; Muranjan, Seema; Toren, Paul C; Parkinson, Andrew

    2012-10-01

    In vitro metabolite profiling and characterization experiments are widely employed in early drug development to support safety studies. Samples from incubations of investigational drugs with liver microsomes or hepatocytes are commonly analyzed by liquid chromatography/mass spectrometry for detection and structural elucidation of metabolites. Advanced mass spectrometers with accurate mass capabilities are becoming increasingly popular for characterization of drugs and metabolites, spurring changes in the routine workflows applied. In the present study, using a generic full-scan high-resolution data acquisition approach with a time-of-flight mass spectrometer combined with postacquisition data mining, we detected and characterized metabonates (false metabolites) in microsomal incubations of several alkylamine drugs. If a targeted approach to mass spectrometric detection (without full-scan acquisition and appropriate data mining) were employed, the metabonates may not have been detected, hence their formation underappreciated. In the absence of accurate mass data, the metabonate formation would have been incorrectly characterized because the detected metabonates manifested as direct cyanide-trapped conjugates or as cyanide-trapped metabolites formed from the parent drugs by the addition of 14 Da, the mass shift commonly associated with oxidation to yield a carbonyl. This study demonstrates that high-resolution mass spectrometry and the associated workflow is very useful for the detection and characterization of unpredicted sample components and that accurate mass data were critical to assignment of the correct metabonate structures. In addition, for drugs containing an alkylamine moiety, the results suggest that multiple negative controls and chemical trapping agents may be necessary to correctly interpret the results of in vitro experiments. PMID:22798552

  18. Toxicological significance of dihydrodiol metabolites

    SciTech Connect

    Hsia, M.T.

    1982-01-01

    Dihydrodiols are often found as the major organic-extractable metabolites of various olefinic or aromatic xenobiotics in many biological samples. Studies on the chemistry of dihydrodiol metabolites have provided insight into the pharmacokinetic behavior and the mode of action of the parent compound. The toxicology of dihydrodiol is more complex than what can be deduced solely on the basis of diminished bioavailability of the epoxide precursor, and the increased hydrophilicity associated with the dihydrodiol moiety. Dihydrodiols can be intrinsically toxic and may even represent metabolically activated species. Some of the dihydrodiol metabolites may still retain sufficient lipophilic character to serve again as substrates for microsomal oxygenases. Because of the tremendous chemical and biological diversity that existed among the various dihydrodiols, more mechanistic studies are needed to examine the toxicological properties of these compounds. It may be premature to conclude dihydrodiol formation as purely a detoxification route for xenobioties.

  19. Deleterious effects of reactive metabolites

    PubMed Central

    2010-01-01

    A number of drugs have been withdrawn from the market or severely restricted in their use because of unexpected toxicities that become apparent only after the launch of new drug entities. Circumstantial evidence suggests that, in most cases, reactive metabolites are responsible for these unexpected toxicities. In this review, a general overview of the types of reactive metabolites and the consequences of their formation are presented. The current approaches to evaluate bioactivation potential of new compounds with particular emphasis on the advantages and limitation of these procedures will be discussed. Reasonable reasons for the excellent safety record of certain drugs susceptible to bioactivation will also be explored and should provide valuable guidance in the use of reactive-metabolite assessments when nominating drug candidates for development. This will, in turn, help us to design and bring safer drugs to the market. PMID:20972370

  20. Microbial production of primary metabolites

    NASA Astrophysics Data System (ADS)

    Demain, Arnold L.

    1980-12-01

    Microbial production of primary metabolites contributes significantly to the quality of life. Through fermentation, microorganisms growing on inexpensive carbon sources can produce valuable products such as amino acids, nucleotides, organic acids, and vitamins which can be added to food to enhance its flavor or increase its nutritive value. The contribution of microorganisms will go well beyond the food industry with the renewed interest in solvent fermentations. Microorganisms have the potential to provide many petroleum-derived products as well as the ethanol necessary for liquid fuel. The role of primary metabolites and the microbes which produce them will certainly increase in importance.

  1. Sphingolipid metabolites in inflammatory disease

    PubMed Central

    Maceyka, Michael; Spiegel, Sarah

    2015-01-01

    Sphingolipids are ubiquitous building blocks of eukaryotic cell membranes. Progress in our understanding of sphingolipid metabolism, state-of-the-art sphingolipidomic approaches and animal models have generated a large body of evidence demonstrating that sphingolipid metabolites, particularly ceramide and sphingosine-1-phosphate, are signalling molecules that regulate a diverse range of cellular processes that are important in immunity, inflammation and inflammatory disorders. Recent insights into the molecular mechanisms of action of sphingolipid metabolites and new perspectives on their roles in regulating chronic inflammation have been reported. The knowledge gained in this emerging field will aid in the development of new therapeutic options for inflammatory disorders. PMID:24899305

  2. Multiresidue determination of pesticides and pesticide metabolites in soil

    SciTech Connect

    Mogadati, P.S.; Rosen, J.D.

    1995-12-31

    Methods for the multiresidue extraction, cleanup and GC/MS determination of 142 pesticides and pesticide metabolites in soil have been developed. The use of solid phase extraction cartridges makes it possible to clean up the soil sufficiently so that the equivalent of 40 mg. soil may be injected onto the GC capillary column without overloading or harming the column. Combining this clean-up method with chemical ionization ion trap detection allowed for very low limits of detection.

  3. Methodological considerations for measuring glucocorticoid metabolites in feathers

    PubMed Central

    Berk, Sara A.; McGettrick, Julie R.; Hansen, Warren K.; Breuner, Creagh W.

    2016-01-01

    In recent years, researchers have begun to use corticosteroid metabolites in feathers (fCORT) as a metric of stress physiology in birds. However, there remain substantial questions about how to measure fCORT most accurately. Notably, small samples contain artificially high amounts of fCORT per millimetre of feather (the small sample artefact). Furthermore, it appears that fCORT is correlated with circulating plasma corticosterone only when levels are artificially elevated by the use of corticosterone implants. Here, we used several approaches to address current methodological issues with the measurement of fCORT. First, we verified that the small sample artefact exists across species and feather types. Second, we attempted to correct for this effect by increasing the amount of methanol relative to the amount of feather during extraction. We consistently detected more fCORT per millimetre or per milligram of feather in small samples than in large samples even when we adjusted methanol:feather concentrations. We also used high-performance liquid chromatography to identify hormone metabolites present in feathers and measured the reactivity of these metabolites against the most commonly used antibody for measuring fCORT. We verified that our antibody is mainly identifying corticosterone (CORT) in feathers, but other metabolites have significant cross-reactivity. Lastly, we measured faecal glucocorticoid metabolites in house sparrows and correlated these measurements with corticosteroid metabolites deposited in concurrently grown feathers; we found no correlation between faecal glucocorticoid metabolites and fCORT. We suggest that researchers should be cautious in their interpretation of fCORT in wild birds and should seek alternative validation methods to examine species-specific relationships between environmental challenges and fCORT. PMID:27335650

  4. Methodological considerations for measuring glucocorticoid metabolites in feathers.

    PubMed

    Berk, Sara A; McGettrick, Julie R; Hansen, Warren K; Breuner, Creagh W

    2016-01-01

    In recent years, researchers have begun to use corticosteroid metabolites in feathers (fCORT) as a metric of stress physiology in birds. However, there remain substantial questions about how to measure fCORT most accurately. Notably, small samples contain artificially high amounts of fCORT per millimetre of feather (the small sample artefact). Furthermore, it appears that fCORT is correlated with circulating plasma corticosterone only when levels are artificially elevated by the use of corticosterone implants. Here, we used several approaches to address current methodological issues with the measurement of fCORT. First, we verified that the small sample artefact exists across species and feather types. Second, we attempted to correct for this effect by increasing the amount of methanol relative to the amount of feather during extraction. We consistently detected more fCORT per millimetre or per milligram of feather in small samples than in large samples even when we adjusted methanol:feather concentrations. We also used high-performance liquid chromatography to identify hormone metabolites present in feathers and measured the reactivity of these metabolites against the most commonly used antibody for measuring fCORT. We verified that our antibody is mainly identifying corticosterone (CORT) in feathers, but other metabolites have significant cross-reactivity. Lastly, we measured faecal glucocorticoid metabolites in house sparrows and correlated these measurements with corticosteroid metabolites deposited in concurrently grown feathers; we found no correlation between faecal glucocorticoid metabolites and fCORT. We suggest that researchers should be cautious in their interpretation of fCORT in wild birds and should seek alternative validation methods to examine species-specific relationships between environmental challenges and fCORT. PMID:27335650

  5. Reactive Arrays of Colorimetric Sensors for Metabolite and Steroid Identification.

    PubMed

    Batres, Gary; Jones, Talia; Johnke, Hannah; Wilson, Mark; Holmes, Andrea E; Sikich, Sharmin

    2014-12-31

    The work described herein examines a rapid mix-and-measure method called DETECHIP suitable for screening of steroids and metabolites. The addition of steroids and metabolites to reactive arrays of colorimetric sensors generated characteristic color "fingerprints" that were used to identify the analyte. A color analysis tool was used to identify the analyte pool that now includes biologically relevant analytes. The mix-and-measure arrays allowed the detection of disease metabolites, orotic acid and argininosuccinic acid; and the steroids androsterone, 1,4-androstadiene, testosterone, stanozolol, and estrone. The steroid 1,4-androstadiene was also detected by this method while dissolved in synthetic urine. Some of the steroids, such as androstadiene, stanozolol, and androsterone were co-dissolved with (2-hydroxypropyl)-β-cyclodextrin in order to increase solubility in aqueous buffered solutions. The colorimetric arrays do not intend to eliminate ELISA or mass spectroscopy based screening, but to possibly provide an alternative analytical detection method for steroids and metabolites. PMID:25019034

  6. Pesticides in ground water: Do atrazine metabolites matter?

    USGS Publications Warehouse

    Liu, S.; Yen, S.T.; Kolpin, D.W.

    1996-01-01

    Atrazine and atrazine-residue (atrazine + two metabolites - deethylatrazine and deisopropylatrazine) concentrations were examined to determine if consideration of these atrazine metabolites substantially adds to our understanding of the distribution of this pesticide in groundwater of the midcontinental United States. The mean of atrazine.residue concentrations was 53 percent greater than that of atrazine alone for those observations above the detection limit (> 0.05 μg/l). Furthermore, a censored regression analysis using atrazine-residue concentrations revealed significant factors not identified when only atrazine concentrations were used. Thus, knowledge of concentrations of these atrazine metabolites is required to obtain a true estimation of risk of using these aquifers as sources for drinking water, and such knowledge also provides information that ultimately may be important for future management policies designed to reduce atrazine concentrations in ground water.

  7. Nontargeted Modification-Specific Metabolomics Investigation of Glycosylated Secondary Metabolites in Tea (Camellia sinensis L.) Based on Liquid Chromatography-High-Resolution Mass Spectrometry.

    PubMed

    Dai, Weidong; Tan, Junfeng; Lu, Meiling; Xie, Dongchao; Li, Pengliang; Lv, Haipeng; Zhu, Yin; Guo, Li; Zhang, Yue; Peng, Qunhua; Lin, Zhi

    2016-09-01

    Glycosylation on small molecular metabolites modulates a series of biological events in plants. However, a large number of glycosides have not been discovered and investigated using -omics approaches. Here, a general strategy named "nontargeted modification-specific metabolomics" was applied to map the glycosylation of metabolites. The key aspect of this method is to adopt in-source collision-induced dissociation to dissociate the glycosylated metabolite, causing a characteristic neutral loss pattern, which acts as an indicator for the glycosylation identification. In an exemplary application in green teas, 120 glucosylated/galactosylated, 38 rhamnosylated, 21 rutinosylated, and 23 primeverosylated metabolites were detected simultaneously. Among them, 61 glycosylated metabolites were putatively identified according to current tea metabolite databases. Thanks to the annotations of glycosyl moieties in advance, the method aids metabolite identifications. An additional 40 novel glycosylated metabolites were tentatively elucidated. This work provides a feasible strategy to discover and identify novel glycosylated metabolites in plants. PMID:27541009

  8. CHARACTERIZATION OF METABOLITES IN SMALL FISH BIOFLUIDS AND TISSUES BY NMR SPECTROSCOPY

    EPA Science Inventory

    Nuclear magnetic resonance (NMR) spectroscopy has been utilized for assessing ecotoxicity in small fish models by means of metabolomics. Two fundamental challenges of NMR-based metabolomics are the detection limit and characterization of metabolites (or NMR resonance assignments...

  9. Concurrent quantification of tryptophan and its major metabolites

    PubMed Central

    Lesniak, Wojciech G.; Jyoti, Amar; Mishra, Manoj K.; Louissaint, Nicolette; Romero, Roberto; Chugani, Diane C.; Kannan, Sujatha; Kannan, Rangaramanujam M.

    2014-01-01

    An imbalance in tryptophan (TRP) metabolites is associated with several neurological and inflammatory disorders. Therefore, analytical methods allowing for simultaneous quantification of TRP and its major metabolites would be highly desirable, and may be valuable as potential biomarkers. We have developed a HPLC method for concurrent quantitative determination of tryptophan, serotonin, 5-hydroxyindoleacetic acid, kynurenine, and kynurenic acid in tissue and fluids. The method utilizes the intrinsic spectroscopic properties of TRP and its metabolites that enable UV absorbance and fluorescence detection by HPLC, without additional labeling. The origin of the peaks related to analytes of interest was confirmed by UV–Vis spectral patterns using a PDA detector and mass spectrometry. The developed methods were validated in rabbit fetal brain and amniotic fluid at gestational day 29. Results are in excellent agreement with those reported in the literature for the same regions. This method allows for rapid quantification of tryptophan and four of its major metabolites concurrently. A change in the relative ratios of these metabolites can provide important insights in predicting the presence and progression of neuroinflammation in disorders such as cerebral palsy, autism, multiple sclerosis, Alzheimer disease, and schizophrenia. PMID:24036037

  10. Primary expectations of secondary metabolites

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Plant secondary metabolites (e.g., phenolics) are important for human health, in addition to the organoleptic properties they impart to fresh and processed foods. Consumer expectations such as appearance, taste, or texture influence their purchasing decisions. Thorough identification of phenolic com...

  11. Natural products: Hunting microbial metabolites

    NASA Astrophysics Data System (ADS)

    Schmidt, Eric W.

    2015-05-01

    Symbiotic bacteria synthesize many specialized small molecules; however, establishing the role these chemicals play in human health and disease has been difficult. Now, the chemical structure and mechanism of the Escherichia coli product colibactin provides insight into the link between this secondary metabolite and colorectal cancer.

  12. METABOLITE PROFILING OF ECHINACEA GENOTYPES

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Echinacea extracts have historically been used as herbal remedies to treat colds, coughs and snake bites. Echinacea products are currently sold as a popular herbal-remedy used for general enhancement of the immune system. However, the genetic variation in metabolites has not been systematically ch...

  13. Quantitating Metabolites in Protein Precipitated Serum Using NMR Spectroscopy

    PubMed Central

    2015-01-01

    Quantitative NMR-based metabolite profiling is challenged by the deleterious effects of abundant proteins in the intact blood plasma/serum, which underscores the need for alternative approaches. Protein removal by ultrafiltration using low molecular weight cutoff filters thus represents an important step. However, protein precipitation, an alternative and simple approach for protein removal, lacks detailed quantitative assessment for use in NMR based metabolomics. In this study, we have comprehensively evaluated the performance of protein precipitation using methanol, acetonitrile, perchloric acid, and trichloroacetic acid and ultrafiltration approaches using 1D and 2D NMR, based on the identification and absolute quantitation of 44 human blood metabolites, including a few identified for the first time in the NMR spectra of human serum. We also investigated the use of a “smart isotope tag,” 15N-cholamine for further resolution enhancement, which resulted in the detection of a number of additional metabolites. 1H NMR of both protein precipitated and ultrafiltered serum detected all 44 metabolites with comparable reproducibility (average CV, 3.7% for precipitation; 3.6% for filtration). However, nearly half of the quantified metabolites in ultrafiltered serum exhibited 10–74% lower concentrations; specifically, tryptophan, benzoate, and 2-oxoisocaproate showed much lower concentrations compared to protein precipitated serum. These results indicate that protein precipitation using methanol offers a reliable approach for routine NMR-based metabolomics of human blood serum/plasma and should be considered as an alternative to ultrafiltration. Importantly, protein precipitation, which is commonly used by mass spectrometry (MS), promises avenues for direct comparison and correlation of metabolite data obtained from the two analytical platforms to exploit their combined strength in the metabolomics of blood. PMID:24796490

  14. Microbial secondary metabolites in homes in association with moisture damage and asthma.

    PubMed

    Kirjavainen, P V; Täubel, M; Karvonen, A M; Sulyok, M; Tiittanen, P; Krska, R; Hyvärinen, A; Pekkanen, J

    2016-06-01

    We aimed to characterize the presence of microbial secondary metabolites in homes and their association with moisture damage, mold, and asthma development. Living room floor dust was analyzed by LC-MS/MS for 333 secondary metabolites from 93 homes of 1-year-old children. Moisture damage was present in 15 living rooms. At 6 years, 8 children had active and 15 lifetime doctor-diagnosed asthma. The median number of different metabolites per house was 17 (range 8-29) and median sum load 65 (4-865) ng/m(2) . Overall 42 different metabolites were detected. The number of metabolites present tended to be higher in homes with mold odor or moisture damage. The higher sum loads and number of metabolites with loads over 10 ng/m(2) were associated with lower prevalence of active asthma at 6 years (aOR 0.06 (95% CI <0.001-0.96) and 0.05 (<0.001-0.56), respectively). None of the individual metabolites, which presence tended (P < 0.2) to be increased by moisture damage or mold, were associated with increased risk of asthma. Microbial secondary metabolites are ubiquitously present in home floor dust. Moisture damage and mold tend to increase their numbers and amount. There was no evidence indicating that the secondary metabolites determined would explain the association between moisture damage, mold, and the development of asthma. PMID:25913237

  15. Relative mass defect filtering of mass spectra: a path to discovery of plant specialized metabolites.

    PubMed

    Ekanayaka, E A Prabodha; Celiz, Mary Dawn; Jones, A Daniel

    2015-04-01

    The rapid identification of novel plant metabolites and assignments of newly discovered substances to natural product classes present the main bottlenecks to defining plant specialized phenotypes. Although mass spectrometry provides powerful support for metabolite discovery by measuring molecular masses, ambiguities in elemental formulas often fail to reveal the biosynthetic origins of specialized metabolites detected using liquid chromatography-mass spectrometry. A promising approach for mining liquid chromatography-mass spectrometry metabolite profiling data for specific metabolite classes is achieved by calculating relative mass defects (RMDs) from molecular and fragment ions. This strategy enabled the rapid recognition of an extensive range of terpenoid metabolites in complex plant tissue extracts and is independent of retention time, abundance, and elemental formula. Using RMD filtering and tandem mass spectrometry data analysis, 24 novel elemental formulas corresponding to glycosylated sesquiterpenoid metabolites were identified in extracts of the wild tomato Solanum habrochaites LA1777 trichomes. Extensive isomerism was revealed by ultra-high-performance liquid chromatography, leading to evidence of more than 200 distinct sesquiterpenoid metabolites. RMD filtering led to the recognition of the presence of glycosides of two unusual sesquiterpenoid cores that bear limited similarity to known sesquiterpenes in the genus Solanum. In addition, RMD filtering is readily applied to existing metabolomics databases and correctly classified the annotated terpenoid metabolites in the public metabolome database for Catharanthus roseus. PMID:25659383

  16. Cytochrome c adducts with PCB quinoid metabolites.

    PubMed

    Li, Miao; Teesch, Lynn M; Murry, Daryl J; Pope, R Marshal; Li, Yalan; Robertson, Larry W; Ludewig, Gabriele

    2016-02-01

    Polychlorinated biphenyls (PCBs) are a group of 209 individual congeners widely used as industrial chemicals. PCBs are found as by-products in dye and paint manufacture and are legacy, ubiquitous, and persistent as human and environmental contaminants. PCBs with fewer chlorine atoms may be metabolized to hydroxy- and dihydroxy-metabolites and further oxidized to quinoid metabolites both in vitro and in vivo. Specifically, quinoid metabolites may form adducts on nucleophilic sites within cells. We hypothesized that the PCB-quinones covalently bind to cytochrome c and, thereby, cause defects in the function of cytochrome c. In this study, synthetic PCB quinones, 2-(4'-chlorophenyl)-1,4-benzoquinone (PCB3-pQ), 4-4'-chlorophenyl)-1,2-benzoquinone (PCB3-oQ), 2-(3', 5'-dichlorophenyl)-1,4-benzoquinone, 2-(3',4', 5'-trichlorophenyl)-1,4-benzoquinone, and 2-(4'-chlorophenyl)-3,6-dichloro-1,4-benzoquinone, were incubated with cytochrome c, and adducts were detected by liquid chromatography-mass spectrometry (LC-MS) and matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI TOF). Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) was employed to separate the adducted proteins, while trypsin digestion and liquid chromatography-tandem mass spectrometry (LC-MS/MS) were applied to identify the amino acid binding sites on cytochrome c. Conformation change of cytochrome c after binding with PCB3-pQ was investigated by SYBYL-X simulation and cytochrome c function was examined. We found that more than one molecule of PCB-quinone may bind to one molecule of cytochrome c. Lysine and glutamic acid were identified as the predominant binding sites. Software simulation showed conformation changes of adducted cytochrome c. Additionally, cross-linking of cytochrome c was observed on the SDS-PAGE gel. Cytochrome c was found to lose its function as electron acceptor after incubation with PCB quinones. These data provide evidence that the covalent

  17. Concentrations of Phthalate Metabolites in Milk, Urine, Saliva, and Serum of Lactating North Carolina Women

    PubMed Central

    Hines, Erin P.; Calafat, Antonia M.; Silva, Manori J.; Mendola, Pauline; Fenton, Suzanne E.

    2009-01-01

    Background Phthalates are ubiquitous in the environment, but concentrations in multiple media from breast-feeding U.S. women have not been evaluated. Objectives The objective of this study was to accurately measure and compare the concentrations of oxidative monoester phthalate metabolites in milk and surrogate fluids (serum, saliva, and urine) of 33 lactating North Carolina women. Methods We analyzed serum, saliva, urine, and milk for the oxidative phthalate metabolites mono(3-carboxypropyl) phthalate, mono(2-ethyl-5-carboxypentyl) phthalate (MECPP), mono(2-ethyl-5-hydroxyhexyl) phthalate, and mono(2-ethyl-5-oxohexyl) phthalate using isotope-dilution high-performance liquid chromatography tandem mass spectroscopy. Because only urine lacks esterases, we analyzed it for the hydrolytic phthalate monoesters. Results We detected phthalate metabolites in few milk (< 10%) and saliva samples. MECPP was detected in > 80% of serum samples, but other metabolites were less common (3–22%). Seven of the 10 urinary metabolites were detectable in ≥ 85% of samples. Monoethyl phthalate had the highest mean concentration in urine. Metabolite concentrations differed by body fluid (urine > serum > milk and saliva). Questionnaire data suggest that frequent nail polish use, immunoglobulin A, and fasting serum glucose and triglyceride levels were increased among women with higher concentrations of urinary and/or serum phthalate metabolites; motor vehicle age was inversely correlated with certain urinary phthalate concentrations. Conclusions Our data suggest that phthalate metabolites are most frequently detected in urine of lactating women and are less often detected in serum, milk, or saliva. Urinary phthalate concentrations reflect maternal exposure and do not represent the concentrations of oxidative metabolites in other body fluids, especially milk. PMID:19165392

  18. Global Perspectives of Fungal Secondary Metabolite Research

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Fungi produce a wide range of unusual metabolites, termed secondary metabolites because they play no role in the normal, basic metabolic pathways used for growth and energy production, etc. Some of these secondary metabolites have antibiotic properties; others are potent toxins that are dangerous w...

  19. Extending metabolome coverage for untargeted metabolite profiling of adherent cultured hepatic cells.

    PubMed

    García-Cañaveras, Juan Carlos; López, Silvia; Castell, José Vicente; Donato, M Teresa; Lahoz, Agustín

    2016-02-01

    MS-based metabolite profiling of adherent mammalian cells comprises several challenging steps such as metabolism quenching, cell detachment, cell disruption, metabolome extraction, and metabolite measurement. In LC-MS, the final metabolome coverage is strongly determined by the separation technique and the MS conditions used. Human liver-derived cell line HepG2 was chosen as adherent mammalian cell model to evaluate the performance of several commonly used procedures in both sample processing and LC-MS analysis. In a first phase, metabolite extraction and sample analysis were optimized in a combined manner. To this end, the extraction abilities of five different solvents (or combinations) were assessed by comparing the number and the levels of the metabolites comprised in each extract. Three different chromatographic methods were selected for metabolites separation. A HILIC-based method which was set to specifically separate polar metabolites and two RP-based methods focused on lipidome and wide-ranging metabolite detection, respectively. With regard to metabolite measurement, a Q-ToF instrument operating in both ESI (+) and ESI (-) was used for unbiased extract analysis. Once metabolite extraction and analysis conditions were set up, the influence of cell harvesting on metabolome coverage was also evaluated. Therefore, different protocols for cell detachment (trypsinization or scraping) and metabolism quenching were compared. This study confirmed the inconvenience of trypsinization as a harvesting technique, and the importance of using complementary extraction solvents to extend metabolome coverage, minimizing interferences and maximizing detection, thanks to the use of dedicated analytical conditions through the combination of HILIC and RP separations. The proposed workflow allowed the detection of over 300 identified metabolites from highly polar compounds to a wide range of lipids. PMID:26769129

  20. Electrophilicities and Protein Covalent Binding of Demethylation Metabolites of Colchicine.

    PubMed

    Guo, Xiucai; Lin, Dongju; Li, Weiwei; Wang, Kai; Peng, Ying; Zheng, Jiang

    2016-03-21

    Colchicine, an alkaloid existing in plants of Liliaceous colchicum, has been widely used in the treatment of gout and familial Mediterranean fever. The administration of colchicine was found to cause liver injury in humans. The mechanisms of colchicine-induced liver toxicity remain unknown. The objectives of this study were to determine the electrophilicities of demethylation metabolites of colchicine and investigate the protein adductions derived from the reactive metabolites of colchicine. Four demethylated colchicine (1-, 2-, 3-, and 10-DMCs), namely, M1-M4, were detected in colchicine-fortified microsomal incubations. Four N-acetyl cysteine (NAC) conjugates (M5-M8) derived from colchicine were detected in the microsomes in the presence of NAC. M5 and M6 were derived from 10-DMC. M7 resulted from the reaction of 2-DMC or 3-DMC with NAC, and M8 originated from 10-DMC. Microsomal protein covalent binding was observed after exposure to colchicine. Two cysteine adducts (CA-1 and CA-2) derived from 10-DMC were found in proteolytically digested microsomal protein samples after incubation with colchicine. The findings allow us to define the chemical property of demethylation metabolites of colchicine and the interaction between protein and the reactive metabolites of colchicine generated in situ. PMID:26845511

  1. Antileishmanial Metabolites from Geosmithia langdonii

    PubMed Central

    2015-01-01

    Antileishmanial bioassay guided fractionation of Geosmithia langdonii has resulted in the isolation and identification of two new compounds (1 and 2) together with 10 known compounds (3–12). The structures of the isolated metabolites were elucidated based on comprehensive 1D and 2D NMR spectroscopic data as well as mass spectrometry. The absolute configuration at C4, C5, and C6 of 2 was determined as R using a modified Mosher esterification method and NOESY correlations. The extracts and the isolated metabolites were evaluated for their antileishmanial activities. Compounds 3, 9, 11, and 12 were found to be active against Leishmania donovani with IC50 values of 6.9, 3.3, 8.5, and 9.2 μM, respectively, while compounds 1, 5, and 10 showed lower activities against L. donovani with IC50 values of 13.0, 47.3, and 34.0 μM, respectively. PMID:25084548

  2. The effects of GA and ABA treatments on metabolite profile of germinating barley.

    PubMed

    Huang, Yuqing; Cai, Shengguan; Ye, Lingzhen; Hu, Hongliang; Li, Chengdao; Zhang, Guoping

    2016-02-01

    Sugar degradation during grain germination is important for malt quality. In malting industry, gibberellin (GA) is frequently used for improvement of malting quality. In this study, the changes of metabolite profiles and starch-degrading enzymes during grain germination, and as affected by GA and abscisic acid (ABA) were investigated using two wild barley accessions XZ72 and XZ95. Totally fifty-two metabolites with known structures were detected and the change of metabolite during germination was time- and genotype dependent. Sugars and amino acids were the most dramatically changed compounds. Addition of GA enhanced the activities of starch-degrading enzymes, and increased most metabolites, especially sugars and amino acids, whereas ABA had the opposite effect. The effect varied with the barley accessions. The current study is the first attempt in investigating the effect of hormones on metabolite profiles in germinating barley grain, being helpful for identifying the factors affecting barley germination or malt quality. PMID:26304431

  3. Different profiles of quercetin metabolites in rat plasma: comparison of two administration methods.

    PubMed

    Kawai, Yoshichika; Saito, Satomi; Nishikawa, Tomomi; Ishisaka, Akari; Murota, Kaeko; Terao, Junji

    2009-03-23

    The bioavailability of polyphenols in human and rodents has been discussed regarding their biological activity. We found different metabolite profiles of quercetin in rat plasma between two administration procedures. A single intragastric administration (50 mg/kg) resulted in the appearance of a variety of metabolites in the plasma, whereas only a major fraction was detected by free access (1% quercetin). The methylated/non-methylated metabolites ratio was much higher in the free access group. Mass spectrometric analyses showed that the fraction from free access contained highly conjugated quercetin metabolites such as sulfo-glucuronides of quercetin and methylquercetin. The metabolite profile of human plasma after an intake of onion was similar to that with intragastric administration in rats. In vitro oxidation of human low-density lipoprotein showed that methylation of the catechol moiety of quercetin significantly attenuated the antioxidative activity. These results might provide information about the bioavailability of quercetin when conducting animal experiments. PMID:19270373

  4. Metabolites from inhalation of aerosolized S-8 synthetic jet fuel in rats.

    PubMed

    Tremblay, Raphael T; Martin, Sheppard A; Fisher, Jeffrey W

    2011-01-01

    Alternative fuels are being considered for civilian and military uses. One of these is S-8, a replacement jet fuel synthesized using the Fischer-Tropsch process, which contains no aromatic compounds and is mainly composed of straight and branched alkanes. Metabolites of S-8 fuel in laboratory animals have not been identified. The goal of this study was to identify metabolic products from exposure to aerosolized S-8 and a designed straight-chain alkane/polyaromatic mixture (decane, undecane, dodecane, tridecane, tetradecane, pentadecane, naphthalene, and 2-methylnaphthalene) in male Fischer 344 rats. Collected blood and tissue samples were analyzed for 70 straight and branched alcohols and ketones ranging from 7 to 15 carbons. No fuel metabolites were observed in the blood, lungs, brain, and fat following S-8 exposure. Metabolites were detected in the liver, urine, and feces. Most of the metabolites were 2- and 3-position alcohols and ketones of prominent hydrocarbons with very few 1- or 4-position metabolites. Following exposure to the alkane mixture, metabolites were observed in the blood, liver, and lungs. Interestingly, heavy metabolites (3-tridecanone, 2-tridecanol, and 2-tetradecanol) were observed only in the lung tissues possibly indicating that metabolism occurred in the lungs. With the exception of these heavy metabolites, the metabolic profiles observed in this study are consistent with previous studies reporting on the metabolism of individual alkanes. Further work is needed to determine the potential metabolic interactions of parent, primary, and secondary metabolites and identify more polar metabolites. Some metabolites may have potential use as biomarkers of exposure to fuels. PMID:21222558

  5. Characterization of oxygenated metabolites of ginsenoside Rg1 in plasma and urine of rat.

    PubMed

    Wang, Jing-Rong; Tong, Tian-Tian; Yau, Lee-Fong; Chen, Cheng-Yu; Bai, Li-Ping; Ma, Jing; Hu, Ming; Liu, Liang; Jiang, Zhi-Hong

    2016-07-15

    This study describes the characterization of oxygenated metabolites of ginsenoside Rg1 in rat urine and plasma. These in vivo metabolites were profiled by using UHPLC-QTOF MS-based method. On the basis of high-resolution MS/MS data, and comparison with chemically synthesized authentic compounds, nine oxygenated metabolites of Rg1 were characterized as vinaginsenosides 21 and 22 (M1 and M2), vinaginsenoside R15 (M3), 6-O-(β-d-glucopyranosyl)-20-O-(β-d-glucopyranosyl) 3β, 6α, 12β, 20(S)-tetrahydroxy-24ξ-hydroxydammar-25-ene (M4 and M5), floralginsenoside A (M7 and M8), floralginsenoside B (M9) and epoxyginsenoside Rg1 (M13), respectively. Among these metabolites, M4, M5 and M13 are new ginsenosides and others were detected as in vivo metabolites of Rg1 for the first time. In addition, a series of oxygenated metabolites of Rh1 and deglycosylated metabolite of Rg1, were observed and characterized by comparing with compounds synthesized by us, which revealed an association between C-20 configuration and the extent of oxidation metabolism. Appearance of all these metabolites in blood stream and urine after i.v. dosing and oral administration of Rg1 was further examined, which clearly showed that mono-oxygenated metabolites of Rg1 were major circulating metabolites at the early stage after dosing. Characterization of exact chemical structures of these circulating metabolites contribute greatly to our understanding of chemical exposure after consumption of ginseng products, and provide valuable information for explaining multiple bioactivities of ginseng products. PMID:26809375

  6. Is oseltamivir (Tamiflu) safe? Re-examining the Tamiflu 'ado' from Japan.

    PubMed

    Okamoto, Etsuji

    2010-02-01

    Reports that oseltamivir causes abnormal behaviors in influenza patients and, thereby, increases the risk of injuries or death alerted the world. Such reports came almost exclusively from Japan, which consumes more than 75% of the world supply, even before the novel influenza pandemic started. The Japanese government will not revoke its warning issued in March 2007 against the use of oseltamivir in teenagers despite the accumulating evidence that such abnormal behaviors are part of generic symptoms of influenza-like delirium and not attributable to certain drugs. The author analyzes the background of the 'ado' by compiling the various sources of information, some of which have not been readily available to the international audience and explains why Japan is incapable of producing firm evidence to draw a definite conclusion. The author also alarms the potential risk of sudden death related to oseltamivir and foresees how the problem may be solved in the future. PMID:20121561

  7. Can we predict the intracellular metabolic state of a cell based on extracellular metabolite data?

    PubMed

    Granucci, Ninna; Pinu, Farhana R; Han, Ting-Li; Villas-Boas, Silas G

    2015-12-01

    The analysis of extracellular metabolites presents many technical advantages over the analysis of intracellular compounds, which made this approach very popular in recent years as a high-throughput tool to assess the metabolic state of microbial cells. However, very little effort has been made to determine the actual relationship between intracellular and extracellular metabolite levels. The secretion of intracellular metabolites has been traditionally interpreted as a consequence of an intracellular metabolic overflow, which is based on the premise that for a metabolite to be secreted, it must be over-produced inside the cell. Therefore, we expect to find a secreted metabolite at increased levels inside the cells. Here we present a time-series metabolomics study of Saccharomyces cerevisiae growing on a glucose-limited chemostat with parallel measurements of intra- and extracellular metabolites. Although most of the extracellular metabolites were also detected in the intracellular samples and showed a typical metabolic overflow behaviour, we demonstrate that the secretion of many metabolites could not be explained by the metabolic overflow theory. PMID:26400772

  8. Associations of cord blood metabolites with early childhood obesity risk

    PubMed Central

    Isganaitis, Elvira; Rifas-Shiman, Sheryl L.; Oken, Emily; Dreyfuss, Jonathan; Gall, Walt; Gillman, Matthew W.; Patti, Mary-Elizabeth

    2015-01-01

    Background/Objective Rapid postnatal weight gain is a potentially modifiable risk factor for obesity and metabolic syndrome. To identify markers of rapid infancy weight gain and childhood obesity, we analyzed the metabolome in cord blood from infants differing in their postnatal weight trajectories. Methods We performed a nested case-control study within Project Viva, a longitudinal cohort of mothers and children. We selected cases (n=26) based on top quartile of change in weight-for-age 0-6 mo and BMI >85th percentile in mid-childhood (median 7.7 years). Controls (n=26) were age- and sex-matched, had normal postnatal weight gain (2nd or 3rd quartile of change in weight-for-age 0-6 mo) and normal mid-childhood weight (BMI 25th-75th percentile). Cord blood metabolites were measured using untargeted LC/MS; individual metabolites and pathways differing between cases vs. controls were compared in categorical analyses. We adjusted metabolites for maternal age, maternal BMI, and breastfeeding duration (linear regression), and assessed whether metabolites improved the ability to predict case-control status (logistic regression). Results Of 415 detected metabolites, 16 were altered in cases vs. controls (T-test, nominal P<0.05). 3 metabolites were related to tryptophan: serotonin, tryptophan betaine, and tryptophyl leucine (46%, 48% and 26% lower in cases, respectively, P<0.05). Mean levels of 2 methyl donors, dimethylglycine and N-acetylmethionine, were also lower in cases (18% and 16% respectively, P=0.01). Moreover, the glutamine:glutamate ratio was reduced by 33% (P<0.05) in cases. Levels of serotonin, tryptophyl leucine, and N-acetylmethionine remained significantly different after adjustment for maternal BMI, age, and breastfeeding. Adding metabolite levels to logistic regression models including only clinical covariates improved the ability to predict case vs. control status. Conclusions Several cord blood metabolites are associated with rapid postnatal weight gain

  9. Mass spectrometry-based imaging of metabolites and proteins.

    PubMed

    Peukert, Manuela; Becker, Michael; Matros, Andrea; Mock, Hans-Peter

    2014-01-01

    Imaging techniques based on mass spectrometry (MS) have become powerful approaches to decipher the spatial distribution of metabolites and proteins. MS imaging (MSI) mostly relies on matrix-assisted laser desorption/ionization coupled to MS detection, but desorption electrospray ionization is also frequently used. Here we describe our current protocols for MALDI-MSI of seed sections and for root tissue. Detailed procedures for cryo-sectioning, matrix application, image capture, mass spectrometry measurement and data analysis are given. PMID:24136526

  10. Monitoring of propofol and its metabolite during total intravenous anesthesia

    NASA Astrophysics Data System (ADS)

    Elizarov, A. Yu.; Ershov, T. D.; Levshankov, A. I.

    2011-12-01

    Intravenous hypnotic propofol and its metabolite are detected in real time during total intravenous anesthesia by an electron ionization mass spectrometer. The mass spectrometer is connected directly to the breathing circuit of an apparatus for inhalational anesthesia. Ratios between the propofol concentrations in expired air and blood serum are measured. It is concluded that real-time noninvasive monitoring of the propofol concentration in blood using electron ionization mass spectrometry is feasible.

