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Sample records for ethyl glucuronide etg

  1. A wearable biochemical sensor for monitoring alcohol consumption lifestyle through Ethyl glucuronide (EtG) detection in human sweat

    NASA Astrophysics Data System (ADS)

    Panneer Selvam, Anjan; Muthukumar, Sriram; Kamakoti, Vikramshankar; Prasad, Shalini

    2016-03-01

    We demonstrate for the first time a wearable biochemical sensor for monitoring alcohol consumption through the detection and quantification of a metabolite of ethanol, ethyl glucuronide (EtG). We designed and fabricated two co-planar sensors with gold and zinc oxide as sensing electrodes. We also designed a LED based reporting for the presence of EtG in the human sweat samples. The sensor functions on affinity based immunoassay principles whereby monoclonal antibodies for EtG were immobilized on the electrodes using thiol based chemistry. Detection of EtG from human sweat was achieved through chemiresistive sensing mechanism. In this method, an AC voltage was applied across the two coplanar electrodes and the impedance across the sensor electrodes was measured and calibrated for physiologically relevant doses of EtG in human sweat. EtG detection over a dose concentration of 0.001–100 μg/L was demonstrated on both glass and polyimide substrates. Detection sensitivity was lower at 1 μg/L with gold electrodes as compared to ZnO, which had detection sensitivity of 0.001 μg/L. Based on the detection range the wearable sensor has the ability to detect alcohol consumption of up to 11 standard drinks in the US over a period of 4 to 9 hours.

  2. A wearable biochemical sensor for monitoring alcohol consumption lifestyle through Ethyl glucuronide (EtG) detection in human sweat.

    PubMed

    Selvam, Anjan Panneer; Muthukumar, Sriram; Kamakoti, Vikramshankar; Prasad, Shalini

    2016-03-21

    We demonstrate for the first time a wearable biochemical sensor for monitoring alcohol consumption through the detection and quantification of a metabolite of ethanol, ethyl glucuronide (EtG). We designed and fabricated two co-planar sensors with gold and zinc oxide as sensing electrodes. We also designed a LED based reporting for the presence of EtG in the human sweat samples. The sensor functions on affinity based immunoassay principles whereby monoclonal antibodies for EtG were immobilized on the electrodes using thiol based chemistry. Detection of EtG from human sweat was achieved through chemiresistive sensing mechanism. In this method, an AC voltage was applied across the two coplanar electrodes and the impedance across the sensor electrodes was measured and calibrated for physiologically relevant doses of EtG in human sweat. EtG detection over a dose concentration of 0.001-100 μg/L was demonstrated on both glass and polyimide substrates. Detection sensitivity was lower at 1 μg/L with gold electrodes as compared to ZnO, which had detection sensitivity of 0.001 μg/L. Based on the detection range the wearable sensor has the ability to detect alcohol consumption of up to 11 standard drinks in the US over a period of 4 to 9 hours.

  3. A wearable biochemical sensor for monitoring alcohol consumption lifestyle through Ethyl glucuronide (EtG) detection in human sweat

    PubMed Central

    Panneer Selvam, Anjan; Muthukumar, Sriram; Kamakoti, Vikramshankar; Prasad, Shalini

    2016-01-01

    We demonstrate for the first time a wearable biochemical sensor for monitoring alcohol consumption through the detection and quantification of a metabolite of ethanol, ethyl glucuronide (EtG). We designed and fabricated two co-planar sensors with gold and zinc oxide as sensing electrodes. We also designed a LED based reporting for the presence of EtG in the human sweat samples. The sensor functions on affinity based immunoassay principles whereby monoclonal antibodies for EtG were immobilized on the electrodes using thiol based chemistry. Detection of EtG from human sweat was achieved through chemiresistive sensing mechanism. In this method, an AC voltage was applied across the two coplanar electrodes and the impedance across the sensor electrodes was measured and calibrated for physiologically relevant doses of EtG in human sweat. EtG detection over a dose concentration of 0.001–100 μg/L was demonstrated on both glass and polyimide substrates. Detection sensitivity was lower at 1 μg/L with gold electrodes as compared to ZnO, which had detection sensitivity of 0.001 μg/L. Based on the detection range the wearable sensor has the ability to detect alcohol consumption of up to 11 standard drinks in the US over a period of 4 to 9 hours. PMID:26996103

  4. Quantification of fatty acid ethyl esters (FAEE) and ethyl glucuronide (EtG) in meconium for detection of alcohol abuse during pregnancy: Correlation study between both biomarkers.

    PubMed

    Cabarcos, Pamela; Tabernero, María Jesús; Otero, José Luís; Míguez, Martha; Bermejo, Ana María; Martello, Simona; De Giovanni, Nadia; Chiarotti, Marcello

    2014-11-01

    This article presents results from 47 meconium samples, which were analyzed for fatty acid ethyl esters (FAEE) and ethyl glucuronide (EtG) for detection of gestational alcohol consumption. A validated microwave assisted extraction (MAE) method in combination with GC-MS developed in the Institute of Forensic Science (Santiago de Compostela) was used for FAEE and the cumulative concentration of ethyl myristate, ethyl palmitate and ethyl stearate with a cut-off of 600ng/g was applied for interpretation. A simple method for identification and quantification of EtG has been evaluated by ultrasonication followed solid phase extraction (SPE). Successful validation parameters were obtained for both biochemical markers of alcohol intake. FAEE and EtG concentrations in meconium ranged between values lower than LOD and 32,892ng/g or 218ng/g respectively. We have analyzed FAEE and EtG in the same meconium aliquot, enabling comparison of the efficiency of gestational ethanol exposure detection. Certain agreement between the two biomarkers was found as they are both a very specific alcohol markers, making it a useful analysis for confirmation.

  5. Detecting alcohol abuse: traditional blood alcohol markers compared to ethyl glucuronide (EtG) and fatty acid ethyl esters (FAEEs) measurement in hair.

    PubMed

    Hastedt, Martin; Büchner, Mara; Rothe, Michael; Gapert, René; Herre, Sieglinde; Krumbiegel, Franziska; Tsokos, Michael; Kienast, Thorsten; Heinz, Andreas; Hartwig, Sven

    2013-12-01

    Alcohol abuse is a common problem in society; however, the technical capabilities of evaluating individual alcohol consumption using objective biomarkers are rather limited at present. In recent years research has focused on alcohol markers using hair analysis but data on performance and reliable cut-off values are still lacking. In this study 169 candidates were tested to compare traditional biomarkers, such as carbohydrate-deficient-transferrin (CDT), gamma glutamyl transferase (GGT), aspartate amino transferase, alanine amino transferase and the mean corpuscular volume of the erythrocytes, with alcohol markers detectable in hair such as ethyl glucuronide (EtG) and fatty acid ethyl esters (FAEEs). This study revealed that EtG, GGT and CDT showed the best results, demonstrating areas under the curve calculated from receiver operating characteristics of 0.941, 0.943 and 0.899 respectively. The lowest false-negative and false-positive rates were obtained by using a combined interpretation system for hair EtG and FAEEs. All markers demonstrated only low to moderate correlations. Optimum cut-off values for differentiation between social and chronic excessive drinking calculated for hair EtG and FAEEs were 28 pg/mg and 0.675 ng/mg, respectively. The critical values published in the "Consensus on Alcohol Markers 2012" by the Society of Hair Testing were confirmed.

  6. High levels of agreement between clinic-based ethyl glucuronide (EtG) immunoassays and laboratory-based mass spectrometry

    PubMed Central

    Leickly, Emily; McDonell, Michael G.; Vilardaga, Roger; Angelo, Frank A.; Lowe, Jessica M.; McPherson, Sterling; Srebnik, Debra; Roll, John M.; Ries, Richard K.

    2015-01-01

    Background Immunoassay urine drug screening cups that detect use for two or more days are commonly used in addiction treatment settings. Until recently, there has been no comparable immunoassay test for alcohol use in these settings. Objectives The aim of this study was to assess the agreement of a commercially available ethyl glucuronide immunoassay (EtG-I) test conducted at an outpatient addiction clinic and lab-based EtG mass spectrometry (EtG-MS) conducted at a drug testing laboratory at three cut-off levels. High agreement between these two measures would support the usefulness of EtG-I as a clinical tool for monitoring alcohol use. Methods Forty adults with co-occurring alcohol dependence and serious mental illnesses submitted 1068 urine samples over a 16-week alcohol treatment study. All samples were tested using EtG-I on a benchtop analyzer and 149 were randomly selected for EtG-MS analysis at a local laboratory. Agreement was defined as the number of samples where EtG-I and EtG-MS were both above or below a specific cut-off level. Agreement was calculated at low cut-off levels (100 and 250 ng/ml), as well as at a higher cut-off level (500 ng/ml) recommended by most by commercial drug testing laboratories. Results Agreement between EtG-I and EtG-MS was high across all cut-off levels (90.6% at 100 ng/ml, and 96.6% at 250 and 500 ng/ml). Conclusions EtG immunoassays conducted at low cut-off levels in point-of-care testing settings have high agreement with lab-based EtG-MS. EtG-I can be considered a useful clinical monitoring tool for alcohol use in community-based addiction treatment settings. PMID:25695340

  7. Commercial Ethyl Glucuronide (EtG) and Ethyl Sulfate (EtS) Testing is not Vulnerable to Incidental Alcohol Exposure in Pregnant Women

    PubMed Central

    Beatty, Jessica R.; Rosano, Thomas G.; Strickler, Ronald C.; Graham, Amy E.; Sokol, Robert J.

    2016-01-01

    Background Ethyl Glucoronide (EtG) and Ethyl Sulfate (EtS) have shown promise as biomarkers for alcohol and may be sensitive enough for use with pregnant women in whom even low-level alcohol use is important. However, there have been reports of over-sensitivity of EtG and EtS to incidental exposure to sources such as alcohol-based hand sanitizer. Further, few studies have evaluated these biomarkers among pregnant women, in whom the dynamics of these metabolites may differ. Objectives This study evaluated whether commercial EtG-EtS testing was vulnerable to high levels of environmental exposure to alcohol in pregnant women. Methods Two separate samples of five nurses—one pregnant and the other postpartum, all of whom reported high levels of alcohol-based hand sanitizer use—provided urine samples before and 4–8 hours after rinsing with alcohol-based mouthwash and using hand sanitizer. The five pregnant nurses provided urine samples before, during, and after an 8-hour nursing shift, during which they repeatedly cleansed with alcohol-based hand sanitizer (mean 33.8 uses). The five postpartum nurses used hand sanitizer repeatedly between baseline and follow-up urine samples. Results No urine samples were positive for EtG-EtS at baseline or follow-up, despite use of mouthwash and—in the pregnant sample—heavy use of hand sanitizer (mean of 33.8 uses) throughout the 8-hour shift. Conclusions/Importance Current, commercially available EtG-EtS testing does not appear vulnerable to even heavy exposure to incidental sources of alcohol among pregnant and postpartum women. PMID:26771303

  8. Testing for ethanol markers in hair: discrepancies after simultaneous quantification of ethyl glucuronide and fatty acid ethyl esters.

    PubMed

    Kintz, P; Nicholson, D

    2014-10-01

    The hair of 97 cases were analysed for ethyl glucuronide (EtG) and fatty acid ethyl esters (FAEE, including ethyl myristate, ethyl palmitate, ethyl oleate and ethyl stearate) according to the Society of Hair Testing guidelines to examine the role of both tests in documenting chronic excessive alcohol drinking, particularly when the results are in contradiction. 27 (27.8%) results were EtG negative and FAEE positive, when applying the SoHT cut-offs, probably due to the use of alcohol-containing hair products. Four cases (4.1%) were EtG positive and FAEE negative that were attributed to the use of herbal lotions containing EtG.

  9. Determination of ethyl glucuronide and fatty acid ethyl esters in hair samples.

    PubMed

    Oppolzer, David; Barroso, Mário; Passarinha, Luís; Gallardo, Eugenia

    2017-04-01

    Hair testing for alcohol biomarkers is an important tool for monitoring alcohol consumption. We propose two methods for assessing alcohol exposure through combined analysis of ethyl glucuronide (EtG) and fatty acid ethyl esters (FAEEs) species (ethyl myristate, palmitate, stearate and oleate) in hair (30 mg). EtG was analysed by liquid chromatography-tandem mass spectrometry, while FAEEs were analysed by gas chromatography-tandem mass spectrometry using electron impact ionization. Both methods were validated according to internationally accepted guidelines. Linearity was proven between 3 and 500 pg/mg for EtG and 30-5000 pg/mg for FAEEs, and the limits of quantification were 3 pg/mg for EtG and 30 pg/mg for each of the four FAEEs. Precision and accuracy were considered adequate, processed EtG samples were found to be stable for up to 96 h left in the injector and processed FAEEs samples for up to 24 h. Matrix effects were not significant. Both methods were applied to the analysis of 15 authentic samples, using the cut-off values proposed by the Society of Hair Testing for interpretation. The results agreed well with the self-reported alcohol consumption in most cases, and demonstrated the suitability of the methods to be applied in routine analysis of alcohol biomarkers, allowing monitoring consumption using low sample amounts.

  10. Voucher-Based Reinforcement for Alcohol Abstinence Using the Ethyl-Glucuronide Alcohol Biomarker

    ERIC Educational Resources Information Center

    McDonell, Michael G.; Howell, Donelle N,; McPherson, Sterling; Cameron, Jennifer M.; Srebnik, Debra; Roll, John M.; Ries, Richard K.

    2012-01-01

    This study assessed the effects of a contingency management (CM) intervention for alcohol consumption in 10 alcohol-dependent participants. An ABCA design was used. Vouchers were provided contingent on results of ethyl glucuronide (EtG) urine tests (an alcohol biomarker with a 2-day detection period) and alcohol breath tests during the C phase.…

  11. Examination of sex differences in fatty acid ethyl ester and ethyl glucuronide hair analysis.

    PubMed

    Gareri, Joey; Rao, Chitra; Koren, Gideon

    2014-06-01

    Clinical studies examining performance of fatty acid ethyl esters (FAEE) and ethyl glucuronide (EtG) in identifying excessive alcohol consumption have been primarily conducted in male populations. An impact of hair cosmetics in producing both false-negative EtG results and false-positive FAEE results has been demonstrated, suggesting a possible bias in female populations. This study evaluates FAEE-positive hair samples (>0.50 ng/mg) from n = 199 female and n = 73 male subjects for EtG. Higher FAEE/EtG concordance was observed amongst male over female subjects. Performance of multiple proposed EtG cut-off levels were assessed; amongst female samples, FAEE/EtG concordance was 36.2% (30 pg/mg), 36.7% (27 pg/mg), and 43.7% (20 pg/mg). Non-coloured hair demonstrated a two-fold increase in concordance (41.8 v. 20.8%) over coloured hair in the female cohort. FAEE levels did not differ between male and female subjects; however they were lower in coloured samples (p = 0.046). EtG was lower in female subjects (p = 0.019) and coloured samples (p = 0.026). A total of n = 111 female samples were discordant. Amongst discordant samples (EtG-negative), 26% had evidence of recent alcohol use including consultation histories (n = 20) and detectable cocaethylene (n = 9); 29% of discordant samples were coloured. False-negative risk with ethyl glucuronide analysis in females was mediated by cosmetic colouring. These findings suggest that combined analysis of FAEE and EtG is optimal when assessing a female population and an EtG cut-off of 20 pg/mg is warranted when using combined analysis. While concordant FAEE/EtG-positive findings constitute clear evidence, discordant FAEE/EtG findings should still be considered suggestive evidence of chronic excessive alcohol consumption.

  12. Ethyl glucuronide, ethyl sulfate, and ethanol in urine after intensive exposure to high ethanol content mouthwash.

    PubMed

    Reisfield, Gary M; Goldberger, Bruce A; Pesce, Amadeo J; Crews, Bridgit O; Wilson, George R; Teitelbaum, Scott A; Bertholf, Roger L

    2011-06-01

    To determine the degree of ethanol absorption and the resultant formation and urinary excretion of its conjugated metabolites following intensive use of high ethanol content mouthwash, 10 subjects gargled with Listerine(®) antiseptic 4 times daily for 3¼ days. First morning void urine specimens were collected on each of the four study days and post-gargle specimens were collected at 2, 4, and 6 h after the final gargle of the study. Urine ethanol, ethyl glucuronide (EtG), ethyl sulfate (EtS), and creatinine were measured. Ethanol was below the positive threshold of 20 mg/dL in all of the urine specimens. EtG was undetectable in all pre-study urine specimens, but two pre-study specimens had detectable EtS (6 and 82 ng/mL; 16 and 83 μg/g creatinine). Only one specimen contained detectable EtG (173 ng/mL; 117 μg/g creatinine). EtS was detected in the urine of seven study subjects, but was not detected in the single specimen that had detectable EtG. The maximum EtS concentrations were 104 ng/mL and 112 μg/g creatinine (in different subjects). Three subjects produced a total of eight (non-baseline) urinary EtS concentrations above 50 ng/mL or 50 μg/g creatinine and three EtS concentrations exceeding 100 ng/mL or 100 μg/g creatinine. In patients being monitored for ethanol use by urinary EtG and EtS concentrations, currently accepted EtG and EtS cutoffs of 500 ng/mL are adequate to distinguish between ethanol consumption and four times daily use of high ethanol content mouthwash.

  13. Levels of ethyl glucuronide and ethyl sulfate in oral fluid, blood, and urine after use of mouthwash and ingestion of nonalcoholic wine.

    PubMed

    Høiseth, Gudrun; Yttredal, Borghild; Karinen, Ritva; Gjerde, Hallvard; Christophersen, Asbjørg

    2010-03-01

    The aim of this study is to investigate the concentrations of ethyl glucuronide (EtG) in oral fluid and both EtG and ethyl sulfate (EtS) in blood and urine following intense use of mouthwash and ingestion of nonalcoholic wine, which are proven to contain 3 mg/L EtG, 1.5 mg/L EtS, and 0.2 g/L ethanol. Twelve subjects participated in a controlled experiment. All subjects ingesting nonalcoholic wine showed urine samples negative for EtG but positive for EtS (Cmax 2.15 mg/L). All four subjects using mouthwash were negative for EtG and EtS in urine. All samples of oral fluid were negative for EtG and all samples of blood were negative for EtG and EtS. This study showed that ingestion of EtG and EtS as components of nonalcoholic wine lead to detection of urine EtS only, suggesting superior bioavailability of orally ingested EtS compared to EtG. This possibility of false-positive EtS results in urine after ingestion of nonalcoholic wine is important to remember when using EtG and EtS as relapse markers for alcohol. Finally, the study showed that a positive EtG or EtS result after accidental alcohol exposure is unlikely in blood and oral fluid.

  14. Stability of ethyl glucuronide in urine, post-mortem tissue and blood samples.

    PubMed

    Schloegl, Haiko; Dresen, Sebastian; Spaczynski, Karin; Stoertzel, Mylène; Wurst, Friedrich Martin; Weinmann, Wolfgang

    2006-03-01

    The stability of ethyl glucuronide (EtG) under conditions of degradation was examined in urine samples of nine volunteers and in post-mortem tissue (liver, skeletal muscle) and blood taken from seven corpses at autopsies. Analysis was performed via LC-MS/MS. EtG concentrations in urine samples ranged from 2.5 to 296.5 mg/l. When stored at 4 degrees C in airtight test tubes, EtG concentrations remained relatively constant; when stored at room temperature (RT) for 5 weeks in ventilated vials, variations of EtG concentrations ranged from a 30% decrease to an 80% increase, with an average of 37.5% increase. Liver and skeletal muscle tissue of three corpses with positive blood alcohol concentrations (BAC; ranging from 0.106 to 0.183 g%) were stored for 4 weeks and analysed periodically. EtG concentrations decreased 27.7% on average in 4 weeks storage at RT but EtG was still detectable in all samples with initial EtG concentrations higher than 1 mug/g. Blood and liver samples of four corpses with negative BACs were stored at RT after addition of 0.1 g% ethanol, and no new formation of EtG was observed.

  15. Influence of thermal hair straightening on ethyl glucuronide content in hair.

    PubMed

    Ettlinger, Jana; Kirchen, Luc; Yegles, Michel

    2014-06-01

    Hair analysis of ethyl glucuronide (EtG) has become a valuable marker for the detection of moderate and chronic alcohol consumption. It has been shown that bleaching and perming may decrease EtG content in hair. So far, no studies exist about the influence of thermal hair straightening on EtG content in hair. Forty-one positive EtG hair samples were treated in vitro with a hair straightener at 200°C. Duration of treatment of 1 min was chosen for this study. After washing, pulverization, incubation in ultrasonic bath, solid-phase extraction, and derivatization with heptafluorobutyric anhydride, EtG was determined by gas chromatography-mass spectrometry - negative ion chemical ionization (GC-MS-NICI). The EtG contents in straightened hair strands were then compared with those in the corresponding untreated strands. In 20 of 41 hair samples, a decrease of EtG content was found ranging from 0.7% to 79.3% (average 20%) whereas in 21 cases an increase was shown ranging from 2.0% to 50.9% (average 15%). The variation of the results seems to depend on hair colour. The decrease may be explained by thermic in vitro destruction of EtG. The increase may be explained by denaturation of the hair matrix by thermal treatment possibly causing a better extraction of EtG during incubation in ultrasonic bath. This in vitro study indicates that thermal hair straightening has an impact on the EtG content in hair. This has to be considered for a correct interpretation of EtG results in hair. However, these results should be confirmed by in vivo studies.

  16. Voucher-based reinforcement for alcohol abstinence using the ethyl-glucuronide alcohol biomarker.

    PubMed

    McDonell, Michael G; Howell, Donelle N; McPherson, Sterling; Cameron, Jennifer M; Srebnik, Debra; Roll, John M; Ries, Richard K

    2012-01-01

    This study assessed the effects of a contingency management (CM) intervention for alcohol consumption in 10 alcohol-dependent participants. An ABCA design was used. Vouchers were provided contingent on results of ethyl glucuronide (EtG) urine tests (an alcohol biomarker with a 2-day detection period) and alcohol breath tests during the C phase. The percentage of negative urines was 35% during the first baseline phase, 69% during the C phase, and 20% during the return-to-baseline phase. Results suggest that EtG urine tests may be a feasible method to deliver CM to promote alcohol abstinence.

  17. [Carbohydrate deficient transferrin and ethyl glucuronide: markers for alcohol use].

    PubMed

    Paling, Erik P; Mostert, Leendert J

    2013-01-01

    In this article, we report on the usefulness of physicians testing for carbohydrate deficient transferrin (CDT) and ethyl glucuronide (EtG) when there are doubts about alcohol use by their patients. A 44-year-old male consulted his general practitioner with depressive symptoms and denied using alcohol. Laboratory examination revealed an elevated CDT value. The latter was caused by chronic alcohol use. The second patient, a 32-year-old female with known alcohol dependence and receiving inpatient treatment at an addiction clinic, came back from leave. She denied having consumed alcohol and her blood alcohol concentration was zero. Examination of her urine showed an elevated EtG/creatinine ratio. This was caused by having had a few drinks during her leave and could not have been caused by using mouthwash or disinfection soap. We describe how to use the results of CDT and EtG testing in the therapeutic process and give recommendations for patient communication before performing these two tests.