  11. Reactive metabolites in early drug development: predictive in vitro tools.

    PubMed

    Pelkonen, Olavi; Pasanen, Markku; Tolonen, Ari; Koskinen, Mikko; Hakkola, Jukka; Abass, Khaled; Laine, Jaana; Hakkinen, Merja; Juvonen, Risto; Auriola, Seppo; Storvik, Markus; Huuskonen, Pasi; Rousu, Timo; Rahikkala, Maiju

    2015-01-01

    Drug metabolism can result in the formation of highly reactive metabolites that are known to play a role in toxicity resulting in a significant proportion of attrition during drug development and clinical use. Thus, the earlier such reactivity was detected, the better. This review summarizes our multi-year project, together with pertinent literature, to examine a battery of in vitro tests capable of detecting the formation of reactive metabolites. Principal prerequisites for such tests were delineated: chemicals known/not known to cause tissue injury and produce reactive metabolites, activation system (mainly human-derived), small- and large-molecular targets (small-molecular trappers, peptides, proteins), analytical techniques (mass spectrometry), and cellular toxicity biomarkers. The current status of in vitro tools to detect reactive intermediates is the following: 1. Small-molecular trapping agents such glutathione or cyanide detect the production of reactive species with high sensitivity by proper MS technique. However, it seems that also putative "negatives" give rise to corresponding adducts. 2. Results from peptide and dG (DNA targeting) trapper studies are generally in line with those of small-molecular trappers, although also important differences exist. These two trapping platforms do not overlap. 3. It is anticipated that the in vitro adduct studies could be fully interpreted only in conjunction with toxicity biomarker (such as the Nrf2 pathway) information from whole cells or tissues. However, while there are tools to characterize the chemical liability and there are correlation between individual/integrated endpoints and toxicity, there are still severe gaps in understanding the mechanisms behind the link between reactive metabolites and adverse effects. PMID:25312212

  12. Metabolites identification of bioactive licorice compounds in rats.

    PubMed

    Wang, Qi; Qian, Yi; Wang, Qing; Yang, Yan-Fang; Ji, Shuai; Song, Wei; Qiao, Xue; Guo, De-An; Liang, Hong; Ye, Min

    2015-11-10

    Licorice (Glycyrrhiza uralensis Fisch.) is one of the most popular herbal medicines worldwide. This study aims to identify the metabolites of seven representative bioactive licorice compounds in rats. These compounds include 22β-acetoxyl glycyrrhizin (1), licoflavonol (2), licoricidin (3), licoisoflavanone (4), isoglycycoumarin (5), semilicoisoflavone B (6), and 3-methoxy-9-hydroxy-pterocarpan (7). After oral administration of 250mg/kg of 1 or 40mg/kg of 2-7 to rats, a total of 16, 43 and 31 metabolites were detected in the plasma, urine and fecal samples, respectively. The metabolites were characterized by HPLC/DAD/ESI-MS(n) and LC/IT-TOF-MS analyses. Particularly, two metabolites of 1 were unambiguously identified by comparing with reference standards, and 22β-acetoxyl glycyrrhizin-6″-methyl ester (1-M2) is a new compound. Compound 1 could be readily hydrolyzed to eliminate the glucuronic acid residue. The phenolic compounds (4-7) mainly undertook phase II metabolism (glucuronidation or sulfation). Most phenolic compounds with an isoprenyl group (chain or cyclized, 2-5) could also undertake hydroxylation reaction. This is the first study on in vivo metabolism of these licorice compounds. PMID:26311472

  13. Synthesis of the alkylated active metabolite of tipidogrel.

    PubMed

    Zhi, Shuang; Xia, Guangping; Liu, Ying; Tao, Zunwei; Chen, Ligong; Liu, Dengke

    2015-04-15

    Tipidogrel (3), an effective anti-platelet drug candidate working by irreversibly inhibiting P2Y12 receptor, holds great promise in overcoming clopidogrel resistance and increasing bioavailability. As a prodrug like other thienopyridines, it metabolizes through thiophene ring opening to form active metabolites 3a and 3b, nevertheless they are easily to form disulfide bond. Derivatization of 3a and 3b via alkylation with MPBr can prevent disulfide conjugation and ensure reliable pharmacokinetic results. Thus, in order to support its pre-clinical studies on efficiencies in the formation of tipidogrel active metabolites, 13a and 13b were synthesized via seven steps of chemosynthesis and incubation with MPBr in rat plasma in vitro. The resulting crude productions were purified by semi-preparative HPLC to give Z configuration 13a and E configuration 13b. In LC-MS/MS spectra, they showed identical fragmentation pattern and retention time with M-13a and M-13b, the MPBr-derivatives of active metabolites of tipidogrel in rats. Thus, 13a and 13b were the anticipated alkylated active metabolite of tipidogrel. In addition, in the nucleophilic substitution of thioacetate with compound 11, besides the anticipated compounds 12a and 12b, their isomers compounds 12c and 12d were detected, whose structures were confirmed and the corresponding mechanism was presented. PMID:25801935

  14. Effects of progesterone and its metabolites on human granulosa cells.

    PubMed

    Pietrowski, D; Gong, Y; Mairhofer, M; Gessele, R; Sator, M

    2014-02-01

    The corpus luteum (CL) is under control of gonadotrophic hormones and produces progesterone, which is necessary for endometrial receptivity. Recent studies have shown that progesterone and its metabolites are involved in cell proliferation and apoptosis of cancer cells. Here weanalyzed the role of progesterone and its meta-bolites on luteinized granulosa cells (LGC) by FACS analysis and quantitative Real-Time PCR. We detected the mRNA of the progesterone metabolizing genes SRD5A1, AKR1C1, and AKR1C2 in LGC. The stimulation of LGC with progesterone or progesterone metabolites did not show any effect on the mRNA expression of these genes. However, a downregulation of Fas expression was found to be accomplished by progesterone and human chorionic gonadotropin. Our findings do not support the concept of an effect of progesterone metabolites on LGCs. However, it suggests an antiapoptotic effect of hCG and progesterone during corpus luteum development by downregulation of Fas. PMID:24136781

  15. Profiling of Serum Metabolites in Canine Lymphoma Using Gas Chromatography Mass Spectrometry

    PubMed Central

    TAMAI, Reo; FURUYA, Masaru; HATOYA, Shingo; AKIYOSHI, Hideo; YAMAMOTO, Ryohei; KOMORI, Yoshiaki; YOKOI, Shin-ichi; TANI, Kenichiro; HIRANO, Yuji; KOMORI, Masayuki; TAKENAKA, Shigeo

    2014-01-01

    ABSTRACT Canine lymphoma is a common cancer that has high rates of complete remission with combination chemotherapy. However, the duration of remission varies based on multiple factors, and there is a need to develop a method for early detection of recurrence. In this study, we compared the metabolites profiles in serum from 21 dogs with lymphoma and 13 healthy dogs using gas chromatography mass spectrometry (GC-MS). The lymphoma group was separated from the control group in an orthogonal projection to latent structure with discriminant analysis (OPLS-DA) plot using ions of m/z 100–600, indicating that the metabolites profiles in lymphoma cases differed from those in healthy dogs. The lymphoma group was also separated from the control group on OPLS-DA plot using 29 metabolites identified in all serum samples. Significant differences were found for 16 of these metabolites with higher levels in the lymphoma group for 15 of the metabolites and lower levels for inositol. An OPLS-DA plot showed separation of the lymphoma and healthy groups using these 16 metabolites only. These results indicate that metabolites profile with GC-MS may be a useful tool for detection of potential biomarker and diagnosis of canine lymphoma. PMID:25131950

  16. Phthalate metabolites in the European eel (Anguilla anguilla) from Mediterranean coastal lagoons.

    PubMed

    Fourgous, C; Chevreuil, M; Alliot, F; Amilhat, E; Faliex, E; Paris-Palacios, S; Teil, M J; Goutte, A

    2016-11-01

    The levels and fate of phthalate metabolites have been poorly evaluated in fish, despite their potential ecotoxicological impacts. The present study aims to characterize the levels of phthalate metabolites in muscle tissue of yellow eels (Anguilla anguilla) from two coastal Mediterranean lagoons, during three sampling periods. Nine phthalate metabolites were detected in >70% of the samples. Slightly higher levels of phthalate metabolites were detected in March and June compared to October, suggesting possible seasonal variations in environmental release and/or phthalate metabolization process by eels. The large sample size (N=117) made it possible to explore correlations between phthalate metabolites' levels and individual parameters, such as body length, age, body condition and hepatic histo-pathologies. Body length and estimated age poorly correlated with phthalate metabolites, suggesting that eels did not accumulate phthalates during growth, contrary to persistent compounds. Eels presented different grades of hepatic fibrosis and lipidosis. A negative correlation was found between the severity of these pathologies in the liver and the sum of phthalate metabolites levels, supporting the hypothesis that eels with damaged liver are less able to metabolize xenobiotics. PMID:27412480

  17. Seed metabolomic study reveals significant metabolite variations and correlations among different soybean cultivars.

    PubMed

    Lin, Hong; Rao, Jun; Shi, Jianxin; Hu, Chaoyang; Cheng, Fang; Wilson, Zoe A; Zhang, Dabing; Quan, Sheng

    2014-09-01

    Soybean [Glycine max (L.) Merr.] is one of the world's major crops, and soybean seeds are a rich and important resource for proteins and oils. While "omics" studies, such as genomics, transcriptomics, and proteomics, have been widely applied in soybean molecular research, fewer metabolomic studies have been conducted for large-scale detection of low molecular weight metabolites, especially in soybean seeds. In this study, we investigated the seed metabolomes of 29 common soybean cultivars through combined gas chromatography-mass spectrometry and ultra-performance liquid chromatography-tandem mass spectrometry. One hundred sixty-nine named metabolites were identified and subsequently used to construct a metabolic network of mature soybean seed. Among the 169 detected metabolites, 104 were found to be significantly variable in their levels across tested cultivars. Metabolite markers that could be used to distinguish genetically related soybean cultivars were also identified, and metabolite-metabolite correlation analysis revealed some significant associations within the same or among different metabolite groups. Findings from this work may potentially provide the basis for further studies on both soybean seed metabolism and metabolic engineering to improve soybean seed quality and yield. PMID:24942044

  18. Screening botanical extracts for quinoid metabolites.

    PubMed

    Johnson, B M; Bolton, J L; van Breemen, R B

    2001-11-01

    Botanical dietary supplements represent a significant share of the growing market for alternative medicine in the USA, where current regulations do not require assessment of their safety. To help ensure the safety of such products, an in vitro assay using pulsed ultrafiltration and LC-MS-MS has been developed to screen botanical extracts for the formation of electrophilic and potentially toxic quinoid species upon bioactivation by hepatic cytochromes P450. Rat liver microsomes were trapped in a flow-through chamber by an ultrafiltration membrane, and samples containing botanical extracts, GSH and NADP(H), were flow-injected into the chamber. Botanical compounds that were metabolized to reactive intermediates formed stable GSH adducts mimicking a common in vivo detoxification pathway. If present in the ultrafiltrate, GSH conjugates were detected using LC-MS-MS with precursor ion scanning followed by additional characterization using product ion scanning and comparison to standard compounds. As expected, no GSH adducts of reactive metabolites were found in extracts of Trifolium pratense L. (red clover), which are under investigation as botanical dietary supplements for the management of menopause. However, extracts of Sassafras albidum (Nutt.) Nees (sassafras), Symphytum officinale L. (comfrey), and Rosmarinus officinalis L. (rosemary), all of which are known to contain compounds that are either carcinogenic or toxic to mammals, produced GSH adducts during this screening assay. Several compounds that formed GSH conjugates including novel metabolites of rosmarinic acid were identified using database searching and additional LC-MS-MS studies. This assay should be useful as a preliminary toxicity screen during the development of botanical dietary supplements. A positive test suggests that additional toxicological studies are warranted before human consumption of a botanical product. PMID:11712913

  19. In Vivo and Real-time Monitoring of Secondary Metabolites of Living Organisms by Mass Spectrometry

    NASA Astrophysics Data System (ADS)

    Hu, Bin; Wang, Lei; Ye, Wen-Cai; Yao, Zhong-Ping

    2013-07-01

    Secondary metabolites are compounds that are important for the survival and propagation of animals and plants. Our current understanding on the roles and secretion mechanism of secondary metabolites is limited by the existing techniques that typically cannot provide transient and dynamic information about the metabolic processes. In this manuscript, by detecting venoms secreted by living scorpion and toad upon attack and variation of alkaloids in living Catharanthus roseus upon stimulation, which represent three different sampling methods for living organisms, we demonstrated that in vivo and real-time monitoring of secondary metabolites released from living animals and plants could be readily achieved by using field-induced direct ionization mass spectrometry.

  20. Exposure measurement of aflatoxins and aflatoxin metabolites in human body fluids. A short review.

    PubMed

    Leong, Yin-Hui; Latiff, Aishah A; Ahmad, Nurul Izzah; Rosma, Ahmad

    2012-05-01

    Aflatoxins are highly toxic secondary fungal metabolites mainly produced by Aspergillus flavus and A. parasiticus. Human exposure to aflatoxins may result directly from ingestion of contaminated foods, or indirectly from consumption of foods from animals previously exposed to aflatoxins in feeds. This paper focuses on exposure measurement of aflatoxins and aflatoxin metabolites in various human body fluids. Research on different metabolites present in blood, urine, breast milk, and other human fluids or tissues including their detection techniques is reviewed. The association between dietary intake of aflatoxins and biomarker measurement is also highlighted. Finally, aspects related to the differences between aflatoxin determination in food versus the biomarker approach are discussed. PMID:23606045

  1. Untargeted Profiling of Tracer-Derived Metabolites Using Stable Isotopic Labeling and Fast Polarity-Switching LC–ESI-HRMS

    PubMed Central

    2014-01-01

    An untargeted metabolomics workflow for the detection of metabolites derived from endogenous or exogenous tracer substances is presented. To this end, a recently developed stable isotope-assisted LC–HRMS-based metabolomics workflow for the global annotation of biological samples has been further developed and extended. For untargeted detection of metabolites arising from labeled tracer substances, isotope pattern recognition has been adjusted to account for nonlabeled moieties conjugated to the native and labeled tracer molecules. Furthermore, the workflow has been extended by (i) an optional ion intensity ratio check, (ii) the automated combination of positive and negative ionization mode mass spectra derived from fast polarity switching, and (iii) metabolic feature annotation. These extensions enable the automated, unbiased, and global detection of tracer-derived metabolites in complex biological samples. The workflow is demonstrated with the metabolism of 13C9-phenylalanine in wheat cell suspension cultures in the presence of the mycotoxin deoxynivalenol (DON). In total, 341 metabolic features (150 in positive and 191 in negative ionization mode) corresponding to 139 metabolites were detected. The benefit of fast polarity switching was evident, with 32 and 58 of these metabolites having exclusively been detected in the positive and negative modes, respectively. Moreover, for 19 of the remaining 49 phenylalanine-derived metabolites, the assignment of ion species and, thus, molecular weight was possible only by the use of complementary features of the two ion polarity modes. Statistical evaluation showed that treatment with DON increased or decreased the abundances of many detected metabolites. PMID:25372979

  2. Novel rapid liquid chromatography tandem masspectrometry method for vemurafenib and metabolites in human plasma, including metabolite concentrations at steady state.

    PubMed

    Vikingsson, Svante; Strömqvist, Malin; Svedberg, Anna; Hansson, Johan; Höiom, Veronica; Gréen, Henrik

    2016-08-01

    A novel, rapid and sensitive liquid chromatography tandem-mass spectrometry method for quantification of vemurafenib in human plasma, that also for the first time allows for metabolite semi-quantification, was developed and validated to support clinical trials and therapeutic drug monitoring. Vemurafenib was analysed by precipitation with methanol followed by a 1.9 min isocratic liquid chromatography tandem masspectrometry analysis using an Acquity BEH C18 column with methanol and formic acid using isotope labelled internal standards. Analytes were detected in multireaction monitoring mode on a Xevo TQ. Semi-quantification of vemurafenib metabolites was performed using the same analytical system and sample preparation with gradient elution. The vemurafenib method was successfully validated in the range 0.5-100 μg/mL according to international guidelines. The metabolite method was partially validated owing to the lack of commercially available reference materials. For the first time concentration levels at steady state for melanoma patients treated with vemurafenib is presented. The low abundance of vemurafenib metabolites suggests that they lack clinical significance. Copyright © 2016 John Wiley & Sons, Ltd. PMID:26683023

  3. Formed and preformed metabolites: facts and comparisons.

    PubMed

    Pang, K Sandy; Morris, Marilyn E; Sun, Huadong

    2008-10-01

    The administration of metabolites arising from new drug entities is often employed in drug discovery to investigate their associated toxicity. It is expected that administration of metabolites can predict the exposure of metabolites originating from the administration of precursor drug. Whether exact and meaningful information can be obtained from this has been a topic of debate. This communication summarizes observations and theoretical relationships based on physiological modelling for the liver, kidney and intestine, three major eliminating organs/tissues. Theoretical solutions based on physiological modelling of organs were solved, and the results suggest that deviations are expected. Here, examples of metabolite kinetics observed mostly in perfused organs that did not match predictions are provided. For the liver, discrepancies in fate between formed and preformed metabolites may be explained by the heterogeneity of enzymes, the presence of membrane barriers and whether transporters are involved. For the kidney, differences have been attributed to glomerular filtration of the preformed but not the formed metabolite. For the intestine, the complexity of segregated flows to the enterocyte and serosal layers and differences in metabolism due to the route of administration are addressed. Administration of the metabolite may or may not directly reflect the toxicity associated with drug use. However, kinetic data on the preformed metabolite will be extremely useful to develop a sound model for modelling and simulations; in-vitro evidence on metabolite handling at the target organ is also paramount. Subsequent modelling and simulation of metabolite data arising from a combined model based on both drug and preformed metabolite data are needed to improve predictions on the behaviours of formed metabolites. PMID:18812018

  4. Identification of catechol and hydroquinone metabolites of 4-monochlorobiphenyl.

    PubMed

    McLean, M R; Bauer, U; Amaro, A R; Robertson, L W

    1996-01-01

    Polychlorinated biphenyls (PCBs) may be metabolically activated to electrophiles, which bind to proteins and nucleic acids. One activation scheme involves the formation of reactive arene oxide intermediates during cytochrome P450-catalyzed hydroxylation. We propose a second activation pathway whereby PCB catechol and hydroquinone metabolites may be oxidized to reactive semiquinones and/or quinones. By employing 4-monochlorobiphenyl (4-MCB) as a model substrate and liver microsomes from rats treated with phenobarbital and 3-methyl-cholanthrene, five monol and three diol metabolites were identified. The major metabolite was 4-chloro-4'-monohydroxybiphenyl, followed by, in decreasing order, 4-chloro-3',4'-dihydroxybiphenyl, unknown B (a monol), 4-chloro-2',3'-dihydroxybiphenyl, 4-chloro-3'-hydroxybiphenyl, 4-chloro-2',5'-dihydroxybiphenyl, unknown A (a monol), and 4-chloro-2'-monohydroxybiphenyl. A trace of a dihydrodiol was detected by GC/MS. To elucidate the source of the diols, 4-MCB and the synthetic monol metabolites 4-chloro-2'-/-3'-/-4'-monohydroxybiphenyls were each employed as substrates in incubations with microsomes from rats treated with phenobarbital, 3-methylcholanthrene, or both inducers. The three diol metabolites were all produced from 4-MCB in incubations with microsomes from 3-methylcholanthrene-treated rats, but incubations with microsomes from phenobarbital-treated rats did not yield detectable amounts of 4-chloro-2',3'-dihydroxybiphenyl. 4-Chloro-2',3'-dihydroxybiphenyl was only found as a product of 4-chloro-2'-monohydroxybiphenyl. The 4-chloro-2',5'-dihydroxybiphenyl was found in extracts of incubations with 4-chloro-2'- and -3'-monohydroxybiphenyls, while the 4-chloro-3',4'-dihydroxybiphenyl was the only product found from 4-chloro-3'- and -4'-monohydroxybiphenyls. No other chlorinated diols were detected by GC/MS. These data suggest that the major route of biosynthesis of the diols was via a second hydroxylation step and not aromatization of

  5. Aspirin metabolites are GPR35 agonists.

    PubMed

    Deng, Huayun; Fang, Ye

    2012-07-01

    Aspirin is widely used as an anti-inflammatory, anti-platelet, anti-pyretic, and cancer-preventive agent; however, the molecular mode of action is unlikely due entirely to the inhibition of cyclooxygenases. Here, we report the agonist activity of several aspirin metabolites at GPR35, a poorly characterized orphan G protein-coupled receptor. 2,3,5-Trihydroxybenzoic acid, an aspirin catabolite, was found to be the most potent GPR35 agonist among aspirin metabolites. Salicyluric acid, the main metabolite of aspirin, was also active. These results suggest that the GPR35 agonist activity of certain aspirin metabolites may contribute to the clinical features of aspirin. PMID:22526472

  6. From known knowns to known unknowns: predicting in vivo drug metabolites.

    PubMed

    Pelkonen, Olavi; Tolonen, Ari; Korjamo, Timo; Turpeinen, Miia; Raunio, Hannu

    2009-05-01

    'It is better to be useful than perfect'. This review attempts to critically cover and assess the currently available approaches and tools to answer the crucial question: Is it possible (and if it is, to what extent is it possible) to predict in vivo metabolites and their abundances on the basis of in vitro and preclinical animal studies? In preclinical drug development, it is possible to produce metabolite patterns from a candidate drug by virtual means (i.e., in silico models), but these are not yet validated. However, they may be useful to cover the potential range of metabolites. In vitro metabolite patterns and apparent relative abundances are produced by various in vitro systems employing tissue preparations (mainly liver) and in most cases using liquid chromatography-mass spectrometry analytical techniques for tentative identification. The pattern of the metabolites produced depends on the enzyme source; the most comprehensive source of drug-metabolizing enzymes is cultured human hepatocytes, followed by liver homogenate fortified with appropriate cofactors. For specific purposes, such as the identification of metabolizing enzyme(s), recombinant enzymes can be used. Metabolite data from animal in vitro and in vivo experiments, despite known species differences, may help pinpoint metabolites that are not apparently produced in in vitro human systems, or suggest alternative experimental approaches. The range of metabolites detected provides clues regarding the enzymes attacking the molecule under study. We also discuss established approaches to identify the major enzymes. The last question, regarding reliability and robustness of metabolite extrapolations from in vitro to in vivo, both qualitatively and quantitatively, cannot be easily answered. There are a number of examples in the literature suggesting that extrapolations are generally useful, but there are only a few systematic and comprehensive studies to validate in vitro-in vivo extrapolations. In

  7. Untargeted MS-based small metabolite identification from the plant leaves and stems of Impatiens balsamina.

    PubMed

    Chua, Lee Suan

    2016-09-01

    The identification of plant metabolites is very important for the understanding of plant physiology including plant growth, development and defense mechanism, particularly for herbal medicinal plants. The metabolite profile could possibly be used for future drug discovery since the pharmacological activities of the indigenous herbs have been proven for centuries. An untargeted mass spectrometric approach was used to identify metabolites from the leaves and stems of Impatiens balsamina using LC-DAD-MS/MS. The putative compounds are mostly from the groups of phenolic, organic and amino acids which are essential for plant growth and as intermediates for other compounds. Alanine appeared to be the main amino acid in the plant because many alanine derived metabolites were detected. There are also several secondary metabolites from the groups of benzopyrones, benzofuranones, naphthoquinones, alkaloids and flavonoids. The widely reported bioactive components such as kaempferol, quercetin and their glycosylated, lawsone and its derivatives were detected in this study. The results also revealed that aqueous methanol could extract flavonoids better than water, and mostly, flavonoids were detected from the leaf samples. The score plots of component analysis show that there is a minor variance in the metabolite profiles of water and aqueous methanolic extracts with 21.5 and 30.5% of the total variance for the first principal component at the positive and negative ion modes, respectively. PMID:27135814

  8. Bioanalytical methods for determination of tamoxifen and its phase I metabolites: a review.

    PubMed

    Teunissen, S F; Rosing, H; Schinkel, A H; Schellens, J H M; Beijnen, J H

    2010-12-17

    The selective estrogen receptor modulator tamoxifen is used in the treatment of early and advanced breast cancer and in selected cases for breast cancer prevention in high-risk subjects. The cytochrome P450 enzyme system and flavin-containing monooxygenase are responsible for the extensive metabolism of tamoxifen into several phase I metabolites that vary in toxicity and potencies towards estrogen receptor (ER) alpha and ER beta. An extensive overview of publications on the determination of tamoxifen and its phase I metabolites in biological samples is presented. In these publications techniques were used such as capillary electrophoresis, liquid, gas and thin layer chromatography coupled with various detection techniques (mass spectrometry, ultraviolet or fluorescence detection, liquid scintillation counting and nuclear magnetic resonance spectroscopy). A trend is seen towards the use of liquid chromatography coupled to mass spectrometry (LC-MS). State-of-the-art LC-MS equipment allowed for identification of unknown metabolites and quantification of known metabolites reaching lower limit of quantification levels in the sub pg mL(-1) range. Although tamoxifen is also metabolized into phase II metabolites, the number of publications reporting on phase II metabolism of tamoxifen is scarce. Therefore the focus of this review is on phase I metabolites of tamoxifen. We conclude that in the past decades tamoxifen metabolism has been studied extensively and numerous metabolites have been identified. Assays have been developed for both the identification and quantification of tamoxifen and its metabolites in an array of biological samples. This review can be used as a resource for method transfer and development of analytical methods used to support pharmacokinetic and pharmacodynamic studies of tamoxifen and its phase I metabolites. PMID:21094378

  9. Benzodiazepines and metabolites from biological fluids by liquid chromatography electrospray tandem mass spectrometry.

    PubMed

    Morris-Kukoski, Cynthia L; Schaff, Jason E; Reda, Louis J

    2012-01-01

    Solid-phase extraction and liquid chromatography-tandem mass spectrometry are invaluable techniques for the determination of benzodiazepines and metabolites in biological matrices. The reason for using tandem mass spectrometry is to increase limits of detection without the need for chemical derivatization. Here we describe a technique for the detection of 26 benzodiazepines and metabolites at a detection limit of approximately 1-2 ng/mL in blood and 1-5 ng/mL in urine when screened using a data-dependent scan method. PMID:22767106

  10. Atrazine and its metabolites as indicators of stream-aquifer interaction in Kansas, USA

    USGS Publications Warehouse

    Townsend, M.A.; Young, D.P.

    2000-01-01

    A survey of atrazine and its metabolites in Kansas ground water indicated that ground-water quality was impacted by stream-aquifer interaction between rivers in the Kansas River basin and their adjacent alluvial aquifers. Atrazine was detected in 19 of the 78 samples. The most common metabolite, deethylatrazine, was detected in 25 samples, 18 of which also had atrazine. The deethylatrazine/atrazine ratio (DAR) of < 1.0 indicates rapid movement of agricultural chemicals to ground water. In this study, 12 of 18 samples had DAR values < 1.0, suggesting rapid recharge to the aquifers. Hydroxyatrazine is seldom detected in ground water. In this study hydroxyatrazine was detected primarily in wells sited in alluvium of rivers. These rivers contain atrazine in varying concentrations. Results of the study suggest that stream-aquifer interaction is a process contributing to the presence of both atrazine and its metabolites in ground water in these areas.A survey of atrazine and its metabolites in Kansas ground water indicated that ground water quality was impacted by stream-aquifer interaction between rivers in the Kansas River basin and their adjacent alluvial aquifers. Atrazine was detected in 19 of the 78 samples. The most common metabolite, deethylatrazine, was detected in 25 samples, 18 of which also had atrazine. The deethylatrazine/attrazine ratio (DAR) of < 1.0 indicates rapid movement of agricultural chemicals to ground water. In this study, 12 of 18 samples had DAR values < 1.0, suggesting rapid recharge to the aquifers. Hydroxyatrazine is seldom detected in ground water. In this study hydroxyatrazine was detected primarily in wells sited in alluvium of rivers. These rivers contain atrazine in varying concentration. Results of the study suggest that stream-aquifer interaction is a process contributing to the presence of both attrazine and its metabolites in ground water in these areas.

  11. Fractionation of the secondary metabolites of orange (Citrus sinensis L.) leaves by fast centrifugal partition chromatography

    Technology Transfer Automated Retrieval System (TEKTRAN)

    There is interest in the detection of changes in secondary metabolites in orange leaves in response to citrus greening disease. Conventional HPLC analysis readily provides detection of major phenolic compounds, but further, more detailed chromatographic analyses show many more compounds, to an exten...

  12. Electrochemical generation of selegiline metabolites coupled to mass spectrometry.

    PubMed

    Mielczarek, Przemyslaw; Smoluch, Marek; Kotlinska, Jolanta H; Labuz, Krzysztof; Gotszalk, Teodor; Babij, Michal; Suder, Piotr; Silberring, Jerzy

    2015-04-10

    The metabolic pathways of selegiline (a drug used for the treatment of early-stage Parkinson's disease) were analyzed by electrochemical oxidation with application of the flow electrochemical cell consisting of three electrodes (ROXY™, Antec, the Netherlands). Two types of working electrodes were applied: glassy carbon (GC) and boron-doped diamond (BDD). The potential applied at working electrode and composition of the solvent were optimized for the best conditions for oxidation and identification processes. All products were directly analyzed on-line by mass spectrometry. For further characterization of electrochemical oxidation products, the novel approach involving reversed phase chromatography linked to mass spectrometry with dielectric barrier discharge ionization (DBDI-MS) was used. In this manuscript, we report a novel technique for simulation of drug metabolism by electrochemical system (EC) connected to liquid chromatography (LC) and dielectric barrier discharge ionization (DBDI) mass spectrometry (MS) for direct on-line detection of electrochemical oxidation products. Here, we linked LC/DBDI-MS system with an electrochemical flow cell in order to study metabolic pathways via identification of drug metabolites generated electrochemically. The DBDI source has never been used before for identification of psychoactive metabolites generated in an electrochemical flow cell. Our knowledge on the biological background of xenobiotics metabolism and its influence on human body is constantly increasing, but still many mechanisms are not explained. Nowadays, metabolism of pharmaceuticals is mainly studied using liver cells prepared from animals or humans. Cytochrome P450, present in microsomes, is primarily responsible for oxidative metabolism of xenobiotics. It was also shown, that breakdown of popular medicines may be successfully simulated by electrochemistry under appropriate conditions. The presented experiments allow for comparison of these two entirely

  13. Identification of Microbial Metabolites Elevating Vitamin Contents in Barley Seeds.

    PubMed

    Yousaf, Anam; Qadir, Abdul; Anjum, Tehmina; Ahmad, Aqeel

    2015-08-19

    The current investigation analyzes metabolites of Acetobacter aceti to explore chemical compounds responsible for the induction of vitamins in barley seeds. A bioactivity guided assay of bacterial extracts and chromatographic analyses of barley produce revealed 13 chemical compounds, which were subjected to principal component analysis (PCA). PCA determined four chemical compounds (i.e., quinolinic acid, pyridoxic acid, p-aminobenzoate, and α-oxobutanoic acid) highly associated with increased quantities of vitamins. Further experimentations confirmed that quinolinic acid and p-aminobenzoate were the most efficient vitamin inducers. The results indicated chloroform/ethanol (4:1) as the best solvent system for the extraction of active compounds from crude metabolites of A. aceti. Significant quantities of mevalonic acid were detected in the extracted fraction, indicating the possible induction of the isoprenoid pathway. Altogether, the current investigation broadens the frontiers in plant-microbe interaction. PMID:26173019

  14. High resolution mass spectrometry to investigate omeprazole and venlafaxine metabolites in wastewater.

    PubMed

    Boix, Clara; Ibáñez, María; Bagnati, Renzo; Zuccato, Ettore; Sancho, Juan V; Hernández, Félix; Castiglioni, Sara

    2016-01-25

    This study reports an investigation of omeprazole and venlafaxine parent substances and metabolites in Italian municipal influent wastewaters (IWWs). These pharmaceuticals were selected because they are widely consumed in Italy, but are poorly detected in waste and surface water. The aim of the study was to identify the most relevant pharmaceuticals metabolites in wastewater in order to improve the prioritization step and choose priority pollutants for environmental monitoring campaigns. This was done by investigating omeprazole, venlafaxine and their main metabolites in 30 IWWs from ten Italian cities and by comparing results with information from pharmacokinetic studies. Analysis was performed by solid phase extraction (SPE) and high-performance liquid chromatography (HPLC) coupled to high resolution mass spectrometry (HRMS). We searched for 23 omeprazole and four venlafaxine metabolites using data-dependent and MS/MS methods. Parent omeprazole was never present in the samples. Six omeprazole metabolites were found in IWWs. Venlafaxine and two metabolites were present in all the samples. The metabolic profiles in Italian IWW agreed with results in IWW from Spain and with urinary excretion profiles from pharmacokinetic studies. Comparing results from different sources was useful to improve the identification of pharmaceuticals metabolites in environmental samples and to focus the attention of future studies on the most relevant compounds. PMID:26476321

  15. Identification of alcohol-dependent clopidogrel metabolites using conventional liquid chromatography/triple quadrupole mass spectrometry

    PubMed Central

    Hu, Zhe-Yi; Laizure, S. Casey; Herring, Vanessa L.; Parker, Robert B.

    2014-01-01

    RATIONALE Clopidogrel (CLO) is a prodrug used to prevent ischemic events in patients undergoing percutaneous coronary intervention or with myocardial infarction. A previous study found ethyl clopidogrel (ECLO) is formed by transesterification of CLO when incubated with alcohol in human liver microsomes. We hypothesize that ECLO will be subject to further metabolism and developed an assay to identify its metabolites. METHODS A liquid chromatography/triple quadrupole mass spectrometry (LC-MS/MS) method was developed to identify metabolites of ECLO. According to the predicted metabolic pathway of ECLO, precursor–product ion pairs were used to screen the possible metabolites of ECLO in human liver S9 fractions. Subsequently, the detected metabolites were characterized by the results of product ion scan. RESULTS In the presence of alcohol, CLO was tranesterified to ECLO, which was further oxidized to form ethylated 2-oxo-clopidogrel and several ethylated thiol metabolites including the ethylated form of the H4 active metabolite. CONCLUSIONS The ECLO formed by transesterification with alcohol is subject to metabolism by CYP450 enzymes producing ethylated forms of 2-oxo-clopidogrel and the active H4 thiol metabolite. PMID:24760569

  16. Targeted serum metabolite profiling and sequential metabolite ratio analysis for colorectal cancer progression monitoring.

    PubMed

    Zhu, Jiangjiang; Djukovic, Danijel; Deng, Lingli; Gu, Haiwei; Himmati, Farhan; Abu Zaid, Mohammad; Chiorean, Elena Gabriela; Raftery, Daniel

    2015-10-01

    Colorectal cancer (CRC) is one of the most prevalent cancers worldwide and a major cause of human morbidity and mortality. In addition to early detection, close monitoring of disease progression in CRC can be critical for patient prognosis and treatment decisions. Efforts have been made to develop new methods for improved early detection and patient monitoring; however, research focused on CRC surveillance for treatment response and disease recurrence using metabolomics has yet to be reported. In this proof of concept study, we applied a targeted liquid chromatography tandem mass spectrometry (LC-MS/MS) metabolic profiling approach focused on sequential metabolite ratio analysis of serial serum samples to monitor disease progression from 20 CRC patients. The use of serial samples reduces patient to patient metabolic variability. A partial least squares-discriminant analysis (PLS-DA) model using a panel of five metabolites (succinate, N2, N2-dimethylguanosine, adenine, citraconic acid, and 1-methylguanosine) was established, and excellent model performance (sensitivity = 0.83, specificity = 0.94, area under the receiver operator characteristic curve (AUROC) = 0.91 was obtained, which is superior to the traditional CRC monitoring marker carcinoembryonic antigen (sensitivity = 0.75, specificity = 0.76, AUROC = 0.80). Monte Carlo cross validation was applied, and the robustness of our model was clearly observed by the separation of true classification models from the random permutation models. Our results suggest the potential utility of metabolic profiling for CRC disease monitoring. PMID:26342311

  17. Complicating factors in safety testing of drug metabolites: Kinetic differences between generated and preformed metabolites

    SciTech Connect

    Prueksaritanont, Thomayant . E-mail: thomayant_prueksaritanont@merck.com; Lin, Jiunn H.; Baillie, Thomas A.

    2006-12-01

    This paper aims to provide a scientifically based perspective on issues surrounding the proposed toxicology testing of synthetic drug metabolites as a means of ensuring adequate nonclinical safety evaluation of drug candidates that generate metabolites considered either to be unique to humans or are present at much higher levels in humans than in preclinical species. We put forward a number of theoretical considerations and present several specific examples where the kinetic behavior of a preformed metabolite given to animals or humans differs from that of the corresponding metabolite generated endogenously from its parent. The potential ramifications of this phenomenon are that the results of toxicity testing of the preformed metabolite may be misleading and fail to characterize the true toxicological contribution of the metabolite when formed from the parent. It is anticipated that such complications would be evident in situations where (a) differences exist in the accumulation of the preformed versus generated metabolites in specific tissues, and (b) the metabolite undergoes sequential metabolism to a downstream product that is toxic, leading to differences in tissue-specific toxicity. Owing to the complex nature of this subject, there is a need to treat drug metabolite issues in safety assessment on a case-by-case basis, in which a knowledge of metabolite kinetics is employed to validate experimental paradigms that entail administration of preformed metabolites to animal models.

  18. A New Paradigm for Known Metabolite Identification in Metabonomics/Metabolomics: Metabolite Identification Efficiency

    PubMed Central

    Everett, Jeremy R.

    2015-01-01

    A new paradigm is proposed for assessing confidence in the identification of known metabolites in metabonomics studies using NMR spectroscopy approaches. This new paradigm is based upon the analysis of the amount of metabolite identification information retrieved from NMR spectra relative to the molecular size of the metabolite. Several new indices are proposed including: metabolite identification efficiency (MIE) and metabolite identification carbon efficiency (MICE), both of which can be easily calculated. These indices, together with some guidelines, can be used to provide a better indication of known metabolite identification confidence in metabonomics studies than existing methods. Since known metabolite identification in untargeted metabonomics studies is one of the key bottlenecks facing the science currently, it is hoped that these concepts based on molecular spectroscopic informatics, will find utility in the field. PMID:25750701

  19. Occurrence of selected pesticides and their metabolites in near-surface aquifers of the midwestern United States

    USGS Publications Warehouse

    Kolpin, D.W.; Michael, Thurman E.; Goolsby, D.A.

    1996-01-01

    The occurrence and distribution of selected pesticides and their metabolites were investigated through the collection of 837 water-quality samples from 303 wells across the Midwest. Results of this study showed that five of the six most frequently detected compounds were pesticide metabolites. Thus, it was common for a metabolite to be found more frequently in groundwater than its parent compound. The metabolite alachlor ethanesulfonic acid (alachlor-ESA; 2-[(2,6-diethylphenyl)(methoxymethyl)amino]-2-oxoethanesulfonic acid) was detected almost 10 times as frequently and at much higher concentrations than its parent compound alachlor (2-chloro-2‘,6‘-diethyl-N-(methoxymethyl)acetamide). The median detectable atrazine (2-chloro-4-ethylamino-6- isopropylamino-s-triazine) concentration was almost half that of atrazine residue (atrazine plus the two atrazine metabolites analyzed). Cyanazine amide [2-chloro-4-(1-carbamoyl-1-methylethylamino)-6-ethylamino-s-triazine] was detected almost twice as frequently as cyanazine (2-chloro-4-ethylamino-6-methylpropionitrileamino-s-triazine). Results show that information on pesticide metabolites is necessary to understand the environmental fate of pesticides. Consequently, if pesticide metabolites are not quantified, the effects of chemical use on groundwater quality would be substantially underestimated. Thus, continued research is needed to identify major degradation pathways for all pesticides and to develop analytical methods to determine their concentrations in water and other environmental media.

  20. Shotgun Metabolomics Approach for the Analysis of Negatively Charged Water-Soluble Cellular Metabolites from Mouse Heart Tissue

    PubMed Central

    Sun, Gang; Yang, Kui; Zhao, Zhongdan; Guan, Shaoping; Han, Xianlin; Gross, Richard W.