  18. Stability of ethyl glucuronide in hair reference materials after accelerated aging.

    PubMed

    Ammann, D; Becker, R; Nehls, I

    2015-12-01

    Two different hair reference materials, one produced from authentic hair displaying an ethyl glucuronide (EtG) content of about 25 pg/mg and one obtained by fortification of blank hair to an EtG level of 85 pg/mg were submitted to accelerated aging between 4 °C and 60 °C for periods between one and 24 months. Subsequently, the EtG content was determined in the aged samples and untreated reference samples stored at -22 °C under repeatability conditions following the so-called isochronous approach. The EtG content remained stable even at 40 °C for 24 months and at 60 °C over six months. This is in contrast to many organic analytes contained in trace concentrations in diverse matrices. A slight but significant increase of the recovered EtG in case of authentic hair samples having been exposed for 24 months between 4 °C and 60 °C may be due to a temperature-driven process that allows increased recoveries of the physiologically embedded EtG.

  19. Combined use of fatty acid ethyl esters and ethyl glucuronide in hair for diagnosis of alcohol abuse: interpretation and advantages.

    PubMed

    Pragst, F; Rothe, M; Moench, B; Hastedt, M; Herre, S; Simmert, D

    2010-03-20

    In this study the combined use of fatty acid ethyl esters (FAEE) and ethyl glucuronide (EtG) for diagnoses of chronically excessive alcohol abuse is investigated at 174 hair samples from driving ability examination, workplace testing and child custody cases for family courts and evaluated with respect to the basics of interpretation. Using the cut-off values of 0.50 ng/mg for FAEE and 25 pg/mg for EtG, both markers were in agreement in 75% of the cases with 103 negative and 28 positive results and there were 30 cases with FAEE positive and EtG negative and 13 cases with FAEE negative and EtG positive. As the theoretical basis of interpretation, the pharmacokinetics of FAEE and EtG is reviewed for all steps between drinking of ethanol to incorporation in hair with particular attention to relationships between alcohol dose and concentrations in hair. It is shown that the concentrations of both markers are essentially determined by the area under the ethanol concentration in blood vs. time curve AUC(EtOH), despite large inter-individual variations. It is demonstrated by calculation of AUC(EtOH) on monthly basis for moderate, risky and heavy drinking that AUC(EtOH) increases very strongly in the range between 60 and 120 g ethanol per day. This specific feature which is caused by the zero-order elimination of ethanol is a favorable prerequisite for a high discrimination power of the hair testing for alcohol abuse. From the consideration of the different profiles of FAEE and EtG along the hair and in agreement with the literature survey, a standardized hair segment 0-3 cm is proposed with cut-off values of 0.5 ng/mg for FAEE and 30 pg/mg for EtG. This improves also the agreement between FAEE and EtG results in the cases of the present study. A scheme for combined interpretation of FAEE and EtG is proposed which uses the levels of abstinence and the double of the cut-off values as criteria in addition to the cut-off's. Considering the large variations in the relationship

  20. Ethyl-glucuronide and ethyl-sulfate in placental and fetal tissues by liquid chromatography coupled with tandem mass spectrometry.

    PubMed

    Morini, Luca; Falcón, Maria; Pichini, Simona; Garcia-Algar, Oscar; Danesino, Paolo; Groppi, Angelo; Luna, Aurelio

    2011-11-01

    The aim of this study was to develop a method for the determination of ethyl-glucuronide (EtG) and ethyl-sulfate (EtS), two direct ethanol metabolites, in early placental and fetal human tissues, as potential biomarkers of transplacental ethanol transfer from the mother to the fetus. Placental and fetal tissue samples were obtained from women undergoing voluntary termination of pregnancy at 12 weeks of gestation. Samples were deproteinized and directly injected into a liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) system. Limits of detection of 13.0 and 23.0 pmol/g and lower limits of quantification of 22.0 and 40.0 pmol/g were reached for EtG and EtS, respectively. Inter- and intraday imprecision and accuracy were always lower than 15%. The method was applied to 70 samples (35 placentas and 35 fetal tissues). Of 35 samples, 4 samples collected from 4 women tested positive for EtG and EtS, always showing higher concentrations for EtG. The placenta/fetal tissue ratio for EtG was 2.9 ± 0.9, whereas EtS showed a ratio of 1.7 ± 0.7. Preliminary results suggest that these metabolites are present in both tissues. Further studies should now corroborate the hypothesis, not yet confirmed, that transplacental transfer of ethanol takes place not only for the parent compound but also for EtG and EtS.

  1. Urine tested positive for ethyl glucuronide and ethyl sulphate after the consumption of "non-alcoholic" beer.

    PubMed

    Thierauf, Annette; Gnann, Heike; Wohlfarth, Ariane; Auwärter, Volker; Perdekamp, Markus Grosse; Buttler, Klaus-Juergen; Wurst, Friedrich M; Weinmann, Wolfgang

    2010-10-10

    In abstinence maintenance programs, for reissuing the driving licence and in workplace monitoring programs abstinence from ethanol and its proof are demanded. Various monitoring programs that mainly use ethyl glucuronide (EtG) as alcohol consumption marker have been established. To abstain from ethanol, but not from the taste of alcoholic beverages, in particular non-alcoholic beer has become more and more popular. In Germany, these "alcohol-free" beverages may still have an ethanol content of up to 0.5vol.% without the duty of declaration. Due to severe negative consequences resulting from positive EtG tests, a drinking experiment with 2.5L of non-alcoholic beer per person was performed to address the question of measurable concentrations of the direct metabolites EtG and EtS (ethyl sulphate) in urine and blood. Both alcohol consumption markers - determined by LC-MS/MS - were found in high concentrations: maximum concentrations in urine found in three volunteers were EtG 0.30-0.87mg/L and EtS 0.04-0.07mg/L, i.e., above the often applied cut-off value for the proof of abstinence of 0.1mg EtG/L. In the urine samples of one further volunteer, EtG and EtS concentrations cumulated over-night and reached up to 14.1mg/L EtG and 16.1mg/L EtS in the next morning's urine. Ethanol concentrations in blood and urine samples were negative (determined by HS-GC-FID and by an ADH-based method).

  2. Determination of ethyl glucuronide in human hair by hydrophilic interaction liquid chromatography-tandem mass spectrometry.

    PubMed

    Yaldiz, Fadile; Daglioglu, Nebile; Hilal, Ahmet; Keten, Alper; Gülmen, Mete Korkut

    2013-10-01

    Ethyl glucuronide (EtG) is a direct metabolite of ethanol and has been utilized as a marker for alcohol intake. This study presents development, validation and application of a new hydrophilic interaction liquid chromatography-tandem mass spectrometry (HILIC-MS/MS) method for the analysis of EtG in human hair samples. The linearity was assessed in the range of 5-2000 pg/mg hair, with a correlation coefficient of >0.99. The method was selective and sensitive, with a limit of detection (LOD) and limit of quantitation (LOQ) of 0.05 pg/mg and 0.18 pg/mg in hair, respectively. Differently from the extraction procedures in the literature, a fast and simple liquid-liquid method was used and highest recoveries and cleanest extracts were obtained. The method was successfully applied to 30 human hair samples which were taken from those who state they consume alcohol. EtG concentrations in the hair samples of alcohol users participated in this study, ranged between 1.34 and 82.73 pg/mg. From the concentration of EtG in hair strands 20 of the 30 subjects can be considered regular moderate drinkers.

  3. The influence of ethanol containing cosmetics on ethyl glucuronide concentration in hair.

    PubMed

    Martins Ferreira, Liliane; Binz, Tina; Yegles, Michel

    2012-05-10

    Ethyl glucuronide (EtG) and fatty acid ethyl esters (FAEE), non-volatile, direct metabolites of ethanol have been shown to be suitable markers for the evaluation of social and chronic excessive alcohol consumption. Previous investigations have shown that the regular use of hair-care products with high alcohol content lead to an increase of FAEE concentration and consequently gave false-positive results for the determination of FAEE in hair. In this study we investigated the influence of a long-term hair treatment with EtOH containing lotion, on the EtG concentrations in hair. In this study 7 volunteer subjects (classified as either rare, social or heavy drinkers) treated the right side of their scalp every day during a one or two month period with a commercial hair tonic (Seborin), which contains 44.0% ethanol (vol%). Collection of hair specimens from both sides of the scalp was done one day before hair treatment, one week and one month after treatment (for 5 subjects also after two months of treatment). A hair segment of 3 centimeters (cm) was cut and then washed with water and acetone, and then pulverized. EtG was quantified by GC/MS after pulverization and 2h of ultrasonication in water, extraction by solid phase extraction using Oasis MAX columns and derivatization with HFBA. Measurements were done in negative chemical ionization mode using EtG-D5 as internal standard. Comparison of EtG concentration in the treated and in the non-treated hair specimens did not show any increase at the different dates of collection for the 7 subjects. In conclusion, these results show that there is no indication for an increase of EtG after use of ethanol containing hair cosmetics.

  4. Utility of urinary ethyl glucuronide analysis in post-mortem toxicology when investigating alcohol-related deaths.

    PubMed

    Sundström, M; Jones, A W; Ojanperä, I

    2014-08-01

    Use and abuse of alcohol are common findings when unnatural deaths are investigated as evidenced by high blood- and urine- alcohol concentrations (BAC and UAC) at autopsy. Because ethanol is metabolized in the liver until the time of death, the autopsy BAC or UAC might be negative even though the deceased had consumed alcohol in the immediate ante-mortem period. Analysis of the non-oxidative metabolite of ethanol [ethyl glucuronide (EtG)] offers a more sensitive test of recent drinking. In this paper, we determined the concentrations of ethanol and EtG in urine samples from 972 consecutive forensic autopsies. In 425 cases (44%) both EtG and ethanol were positive, which supports ante-mortem drinking. In 342 cases (35%), both EtG and ethanol was negative, which speaks against any consumption of alcohol just before death. In 181 cases, ethanol was negative in urine (<0.2 g/kg), whereas EtG was positive (>0.5 mg/L), which points towards ingestion of alcohol some time before death. In these cases, mean and median concentrations of EtG were 53.2 mg/L and 23.7 mg/L, respectively, although there was no mention of alcohol on 131 of the death certificates. Alcohol was mentioned on death certificates as an underlying or immediate cause of death or a contributing factor in 435 (45%) cases, which rose to 566 (58%) cases when positive EtG results were included. This article demonstrates the usefulness of EtG analysis in routine post-mortem toxicology when ante-mortem drinking and alcohol-related deaths are investigated.

  5. Determination of ethyl glucuronide in hair to assess excessive alcohol consumption in a student population.

    PubMed

    Oppolzer, David; Barroso, Mário; Gallardo, Eugenia

    2016-03-01

    Hair analysis for ethyl glucuronide (EtG) was used to evaluate the pattern of alcohol consumption amongst the Portuguese university student population. A total of 975 samples were analysed. For data interpretation, the 2014 guidelines from the Society of Hair Testing (SoHT) for the use of alcohol markers in hair for the assessment of both abstinence and chronic excessive alcohol consumption were considered. EtG concentrations were significantly higher in the male population. The effect of hair products and cosmetics was evaluated by analysis of variance (ANOVA), and significant lower concentrations were obtained when conditioner or hair mask was used or when hair was dyed. Based on the analytical data and information obtained in the questionnaires from the participants, receiver operating characteristic (ROC) curves were constructed in order to determine the ideal cut-offs for our study population. Optimal cut-off values were estimated at 7.3 pg/mg for abstinence or rare occasional drinking control and 29.8 pg/mg for excessive consumption. These values are very close to the values suggested by the SoHT, proving their adequacy to the studied population. Overall, the obtained EtG concentrations demonstrate that participants are usually well aware of their consumption pattern, correlating with the self-reported consumed alcohol quantity, consumption habits and excessive consumption close to the time of hair sampling.