    2010-01-01

    A shotgun metabolomics approach using MALDI-TOF/TOF mass spectrometry was developed for the rapid analysis of negatively charged water-soluble cellular metabolites. Through the use of neutral organic solvents to inactivate endogenous enzyme activities (i.e., methanol/chloroform/H2O extraction), in conjunction with a matrix having minimal background noise (9-amnioacridine), a set of multiplexed conditions was developed that allowed identification of 285 peaks corresponding to negatively charged metabolites from mouse heart extracts. Identification of metabolite peaks was based on mass accuracy and was confirmed by tandem mass spectrometry for 90 of the identified metabolite peaks. Through multiplexing ionization conditions, new suites of metabolites could be ionized and “spectrometric isolation” of closely neighboring peaks for subsequent tandem mass spectrometric interrogation could be achieved. Moreover, assignments of ions from isomeric metabolites and quantitation of their relative abundance was achieved in many cases through tandem mass spectrometry by identification of diagnostic fragmentation ions (e.g., discrimination of ATP from dGTP). The high sensitivity of this approach facilitated the detection of extremely low abundance metabolites including important signaling metabolites such as IP3, cAMP, and cGMP. Collectively, these results identify a multiplexed MALDI-TOF/TOF MS approach for analysis of negatively charged metabolites in mammalian tissues. PMID:17665876

  1. Evolution of blood-brain barrier damage associated with changes in brain metabolites following acute ischemia.

    PubMed

    Yan, Gen; Xuan, Yinghua; Dai, Zhuozhi; Zhang, Guishan; Xu, Haiyun; Mikulis, David; Wu, Renhua

    2015-11-11

    Stroke is a serious medical condition that requires emergency care. In the case of ischemic stroke, ischemia may lead to damage to the blood-brain barrier (BBB); the damage in turn may exacerbate the condition. Therefore, noninvasive detection of BBB damage represents a challenge for experimental and clinical researchers. In this study, we assessed the onset of BBB disruption by means of T1-weighted images with administration of the contrast enhancement agent gadolinium-diethylenetriamine pentaacetic acid (Gd-DTPA) and related BBB breakdown to brain metabolite changes in proton magnetic resonance spectrum (H-MRS) in the infarcted site following middle cerebral artery occlusion (MCAO) in rats. It was shown that MCAO for 30 min and 1.5 h caused no Gd-DTPA signal change in the T1-weighted images, whereas MCAO for 1 h significantly altered some of H-MRS brain metabolites, suggesting that brain metabolite changes occurred earlier than BBB damage after ischemic stroke. MCAO for 2 h caused BBB breakdown, which was related to changes in the levels of some brain metabolites detected by H-MRS. Between the second and the third hour after MCAO, brain metabolite changes continued as the result of BBB breakdown and the concurrent overperfusion to the infarcted site, which may ameliorate the metabolite changes, thus compensating for the functional failures of the brain after stroke. PMID:26366833

  2. A new sulphate metabolite as a long-term marker of metandienone misuse.

    PubMed

    Gómez, C; Pozo, O J; Garrostas, L; Segura, J; Ventura, R

    2013-12-11

    Metandienone is one of the most frequently detected anabolic androgenic steroids in sports drug testing. Metandienone misuse is commonly detected by monitoring different metabolites excreted free or conjugated with glucuronic acid using gas chromatography mass spectrometry (GC-MS) and liquid chromatography tandem mass spectrometry (LC-MS/MS) after hydrolysis with β-glucuronidase and liquid-liquid extraction. It is known that several metabolites are the result of the formation of sulphate conjugates in C17, which are converted to their 17-epimers in urine. Therefore, sulphation is an important phase II metabolic pathway of metandienone that has not been comprehensively studied. The aim of this work was to evaluate the sulphate fraction of metandienone metabolism by LC-MS/MS. Seven sulphate metabolites were detected after the analysis of excretion study samples by applying different neutral loss scan, precursor ion scan and SRM methods. One of the metabolites (M1) was identified and characterised by GC-MS/MS and LC-MS/MS as 18-nor-17β-hydroxymethyl-17α-methylandrost-1,4,13-triene-3-one sulphate. M1 could be detected up to 26 days after the administration of a single dose of metandienone (5 mg), thus improving the period in which the misuse can be reported with respect to the last long-term metandienone metabolite described (18-nor-17β-hydroxymethyl-17α-methylandrost-1,4,13-triene-3-one excreted in the glucuronide fraction). PMID:24055830

  3. A guide to the identification of metabolites in NMR-based metabonomics/metabolomics experiments

    PubMed Central

    Dona, Anthony C.; Kyriakides, Michael; Scott, Flora; Shephard, Elizabeth A.; Varshavi, Dorsa; Veselkov, Kirill; Everett, Jeremy R.

    2016-01-01

    Metabonomics/metabolomics is an important science for the understanding of biological systems and the prediction of their behaviour, through the profiling of metabolites. Two technologies are routinely used in order to analyse metabolite profiles in biological fluids: nuclear magnetic resonance (NMR) spectroscopy and mass spectrometry (MS), the latter typically with hyphenation to a chromatography system such as liquid chromatography (LC), in a configuration known as LC–MS. With both NMR and MS-based detection technologies, the identification of the metabolites in the biological sample remains a significant obstacle and bottleneck. This article provides guidance on methods for metabolite identification in biological fluids using NMR spectroscopy, and is illustrated with examples from recent studies on mice. PMID:27087910

  4. Synthesis and identification of hydroxylated metabolites of the anti-estrogenic agent cyclofenil.

    PubMed

    Gärtner, Peter; Hofbauer, Karin; Reichel, Christian; Geisendorfer, Thomas; Gmeiner, Günter

    2008-07-01

    The detection of metabolites of the anti-estrogenic substance cyclofenil, listed on the World Anti-Doping Agency (WADA) Prohibited List since 2004 is described. Target substances are hydroxylated metabolites, bearing an aliphatic hydroxyl group either in the 2-, 3- or 4-position of the aliphatic ring, in addition to the phenolic functions on the aromatic rings. Structural identification used NMR as well as high-resolution mass spectrometry after nano-electrospray ionisation (ESI). Unambiguous detection of all three synthesised cyclofenil metabolites M1-M3 was done using gas chromatography for separation and electron ionisation mass spectrometry for detection of the per-silylated compounds in comparison with a reference urine deriving from an excretion study within the WADA 2007 Educational Programme. PMID:18576435

  5. Highly sensitive simultaneous quantification of estrogenic tamoxifen metabolites and steroid hormones by LC-MS/MS.

    PubMed

    Johänning, Janina; Heinkele, Georg; Precht, Jana C; Brauch, Hiltrud; Eichelbaum, Michel; Schwab, Matthias; Schroth, Werner; Mürdter, Thomas E

    2015-09-01

    Tamoxifen is a mainstay in the treatment of estrogen receptor-positive breast cancer and is metabolized to more than 30 different compounds. Little is known about in vivo concentrations of estrogenic metabolites E-metabolite E, Z-metabolite E, and bisphenol and their relevance for tamoxifen efficacy. Therefore, we developed a highly sensitive HPLC-ESI-MS/MS quantification method for tamoxifen metabolites bisphenol, E-metabolite E, and Z-metabolite E as well as for the sex steroid hormones estradiol, estrone, testosterone, androstenedione, and progesterone. Plasma samples were subjected to protein precipitation followed by solid phase extraction. Upon derivatization with 3-[(N-succinimide-1-yl)oxycarbonyl]-1-methylpyridinium iodide, all analytes were separated on a sub-2-μm column with a gradient of acetonitrile in water with 0.1 % of formic acid. Analytes were detected on a triple-quadrupole mass spectrometer with positive electrospray ionization in the multiple reaction monitoring mode. Our method demonstrated high sensitivity, accuracy, and precision. The lower limits of quantification were 12, 8, and 25 pM for bisphenol, E-metabolite E, and Z-metabolite E, respectively, and 4 pM for estradiol and estrogen, 50 pM for testosterone and androstenedione, and 25 pM for progesterone. The method was applied to plasma samples of postmenopausal patients taken at baseline and under tamoxifen therapy. Graphical Abstract Sample preparation and derivatization for highly sensitive quantification of estrogenic tamoxifen metabolites and steroid hormones by HPLC-MS/MS. PMID:26206706

  6. Temperament Type Specific Metabolite Profiles of the Prefrontal Cortex and Serum in Cattle

    PubMed Central

    Brand, Bodo; Hadlich, Frieder; Brandt, Bettina; Schauer, Nicolas; Graunke, Katharina L.; Langbein, Jan; Repsilber, Dirk; Ponsuksili, Siriluk; Schwerin, Manfred

    2015-01-01

    In the past decade the number of studies investigating temperament in farm animals has increased greatly because temperament has been shown not only to affect handling but also reproduction, health and economically important production traits. However, molecular pathways underlying temperament and molecular pathways linking temperament to production traits, health and reproduction have yet to be studied in full detail. Here we report the results of metabolite profiling of the prefrontal cortex and serum of cattle with distinct temperament types that were performed to further explore their molecular divergence in the response to the slaughter procedure and to identify new targets for further research of cattle temperament. By performing an untargeted comprehensive metabolite profiling, 627 and 1097 metabolite features comprising 235 and 328 metabolites could be detected in the prefrontal cortex and serum, respectively. In total, 54 prefrontal cortex and 51 serum metabolite features were indicated to have a high relevance in the classification of temperament types by a sparse partial least square discriminant analysis. A clear discrimination between fearful/neophobic-alert, interested-stressed, subdued/uninterested-calm and outgoing/neophilic-alert temperament types could be observed based on the abundance of the identified relevant prefrontal cortex and serum metabolites. Metabolites with high relevance in the classification of temperament types revealed that the main differences between temperament types in the response to the slaughter procedure were related to the abundance of glycerophospholipids, fatty acyls and sterol lipids. Differences in the abundance of metabolites related to C21 steroid metabolism and oxidative stress indicated that the differences in the metabolite profiles of the four extreme temperament types could be the result of a temperament type specific regulation of molecular pathways that are known to be involved in the stress and fear response

  7. Widespread occurrence of neuro-active pharmaceuticals and metabolites in 24 Minnesota rivers and wastewaters.

    PubMed

    Writer, Jeffrey H; Ferrer, Imma; Barber, Larry B; Thurman, E Michael

    2013-09-01

    Concentrations of 17 neuro-active pharmaceuticals and their major metabolites (bupropion, hydroxy-bupropion, erythro-hydrobupropion, threo-hydrobupropion, carbamazepine, 10,11,-dihydro-10,11,-dihydroxycarbamazepine, 10-hydroxy-carbamazepine, citalopram, N-desmethyl-citalopram, fluoxetine, norfluoxetine, gabapentin, lamotrigine, 2-N-glucuronide-lamotrigine, oxcarbazepine, venlafaxine and O-desmethyl-venlafaxine), were measured in treated wastewater and receiving surface waters from 24 locations across Minnesota, USA. The analysis of upstream and downstream sampling sites indicated that the wastewater treatment plants were the major source of the neuro-active pharmaceuticals and associated metabolites in surface waters of Minnesota. Concentrations of parent compound and the associated metabolite varied substantially between treatment plants (concentrations±standard deviation of the parent compound relative to its major metabolite) as illustrated by the following examples; bupropion and hydrobupropion 700±1000 ng L(-1), 2100±1700 ng L(-1), carbamazepine and 10-hydroxy-carbamazepine 480±380 ng L(-1), 360±400 ng L(-1), venlafaxine and O-desmethyl-venlafaxine 1400±1300 ng L(-1), 1800±2300 ng L(-1). Metabolites of the neuro-active compounds were commonly found at higher or comparable concentrations to the parent compounds in wastewater effluent and the receiving surface water. Neuro-active pharmaceuticals and associated metabolites were detected only sporadically in samples upstream from the effluent outfall. Metabolite to parent ratios were used to evaluate transformation, and we determined that ratios in wastewater were much lower than those reported in urine, indicating that the metabolites are relatively more labile than the parent compounds in the treatment plants and in receiving waters. The widespread occurrence of neuro-active pharmaceuticals and metabolites in Minnesota effluents and surface waters indicate that this is likely a global environmental issue

  8. Human hydroxylated metabolites of BDE-47 and BDE-99 are glucuronidated and sulfated in vitro.

    PubMed

    Erratico, Claudio; Zheng, Xiaobo; Ryden, Andreas; Marsh, Goran; Maho, Walid; Covaci, Adrian

    2015-07-16

    Polybrominated diphenyl ethers (PBDEs) were used worldwide as additive flame retardants and are classified as persistent, bioaccumulable and toxic environmental pollutants. In humans, the hydroxylated metabolites of 2,2',4,4'-tetrabromodiphenyl ether (BDE-47) and 2,2',4,4',5-pentabromodiphenyl ether (BDE-99) formed in vitro have also been detected in vivo. To further characterize the metabolism of BDE-47 and BDE-99 and to identify candidate markers for monitoring the human exposure to PBDEs using non-invasive approaches, glucuronidation and sulfation of hydroxylated metabolites of BDE-47 and BDE-99 were investigated using human liver microsomes and cytoplasm, respectively. The formed Phase II metabolites were analyzed by liquid chromatography-tandem mass spectrometry using a novel approach to develop analytical methods in absence of authentic standards. All available standards for hydroxylated metabolites of BDE-47 and BDE-99 were glucuronidated and sulfated, showing that glucuronidation and sulfation are part of the metabolism pathway of BDE-47 and BDE-99 in vitro. The major glucuronidated and sulfated analogs of hydroxylated metabolites of BDE-47 were (a) 2,4-DBP-Gluc and 5-Gluc-BDE-47, and (b) 2'-Sulf-BDE-28, 4-Sulf-BDE-42 and 3-Sulf-BDE-47, respectively. The major glucuronidated and sulfated analogs of hydroxylated metabolites of BDE-99 were (a) 2,4,5-TBP-Gluc and 6'-Gluc-BDE-99, and (b) 3'-Sulf-BDE-99 and 5'-Sulf-BDE-99, respectively. Apparent Km values associated with the formation of sulfated metabolites of BDE-47 and BDE-99 were ten times lower than those of the corresponding glucuronidated metabolites, suggesting that sulfated rather than glucuronidated metabolites of OH-PBDEs might be used as markers of human exposure to PBDEs using a non-invasive approach based on urine sample collection. PMID:25956475

  9. Identification of novel hydrazine metabolites by 15N-NMR.

    PubMed

    Preece, N E; Nicholson, J K; Timbrell, J A

    1991-05-01

    15N-NMR has been used to study the metabolism of hydrazine in rats in vivo. Single doses of [15N2]hydrazine (2.0 mmol/kg: 98.6% g atom) were administered to rats and urine collected for 24 hr over ice. A number of metabolites were detected by 15N-NMR analysis of lyophilized urine. Ammonia was detected as a singlet at 0 ppm and unchanged [15N2]hydrazine was present in the urine detectable as a singlet at 32 ppm. Peaks were observed at 107 and 110 ppm which were identified as being due to the hydrazido nitrogen of acetylhydrazine and diacetylhydrazine, respectively. A resonance at 85 ppm was ascribed to carbazic acid, resulting from reaction of hydrazine with carbon dioxide. A singlet detected at 316 ppm was thought to be due to the hydrazono nitrogen of the pyruvate hydrazone. The resonance at 56 ppm was assigned to 15N-enriched urea, this together with the presence of ammonia indicates that the N-N bond of hydrazine is cleaved in vivo, possibly by N-oxidation, and the resultant ammonia is incorporated into urea. A doublet centred at 150 ppm and a singlet at 294 ppm were assigned to a metabolite which results from cyclization of the 2-oxoglutarate hydrazone. Therefore 15N-NMR spectroscopic analysis of urine has yielded significant new information on the metabolism of hydrazine. PMID:2018564

  10. NMR metabolomics profiling of blood plasma mimics shows that medium- and long-chain fatty acids differently release metabolites from human serum albumin

    NASA Astrophysics Data System (ADS)

    Jupin, M.; Michiels, P. J.; Girard, F. C.; Spraul, M.; Wijmenga, S. S.

    2014-02-01

    Metabolite profiling by NMR of body fluids is increasingly used to successfully differentiate patients from healthy individuals. Metabolites and their concentrations are direct reporters of body biochemistry. However, in blood plasma the NMR-detected free-metabolite concentrations are also strongly affected by interactions with the abundant plasma proteins, which have as of yet not been considered much in metabolic profiling. We previously reported that many of the common NMR-detected metabolites in blood plasma bind to human serum albumin (HSA) and many are released by fatty acids present in fatted HSA. HSA is the most abundant plasma protein and main transporter of endogenous and exogenous metabolites. Here, we show by NMR how the two most common fatty acids (FAs) in blood plasma - the long-chain FA, stearate (C18:0) and medium-chain FA, myristate (C14:0) - affect metabolite-HSA interaction. Of the set of 18 common NMR-detected metabolites, many are released by stearate and/or myristate, lactate appearing the most strongly affected. Myristate, but not stearate, reduces HSA-binding of phenylalanine and pyruvate. Citrate signals were NMR invisible in the presence of HSA. Only at high myristate-HSA mole ratios 11:1, is citrate sufficiently released to be detected. Finally, we find that limited dilution of blood-plasma mimics releases HSA-bound metabolites, a finding confirmed in real blood plasma samples. Based on these findings, we provide recommendations for NMR experiments for quantitative metabolite profiling.

  11. 40 CFR 159.179 - Metabolites, degradates, contaminants, and impurities.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... chemical properties of the metabolite or degradate. (B) Data regarding structurally analogous chemicals. (C) Data regarding chemical reactivity of the metabolite or degradate and structurally analogous...

  12. 40 CFR 159.179 - Metabolites, degradates, contaminants, and impurities.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... chemical properties of the metabolite or degradate. (B) Data regarding structurally analogous chemicals. (C) Data regarding chemical reactivity of the metabolite or degradate and structurally analogous...

  13. 40 CFR 159.179 - Metabolites, degradates, contaminants, and impurities.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... chemical properties of the metabolite or degradate. (B) Data regarding structurally analogous chemicals. (C) Data regarding chemical reactivity of the metabolite or degradate and structurally analogous...

  14. Profiling a gut microbiota-generated catechin metabolite's fate in human blood cells using a metabolomic approach.

    PubMed

    Mülek, Melanie; Fekete, Agnes; Wiest, Johannes; Holzgrabe, Ulrike; Mueller, Martin J; Högger, Petra

    2015-10-10

    The microbial catechin metabolite δ-(3,4-dihydroxy-phenyl)-γ-valerolactone (M1) has been found in human plasma samples after intake of maritime pine bark extract (Pycnogenol). M1 has been previously shown to accumulate in endothelial and blood cells in vitro after facilitated uptake and to exhibit anti-inflammatory activity. The purpose of the present research approach was to systematically and comprehensively analyze the metabolism of M1 in human blood cells in vitro and in vivo. A metabolomic approach that had been successfully applied for drug metabolite profiling was chosen to detect 19 metabolite peaks of M1 which were subsequently further analyzed and validated. The metabolites were categorized into three levels of identification according to the Metabolomics Standards Initiative with six compounds each confirmed at levels 1 and 2 and seven putative metabolites at level 3. The predominant metabolites were glutathione conjugates which were rapidly formed and revealed prolonged presence within the cells. Although a formation of an intracellular conjugate of M1 and glutathione (M1-GSH) was already known two GSH conjugate isomers, M1-S-GSH and M1-N-GSH were observed in the current study. Additionally detected organosulfur metabolites were conjugates with oxidized glutathione and cysteine. Other biotransformation products constituted the open-chained ester form of M1 and a methylated M1. Six of the metabolites determined in in vitro assays were also detected in blood cells in vivo after ingestion of the pine bark extract by two volunteers. The present study provides the first evidence that multiple and structurally heterogeneous polyphenol metabolites can be generated in human blood cells. The bioactivity of the M1 metabolites and their contribution to the previously determined anti-inflammatory effects of M1 now need to be elucidated. PMID:26025814

  15. Application of mass spectrometry for metabolite identification.

    PubMed

    Ma, Shuguang; Chowdhury, Swapan K; Alton, Kevin B

    2006-06-01

    Metabolism studies play a pivotal role in drug discovery and development. Characterization of metabolic "hot-spots" as well as reactive and pharmacologically active metabolites is critical to designing new drug candidates with improved metabolic stability, toxicological profile and efficacy. Metabolite identification in the preclinical species used for safety evaluation is required in order to determine whether human metabolites have been adequately tested during non-clinical safety assessment. From an instrumental standpoint, high performance liquid chromatography (HPLC) coupled with mass spectrometry (MS) dominates all analytical tools used for metabolite identification. The general strategies employed for metabolite identification in both drug discovery and drug development settings together with sample preparation techniques are reviewed herein. These include a discussion of the various ionization methods, mass analyzers, and tandem mass spectrometry (MS/MS) techniques that are used for structural characterization in a modern drug metabolism laboratory. Mass spectrometry-based techniques, such as stable isotope labeling, on-line H/D exchange, accurate mass measurement to enhance metabolite identification and recent improvements in data acquisition and processing for accelerating metabolite identification are also described. Rounding out this review, we offer additional thoughts about the potential of alternative and less frequently used techniques such as LC-NMR/MS, CRIMS and ICPMS. PMID:16787159

  16. Absorption properties of micellar lipid metabolites into Caco2 cells.

    PubMed

    Tsuzuki, Wakako

    2007-07-01

    To elucidate the absorption characteristics of dietary lipids in the human intestine, we investigated the cellular uptake of lipid metabolites using a differential monolayer of the Caco2 cells. As lipid metabolites, several free fatty acids and 2-monoacylglycerols, were formed a mixed micelle by bile salts and lysophospholipids and they were supplied to the Caco2 cells. To estimate the effect of the mixed micelles on the permeability of cells' membranes during incubation with the mixed micelles, the transepitherial electrical resistance (TEER) value was monitored, and no pronounced changes of TEER was detected. This suggested that mixed micelles did not affect their cellular properties of the barrier measured by TEER. The lipid metabolites transferred from the mixed micelle into the Caco2 cells were determined quantitatively by an enzymatic colorimetric method and were done by thin layer chromatography (TLC) for a species of acylglycerols. These highly sensitive methods enabled us to monitor the transepithelial transports of various kinds of non-isotope-labeled various lipid metabolites. Newly re-synthesized triacylglycerols were accumulated in Caco2 cells after 30 min incubation with the mixed micelles, and their amounts increased gradually for 4 h. The secretion of re-esterified triacylglycerols into a basolateral medium from the Caco2 cells began at 2 h after the mixed micelles were added to the apical medium. The intake of external lipid metabolites by the Caco2 cells were evaluated by an initial 2-h incubation with the mixed micelles. For example, 2-monomyristin and 2-monopalmitin were more rapidly transferred into the Caco2 cells from the mixed micelles than 2-monocaprin was. On the other hand, the absorption rates of capric acid, lauric acid and myristic acid by the cells were larger than those of stearic acid and oleic acid. It revealed that the side-chain structure of these lipid metabolites affected their absorption by the Caco2 cells. The results of this

  17. Metabolism and metabolites of polychlorinated biphenyls (PCBs)

    PubMed Central

    Grimm, FA; Hu, D; Kania-Korwel, I; Lehmler, HJ; Ludewig, G; Hornbuckle, KC; Duffel, MW; Bergman, A; Robertson, LW

    2015-01-01

    The metabolism of polychlorinated biphenyls (PCBs) is complex and has an impact on toxicity and thereby assessment of PCB risks. A large number of reactive and stable metabolites are formed in the processes of biotransformation in biota in general and in humans in particular. The aim of this document is to provide an overview of PCB metabolism and to identify metabolites of concern and their occurrence. Emphasis is given to mammalian metabolism of PCBs and their hydroxyl, methylsulfonyl, and sulfated metabolites, especially those that persist in human blood. Potential intracellular targets and health risks are also discussed. PMID:25629923

  18. Pharmacokinetics of rectal diclofenac and its hydroxy metabolites in man.

    PubMed

    Landsdorp, D; Vree, T B; Janssen, T J; Guelen, P J

    1990-07-01

    Diclofenac 100 mg suppository is well absorbed from the gastrointestinal tract of six human volunteers. Plasma concentrations of 4'-hydroxydiclofenac and diclofenac could be measured; the concentrations of the three other metabolites were below the detection limit of the HPLC analysis. The apparent half-lives of diclofenac and 4'-OH-diclofenac are, respectively 1.3 +/- 0.3 h and 4.3 +/- 1.0 h. When the t 1/2 values are derived from the renal excretion rate-time profiles, they are as follows: diclofenac 1.8 +/- 0.9 h, 3'-OH-, and 5-OH-diclofenac 2.3 +/- 1.0 h and 2.5 +/- 0.4 h, respectively, while those of 4'-OH- and 4',5-diOH-diclofenac are, respectively 3.6 +/- 0.5 h and 3.1 +/- 1.3 h. Diclofenac is excreted for 13.6 +/- 6.5%, its renal clearance (Clr) = 3.23 +/- 1.03 ml/min. The main metabolite excreted in the urine is 4'-OH-diclofenac (27.2 +/- 12% dose; Clr = 6.14 +/- 4.04 ml/min). The total body clearance of the parent drug and the apparent total body clearance of the main metabolite are similar, 28.0 +/- 11.9 l/h and 27.5 +/- 10.9 l/h. PMID:2387653

  19. Behavior of alkylphenol polyethoxylate metabolites during soil aquifer treatment.

    PubMed

    Montgomery-Brown, John; Drewes, Jörg E; Fox, Peter; Reinhard, Martin

    2003-09-01

    The attenuation of alkylphenol polyethoxylates (APEOs) metabolites was studied at a soil aquifer treatment (SAT) site located in Arizona, USA. Two parcels of water were monitored during infiltration; one parcel was predominantly oxic while the other was predominantly anoxic. In this study, only alkylphenol ethoxycarboxylates (APECs) and carboxyalkylphenol ethoxycarboxylates (CAPECs) were detected, no short-chained APEOs were observed-even under anoxic conditions. APEO metabolites were rapidly (<7 days) removed under both aerobic and anoxic conditions. In general, the length of the ethoxycarboxylate chain decreases with depth--at depths greater than 3m, only alkylphenoxy acetic acids (AP1ECs), carboxyalkylphenoxy acetic acids (CAP1ECs), and alkylphenols (APs) remain. Under aerobic conditions, octylphenol and nonylphenol concentrations decreased by approximately 80% (w/w) within 3m of the ground surface. Under anoxic conditions however, alkylphenol concentrations increased by approximately 200% during the first 1.5m and then decreased during the next 1.5m; overall, under anoxic conditions, alkylphenol concentrations increased by approximately 38% within 3m. During infiltration, APEC and CAPEC concentrations decrease by more than 95% within 3m of SAT. Alternate flooding and drying cycles appear to enhance overall APEO metabolite removal efficiencies. PMID:12867334

  20. Secondary metabolites from Rubiaceae species.

    PubMed

    Martins, Daiane; Nunez, Cecilia Veronica

    2015-01-01

    This study describes some characteristics of the Rubiaceae family pertaining to the occurrence and distribution of secondary metabolites in the main genera of this family. It reports the review of phytochemical studies addressing all species of Rubiaceae, published between 1990 and 2014. Iridoids, anthraquinones, triterpenes, indole alkaloids as well as other varying alkaloid subclasses, have shown to be the most common. These compounds have been mostly isolated from the genera Uncaria, Psychotria, Hedyotis, Ophiorrhiza and Morinda. The occurrence and distribution of iridoids, alkaloids and anthraquinones point out their chemotaxonomic correlation among tribes and subfamilies. From an evolutionary point of view, Rubioideae is the most ancient subfamily, followed by Ixoroideae and finally Cinchonoideae. The chemical biosynthetic pathway, which is not so specific in Rubioideae, can explain this and large amounts of both iridoids and indole alkaloids are produced. In Ixoroideae, the most active biosysthetic pathway is the one that produces iridoids; while in Cinchonoideae, it produces indole alkaloids together with other alkaloids. The chemical biosynthetic pathway now supports this botanical conclusion. PMID:26205062

  1. The profiling and identification of the metabolites of (+)-catechin and study on their distribution in rats by HPLC-DAD-ESI-IT-TOF-MS(n) technique.

    PubMed

    Liang, Jing; Xu, Feng; Zhang, Ya-Zhou; Zang, Xin-Yu; Wang, Dan; Shang, Ming-Ying; Wang, Xuan; Chui, De-Hua; Cai, Shao-Qing

    2014-03-01

    (+)-Catechin, a potential beneficial compound to human health, is widely distributed in plants and foods. A high-performance liquid chromatography with diode array detector and combined with electrospray ionization ion trap time-of-flight multistage mass spectrometry method was applied to profile and identify the metabolites of (+)-catechin in rats and to study the distribution of these metabolites in rat organs for the first time. In total, 51 phase II metabolites (44 new) and three phase I metabolites were tentatively identified, comprising 16 (+)-catechin conjugates, 14 diarylpropan-2-ol metabolites, 6 phenyl valerolactone metabolites and 18 aromatic acid metabolites. Further, 19 phase II metabolites were new compounds. The in vivo metabolic reactions of (+)-catechin in rats were found to be ring-cleavage, sulfation, glucuronidation, methylation, dehydroxylation and dehydrogenation. The numbers of detected metabolites in urine, plasma, small intestine, kidney, liver, lung, heart, brain and spleen were 53, 23, 27, 9, 7, 5, 3, 2 and 1, respectively. This indicated that small intestine, kidney and liver were the major organs for the distribution of (+)-catechin metabolites. In addition, eight metabolites were found to possess bioactivities according to literature. These results are very helpful for better comprehension of the in vivo metabolism of (+)-catechin and its pharmacological actions, and also can give strong indications on the effective forms of (+)-catechin in vivo. PMID:24105958

  2. Two-dimensional liquid chromatography/mass spectrometry/mass spectrometry separation of water-soluble metabolites.

    PubMed

    Fairchild, Jacob N; Horvath, Krisztian; Gooding, Jessica R; Campagna, Shawn R; Guiochon, Georges

    2010-12-24

    Off-line two-dimensional liquid chromatography with tandem mass spectrometry detection (2D-LC/MS-MS) was used to separate a set of metabolomic species. Water-soluble metabolites were extracted from Escherichia coli and Saccharomyces cerevisae cultures and were immediately analyzed using strong cation exchange (SCX)-hydrophilic interaction chromatography (HILIC). Metabolite mixtures are well-suited for multidimensional chromatography as the range of components varies widely with respect to polarity and chemical makeup. Some currently used methods employ two different separations for the detection of positively and negatively ionized metabolites by mass spectrometry. Here we developed a single set of chromatographic conditions for both ionization modes and were able to detect a total of 141 extracted metabolite species, with an overall peak capacity of ca. 2500. We show that a single two-dimensional separation method is sufficient and practical when a pair or more of unidimensional separations are used in metabolomics. PMID:21094946

  3. Concentrations of dimethylaniline and other metabolites in milk and tissues of dairy cows treated with lidocaine.

    PubMed

    Hoogenboom, Ron L A P; Zuidema, Tina; Essers, Martien; van Vuuren, Ad M; van Wikselaar, Piet G; van Eijkeren, Jan C H; Mengelers, Marcel J B; Zeilmaker, Marco J; Bulder, Astrid S

    2015-01-01

    Lidocaine is a topical anaesthetic drug used in dairy cows for laparotomy (caesarean section, abomasal displacement). Because there are no registered drugs for this indication, it can be applied under the so-called Cascade rules (off-label use), with the restriction that the off-label withdrawal periods of 7 days for milk and 28 days for meat are taken into account. In animals, lidocaine is rapidly metabolised into various metabolites, one being 2,6-dimethylaniline (DMA) which is reported to possess carcinogenic and mutagenic properties and detected also in milk. To investigate whether the off-label withdrawal periods are long enough to exclude the presence of lidocaine and DMA, and potential other metabolites, in edible products, a study was performed with eight dairy cows treated with lidocaine by injection in the abdominal muscles. At various time points blood samples, milk and urine were collected. Four animals were slaughtered 3.5 h after treatment, the other four after 48.5 h. The injection site, meat, liver and kidney were analysed for levels of lidocaine, DMA, monoethylglycinexylidide (MEGX) and 3-OH-lidocaine. It was shown that DMA is an important metabolite in dairy cows and can be detected in both meat and milk. In addition, also MEGX, 3-OH-lidocaine and three other metabolites were identified and to some extent quantified. These metabolites were 4-OH-lidocaine, lidocaine-N-oxide and 4-hydroxy-DMA. The latter compound was the most important metabolite in urine. However, levels in milk and meat decreased rapidly after the application. Overall, it can be concluded that the off-label withdrawal times of 7 and 28 days for milk and meat, respectively, guarantee the absence of detectable levels of lidocaine and metabolites. PMID:26118443

  4. Metabolites of synthetic cannabinoids in hair-proof of consumption or false friends for interpretation?

    PubMed

    Franz, Florian; Angerer, Verena; Hermanns-Clausen, Maren; Auwärter, Volker; Moosmann, Bjoern

    2016-05-01

    The detection of drug metabolites in hair is widely accepted as a proof for systemic uptake of the drug, unless the metabolites can be formed as artefacts. However, regarding synthetic cannabinoids, not much is known about mechanisms of incorporation into hair. For a correct interpretation concerning hair findings of these compounds and their metabolites, it is necessary to identify the different routes of incorporation and to assess their contribution to analytical findings. This study presents the results of the LC-ESI-MS/MS analysis of an authentic hair sample taken from a patient with a known history of heavy consumption of synthetic cannabinoids. In the authentic hair sample, 5F-PB-22 and AB-CHMINACA as well as their main metabolites 5F-PB-22 3-carboxyindole, PB-22 5-OH-pentyl, and AB-CHMINACA valine were detected in all segments, comprising segments grown in a time period where the substances had not been distributed on the 'legal high' market. To enable interpretation of the results regarding the distribution of the detected analytes along the hair shaft, the stability of 5F-PB-22 and AB-CHMINACA in hair matrix and under thermal stress was assessed. The stability tests revealed that the three 'metabolites' are also formed in externally contaminated hair after storage of the samples under different conditions. In addition, 5F-PB-22 3-carboxyindole and AB-CHMINACA valine were identified as degradation products in smoke condensate. Therefore, interpretation of 'metabolite' findings of compounds comprising chemically labile amide/ester bonds or 5-fluoro-pentyl side chains should be carried out with utmost care, taking into account the different mechanisms of formation and incorporation into hair. Graphical Abstract Degradation processes leading to artefacts identical with main metabolites of synthetic cannabinoids. PMID:26935933

  5. A targeted proteomics approach to the identification of peptides modified by reactive metabolites.

    PubMed

    Tzouros, Manuel; Pähler, Axel

    2009-05-01

    Covalent binding of reactive metabolites is generally accepted as one underlying mechanism of drug-induced toxicity. However, identification of protein targets by reactive metabolites still remains a challenge due to their low abundance. Here, we report the development of a highly selective proteomics workflow for the targeted identification of peptides modified by reactive metabolites. An equimolar mixture of non- and radiolabeled furan containing 2-amino-pyrimidine drug candidate (1 and 14C(1)-1) along with rat liver microsomes were used for the in vitro generation of reactive metabolites. Liver microsomal proteins were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, modified protein bands were highlighted by autoradiography and in-gel digested, and peptides were fractionated by strong cation exchange chromatography. Fractions enriched in modified peptides, as determined by radioactivity levels, were subjected to nanoLC-MS/MS and unambiguously detected based on their unique 12C/14C MS isotope pattern fingerprint. The peptide detection step could be automated using isotope pattern recognition software. Peptide sequencing, identification of the site of modification, and reactive metabolite characterization were achieved by MS2 and MS3 experiments using high-resolution and accurate mass detection. This approach led to the identification of four modified peptides originating from three drug-metabolizing enzymes, MGST1, FMO1, and P450 2C11. These revealed modifications by five different metabolite structures. This approach is generally suitable for the identification and characterization of modified proteins and metabolite structures involved in covalent binding and may serve as a valuable tool to link protein targets with clinically relevant toxicities. PMID:19317514

  6. Identification and quantification of metabolites of AB-CHMINACA in a urine specimen of an abuser.

    PubMed

    Wurita, Amin; Hasegawa, Koutaro; Minakata, Kayoko; Gonmori, Kunio; Nozawa, Hideki; Yamagishi, Itaru; Suzuki, Osamu; Watanabe, Kanako

    2016-03-01

    We experienced an autopsy case in which the cause of death was judged as poisoning by multiple new psychoactive substances, including AB-CHMINACA, 5-fluoro-AMB and diphenidine [Forensic Toxicol. 33 (2015): 45-53]. Although unchanged AB-CHMINACA could be detected from 8 solid tissues, it could neither be detected from blood nor urine specimens. In this article, we obtained eight kinds of reference standards of AB-CHMINACA metabolites from a commercial source. The AB-CHMINACA metabolites from the urine specimen of the abuser were extracted by a modified QuEChERS method and analyzed by liquid chromatography-tandem mass spectrometry before and after hydrolysis with β-glucuronidase. Among the eight AB-CHMINACA metabolites tested, only 2 metabolites could be identified in the urine specimen of the deceased. After hydrolysis with β-glucuronidase, the concentrations of the two metabolites were not increased, suggesting that the metabolites were not in the conjugated forms. The metabolites detected were 4-hydroxycyclohexylmethyl AB-CHMINACA (M1), followed by N-[[1-(cyclohexylmethyl)-1H-indazol-3-yl]carbonyl]-l-valine (M3). Their concentrations were 52.8±3.44 and 41.3±5.04ng/ml (n=10) for M1 and M3, respectively. Although there is one preceding report showing the estimations of metabolism of AB-CHMINACA without reference standards, this is the first report dealing with exact identification using reference standards, and quantification of M1 and M3 in an authentic urine specimen. PMID:26257317

  7. Disposition of xenobiotic chemicals and metabolites in marine organisms

    SciTech Connect

    Varanasi, U.; Stein, J.E. )

    1991-01-01

    Studies with several bottom fish species from urban waterways show that of the identified xenobiotic chemicals in bottom sediments, polycyclic aromatic hydrocarbons (PAHs) are the most strongly associated with the prevalence of liver lesions, including neoplasms. Accordingly, there is concern about the transfer of contaminants, such as PAHs, from aquatic species to humans. Because PAHs exert their toxicity only after being biotransformed, increasing attention has been focused on the ability of aquatic organisms to metabolize these chemicals. Overall, the results of both laboratory and field studies show that generally low levels of a few low molecular weight PAHs may be present in edible tissue of fish from contaminated areas and that high molecular weight PAHs, such as the carcinogen benzo(a)pyrene, will rarely be detected because of extensive metabolism. Additionally, the results from a few studies suggest that even though interactions between xenobiotics can affect both biochemical and physiological systems to alter the disposition of PAHs in fish, these interactions do not markedly change the relative proportions of metabolites to parent PAH in tissues. Thus, these studies clearly demonstrate that to obtain some insight into the questions of whether there is any risk to human health from consuming fish and crustaceans from urban areas, techniques must be developed that measure metabolites of carcinogens, such as PAHs, in edible tissue. Initial attempts may focus on semiquantitative methods that permit rapid assessment of the level of metabolites in edible tissues of fish and crustaceans from many urban areas. Based on information from such screening studies, further refinement in methodology leading to identification of specific compounds may be needed because certain metabolites may not be as toxic or carcinogenic as others.

  8. Distribution of terfenadine and its metabolites in locusts studied by desorption electrospray ionization mass spectrometry imaging.

    PubMed

    Olsen, Line Rørbæk; Hansen, Steen Honoré; Janfelt, Christian

    2015-03-01

    Desorption electrospray ionization (DESI) mass spectrometry (MS) imaging was used to image locusts dosed with the antihistamine drug terfenadine. The study was conducted in order to elucidate a relatively high elimination rate of terfenadine from the locust hemolymph. In this one of the few MS imaging studies on insects, a method for cryosectioning of whole locusts was developed, and the distributions of a number of endogenous compounds are reported, including betaine and a number of amino acids and phospholipids. Terfenadine was detected in the stomach region and the intestine walls, whereas three different metabolites-terfenadine acid (fexofenadine), terfenadine glucoside, and terfenadine phosphate-were detected in significantly smaller amounts and only in the unexcreted feces in the lower part of the intestine. The use of MS/MS imaging was necessary in order to detect the metabolites. With use of DESI-MS imaging, no colocalization of the drug and the metabolites was observed, suggesting a very rapid excretion of metabolites into the feces. Additional liquid chromatography-MS investigations were performed on hemolymph and feces and showed some abundance of terfenadine and the three metabolites, although at low levels, in both the hemolymph and the feces. PMID:25404166

  9. Coping with shrub secondary metabolites by ruminants

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Rangelands throughout the world contain varying but often substantial proportions of shrubs. Shrubs are generally heavily chemically defended, and herbivores must either contend with their plant secondary metabolites (PSM) or avoid a significant component of the available forage. Browsing ruminants ...

  10. The Significance of Lichens and Their Metabolites

    NASA Astrophysics Data System (ADS)

    Huneck, S.

    Lichens, symbiontic organisms of fungi and algae, synthesize numerous metabolites, the "lichen substances," which comprise aliphatic, cycloaliphatic, aromatic, and terpenic compounds. Lichens and their metabolites have a manifold biological activity: antiviral, antibiotic, antitumor, allergenic, plant growth inhibitory, antiherbivore, and enzyme inhibitory. Usnic acid, a very active lichen substance is used in pharmaceutical preparations. Large amounts of Pseudevernia furfuracea and Evernia prunastri are processed in the perfume industry, and some lichens are sensitive reagents for the evaluation of air pollution.

  11. Cellular toxicity of nicotinamide metabolites.

    PubMed

    Rutkowski, Bolesław; Rutkowski, Przemysław; Słomińska, Ewa; Smolenski, Ryszard T; Swierczyński, Julian

    2012-01-01

    There are almost 100 different substances called uremic toxins. Nicotinamide derivatives are known as new family of uremic toxins. These uremic compounds play a role in an increased oxidative stress and disturbances in cellular repair processes by inhibiting poly (ADP-ribose) polymerase activity. New members of this family were discovered and described. Their toxic properties were a subject of recent studies. This study evaluated the concentration of 4-pyridone-3-carboxamid-1-β-ribonucleoside-triphosphate (4PYTP) and 4-pyridone-3-carboxamid-1-β-ribonucleoside-monophosphate (4PYMP) in erythrocytes of patients with chronic renal failure. Serum and red blood cells were collected from chronic renal failure patients on conservative treatment, those treated with hemodialysis, and at different times from those who underwent kidney transplantation. Healthy volunteers served as a control group. Nicotinamide metabolites were determined using liquid chromatography with mass spectrometry based on originally discovered and described method. Three novel compounds were described: 4-pyridone-3-carboxamid-1-β-ribonucleoside (4PYR), 4PYMP, and 4PYTP. 4PYR concentration was elevated in the serum, whereas 4PYMP and 4PYTP concentrations were augmented in erythrocytes of dialysis patients. Interestingly, concentrations of these compounds were less elevated during the treatment with erythropoietin-stimulating agents (ESAs). After successful kidney transplantation, concentrations of 4PYR and 4PYMP normalized according to the graft function, whereas that of 4PYTP was still elevated. During the incubation of erythrocytes in the presence of 4PYR, concentration of 4PYMP rose very rapidly while that of 4PYTP increased slowly. Therefore, we hypothesized that 4PYR, as a toxic compound, was actively absorbed by erythrocytes and metabolized to the 4PYMP and 4PYTP, which may interfere with function and life span of these cells. PMID:22200423

  12. Metabolism of methyleugenol in liver microsomes and primary hepatocytes: pattern of metabolites, cytotoxicity, and DNA-adduct formation.

    PubMed

    Cartus, Alexander T; Herrmann, Kristin; Weishaupt, Lucas W; Merz, Karl-Heinz; Engst, Wolfram; Glatt, Hansruedi; Schrenk, Dieter

    2012-09-01

    Methyleugenol (1) is a constituent of many foods, in particular of herbal spices, and is used as flavoring agent in foodstuffs and as fragrance in cosmetics. 1 has been found to be carcinogenic in rodents, its metabolite, 1-hydroxymethyleugenol (2) acting as proximate DNA-binding carcinogen. We incubated 1 with liver microsomes of rat, bovine, and human origin. We found 2, 3-hydroxymethylisoeugenol (3), and 6-hydroxymethyleugenol (4) as major metabolites, and 1-oxomethyleugenol (5), 3-oxomethylisoeugenol (6), eugenol (9), chavibetol (11), and (RS)-2,3-dihydroxy-2,3-dihydromethyleugenol (7) as minor metabolites. Methyleugenol-2,3-epoxide (8), probably the precursor of 7, could not be detected. Incubations with synthetic metabolites were applied in order to uncover metabolic pathways. Incubations with primary rat hepatocytes revealed mainly nonconjugated 2 and conjugated 4, and minor amounts of partly conjugated 7 and conjugated 9 + 11. The "reactive metabolites" 3, 5, 6, and 8 were not detectable, possibly due to rapid reaction with cellular macromolecules. The highest cytotoxicity (resazurin reduction assay and lactate dehydrogenase leakage assay) was observed for the main metabolite 2 and its secondary metabolite 5 with EC(50) values of 50 and 10 µM, respectively. Deoxyadenosine or deoxyguanosine adducts were formed by incubating 1 or metabolites with rat hepatocytes. The rank order of adduct formation was 2 > 1 > 3 > 6, whereas 4, 5, and 8 were inactive. In conclusion, we present a virtually complete pattern of microsomal (rat, bovine, and human) and hepatocellular (rat) metabolites of 1 suggesting the formation of several reactive metabolites possibly involved in carcinogenicity, organ toxicity, and immune reactions. PMID:22610610

  13. Metabolites of cannabidiol identified in human urine.

    PubMed

    Harvey, D J; Mechoulam, R

    1990-03-01

    1. Urine from a dystonic patient treated with cannabidiol (CBD) was examined by g.l.c.-mass spectrometry for CBD metabolites. Metabolites were identified as their trimethylsilyl (TMS), [2H9]TMS, and methyl ester/TMS derivatives and as the TMS derivatives of the product of lithium aluminium deuteride reduction. 2. Thirty-three metabolites were identified in addition to unmetabolized CBD, and a further four metabolites were partially characterized. 3. The major metabolic route was hydroxylation and oxidation at C-7 followed by further hydroxylation in the pentyl and propenyl groups to give 1"-, 2"-, 3"-, 4"- and 10-hydroxy derivatives of CBD-7-oic acid. Other metabolites, mainly acids, were formed by beta-oxidation and related biotransformations from the pentyl side-chain and these were also hydroxylated at C-6 or C-7. The major oxidized metabolite was CBD-7-oic acid containing a hydroxyethyl side-chain. 4. Two 8,9-dihydroxy compounds, presumably derived from the corresponding epoxide were identified. 5. Also present were several cyclized cannabinoids including delta-6- and delta-1-tetrahydrocannabinol and cannabinol. 6. This is the first metabolic study of CBD in humans; most observed metabolic routes were typical of those found for CBD and related cannabinoids in other species. PMID:2336840

  14. Integrating Multiple Analytical Datasets to Compare Metabolite Profiles of Mouse Colonic-Cecal Contents and Feces

    PubMed Central

    Zeng, Huawei; Grapov, Dmitry; Jackson, Matthew I.; Fahrmann, Johannes; Fiehn, Oliver; Combs, Gerald F.

    2015-01-01

    The pattern of metabolites produced by the gut microbiome comprises a phenotype indicative of the means by which that microbiome affects the gut. We characterized that phenotype in mice by conducting metabolomic analyses of the colonic-cecal contents, comparing that to the metabolite patterns of feces in order to determine the suitability of fecal specimens as proxies for assessing the metabolic impact of the gut microbiome. We detected a total of 270 low molecular weight metabolites in colonic-cecal contents and feces by gas chromatograph, time-of-flight mass spectrometry (GC-TOF) and ultra-high performance liquid chromatography, quadrapole time-of-flight mass spectrometry (UPLC-Q-TOF). Of that number, 251 (93%) were present in both types of specimen, representing almost all known biochemical pathways related to the amino acid, carbohydrate, energy, lipid, membrane transport, nucleotide, genetic information processing, and cancer-related metabolism. A total of 115 metabolites differed significantly in relative abundance between both colonic-cecal contents and feces. These data comprise the first characterization of relationships among metabolites present in the colonic-cecal contents and feces in a healthy mouse model, and shows that feces can be a useful proxy for assessing the pattern of metabolites to which the colonic mucosum is exposed. PMID:26378591

  15. Bacterial dynamics and metabolite changes in solid-state acetic acid fermentation of Shanxi aged vinegar.

    PubMed

    Li, Sha; Li, Pan; Liu, Xiong; Luo, Lixin; Lin, Weifeng

    2016-05-01

    Solid-state acetic acid fermentation (AAF), a natural or semi-controlled fermentation process driven by reproducible microbial communities, is an important technique to produce traditional Chinese cereal vinegars. Highly complex microbial communities and metabolites are involved in traditional Chinese solid-state AAF, but the association between microbiota and metabolites during this process are still poorly understood. In this study, we performed amplicon 16S rRNA gene sequencing on the Illumina MiSeq platform, PCR-denaturing gradient gel electrophoresis, and metabolite analysis to trace the bacterial dynamics and metabolite changes under AAF process. A succession of bacterial assemblages was observed during the AAF process. Lactobacillales dominated all the stages. However, Acetobacter species in Rhodospirillales were considerably accelerated during AAF until the end of fermentation. Quantitative PCR results indicated that the biomass of total bacteria showed a "system microbe self-domestication" process in the first 3 days, and then peaked at the seventh day before gradually decreasing until the end of AAF. Moreover, a total of 88 metabolites, including 8 organic acids, 16 free amino acids, and 66 aroma compounds were detected during AAF. Principal component analysis and cluster analyses revealed the high correlation between the dynamics of bacterial community and metabolites. PMID:26754813

  16. Piceatannol and its metabolite, isorhapontigenin, induce SIRT1 expression in THP-1 human monocytic cell line.

    PubMed

    Kawakami, Shinpei; Kinoshita, Yosuke; Maruki-Uchida, Hiroko; Yanae, Koji; Sai, Masahiko; Ito, Tatsuhiko

    2014-11-01

    Piceatannol is a phytochemical that is present in large amounts in passion fruit (Passiflora edulis) seeds, and is an analog of resveratrol. Recently, the absorption and metabolism of piceatannol were investigated in rats, and isorhapontigenin, O-methyl piceatannol, was detected as a piceatannol metabolite in rat plasma. To elucidate the function of piceatannol and its metabolites, we investigated the expression of sirtuin 1 (SIRT1) in THP-1 monocytic cells after treatment with piceatannol and its metabolites, and compared their effects with those of resveratrol and its metabolites. Piceatannol and resveratrol upregulated the expression levels of SIRT1 mRNA and SIRT1 protein. An extract of passion fruit seeds, which contained high levels of piceatannol, also upregulated SIRT1 mRNA expression. As for the metabolites, isorhapontigenin upregulated SIRT1 mRNA expression, whereas resveratrol glucuronides and sulfate did not affect SIRT1 expression. These findings indicate that after intake of piceatannol, not only piceatannol itself, but also its metabolite, isorhapontigenin, contributed to the upregulation of SIRT1 expression. PMID:25360511

  17. Elucidation of Drug Metabolite Structural Isomers Using Molecular Modeling Coupled with Ion Mobility Mass Spectrometry.

    PubMed

    Reading, Eamonn; Munoz-Muriedas, Jordi; Roberts, Andrew D; Dear, Gordon J; Robinson, Carol V; Beaumont, Claire

    2016-02-16

    Ion mobility-mass spectrometry (IM-MS) in combination with molecular modeling offers the potential for small molecule structural isomer identification by measurement of their gas phase collision cross sections (CCSs). Successful application of this approach to drug metabolite identification would facilitate resource reduction, including animal usage, and may benefit other areas of pharmaceutical structural characterization including impurity profiling and degradation chemistry. However, the conformational behavior of drug molecules and their metabolites in the gas phase is poorly understood. Here the gas phase conformational space of drug and drug-like molecules has been investigated as well as the influence of protonation and adduct formation on the conformations of drug metabolite structural isomers. The use of CCSs, measured from IM-MS and molecular modeling information, for the structural identification of drug metabolites has also been critically assessed. Detection of structural isomers of drug metabolites using IM-MS is demonstrated and, in addition, a molecular modeling approach has been developed offering rapid conformational searching and energy assessment of candidate structures which agree with experimental CCSs. Here it is illustrated that isomers must possess markedly dissimilar CCS values for structural differentiation, the existence and extent of CCS differences being ionization state and molecule dependent. The results present that IM-MS and molecular modeling can inform on the identity of drug metabolites and highlight the limitations of this approach in differentiating structural isomers. PMID:26752623

  18. Metabolomics reveals significant variations in metabolites and correlations regarding the maturation of walnuts (Juglans regia L.)

    PubMed Central

    Rao, Guodong; Sui, Jinkai

    2016-01-01

    ABSTRACT The content of walnut metabolites is related to its nutritive value and physiological characteristics, however, comprehensive information concerning the metabolome of walnut kernels is limited. In this study we analyzed the metabolites of walnut kernels at five developmental stages from filling to ripening using GC-MS-based untargeted metabolomics; of a total 252 peaks identified, 85 metabolites were positively identified. Further statistical analysis revealed that these 85 metabolites covered different types of metabolism pathways. PCA scores revealed that the metabolic compositions of the embryo are different at each stage, while the metabolic composition of the endotesta could not be significantly separated into distinct groups. Additionally, 7225 metabolite-metabolite correlations were detected in walnut kernel by a Pearson correlation coefficient approach; during screening of the calculated correlations, 463 and 1047 were determined to be significant with r2≥0.49 and had a false discovery rate (FDR) ≤0.05 in endotesta and embryo, respectively. This work provides the first comprehensive metabolomic study of walnut kernels and reveals that most of the carbohydrate and protein-derived carbon was transferred into other compounds, such as fatty acids, during the maturation of walnuts, which may potentially provide the basis for further studies on walnut kernel metabolism. PMID:27215321

  19. Changes in cerebral metabolites in obstructive sleep apnea: a systemic review and meta-analysis.

    PubMed

    Xia, Yunyan; Fu, Yiqun; Xu, Huajun; Guan, Jian; Yi, Hongliang; Yin, Shankai

    2016-01-01

    Cognitive impairment is associated with changes in cerebral metabolites in patients with obstructive sleep apnea (OSA). Several studies have used magnetic resonance spectroscopy (MRS) to detect variations in cerebral metabolites; however, the results have been inconsistent. This meta-analysis summarizes the differences in cerebral metabolites between patients with OSA and controls. Two electronic databases, PubMed and Embase, were searched for articles (published before March 31, 2016) describing studies that used MRS to evaluate the cerebral metabolite changes. The overall effects were measured using the weighted mean difference with a 95% confidence interval. Subgroup analysis and sensitivity analysis were used to explore the sources of between-study heterogeneity and the stability of the results. Publication bias was also evaluated. Thirteen studies were ultimately included. In the hippocampus, the N-acetylaspartate (NAA)/creatine ratio was lower in patients with OSA. In the frontal lobe, only the NAA/choline ratio was lower in patients with OSA. Cerebral metabolites are significantly altered in the hippocampus in patients with OSA. Further clinical studies are needed to explore the underlying mechanisms between OSA and the changes in cerebral metabolites in the brain. PMID:27349417

  20. Metabolomics reveals significant variations in metabolites and correlations regarding the maturation of walnuts (Juglans regia L.).

    PubMed

    Rao, Guodong; Sui, Jinkai; Zhang, Jianguo

    2016-01-01

    The content of walnut metabolites is related to its nutritive value and physiological characteristics, however, comprehensive information concerning the metabolome of walnut kernels is limited. In this study we analyzed the metabolites of walnut kernels at five developmental stages from filling to ripening using GC-MS-based untargeted metabolomics; of a total 252 peaks identified, 85 metabolites were positively identified. Further statistical analysis revealed that these 85 metabolites covered different types of metabolism pathways. PCA scores revealed that the metabolic compositions of the embryo are different at each stage, while the metabolic composition of the endotesta could not be significantly separated into distinct groups. Additionally, 7225 metabolite-metabolite correlations were detected in walnut kernel by a Pearson correlation coefficient approach; during screening of the calculated correlations, 463 and 1047 were determined to be significant with r(2)≥0.49 and had a false discovery rate (FDR) ≤0.05 in endotesta and embryo, respectively. This work provides the first comprehensive metabolomic study of walnut kernels and reveals that most of the carbohydrate and protein-derived carbon was transferred into other compounds, such as fatty acids, during the maturation of walnuts, which may potentially provide the basis for further studies on walnut kernel metabolism. PMID:27215321

  1. Changes in cerebral metabolites in obstructive sleep apnea: a systemic review and meta-analysis

    PubMed Central

    Xia, Yunyan; Fu, Yiqun; Xu, Huajun; Guan, Jian; Yi, Hongliang; Yin, Shankai

    2016-01-01

    Cognitive impairment is associated with changes in cerebral metabolites in patients with obstructive sleep apnea (OSA). Several studies have used magnetic resonance spectroscopy (MRS) to detect variations in cerebral metabolites; however, the results have been inconsistent. This meta-analysis summarizes the differences in cerebral metabolites between patients with OSA and controls. Two electronic databases, PubMed and Embase, were searched for articles (published before March 31, 2016) describing studies that used MRS to evaluate the cerebral metabolite changes. The overall effects were measured using the weighted mean difference with a 95% confidence interval. Subgroup analysis and sensitivity analysis were used to explore the sources of between-study heterogeneity and the stability of the results. Publication bias was also evaluated. Thirteen studies were ultimately included. In the hippocampus, the N-acetylaspartate (NAA)/creatine ratio was lower in patients with OSA. In the frontal lobe, only the NAA/choline ratio was lower in patients with OSA. Cerebral metabolites are significantly altered in the hippocampus in patients with OSA. Further clinical studies are needed to explore the underlying mechanisms between OSA and the changes in cerebral metabolites in the brain. PMID:27349417

  2. Ardipusilloside-I Metabolites from Human Intestinal Bacteria and Their Antitumor Activity.

    PubMed

    Cao, Wei-Yu; Wang, Ya-Nan; Wang, Peng-Yuan; Lei, Wan; Feng, Bin; Wang, Xiao-Juan

    2015-01-01

    Ardipusilloside-I (ADS-I) is a triterpenoid saponin extracted from Ardisia pusilla DC, and has been demonstrated to have potent antitumor activity. However, ADS-I metabolism in humans has not been investigated. In this study, we studied the biotransformation of ADS-I in human intestinal bacteria, and examined the in vitro antitumor activity of the major metabolites. Ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) was used to detect ADS-I biotransformation products, and their chemical structures were identified by high performance liquid chromatography-nuclear magnetic resonance (HPLC-NMR). The antitumor activity of the major metabolites was determined by the MTT assay. Here, we show that main reaction seen in the metabolism of ADS-I in human intestinal bacteria was deglycosylation, which produced a total of four metabolites. The structures of the two major metabolites M1 and M2 were confirmed by using NMR. MTT assay showed that ADS-I metabolites M1 and M2 have the same levels of inhibitory activities as ADS-I in cultured SMMC-7721 cells and MCF-7 cells. In conclusion, this study demonstrates deglycosylation as a primary pathway of ADS-I metabolism in human intestinal bacteria, and suggests that the pharmacological activity of ADS-I may be mediated, at least in part, by its metabolites. PMID:26610438

  3. Evaluating enzymes that generate genotoxic benzo[a]pyrene metabolites using sensor arrays.

    PubMed

    Wang, Bingquan; Jansson, Ingela; Schenkman, John B; Rusling, James F

    2005-03-01

    Arrays with individually addressable, demountable electrodes coated with ultrathin DNA/enzyme films were evaluated to estimate relative rates of genotoxic bioactivation of benzo[a]pyrene (BP) for several different enzymes simultaneously. Specifically, cytochrome (cyt) P450cam, cyt P40 1A2, and myoglobin in the array were activated with H2O2 to metabolize BP to genotoxic metabolites. DNA damage by the metabolites was detected by increases in square wave voltammetric oxidation peaks using Ru(bpy)3(2+) as catalyst. Cyt P450cam and cyt P450 1A2 showed 3-fold higher activity for genotoxic bioactivation of BP than myoglobin. The ability of the arrays to generate and detect metabolite-based DNA damage simultaneously for several enzymes is a rapid and promising approach to identify and characterize enzymes involved in genotoxicity of drugs and pollutants. PMID:15732919

  4. Novel and Distinct Metabolites Identified Following a Single Oral Dose of α- or γ-Hexabromocyclododecane in Mice

    PubMed Central

    Szabo, David T.; Huwe, Janice; Diliberto, Janet; Birnbaum, Linda S.

    2013-01-01

    The metabolism of α- and γ-hexabromocyclododecane (HBCD) was investigated in adult C57BL/6 female mice. α- or γ-[14C]HBCD (3 mg/kg bw) was orally administered with subsequent urine and feces collection for 4 consecutive days; a separate group of mice were dosed and sacrificed 3 hours post-exposure to investigate tissue metabolite levels. Extractable and non-extractable HBCD metabolites were quantitated in liver, blood, fat, brain, bile, urine and feces and characterized by LC/MS (ESI-). Metabolites identified were distinct between the two stereoisomers. In mice exposed to α-HBCD, four hydroxylated metabolites were detected in fecal extracts, and one of these metabolite isomers was consistently characterized in liver, brain, and adipose tissue extracts. In contrast, mice exposed to γ-HBCD contained multiple isomers of monohydroxy-pentabromocyclododecene, dihydroxy-pentabromocyclododecene, and dihydroxy-pentabromocyclododecadiene in the feces while only a single monohydroxy-pentabromocyclododecane metabolite was measured in liver and adipose tissue. Both stereoisomers were transformed to metabolites which formed covalent bonds to proteins and/or lipids in the gut as evidenced by high fecal non-extractables. Although the potential toxicity of these free and bound metabolites remains to be determined, the presence of distinct metabolic products from the two main HBCD stereoisomers should allow biomarkers to be selected that may aid in characterizing sources of HBCD exposure. PMID:23171393

  5. Volatile Metabolites of Pathogens: A Systematic Review

    PubMed Central

    Bos, Lieuwe D. J.; Sterk, Peter J.; Schultz, Marcus J.

    2013-01-01

    Ideally, invading bacteria are detected as early as possible in critically ill patients: the strain of morbific pathogens is identified rapidly, and antimicrobial sensitivity is known well before the start of new antimicrobial therapy. Bacteria have a distinct metabolism, part of which results in the production of bacteria-specific volatile organic compounds (VOCs), which might be used for diagnostic purposes. Volatile metabolites can be investigated directly in exhaled air, allowing for noninvasive monitoring. The aim of this review is to provide an overview of VOCs produced by the six most abundant and pathogenic bacteria in sepsis, including Staphylococcus aureus, Streptococcus pneumoniae, Enterococcus faecalis, Pseudomonas aeruginosa, Klebsiella pneumoniae, and Escherichia coli. Such VOCs could be used as biological markers in the diagnostic approach of critically ill patients. A systematic review of existing literature revealed 31 articles. All six bacteria of interest produce isopentanol, formaldehyde, methyl mercaptan, and trimethylamine. Since humans do not produce these VOCs, they could serve as biological markers for presence of these pathogens. The following volatile biomarkers were found for identification of specific strains: isovaleric acid and 2-methyl-butanal for Staphylococcus aureus; 1-undecene, 2,4-dimethyl-1-heptane, 2-butanone, 4-methyl-quinazoline, hydrogen cyanide, and methyl thiocyanide for Pseudomonas aeruginosa; and methanol, pentanol, ethyl acetate, and indole for Escherichia coli. Notably, several factors that may effect VOC production were not controlled for, including used culture media, bacterial growth phase, and genomic variation within bacterial strains. In conclusion, VOCs produced by bacteria may serve as biological markers for their presence. Goal-targeted studies should be performed to identify potential sets of volatile biological markers and evaluate the diagnostic accuracy of these markers in critically ill patients. PMID

  6. Degradation of chloroacetanilide herbicides: The prevalence of sulfonic and oxanilic acid metabolites in Iowa groundwaters and surface waters

    USGS Publications Warehouse

    Kalkhoff, S.J.; Kolpin, D.W.; Thurman, E.M.; Ferrer, I.; Barcelo, D.

    1998-01-01

    Water samples were collected from 88 municipal wells throughout Iowa during the summer and were collected monthly at 12 stream sites in eastern Iowa from March to December 1996 to study the occurrence of the sulfonic and oxanilic metabolites of acetochlor, alachlor, and metolachlor. The sulfonic and oxanilic metabolites were present in almost 75% of the groundwater samples and were generally present from 3 to 45 times more frequently than their parent compounds. In groundwater, the median value of the summed concentrations of acetochlor, alachlor, and metolachlor was less than 0.05 μg/L, and the median value of the summed concentrations of the six metabolites was 1.2 μg/L. All surface water samples contained at least one detectable metabolite compound. Individual metabolites were detected from 2 to over 100 times more frequently than the parent compounds. In surface water, the median value of the summed concentrations of the three parent compounds was 0.13 μg/L, and the median value of the summed concentrations of the six metabolites was 6.4 μg/L. These data demonstrate the importance of analyzing both parent compounds and metabolites to more fully understand the environmental fate and transport of herbicides in the hydrologic system.

  7. Multifaceted Investigation of Metabolites During Nitrogen Fixation in Medicago via High Resolution MALDI-MS Imaging and ESI-MS

    PubMed Central

    Gemperline, Erin; Jayaraman, Dhileepkumar; Maeda, Junko; Ané, Jean-Michel; Li, Lingjun

    2014-01-01

    Legumes have developed the unique ability to establish a symbiotic relationship with soil bacteria known as rhizobia. This interaction results in the formation of root nodules in which rhizobia thrive and reduce atmospheric dinitrogen into plant-usable ammonium through biological nitrogen fixation (BNF). Due to the availability of genetic information for both of the symbiotic partners, the Medicago truncatula–Sinorhizobium meliloti association is an excellent model for examining the BNF process. Although metabolites are important in this symbiotic association, few studies have investigated the array of metabolites that influence this process. Of these studies, most target only a few specific metabolites, the roles of which are either well known or are part of a well-characterized metabolic pathway. Here, we used a multifaceted mass spectrometric (MS) approach to detect and identify the key metabolites that are present during BNF using the Medicago truncatula–Sinorhizobium meliloti association as the model system. High mass accuracy and high resolution matrix-assisted laser desorption/ionization (MALDI) and electrospray ionization (ESI) Orbitrap instruments were used in this study and provide complementary results for more in-depth characterization of the nitrogen-fixation process. We used well-characterized plant and bacterial mutants to highlight differences between the metabolites that are present in functional vs. non-functional nodules. Our study highlights the benefits of using a combination of mass spectrometric techniques to detect differences in metabolite composition and the distributions of these metabolites in plant biology. PMID:25323862

  8. Determination of rate constants and half-lives for the simultaneous biodegradation of several cyanobacterial metabolites in Australian source waters.

    PubMed

    Ho, Lionel; Tang, Tim; Hoefel, Daniel; Vigneswaran, Bala

    2012-11-01

    The fate of five cyanobacterial metabolites was assessed in water sourced from Lake Burragorang (Warragamba Dam) in New South Wales, Australia. All of the studied metabolites were shown to be biodegradable in this water source. For some metabolites, biodegradation was influenced by factors, including temperature, location (within the water body) and seasonal variations. The biodegradation of the metabolites was shown to follow pseudo-first-order kinetics with rate constants ranging from 8.0 × 10(-4) to 1.3 × 10(-2) h(-1). Half-lives of the metabolites were also estimated and ranged from 2.2 to 36.1 d. The order of ease of biodegradability in this water source followed the trend: microcystin-LR ≥ cylindrospermopsin > saxitoxins > geosmin ≥ 2-methylisoborneol. The lack of detection of the mlrA gene during microcystin biodegradation suggests that these toxins may be degraded via a different pathway. While no metabolite-degrading organisms were isolated in this study, the inoculation of previously isolated geosmin- and microcystin-degrading bacteria into Lake Burragorang water resulted in efficient biodegradation of the respective metabolites. For example, microcystin-degrading isolate TT25 was able to degrade three microcystin variants to concentrations below analytical detection within 24 h, suggesting that inoculation of such bacteria has the potential to enhance biodegradation in Lake Burragorang. PMID:22921397

  9. Multifaceted Investigation of Metabolites During Nitrogen Fixation in Medicago via High Resolution MALDI-MS Imaging and ESI-MS

    NASA Astrophysics Data System (ADS)

    Gemperline, Erin; Jayaraman, Dhileepkumar; Maeda, Junko; Ané, Jean-Michel; Li, Lingjun

    2015-01-01

    Legumes have developed the unique ability to establish a symbiotic relationship with soil bacteria known as rhizobia. This interaction results in the formation of root nodules in which rhizobia thrive and reduce atmospheric dinitrogen into plant-usable ammonium through biological nitrogen fixation (BNF). Owing to the availability of genetic information for both of the symbiotic partners, the Medicago truncatula- Sinorhizobium meliloti association is an excellent model for examining the BNF process. Although metabolites are important in this symbiotic association, few studies have investigated the array of metabolites that influence this process. Of these studies, most target only a few specific metabolites, the roles of which are either well known or are part of a well-characterized metabolic pathway. Here, we used a multifaceted mass spectrometric (MS) approach to detect and identify the key metabolites that are present during BNF using the Medicago truncatula- Sinorhizobium meliloti association as the model system. High mass accuracy and high resolution matrix-assisted laser desorption/ionization (MALDI) and electrospray ionization (ESI) Orbitrap instruments were used in this study and provide complementary results for more in-depth characterization of the nitrogen-fixation process. We used well-characterized plant and bacterial mutants to highlight differences between the metabolites that are present in functional versus nonfunctional nodules. Our study highlights the benefits of using a combination of mass spectrometric techniques to detect differences in metabolite composition and the distributions of these metabolites in plant biology.

  10. Salmonella Typhi and Salmonella Paratyphi A elaborate distinct systemic metabolite signatures during enteric fever

    PubMed Central

    Näsström, Elin; Vu Thieu, Nga Tran; Dongol, Sabina; Karkey, Abhilasha; Voong Vinh, Phat; Ha Thanh, Tuyen; Johansson, Anders; Arjyal, Amit; Thwaites, Guy; Dolecek, Christiane; Basnyat, Buddha; Baker, Stephen; Antti, Henrik

    2014-01-01

    The host–pathogen interactions induced by Salmonella Typhi and Salmonella Paratyphi A during enteric fever are poorly understood. This knowledge gap, and the human restricted nature of these bacteria, limit our understanding of the disease and impede the development of new diagnostic approaches. To investigate metabolite signals associated with enteric fever we performed two dimensional gas chromatography with time-of-flight mass spectrometry (GCxGC/TOFMS) on plasma from patients with S. Typhi and S. Paratyphi A infections and asymptomatic controls, identifying 695 individual metabolite peaks. Applying supervised pattern recognition, we found highly significant and reproducible metabolite profiles separating S. Typhi cases, S. Paratyphi A cases, and controls, calculating that a combination of six metabolites could accurately define the etiological agent. For the first time we show that reproducible and serovar specific systemic biomarkers can be detected during enteric fever. Our work defines several biologically plausible metabolites that can be used to detect enteric fever, and unlocks the potential of this method in diagnosing other systemic bacterial infections. DOI: http://dx.doi.org/10.7554/eLife.03100.001 PMID:24902583

  11. Identification of signatory secondary metabolites during mycoparasitism of Rhizoctonia solani by Stachybotrys elegans

    PubMed Central

    Chamoun, Rony; Aliferis, Konstantinos A.; Jabaji, Suha

    2015-01-01

    Stachybotrys elegans is able to parasitize the fungal plant pathogen Rhizoctonia solani AG-3 following a complex and intimate interaction, which, among others, includes the production of cell wall-degrading enzymes, intracellular colonization, and expression of pathogenic process encoding genes. However, information on the metabolome level is non-existent during mycoparasitism. Here, we performed a direct-infusion mass spectrometry (DIMS) metabolomics analysis using an LTQ Orbitrap analyzer in order to detect changes in the profiles of induced secondary metabolites of both partners during this mycoparasitic interaction 4 and 5 days following its establishment. The diketopiperazine(s) (DKPs) cyclo(S-Pro-S-Leu)/cyclo(S-Pro-S-Ile), ethyl 2-phenylacetate, and 3-nitro-4-hydroxybenzoic acid were detected as the primary response of Rhizoctonia 4 days following dual-culturing with Stachybotrys, whereas only the latter metabolite was up-regulated 1 day later. On the other hand, trichothecenes and atranones were mycoparasite-derived metabolites identified during mycoparasitism 4 and 5 days following dual-culturing. All the above secondary metabolites are known to exhibit bioactivity, including fungitoxicity, and represent key elements that determine the outcome of the interaction being studied. Results could be further exploited in programs for the evaluation of the bioactivity of these metabolites per se or their chemical analogs, and/or genetic engineering programs to obtain more efficient mycoparasite strains with improved efficacy and toxicological profiles. PMID:25972848

  12. Levels of Pesticides and Their Metabolites in Wistar Rat Amniotic Fluids and Maternal Urine upon Gestational Exposure

    PubMed Central

    Bossi, Rossana; Vinggaard, Anne Marie; Taxvig, Camilla; Boberg, Julie; Bonefeld-Jørgensen, Eva Cecilie

    2013-01-01

    Concentrations of pesticides and selected metabolites in rat urine and amniotic fluid were determined as biomarker upon oral administration of Wistar rats to two pesticide mixtures consisting of three to five pesticides (bitertanol, propiconazole, cypermethrin, malathion, and terbuthylazine). The pesticides and their metabolites were found in rat amniotic fluid and urine, generally in dose-response concentrations in relation to dosage. The measurement of the substances in the amniotic fluid indicated that the fetus was exposed to the pesticides as well as their metabolites. Moreover, the pesticides detected in urine demonstrated the exposure as well as the ability of the rat to excrete these compounds. PMID:23736656

  13. Reference range levels of polycyclic aromatic hydrocarbons in the US population by measurement of urinary monohydroxy metabolites

    SciTech Connect

    Grainger, James . E-mail: jag2@cdc.gov; Huang, Wenlin; Patterson, Donald G.; Turner, Wayman E.; Pirkle, James; Caudill, Samuel P.; Wang, Richard Y.; Needham, Larry L.; Sampson, Eric J.

    2006-03-15

    We developed a gas chromatography isotope-dilution high-resolution mass spectrometry (GC/Id-HRMS) method for measuring 14 polycyclic aromatic hydrocarbon (PAH) metabolites representing seven parent PAHs in 3 mL of urine at low parts-per-trillion levels. PAH levels were determined in urine samples collected in 1999 and 2000 from approximately 2400 participants in the National Health and Nutrition Examination Survey, and, for the first time, reference range values were calculated for these metabolites in the US population. Using this GC/ID-HRMS method, we found detectable concentrations for monohydroxy metabolite isomers of fluorene, phenanthrene, fluoranthene, pyrene, and chrysene, benzo[c]phenanthrene, and benz[a]anthracene. Some monohydroxy metabolite isomers of benzo[c]phenanthrene, chrysene, and benz[a]anthracene exhibited low detection frequencies that did not allow for geometric mean calculations. Our study results enabled us to establish a reference range for the targeted PAHs in the general US population.

  14. [Secondary Metabolites from Marine Microorganisms. I. Secondary Metabolites from Marine Actinomycetes].

    PubMed

    Orlova, T I; Bulgakova, V G; Polin, A N

    2015-01-01

    Review represents data on new active metabolites isolated from marine actinomycetes published in 2007 to 2014. Marine actinomycetes are an unlimited source of novel secondary metabolites with various biological activities. Among them there are antibiotics, anticancer compounds, inhibitors of biochemical processes. PMID:26863742

  15. Mycotoxin detection.

    PubMed

    Anfossi, Laura; Giovannoli, Cristina; Baggiani, Claudio

    2016-02-01

    Mycotoxins are toxic metabolites of certain fungi that growth on a variety of crops, pre-harvest, during and post-harvest. Because of their toxicity, maximum admissible levels of mycotoxins are regulated worldwide and monitoring of their occurrence in several commodities is mandatory for assuring food safety and consumers' health protection. Analytical methods for mycotoxins include immunochemical-based techniques that principally apply for routinely controls and rapid, on-site detection, and chromatographic-based techniques that provide sensitive, accurate and selective determination of known mycotoxins, besides identification of new or modified compounds through tandem mass spectrometric detectors. PMID:26723009

  16. Metabolite fingerprinting of exudates from carrot roots grown under phosphorus stress

    Technology Transfer Automated Retrieval System (TEKTRAN)

    To see if differences in the metabolite content of exudates obtained from cultured carrot roots grown in the presence and absence of phosphorus could be detected, crude samples were analyzed via Fourier Transform Ion Cyclotron Mass Spectrometry (FTMS). The highly accurate masses and elemental compo...

  17. Stereoselective bioaccumulation and metabolite formation of triadimefon in Tubifex tubifex.

    PubMed

    Liu, Tiantian; Diao, Jinling; Di, Shanshan; Zhou, Zhiqiang

    2014-06-17

    Triadimefon, a chiral fungicide, could be metabolized to triadimenol which has two chiral centers. In this work, Tubifex tubifex was exposed to triadimefon through the aqueous and soil phase to explore the relative importance of the routes of uptake. Bioaccumulation of triadimefon in tubifex was detected in both treatments, and the kinetics of the accumulation processes were significantly different in these two experiments. In spiked water treatment, (S)-triadimefon was preferentially accumulated over the (R)-triadimefon, whereas the enantioselective bioaccumulation was not detected in the spiked soil microenvironment. Simultaneously, four stereoisomers of triadimenol were also found in the tubifex tissue. Although the amount of these stereoisomers were different from each other with relatively more accumulation of the most fungi-toxic stereoisomer (1S,2R), the abundance ratios in the two exposure treatments were similar at the same sampling, following the order (1S,2S) > (1R,2S) > (1R,2R) > (1S,2R). The bioaccumulation factor was calculated for parent compound triadimefon and metabolite enrichment factor for metabolite. The results showed that both uptake routes, epidermal contact in the aqueous phase and ingestion of solid particles in soil, were important to the bioaccumulation of the triadimefon and triadimenol in tubifex. PMID:24846121

  18. Exploring the transport of plant metabolites using positron emitting radiotracers

    PubMed Central

    Kiser, Matthew R.; Reid, Chantal D.; Crowell, Alexander S.; Phillips, Richard P.; Howell, Calvin R.