  6. Ethyl glucuronide concentrations in hair: a controlled alcohol-dosing study in healthy volunteers.

    PubMed

    L Crunelle, Cleo; Cappelle, Delphine; Yegles, Michel; De Doncker, Mireille; Michielsen, Peter; Dom, Geert; van Nuijs, Alexander L N; Maudens, Kristof E; Covaci, Adrian; Neels, Hugo

    2016-03-01

    Ethyl glucuronide (EtG) is a minor phase II metabolite of alcohol that accumulates in hair. It has been established as a sensitive marker to assess the retrospective consumption of alcohol over recent months using a cut-off of ≥7 pg/mg hair to assess repeated alcohol consumption. The primary aim was to assess whether amounts of alcohol consumed correlated with EtG concentrations in hair. Additionally, we investigated whether the current applied cut-off value of 7 pg/mg hair was adequate to assess the regular consumption of low-to-moderate amounts of alcohol. A prospective controlled alcohol-dosing study in 30 healthy individuals matched on age and gender. Individuals were instructed to drink no alcohol (N = 10), 100 g alcohol per week (N = 10) or 150 g alcohol per week (N = 10) for 12 consecutive weeks, before and after which hair was collected. Throughout the study, compliance to daily alcohol consumption was assessed by analyzing urine EtG three times weekly. Participants in the non-drinking group had median EtG concentrations of 0.5 pg/mg hair (interquartile range (IQR) 1.7 pg/mg; range < 0.21-4.5 pg/mg). Participants consuming 100 and 150 g alcohol per week showed median EtG concentrations of 5.6 pg/mg hair (IQR 4.7 pg/mg; range 2.0-9.8 pg/mg) and 11.3 pg/mg hair (IQR 5.0 pg/mg; range 7.7-38.9 pg/mg), respectively. Hair EtG concentrations between the three study groups differed significantly from one another (p < 0.001). Hair EtG concentrations can be used to differentiate between repeated (low-to-moderate) amounts of alcohol consumed over a long time period. For the assessment of repeated alcohol use, we propose that the current cut-off of 7 pg/mg could be re-evaluated.

  7. Immunoassay for ethyl glucuronide in vitreous humor: a new tool for postmortem diagnostics of alcohol use.

    PubMed

    Rainio, Juha; Kultti, Johanna; Kangastupa, Päivikki; Tuomi, Heidi; Ahola, Sanna; Karhunen, Pekka J; Helander, Anders; Niemelä, Onni

    2013-03-10

    Although excessive alcohol consumption plays a major role in fatal events, the role of alcohol use as a possible contributing factor at the time of death is not easy to establish due to lack of suitable biomarkers for postmortem analyses. We used an immunological approach to measure ethyl glucuronide (EtG) concentrations from vitreous humor (VH) and serum from 58 individuals representing a forensic autopsy population of cases with either a well-documented history of excessive alcohol use (n=37) or cases without such history (n=21), according to medical and police records and blood alcohol determinations (BAC). The immunoassay was based on the Microgenics DRI-EtG EIA reagents applied on an automated Abbott Architect c8000 clinical chemistry analyzer. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) determination of EtG and ethyl sulfate (EtS) was used as a reference method. At a cut-off of 0.3mg/l for VH-EtG, the immunoassay correctly identified 92% of the cases with a history of excessive alcohol use, whereas the BAC was positive (cut-off 10mg/dl) in 68% of the cases. A significant correlation emerged between VH-EtG and serum EtG (r=0.77, p<0.001) and between VH-EtG and BAC (r=0.62, p<0.001), although VH-EtG was frequently elevated also in cases with no detectable BAC. The EtG immunoassay showed a strong correlation with the LC-MS/MS reference method (r=0.94, p<0.001) and there was 100% agreement in the frequency of marker positive and negative findings between the immunoassay EtG results and the LC-MS/MS analysis of EtG and EtS. The present data indicate that the immunoassay for VH-EtG is a useful forensic tool for screening of antemortem alcohol use.

  8. Distribution Pattern of Ethyl Glucuronide and Caffeine Concentrations Over the Scalp of a Single Person in a Forensic Context.

    PubMed

    Meier, Ulf; Briellmann, Thomas; Scheurer, Eva; Dussy, Franz

    2017-03-11

    The distribution of analyte concentrations in hair across the scalp has not been thoroughly investigated. Differences in concentrations depending on sampling location are problematic, especially when measuring a second strand to confirm the result of the first measurement. Aiming at a better understanding of the concentration differences, the distribution of ethyl glucuronide (EtG) and caffeine concentrations in hair across the entire head of one test subject was investigated by dividing the scalp completely into regions of ca. 2 x 2 cm area, yielding a total of 104 samples. For the quantification of EtG, a novel LC-MS(3) /MRM method was developed and validated with a limit of detection and limit of quantification of 2 and 4 pg/mg, respectively. Large variations of the concentration across the head were found, with a factor of ca. 3.0 and 10.6 for EtG and caffeine, respectively. These differences could not be attributed to measurement error alone. The concentrations were projected onto the subjects head, and concentration patterns were identified for EtG and caffeine. When examining multiple strands from within one 2 x 2 cm sampling area, the strands showed similar concentrations. Segmental analysis of selected 3 cm strands showed decreasing concentrations of EtG and caffeine from proximal to distal end, possibly due to wash out of the analytes.

  9. The influence of cleansing shampoos on ethyl glucuronide concentration in hair analyzed with an optimized and validated LC-MS/MS method.

    PubMed

    Binz, Tina M; Baumgartner, Markus R; Kraemer, Thomas

    2014-11-01

    Ethyl glucuronide (EtG) is widely used as a marker for assessment of alcohol consumption behavior. In this study the influence of special cleansing shampoos on ethyl glucuronide concentrations in hair was investigated. For that purpose an optimized LC-MS/MS method was developed using a Hypercarb™ porous graphitic carbon (PGC) column and validated according to the guidelines of the German Society of Toxicological and Forensic Chemistry (GTFCh). Twenty-five hair samples of persons with known alcohol consumption behavior were investigated (21 positive samples and 4 blank samples). The hair samples were divided into two strands of hair and were analyzed after treatment with one out of four cleansing shampoos and without shampoo treatment. EtG concentrations in hair did not show any significant differences after a single application of the different cleansing shampoos. EtG was still detectable in all the positive hair samples without significant concentration change. These results clearly demonstrated that a single application of the tested cleansing shampoos did not remove EtG from hair and therefore had no influence on EtG concentration in analytical hair analysis.

  10. Ethyl glucuronide in hair and fingernails as a long-term alcohol biomarker

    PubMed Central

    Berger, Lisa; Fendrich, Michael; Jones, Joseph; Fuhrmann, Daniel; Plate, Charles; Lewis, Douglas

    2014-01-01

    Aims This study aimed to evaluate the performance of ethyl glucuronide (EtG) in hair and fingernails as a long-term alcohol biomarker. Design Cross-sectional survey with probability sampling. Setting Midwestern United States. Participants Participants were 606 undergraduate college students between the ages of 18 and 25 years at the time of selection for potential study participation. Measurements EtG concentrations in hair and fingernails were measured by liquid chromatography-tandem mass spectrometry at three thresholds [30 picograms (pg) per milligram (mg); 20 pg/mg; and 8 pg/mg]. Any weekly alcohol use, increasing-risk drinking and high-risk drinking on average during the past 12 weeks was assessed by participant interview using the time-line follow-back method. Findings In both hair and fingernails at all three EtG thresholds, sensitivity was greatest for the high-risk drinking group [hair: 0.43, confidence interval (CI) = 0.17, 0.69 at 30 pg/mg, 0.71, CI = 0.47, 0.95 at 20 pg/mg; 0.93, CI = 0.79, 1.00 at 8 pg/mg; fingernails: 1.00, CI = 1.00–1.00 at 30, 20 and 8 pg/mg] and specificity was greatest for any alcohol use (hair: 1.00, CI = 1.00, 1.00 at 30 and 20 pg/mg; 0.97, CI = 0.92–0.99 at 8 pg/mg; fingernails: 1.00, CI = 1.00–1.00 at 30, 20 and 8 pg/mg). Areas under the receiver operating characteristic curves were significantly higher for EtG concentration in fingernails than hair for any weekly alcohol use (P = 0.02, DeLong test, two-tailed) and increasing-risk drinking (P = 0.02, DeLong test, two-tailed). Conclusions Ethyl glucuronide, especially in fingernails, may have potential as a quantitative indicator of alcohol use. PMID:24524319

  11. Practical experiences in application of hair fatty acid ethyl esters and ethyl glucuronide for detection of chronic alcohol abuse in forensic cases.

    PubMed

    Suesse, S; Pragst, F; Mieczkowski, T; Selavka, C M; Elian, A; Sachs, H; Hastedt, M; Rothe, M; Campbell, J

    2012-05-10

    This article presents results from 1872 hair samples, which were analyzed for fatty acid ethyl esters (FAEEs) and ethyl glucuronide (EtG). The results were evaluated in the context of self-reported drinking behavior, the use of hair cosmetics, the gender of the sample donors and hair sample length. For comparison, CDT and GGT in serum were available in 477 and 454 cases, respectively. A number of alcohol abstainers or low moderate drinkers and excessive drinkers were selected for assessment of cut-offs for FAEEs in the proximal 6cm hair segments and for EtG in the proximal 3cm hair segments. Cut-off values were assessed by ROC analysis. It was found that the cut-offs of 1.0ng/mg FAEE and 30pg/mg EtG presently used for excessive drinking lead to a low portion of false positives (4% and 3% respectively) but to a higher portion of false negatives (23% and 25% respectively). Comparison of the mean and medium concentrations in samples without any reported hair cosmetics (N=1079) and in samples with reported use of hair spray (N=79) showed an increase by the factor of about two for FAEE but no significant difference for EtG. Mean values of EtG were decreased by 80% in bleached samples (N=164) and by 63% in dyed samples (N=96). There was no significant effect of bleaching and dyeing on FAEE. Hair gel and hair wax, oil or grease showed no significant effect on both FAEE and EtG. With respect to gender and investigated hair length ambiguous results were obtained because of major differences in the compared subpopulations of male with higher alcohol consumption and mainly shorter hair, and less drinking female with longer hair. For excessive drinkers FAEEs in the 0-6cm hair segment and EtG in the 0-3cm segment decreased with increasing time of reported abstinence before sample collection. These drinkers attain the level of teetotalers only after more than 10 months of abstinence. In comparison to scalp hair, FAEEs recovered from armpit hair and leg hair were lower and from

  12. Improved liquid chromatography-tandem mass spectrometric method for the determination of ethyl glucuronide concentrations in hair: applications to forensic cases.

    PubMed

    Imbert, Laurent; Gaulier, Jean-Michel; Dulaurent, Sylvain; Morichon, Julien; Bevalot, Fabien; Izac, Paul; Lachâtre, Gérard

    2014-01-01

    Ethyl glucuronide (EtG) is a direct marker of ethanol consumption, and its assay in hair is an efficient tool for chronic alcoholism diagnosis. In 2012, the Society of Hair Testing proposed a new consensus for hair concentrations interpretation, strongly advising the use of analytical methods providing a limit of quantification of less than 3 pg/mg. The present work describes the optimization and validation of a previously developed liquid chromatography-tandem mass spectrometric method in order to comply with this recommendation. The concentration range of this improved method is from 3 to 1,000 pg/mg. Some cases are then described to illustrate the usefulness of hair EtG: a forensic post-mortem case and two cases of suspension of driving licences. Finally, hair samples of some teetotallers (n = 10) have been analyzed, which allowed neither to quantitate nor to detect any trace of EtG.