    2008-01-01

    Short-lived positron-emitting radiotracer techniques provide time-dependent data that are critical for developing models of metabolite transport and resource distribution in plants and their microenvironments. Until recently these techniques were applied to measure radiotracer accumulation in coarse regions along transport pathways. The recent application of positron emission tomography (PET) techniques to plant research allows for detailed quantification of real-time metabolite dynamics on previously unexplored spatial scales. PET provides dynamic information with millimeter-scale resolution on labeled carbon, nitrogen, and water transport over a small plant-size field of view. Because details at the millimeter scale may not be required for all regions of interest, hybrid detection systems that combine high-resolution imaging with other radiotracer counting technologies offer the versatility needed to pursue wide-ranging plant physiological and ecological research. In this perspective we describe a recently developed hybrid detection system at Duke University that provides researchers with the flexibility required to carry out measurements of the dynamic responses of whole plants to environmental change using short-lived radiotracers. Following a brief historical development of radiotracer applications to plant research, the role of radiotracers is presented in the context of various applications at the leaf to the whole-plant level that integrates cellular and subcellular signals and∕or controls. PMID:19404430

  19. Secondary metabolites in fungus-plant interactions

    PubMed Central

    Pusztahelyi, Tünde; Holb, Imre J.; Pócsi, István

    2015-01-01

    Fungi and plants are rich sources of thousands of secondary metabolites. The genetically coded possibilities for secondary metabolite production, the stimuli of the production, and the special phytotoxins basically determine the microscopic fungi-host plant interactions and the pathogenic lifestyle of fungi. The review introduces plant secondary metabolites usually with antifungal effect as well as the importance of signaling molecules in induced systemic resistance and systemic acquired resistance processes. The review also concerns the mimicking of plant effector molecules like auxins, gibberellins and abscisic acid by fungal secondary metabolites that modulate plant growth or even can subvert the plant defense responses such as programmed cell death to gain nutrients for fungal growth and colonization. It also looks through the special secondary metabolite production and host selective toxins of some significant fungal pathogens and the plant response in form of phytoalexin production. New results coming from genome and transcriptional analyses in context of selected fungal pathogens and their hosts are also discussed. PMID:26300892

  20. Metabolite profiles during oral glucose challenge.

    PubMed

    Ho, Jennifer E; Larson, Martin G; Vasan, Ramachandran S; Ghorbani, Anahita; Cheng, Susan; Rhee, Eugene P; Florez, Jose C; Clish, Clary B; Gerszten, Robert E; Wang, Thomas J

    2013-08-01

    To identify distinct biological pathways of glucose metabolism, we conducted a systematic evaluation of biochemical changes after an oral glucose tolerance test (OGTT) in a community-based population. Metabolic profiling was performed on 377 nondiabetic Framingham Offspring cohort participants (mean age 57 years, 42% women, BMI 30 kg/m(2)) before and after OGTT. Changes in metabolite levels were evaluated with paired Student t tests, cluster-based analyses, and multivariable linear regression to examine differences associated with insulin resistance. Of 110 metabolites tested, 91 significantly changed with OGTT (P ≤ 0.0005 for all). Amino acids, β-hydroxybutyrate, and tricarboxylic acid cycle intermediates decreased after OGTT, and glycolysis products increased, consistent with physiological insulin actions. Other pathways affected by OGTT included decreases in serotonin derivatives, urea cycle metabolites, and B vitamins. We also observed an increase in conjugated, and a decrease in unconjugated, bile acids. Changes in β-hydroxybutyrate, isoleucine, lactate, and pyridoxate were blunted in those with insulin resistance. Our findings demonstrate changes in 91 metabolites representing distinct biological pathways that are perturbed in response to an OGTT. We also identify metabolite responses that distinguish individuals with and without insulin resistance. These findings suggest that unique metabolic phenotypes can be unmasked by OGTT in the prediabetic state. PMID:23382451

  1. Recent Advances in Mass Spectrometry for the Identification of Neuro-chemicals and their Metabolites in Biofluids

    PubMed Central

    Kailasa, Suresh Kumar; Wu, Hui-Fen

    2013-01-01

    Recently, mass spectrometric related techniques have been widely applied for the identification and quantification of neurochemicals and their metabolites in biofluids. This article presents an overview of mass spectrometric techniques applied in the detection of neurological substances and their metabolites from biological samples. In addition, the advances of chromatographic methods (LC, GC and CE) coupled with mass spectrometric techniques for analysis of neurochemicals in pharmaceutical and biological samples are also discussed. PMID:24381533

  2. Qualitative profiling and quantification of neonicotinoid metabolites in human urine by liquid chromatography coupled with mass spectrometry.

    PubMed

    Taira, Kumiko; Fujioka, Kazutoshi; Aoyama, Yoshiko

    2013-01-01

    Neonicotinoid pesticides have been widely applied for the production of fruits and vegetables, and occasionally detected in conventionally grown produce. Thus oral exposure to neonicotinoid pesticides may exist in the general population; however, neonicotinoid metabolites in human body fluids have not been investigated comprehensively. The purpose of this study is the qualitative profiling and quantitative analysis of neonicotinoid metabolites in the human spot urine by liquid chromatography coupled with mass spectrometry (LC/MS). Human urine samples were collected from three patients suspected of subacute exposure to neonicotinoid pesticides. A qualitative profiling of urinary metabolites was performed using liquid chromatography/time-of-flight mass spectrometry (LC/TOFMS) with a database of nominal molecular weights of 57 known metabolites of three neonicotinoid pesticides (acetamiprid, Imidacloprid, and clothianidin), as well as the parent compounds. Then a quantitative analysis of selected urinary metabolites was performed using liquid chromatography/tandem mass spectrometry (LC/MS/MS) with a standard pesticide and metabolite, which were detected by the qualitative profiling. The result of qualitative profiling showed that seven metabolites, i.e. an acetamiprid metabolite, N-desmethyl-acetamiprid; three Imidacloprid metabolites, 5-hydroxy-Imidacloprid, 4,5-dihydroxy-imidacloprid, 4,5-dehydro-Imidacloprid; a common metabolite of acetamiprid and Imidacloprid, N-(6-chloronicotinoyl)-glycine; and two clothianidin metabolites, N-desmethyl-clothianidin, N-(2-(methylsulfanyl)thiazole-5-carboxyl)-glycine, as well as acetamiprid, were detected in the urine of three cases. The result of the quantitative analysis showed N-desmethyl-acetamiprid was determined in the urine of one case, which had been collected on the first visit, at a concentration of 3.2 ng/mL. This is the first report on the qualitative and quantitative detection of N-desmethyl-acetamiprid in the human

  3. Qualitative Profiling and Quantification of Neonicotinoid Metabolites in Human Urine by Liquid Chromatography Coupled with Mass Spectrometry

    PubMed Central

    Taira, Kumiko; Fujioka, Kazutoshi; Aoyama, Yoshiko

    2013-01-01

    Neonicotinoid pesticides have been widely applied for the production of fruits and vegetables, and occasionally detected in conventionally grown produce. Thus oral exposure to neonicotinoid pesticides may exist in the general population; however, neonicotinoid metabolites in human body fluids have not been investigated comprehensively. The purpose of this study is the qualitative profiling and quantitative analysis of neonicotinoid metabolites in the human spot urine by liquid chromatography coupled with mass spectrometry (LC/MS). Human urine samples were collected from three patients suspected of subacute exposure to neonicotinoid pesticides. A qualitative profiling of urinary metabolites was performed using liquid chromatography/time-of-flight mass spectrometry (LC/TOFMS) with a database of nominal molecular weights of 57 known metabolites of three neonicotinoid pesticides (acetamiprid, Imidacloprid, and clothianidin), as well as the parent compounds. Then a quantitative analysis of selected urinary metabolites was performed using liquid chromatography/tandem mass spectrometry (LC/MS/MS) with a standard pesticide and metabolite, which were detected by the qualitative profiling. The result of qualitative profiling showed that seven metabolites, i.e. an acetamiprid metabolite, N-desmethyl-acetamiprid; three Imidacloprid metabolites, 5-hydroxy-Imidacloprid, 4,5-dihydroxy-imidacloprid, 4,5-dehydro-Imidacloprid; a common metabolite of acetamiprid and Imidacloprid, N-(6-chloronicotinoyl)-glycine; and two clothianidin metabolites, N-desmethyl-clothianidin, N-(2-(methylsulfanyl)thiazole-5-carboxyl)-glycine, as well as acetamiprid, were detected in the urine of three cases. The result of the quantitative analysis showed N-desmethyl-acetamiprid was determined in the urine of one case, which had been collected on the first visit, at a concentration of 3.2 ng/mL. This is the first report on the qualitative and quantitative detection of N-desmethyl-acetamiprid in the human

  4. Monitoring microbial metabolites using an inductively coupled resonance circuit

    PubMed Central

    Karnaushenko, Daniil; Baraban, Larysa; Ye, Dan; Uguz, Ilke; Mendes, Rafael G.; Rümmeli, Mark H.; de Visser, J. Arjan G. M.; Schmidt, Oliver G.; Cuniberti, Gianaurelio; Makarov, Denys

    2015-01-01

    We present a new approach to monitor microbial population dynamics in emulsion droplets via changes in metabolite composition, using an inductively coupled LC resonance circuit. The signal measured by such resonance detector provides information on the magnetic field interaction with the bacterial culture, which is complementary to the information accessible by other detection means, based on electric field interaction, i.e. capacitive or resistive, as well as optical techniques. Several charge-related factors, including pH and ammonia concentrations, were identified as possible contributors to the characteristic of resonance detector profile. The setup enables probing the ionic byproducts of microbial metabolic activity at later stages of cell growth, where conventional optical detection methods have no discriminating power. PMID:26264183

  5. Monitoring microbial metabolites using an inductively coupled resonance circuit

    NASA Astrophysics Data System (ADS)

    Karnaushenko, Daniil; Baraban, Larysa; Ye, Dan; Uguz, Ilke; Mendes, Rafael G.; Rümmeli, Mark H.; de Visser, J. Arjan G. M.; Schmidt, Oliver G.; Cuniberti, Gianaurelio; Makarov, Denys

    2015-08-01

    We present a new approach to monitor microbial population dynamics in emulsion droplets via changes in metabolite composition, using an inductively coupled LC resonance circuit. The signal measured by such resonance detector provides information on the magnetic field interaction with the bacterial culture, which is complementary to the information accessible by other detection means, based on electric field interaction, i.e. capacitive or resistive, as well as optical techniques. Several charge-related factors, including pH and ammonia concentrations, were identified as possible contributors to the characteristic of resonance detector profile. The setup enables probing the ionic byproducts of microbial metabolic activity at later stages of cell growth, where conventional optical detection methods have no discriminating power.

  6. Analysis of cocaine and metabolites in hair: validation and application of measurement of hydroxycocaine metabolites as evidence of cocaine ingestion.

    PubMed

    Schaffer, Michael; Cheng, Chen-Chih; Chao, Oscar; Hill, Virginia; Matsui, Paul

    2016-03-01

    An LC/MS/MS method to identify and quantitate in hair the minor metabolites of cocaine-meta-, para-, and ortho-hydroxy cocaine-was developed and validated. Analysis was performed on a triple quadrupole ABSciex API 3000 MS equipped with an atmospheric pressure ionization source via an IonSpray (ESI). For LC, a series 200 micro binary pump with a Perkin Elmer Model 200 autosampler was used. The limit of detection (LOD) and limit of quantification (LOQ) were 0.02 ng/10 mg hair, with linearity from 0.02 to 10 ng/10 mg hair. Concentrations of the para isomer in extensively washed hair samples were in the range of 1-2 % of the cocaine in the sample, while the concentrations of the ortho form were considerably less. The method was used to analyze large numbers of samples from two populations: workplace and criminal justice. In vitro experiments to determine if deodorants or peroxide-containing cosmetic treatments could result in the presence of these metabolites in hair showed that this does not occur with extensively washed hair. Presence of hydroxycocaines, when detected after aggressive washing of the hair samples, provides a valuable additional indicator of ingestion of cocaine rather than mere environmental exposure. PMID:26873203

  7. Determining protein adducts of fipexide: mass spectrometry based assay for confirming the involvement of its reactive metabolite in covalent binding.

    PubMed

    Sleno, Lekha; Varesio, Emmanuel; Hopfgartner, Gérard

    2007-01-01

    Fipexide is a nootropic drug, withdrawn from the market due to its idiosyncratic drug reactions causing adverse effects in man. Previous work on its metabolites has identified several potential reactive metabolites which could be implicated in protein binding. Here, we investigated the formation of these metabolites in rat and human hepatocytes. Based on these results, the o-quinone of fipexide (FIP), formed via the demethylenation reaction through a catechol intermediate, was chosen for further investigation. Studies were then pursued in order to relate this metabolite to protein binding, and thus better understand potential mechanisms for the toxicity of the parent compound. An assay was developed for determining the fipexide catechol-cysteine adduct in the microsomal protein fractions following in vitro incubations. This method digests the entire protein fraction into amino acids, followed by the detection of the Cys-metabolite adduct by liquid chromatography/mass spectrometry (LC/MS). We have designed a strategy where drug metabolism taking place in microsomal incubations and involved in protein binding can be assessed after the proteins have been digested, with the detection of the specific amino acid adduct. In this study, the structure of the fipexide adduct was hypothesized using knowledge previously gained in glutathione and N-acetylcysteine trapping experiments. Acetaminophen was used as a positive control for detecting a drug metabolite-cysteine adduct by LC/MS. This approach has the potential to be applicable as a protein-binding assay in early drug discovery without the need for radioactive compounds. PMID:18022964

  8. High-performance liquid chromatography and immunoassay techniques for monitoring urinary metabolites of polycyclic aromatic hydrocarbons

    SciTech Connect

    Chamberlain, V.C.

    1988-01-01

    The purpose of this research was to investigate the feasibility of using high performance liquid chromatography (HPLC) and enzyme linked immunosorbent assays (ELISA) as sensitive techniques for monitoring polycyclic aromatic hydrocarbon (PAH) metabolites in human urine. The method was tested using synthesized PAH conjugates as positive markers. Results showed that a PAH conjugate, S-(9,10-dihydro-9-hydroxy-10-phenanthryl)N-acetyl cysteine (PHONAC), present in HPLC effluent could be detected by ELISA at picomole levels, well below the sensitivity of the HPLC UV detector. Analyses of urine from mice dosed with phenanthrene demonstrated that a substance detected by HPLC which was not detected in ELISA tests was the principal phenanthrene metabolite. This substance was not hydrolysed by Beta-glucuronidase. PHONAC was detected by ELISA in mouse urine extracts subjected to HPLC.

  9. Solid-phase extraction and HPLC analysis of kebuzone and its metabolites in blood.

    PubMed

    Klimes, J; Sochor, J; Sedlacek, J

    1996-09-01

    A HPLC method for quantification of kebuzone and its metabolites in whole blood was developed. The compounds and the internal standard were isolated from blood by solid-phase extraction on a C-18 cartridge. A blood sample was to be hemolyzed before extraction. HPLC was performed on a C-18 column with the mobile phase composed of methanol/water acidified to pH 2.7 and UV absorbance detection at 247 nm. This method has been successfully applied to a pharmacokinetic study of kebuzone and its metabolites in rabbits. PMID:8878255

  10. Transporter and its engineering for secondary metabolites.

    PubMed

    Lv, Huajun; Li, Jianhua; Wu, Yingying; Garyali, Sanjog; Wang, Yong

    2016-07-01

    Secondary metabolites possess a lot of biological activities, and to achieve their functions, transmembrane transportation is crucial. Elucidation of their transport mechanisms in the cell is critical for discovering ways to improve the production. Here, we have summarized the recent progresses for representative secondary metabolite transporters and also the strategies for uncovering the transporter systems in plants and microbes. We have also discussed the transporter engineering strategies being utilized for improving the heterologous natural product production, which exhibits promising future under the guide of synthetic biology. PMID:27209041

  11. [Autism and Autism-associated Metabolites].

    PubMed

    Watanabe, Kunitomo

    2016-06-01

    Gene-microbiota interactions are now proposed to be a special case of gene-environmental interaction. Preclinical and clinical data summarized in this article reveal that a specific serum metabolite, associated with alterations in gut microbiome composition, might have an emerging role in the onset and pathogenesis of autism. Altered level of this specified metabolite may induce perturbations in the epigenome and modulate the expression of key disease susceptible genes in neurons and their associated cells during critical periods of neurodevelopment. The gut microbiota itself is now regarded as a reservoir for environmental epigenetic factors. PMID:27279160

  12. Streptomyces metabolites in divergent microbial interactions.

    PubMed

    Takano, Hideaki; Nishiyama, Tatsuya; Amano, Sho-ichi; Beppu, Teruhiko; Kobayashi, Michihiko; Ueda, Kenji

    2016-03-01

    Streptomyces and related bacteria produce a wide variety of secondary metabolites. Of these, many compounds have industrial applications, but the question of why this group of microorganism produces such various kinds of biologically active substances has not yet been clearly answered. Here, we overview the results from our studies on the novel function and role of Streptomyces metabolites. The diverged action of negative and positive influences onto the physiology of various microorganisms infers the occurrence of complex microbial interactions due to the effect of small molecules produced by Streptomyces. The interactions may serve as a basis for the constitution of biological community. PMID:26408311

  13. Multi-mycotoxin screening reveals the occurrence of 139 different secondary metabolites in feed and feed ingredients.

    PubMed

    Streit, Elisabeth; Schwab, Christina; Sulyok, Michael; Naehrer, Karin; Krska, Rudolf; Schatzmayr, Gerd

    2013-03-01

    The development of liquid chromatography-mass spectrometry (LC-MS)/mass spectrometry (MS) methods for the simultaneous detection and quantification of a broad spectrum of mycotoxins has facilitated the screening of a larger number of samples for contamination with a wide array of less well-known "emerging" mycotoxins and other metabolites. In this study, 83 samples of feed and feed raw materials were analysed. All of them were found to contain seven to 69 metabolites. The total number of detected metabolites amounts to 139. Fusarium mycotoxins were most common, but a number of Alternaria toxins also occurred very often. Furthermore, two so-called masked mycotoxins (i.e., mycotoxin conjugates), namely deoxynivalenol-3-glucoside (75% positives) and zearalenone-4-sulfate (49% positives), were frequently detected. Although the observed median concentrations of the individual analytes were generally in the low μg/kg range, evaluating the toxicological potential of a given sample is difficult. Toxicity data on less well-known mycotoxins and other detected metabolites are notoriously scarce, as an overview on the available information on the most commonly detected metabolites shows. Besides, the possible synergistic effects of co-occurring substances have to be considered. PMID:23529186

  14. Multi-Mycotoxin Screening Reveals the Occurrence of 139 Different Secondary Metabolites in Feed and Feed Ingredients

    PubMed Central

    Streit, Elisabeth; Schwab, Christina; Sulyok, Michael; Naehrer, Karin; Krska, Rudolf; Schatzmayr, Gerd

    2013-01-01

    The development of liquid chromatography-mass spectrometry (LC-MS)/mass spectrometry (MS) methods for the simultaneous detection and quantification of a broad spectrum of mycotoxins has facilitated the screening of a larger number of samples for contamination with a wide array of less well-known “emerging” mycotoxins and other metabolites. In this study, 83 samples of feed and feed raw materials were analysed. All of them were found to contain seven to 69 metabolites. The total number of detected metabolites amounts to 139. Fusarium mycotoxins were most common, but a number of Alternaria toxins also occurred very often. Furthermore, two so-called masked mycotoxins (i.e., mycotoxin conjugates), namely deoxynivalenol-3-glucoside (75% positives) and zearalenone-4-sulfate (49% positives), were frequently detected. Although the observed median concentrations of the individual analytes were generally in the low μg/kg range, evaluating the toxicological potential of a given sample is difficult. Toxicity data on less well-known mycotoxins and other detected metabolites are notoriously scarce, as an overview on the available information on the most commonly detected metabolites shows. Besides, the possible synergistic effects of co-occurring substances have to be considered. PMID:23529186

  15. Presence and fate of carbamazepine, oxcarbazepine, and seven of their metabolites at wastewater treatment plants.

    PubMed

    Leclercq, Marie; Mathieu, Olivier; Gomez, Elena; Casellas, Claude; Fenet, Hélène; Hillaire-Buys, Dominique

    2009-04-01

    Many pharmaceuticals are excreted in wastewater as parent substances or metabolites subsequent to therapeutic or diagnostic application in medical care. This includes the antiepileptic carbamazepine, which is not removed during conventional wastewater treatment and was found to be ubiquitous in the aquatic environment. Some carbamazepine metabolites have also been found in treated wastewater, but only five of them have been studied to date. However, at least 30 carbamazepine metabolites have been identified in humans, including some pharmacologically active or genotoxic compounds. Oxcarbazepine, an antiepileptic which is increasingly used, generates metabolites common to those of carbamazepine. The present work focuses on the presence of carbamazepine, oxcarbazepine, and seven of their metabolites (carbamazepine-10,11-epoxide, 10-hydroxy-10,11-dihydrocarbamazepine, 10,11-dihydro-10,11-trans-dihydroxycarbamazepine, 2-hydroxy-carbamazepine, iminostilbene, acridine, and acridone) at three different treatment plants (conventional activated sludge, trickling filter, and stabilization ponds) selected in France. The main aim of this work was to identify selected compounds in wastewater after therapeutic use and to measure concentrations in influents and effluents at the three wastewater treatment plants. Except for iminostilbene, all of these compounds were detected in wastewater. The metabolite common to carbamazepine and oxcarbazepine, i.e., 10,11-dihydro-10,11-trans-dihydroxycarbamazepine, was detected at a higher concentration than the parent substances in wastewater. The presence of parent molecules was noted in inlet and outlet water samples. Carbamazepine, as expected, was not removed by conventional activated sludge treatment. Nevertheless, in a wastewater treatment plant with a 78-day hydraulic retention time, a 73% decrease in carbamazepine concentration was observed. For the first time, oxcarbazepine was found in environmental samples. A decrease in

  16. Urinary Metabolites of Organophosphate and Pyrethroid Pesticides and Behavioral Problems in Canadian Children

    PubMed Central

    Oulhote, Youssef

    2013-01-01

    Background: Exposure to organophosphate pesticides has been associated with neurobehavioral deficits in children, although data on low levels of exposure experienced by the general population are sparse. Pyrethroids are insecticides rapidly gaining popularity, and epidemiological evidence on their potential effects is lacking. Objective: We examined the association between exposure to organophosphate and pyrethroid pesticides, indicated by urinary metabolites, and parentally reported behavioral problems in children. Methods: We used data on children 6–11 years of age from the Canadian Health Measures Survey (2007–2009). We used logistic regressions to estimate odds ratios (ORs) for high scores on the Strengths and Difficulties Questionnaire (SDQ), which may indicate behavioral problems, in association with concentrations of pyrethroid and organophosphate metabolites in the urine of 779 children, adjusting for covariates (sex, age, race/ethnicity, income, parental education, blood lead levels, maternal smoking during pregnancy, and others). Results: At least one urinary metabolite for organophosphates was detected in 91% of children, and for pyrethroids in 97% of children. Organophosphate metabolites were not significantly associated with high SDQ scores. The pyrethroid metabolite cis-DCCA [3-(2,2-dichlorovinyl)-2,2-dimethylycyclopropane carboxylic acid] was significantly associated with high scores for total difficulties on the SDQ (OR for a 10-fold increase = 2.0; 95% CI: 1.1, 3.6), and there was a nonsignificant association with trans-DCCA (OR = 1.6; 95% CI: 0.9, 3.0). Conclusion: In contrast with previous studies, we did not observe an association between exposure to organophosphate pesticides and behavioral scores in children. However, some pyrethroid urinary metabolites were associated with a high level of parent-reported behavioral problems. Longitudinal studies should be conducted on the potential risks of pyrethroids. Citation: Oulhote Y, Bouchard MF

  17. Manual therapy ameliorates delayed-onset muscle soreness and alters muscle metabolites in rats

    PubMed Central

    Urakawa, Susumu; Takamoto, Kouichi; Nakamura, Tomoya; Sakai, Shigekazu; Matsuda, Teru; Taguchi, Toru; Mizumura, Kazue; Ono, Taketoshi; Nishijo, Hisao

    2015-01-01

    Delayed-onset muscle soreness (DOMS) can be induced by lengthening contraction (LC); it can be characterized by tenderness and movement-related pain in the exercised muscle. Manual therapy (MT), including compression of exercised muscles, is widely used as physical rehabilitation to reduce pain and promote functional recovery. Although MT is beneficial for reducing musculoskeletal pain (i.e. DOMS), the physiological mechanisms of MT remain unclear. In the present study, we first developed an animal model of MT in DOMS; LC was applied to the rat gastrocnemius muscle under anesthesia, which induced mechanical hyperalgesia 2–4 days after LC. MT (manual compression) ameliorated mechanical hyperalgesia. Then, we used capillary electrophoresis time-of-flight mass spectroscopy (CE-TOFMS) to investigate early effects of MT on the metabolite profiles of the muscle experiencing DOMS. The rats were divided into the following three groups; (1) normal controls, (2) rats with LC application (LC group), and (3) rats undergoing MT after LC (LC + MT group). According to the CE-TOFMS analysis, a total of 171 metabolites were detected among the three groups, and 19 of these metabolites were significant among the groups. Furthermore, the concentrations of eight metabolites, including branched-chain amino acids, carnitine, and malic acid, were significantly different between the LC + MT and LC groups. The results suggest that MT significantly altered metabolite profiles in DOMS. According to our findings and previous data regarding metabolites in mitochondrial metabolism, the ameliorative effects of MT might be mediated partly through alterations in metabolites associated with mitochondrial respiration. PMID:25713324

  18. Urinary corticosterone metabolite responses to capture and captivity in the cane toad (Rhinella marina).

    PubMed

    Narayan, Edward J; Cockrem, John F; Hero, Jean-Marc

    2011-09-01

    Urinary corticosterone metabolite responses to capture have recently been shown for the first time in amphibians, and in the present study urinary corticosterone metabolite responses to capture and to confinement in captivity were measured in adult cane toads (Rhinella marina) in Queensland, Australia. An adrenocorticotropic hormone (ACTH) challenge was used to provide a biological validation for urinary corticosterone metabolite concentrations measured by radioimmunoassay (RIA). Urinary corticosterone metabolite increased 1-2 days after ACTH but not saline injection and then returned to initial values, indicating that the RIA could detect changes in corticosterone secretion in toads. Urinary corticosterone metabolite responses to short-term capture and restraint in plastic bags were first apparent 2h after capture of wild toads. Toads held communally in captivity for 5 days had elevated urinary corticosterone metabolite concentrations. Mean corticosterone concentrations declined significantly after a further 7 days in individual housing chambers. There was no sex difference in urinary corticosterone metabolite responses of toads to ACTH challenge, short-term capture or captivity. The relative amount of variation in the mean corticosterone responses was quantified by calculating coefficients of variation (CV) for each mean corticosterone response. Mean corticosterone at 0 min was more variable for captive toads than wild toads. Furthermore, initial corticosterone concentrations (0 min) were more variable than concentrations during the ACTH challenge, short-term capture and captivity. There was little change in the amount of variation of mean corticosterone levels between male and female toads with increasing time in captivity (12-29 days). This study has shown individual corticosterone responses of amphibians for the first-time, and has provided a novel method for quantifying the relative amount of variation in amphibian corticosterone responses. PMID:21756910

  19. Measurement of Blood Thiamine Metabolites for Alzheimer's Disease Diagnosis

    PubMed Central

    Pan, Xiaoli; Fei, Guoqiang; Lu, Jingwen; Jin, Lirong; Pan, Shumei; Chen, Zhichun; Wang, Changpeng; Sang, Shaoming; Liu, Huimin; Hu, Weihong; Zhang, Hua; Wang, Hui; Wang, Zhiliang; Tan, Qiong; Qin, Yan; Zhang, Qunying; Xie, Xueping; Ji, Yong; Cui, Donghong; Gu, Xiaohua; Xu, Jun; Yu, Yuguo; Zhong, Chunjiu

    2015-01-01

    Background Brain glucose hypometabolism is an invariant feature and has significant diagnostic value for Alzheimer's disease. Thiamine diphosphate (TDP) is a critical coenzyme for glucose metabolism and significantly reduced in brain and blood samples of patients with Alzheimer's disease (AD). Aims To explore the diagnostic value of the measurement of blood thiamine metabolites for AD. Methods Blood TDP, thiamine monophosphate, and thiamine levels were detected using high performance liquid chromatography (HPLC). The study included the exploration and validation phases. In the exploration phase, the samples of 338 control subjects and 43 AD patients were utilized to establish the models for AD diagnosis assayed by receiver operating characteristic (ROC) curve, including the variable γ that represents the best combination of thiamine metabolites and age to predict the possibility of AD. In the validation phase, the values of models were further tested for AD diagnosis using samples of 861 control subjects, 81 AD patients, 70 vascular dementia patients, and 13 frontotemporal dementia patients. Results TDP and the γ exhibited significant and consistent values for AD diagnosis in both exploration and validation phases. TDP had 0.843 and 0.837 of the areas under ROC curve (AUCs), 77.4% and 81.5% of sensitivities, and 78.1% and 77.2% of specificities respectively in the exploration and validation phases. The γ had 0.938 and 0.910 of AUCs, 81.4% and 80.2% of sensitivities, and 90.5% and 87.2% of specificities respectively in the exploration and validation phases. TDP and the γ can effectively distinguish AD from vascular dementia (64.3% for TDP, 67.1% for γ) and frontotemporal dementia (84.6% for TDP, 100.0% for γ). Interpretation. The measurement of blood thiamine metabolites by HPLC is an ideal diagnostic test for AD with inexpensive, easy to perform, noninvasive merits. PMID:26870826

  20. Human Colon Microbiota Transform Polycyclic Aromatic Hydrocarbons to Estrogenic Metabolites

    PubMed Central

    Van de Wiele, Tom; Vanhaecke, Lynn; Boeckaert, Charlotte; Peru, Kerry; Headley, John; Verstraete, Willy; Siciliano, Steven

    2005-01-01

    Ingestion is an important exposure route for polycyclic aromatic hydrocarbons (PAHs) to enter the human body. Although the formation of hazardous PAH metabolites by human biotransformation enzymes is well documented, nothing is known about the PAH transformation potency of human intestinal microbiota. Using a gastrointestinal simulator, we show that human intestinal microbiota can also bioactivate PAHs, more in particular to estrogenic metabolites. PAH compounds are not estrogenic, and indeed, stomach and small intestine digestions of 62.5 nmol naphthalene, phenanthrene, pyrene, and benzo(a)pyrene showed no estrogenic effects in the human estrogen receptor bioassay. In contrast, colon digests of these PAH compounds displayed estrogenicity, equivalent to 0.31, 2.14, 2.70, and 1.48 nmol 17α-ethynylestradiol (EE2), respectively. Inactivating the colon microbiota eliminated these estrogenic effects. Liquid chromatography–mass spectrometry analysis confirmed the microbial PAH transformation by the detection of PAH metabolites 1-hydroxypyrene and 7-hydroxybenzo(a)pyrene in colon digests of pyrene and benzo(a)pyrene. Furthermore, we show that colon digests of a PAH-contaminated soil (simulated ingestion dose of 5 g/day) displayed estrogenic activity equivalent to 0.58 nmol EE2, whereas stomach or small intestine digests did not. Although the matrix in which PAHs are ingested may result in lower exposure concentrations in the gut, our results imply that the PAH bioactivation potency of colon microbiota is not eliminated by the presence of soil. Moreover, because PAH toxicity is also linked to estrogenicity of the compounds, the PAH bioactivation potency of colon microbiota suggests that current risk assessment may underestimate the risk from ingested PAHs. PMID:15626640

  1. Disposition of xenobiotic chemicals and metabolites in marine organisms.

    PubMed Central

    Varanasi, U; Stein, J E

    1991-01-01

    Studies with several bottom fish species from urban waterways show that of the identified xenobiotic chemicals in bottom sediments, polycylic aromatic hydrocarbons (PAHs) are the most strongly associated with the prevalence of liver lesions, including neoplasms. Accordingly, there is concern about the transfer of contaminants, such as PAHs, from aquatic species to humans. Because PAHs exert their toxicity only after being biotransformed, increasing attention has been focused on the ability of aquatic organisms to metabolize these chemicals. Overall, the results of both laboratory and field studies show that generally low levels (nanograms per gram wet weight) of a few low molecular weight PAHs may be present in edible tissue of fish from contaminated areas and that high molecular weight PAHs, such as the carcinogen benzo(a)pyrene, will rarely be detected because of extensive metabolism. Additionally, the results from a few studies suggest that even though interactions between xenobiotics can affect both biochemical and physiological systems to alter the disposition of PAHs in fish, these interactions do not markedly change the relative proportions of metabolites to parent PAH in tissues. Thus, these studies clearly demonstrate that to obtain some insight into the questions of whether there is any risk to human health from consuming fish and crustaceans from urban areas, techniques must be developed that measure metabolites of carcinogens, such as PAHs, in edible tissue. Initial attempts may focus on semiquantitative methods that permit rapid assessment of the level of metabolites in edible tissues of fish and crustaceans from many urban areas.(ABSTRACT TRUNCATED AT 250 WORDS) Images FIGURE 4. FIGURE 4. FIGURE 4. PMID:2050086

  2. HPLC Determination and MS Confirmation of Malachite Green, Gentian Violet, and Their Leuco Metabolites in Catfish Muscle

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Residues of malachite green (MG), gentian violet (GV), and their leuco metabolites in catfish muscle were individually determined by HPLC using visible and fluorescence detectors. This detection scheme obviated a PbO2 column that converts leuco forms to chromatic forms for visible detection, thus el...

  3. Metabolite Profiling of Sugarcane Genotypes and Identification of Flavonoid Glycosides and Phenolic Acids.

    PubMed

    Coutinho, Isabel D; Baker, John M; Ward, Jane L; Beale, Michael H; Creste, Silvana; Cavalheiro, Alberto J

    2016-06-01

    Sugarcane is an important agricultural crop in the economy of tropical regions, and Brazil has the largest cultivated acreage in the world. Sugarcane accumulates high levels of sucrose in its stalks. Other compounds produced by sugarcane are currently not of economic importance. To explore potential coproducts, we have studied the chemical diversity of sugarcane genotypes, via metabolite profiling of leaves by NMR and LC-DAD-MS. Metabolites were identified via in-house and public databases. From the analysis of 60 HPLC-fractionated extracts, LC-DAD-MS detected 144 metabolites, of which 56 were identified (MS-MS and (1)H NMR), including 19 phenolics and 25 flavones, with a predominance of isomeric flavone C-glycosides. Multivariate analysis of the profiles from genotypes utilized in Brazilian breeding programs revealed clustering according to sugar, phenolic acid, and flavone contents. PMID:27152527

  4. The determination of quizalofop-p-tefuryl, Pantera, and metabolites in soils using GC/MSD

    SciTech Connect

    Parkins, M.D.; Bruns, G.

    1995-12-01

    A published procedure for the analysis of herbicides in soil and sediment was adapted for determining residues of Pantera, quizalofop-p-tefuryl, and its metabolites. Soil is extracted by shaking with a solvent mixture of acetone/0.05N HCL, followed by 0.1 N KOH. The extracts are combined, acidified and then partitioned with methylene chloride. The organic phase is concentrated to 1.0 mL. One half the sample is reacted with diazomethane and analyzed for parent compound and the methyl derivative of the acid metabolite, quizalofop, by GC/MSD with no further workup. The other half is reacted with diazomethane using an elevated temperature, to form volatile methyl derivatives of the metabolites; chlorohydroxyquinoxaline, and chloroquinoxaline phenol. Recoveries were determined at 0.02 ppm, the level of detection, and at 0.20 ppm. The average recovery value for all analytes was greater than 90%.

  5. Utilizing DART Mass Spectrometry to Pinpoint Halogenated Metabolites from a Marine Invertebrate-Derived Fungus

    PubMed Central

    Watts, Katharine R.; Loveridge, Steven T.; Tenney, Karen; Media, Joseph; Valeriote, Frederick A.; Crews, Phillip

    2011-01-01

    Prenylated indole alkaloids are a diverse group of fungal secondary metabolites and represent an important biosynthetic class. In this study we have identified new halogenated prenyl-indole alkaloids from an invertebrate-derived Malbranchea graminicola strain. Using Direct Analysis in Real Time (DART) Mass Spectrometry, these compounds were initially detected from spores of the fungus grown on agar plates, without the need for any organic extraction. Subsequently, the metabolites were isolated from liquid culture in artificial seawater. The structures of two novel chlorinated metabolites, named (−)-spiromalbramide and (+)-isomalbrancheamide B, provide additional insights into the assembly of the malbrancheamide compound family. Remarkably, two new brominated analogs, (+)-malbrancheamide C and (+)-isomalbrancheamide C, were produced by enriching the growth medium with bromine salts. PMID:21682275

  6. Chemosensation of bacterial secondary metabolites modulates neuroendocrine signaling and behavior of C. elegans.

    PubMed

    Meisel, Joshua D; Panda, Oishika; Mahanti, Parag; Schroeder, Frank C; Kim, Dennis H

    2014-10-01

    Discrimination between pathogenic and beneficial microbes is essential for host organism immunity and homeostasis. Here, we show that chemosensory detection of two secondary metabolites produced by Pseudomonas aeruginosa modulates a neuroendocrine signaling pathway that promotes avoidance behavior in the simple animal host Caenorhabditis elegans. Secondary metabolites phenazine-1-carboxamide and pyochelin activate a G-protein-signaling pathway in the ASJ chemosensory neuron pair that induces expression of the neuromodulator DAF-7/TGF-β. DAF-7, in turn, activates a canonical TGF-β signaling pathway in adjacent interneurons to modulate aerotaxis behavior and promote avoidance of pathogenic P. aeruginosa. Our data provide a chemical, genetic, and neuronal basis for how the behavior and physiology of a simple animal host can be modified by the microbial environment and suggest that secondary metabolites produced by microbes may provide environmental cues that contribute to pathogen recognition and host survival. PMID:25303524

  7. Chemosensation of Bacterial Secondary Metabolites Modulates Neuroendocrine Signaling and Behavior of C. elegans

    PubMed Central

    Meisel, Joshua D.; Panda, Oishika; Mahanti, Parag; Schroeder, Frank C.; Kim, Dennis H.

    2014-01-01

    Summary Discrimination among pathogenic and beneficial microbes is essential for host organism immunity and homeostasis. Here, we show that chemosensory detection of two secondary metabolites produced by Pseudomonas aeruginosa modulates a neuroendocrine signaling pathway that promotes avoidance behavior in the simple animal host Caenorhabditis elegans. Secondary metabolites phenazine-1-carboxamide and pyochelin activate a G protein-signaling pathway in the ASJ chemosensory neuron pair that induces expression of the neuromodulator DAF-7/TGF-β. DAF-7, in turn, activates a canonical TGF-β signaling pathway in adjacent interneurons to modulate aerotaxis behavior and promote avoidance of pathogenic P. aeruginosa. Our data provide a chemical, genetic, and neuronal basis for how the behavior and physiology of a simple animal host can be modified by the microbial environment, and suggest that secondary metabolites produced by microbes may provide environmental cues that contribute to pathogen recognition and host survival. PMID:25303524

  8. Metabolite profiling of RCS-4, a novel synthetic cannabinoid designer drug, using human hepatocyte metabolism and TOF-MS

    PubMed Central

    Gandhi, Adarsh S; Zhu, Mingshe; Pang, Shaokun; Wohlfarth, Ariane; Scheidweiler, Karl B; Huestis, Marilyn A

    2014-01-01

    Background Since 2009, scheduling legislation of synthetic cannabinoids prompted new compound emergence to circumvent legal restrictions. 2-(4-methoxyphenyl)-1-(1-pentyl-indol-3-yl)methanone (RCS-4) is a potent cannabinoid receptor agonist sold in herbal smoking blends. Absence of parent synthetic cannabinoids in urine suggests the importance of metabolite identification for detecting RCS-4 consumption in clinical and forensic investigations. Materials & methods & Results With 1 h human hepatocyte incubation and TOF high-resolution MS, we identified 18 RCS-4 metabolites, many not yet reported. Most metabolites were hydroxylated with or without demethylation, carboxylation and dealkylation followed by glucuronidation. One additional sulfated metabolite was also observed. O-demethylation was the most common biotransformation and generated the major metabolite. Conclusion For the first time, we present a metabolic scheme of RCS-4 obtained from human hepatocytes, including Phase I and II metabolites. Metabolite structural information and associated high-resolution mass spectra can be employed for developing clinical and forensic laboratory RCS-4 urine screening methods. PMID:25046048

  9. Production of antioxidant and antitumor metabolites by submerged cultures of Inonotus obliquus cocultured with Phellinus punctatus.

    PubMed

    Zheng, Weifa; Zhao, Yanxia; Zheng, Xin; Liu, Yubing; Pan, Shenyuan; Dai, Yucheng; Liu, Fuming

    2011-01-01

    While Inonotus obliquus produces a diverse range of bioactive metabolites in its natural habitats, it accumulates less in its submerged cultures. We show here that coculture of I. obliquus with Phellinus punctatus resulted in less production of mycelial biomass but an increased accumulation of phenolic compounds, melanins, and lanostane-type triterpenoids. Metabolites increased in production by coculture include phelligridin C, phelligridin H, methyl inoscavin A, inoscavin C, inoscavin B, davallialactone, methyl davallialactone, foscoparianol D, 21,24-cyclopentalanosta-3β,21,25-triol-8-en, lanosta-7,9(11),23-triene-3β,22,25-triol, and inotodisaccharide and melanins. Metabolites from coculture also showed an increased potential for scavenging free radicals and inhibiting the proliferation of HeLa 229 cells. Davallialactone, methyl davallialactone, and minor phenolic components are the major contributors for scavenging DPPH and hydroxyl radical in monoculture, and phelligridin C, phelligridin H, methyl inoscavin A, inoscavin C, methyl davallialactone, foscoparianol D, and inotodisaccharide are those for scavenging the tested radicals in coculture. Lanostane-type triterpenoids indicated limited roles in scavenging free radicals. Nearly all the detected metabolites correlate positively with inhibiting proliferation of HeLa 229 cells. Thus, coculture of I. obliquus with other fungi seems to be a cost-effective strategy for upregulating biosynthesis of bioactive metabolites. PMID:20830471

  10. Mass Spectrometry-Based Metabolite Profiling in the Mouse Liver following Exposure to Ultraviolet B Radiation

    PubMed Central

    Park, Hye Min; Shon, Jong Cheol; Lee, Mee Youn; Liu, Kwang-Hyeon; Kim, Jeong Kee; Lee, Sang Jun; Lee, Choong Hwan

    2014-01-01

    Although many studies have been performed on the effects of ultraviolet (UV) radiation on the skin, only a limited number of reports have investigated these effects on non-skin tissue. This study aimed to describe the metabolite changes in the liver of hairless mice following chronic exposure to UVB radiation. We did not observe significant macroscopic changes or alterations in hepatic cholesterol and triglyceride levels in the liver of UVB-irradiated mice, compared with those for normal mice. In this study, we detected hepatic metabolite changes by UVB exposure and identified several amino acids, fatty acids, nucleosides, carbohydrates, phospholipids, lysophospholipids, and taurine-conjugated cholic acids as candidate biomarkers in response to UVB radiation in the mouse liver by using various mass spectrometry (MS)-based metabolite profiling including ultra-performance liquid chromatography-quadrupole time-of-flight (TOF)-MS, gas chromatography-TOF-MS and nanomate LTQ-MS. Glutamine exhibited the most dramatic change with a 5-fold increase in quantity. The results from altering several types of metabolites suggest that chronic UVB irradiation may impact significantly on major hepatic metabolism processes, despite the fact that the liver is not directly exposed to UVB radiation. MS-based metabolomic approach for determining regulatory hepatic metabolites following UV irradiation will provide a better understanding of the relationship between internal organs and UV light. PMID:25275468

  11. Identification and distribution of four metabolites of geniposide in rats with adjuvant arthritis.

    PubMed

    Chen, Jian; Wu, Hong; Dai, Miao-Miao; Li, Hui; Chen, Jin-Yun; Hu, Shun-Li

    2014-09-01

    Geniposide (GE), also called Jasminoidin, is the major active ingredient of Gardenia jasminoides Ellis (GJ) fruit, which has long been used in traditional Chinese medicine (TCM). Growing evidences suggested that GE has a great potentiality for treating rheumatoid arthritis (RA). However, GE is rapidly metabolized, and we know little about its availability or metabolites in tissues. To elucidate the distribution of GE and its metabolites in tissues, three groups of adjuvant arthritis (AA) rats were given GE (33, 66 and 120 mg/kg) from days 18 to 24, and the biotransformation of GE in plasma, liver, spleen, synovium, urine and mesenteric lymph node (MLN) of rats was investigated by a novel approach named Information-Dependent Acquisition (IDA)-Mediated LC-MS/MS method. As a result, GE and its four major metabolites were detected as follows: GE, G1, G2 in plasma; GE, G2 in MLNs; only GE in liver and synovium; GE, G2, G3 and G4 in spleen; and GE, G1, G2 and G4 in urine. In total four metabolites (G1-G4) involved in the in vivo metabolism processes were identified. The results of this work have demonstrated the IDA-Mediated LC-MS/MS could screen rapidly and reliably the characterization of metabolites from iridoid compounds. PMID:24910002

  12. Sex, stress and social status: patterns in fecal testosterone and glucocorticoid metabolites in male Ethiopian wolves.

    PubMed

    van Kesteren, Freya; Sillero-Zubiri, Claudio; Millar, Robert; Argaw, Kifle; Macdonald, David W; Paris, Monique

    2012-10-01

    Ethiopian wolves, Canis simensis, live in large multi-male family packs, where males are philopatric and do not disperse. Within a pack, mating and breeding is largely monopolized by the dominant male and female, although extra-pack copulations are common, and subordinate males may sire pups in neighboring packs. Regardless of paternity, all males in a pack help rear the pups. We non-invasively studied patterns in fecal testosterone and glucocorticoid metabolite concentrations using radioimmunoassays of fecal samples collected from nine wild male Ethiopian wolves between August 2007 and February 2008. We tested the predictions of the Challenge Hypothesis, namely that fecal testosterone metabolite concentrations would be higher during the annual mating season, which is the portion of the reproductive cycle when mating and increased aggression typically occur, and lower when there were pups in the pack for which to care. Contrary to the predictions of the Challenge Hypothesis, we did not detect patterns in fecal testosterone metabolite concentrations associated with reproductive stage during our study period. Similarly, we found no patterns associated with reproductive stage in male fecal glucocorticoid metabolite concentrations. Dominant males had higher average fecal testosterone and glucocorticoid metabolite concentrations than did subordinates, which may be related to higher rates of aggression and mate guarding in dominant males of group-living canids, a pattern also reported in African wild dogs, Lycaon pictus. PMID:22841807

  13. Metabolite profiling reveals new insights into the regulation of serum urate in humans.

    PubMed

    Albrecht, Eva; Waldenberger, Melanie; Krumsiek, Jan; Evans, Anne M; Jeratsch, Ulli; Breier, Michaela; Adamski, Jerzy; Koenig, Wolfgang; Zeilinger, Sonja; Fuchs, Christiane; Klopp, Norman; Theis, Fabian J; Wichmann, H-Erich; Suhre, Karsten; Illig, Thomas; Strauch, Konstantin; Peters, Annette; Gieger, Christian; Kastenmüller, Gabi; Doering, Angela; Meisinger, Christa

    2014-01-01

    Serum urate, the final breakdown product of purine metabolism, is causally involved in the pathogenesis of gout, and implicated in cardiovascular disease and type 2 diabetes. Serum urate levels highly differ between men and women; however the underlying biological processes in its regulation are still not completely understood and are assumed to result from a complex interplay between genetic, environmental and lifestyle factors. In order to describe the metabolic vicinity of serum urate, we analyzed 355 metabolites in 1,764 individuals of the population-based KORA F4 study and constructed a metabolite network around serum urate using Gaussian Graphical Modeling in a hypothesis-free approach. We subsequently investigated the effect of sex and urate lowering medication on all 38 metabolites assigned to the network. Within the resulting network three main clusters could be detected around urate, including the well-known pathway of purine metabolism, as well as several dipeptides, a group of essential amino acids, and a group of steroids. Of the 38 assigned metabolites, 25 showed strong differences between sexes. Association with uricostatic medication intake was not only confined to purine metabolism but seen for seven metabolites within the network. Our findings highlight pathways that are important in the regulation of serum urate and suggest that dipeptides, amino acids, and steroid hormones are playing a role in its regulation. The findings might have an impact on the development of specific targets in the treatment and prevention of hyperuricemia. PMID:24482632

  14. Synthesis and composition of vitamin D-3 metabolites in Solanum malacoxylon.

    PubMed

    Esparza, M S; Vega, M; Boland, R L

    1982-12-17

    The synthesis of vitamin D-3 hydroxylated metabolites in Solanum malacoxylon was investigated. When crude leaf homogenates and subcellular fractions were incubated with [3H]vitamin D-3 and [3H]25-hydroxy-vitamin D-3 under conditions described for animal vitamin D-3-25-hydroxylase and 25-hydroxy-vitamin D-3-1 alpha-hydroxylase, respectively, labelled metabolites identified on the basis of their chromatographic properties as 25-hydroxy-vitamin D-3 and 1,25-dihydroxy-vitamin D-3 were formed. Other unidentified product metabolites were also detected. Vitamin D-3-25-hydroxylase activity was localized in microsomes and 25-hydroxy-vitamin D-3-1 alpha-hydroxylase in mitochondria and microsomes. Chromatography of sterols isolated from leaf extracts preincubated with beta-glucosidase on Sephadex LH-20 columns permitted the isolation of three biologically active fractions with elution properties similar to vitamin D-3, 25-hydroxy-vitamin D-3 and 1,25-dihydroxy-vitamin D-3, respectively. Ultraviolet spectra characteristic of vitamin D-3 and its metabolites were obtained after purification of the fractions by TLC. Co-chromatography of individual fractions with authentic metabolites on TLC provided further evidence that the plant contains vitamin D-3, 25-hydroxy-vitamin D-3 and 1,25-dihydroxy-vitamin D-3 as glucoside derivatives. These results suggest that a similar pathway of vitamin D-3 hydroxylation as in animals may be operative in S. malacoxylon. PMID:6295509

  15. Determination of hexazinone and its metabolites in groundwater by capillary electrophoresis.

    PubMed

    Kubilius, D T; Bushway, R J

    1998-01-16

    A micellar electrokinetic chromatographic method was developed to separate and quantify hexazinone and metabolites C, A1, E, B and D in groundwater. Hexazinone and its metabolites were extracted from water using Supelclean ENVI-Carb solid-phase extraction tubes. Quantitation was performed using UV photodiode detection at 220, 225, 230 and 247 nm. Intra-assay and inter-assay reproducibility studies run at 0.5, 1.0, 2.0 and 5.0 ppb indicated the procedure was reproducible. Groundwater samples collected from US Geological Survey monitoring wells were analyzed for hexazinone and its metabolites by CE. A comparison was made between CE and an established HPLC method of the hexazinone and metabolite B. The linear regression for hexazinone was y = 1.007x + 0.219 with a correlation coefficient of 0.96 while the linear regression for metabolite B was y = 1.100x-0.057 with a correlation coefficient of 0.91. PMID:9474788

  16. Biliary PAH metabolites and the hepatosomatic index of brown bullheads from Lake Erie tributaries

    USGS Publications Warehouse

    Yang, X.; Baumann, P.C.

    2006-01-01

    In studies designed to investigate the environmental exposure of fish in Lake Erie tributaries, a benthic fish, the brown bullhead (Ameiurus nebulosus), was collected from the industrially contaminated Detroit River, Ottawa River, Black River, Cuyahoga River-harbor and -upstream, Ashtabula River, Buffalo River, and Niagara River, and the non-industrialized Old Woman Creek during 1997-2000. Biliary benzo[a]pyrene (B[a]P)- and naphthalene (NAPH)-type metabolites and the hepatosomatic index (HSI) were measured in fish and compared between different sites. Fish from all of the contaminated sites except Niagara River had significantly higher concentrations of both types of polycyclic aromatic hydrocarbon (PAH) metabolites than fish from the Old Woman Creek. Concentrations of PAH metabolites in bile of fish were positively associated with concentrations of PAHs in sediments, supporting the use of bile metabolites as a measure of PAH exposure. Relatively low concentrations of PAHs detected in fish bile and sediments of the Niagara River, which had undergone extensive remediation, suggested a lowered PAH exposure for fish at this site. No apparent trend was observed in HSI between the industrialized and non-industrialized sites. This study demonstrates that biliary PAH metabolites are an effective indicator of exposure of fish to PAHs. However, because factors other than contamination could also affect the liver size of wild fish, HSI alone may be not a reliable biomarker for assessing contaminant stress. ?? 2005 Elsevier Ltd. All rights reserved.

  17. Application of dried blood spot cards to determine olive oil phenols (hydroxytyrosol metabolites) in human blood.

    PubMed

    de Las Hazas, María Carmen López; Motilva, Maria José; Piñol, Carme; Macià, Alba

    2016-10-01

    In this study, a fast and simple blood sampling and sample pre-treatment method based on the use of the dried blood spot (DBS) cards and ultra-performance liquid chromatography coupled to tandem mass spectrometry (UPLC-MS/MS) for the quantification of olive oil phenolic metabolites in human blood was developed and validated. After validation, the method was applied to determine hydroxytyrosol metabolites in human blood samples after the acute intake of an olive oil phenolic extract. Using the FTA DMPK-A DBS card under optimum conditions, with 20µL as the blood solution volume, 100µL of methanol/Milli-Q water (50/50, v/v) as the extraction solvent and 7 disks punched out from the card, the main hydroxytyrosol metabolites (hydroxytyrosol-3-O-sulphate and hydroxytyrosol acetate sulphate) were identified and quantified. The developed methodology allowed detecting and quantifying the generated metabolites at low μM levels. The proposed method is a significant improvement over existing methods to determine phenolic metabolites circulating in blood and plasma samples, thus making blood sampling possible with the volunteer pricking their own finger, and the subsequent storage of the blood in the DBS cards prior to chromatographic analysis. PMID:27474297

  18. Serum albumin complexation of acetylsalicylic acid metabolites.

    PubMed

    Jurkowski, Wiktor; Porebski, Grzegorz; Obtułowicz, Krystyna; Roterman, Irena

    2009-06-01

    One possible origin of the type I hypersensitivity reaction is reaction of drugs such as acetylsalicylic acid and its metabolites being complexed with human serum albumin. Albumin, being transporting molecule abundant in blood plasma is able to bind large array of ligands varying from small single carbon particles to long hydrophobic tailed lipidic acids (e.g. myristic acid). This non specificity is possible because of multi domain scaffold and large flexibility of inter-domain loops, which results in serious reorientation of domains. Hypothesis that acetylsalicylic acid metabolites may play indirect role in activation of allergic reaction has been tested. Binding of acetylsalicylic acid metabolites in intra-domain space causes significant increase of liability of domains IIIA and IIIB. One of metabolites, salicyluric acid, once is bound causes distortion and partial unfolding of helices in domains IA, IIB and IIIB. Changed are both directions and amplitude of relative motions as well as intra-domain distances. In result albumin is able to cross-link of adjacent IgE receptors which subsequently starts allergic reaction. PMID:19689242

  19. Aspirin-triggered metabolites of EFAs.

    PubMed

    Makriyannis, Alexandros; Nikas, Spyros P

    2011-10-28

    Aspirin triggers the biosynthesis of oxygenated metabolites from arachidonic, eicosapentaenoic, and docosahexaenoic (DHA) acids. In a preceding issue, Serhan et al. (2011) describe a novel aspirin-triggered DHA pathway for the biosynthesis of a potent anti-inflammatory and proresolving molecule. PMID:22035788

  20. Microbial metabolism part 13 metabolites of hesperetin

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The fungal culture, Mucor ramannianus (ATCC 2628) transformed hesperitin to four metabolites: 4'-methoxy -5, 7, 8, 3'-tetrahydroxyflavanone (8-hydroxyhesperetin), 5, 7, 3', 4'-tetrahydroxyflavanone (eriodictyol), 4'-methoxy-5, 3'-dihydroxyflavanone 7-sulfate (hesperetin 7-sulfate) and 5, 7, 3'-tri...

  1. Eleven microbial metabolites of 6-hydroxyflavanone

    Technology Transfer Automated Retrieval System (TEKTRAN)

    6-Hydroxyflavanone (1) when fermented with fungal culture Cunninghamella blakesleeana (ATCC 8688a) yielded flavanone 6-O-ß-D-glucopyranoside (2), flavanone 6-sulfate (3), and 6-hydroxyflavanone 7-sulfate (4). Aspergillus alliaceus (ATCC 10060) also transformed 1 to metabolite 3 as well as 4'-hydrox...

  2. Spatio-Temporal Metabolite Profiling of the Barley Germination Process by MALDI MS Imaging.

    PubMed

    Gorzolka, Karin; Kölling, Jan; Nattkemper, Tim W; Niehaus, Karsten

    2016-01-01

    MALDI mass spectrometry imaging was performed to localize metabolites during the first seven days of the barley germination. Up to 100 mass signals were detected of which 85 signals were identified as 48 different metabolites with highly tissue-specific localizations. Oligosaccharides were observed in the endosperm and in parts of the developed embryo. Lipids in the endosperm co-localized in dependency on their fatty acid compositions with changes in the distributions of diacyl phosphatidylcholines during germination. 26 potentially antifungal hordatines were detected in the embryo with tissue-specific localizations of their glycosylated, hydroxylated, and O-methylated derivates. In order to reveal spatio-temporal patterns in local metabolite compositions, multiple MSI data sets from a time series were analyzed in one batch. This requires a new preprocessing strategy to achieve comparability between data sets as well as a new strategy for unsupervised clustering. The resulting spatial segmentation for each time point sample is visualized in an interactive cluster map and enables simultaneous interactive exploration of all time points. Using this new analysis approach and visualization tool germination-dependent developments of metabolite patterns with single MS position accuracy were discovered. This is the first study that presents metabolite profiling of a cereals' germination process over time by MALDI MSI with the identification of a large number of peaks of agronomically and industrially important compounds such as oligosaccharides, lipids and antifungal agents. Their detailed localization as well as the MS cluster analyses for on-tissue metabolite profile mapping revealed important information for the understanding of the germination process, which is of high scientific interest. PMID:26938880

  3. Profiles of metabolites and gene expression in rats with chemically induced hepatic necrosis.

    PubMed

    Heijne, Wilbert H M; Lamers, Robert-Jan A N; van Bladeren, Peter J; Groten, John P; van Nesselrooij, Joop H J; van Ommen, Ben

    2005-01-01

    This study investigated whether integrated analysis of transcriptomics and metabolomics data increased the sensitivity of detection and provided new insight in the mechanisms of hepatotoxicity. Metabolite levels in plasma or urine were analyzed in relation to changes in hepatic gene expression in rats that received bromobenzene to induce acute hepatic centrilobular necrosis. Bromobenzene-induced lesions were only observed after treatment with the highest of 3 dose levels. Multivariate statistical analysis showed that metabolite profiles of blood plasma were largely different from controls when the rats were treated with bromobenzene, also at doses that did not elicit histopathological changes. Changes in levels of genes and metabolites were related to the degree of necrosis, providing putative novel markers of hepatotoxicity. Levels of endogenous metabolites like alanine, lactate, tyrosine and dimethylglycine differed in plasma from treated and control rats. The metabolite profiles of urine were found to be reflective of the exposure levels. This integrated analysis of hepatic transcriptomics and plasma metabolomics was able to more sensitively detect changes related to hepatotoxicity and discover novel markers. The relation between gene expression and metabolite levels was explored and additional insight in the role of various biological pathways in bromobenzene-induced hepatic necrosis was obtained, including the involvement of apoptosis and changes in glycolysis and amino acid metabolism. The complete Table 2 is available as a supplemental file online at http://taylorandfrancis.metapress.com/openurlasp?genre=journal&issn=0192-6233. To access the file, click on the issue link for 33(4), then select this article. A download option appears at the bottom of this abstract. In order to access the full article online, you must either have an individual subscription or a member subscription accessed through www.toxpath.org. PMID:16036859

  4. Spatio-Temporal Metabolite Profiling of the Barley Germination Process by MALDI MS Imaging

    PubMed Central

    Gorzolka, Karin; Kölling, Jan; Nattkemper, Tim W.; Niehaus, Karsten

    2016-01-01

    MALDI mass spectrometry imaging was performed to localize metabolites during the first seven days of the barley germination. Up to 100 mass signals were detected of which 85 signals were identified as 48 different metabolites with highly tissue-specific localizations. Oligosaccharides were observed in the endosperm and in parts of the developed embryo. Lipids in the endosperm co-localized in dependency on their fatty acid compositions with changes in the distributions of diacyl phosphatidylcholines during germination. 26 potentially antifungal hordatines were detected in the embryo with tissue-specific localizations of their glycosylated, hydroxylated, and O-methylated derivates. In order to reveal spatio-temporal patterns in local metabolite compositions, multiple MSI data sets from a time series were analyzed in one batch. This requires a new preprocessing strategy to achieve comparability between data sets as well as a new strategy for unsupervised clustering. The resulting spatial segmentation for each time point sample is visualized in an interactive cluster map and enables simultaneous interactive exploration of all time points. Using this new analysis approach and visualization tool germination-dependent developments of metabolite patterns with single MS position accuracy were discovered. This is the first study that presents metabolite profiling of a cereals’ germination process over time by MALDI MSI with the identification of a large number of peaks of agronomically and industrially important compounds such as oligosaccharides, lipids and antifungal agents. Their detailed localization as well as the MS cluster analyses for on-tissue metabolite profile mapping revealed important information for the understanding of the germination process, which is of high scientific interest. PMID:26938880

  5. Pesticides and their metabolites in wells of Suffolk County, New York, 1998

    USGS Publications Warehouse

    Phillips, Patrick J.; Eckhardt, D.A.; Terracciano, S.A.; Rosenmann, Larry

    1999-01-01

    Five insecticide residues and 20 herbicide residues were detected in water samples collected from 50 shallow wells screened in the surficial sand and gravel aquifer in Suffolk County, Long Island in areas with known or suspected residues. Laboratory analytical methods with extremely low detection limits - from 0.001 to 0.2 ?g/L (micrograms per liter) - were used to analyze the samples for 60 pesticide residues. Forty-four of the samples contained at least one pesticide residue, and some samples contained as many as 11 different pesticides or pesticide metabolites. Only four water- quality standards were exceeded in the samples collected in this study. Dieldrin exceeded the New York State Class GA standard (0.004 ?g/L) in samples from eight wells. The Federal and New York State Maximum Contaminant Level for simazine (4 ?g/L) was exceeded in samples from two wells, and the State Class GA standard for simazine (0.5 ?g/L) was exceeded in samples from six wells. Federal water-quality standards have not been established for many of the compounds detected in this study, including herbicide metabolites. Maximum concentrations of four herbicide metabolites -metolachlor ESA (ethanesulfonic acid), metolachlor OA (oxanilic acid), and the alachlor metabolites alachlor ESA and alachlor OA -exceeded 20 ?g/L. The maximum concentration of one herbicide (tebuthiuron) exceeded 10 ?g/L, and the maximum concentration of three herbicides (simazine, metolachlor, and atrazine) and one herbicide metabolite (deisopropylatrazine) ranged from 1 to 10 ?g/L. The herbicide metolachlor, which is used on potato fields in Suffolk County, and its metabolites (metolachlor ESA and metolachlor OA) were most frequently detected in samples from agricultural areas. The herbicides simazine and tebuthiuron, which were used in utility rights-of-way, and the simazine metabolite deisopropylatrazine were detected at concentrations greater than 0.05 ?g/L most frequently in samples from residential and mixed land

  6. Analysis of Fusarium avenaceum metabolites produced during wet apple core rot.

    PubMed

    Sørensen, Jens Laurids; Phipps, Richard Kerry; Nielsen, Kristian Fog; Schroers, Hans-Josef; Frank, Jana; Thrane, Ulf

    2009-02-25

    Wet apple core rot (wACR) is a well-known disease of susceptible apple cultivars such as Gloster, Jona Gold, and Fuji. Investigations in apple orchards in Slovenia identified Fusarium avenaceum, a known producer of several mycotoxins, as the predominant causal agent of this disease. A LC-MS/MS method was developed for the simultaneous detection of thirteen F. avenaceum metabolites including moniliformin, acuminatopyrone, chrysogine, chlamydosporol, antibiotic Y, 2-amino-14,16-dimethyloctadecan-3-ol (2-AOD-3-ol), aurofusarin, and enniatins A, A1, B, B1, B2, and B3 from artificially and naturally infected apples. Levels of moniliformin, antibiotic Y, aurofusarin, and enniatins A, A1, B, and B1 were quantitatively examined in artificially inoculated and naturally infected apples, whereas the remaining metabolites were qualitatively detected. Metabolite production was examined in artificially inoculated apples after 3, 7, 14, and 21 days of incubation. Most metabolites were detected after 3 or 7 days and reached significantly high levels within 14 or 21 days. The highest levels of moniliformin, antibiotic Y, aurofusarin, and the combined sum of enniatins A, A1, B, and B1 were 7.3, 5.7, 152, and 12.7 microg g(-1), respectively. Seventeen of twenty naturally infected apples with wACR symptoms contained one or more of the metabolites. Fourteen of these apples contained moniliformin, antibiotic Y, aurofusarin, and enniatins in levels up to 2.9, 51, 167, and 3.9 microg g(-1), respectively. Acuminatopyrone, chrysogine, chlamydosporol, and 2-AOD-3-ol were detected in 4, 11, 4, and 10 apples, respectively. During wet apple core rot, F. avenaceum produced high amounts of mycotoxins, which may pose a risk for consumers of apple or processed apple products. PMID:19170495

  7. Detection of Rare Drug Resistance Mutations by Digital PCR in a Human Influenza A Virus Model System and Clinical Samples

    PubMed Central

    Bushell, Claire A.; Grant, Paul R.; Cowen, Simon; Gutierrez-Aguirre, Ion; O'Sullivan, Denise M.; Žel, Jana; Milavec, Mojca; Foy, Carole A.; Nastouli, Eleni; Garson, Jeremy A.; Huggett, Jim F.

    2015-01-01

    Digital PCR (dPCR) is being increasingly used for the quantification of sequence variations, including single nucleotide polymorphisms (SNPs), due to its high accuracy and precision in comparison with techniques such as quantitative PCR (qPCR) and melt curve analysis. To develop and evaluate dPCR for SNP detection using DNA, RNA, and clinical samples, an influenza virus model of resistance to oseltamivir (Tamiflu) was used. First, this study was able to recognize and reduce off-target amplification in dPCR quantification, thereby enabling technical sensitivities down to 0.1% SNP abundance at a range of template concentrations, a 50-fold improvement on the qPCR assay used routinely in the clinic. Second, a method was developed for determining the false-positive rate (background) signal. Finally, comparison of dPCR with qPCR results on clinical samples demonstrated the potential impact dPCR could have on clinical research and patient management by earlier (trace) detection of rare drug-resistant sequence variants. Ultimately this could reduce the quantity of ineffective drugs taken and facilitate early switching to alternative medication when available. In the short term such methods could advance our understanding of microbial dynamics and therapeutic responses in a range of infectious diseases such as HIV, viral hepatitis, and tuberculosis. Furthermore, the findings presented here are directly relevant to other diagnostic areas, such as the detection of rare SNPs in malignancy, monitoring of graft rejection, and fetal screening. PMID:26659206

  8. Detection of Rare Drug Resistance Mutations by Digital PCR in a Human Influenza A Virus Model System and Clinical Samples.

    PubMed

    Whale, Alexandra S; Bushell, Claire A; Grant, Paul R; Cowen, Simon; Gutierrez-Aguirre, Ion; O'Sullivan, Denise M; Žel, Jana; Milavec, Mojca; Foy, Carole A; Nastouli, Eleni; Garson, Jeremy A; Huggett, Jim F

    2016-02-01

    Digital PCR (dPCR) is being increasingly used for the quantification of sequence variations, including single nucleotide polymorphisms (SNPs), due to its high accuracy and precision in comparison with techniques such as quantitative PCR (qPCR) and melt curve analysis. To develop and evaluate dPCR for SNP detection using DNA, RNA, and clinical samples, an influenza virus model of resistance to oseltamivir (Tamiflu) was used. First, this study was able to recognize and reduce off-target amplification in dPCR quantification, thereby enabling technical sensitivities down to 0.1% SNP abundance at a range of template concentrations, a 50-fold improvement on the qPCR assay used routinely in the clinic. Second, a method was developed for determining the false-positive rate (background) signal. Finally, comparison of dPCR with qPCR results on clinical samples demonstrated the potential impact dPCR could have on clinical research and patient management by earlier (trace) detection of rare drug-resistant sequence variants. Ultimately this could reduce the quantity of ineffective drugs taken and facilitate early switching to alternative medication when available. In the short term such methods could advance our understanding of microbial dynamics and therapeutic responses in a range of infectious diseases such as HIV, viral hepatitis, and tuberculosis. Furthermore, the findings presented here are directly relevant to other diagnostic areas, such as the detection of rare SNPs in malignancy, monitoring of graft rejection, and fetal screening. PMID:26659206

  9. Metabolites of tobacco smoking and colorectal cancer risk.

    PubMed

    Cross, Amanda J; Boca, Simina; Freedman, Neal D; Caporaso, Neil E; Huang, Wen-Yi; Sinha, Rashmi; Sampson, Joshua N; Moore, Steven C

    2014-07-01

    Colorectal cancer is not strictly considered a tobacco-related malignancy, but modest associations have emerged from large meta-analyses. Most studies, however, use self-reported data, which are subject to misclassification. Biomarkers of tobacco exposure may reduce misclassification and provide insight into metabolic variability that potentially influences carcinogenesis. Our aim was to identify metabolites that represent smoking habits and individual variation in tobacco metabolism, and investigate their association with colorectal cancer. In a nested case-control study of 255 colorectal cancers and 254 matched controls identified in the Prostate, Lung, Colorectal and Ovarian cancer screening trial, baseline serum was used to identify metabolites by ultra-high-performance liquid-phase chromatography and mass spectrometry, as well as gas chromatography with tandem mass spectrometry. Odds ratios (OR) and 95% confidence intervals (CI) were estimated by logistic regression. Self-reported current smoking was associated with serum cotinine, O-cresol sulfate and hydroxycotinine. Self-reported current smoking of any tobacco (OR = 1.90, 95% CI: 1.02-3.54) and current cigarette smoking (OR = 1.51, 95% CI: 0.75-3.04) were associated with elevated colorectal cancer risks, although the latter was not statistically significant. Individuals with detectable levels of hydroxycotinine had an increased colorectal cancer risk compared with those with undetectable levels (OR = 2.68, 95% CI: 1.33-5.40). Although those with detectable levels of cotinine had a suggestive elevated risk of this malignancy (OR = 1.81, 95% CI: 0.98-3.33), those with detectable levels of O-cresol sulfate did not (OR = 1.16, 95% CI: 0.57-2.37). Biomarkers capturing smoking behavior and metabolic variation exhibit stronger associations with colorectal cancer than self-report, providing additional evidence for a role for tobacco in this malignancy. PMID:24648381

  10. Persistence of DDT and its metabolites in a farm pond

    USGS Publications Warehouse

    Bridges, W.R.; Kallman, B.J.; Andrews, A.K.

    1963-01-01

    A farm pond near Morrison, Colorado, was treated with 0.02 p.p.m. of DDT in June 1961. The persistence and distribution of the insecticide in materials sampled from the aquatic environment were studied until November 1962. Detectable amounts of DDT were not found in the water after 3 weeks. Residues in the mud had declined within 8 weeks after the treatment to levels not significantly higher than pre-treatment levels, but a sample of vegetation still contained relatively high levels of residues. From this time until the second summer, sufficient vegetation was not present to provide a sample for chemical analysis. A new crop of vegetation sampled 1 year after the treatment contained residues approximating pre-treatment levels. Fish accumulated 3 to 4 p.p.m. of DDT and its metabolites within 1 month after the treatment. The residue levels slowly declined after this, but when the study was terminated, 2 to 3 p.p.m. of the metabolites DDD and DDE still remained in the fish. The highest residue levels measured in crayfish were about one-half of those found in fish. Some mortality of the more susceptible fish and invertebrates occurred as a result of the DDT treatment; however, severe adverse effects were not demonstrated.

  11. METLIN: MS/MS metabolite data from the MAGGIE Project

    DOE Data Explorer

    METLIN is a metabolite database for metabolomics containing over 50,000 structures, it also represents a data management system designed to assist in a broad array of metabolite research and metabolite identification by providing public access to its repository of current and comprehensive MS/MS metabolite data. An annotated list of known metabolites and their mass, chemical formula, and structure are available on the METLIN website. Each metabolite is conveniently linked to outside resources such as the the Kyoto Encyclopedia of Genes and Genomes (KEGG) for further reference and inquiry. MS/MS data is also available on many of the metabolites. The list is expanding continuously as more metabolite information is being deposited and discovered. [from http://metlin.scripps.edu/] Metlin is a component of the MAGGIE Project. MAGGIE is funded by the DOE Genomics: GTL and is an acronym for "Molecular Assemblies, Genes, and Genomics Integrated Efficiently."

  12. ALGAL METABOLITE INFLUENCE ON BLOOM SEQUENCE IN EUTROPHIED FRESHWATER PONDS

    EPA Science Inventory

    The extracellular metabolites of planktonic bloom dominant algae play a most significant role in the determination of bloom sequence in a eutrophied freshwater pond. Certain extracellular metabolites of planktonic blue-green algae substantially inhibit the growth of planktonic di...

  13. Discovering the secondary metabolite potential encoded within Entomopathogenic Fungi

    Technology Transfer Automated Retrieval System (TEKTRAN)

    This article discusses the secondary metabolite potential of the insect pathogens Metarhizium and Beauveria, including a bioinformatics analysis of secondary metabolite genes for which no products are yet identified....

  14. Metabolite Analysis and Histology on the Exact Same Tissue: Comprehensive Metabolomic Profiling and Metabolic Classification of Prostate Cancer.

    PubMed

    Huan, Tao; Troyer, Dean A; Li, Liang

    2016-01-01

    We report a method of metabolomic profiling of intact tissue based on molecular preservation by extraction and fixation (mPREF) and high-performance chemical isotope labeling (CIL) liquid chromatography mass spectrometry (LC-MS). mPREF extracts metabolites by aqueous methanol from tissue biopsies without altering tissue architecture and thus conventional histology can be performed on the same tissue. In a proof-of-principle study, we applied dansylation LC-MS to profile the amine/phenol submetabolome of prostate needle biopsies from 25 patient samples derived from 16 subjects. 2900 metabolites were consistently detected in more than 50% of the samples. This unprecedented coverage allowed us to identify significant metabolites for differentiating tumor and normal tissues. The panel of significant metabolites was refined using 36 additional samples from 18 subjects. Receiver Operating Characteristic (ROC) analysis showed area-under-the-curve (AUC) of 0.896 with sensitivity of 84.6% and specificity of 83.3% using 7 metabolites. A blind study of 24 additional validation samples gave a specificity of 90.9% at the same sensitivity of 84.6%. The mPREF extraction can be readily implemented into the existing clinical workflow. Our method of combining mPREF with CIL LC-MS offers a powerful and convenient means of performing histopathology and discovering or detecting metabolite biomarkers in the same tissue biopsy. PMID:27578275

  15. Metabolite Analysis and Histology on the Exact Same Tissue: Comprehensive Metabolomic Profiling and Metabolic Classification of Prostate Cancer

    PubMed Central

    Huan, Tao; Troyer, Dean A.; Li, Liang

    2016-01-01

    We report a method of metabolomic profiling of intact tissue based on molecular preservation by extraction and fixation (mPREF) and high-performance chemical isotope labeling (CIL) liquid chromatography mass spectrometry (LC-MS). mPREF extracts metabolites by aqueous methanol from tissue biopsies without altering tissue architecture and thus conventional histology can be performed on the same tissue. In a proof-of-principle study, we applied dansylation LC-MS to profile the amine/phenol submetabolome of prostate needle biopsies from 25 patient samples derived from 16 subjects. 2900 metabolites were consistently detected in more than 50% of the samples. This unprecedented coverage allowed us to identify significant metabolites for differentiating tumor and normal tissues. The panel of significant metabolites was refined using 36 additional samples from 18 subjects. Receiver Operating Characteristic (ROC) analysis showed area-under-the-curve (AUC) of 0.896 with sensitivity of 84.6% and specificity of 83.3% using 7 metabolites. A blind study of 24 additional validation samples gave a specificity of 90.9% at the same sensitivity of 84.6%. The mPREF extraction can be readily implemented into the existing clinical workflow. Our method of combining mPREF with CIL LC-MS offers a powerful and convenient means of performing histopathology and discovering or detecting metabolite biomarkers in the same tissue biopsy. PMID:27578275

  16. Characterization of human metabolism and disposition of levo-tetrahydropalmatine: Qualitative and quantitative determination of oxidative and conjugated metabolites.

    PubMed

    Xiao, Weibin; Shen, Guolin; Zhuang, Xiaomei; Ran, Xiaorong; Zhu, Mingshe; Li, Hua

    2016-09-01

    Levo-tetrahydropalmatine (l-THP) is a tetrahydroprotoberberine isoquinoline alkaloid and has been used as an analgesic agent in China for over 50 years. Recent studies revealed that l-THP was effective in the treatment of drug addiction. However, the plasma metabolic profile, mass balance and clearance pathways of l-THP in human remain unknown. In the present study, an analytical strategy was developed for qualitative and quantitative investigation of metabolism and disposition of l-THP in human. Detection and structural characterization of l-THP metabolites were performed using liquid chromatography-quadrupole time-of-flight mass spectrometry. Selected major metabolites in plasma, urine and feces determined by liquid chromatography with UV detection were further quantified using a triple quadruple mass spectrometry and reference standards. A total of 20 metabolites were identified, most of which were formed via demethylation, mono-hydroxylation, and glucuronidation and sulfonation of desmethyl metabolites. Five major metabolites accounted for over 10% of the parent drug concentration in plasma. Major urinary and fecal metabolites and the parent drug that were monitored for 72h accounted for 46.3% of the dose, while only 0.16% of the dose was the unchanged drug. Multiple demethylations followed by glucuronide and sulfate conjugations and renal excretion were the major drug clearance pathways of l-THP in human. PMID:27343900

  17. Identification of the urinary metabolites of 4-bromoaniline and 4-bromo-[carbonyl-13C]-acetanilide in rat.

    PubMed

    Scarfe, G B; Nicholson, J K; Lindon, J C; Wilson, I D; Taylor, S; Clayton, E; Wright, B

    2002-04-01

    1. The urinary excretion of 4-bromoaniline and its [carbonyl-(13)C]-labelled N-acetanilide, together with their corresponding metabolites, have been investigated in the rat following i.p. administration at 50 mg kg(-1). 2. Metabolite profiling was performed by reversed-phase HPLC with UV detection, whilst identification was performed using a combination of enzymic hydrolysis and directly coupled HPLC-NMR-MS analysis. The urinary metabolite profile was quantitatively and qualitatively similar for both compounds with little of either excreted unchanged. 3. The major metabolite present in urine was 2-amino-5-bromophenylsulphate, but, in addition, a number of metabolites with modification of the N-acetyl moiety were identified (from both the [(13)C]-acetanilide or produced following acetylation of the free bromoaniline). 4. For 4-bromoacetanilide, N-deacetylation was a major route of metabolism, but despite the detection of the acetanilide following the administration of the free aniline, there was no evidence of reacetylation (futile deacetylation). 5. Metabolites resulting from the oxidation of the acetyl group included a novel glucuronide of an N-glycolanilide, an unusual N-oxanilic acid and a novel N-acetyl cysteine conjugate. PMID:12028665

  18. Activation and Products of the Cryptic Secondary Metabolite Biosynthetic Gene Clusters by Rifampin Resistance (rpoB) Mutations in Actinomycetes

    PubMed Central

    Tanaka, Yukinori; Kasahara, Ken; Hirose, Yutaka; Murakami, Kiriko; Kugimiya, Rie

    2013-01-01

    A subset of rifampin resistance (rpoB) mutations result in the overproduction of antibiotics in various actinomycetes, including Streptomyces, Saccharopolyspora, and Amycolatopsis, with H437Y and H437R rpoB mutations effective most frequently. Moreover, the rpoB mutations markedly activate (up to 70-fold at the transcriptional level) the cryptic/silent secondary metabolite biosynthetic gene clusters of these actinomycetes, which are not activated under general stressful conditions, with the exception of treatment with rare earth elements. Analysis of the metabolite profile demonstrated that the rpoB mutants produced many metabolites, which were not detected in the wild-type strains. This approach utilizing rifampin resistance mutations is characterized by its feasibility and potential scalability to high-throughput studies and would be useful to activate and to enhance the yields of metabolites for discovery and biochemical characterization. PMID:23603745

  19. DnsID in MyCompoundID for rapid identification of dansylated amine- and phenol-containing metabolites in LC-MS-based metabolomics.