  13. Validation of a novel method to identify in utero ethanol exposure: simultaneous meconium extraction of fatty acid ethyl esters, ethyl glucuronide, and ethyl sulfate followed by LC-MS/MS quantification.

    PubMed

    Himes, Sarah K; Concheiro, Marta; Scheidweiler, Karl B; Huestis, Marilyn A

    2014-03-01

    Presence of fatty acid ethyl esters (FAEE), ethyl glucuronide (EtG), and ethyl sulfate (EtS) in meconium, the first neonatal feces, identifies maternal alcohol consumption during pregnancy. Current meconium alcohol marker assays require separate analyses for FAEE and EtG/EtS. We describe development and validation of the first quantitative liquid chromatography tandem mass spectrometry assay for 9 FAEEs, EtG, and EtS in 100 mg meconium. For the first time, these alcohol markers are analyzed in the same meconium aliquot, enabling comparison of the efficiency of gestational ethanol exposure detection. 100 mg meconium was homogenized in methanol and centrifuged. The supernatant was divided, and applied to two different solid phase extraction columns for optimized analyte recovery. Limits of quantification for ethyl laurate, myristate, linolenate, palmitoleate, arachidonate, linoleate, palmitate, oleate, and stearate ranged from 25-50 ng/g, with calibration curves to 2,500-5,000 ng/g. EtG and EtS linear dynamic ranges were 5-1,000 and 2.5-500 ng/g, respectively. Mean bias and between-day imprecision were <15 %. Extraction efficiencies were 51.2-96.5 %. Matrix effects ranged from -84.7 to 16.0 %, but were compensated for by matched deuterated internal standards when available. All analytes were stable (within ±20 % change from baseline) in 3 authentic positive specimens, analyzed in triplicate, after 3 freeze/thaw cycles (-20 °C). Authentic EtG and EtS also were stable after 12 h at room temperature and 72 h at 4 °C; some FAEE showed instability under these conditions, although there was large inter-subject variability. This novel method accurately detects multiple alcohol meconium markers and enables comparison of markers for maternal alcohol consumption.

  14. Validation of a novel method to identify in utero ethanol exposure: simultaneous meconium extraction of fatty acid ethyl esters, ethyl glucuronide, and ethyl sulfate followed by LC-MS/MS quantification

    PubMed Central

    Himes, Sarah K.; Concheiro, Marta; Scheidweiler, Karl B.

    2015-01-01

    Presence of fatty acid ethyl esters (FAEE), ethyl glucuronide (EtG), and ethyl sulfate (EtS) in meconium, the first neonatal feces, identifies maternal alcohol consumption during pregnancy. Current meconium alcohol marker assays require separate analyses for FAEE and EtG/EtS. We describe development and validation of the first quantitative liquid chromatography tandem mass spectrometry assay for 9 FAEEs, EtG, and EtS in 100 mg meconium. For the first time, these alcohol markers are analyzed in the same meconium aliquot, enabling comparison of the efficiency of gestational ethanol exposure detection. 100 mg meconium was homogenized in methanol and centrifuged. The supernatant was divided, and applied to two different solid phase extraction columns for optimized analyte recovery. Limits of quantification for ethyl laurate, myristate, linolenate, palmitoleate, arachidonate, linoleate, palmitate, oleate, and stearate ranged from 25–50 ng/g, with calibration curves to 2,500–5,000 ng/g. EtG and EtS linear dynamic ranges were 5–1,000 and 2.5–500 ng/g, respectively. Mean bias and between-day imprecision were <15 %. Extraction efficiencies were 51.2–96.5 %. Matrix effects ranged from −84.7 to 16.0 %, but were compensated for by matched deuterated internal standards when available. All analytes were stable (within ±20 % change from baseline) in 3 authentic positive specimens, analyzed in triplicate, after 3 freeze/thaw cycles (−20 °C). Authentic EtG and EtS also were stable after 12 h at room temperature and 72 h at 4 °C; some FAEE showed instability under these conditions, although there was large inter-subject variability. This novel method accurately detects multiple alcohol meconium markers and enables comparison of markers for maternal alcohol consumption. PMID:24408304

  15. Urine ethyl glucuronide and ethyl sulphate using liquid chromatography-tandem mass spectrometry in a routine clinical laboratory.

    PubMed

    Armer, Jane M; Allcock, Rebecca L

    2017-01-01

    Background Detection of alcohol consumption in clients undergoing treatment for alcohol dependence can be difficult. The ethanol metabolites ethyl glucuronide and ethyl sulphate are detectable for longer in urine than either breath ethanol or urine ethanol. Our aim was to develop a liquid chromatography-tandem mass spectrometry method for urine ethyl glucuronide and ethyl sulphate for use in a routine clinical laboratory and define clinical cut-offs in a large population who had not consumed alcohol for at least two weeks. Methods Urine samples were diluted in 0.05% formic acid in HPLC grade water and then directly injected onto a Waters Acquity ultra high performance liquid chromatography coupled to a Waters TQ Detector. Eighty participants were recruited who had not consumed alcohol for at least two weeks to define cut-offs for urine ethyl glucuronide and ethyl sulphate. Samples and alcohol diaries were also collected from 12 alcohol-dependent clients attending a treatment programme. Results The assay was validated with a lower limit of quantitation of 0.20 mg/L for ethyl glucuronide and 0.04 mg/L for ethyl sulphate. Accuracy, precision, linearity and recovery were acceptable. Cut-offs were established for ethyl glucuronide, ethyl sulphate and ethyl sulphate/creatinine ratio (≤0.26 mg/L, ≤0.22 mg/L and ≤0.033 mg/mmol, respectively) in a non-drinking population. The validated cut-offs correctly identified clients in alcohol treatment who were continuing to drink alcohol. Conclusions A simple liquid chromatography-tandem mass spectrometry method for urine ethyl glucuronide and ethyl sulphate has been validated and cut-offs defined using 80 participants who had not consumed alcohol for at least two weeks. This is the largest study to date to define cut-offs for ethyl glucuronide, ethyl sulphate and ethyl sulphate/creatinine ratio.

  16. Degradation of the ethyl glucuronide content in hair by hydrogen peroxide and a non-destructive assay for oxidative hair treatment using infra-red spectroscopy.

    PubMed

    Ammann, Dominic; Becker, Roland; Kohl, Anka; Hänisch, Jessica; Nehls, Irene

    2014-11-01

    The assessment of quantification results of the alcohol abuse marker ethyl glucuronide (EtG) in hair in comparison to the cut-off values for the drinking behavior may be complicated by cosmetic hair bleaching. Thus, the impact of increasing exposure to hydrogen peroxide on the EtG content of hair was investigated. Simultaneously, the change of absorbance in the range of 1000-1100 cm(-1) indicative for the oxidation of cystine was investigated non-destructively by attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR) using pulverized portions of the respective hair samples. Hair samples treated with hydrogen peroxide consistently displayed a significantly increased absorbance at 1040 cm(-1) associated with the formation of cysteic acid. The EtG content decreased significantly if the hair was treated with alkaline hydrogen peroxide as during cosmetic bleaching. It could be shown that ATR-FTIR is capable of detecting an exposure to hydrogen peroxide when still no brightening was visible and already before the EtG content deteriorated significantly. Thus, hair samples suspected of having been exposed to oxidative treatment may be checked non-destructively by a readily available technique. This assay is also possible retrospectively after EtG extraction and using archived samples.

  17. Impact of the grinding process on the quantification of ethyl glucuronide in hair using a validated UPLC-ESI-MS-MS method.

    PubMed

    Kummer, Natalie; Wille, Sarah M R; Di Fazio, Vincent; Ramírez Fernández, Maria Del Mar; Yegles, Michel; Lambert, Willy E E; Samyn, Nele

    2015-01-01

    The Society of Hair Testing (SoHT) has provided cutoffs for the quantification of ethyl glucuronide (EtG) in hair to indicate occasional or chronic/excessive alcohol consumption. Although several sensitive methods have been reported, past proficiency test results show a lack of reproducibility. An ultra-performance liquid chromatographic mass spectrometric method (LLOQ of 10 pg EtG/mg hair) has been validated according to the international guidelines, including the successful participation in five proficiency tests. This method was subsequently used to evaluate the impact of different grinding conditions (cut, weakly or extensively pulverized hair samples) on the final measured EtG concentration. Hair from alcohol consumers (n = 2) and commercially available quality control samples (QCs) (n = 2) was used. For the QCs, extensive pulverization led to a significantly higher amount of measured EtG. In the hair samples obtained from volunteers, cut or weakly pulverized hair resulted in EtG concentrations below the LLOQ, while the mean concentrations of 14 and 40 pg EtG/mg hair were determined after extensive pulverization. Differences in sample preparation could partially explain inter-laboratory variability. As the differences in results can lead to a different interpretation even when applying the SoHT cutoffs, it is of interest to standardize sample preparation techniques in the field of EtG hair testing.

  18. Microwave-assisted extraction: a simpler and faster method for the determination of ethyl glucuronide in hair by gas chromatography-mass spectrometry.

    PubMed

    Alvarez, Iván; Bermejo, Ana María; Tabernero, María Jesús; Fernández, Purificación; Cabarcos, Pamela; López, Patricia

    2009-02-01

    Alcohol is the most frequently abused "addictive substance" that causes serious social problems throughout the world; thus, alcoholism is of particular interest in clinical and forensic medicine. Alcohol biomarkers are physiological indicators of alcohol exposure or ingestion and may reflect the presence of an alcohol use disorder. The glucuronide conjugation is a minor pathway of ethanol metabolism. Ethyl glucuronide (EtG) is a marker of recent alcohol consumption that detects alcohol use reliably over a definite time period. The present paper describes a new method for the determination of EtG in hair. It is based both in the microwave-assisted extraction (MAE), to extract the analyte from hair samples, and gas chromatography-mass spectrometry (GC-MS), to identify and quantify the EtG in selected ion monitoring (SIM) mode. The method was applied to 15 hair samples from occasional alcohol users, obtaining positive results in all cases. It was fully validated, including a linear range (0.3-10 ng/mg) and the main precision parameters. In summary, the use of microwave-assisted extraction turned out to be a substantially simpler, faster, and a more sensitive procedure than any other conventional sample preparations.

  19. Do drug users use less alcohol than non-drug users? A comparison of ethyl glucuronide concentrations in hair between the two groups in medico-legal cases.

    PubMed

    Paul, Richard; Kingston, Robert; Tsanaclis, Lolita; Berry, Anthony; Guwy, Alan

    2008-03-21

    Two groups were selected from the remainder of hair samples that had been tested for drugs at TrichoTech for medico-legal cases: samples that tested negative (drug-negative group; N=42, age 33.4+/-7.2 years) and samples that tested positive for drugs (drug-positive group; N=57, age 32.5+/-8.8 years). A rapid, simple method to detect the ethanol metabolite, ethyl glucuronide (EtG) in hair has been developed. The hair samples were sectioned, and then submitted to overnight sonication in water. Samples then underwent SPE using anion exchange cartridges, followed by derivatisation with N,O-bis[trimethylsilyl]trifluoroacetamide (BSTFA), before confirmation by GC-MS/MS. The assay produced excellent linearity and sensitivity over the calibration range 0.02-1.0 ng/mg, assuming a 10 mg hair sample. The mean age of the two groups was not statistically different (p=0.575, Student t-test), indicating a homogeneous group. Twelve of the 57 (21.0%) hair samples of the drug-positive group tested positive for EtG, and 17 of the 42 (40.5%) hair samples of the drug-negative group tested positive for EtG. The mean concentration of EtG in the drug-positive group was 0.011 ng/mg compared to 0.107 ng/mg in the drug-negative group. When the full results of this study were subjected to statistical analysis it was shown that EtG levels in the drug-negative group were statistically higher than those found in the drug-positive group (p<0.05). This preliminary finding may be of use in the study of addiction and adds valuable data to previous studies regarding the use of EtG as a valuable marker for alcohol levels in hair.