    PubMed

    Huan, Tao; Wu, Yiman; Tang, Chenqu; Lin, Guohui; Li, Liang

    2015-10-01

    High-performance chemical isotope labeling (CIL) liquid chromatography-mass spectrometry (LC-MS) is an enabling technology based on rational design of labeling reagents to target a class of metabolites sharing the same functional group (e.g., all the amine-containing metabolites or the amine submetabolome) to provide concomitant improvements in metabolite separation, detection, and quantification. However, identification of labeled metabolites remains to be an analytical challenge. In this work, we describe a library of labeled standards and a search method for metabolite identification in CIL LC-MS. The current library consists of 273 unique metabolites, mainly amines and phenols that are individually labeled by dansylation (Dns). Some of them produced more than one Dns-derivative (isomers or multiple labeled products), resulting in a total of 315 dansyl compounds in the library. These metabolites cover 42 metabolic pathways, allowing the possibility of probing their changes in metabolomics studies. Each labeled metabolite contains three searchable parameters: molecular ion mass, MS/MS spectrum, and retention time (RT). To overcome RT variations caused by experimental conditions used, we have developed a calibration method to normalize RTs of labeled metabolites using a mixture of RT calibrants. A search program, DnsID, has been developed in www.MyCompoundID.org for automated identification of dansyl labeled metabolites in a sample based on matching one or more of the three parameters with those of the library standards. Using human urine as an example, we illustrate the workflow and analytical performance of this method for metabolite identification. This freely accessible resource is expandable by adding more amine and phenol standards in the future. In addition, the same strategy should be applicable for developing other labeled standards libraries to cover different classes of metabolites for comprehensive metabolomics using CIL LC-MS. PMID:26327437

  20. Determination of metabolites of cytochrome P-450 model systems using high-performance liquid chromatography.

    PubMed

    Esclade, L; Guillochon, D; Thomas, D

    1985-06-14

    High-performance liquid chromatographic techniques were developed for the simultaneous detection of metabolites in a cytochrome P-450 model system composed of NADH, haemoglobin and methylene blue. Monohydroxylated metabolites were determined following aniline, acetanilide and phenol hydroxylations. 4-Aminoantipyrine, 7-hydroxycoumarin and p-nitrophenol were determined after dealkylation of 4-N,N-dimethylamino-antipyrine, 7-ethoxycoumarin and p-nitroanisole. These substrates are commonly used for measuring cytochrome P-450 activities. Treatment of the samples was minimal, consisting of a simple deproteinization, and did not involve any organic extraction. Separations were carried out on reversed-phase columns and the products were detected by UV adsorption. Separations were completed in less than 15 min and the detection limits were between 0.5 and 4 microM. PMID:3875625

  1. Identification and characterization of in vivo metabolites of asulacrine using advanced mass spectrophotometry technique in combination with improved data mining strategy.

    PubMed

    Afzal, Attia; Zhong, Yunxi; Sarfraz, Muhammad; Peng, Ying; Sheng, Longsheng; Wu, Zimei; Sun, Jianguo; Wang, Guangji

    2016-04-29

    Asulacrine (ASL) is a broad-spectrum, antitumor drug whose data are promising for the treatment of breast and lung cancers; however, a high incidence of phlebitis hampered its further development. Phlebitis is associated with generation of reactive species. Asulacrine donates electrons and produces oxidative stress in chemical reactions. It was expected that ASL would actively metabolize to oxidized products through reactive intermediates and produce more products in vivo than reported and thus cause phlebitis. A comprehensive study was planned to investigate in vivo metabolism of ASL, using high-resolution mass spectrometry LC/IT-TOF MS in positive mode. Metabolites were detected by different software by applying annotated detection strategy. The possible metabolites and their product ions were simultaneously detected by segmented data acquisition to get accurate mass values. Segmented data acquisition improved signal-to-noise (S/N) ratio, which was helpful to detect metabolites and their fragments even when present in trace amounts. A total of 21 metabolites were detected in gender-based biological fluids and characterized by comparing their accurate mass values, fragmentation patterns, and relative retention times with that of ASL. Among previously reported glucuronosylation metabolites, some oxidation, hydroxylation, carboxylation, demethylation, hydrogenation, glutamination, and acetylcysteine conjugation were detected for the first time. Twenty metabolites were tentatively identified by using the annotated strategy for data acquisition and post-data mining. PMID:27040513

  2. Rapidly improved determination of metabolites from biological data sets using the high-efficient TransOmics tool.

    PubMed

    Zhang, Aihua; Zhou, Xiaohang; Zhao, Hongwei; Guan, Yu; Zou, Shiyu; Zhou, Shiyu; Yan, Guang-li; Ma, Chung Wah; Ma, Zhonghua; Liu, Qi; Wang, Xijun

    2014-08-01

    Metabolomics is a new approach based on the systematic study of the full complement of metabolites in a biological sample. Extracting biomedical information from large datasets is of considerable complexity. Furthermore, the traditional method of assessing metabolomics data is not only time-consuming but it is often subjective work. Here we used sensitive ultra-performance LC-ESI/Q-TOF high-definition mass spectrometry (UPLC-ESI-Q-TOF-MS) in positive ion mode coupled with a new developed software program TransOmics for widely untargeted metabolomics, which incorporates novel nonlinear alignment, deconvolution, matched filtration, peak detection, and peak matching to characterize metabolites as a case study. The TransOmics method can facilitate prioritization of the data and greatly increase the probability of identifying metabolites related to the phenotype of interest. By this means, 17 urinary differential metabolites were identified (less than 10 min) involving the key metabolic pathways including tyrosine metabolism, glutathione metabolism, phenylalanine metabolism, ascorbate and aldarate metabolism, arginine and proline metabolism, and so forth. Metabolite identification has also been significantly improved, using the correlation peak patterns in contrast to a reference metabolite panel. It can detect and identify metabolites automatically and remove background noise, and also provides a user-friendly graphical interface to apply principal component analyses, correlation analysis and compound statistics. This investigation illustrates that metabolomics combined with the proposed bioinformatic approach (based on TransOmics) is important to elucidate the developing biomarkers and the physiological mechanism of disease, and has opened the door for the development of a new genre of metabolite identification methods. PMID:24889752

  3. Identification of in vitro metabolites of ethylphenidate by liquid chromatography coupled to quadrupole time-of-flight mass spectrometry.

    PubMed

    Negreira, Noelia; Erratico, Claudio; van Nuijs, Alexander L N; Covaci, Adrian

    2016-01-01

    Ethylphenidate is a new potent synthetic psychoactive drug, structurally related to methylphenidate. Using human liver microsomes and cytosol, we have investigated for the first time the Phase-I and Phase-II in vitro metabolism of ethylphenidate. The structure of the metabolites was elucidated by hybrid quadrupole time-of-flight mass spectrometry. Overall, seven Phase-I, but no Phase-II metabolites were detected. Ethylphenidate underwent hydroxylation forming two primary mono-hydroxylated metabolites and, subsequently, dehydration and ring opening with an additional hydroxylation, forming secondary metabolites. The involvement of different human cytochrome P450 (CYP) enzymes in the formation of ethylphenidate metabolites was investigated using a panel of human recombinant CYPs (rCYPs). rCYP2C19 was the most active recombinant enzyme involved in the formation of all seven ethylphenidate metabolites detected, although other rCYPs (rCYP1A2, rCYP2B6, rCYPC9, rCYP2D6, and rCYP3A4, but not rCYP2E1) played a role in the metabolism of ethylphenidate. All metabolites identified in the present study can be considered as potential specific biomarkers of ethylphenidate in toxicological studies. Additionally, ritalinic acid and methylphenidate were formed by non-enzymatic hydrolysis and trans-esterification, and, therefore, they cannot be considered as (oxidative) metabolites of ethylphenidate. The presence of methylphenidate and ritalinic acid cannot be exclusively associated to the use of ethylphenidate, since methylphenidate is a drug itself and ritanilic acid can be formed from both ethylphenidate and methylphenidate. PMID:26454340

  4. Widespread occurrence of neuro-active pharmaceuticals and metabolites in 24 Minnesota rivers and wastewaters

    USGS Publications Warehouse

    Writer, Jeffrey; Ferrer, Imma; Barber, Larry B.; Thurman, E. Michael

    2013-01-01

    Concentrations of 17 neuro-active pharmaceuticals and their major metabolites (bupropion, hydroxy-bupropion, erythro-hydrobupropion, threo-hydrobupropion, carbamazepine, 10,11,-dihydro-10,11,-dihydroxycarbamazepine, 10-hydroxy-carbamazepine, citalopram, N-desmethyl-citalopram, fluoxetine, norfluoxetine, gabapentin, lamotrigine, 2-N-glucuronide-lamotrigine, oxcarbazepine, venlafaxine and O-desmethyl-venlafaxine), were measured in treated wastewater and receiving surface waters from 24 locations across Minnesota, USA. The analysis of upstream and downstream sampling sites indicated that the wastewater treatment plants were the major source of the neuro-active pharmaceuticals and associated metabolites in surface waters of Minnesota. Concentrations of parent compound and the associated metabolite varied substantially between treatment plants (concentrations ± standard deviation of the parent compound relative to its major metabolite) as illustrated by the following examples; bupropion and hydrobupropion 700 ± 1000 ng L−1, 2100 ± 1700 ng L−1, carbamazepine and 10-hydroxy-carbamazepine 480 ± 380 ng L−1, 360 ± 400 ng L−1, venlafaxine and O-desmethyl-venlafaxine 1400 ± 1300 ng L−1, 1800 ± 2300 ng L−1. Metabolites of the neuro-active compounds were commonly found at higher or comparable concentrations to the parent compounds in wastewater effluent and the receiving surface water. Neuro-active pharmaceuticals and associated metabolites were detected only sporadically in samples upstream from the effluent outfall. Metabolite to parent ratios were used to evaluate transformation, and we determined that ratios in wastewater were much lower than those reported in urine, indicating that the metabolites are relatively more labile than the parent compounds in the treatment plants and in receiving waters. The widespread occurrence of neuro-active pharmaceuticals and metabolites in Minnesota effluents and surface waters indicate that

  5. Noninvasive quantitation of phosphorus metabolites in human tissue by NMR spectroscopy

    NASA Astrophysics Data System (ADS)

    Roth, K.; Hubesch, B.; Meyerhoff, D. J.; Naruse, S.; Gober, J. R.; Lawry, T. J.; Boska, M. D.; Matson, G. B.; Weiner, M. W.

    Quantitation of metabolite concentrations by NMR spectroscopy is complicated by the need to determine the volume from which signals are detected, and by the need to obtain the relative sensitivity of detection within this volume. The use of coils with inhomogeneous B1 fields further complicates these problems. In order to quantify metabolite concentrations using 31P NMR spectroscopy, an external reference of hexamethyl phosphoroustriamide was used. Studies were performed on phantoms, using either a surface coil or a Helmholtz head coil to confirm the accuracy of both the ISIS volume selection technique and the use of an external reference. The limitations of this method are related to contamination and signal loss inherent in the ISIS technique and difficulties with integration of broad overlapping peaks. The method was applied to seven normal human subjects. The integrals for metabolite signals in normal brain and calf muscle were determined by using NMRI software. The T1 values of the signals of all phosphorus metabolites in the selected volume were measured in order to correct for saturation effects. The concentrations for PCr, P i, and ATP were 4.9, 2.0, and 2.5 m M in brain and 36.5, 5.7, and 7.3 m M in muscle. These results are in good agreement with those reported for animals, demonstrating the validity of this quantitation technique.

  6. Facilitated uptake of a bioactive metabolite of maritime pine bark extract (pycnogenol) into human erythrocytes.

    PubMed

    Kurlbaum, Max; Mülek, Melanie; Högger, Petra

    2013-01-01

    Many plant secondary metabolites exhibit some degree of biological activity in humans. It is a common observation that individual plant-derived compounds in vivo are present in the nanomolar concentration range at which they usually fail to display measurable activity in vitro. While it is debatable that compounds detected in plasma are not the key effectors of bioactivity, an alternative hypothesis may take into consideration that measurable concentrations also reside in compartments other than plasma. We analysed the binding of constituents and the metabolite δ-(3,4-dihydroxy-phenyl)-γ-valerolactone (M1), that had been previously detected in plasma samples of human consumers of pine bark extract Pycnogenol, to human erythrocytes. We found that caffeic acid, taxifolin, and ferulic acid passively bind to red blood cells, but only the bioactive metabolite M1 revealed pronounced accumulation. The partitioning of M1 into erythrocytes was significantly diminished at higher concentrations of M1 and in the presence of glucose, suggesting a facilitated transport of M1 via GLUT-1 transporter. This concept was further supported by structural similarities between the natural substrate α-D-glucose and the S-isomer of M1. After cellular uptake, M1 underwent further metabolism by conjugation with glutathione. We present strong indication for a transporter-mediated accumulation of a flavonoid metabolite in human erythrocytes and subsequent formation of a novel glutathione adduct. The physiologic role of the adduct remains to be elucidated. PMID:23646194

  7. Expanded separation technique for chlorophyll metabolites in Oriental tobacco leaf using non aqueous reversed phase chromatography.

    PubMed

    Ishida, Naoyuki

    2011-08-26

    An improved separation method for chlorophyll metabolites in Oriental tobacco leaf was developed. While Oriental leaf still gives the green color even after the curing process, little attention has been paid to the detailed composition of the remaining green pigments. This study aimed to identify the green pigments using non aqueous reversed phase chromatography (NARPC). To this end, liquid chromatograph (LC) equipped with a photo diode array detector (DAD) and an atmospheric pressure chemical ionization/mass spectrometer (APCI/MSD) was selected, because it is useful for detecting low polar non-volatile compounds giving green color such as pheophytin a. Identification was based on the wavelength spectrum, mass spectrum and retention time, comparing the analytes in Oriental leaf with the commercially available and synthesized components. Consequently, several chlorophyll metabolites such as hydroxypheophytin a, solanesyl pheophorbide a and solanesyl hydroxypheophorbide a were newly identified, in addition to typical green pigments such as chlorophyll a and pheophytin a. Chlorophyll metabolites bound to solanesol were considered the tobacco specific components. NARPC expanded the number of detectable low polar chlorophyll metabolites in Oriental tobacco leaf. PMID:21782189

  8. MALDI-Mass Spectrometric Imaging for the Investigation of Metabolites in Medicago truncatula Root Nodules

    PubMed Central

    Gemperline, Erin; Li, Lingjun

    2014-01-01

    Most techniques used to study small molecules, such as pharmaceutical drugs or endogenous metabolites, employ tissue extracts which require the homogenization of the tissue of interest that could potentially cause changes in the metabolic pathways being studied1. Mass spectrometric imaging (MSI) is a powerful analytical tool that can provide spatial information of analytes within intact slices of biological tissue samples1-5. This technique has been used extensively to study various types of compounds including proteins, peptides, lipids, and small molecules such as endogenous metabolites. With matrix-assisted laser desorption/ionization (MALDI)-MSI, spatial distributions of multiple metabolites can be simultaneously detected. Herein, a method developed specifically for conducting untargeted metabolomics MSI experiments on legume roots and root nodules is presented which could reveal insights into the biological processes taking place. The method presented here shows a typical MSI workflow, from sample preparation to image acquisition, and focuses on the matrix application step, demonstrating several matrix application techniques that are useful for detecting small molecules. Once the MS images are generated, the analysis and identification of metabolites of interest is discussed and demonstrated. The standard workflow presented here can be easily modified for different tissue types, molecular species, and instrumentation. PMID:24637669

  9. Identification, quantification, and relative concentrations of carotenoids and their metabolites in human milk and serum.

    PubMed

    Khachik, F; Spangler, C J; Smith, J C; Canfield, L M; Steck, A; Pfander, H

    1997-05-15

    Thirty-four carotenoids, including 13 geometrical isomers and eight metabolites, in breast milk and serum of three lactating mothers have been separated, identified, quantified, and compared by high-performance liquid chromatography (HPLC)-photodiode array (PDA) detection-mass spectrometry (MS). Among the metabolites were two oxidation products of lycopene and four of lutein/ zeaxanthin. In addition, two metabolites of lutein, formed as a result of dehydration of this dihydroxycarotenoid under acidic conditions similar to those of the stomach, have also been identified in plasma and breast milk. The oxidative metabolites of lycopene with a novel five-membered-ring end group have been identified as epimeric 2,6-cyclolycopene-1,5-diols by comparison of their HPLC-UV/visible-MS profiles with those of fully characterized (1H- and 13C-NMR spectroscopy) synthetic compounds. The HPLC procedures employed also detected vitamin A, two forms of vitamin E (gamma- and alpha-tocopherol), and two non-carotenoid food components, i.e., piperine and caffeine, in serum and breast milk. PMID:9164160

  10. Alkylphenol Polyethoxylate Metabolite Behavior During Short-Term Soil Aquifer Treatment

    NASA Astrophysics Data System (ADS)

    Reinhard, M.

    2002-12-01

    The attenuation of alkylphenol polyethoxylate (APEO) metabolites was studied at a soil aquifer treatment (SAT) facility located in Mesa, Arizona, USA. SAT is a technique commonly used in arid environments to augment groundwater supplies. In SAT, municipal wastewater is discharged into basins and allowed to infiltrate into the subsurface; the basins are most often filled for several days and then allowed to dry out. During SAT the quality of the recharged water is substantially improved. Because this water may eventually be used to augment drinking water supplies, there is a concern whether organic contaminants survive SAT. APEO metabolites are among the most frequently detected anthropogenic contaminants in the environment. The ubiquitous presence of these compounds may be of concern because they are relatively recalcitrant, can sorb and accumulate in soils and sediments, can bioaccumulate in plants and animals, and can be estrogenic to wildlife at low concentrations. In this study, two parcels of water were monitored during SAT -- one aerobic, the other anaerobic. During infiltration, under aerobic and anaerobic conditions, both alkylphenol ethoxycarboxylates (APECs) and carboxyalkylphenol ethoxycarboxylates (CAPECs) were substantially attenuated (> 90%) within 3 m. As expected, nonylphenol was removed under aerobic conditions, but produced under anaerobic conditions. Interestingly, no short-chained APEOs were detected. The rapid attenuation of CAPECs was surprising, as other researchers have found these metabolites to be very persistent. During infiltration, APEO metabolites with the longest ethoxycarboxylate side chain are attenuated fastest. Unlike several recent studies, alkylphenoxyacetic acids (AP1ECs) and carboxyalkylphenoxyacetic acids (CAP1ECs) were almost twice as abundant as alkylphenoxyethoxyacetic acids (AP2ECs) and carboxyalkylphenoxyethoxyacetic acids (CAP2ECs). Nonylphenol concentrations in both the wastewater and effluent SAT water were > 10

  11. Nitric oxide metabolites in patients with asthma: induced sputum versus blood.

    PubMed

    Jang, A S; Choi, I S

    1999-12-01

    Nitric oxide (NO) plays an important role in physiological regulation of the airways. The monitoring of airway inflammation has being observed in bronchial asthma directly, by sputum examination, and indirectly, by measurements in peripheral blood. To investigate the diagnostic value of these two methods, we compared NO metabolites in induced sputum and serum obtained in patients with asthma and control subjects. Hypertonic saline induced sputum and serum were obtained in 13 patients with asthma and 10 control subjects. NO metabolite level was assayed by using modified Griess reaction. Eosinophil cationic protein (ECP) was measured by fluoroimmunoassay, and detected interleukin (IL)-5 by a sandwich ELISA. The accuracy of the tests was measured by plotting the data in receiver operating characteristic (ROC) curves and comparing the area under the curve for NO metabolites. Asthmatic patients, compared with control subjects, had significantly higher NO metabolites in induced sputum (1252.5+/-203.3 mol l(-1) vs. 557.2+/-101.5 mol l(-1), P<0.01) but not in serum. IL-5 in induced sputum was detected more frequently in patients with asthma than in control subjects [11/13 (84.6%) vs. 1/10 (10%), P<0.01]. Asthmatic patients, compared with control subjects, had significantly higher ECP concentration in induced sputum (1270.0+/-197.9 g l vs. 154.6+/-47.4 g l(-1), P<0.01). There were significant positive correlations between NO metabolites in induced sputum and eosinophils, ECP in induced sputum (r=0.58 P<0.05; r=0.64, P<0.01) in patients with asthma but not in serum. The area under the ROC curve showed that NO metabolites in induced sputum (0.78) are more accurate marker than NO metabolites in serum (0.53) (P<0.05). These findings suggest that NO metabolites in induced sputum is a more valuable indicator to monitor asthmatic airway inflammation than those in serum. PMID:10653055

  12. Gastrointestinal distribution of albendazole metabolites following netobimin administration to cattle: relationship with plasma disposition kinetics.

    PubMed

    Lanusse, C E; Gascon, L H; Prichard, R K

    1993-03-01

    The gastrointestinal (GI) distribution and plasma disposition kinetics of albendazole (ABZ) metabolites after oral administration of netobimin (NTB) to cattle were studied. Eight Holstein steers (150-180 kg) were surgically fitted with permanent cannulae in the rumen, abomasum and ileum. After post-surgical recovery, the animals were treated orally with a suspension of netobimin zwitterion (400 mg/ml) at 20 mg/kg. Jugular blood and ruminal, abomasal and ileal fluid samples were taken serially over a 96 h period and analysed by HPLC for NTB and its metabolites, including ABZ, ABZ sulphoxide (ABZSO), ABZ sulphone (ABZSO2) and amino-albendazole sulphone (NH2ABZSO2). NTB parent drug was only found in the GI tract and for only 12-18 h post-treatment. ABZSO and ABZSO2 were the main metabolites found in plasma, being present for 30-36 h. These metabolites were exchanged between plasma and different GI fluids and were greatly concentrated in the abomasum. This phenomenon may account for the presence of ABZ, ABZSO and ABZSO2 in the GI tract for 72 h post-treatment despite the fact that ABZ was not detected in plasma and ABZSO and ABZSO2 were detected for only 30-36 h in plasma. The presence of ABZ and ABZSO in the abomasum and intestine for this extended period of time is probably relevant for anthelmintic efficacy against GI parasites. The NH2 ABZSO2 metabolite was detected in plasma, abomasum and ileum and its disposition kinetics were characterized for the first time. PMID:8478999

  13. Secondary Metabolites of Hypericum leptophyllum Hochst., an Endemic Turkish Species

    PubMed Central

    Camas, Necdet; Radusiene, Jolita; Stanius, Zydrunas; Caliskan, Omer; Cirak, Cuneyt

    2012-01-01

    In the present study, the presence of the phloroglucinol derivative hyperforin, the naphthodianthrones hypericin and pseudohypericin, the phenylpropane chlorogenic acid and the flavonoids rutin, hyperoside, kaempferol, isoquercetine, quercitrine, and quercetine was investigated in Hypericum leptophyllum Hochst., an endemic Turkish species for the first time. The aerial parts representing a total of 30 individuals were collected at full flowering and dissected into floral, leaf, and stem tissues. After being dried at room temperature, the plant materials were assayed for secondary metabolite concentrations by HPLC. Aerial plant parts accumulated chlorogenic acid, hyperoside, isoquercetine, quercitrine, and quercetine, but they did not accumulate hyperforin, hypericin, pseudohypericin, rutin, and kaempferol. Accumulation levels of the detected compounds varied with plant tissues. Such kind of data could be useful for elucidation of the chemotaxonomical significance of the corresponding compounds and phytochemical evaluation of this endemic species. PMID:22649295

  14. Secondary metabolites of Hypericum leptophyllum Hochst., an endemic Turkish species.

    PubMed

    Camas, Necdet; Radusiene, Jolita; Stanius, Zydrunas; Caliskan, Omer; Cirak, Cuneyt

    2012-01-01

    In the present study, the presence of the phloroglucinol derivative hyperforin, the naphthodianthrones hypericin and pseudohypericin, the phenylpropane chlorogenic acid and the flavonoids rutin, hyperoside, kaempferol, isoquercetine, quercitrine, and quercetine was investigated in Hypericum leptophyllum Hochst., an endemic Turkish species for the first time. The aerial parts representing a total of 30 individuals were collected at full flowering and dissected into floral, leaf, and stem tissues. After being dried at room temperature, the plant materials were assayed for secondary metabolite concentrations by HPLC. Aerial plant parts accumulated chlorogenic acid, hyperoside, isoquercetine, quercitrine, and quercetine, but they did not accumulate hyperforin, hypericin, pseudohypericin, rutin, and kaempferol. Accumulation levels of the detected compounds varied with plant tissues. Such kind of data could be useful for elucidation of the chemotaxonomical significance of the corresponding compounds and phytochemical evaluation of this endemic species. PMID:22649295

  15. Comparative pharmacokinetics of netobimin metabolites in pregnant ewes.

    PubMed

    Cristòfol, C; Franquelo, C; Navarro, M; Carretero, A; Ruberte, J; Arboix, M

    1997-01-01

    The pharmacokinetics of netobimin (NTB) metabolites has been investigated in ewes. Non-pregnant ewes and ewes in the first and last third of pregnancy were dosed orally with 20 mg kg bodyweight of NTB. Blood samples were collected from the jugular vein from 30 minutes to 72 hours after administration and plasma samples were analysed by high performance liquid chromatography. Neither NTB nor albendazole (ABZ) were detected in any of the samples analysed. No statistically significant differences were found between the pharmacokinetic parameters of albendazole suphoxide (ABZSO) and albendazole sulphone (ABZSO2) among the three groups of ewes. The peak plasma concentrations (Cmax) ABZSO and ABZSO2 were reached about 10 and 20 hours respectively after administration in all three groups. The ratios of ABZSO/ABZSO2 for Cmax and the areas under the curve (AUCzero-infinity) were 6 and 3, respectively, in each group and suggest a low rate of oxidation of sulfoxide to sulphone. PMID:9243708

  16. SMURF: genomic mapping of fungal secondary metabolite clusters

    PubMed Central

    Khaldi, Nora; Seifuddin, Fayaz T.; Turner, Geoff; Haft, Daniel; Nierman, William C.; Wolfe, Kenneth H.; Fedorova, Natalie D.

    2010-01-01

    Fungi produce an impressive array of secondary metabolites (SMs) including mycotoxins, antibiotics and pharmaceuticals. The genes responsible for their biosynthesis, export, and transcriptional regulation are often found in contiguous gene clusters. To facilitate annotation of these clusters in sequenced fungal genomes, we developed the web-based software SMURF (www.jcvi.org/smurf/) to systematically predict clustered SM genes based on their genomic context and domain content. We applied SMURF to catalog putative clusters in 27 publicly available fungal genomes. Comparison with genetically characterized clusters from six fungal species showed that SMURF accurately recovered all clusters and detected additional potential clusters. Subsequent comparative analysis revealed the striking biosynthetic capacity and variability of the fungal SM pathways and the correlation between unicellularity and the absence of SMs. Further genetics studies are needed to experimentally confirm these clusters. PMID:20554054

  17. An In Vitro Control Mechanism for Potato Stress Metabolite Biosynthesis

    PubMed Central

    Alves, Leo M.; Kalan, Edwin B.; Heisler, Edward G.

    1981-01-01

    Ethylene/oxygen (E/O2) elevates sesquiterpenoid stress metabolite (SSM) levels in potato (Solanum tuberosum L.) tuber tissue which is reacting hypersensitively. To determine whether E/O2 retards SSM turnover, a measured amount of rishitin was applied to tuber tissue which was then incubated in air or E/O2, and rishitin disappearance was monitored. No difference in the rate of rishitin disappearance was detected between air and E/O2 incubations. However, tissue treated with rishitin and incubated in E/O2 accumulated intermediates of the katahdinone and phytuberin pathways. This was not the case in rishitin-air treatments. These results suggest the dual involvement of ethylene and SSM intermediates in the regulation of the biosynthesis of SSM, compounds which may serve as phytoalexins. PMID:16662127

  18. An in vitro control mechanism for potato stress metabolite biosynthesis.

    PubMed

    Alves, L M; Kalan, E B; Heisler, E G

    1981-12-01

    Ethylene/oxygen (E/O(2)) elevates sesquiterpenoid stress metabolite (SSM) levels in potato (Solanum tuberosum L.) tuber tissue which is reacting hypersensitively. To determine whether E/O(2) retards SSM turnover, a measured amount of rishitin was applied to tuber tissue which was then incubated in air or E/O(2), and rishitin disappearance was monitored. No difference in the rate of rishitin disappearance was detected between air and E/O(2) incubations. However, tissue treated with rishitin and incubated in E/O(2) accumulated intermediates of the katahdinone and phytuberin pathways. This was not the case in rishitin-air treatments. These results suggest the dual involvement of ethylene and SSM intermediates in the regulation of the biosynthesis of SSM, compounds which may serve as phytoalexins. PMID:16662127

  19. Cocaine metabolite (benzoylecgonine) in hair and urine of drug users.

    PubMed

    Martinez, F; Poet, T S; Pillai, R; Erickson, J; Estrada, A L; Watson, R R

    1993-01-01

    Two methods of drug detection, urinalysis and hair analysis, were compared with respect to the efficiency of identification of drug use in a population of men living on the Arizona-Mexico border. The standard curve of cannabinoids in urine was linear to 20 ng/mL. The GC/MS levels for all cannabinoids combined in urine were very similar to that obtained by radioimmunoassay (RIA), 91% concordance. Similar results were obtained from samples analyzed dually for the cocaine metabolite benzoylecgonine (BE) after spiking. As determined by RIA of urine, 74% of the subjects were positive for cannabinoids. The majority were in the range of 100-1000 ng/mg creatinine. The pattern of excretion of THC metabolites with respect to the verbally reported time of first use was fairly normal, with the peak rate of elimination 13-24 hours following the last reported use. Washed hair samples were extracted by overnight acid hydrolysis. Urine samples and neutralized hair extracts were analyzed for cocaine and BE by RIA. Of the hair samples, 55% contained cocaine/BE, as compared with only 4.3% of the urine samples. Most hair samples contained cocaine/BE in the range of 25-100 ng/sample (100 mg hair). All hair samples testing negative for cocaine/BE by RIA also tested negative by GC/MS, and four samples containing the highest amounts of cocaine and BE by RIA were similarly found to contain the highest amounts by GC/MS. Hair analysis, therefore, gives a wider window of detection of drug use than does urinalysis and shows merit in the confirmation of cocaine use in small clinical research studies. PMID:8336486

  20. Metabolite specific proton magnetic resonance imaging

    SciTech Connect

    Hurd, R.E.; Freeman, D.M.

    1989-06-01

    An imaging method is described that makes use of proton double quantum nuclear magnetic resonance (NMR) to construct images based on selected metabolites such as lactic acid. The optimization of the method is illustrated in vitro, followed by in vivo determination of lactic acid distribution in a solid tumor model. Water suppression and editing of lipid signals are such that two-dimensional spectra of lactic acid may be obtained from a radiation-induced fibrosarcoma (RIF-1) tumor in under 1 min and lactic acid images from the same tumor in under 1 hr at 2.0 T. This technique provides a fast and reproducible method at moderate magnetic field strength for mapping biologically relevant metabolites.

  1. Gut microbiota, metabolites and host immunity.

    PubMed

    Rooks, Michelle G; Garrett, Wendy S

    2016-05-27

    The microbiota - the collection of microorganisms that live within and on all mammals - provides crucial signals for the development and function of the immune system. Increased availability of technologies that profile microbial communities is facilitating the entry of many immunologists into the evolving field of host-microbiota studies. The microbial communities, their metabolites and components are not only necessary for immune homeostasis, they also influence the susceptibility of the host to many immune-mediated diseases and disorders. In this Review, we discuss technological and computational approaches for investigating the microbiome, as well as recent advances in our understanding of host immunity and microbial mutualism with a focus on specific microbial metabolites, bacterial components and the immune system. PMID:27231050

  2. Metabolites of a blocked chloramphenicol producer.

    PubMed

    Lewis, Elizabeth A; Adamek, Tamara L; Vining, Leo C; White, Robert L

    2003-01-01

    Addition of p-aminophenylalanine (4), an advanced biosynthetic precursor of the antibiotic chloramphenicol (5), to a Streptomyces venezuelae pabAB mutant (VS629) restored chloramphenicol production and led to formation of the non-chlorinated analogue corynecin II (6) and four acetanilide derivatives: p-(acetylamino)phenylalanine (7), p-(acetylamino)benzyl alcohol (13), p-(acetylamino)benzoic acid (14), and p-(acetylamino)phenol (acetaminophen, 16). Metabolite structures were deduced from NMR and MS-MS data and established by chromatographic and spectroscopic comparisons with authentic samples. Reference compound 13 was synthesized by reducing the acid chloride of 14. Shunt pathways are proposed to account for the formation of the metabolites from p-aminophenylalanine. PMID:12542347

  3. Emerging role of thyroid hormone metabolites.

    PubMed

    Gnocchi, D; Steffensen, K R; Bruscalupi, G; Parini, P

    2016-07-01

    Thyroid hormones (THs) are essential for the regulation of development and metabolism in key organs. THs produce biological effects both by directly affecting gene expression through the interaction with nuclear receptors (genomic effects) and by activating protein kinases and/or ion channels (short-term effects). Such activations can be either direct, in the case of ion channels, or mediated by membrane or cytoplasmic receptors. Short-term-activated signalling pathways often play a role in the regulation of genomic effects. Several TH intermediate metabolites, which were previously considered without biological activity, have now been associated with a broad range of actions, mostly attributable to short-term effects. Here, we give an overview of the physiological roles and mechanisms of action of THs, focusing on the emerging position that TH metabolites are acquiring as important regulators of physiology and metabolism. PMID:26748938

  4. Novel bioactive metabolites of dipyrone (metamizol).

    PubMed

    Rogosch, Tobias; Sinning, Christian; Podlewski, Agnes; Watzer, Bernhard; Schlosburg, Joel; Lichtman, Aron H; Cascio, Maria G; Bisogno, Tiziana; Di Marzo, Vincenzo; Nüsing, Rolf; Imming, Peter

    2012-01-01

    Dipyrone is a common antipyretic drug and the most popular non-opioid analgesic in many countries. In spite of its long and widespread use, molecular details of its fate in the body are not fully known. We administered dipyrone orally to mice. Two unknown metabolites were found, viz. the arachidonoyl amides of the known major dipyrone metabolites, 4-methylaminoantipyrine (2) and 4-aminoantipyrine (3). They were identified by ESI-LC-MS/MS after extraction from the CNS, and comparison with reference substances prepared synthetically. The arachidonoyl amides were positively tested for cannabis receptor binding (CB(1) and CB(2)) and cyclooxygenase inhibition (COX-1 and COX-2 in tissues and as isolated enzymes), suggesting that the endogenous cannabinoid system may play a role in the effects of dipyrone against pain. PMID:22172309

  5. Biologically Active Metabolites Synthesized by Microalgae

    PubMed Central

    de Morais, Michele Greque; Vaz, Bruna da Silva; de Morais, Etiele Greque; Costa, Jorge Alberto Vieira

    2015-01-01

    Microalgae are microorganisms that have different morphological, physiological, and genetic traits that confer the ability to produce different biologically active metabolites. Microalgal biotechnology has become a subject of study for various fields, due to the varied bioproducts that can be obtained from these microorganisms. When microalgal cultivation processes are better understood, microalgae can become an environmentally friendly and economically viable source of compounds of interest, because production can be optimized in a controlled culture. The bioactive compounds derived from microalgae have anti-inflammatory, antimicrobial, and antioxidant activities, among others. Furthermore, these microorganisms have the ability to promote health and reduce the risk of the development of degenerative diseases. In this context, the aim of this review is to discuss bioactive metabolites produced by microalgae for possible applications in the life sciences. PMID:26339647

  6. [Basidiomycetes: A new source of secondary metabolites.].

    PubMed

    Brizuela, M A; García, L; Pérez, L; Mansur, M

    1998-06-01

    The area of natural products research is the most rapidly growing field of organic chemistry, due to the great technical developments in the isolation and identification techniques. Today, near to one million natural products -isolated from the most diverse living things- are known. Microorganisms are among the least-studied of these. Nevertheless, they offer large possibilities for the discovery of new structures and biological activities. Among the microorganisms, the Basidiomycetes present a production capacity and a range of biologically active metabolites, which have scarcely been investigated. The wide spectrum of natural products with biological activity produced by Basidiomycetes includes antimicrobial agents, antifungal, antiviral and cytotoxic activities, enzymes, plant growth regulators and flavors. These metabolites are generally grouped by their chemical origin, and the relationship between chemical structure and the different biological activities reported. The main objective of this review is to bring an updated revision of the numerous and interesting biosynthetic pathways from basidiomycetes. PMID:17655412

  7. Heterogeneous distribution of metabolites across plant species

    NASA Astrophysics Data System (ADS)

    Takemoto, Kazuhiro; Arita, Masanori

    2009-07-01

    We investigate the distribution of flavonoids, a major category of plant secondary metabolites, across species. Flavonoids are known to show high species specificity, and were once considered as chemical markers for understanding adaptive evolution and characterization of living organisms. We investigate the distribution among species using bipartite networks, and find that two heterogeneous distributions are conserved among several families: the power-law distributions of the number of flavonoids in a species and the number of shared species of a particular flavonoid. In order to explain the possible origin of the heterogeneity, we propose a simple model with, essentially, a single parameter. As a result, we show that two respective power-law statistics emerge from simple evolutionary mechanisms based on a multiplicative process. These findings provide insights into the evolution of metabolite diversity and characterization of living organisms that defy genome sequence analysis for different reasons.

  8. Preparative Microfluidic Electrosynthesis of Drug Metabolites

    PubMed Central

    2013-01-01

    In vivo, a drug molecule undergoes its first chemical transformation within the liver via CYP450-catalyzed oxidation. The chemical outcome of the first pass hepatic oxidation is key information to any drug development process. Electrochemistry can be used to simulate CYP450 oxidation, yet it is often confined to the analytical scale, hampering product isolation and full characterization. In an effort to replicate hepatic oxidations, while retaining high throughput at the preparative scale, microfluidic technology and electrochemistry are combined in this study by using a microfluidic electrochemical cell. Several commercial drugs were subjected to continuous-flow electrolysis. They were chosen for their various chemical reactivity: their metabolites in vivo are generated via aromatic hydroxylation, alkyl oxidation, glutathione conjugation, or sulfoxidation. It is demonstrated that such metabolites can be synthesized by flow electrolysis at the 10 to 100 mg scale, and the purified products are fully characterized. PMID:24900614

  9. Metabolic regulation and overproduction of primary metabolites

    PubMed Central

    Sanchez, Sergio; Demain, Arnold L.

    2008-01-01

    Summary Overproduction of microbial metabolites is related to developmental phases of microorganisms. Inducers, effectors, inhibitors and various signal molecules play a role in different types of overproduction. Biosynthesis of enzymes catalysing metabolic reactions in microbial cells is controlled by well‐known positive and negative mechanisms, e.g. induction, nutritional regulation (carbon or nitrogen source regulation), feedback regulation, etc. The microbial production of primary metabolites contributes significantly to the quality of life. Fermentative production of these compounds is still an important goal of modern biotechnology. Through fermentation, microorganisms growing on inexpensive carbon and nitrogen sources produce valuable products such as amino acids, nucleotides, organic acids and vitamins which can be added to food to enhance its flavour, or increase its nutritive values. The contribution of microorganisms goes well beyond the food and health industries with the renewed interest in solvent fermentations. Microorganisms have the potential to provide many petroleum‐derived products as well as the ethanol necessary for liquid fuel. Additional applications of primary metabolites lie in their impact as precursors of many pharmaceutical compounds. The roles of primary metabolites and the microbes which produce them will certainly increase in importance as time goes on. In the early years of fermentation processes, development of producing strains initially depended on classical strain breeding involving repeated random mutations, each followed by screening or selection. More recently, methods of molecular genetics have been used for the overproduction of primary metabolic products. The development of modern tools of molecular biology enabled more rational approaches for strain improvement. Techniques of transcriptome, proteome and metabolome analysis, as well as metabolic flux analysis. have recently been introduced in order to identify new and

  10. Three new metabolites from Botrytis cinerea.

    PubMed

    Wang, Tian-Shan; Zhou, Jin-Yan; Tan, Hong

    2008-01-01

    Three new metabolites, gamma-abscisolactone (1), botrytisic acids A (3) and B (4) were isolated from the fermentation broth of Botrytis cinerea TB-3-H8. Their structures were elucidated on the basis of MS, IR, UV, and NMR spectroscopic data. Compound 2 was isolated from natural resource for the first time. The structure of 1 was further confirmed by single-crystal X-ray diffraction (CCDC-265897). PMID:19003608

  11. Mass spectrometric identification and characterization of a new long-term metabolite of metandienone in human urine.

    PubMed

    Schänzer, Wilhelm; Geyer, Hans; Fusshöller, Gregor; Halatcheva, Natalia; Kohler, Maxie; Parr, Maria-Kristina; Guddat, Sven; Thomas, Andreas; Thevis, Mario

    2006-01-01

    Anabolic-androgenic steroids are some of the most frequently detected drugs in amateur and professional sports. Doping control laboratories have developed numerous assays enabling the determination of administered drugs and/or their metabolic products that allow retrospectives with respect to pharmacokinetics and excretion profiles of steroids and their metabolites. A new metabolite generated from metandienone has been identified as 18-nor-17beta-hydroxymethyl,17alpha-methyl-androst-1,4,13-trien-3-one in excretion study urine samples providing a valuable tool for the long-term detection of metandienone abuse by athletes in sports drug testing. The metabolite was characterized using gas chromatography/(tandem) mass spectrometry, liquid chromatography/tandem mass spectrometry and liquid chromatography/high-resolution/high-accuracy (tandem) mass spectrometry by characteristic fragmentation patterns representing the intact 3-keto-1,4-diene structure in combination with typical product ions substantiating the proposed C/D-ring structure of the steroid metabolite. In addition, structure confirmation was obtained by the analysis of excretion study urine specimens obtained after administration of 17-CD(3)-labeled metandienone providing the deuterated analogue to the newly identified metabolite. 18-Nor-17beta-hydroxymethyl,17alpha-methyl-androst-1,4,13-trien-3-one was determined in metandienone administration study urine specimens up to 19 days after application of a single dose of 5 mg, hence providing an extended detection period compared with commonly employed strategies. PMID:16804957

  12. Analysis of paralytic shellfish toxins and their metabolites in shellfish from the North Yellow Sea of China.

    PubMed

    Li, A; Ma, J; Cao, J; Wang, Q; Yu, R; Thomas, K; Quilliam, M A

    2012-01-01

    Samples of toxic scallop (Patinopecten yessoensis) and clam (Saxidomus purpuratus) collected on the northern coast of China from 2008 to 2009 were analysed. High-performance liquid chromatography with post-column oxidation and fluorescence detection was used to determine the profile of the main paralytic shellfish poisoning (PSP) toxins in these samples and their total toxicity. Hydrophilic interaction liquid ion chromatography with mass spectrometric detection confirmed the toxin profile and detected several metabolites in the shellfish. Results show that C1/2 toxins were the most dominant toxins in the scallop and clam samples. However, GTX1/4 and GTX2/3 were also present. M1 was the predominant metabolite in all the samples, but M3 and M5 were also identified, along with three previously unreported presumed metabolites, M6, M8 and M10. The results indicate that the biotransformation of toxins was species specific. It was concluded that the reductive enzyme in clams is more active than in scallops and that an enzyme in scallops is more apt to catalyse hydrolysis of both the sulfonate moiety at the N-sulfocabamoyl of C toxins and the 11-hydroxysulfate of C and GTX toxins to produce metabolites. This is the first report of new metabolites of PSP toxins in scallops and clams collected in China. PMID:22827221

  13. Personal care product use and urinary levels of phthalate metabolites in Mexican women.

    PubMed

    Romero-Franco, Michelle; Hernández-Ramírez, Raúl U; Calafat, Antonia M; Cebrián, Mariano E; Needham, Larry L; Teitelbaum, Susan; Wolff, Mary S; López-Carrillo, Lizbeth

    2011-07-01

    Sources of phthalates other than Polyvinyl chloride (PVC) related products are scarcely documented in Mexico. The objective of our study was to explore the association between urinary levels of nine phthalate metabolites and the use of personal care products. Subjects included 108 women who participated as controls in an ongoing population-based case-control study of environmental factors and genetic susceptibility to breast cancer in northern Mexico. Direct interviews were performed to inquire about sociodemographic characteristics, reproductive history, use of personal care products, and diet. Phthalate metabolites measured in urine by high performance liquid chromatography-isotope dilution tandem mass spectrometry were monoethyl phthalate (MEP), monobenzyl phthalate (MBzP), mono-n-butyl phthalate (MBP), mono-isobutyl phthalate (MiBP), mono-3-carboxypropyl phthalate (MCPP) as well as mono-2-ethylhexyl phthalate (MEHP), mono-2-ethyl-5-oxohexyl phthalate (MEOHP), mono-2-ethyl-5-hydroxyhexyl phthalate (MEHHP), mono-2-ethyl-5-carboxypentyl phthalate (MECPP) that are metabolites of di-ethylhexyl phthalate (DEHP). Detectable urinary concentrations of phthalate metabolites varied from 75% (MEHP) to 100% (MEP, MBP, MEOHP, MEHHP and MECPP). Medians of urinary concentrations of some phthalate metabolites were significantly higher among users of the following personal care products compared to nonusers: body lotion (MEHHP, MECPP and sum of DEHP metabolites (ΣDEHP)), deodorant (MEHP and ΣDEHP), perfume (MiBP), anti-aging facial cream (MEP, MBP and MCPP) and bottled water (MCPP, MEHHP and MEOHP). Urinary concentrations of MEP showed a positive relationship with the number of personal care products used. Our results suggest that the use of some personal care products contributes to phthalate body burden that deserves attention due to its potential health impact. PMID:21429583

  14. Elevated cytokinin content in ipt transgenic creeping bentgrass promotes drought tolerance through regulating metabolite accumulation.

    PubMed

    Merewitz, Emily B; Du, Hongmei; Yu, Wenjuan; Liu, Yimin; Gianfagna, Thomas; Huang, Bingru

    2012-02-01

    Increased endogenous plant cytokinin (CK) content through transformation with an adenine isopentyl transferase (ipt) gene has been associated with improved plant drought tolerance. The objective of this study is to determine metabolic changes associated with elevated CK production in ipt transgenic creeping bentgrass (Agrostis stolonifera L.) with improved drought tolerance. Null transformants (NTs) and plants transformed with ipt controlled by a stress- or senescence-activated promoter (SAG12-ipt) were exposed to well-watered conditions or drought stress by withholding irrigation in an environmental growth chamber. Physiological analysis confirmed that the SAG12-ipt line (S41) had improved drought tolerance compared with the NT plants. Specific metabolite changes over the course of drought stress and differential accumulation of metabolites in SAG12-ipt plants compared with NT plants at the same level of leaf relative water content (47% RWC) were identified using gas chromatography-mass spectroscopy. The metabolite profiling analysis detected 45 metabolites differentially accumulated in response to ipt expression or drought stress, which included amino acids, carbohydrates, organic acids, and organic alcohols. The enhanced drought tolerance of SAG12-ipt plants was associated with the maintenance of accumulation of several metabolites, particularly amino acids (proline, γ-aminobutyric acid, alanine, and glycine) carbohydrates (sucrose, fructose, maltose, and ribose), and organic acids that are mainly involved in the citric acid cycle. The accumulation of these metabolites could contribute to improved drought tolerance due to their roles in the stress response pathways such as stress signalling, osmotic adjustment, and respiration for energy production. PMID:22131157

  15. Tissue distribution of berberine and its metabolites after oral administration in rats.

    PubMed

    Tan, Xiang-Shan; Ma, Jing-Yi; Feng, Ru; Ma, Chao; Chen, Wen-Jing; Sun, Yu-Peng; Fu, Jie; Huang, Min; He, Chi-Yu; Shou, Jia-Wen; He, Wen-Yi; Wang, Yan; Jiang, Jian-Dong

    2013-01-01

    Berberine (BBR) has been confirmed to have multiple bioactivities in clinic, such as cholesterol-lowering, anti-diabetes, cardiovascular protection and anti- inflammation. However, BBR's plasma level is very low; it cannot explain its pharmacological effects in patients. We consider that the in vivo distribution of BBR as well as of its bioactive metabolites might provide part of the explanation for this question. In this study, liquid chromatography coupled to ion trap time-of-flight mass spectrometry (LC/MS(n)-IT-TOF) as well as liquid chromatography that coupled with tandem mass spectrometry (LC-MS/MS) was used for the study of tissue distribution and pharmacokinetics of BBR in rats after oral administration (200 mg/kg). The results indicated that BBR was quickly distributed in the liver, kidneys, muscle, lungs, brain, heart, pancreas and fat in a descending order of its amount. The pharmacokinetic profile indicated that BBR's level in most of studied tissues was higher (or much higher) than that in plasma 4 h after administration. BBR remained relatively stable in the tissues like liver, heart, brain, muscle, pancreas etc. Organ distribution of BBR's metabolites was also investigated paralleled with that of BBR. Thalifendine (M1), berberrubine (M2) and jatrorrhizine (M4), which the metabolites with moderate bioactivity, were easily detected in organs like the liver and kidney. For instance, M1, M2 and M4 were the major metabolites in the liver, among which the percentage of M2 was up to 65.1%; the level of AUC (0-t) (area under the concentration-time curve) for BBR or the metabolites in the liver was 10-fold or 30-fold higher than that in plasma, respectively. In summary, the organ concentration of BBR (as well as its bioactive metabolites) was higher than its concentration in the blood after oral administration. It might explain BBR's pharmacological effects on human diseases in clinic. PMID:24205048

  16. Production of secondary metabolites by some terverticillate penicillia on carbohydrate-rich and meat substrates.

    PubMed

    Núñez, Félix; Westphal, Carmen D; Bermúdez, Elena; Asensio, Miguel A

    2007-12-01

    Most terverticillate penicillia isolated from dry-cured meat products are toxigenic, but their ability to produce hazardous metabolites on meat-based substrates is not well known. The production of extrolites by selected terverticillate penicillia isolated from dry-cured ham has been studied on carbohydrate-rich media (malt extract agar, Czapek yeast autolysate agar, rice extract agar, and rice), meat extract triolein salt agar, and ham slices. Chloroform extracts from the selected strains grown on malt extract agar were toxic for the brine shrimp (Artemia salina) larvae and VERO cells at a concentration of 2 mg/ml, but 0.02 mg/ml produced no toxic effect. Analysis by high-pressure liquid chromatography (HPLC) coupled with photodiode array detection (DAD) or with mass spectrometry (MS) and an atmospheric pressure chemical ionization (APCI) source revealed different biologically active metabolites: cyclopiazonic acid and rugulovasine A from Penicillium commune; verrucosidin, anacine, puberuline, verrucofortine, and viridicatols from Penicillium polonicum; arisugacin and viridicatols from Penicillium echinulatum; and compactin and viridicatols from Penicillium solitum. Most of these metabolites, including the amino acid-derived compounds, were produced in the media containing high levels of carbohydrates. High concentrations of nitrogen compounds in the medium does not imply a greater production of the metabolites studied, not even those derived from the amino acids. However, molds growing on dry-cured ham are able to synthesize limited amounts of some secondary metabolites, a fact not previously reported. The combination of HPLC coupled with DAD and MS-APCI was useful for identification of closely related terverticillate Penicillium species from dry-cured ham. These techniques could be used to characterize the risk associated with the potential production of secondary metabolites in cured meats. PMID:18095438

  17. Abundant Rodent Furan-Derived Urinary Metabolites Are Associated with Tobacco Smoke Exposure in Humans.

    PubMed

    Grill, Alex E; Schmitt, Thaddeus; Gates, Leah A; Lu, Ding; Bandyopadhyay, Dipankar; Yuan, Jian-Min; Murphy, Sharon E; Peterson, Lisa A

    2015-07-20

    Furan, a possible human carcinogen, is found in heat treated foods and tobacco smoke. Previous studies have shown that humans are capable of converting furan to its reactive metabolite, cis-2-butene-1,4-dial (BDA), and therefore may be susceptible to furan toxicity. Human risk assessment of furan exposure has been stymied because of the lack of mechanism-based exposure biomarkers. Therefore, a sensitive LC-MS/MS assay for six furan metabolites was applied to measure their levels in urine from furan-exposed rodents as well as in human urine from smokers and nonsmokers. The metabolites that result from direct reaction of BDA with lysine (BDA-N(α)-acetyllysine) and from cysteine-BDA-lysine cross-links (N-acetylcysteine-BDA-lysine, N-acetylcysteine-BDA-N(α)-acetyllysine, and their sulfoxides) were targeted in this study. Five of the six metabolites were identified in urine from rodents treated with furan by gavage. BDA-N(α)-acetyllysine, N-acetylcysteine-BDA-lysine, and its sulfoxide were detected in most human urine samples from three different groups. The levels of N-acetylcysteine-BDA-lysine sulfoxide were more than 10 times higher than that of the corresponding sulfide in many samples. The amount of this metabolite was higher in smokers relative to that in nonsmokers and was significantly reduced following smoking cessation. Our results indicate a strong relationship between BDA-derived metabolites and smoking. Future studies will determine if levels of these biomarkers are associated with adverse health effects in humans. PMID:26114498

  18. Urinary Phthalate Metabolites in Relation to Biomarkers of Inflammation and Oxidative Stress: NHANES 1999-2006

    PubMed Central

    Ferguson, Kelly K.; Loch-Caruso, Rita; Meeker, John D.

    2011-01-01

    Phthalate esters are a class of compounds utilized extensively in widely-distributed consumer goods, and have been associated with various adverse health outcomes in previous epidemiologic research. Some of these health outcomes may be the result of phthalate-induced increases in oxidative stress or inflammation, which has been demonstrated in animal studies. The aim of this study was to explore the relationship between urinary phthalate metabolite concentrations and serum markers of inflammation and oxidative stress (C-reactive protein (CRP) and gamma glutamyltransferase (GGT), respectively). Subjects were participants in the National Health and Nutrition Examination Survey (NHANES) between the years 1999 and 2006. In multivariable linear regression models, we observed significant positive associations between CRP and mono-benzyl phthalate (MBzP) and mono-isobutyl phthalate (MiBP). There were CRP elevations of 6.0% (95% confidence interval (CI) 1.7% to 10.8%) and 8.3% (95% CI 2.9% to 14.0%) in relation to interquartile range (IQR) increases in urinary MBzP and MiBP, respectively. GGT was positively associated with mono(2-ethylhexyl) phthalate (MEHP) and an MEHP% variable calculated from the proportion of MEHP in comparison to other di(2-ethylhexyl) phthalate (DEHP) metabolites. IQR increases in MEHP and MEHP% were associated with 2.5% (95%CI 0.2% to 4.8%) and 3.7% (95%CI 1.7% to 5.7%) increases in GGT, respectively. CRP and GGT were also inversely related to several phthalate metabolites, primarily oxidized metabolites. In conclusion, several phthalate monoester metabolites that are detected in a high proportion of urine samples from the US general population are associated with increased serum markers of inflammation and oxidative stress. On the other hand, several oxidized phthalate metabolites were inversely associated with these markers. These relationships deserve further exploration in both experimental and observational studies. PMID:21349512

  19. Phthalate Metabolites, Consumer Habits and Health Effects.

    PubMed

    Wallner, Peter; Kundi, Michael; Hohenblum, Philipp; Scharf, Sigrid; Hutter, Hans-Peter

    2016-01-01

    Phthalates are multifunctional chemicals used in a wide variety of consumer products. The aim of this study was to investigate whether levels of urinary phthalate metabolites in urine samples of Austrian mothers and their children were associated with consumer habits and health indicators. Within an Austrian biomonitoring survey, urine samples from 50 mother-child pairs of five communities (two-stage random stratified sampling) were analysed. The concentrations of 14 phthalate metabolites were determined, and a questionnaire was administered. Monoethyl phthalate (MEP), mono-n-butyl phthalate (MnBP), mono-isobutyl phthalate (MiBP), monobenzyl phthalate (MBzP), mono-(2-ethylhexyl) phthalate (MEHP), mono-(2-ethyl-5-hydroxyhexyl) phthalate (5OH-MEHP), mono-(2-ethyl-5-oxohexyl) phthalate (5oxo-MEHP), mono-(5-carboxy-2-ethylpentyl) phthalate (5cx-MEPP), and 3-carboxy-mono-propyl phthalate (3cx-MPP) could be quantified in the majority of samples. Significant correlations were found between the use of hair mousse, hair dye, makeup, chewing gum, polyethylene terephthalate (PET) bottles and the diethyl phthalate (DEP) metabolite MEP. With regard to health effects, significant associations of MEP in urine with headache, repeated coughing, diarrhoea, and hormonal problems were observed. MBzP was associated with repeated coughing and MEHP was associated with itching. PMID:27428989

  20. Phthalate Metabolites, Consumer Habits and Health Effects

    PubMed Central

    Wallner, Peter; Kundi, Michael; Hohenblum, Philipp; Scharf, Sigrid; Hutter, Hans-Peter

    2016-01-01

    Phthalates are multifunctional chemicals used in a wide variety of consumer products. The aim of this study was to investigate whether levels of urinary phthalate metabolites in urine samples of Austrian mothers and their children were associated with consumer habits and health indicators. Within an Austrian biomonitoring survey, urine samples from 50 mother-child pairs of five communities (two-stage random stratified sampling) were analysed. The concentrations of 14 phthalate metabolites were determined, and a questionnaire was administered. Monoethyl phthalate (MEP), mono-n-butyl phthalate (MnBP), mono-isobutyl phthalate (MiBP), monobenzyl phthalate (MBzP), mono-(2-ethylhexyl) phthalate (MEHP), mono-(2-ethyl-5-hydroxyhexyl) phthalate (5OH-MEHP), mono-(2-ethyl-5-oxohexyl) phthalate (5oxo-MEHP), mono-(5-carboxy-2-ethylpentyl) phthalate (5cx-MEPP), and 3-carboxy-mono-propyl phthalate (3cx-MPP) could be quantified in the majority of samples. Significant correlations were found between the use of hair mousse, hair dye, makeup, chewing gum, polyethylene terephthalate (PET) bottles and the diethyl phthalate (DEP) metabolite MEP. With regard to health effects, significant associations of MEP in urine with headache, repeated coughing, diarrhoea, and hormonal problems were observed. MBzP was associated with repeated coughing and MEHP was associated with itching. PMID:27428989

  1. Oseltamivir (tamiflu) induced depressive episode in a female adolescent.

    PubMed

    Chung, Sungho; Joung, Yoo Sook

    2010-12-01

    Oseltamivir was developed for prophylactic and therapeutic use against influenza, specifically targeting the viral enzyme's highly-conserved active site. In recent years, there have been case reports of neuropsychiatric events during or after oseltamivir treatment, in Japan and other countries. However, a search of the literature revealed no such cases in South Korea. We present the case of a 15-year-old female adolescent diagnosed with depressive episode after taking oseltamivir. Oseltamivir is generally well tolerated. Its most frequent adverse effects include nausea and vomiting, diarrhea, and abdominal pain. In influenza patients taking oseltamivir, neuropsychiatric adverse events include delirium, behavioral disturbance, suicide, delusion, panic attack, convulsion, depressed mood, loss of consciousness, etc. Reportedly, such neuropsychiatric adverse events were more common in children than in adults and generally occurred within 48 hours of administration. Here, we report a retrospective review case of an oseltamivir-related neuropsychiatric event in a female adolescent in South Korea. PMID:21253416

  2. The WEIZMASS spectral library for high-confidence metabolite identification.

    PubMed

    Shahaf, Nir; Rogachev, Ilana; Heinig, Uwe; Meir, Sagit; Malitsky, Sergey; Battat, Maor; Wyner, Hilary; Zheng, Shuning; Wehrens, Ron; Aharoni, Asaph

    2016-01-01

    Annotation of metabolites is an essential, yet problematic, aspect of mass spectrometry (MS)-based metabolomics assays. The current repertoire of definitive annotations of metabolite spectra in public MS databases is limited and suffers from lack of chemical and taxonomic diversity. Furthermore, the heterogeneity of the data prevents the development of universally applicable metabolite annotation tools. Here we present a combined experimental and computational platform to advance this key issue in metabolomics. WEIZMASS is a unique reference metabolite spectral library developed from high-resolution MS data acquired from a structurally diverse set of 3,540 plant metabolites. We also present MatchWeiz, a multi-module strategy using a probabilistic approach to match library and experimental data. This strategy allows efficient and high-confidence identification of dozens of metabolites in model and exotic plants, including metabolites not previously reported in plants or found in few plant species to date. PMID:27571918

  3. Relationships among nitric oxide metabolites and pulses of a PGF2α metabolite during and after luteolysis in mares.

    PubMed

    Ginther, O J; Wolf, C A; Baldrighi, J M; Greene, J M

    2015-07-15

    Hourly circulating concentrations of a PGF2α metabolite (PGFM), progesterone (P4), and LH were obtained from a reported project, and concentrations of nitric oxide (NO) metabolites (NOMs; nitrates and nitrites) were determined in eight mares. Unlike the reported project, hormone concentrations were normalized to the peak of the first PGFM pulse of luteolysis (early luteolysis), second PGFM pulse (late luteolysis), and a pulse after luteolysis. The duration of luteolysis was 23.1 ± 1.0 hours, and the peak of the first and second PGFM pulses occurred 6.5 ± 0.9 and 14.8 ± 0.8 hours after the beginning of luteolysis. Concentration of P4 decreased progressively within and between the PGFM pulses Changes were not detected in LH concentration in association with the PGFM pulses. Concentration of NOMs was greater (P < 0.05) at the peak of the PGFM pulse during early luteolysis (88.8 ± 15.0 μg/mL) than during late luteolysis (58.8 ± 9.0 μg/mL). Concentration of NOMs began to decrease (P < 0.05) 4 hours before the peak of the PGFM pulse of early luteolysis. Concentration began to increase (P < 0.05) an hour after the peak of the PGFM pulse of late luteolysis. An NOM decrease and increase was not detected during the PGFM pulse after luteolysis. On a temporal basis, results indicated that NO either is not required for luteolysis in mares or has a role in or responds only during late luteolysis. A caveat is that the relative contribution of the CL versus other body tissues to circulating concentrations of NOMs in mares has not been determined. PMID:25910877

  4. Characterization and quantitation of urinary metabolites of [1,2,3-13C]acrylamide in rats and mice using 13C nuclear magnetic resonance spectroscopy.

    PubMed

    Sumner, S C; MacNeela, J P; Fennell, T R

    1992-01-01

    Acrylamide, widely used for the production of polymers and as a grouting agent, causes neurotoxic effects in humans and neurotoxic, genotoxic, reproductive, and carcinogenic effects in laboratory animals. In this study, 13C NMR spectroscopy was used to detect metabolites of acrylamide directly in the urine of rats and mice following administration of [1,2,3-13C]acrylamide (50 mg/kg po). Two-dimensional NMR experiments were used to correlate carbon signals for each metabolite in the urine samples and to determine the number of hydrogens attached to each carbon. Metabolite structures were identified from the NMR data together with calculated values of shift for biochemically feasible metabolites and by comparison with standards. The metabolites assigned in rat and mouse urine are N-acetyl-S-(3-amino-3-oxopropyl)cysteine, N-acetyl-S-(3-amino-2-hydroxy-3-oxopropyl)cysteine, N-acetyl-S-(1-carbamoyl-2-hydroxy-ethyl)cysteine, glycidamide, and 2,3-dihydroxypropionamide. These metabolites arise from direct conjugation of acrylamide with glutathione or from oxidation to the epoxide, glycidamide, and further metabolism. Acrylamide was also detected in the urine. Quantitation was carried out by integrating the metabolite carbon signals with respect to that of dioxane added at a known concentration. The major metabolite for both the rat (70% of total metabolites excreted) and the mouse (40%) was formed from direct conjugation of acrylamide with glutathione. The remaining metabolites for the rat (30%) and mouse (60%) are derived from glycidamide. The species differences in extent of metabolism through glycidamide may have important consequences for the toxic and carcinogenic effects of acrylamide. PMID:1581543

  5. Determination of pesticides and their metabolites in processed cereal samples.

    PubMed

    González-Curbelo, M Á; Hernández-Borges, J; Borges-Miquel, T M; Rodríguez-Delgado, M Á

    2012-01-01

    Fifteen pesticides including some of their metabolites (disulfoton sulfoxide, ethoprophos, cadusafos, dimethoate, terbufos, disulfoton, chlorpyrifos-methyl, malaoxon, fenitrothion, pirimiphos-methyl, malathion, chlorpyrifos, terbufos sulfone, disulfoton sulfone and fensulfothion) were analysed in milled toasted wheat and maize as well as in wheat flour and baby cereals. The QuEChERS (quick, easy, cheap, effective, rugged and safe) methodology was used and its dispersive solid-phase extraction procedure was optimised by means of an experimental design with the aim of reducing the amount of co-extracted lipids and obtaining a clean extract. Gas chromatography with nitrogen phosphorus detection were used as the separation and detection techniques, respectively. The method was validated in terms of selectivity, recoveries, calibration, precision and accuracy as well as matrix effects. Limits of detection were between 0.07 and 34.8 µg kg(-1) with recoveries in the range of 71-110% (relative standard deviations were below 9%). A total of 40 samples of different origin were analysed. Residues of pirimiphos-methyl were found in six of the samples at concentrations in the range 0.08-0.47 mg kg(-1), which were below the MRLs established for this pesticide in cereal grains. Tandem mass spectrometry confirmation was also carried out in order to identify unequivocally the presence of this pesticide. PMID:22043870

  6. Mutagenic activity of austocystins - secondary metabolites of Aspergillus ustus

    SciTech Connect

    Kfir, R.; Johannsen, E.; Vleggaar, R.

    1986-11-01

    Mycotoxins constitute a group of toxic secondary fungal metabolites. Fungi that produce these toxins frequently contaminate food and feed, creating a potential threat to human and animal health. Biological activities of mycotoxins include, amongst others: toxicity, mutagenicity and carcinogenicity, which can be expressed with or without metabolic activation. Austocystins are similar in structure to aflatoxin B/sup 1/ and are probably synthesized in a similar manner. The Ames Salmonella test, a widely accepted method employed for the detection of mutagenic activity of various chemical compounds was used for testing the mutagenic activity of different mycotoxins. As aflatoxin B/sup 1/ was found by the Ames test to be highly mutagenic, the same test was applied for the study of possible mutagenicity of the austocystins. The mutagenic activity of these compounds was studied with and without metabolic activation using two tester strains of S. typhimurium, one capable of detecting frame shift mutation (strain TA98) and the other capable of detecting base pair substitution (strain TA100).

  7. Identification of in vivo and in vitro metabolites of triptolide by liquid chromatography-tandem mass spectrometry.

    PubMed

    Peng, Zhi-hong; Wang, Jun-jun; Du, Peng; Chen, Yong

    2012-11-01

    Triptolide, a major active constituent of Tripterygium wilfordii Hook F, has multiple pharmacological activities. In this work, a rapid, sensitive and specific liquid chromatography coupled to an ion trap mass spectrometer (MS) with electrospray ionization (ESI) interface has been developed for identification of triptolide and some of its metabolites in rat urine after oral administration of a single dose (0.6 mg/kg) of triptolide to healthy rats, as well as some metabolites in vitro after incubation with rat liver microsome (RLM) and rat intestinal flora, respectively. All samples were separated on a reversed-phase C18 column using a mobile phase of acetonitrile/water (70:30, v/v) and detected by an on-line MS(n) detector. Identification and structural elucidation of the selected metabolites were performed by comparing their full scan MS(n) spectra with those of the parent drug. In this paper we identified ten metabolites in rat urine, four metabolites in RLM incubation solution and one metabolite in rat intestinal flora incubation solution, after drug administration. The metabolic reactions of triptolide that we observed in vivo were hydrolysis reaction, hydroxylation reaction, and the conjugate reaction with sulfate, glucuronide and GSH, respectively. The in vitro metabolic reactions of triptolide observed were hydrolysis and hydroxylation reactions. PMID:22824635

  8. Determination of Urinary PAH Metabolites Using DLLME Hyphenated to Injector Port Silylation and GC-MS-MS.

    PubMed

    Gupta, Manoj Kumar; Jain, Rajeev; Singh, Pratibha; Ch, Ratnasekhar; Mudiam, Mohana Krishna Reddy

    2015-06-01

    Polycyclic aromatic hydrocarbons (PAHs) are ubiquitous environmental pollutants and well-known carcinogens. Hydroxy derivatives of PAH are considered as biomarkers of PAH exposure, and there is a need to measure these metabolites at low concentrations. So, a precise and eco-friendly analytical method has been developed for rapid determination of PAH metabolites. For the first time, a new analytical method based on coupling of dispersive liquid-liquid microextraction (DLLME) with auto-injector port silylation (auto-IPS) followed by gas chromatography-tandem mass spectrometry (GC-MS-MS) analysis is reported for the analysis of seven urinary PAH metabolites. Factors affecting DLLME and IPS, such as type and volume of extraction and disperser solvent, pH, ionic strength, injector port temperature, volume of N,O-bis(trimethylsilyl)trifluoroacetamide and type of solvent were investigated. Under optimized conditions, the limit of detection and limit of quantification were found to be in the range of 1-9 and 3-29 ng/mL, respectively. Satisfactory recoveries of metabolites in urine samples in the range of 87-95% were found. The developed method has been successfully applied for the determination of PAH metabolites in urine samples of exposed workers. DLLME-auto-IPS-GC-MS-MS method is time, labor, solvent and reagent saving, which can be routinely used for the analysis of urinary PAH metabolites. PMID:25845603

  9. Stable Isotope Labeling Strategy for Curcumin Metabolite Study in Human Liver Microsomes by Liquid Chromatography-Tandem Mass Spectrometry

    NASA Astrophysics Data System (ADS)

    Gao, Dan; Chen, Xiaowu; Yang, Xiaomei; Wu, Qin; Jin, Feng; Wen, Hongliang; Jiang, Yuyang; Liu, Hongxia

    2015-04-01

    The identification of drug metabolites is very important in drug development. Nowadays, the most widely used methods are isotopes and mass spectrometry. However, the commercial isotopic labeled reagents are usually very expensive, and the rapid and convenient identification of metabolites is still difficult. In this paper, an 18O isotope labeling strategy was developed and the isotopes were used as a tool to identify drug metabolites using mass spectrometry. Curcumin was selected as a model drug to evaluate the established method, and the 18O labeled curcumin was successfully synthesized. The non-labeled and 18O labeled curcumin were simultaneously metabolized in human liver microsomes (HLMs) and analyzed by liquid chromatography/mass spectrometry (LC-MS). The two groups of chromatograms obtained from metabolic reaction mixture with and without cofactors were compared and analyzed using Metabolynx software (Waters Corp., Milford, MA, USA). The mass spectra of the newly appearing chromatographic peaks in the experimental sample were further analyzed to find the metabolite candidates. Their chemical structures were confirmed by tandem mass spectrometry. Three metabolites, including two reduction products and a glucuronide conjugate, were successfully detected under their specific HLMs metabolic conditions, which were in accordance with the literature reported results. The results demonstrated that the developed isotope labeling method, together with post-acquisition data processing using Metabolynx software, could be used for fast identification of new drug metabolites.

  10. Identification of fipronil metabolites by time-of-flight mass spectrometry for application in a human exposure study.

    PubMed

    McMahen, Rebecca L; Strynar, Mark J; Dagnino, Sonia; Herr, David W; Moser, Virginia C; Garantziotis, Stavros; Andersen, Erik M; Freeborn, Danielle L; McMillan, Larry; Lindstrom, Andrew B

    2015-05-01

    Fipronil is a phenylpyrazole insecticide commonly used in residential and agricultural applications. To understand more about the potential risks for human exposure associated with fipronil, urine and serum from dosed Long Evans adult rats (5 and 10mg/kg bw) were analyzed to identify metabolites as potential biomarkers for use in human biomonitoring studies. Urine from treated rats was found to contain seven unique metabolites, two of which had not been previously reported-M4 and M7 which were putatively identified as a nitroso compound and an imine, respectively. Fipronil sulfone was confirmed to be the primary metabolite in rat serum. The fipronil metabolites identified in the respective matrices were then evaluated in matched human urine (n=84) and serum (n=96) samples from volunteers with no known pesticide exposures. Although no fipronil or metabolites were detected in human urine, fipronil sulfone was present in the serum of approximately 25% of the individuals at concentrations ranging from 0.1 to 4ng/mL. These results indicate that many fipronil metabolites are produced following exposures in rats and that fipronil sulfone is a useful biomarker in human serum. Furthermore, human exposure to fipronil may occur regularly and require more extensive characterization. PMID:25687022

  11. Tissue-Specific Distribution of Secondary Metabolites in Rapeseed (Brassica napus L.)

    PubMed Central

    Fang, Jingjing; Reichelt, Michael; Hidalgo, William; Agnolet, Sara; Schneider, Bernd

    2012-01-01

    Four different parts, hypocotyl and radicle (HR), inner cotyledon (IC), outer cotyledon (OC), seed coat and endosperm (SE), were sampled from mature rapeseed (Brassica napus L.) by laser microdissection. Subsequently, major secondary metabolites, glucosinolates and sinapine, as well as three minor ones, a cyclic spermidine conjugate and two flavonoids, representing different compound categories, were qualified and quantified in dissected samples by high-performance liquid chromatography with diode array detection and mass spectrometry. No qualitative and quantitative difference of glucosinolates and sinapine was detected in embryo tissues (HR, IC and OC). On the other hand, the three minor compounds were observed to be distributed unevenly in different rapeseed tissues. The hypothetic biological functions of the distribution patterns of different secondary metabolites in rapeseed are discussed. PMID:23133539

  12. Methods for isolating, identifying, and quantifying anthocyanin metabolites in clinical samples.

    PubMed

    de Ferrars, Rachel M; Czank, Charles; Saha, Shikha; Needs, Paul W; Zhang, Qingzhi; Raheem, K Saki; Botting, Nigel P; Kroon, Paul A; Kay, Colin D

    2014-10-21

    The metabolic fate of anthocyanins until recently was relatively unknown, primarily as a result of their instability at physiological pH and a lack of published methods for isolating and identifying their metabolites from biological samples. The aim of the present work was to establish methods for the extraction and quantification of anthocyanin metabolites present in urine, serum, and fecal samples. 35 commercial and 10 synthetic analytes, including both known and predicted human and microbial metabolites of anthocyanins, were obtained as reference standards. HPLC and MS/MS conditions were optimized for organic modifier, ionic modifier, mobile phase gradient, flow rate, column type, MS source, and compound dependent parameters. The impact of sorbent, solvent, acid, preservative, elution, and evaporation on solid phase extraction (SPE) efficiency was also explored. The HPLC-MS/MS method validation demonstrated acceptable linearity (R(2), 0.997 ± 0.002) and sensitivity (limits of detection (LODs): urine, 100 ± 375 nM; serum, 104 ± 358 nM; feces 138 ± 344 nM), and the final SPE methods provided recoveries of 88.3 ± 17.8% for urine, 86.5 ± 11.1% for serum, and 80.6 ± 20.9% for feces. The final methods were applied to clinical samples derived from an anthocyanin intervention study, where 36 of the 45 modeled metabolites were detected within urine, plasma, or fecal samples. The described methods provide suitable versatility for the identification and quantification of an extensive series of anthocyanin metabolites for use in future clinical studies exploring absorption, distribution, metabolism, and elimination. PMID:24828315

  13. Bioactivation of clopidogrel and prasugrel: factors determining the stereochemistry of the thiol metabolite double bond.

    PubMed

    Dansette, Patrick M; Levent, Dan; Hessani, Assia; Mansuy, Daniel

    2015-06-15

    The antithrombotics of the tetrahydrothienopyridine series, clopidogrel and prasugrel, are prodrugs that must be metabolized in two steps to become pharmacologically active. The first step is the formation of a thiolactone metabolite. The second step is a further oxidation with the formation of a thiolactone sulfoxide whose hydrolytic opening leads to a sulfenic acid that is eventually reduced into the corresponding active cis thiol. Very few data were available on the formation of the isomer of the active cis thiol having a trans configuration of the double bond, the most striking result in that regard being that both cis and trans thiols were formed upon the metabolism of clopidogrel by human liver microsomes in the presence of glutathione (GSH), whereas only the cis thiol was detected in the sera of patients treated with this drug. This article shows that trans thiols are also formed upon the microsomal metabolism of prasugrel or its thiolactone metabolite in the presence of GSH and that metabolites having the trans configuration of the double bond are only formed when microsomal incubations are done in the presence of thiols, such as GSH, N-acetyl-cysteine, and mercaptoethanol. Intermediate formation of thioesters resulting from the reaction of GSH with the thiolactone sulfoxide metabolite appears to be responsible for trans thiol formation. Addition of human liver cytosol to the microsomal incubations led to a dramatic decrease of the formation of the trans thiol metabolites. These data suggest that cytosolic esterases would accelerate the hydrolytic opening of thiolactone sulfoxide intermediates and disfavor the formation of thioesters resulting from the reaction of these intermediates with GSH that is responsible for trans isomer formation. This would explain why trans thiols have not been detected in the sera of patients treated with clopidogrel. PMID:25970225

  14. The environmental occurrence and effect of alkylphenol polyethoxylates and their metabolites in Taiwan

    NASA Astrophysics Data System (ADS)

    Ding, W.

    2009-12-01

    Alkylphenol polyethoxylates (APEOs) are widely used nonionic surfactants in domestic, agricultural and household applications, which have been commonly found in wastewater discharges and in sewage treatment plant effluents. Degradation of APEOs in wastewater or in the environment generates more persistent pollutants, including alkylphenols (APs, such as 4-nonylphenol isomers (4-NPs) and 4-t-octylphenol (4-t-OP)) and shortened ethoxy chain APEO residues (such as AP1~3EO). These metabolites of APEOs are of interest in the field of environmental monitoring because of the volume of these substances used and their activity as either endocrine disruptors or as persistent pollutants. APEOs are mass-produced and used widely in Taiwan. Large quantities of these metabolites in wastewater are discharged into the rivers directly because Taiwan’s municipal and industrial wastewater treatment facilities are deficient. However, the occurrence and fate of these metabolites are unclear and can potentially affect the aquatic environment and public health in Taiwan. Determination of APEOs and their metabolites have been performed for household detergents, various surface water, soil, sediments, biota, foodstuffs and even in breast milk. APEOs and their metabolites were detected in all media analyzed and in all environmental samples. The relatively high concentrations detected in oysters and snails provide evidence for bioaccumulation of APs. The presence of APs in breast milk implies that APs enter the food chain in local biota after long chain APEOs were biodegraded. There are also some indications that the plastic wrappings and containers for foodstuffs sold in Taiwan may contain NP or tris(nonylphenol) phosphate (TNPP) used as plasticizers or antioxidants. In addition, possible sources of APs may come from the extensive use of pesticides containing APEO as emulsifiers in agriculture.

  15. Age-Related Differences in the Disposition of Nicotine and Metabolites in Rat Brain and Plasma

    PubMed Central

    2013-01-01

    Introduction: Studies have evaluated