  20. Effect of succinic acid and tween-80 on glucuronidation of 2-ethyl-6-methyl-3-hydroxypyridine.

    PubMed

    Baranov, P A; Kravtsova, O U; Sariev, A K; Sherdev, V P

    2008-07-01

    We studied the effect of succinic acid on the process of glucuronidation of 2-ethyl-6-methyl-3-hydroxypyridine after peroral and intraperitoneal administration in the form of succinate or a base. Since the basic form of 2-ethyl-6-methyl-3-hydroxypyridine is insoluble in water, it was administered in 5% Tween-80. It was necessary to evaluate also the effect of Tween-80 on glucuronidation of 2-ethyl-6-methyl-3-hydroxypyridine in different administration routes. Quantitative assay of glucuronidated fractions was performed by the method of reversed-phase HPLC with fluorometrical detection. The detection limit for this method was 10 ng/ml. We confirmed that the major excretion pathway for 2-ethyl-6-methyl-3-hydroxypyridine is conjugation with glucuronic acid. It was found that succinic acid increased excretion of glucuronidated metabolite after both peroral and intraperitoneal administration of 2-ethyl-6-methyl-3-hydroxypyridine in the form of succinate and base in 5% Tween-80. The effect of Tween-80 was detected only after peroral administration, which was probably related to its effect on absorption of this compound. Tween-80 increased excretion of glucuronate after peroral administration of 2-ethyl-6-methyl-3-hydroxypyridine in the form of succinate and in 5% Tween solution.

  1. Determination of ethyl glucuronide in hair samples of Chinese people by protein precipitation (PPT) and large volume injection-gas chromatography-tandem mass spectrometry (LVI-GC/MS/MS).

    PubMed

    Shi, Yan; Shen, Baohua; Xiang, Ping; Yan, Hui; Shen, Min

    2010-11-15

    Ethyl glucuronide (EtG) has been shown to be a suitable marker of excessive alcohol consumption. Determination of EtG in hair samples may help to differentiate social drinkers from alcoholics, and this testing can be widely used in forensic science, treatment programs, workplaces, military bases as well as driving ability test to provide legal proof of drinking. A method for determination of EtG in hair samples using large volume injection-gas chromatography-tandem mass spectrometry (LVI-GC/MS/MS) was developed and validated. Hair samples (in 1 mL deionized water) were ultrasonicated for 1h and incubated overnight; these samples were then deproteinated to remove impurities and derivatisated with 15 μL of pyridine and 30 μL of BSTFA. EtG was detected using GC/MS/MS in multiple-reaction monitoring mode. This method exhibited good linearity: y=0.0036 x+0.0437, R²=0.9993, the limit of detection and the limit of quantification were 5 pg/mg and 10 pg/mg, respectively. The extraction recoveries were more than 60%, and the inter-day and intra-day relative standard deviations (RSD) were less than 15%. This method has been applied to the analysis of EtG in hair samples from 21 Chinese subjects. The results for samples obtained from all of those who were teetotallers were negative, and the results for the other 15 samples ranged from 10 to 78 pg/mg, except for one negative sample. These data are the basis for interpretation of alcohol abuse.

  2. Liquid Chromatography-Tandem Mass Spectrometry Assay to Detect Ethyl Glucuronide in Human Fingernail: Comparison to Hair and Gender Differences

    PubMed Central

    Jones, Joseph; Jones, Mary; Plate, Charles; Lewis, Douglas; Fendrich, Michael; Berger, Lisa; Fuhrmann, Daniel

    2015-01-01

    Over the past decade, the use of hair specimens for the long-term detection of the alcohol biomarker ethyl glucuronide has been increasing in popularity and usage. We evaluated the usefulness of fingernail clippings as a suitable alternative to hair for ethyl glucuronide detection. A liquid chromatography-tandem mass spectrometry method for the detection of ethyl glucuronide in fingernail clippings was fully validated and used to analyze the hair and/or fingernail specimens of 606 college-aged study participants. The limit of detection was 2 pg/mg, the limit of quantitation was 8 pg/mg and the method was linear from 8 to 2000 pg/mg. Intra- and inter-assay imprecision studies at three different concentrations (20, 40, 200 pg/mg) were all within 7.8% and all intra- and inter-assay bias studies at these levels were within 115.1% of target concentration. Ethyl glucuronide levels in fingernail (mean = 29.1 ± 55.6 pg/mg) were higher than ethyl glucuronide levels in hair (mean = 9.48 ± 22.3 pg/mg) and a correlation of the matched pairs was observed (r = 0.552, P < 0.01, n = 529). Evaluating each gender separately revealed that the correlation of male fingernail to male hair was large and significant (r = 0.782, P < 0.01, n = 195) while female hair to female fingernail was small yet significant (r = 0.249, P < 0.01, n = 334). The study results demonstrated that fingernail may be a suitable alternative to hair for ethyl glucuronide detection and may be the preferred sample type due to the lack of a gender bias. PMID:27134762

  3. Positive EtG findings in hair as a result of a cosmetic treatment.

    PubMed

    Sporkert, Frank; Kharbouche, Hicham; Augsburger, Marc P; Klemm, Clementine; Baumgartner, Markus R

    2012-05-10

    In a case of a driving ability assessment, hair analysis for ethyl glucuronide (EtG) was requested by the authorities. The person concerned denied alcohol consumption and did not present any clinical sign of alcoholism. However, EtG was found in concentrations of up to 910pg/mg in hair from different sampling dates suggesting an excessive drinking behavior. The person declared to use a hair lotion on a regularly base. To evaluate a possible effect of the hair lotion, prospective blood and urine controls as well as hair sampling of scalp and pubic hair were performed. The traditional clinical biomarkers of ethanol consumption, CDT and GGT, were inconspicuous in three blood samples taken. EtG was not detected in all collected urine samples. The hair lotion was transmitted to our laboratory. The ethanol concentration in this lotion was determined with 35g/L. The EtG immunoassay gave a positive result indicating EtG, which could be confirmed by GC-MS/MS-NCI. In a follow-up experiment the lotion was applied to the hair of a volunteer over a period of six weeks. After this treatment, EtG could be measured in the hair at a concentration of 72pg/mg suggesting chronic and excessive alcohol consumption. Overnight incubation of EtG free hair in the lotion yielded an EtG concentration of 140pg/mg. In the present case, the positive EtG hair findings could be interpreted as the result of an EtG containing hair care product. To our knowledge, the existence of such a product has not yet been reported, and it is exceptionally unusual to find EtG in cosmetics. Therefore, external sources for hair contamination should always be taken into account when unusual cosmetic treatment is mentioned. In those cases, it is recommended to analyze the hair product for a possible contamination with EtG. The analysis of body hair can help to reveal problems occurring from cosmetic treatment of head hair. As a consequence, the assessment of drinking behavior should be based on more than one

  4. Inhibition of bacterial degradation of EtG by collection as dried urine spots (DUS).

    PubMed

    Redondo, Ana Hernández; Körber, Christiane; König, Stefan; Längin, Andreas; Al-Ahmad, Ali; Weinmann, Wolfgang

    2012-03-01

    Ethyl glucuronide (EtG) and ethyl sulfate (EtS) are direct alcohol consumption markers widely used nowadays for clinical and forensic applications. They are detectable in blood and urine even after consumption of trace amounts of ethanol and for a longer time frame, being detectable even when no more ethanol is present. The instability of EtG against bacterial degradation in contaminated urine samples and/or the possible postcollection synthesis of this metabolite in samples containing, e.g., Escherichia coli and ethanol, may cause false identification of alcohol uptake. Therefore, it is of paramount importance to constrict these error sources by inhibition of any bacterial growth causing hydrolization or synthesis of EtG. This study evaluates a new method of collecting urine samples on filter paper, dried urine spots (DUS), for simultaneous detection of EtG, EtS and creatinine, having the great advantage of inhibiting bacterial activity. In addition, a method validation for the determination of EtG and EtS in DUS was performed according to the FDA guidelines. Sterile-filtered urine was spiked with EtG and EtS, inoculated with E. coli and incubated. Liquid and dried urine samples were collected after various time intervals up to 96 h. Liquid samples were frozen immediately after collection, whereas aliquots for DUS were pipetted onto filter paper, allowed to dry and stored at RT until analysis 1 week after. The specimens were analyzed by LC-ESI-MS/MS. As expected, degradation of EtG, but not of EtS, was observed in contaminated liquid urine samples. However, the specimens collected on filter paper and stored at RT showed no degradation during storage. Therefore, collecting urine samples on filter paper for EtG and EtS analysis turns out to be a reliable method to avoid bacterial degradation of EtG and EtS, and consequently, stabilization of these ethanol metabolites is achieved. In addition, simultaneous measurement of creatinine content as an indicator of urine

  5. A novel, simultaneous extraction of FAEE and EtG from meconium and analysis by LC-MS/MS.

    PubMed

    Vaiano, Fabio; Favretto, Donata; Palumbo, Diego; Cooper, Gail; Mactier, Helen; Busardò, Francesco P; Mari, Francesco; Bertol, Elisabetta

    2016-04-01

    Fatty acid ethyl esters (FAEEs) and ethyl-glucuronide (EtG) in meconium have been widely studied as biomarkers of maternal alcohol consumption during pregnancy. Many analytical approaches have been proposed for their analysis, mostly consisting of separated extraction procedures requiring the use of two meconium aliquots. This study aimed to validate a new analytical procedure for the simultaneous extraction of FAEEs and EtG from a meconium aliquot through a single solid-phase extraction (SPE) applied to 242 anonymized samples of meconium. Targeted FAEEs were: ethyl-myristate (Myr), ethyl-palmitate (Pal), ethyl-oleate (Ole) and ethyl-stearate (Ste). Two hundred milligrams of meconium was sonicated with acetonitrile, and a single SPE performed by means of aminopropyl columns. FAEEs were eluted with hexane, followed by EtG elution with water. Both the mixtures were dried, recovered, and analyzed by liquid chromatography-tandem mass spectrometry using C8 (FAEEs) and C18 (EtG) columns. Transitions were: m/z 257 → 57,88, Myr; m/z 262 → 57,88, Myr-d5; m/z 285 → 57, 72, Pal; m/z 290 → 57,258, Pal-d5; m/z 311 → 72,114, Ole; m/z 316 → 72,265, Ole-d5; m/z 257 → 57,72 Ste; m/z 318 → 57,286, Ste-d5; m/z 221 → 75,85, EtG; m/z 226 → 75,85, EtG-d5. Lower limit of quantification range was 10-15 ng/g for FAEEs and 10 ng/g for EtG. Linearity was evaluated for different concentration ranges; the mean coefficients of determination (R (2)) were above 0.9961. Precision and accuracy for FAEEs and EtG were consistently ≤20 % and ±20 %, respectively. Ion suppression was observed for all the analytes. Matrix effect did not significantly affect the analyses. Recovery efficiency was 93 % for EtG and 75-85 % for FAEEs.

  6. Using Ethyl Glucuronide in Urine to Detect Light and Heavy Drinking in Alcohol Dependent Outpatients

    PubMed Central

    McDonell, Michael G.; Skalisky, Jordan; Leickly, Emily; McPherson, Sterling; Battalio, Samuel; Nepom, Jenny R.; Srebnik, Debra; Roll, John; Ries, Richard K.

    2015-01-01

    Aims This study investigated which ethyl glucuronide immunoassay (EtG-I) cutoff best detects heavy versus light drinking over five days in alcohol dependent outpatients. Methods A total of 121 adults with alcohol use disorders and co-occurring psychiatric disorders taking part in an alcohol treatment study. Participants provided self-reported drinking data and urine samples three time per week for 16-weeks (total samples = 2761). Agreement between low (100 ng/mL, 200 ng/mL), and moderate (500 ng/mL) EtG-I cutoffs and light (women ≤3 standard drinks, men ≤ 4 standard drinks) and heavy drinking (women >3, men >4 standard drinks) were calculated over one to five days. Results The 100 ng/mL cutoff detected >76% of light drinking for two days, and 66% at five days. The 100 ng/mL cutoff detected 84% (1 day) to 79% (5 days) of heavy drinking. The 200 ng/mL cutoff detected >55% of light drinking across five days and >66% of heavy drinking across five days. A 500 ng/mL cutoff identified 68% of light drinking and 78% of heavy drinking for one day, with detection of light (2–5 days <58%) and heavy drinking (2–5 days <71%) decreasing thereafter. Relative to 100 ng/mL, the 200 ng/mL and 500 ng/mL cutoffs were less likely to result in false positives. Conclusions An EtG-I cutoff of 100 ng/mL is most likely to detect heavy drinking for up to five days and any drinking during the previous two days. Cutoffs of ≥ 500 ng/mL are likely to only detect heavy drinking during the previous day. PMID:26475403

  7. Involvement of UDP-glucuronosyltransferases UGT1A9 and UGT2B7 in ethanol glucuronidation, and interactions with common drugs of abuse.

    PubMed

    Al Saabi, Alaa; Allorge, Delphine; Sauvage, François-Ludovic; Tournel, Gilles; Gaulier, Jean-Michel; Marquet, Pierre; Picard, Nicolas

    2013-03-01

    Ethyl glucuronide (EtG) determination is increasingly used in clinical and forensic toxicology to document ethanol consumption. The enzymes involved in EtG production, as well as potential interactions with common drugs of abuse, have not been extensively studied. Activities of human liver (HLM), kidney (HKM), and intestinal (HIM) microsomes, as well as of 12 major human recombinant UDP-glucuronosyltransferases (UGTs), toward ethanol (50 and 500 mM) were evaluated in vitro using liquid chromatography-tandem mass spectrometry. Enzyme kinetic parameters were determined for pooled microsomes and recombinant UGTs with significant activity. Individual contributions of UGTs were estimated using the relative activity factor approach, proposed for scaling activities obtained with cDNA-expressed enzymes to HLM. Interaction of morphine, codeine, lorazepam, oxazepam, nicotine, cotinine, cannabinol, and cannabidiol (5, 10, 15 mg/l) with ethanol (1.15, 4.6, 11.5 g/l; i.e., 25, 100, 250 mM) glucuronidation was assessed using pooled HLM. Ethanol glucuronidation intrinsic clearance (Cl(int)) was 4 and 12.7 times higher for HLM than for HKM and HIM, respectively. All recombinant UGTs, except UGT1A1, 1A6, and 1A10, produced EtG in detectable amounts. UGT1A9 and 2B7 were the most active enzymes, each accounting for 17 and 33% of HLM Cl(int), respectively. Only cannabinol and cannabidiol significantly affected ethanol glucuronidation. Cannabinol increased ethanol glucuronidation in a concentration-dependent manner, whereas cannabidiol significantly inhibited EtG formation in a noncompetitive manner (IC(50) = 1.17 mg/l; inhibition constant (K(i)) = 3.1 mg/l). UGT1A9 and 2B7 are the main enzymes involved in ethanol glucuronidation. In addition, our results suggest that cannabinol and cannabidiol could significantly alter ethanol glucuronidation.

  8. Cancer chemotherapy: a SN-38 (7-ethyl-10-hydroxycamptothecin) glucuronide prodrug for treatment by a PMT (Prodrug MonoTherapy) strategy.

    PubMed

    Angenault, Stéphane; Thirot, Sylvie; Schmidt, Frédéric; Monneret, Claude; Pfeiffer, Bruno; Renard, Pierre

    2003-03-10

    A glucuronide-based prodrug of SN-38 (7-ethyl-10-hydroxycamptothecin) has been synthesized for use in a Prodrug MonoTherapy Strategy (PMT). Since this prodrug is significantly less cytotoxic than SN-38 itself and efficiently releases the drug in vitro in the presence of beta-D-glucuronidase, it can be considered as an appropriate candidate for cancer treatment by a PMT strategy.

  9. Interpretation of group-level factors from a large population dataset in the determination of ethyl glucuronide in hair.

    PubMed

    Salomone, Alberto; Pirro, Valentina; Lombardo, Tonia; Di Corcia, Daniele; Pellegrino, Sergio; Vincenti, Marco

    2015-05-01

    Ethyl glucuronide in hair (HEtG) is the most accredited marker to prove chronic alcohol abuse. In this study, we evaluate the comprehensive results of HEtG determination obtained during four years of activity (2009-2013) in our laboratory (Northwestern Italy) - across a large cohort of subjects (over 20 000 subjects mostly undergoing medical examination for driving re-licensing) - to provide a general perspective on HEtG analysis and dependence on group-level factors (e.g. age, gender, site and period of hair collection) that could bias the analytical results. HEtG was measured by liquid chromatography-tandem mass spectrometry. About 12% of the subjects presented HEtG concentrations over 30 pg/mg. Upon non-parametric hypothesis tests, distributions of HEtG in independent populations categorized by age proved statistically different, while no differences were found by considering gender, BMI, and site of sampling (head vs. chest hair). A 'seasonality' factor was evaluated by comparing periods of collection approximately representing the hair growth in winter, spring, summer and autumn, and a seasonal trend was observed showing the highest HEtG levels in winter (16.7%) and minimum levels in summer (8.3%). The experimental HEtG distributions confirm that chest hair sampling can be trusted as an alternative to scalp. Furthermore, among biological and external factors, age and season of sampling may significantly influence the measured HEtG concentration, and this potential source of bias should be taken into account when the results are interpreted.

  10. EtG and EtS in Autopsy Blood Samples With and Without Putrefaction Using UPLC-MS-MS.

    PubMed

    Hegstad, Solfrid; Kristoffersen, Lena; Liane, Veronica H; Spigset, Olav

    2017-03-01

    Analytical challenges related to postmortem specimens are well known. The degree of putrefaction of the corpse will influence the quality of the blood samples, and both the efficiency of sample preparation and the subsequent chromatographic performance can be affected. An ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS-MS) method was developed and validated for the determination of ethyl glucuronide (EtG) and ethyl sulfate (EtS) in postmortem whole blood. Sample preparation prior to UPLC-MS-MS analysis consisted of protein precipitation and filtration through a phospholipid removal plate. Chromatography was achieved using an HSS T3 column and gradient elution with formic acid in water in combination with methanol. The injection volume was 0.5 µL. Negative electrospray ionization was performed in the multiple reaction monitoring mode. Two transitions were monitored for the analytes and one for the internal standards. The between-assay relative standard deviations were in the range of 1.7-7.0% and the limits of quantification were 0.025 and 0.009 mg/L for EtG and EtS, respectively. Recovery was 51-55% and matrix effects ranged from 98% to 106% (corrected with internal standard). Blood samples from nine autopsy cases with various extents of putrefaction were analyzed. The sample preparation efficiently removed the phospholipids from the blood specimens. The samples were clean and the analytical quality of the chromatographic performance was satisfactory for both analytes irrespective of the degree of putrefaction.

  11. Evaluation of KIMS immunoassays on a cobas c 501 analyzer for drugs of abuse and ethyl glucuronide testing in urine for forensic abstinence control.

    PubMed

    Neukamm, Merja A; Bahrami, Arsham; Auwärter, Volker; Mehne, Felix M P; Höss, Eva

    2016-12-26

    For the medico-psychological assessment (MPA) during driving licence re-granting in Germany, abstinence control including urine samples is required. In these programmes, even small amounts of markers for drug or alcohol abuse have to be detected. Thus, the concentrations of the target compounds are very low, and, in consequence, the sensitivity of the applied screening method has to be much higher than for clinical use. Modified drugs of abuse and ethyl glucuronide immunoassays on a Roche cobas c 501 analyzer were evaluated for precision, accuracy, onboard calibration stability, cross reactivity, sensitivity, and specificity using authentic urine samples. Precision (intra-day and inter-day relative standard deviation (RSD) and accuracy (bias) at three concentrations were 12% or lower for all parameters. The calibrations remained stable (deviations <25%) for at least 28 days for all assays except amphetamines (21 days). Satisfactory cross reactivity was determined for the relevant analytes and also for several new psychoactive substances (NPS). The sensitivity was 100% for all parameters except methadone metabolite EDDP (92%) and fully met the sensitivity criteria for MPA urine testing. The presented kinetic interaction of microparticles in a solution (KIMS) immunoassays on a cobas c 501 thus provide a new method to reliably detect drug or alcohol consumption in abstinence control programmes requiring high sensitivity. Copyright © 2016 John Wiley & Sons, Ltd.

  12. Gyrokinetic simulations of ETG Turbulence*

    NASA Astrophysics Data System (ADS)

    Nevins, William

    2005-10-01

    Recent gyrokinetic simulations of electron temperature gradient (ETG) turbulence [1,2] produced different results despite similar plasma parameters. Ref.[1] differs from Ref.[2] in that [1] eliminates magnetically trapped particles ( r/R=0 ), while [2] retains magnetically trapped particles ( r/R 0.18 ). Differences between [1] and [2] have been attributed to insufficient phase-space resolution and novel physics associated with toroidicity and/or global simulations[2]. We have reproduced the results reported in [2] using a flux-tube, particle-in-cell (PIC) code, PG3EQ[3], thereby eliminating global effects as the cause of the discrepancy. We observe late-time decay of ETG turbulence and the steady-state heat transport in agreement with [2], and show this results from discrete particle noise. Discrete particle noise is a numerical artifact, so both the PG3EQ simulations reported here and those reported in Ref.[2] have little to say about steady-state ETG turbulence and the associated anomalous electron heat transport. Our attempts to benchmark PIC and continuum[4] codes at the plasma parameters used in Ref.[2] produced very large, intermittent transport. We will present an alternate benchmark point for ETG turbulence, where several codes reproduce the same transport levels. Parameter scans about this new benchmark point will be used to investigate the parameter dependence of ETG transport and to elucidate saturation mechanisms proposed in Refs.[1,2] and elsewhere[5-7].*In collaboration with A. Dimits (LLNL), J. Candy, C. Estrada-Mila (GA), W. Dorland (U of MD), F. Jenko, T. Dannert (Max-Planck Institut), and G. Hammett (PPPL). Work at LLNL performed for US DOE under Contract W7405-ENG-48.[1] F. Jenko and W. Dorland, PRL 89, 225001 (2002).[2] Z. Lin et al, 2004 Sherwood Mtg.; 2004 TTF Mtg.; Fusion Energy 2004 (IAEA, Vienna, 2005); Bull. Am. Phys. Soc. (November, 2004); 2005 TTF Mtg.; 2005 Sherwood Mtg.; Z. Lin, et al, Phys. Plasmas 12, 056125 (2005). [3] A.M. Dimits

  13. Iso-glucuronides.

    PubMed

    Dickinson, Ronald G

    2011-03-01

    This review on isomers or acyl glucuronides (iso-glucuronides) updates earlier reviews, and attempts to place in context the advances that have been made, especially over the last 15 years. The essential chemistry behind the intramolecular acyl migration and anomerization reactions of acyl glucuronides has been appreciated for 30 years. The great advances in the past 15 years have been in understanding the dynamics and kinetics of these processes in vitro, using highly sophisticated modern technology, e.g. LC-NMR, LC-MS/MS. In this way, earlier assumptions on kinetics and identification of migration isomers and anomers have come under intense review and update. Extensive structure-activity relationships, involving electronic and steric characteristics of an acyl glucuronide and its possible 7 isomers (excluding transient open-chain species) have been delineated. The covalent modification of endogenous proteins and other macromolecules has been further explored, though direct linkage between such modification and toxic sequelae remains elusive. An alternative view of acyl glucuronides and iso-glucuronides as just xenobiotics has perhaps added the dimension that acyl glucuronidation (and attendant formation of iso-glucuronides) does not necessarily mean that glucuronidation of the aglycone has ended metabolic sequences in vivo.

  14. Flavonoid glucuronides from Helicteres isora.

    PubMed

    Kamiya, K; Saiki, Y; Hama, T; Fujimoto, Y; Endang, H; Umar, M; Satake, T

    2001-05-01

    Five flavonoid glucuronides were obtained from the fruit of Helicteres isora, three of which were previously unknown compounds: isoscutellarein 4'-methyl ether 8-O-beta-D-glucuronide 6"-n-butyl ester. isoscutellarein 4'-methyl ether 8-O-beta-D-glucuronide 2", 4"-disulfate and isoscutellarein 8-O-beta-D-glucuronide 2",4"-disulfate. The structures were determined on the basis of spectroscopy and hydrolysis experiments.

  15. Dimensionless parameters, scaling laws, and the implications for ETG

    SciTech Connect

    Castle, G.G.

    1995-04-20

    ETG will be useful in resolving several physical issues relevant to Spherical Tokamak Reactor concepts. First, it will provide a test of whether transport is Bohm or gyro-Bohm in nature. The second point is that ETG will operate in a completely different range of {rho}* space from other high performance machines, opening up a previously inaccessible region of parameter space. ETG is also a (very) high-{beta} machine. It would be the only device that would have all of its parameters except {rho}* similar to those of a Spherical tokamak Reactor. If it turns out that the transport scales definitively as either Bohm or gyro-Bohm, then extrapolation to reactor conditions with significantly lower values of {rho}* would become more credible. It is also shown that in general one cannot obtain a power law relation in the dimensionless variables for the confinement tim from a power law fit to the engineering variables. It is shown, however, that if T{sub i}/T{sub e} and n{sub i}/n{sub e} are constant or if a modified definition of certain dimensionless variables is adopted, then such a power law conversion is possible.

  16. Identification of glucuronide metabolites of d- and 1-methadone in bile from the isolated perfused rat liver.

    PubMed

    Lynn, R K; Leger, R M; Gerber, N

    1976-09-01

    Three hydroxylated metabolites of methadone, 2-ethyl-3-(dihydroxyphenyl)-5-methyl-3-phenylpyrroline (di-HO-EMDP), 2-ethyl-3-(hydroxy-methoxy-phenyl)-5-methyl-3-phenylpyrroline (HO-MeO-EMDP) and 2-ethyl-3-(hydroxy-phenyl)-5-methyl-3-phenylpyrroline (HO-EMDP) were excreted as O-glucuronide conjugates in bile from the isolated perfused rat liver following the addition of either d- or 1-methadone to the perfusate. These glucuronide metabolites were identified as intact molecules by combined gas chromatographic-mass spectrometric (GC-MS) analysis of their permethylated derivatives.

  17. Isolation and identification of androstanediol glucuronide from human plasma.

    PubMed

    Rao, P N; Burdett, J E; Moore, P H; Horton, R

    1987-11-01

    [3H]Dihydrotestosterone (50 microCi) was infused into normal men and women for 8 h. It was previously shown that this was sufficient time for this material to reach a steady state. Venous plasma was obtained at 6 and 8 h, pooled, and the unconjugated steroids removed by ether extraction. The remaining plasma was adjusted to pH 4.9 and the steroid conjugate was extracted first with ethyl acetate and then with an ether-ethanol mixture. The extracts were combined and taken to dryness. Steroid sulfates were solvolyzed using dioxane, and the mixture partitioned between ether and 1% NaOH. The aqueous phase was acidified and added to an XAD-2 column, washed with water, and the glucuronide fraction eluted with methanol. The solvent was concentrated and the methanol extract was passed through a C18 Sep-Pak, filtered through an Acrodisc CR and then subjected to gradient high performance liquid chromatography [HPLC] (Nova-Pak C18, KH2PO4, pH 3, and methanol). The fractions containing steroid glucuronides were collected and esterified with diazomethane and then acetylated with acetic anhydride in pyridine. The glucuronide triacetyl methyl ester (GAME) derivatives were then run in a second HPLC system (3 Lichrosorb 5 mu columns, 4 mm x 25 cm) using a gradient of ethanol-heptane and heptane. We clearly established that this system separates 3 alpha-diol GAME conjugated at the 17 and 3 positions (44 vs 50 min) with authentic samples previously synthesized in our laboratory. We concluded that the pooled plasma contained only the 17-GAME conjugate. No significant activity of the 3-glucuronide was detected. The natural compound in circulation, therefore, is 5 alpha-androstane-3 alpha, 17 beta-diol 17-glucuronide.

  18. The UV window on counter rotating ETGs: insight from SPH simulations with chemo-photometric implementation

    NASA Astrophysics Data System (ADS)

    Bettoni, D.; Mazzei, P.; Rampazzo, R.; Marino, A.; Galletta, G.; Buson, L. M.

    2014-11-01

    The Galaxy Evolution Explorer ( GALEX) detected ultraviolet emission in about 50 % of multi-spin early-type galaxies (ETGs), suggesting the occurrence of a recent rejuvenation episode connected to the formation of these kinematical features. With the aim at investigating the complex evolutionary scenario leading to the formation of counter rotating ETGs (CR-ETGs) we use our Smooth Particle Hydrodynamic (SPH) code with chemo-photometric implementation. We discuss here the UV evolutionary path of two CR-ETGs, NGC 3593 and NGC 5173, concurrently best fitting their global observed properties, i.e., morphology, dynamics, as well as their total B-band absolute magnitude and spectral energy distribution (SED) extended over three orders of magnitude in wavelength. These simulations correspond to our predictions about the target evolution which we follow in the color-magnitude diagram (CMD), near-UV (NUV) versus r-band absolute magnitude, as a powerful diagnostic tool to emphasize rejuvenation episodes.

  19. ETG turbulence simulation of tokamak edge plasmas via 3+1 gyrofluid code

    NASA Astrophysics Data System (ADS)

    Xi, P. W.; Xu, X. Q.; Dimits, A.; Umansky, M.; Joseph, I.; Kim, S. S.

    2012-10-01

    To study ETG driven turbulence at H-mode pedestal, which is important for the magnetic reconnection of ELM dynamics via ETG-MHD interaction, a 3+1 gyrofluid code is developed under BOUT++ framework. Four evolving quantities are density, parallel velocity, parallel pressure and perpendicular pressure for electron and adiabatic ion is used. Gyro-average is done by utilizing Pad'e approximation and parallel Landau closure for Landau damping is implemented by using a newly developed non-Fourier method. By calculating the ETG mode growth rate and real frequency for the ETG cyclone equilibrium, our code is benchmarked with gyrokinetic codes. We also calculated the electron heat transport level at turbulence saturation phase for both cyclone case and H-mode pedestal. Because the pedestal width is typically ten times larger than ETG simulation domain, the three different region of pedestal, i.e. pedestal top, peak gradient region and pedestal bottom, are simulated separately. The dramatic difference on magnetic shear and temperature length scale of these three regions lead to different ETG linear and nonlinear behaviors.

  20. Autism and Phthalate Metabolite Glucuronidation

    ERIC Educational Resources Information Center

    Stein, T. Peter; Schluter, Margaret D.; Steer, Robert A.; Ming, Xue

    2013-01-01

    Exposure to environmental chemicals may precipitate autism spectrum disorders (ASD) in genetically susceptible children. Differences in the efficiency of the glucuronidation process may substantially modulate substrate concentrations and effects. To determine whether the efficiency of this pathway is compromised in children with ASD, we measured…

  1. Glucuronidation of lipophilic substrates: preparation of 3-benzo[a]pyrenyl-beta-D-glucopyranosiduronic acid in multimilligram quantities by microsomal UDP-glucuronyl transferase.

    PubMed

    Johnson, D B; Swanson, M J; Barker, C W; Fanska, C B; Murrill, E E

    1979-01-01

    A convenient method for the enzymic conversion of multimilligram quantities of 3-hydroxybenzo[a]pyrene to 3-benzo[a]pyrenyl-beta-D-glucopyranosiduronic acid in 90% yield is described. Commercially available freeze-dried rabbit liver microsomes were incubated in the presence of UDPGA, 3-hydroxybenzo[a]pyrene, and Triton X-100 detergent (Figure 1). The course of the biosynthetic reaction was followed by fluorimetry. The glucuronide product was extracted from the acidified incubation supernate with ethyl acetate and the acid function of the glucuronide was utilized in an acid-base extraction procedure to purify the glucuronide from biological and unreacted starting material. The glucuronide precipitated from ethyl acetate and was collected by centrifugation. High pressure liquid chromatography and spectroscopic techniques were used to verify the structure and purity of 3-benzo[a]pyrenyl-beta-D-glucopyranosiduronic acid.

  2. Scaling of Electron Thermal Conductivity during the Transition between Slab and Mixed Slab-Toroidal ETG Mode

    NASA Astrophysics Data System (ADS)

    Sokolov, Vladimir; Balbaky, Abed; Sen, Amiya K.

    2015-11-01

    Transition from the slab to the toroidal branch of the electron temperature gradient (ETG) mode has been successfully achieved in a basic experiment in Columbia Linear Machine CLM. We found a modest increase in saturated ETG potential fluctuations (~ 2 ×) and a substantial increase in the power density of individual mode peaks (~ 4 - 5 ×) with increased levels of curvature. We have obtained a set of experimental scalings for electron thermal conductivity χ⊥e as a function of the inverse radius of curvature Rc-1 for different fluctuation levels of the initial slab ETG mode. We found that thermal conductivity for pure slab modes is larger than it is for mixed slab-toroidal ETG modes with the same level of mode fluctuation. This effective reduction in diffusive transport can be partly explained by the flute nature of the toroidal ETG mode. This research was supported by the Department of Electrical Engineering of Columbia University.

  3. A Preliminary Basic Experiment on the Production and Identification of Toroidal ETG Modes

    NASA Astrophysics Data System (ADS)

    Sokolov, Vladimir; Balbaky, Abed; Sen, Amiya K.

    2012-10-01

    Production and identification of the slab branch of the Electron Temperature Gradient (ETG) mode and the measurement of the consequent electron thermal conductivity have been successfully made in a basic experiment in Columbia Linear Machine (CLM) [1,2]. A preliminary experiment on the transition of the slab mode to the toroidal (curvature) branch of ETG mode in CLM is reported. CLM was operated in the mirror configuration with cell length (50cm-100cm) and mirror ratio (1-2.2). The radius of curvature is Rc˜1.3m and the critical ratio for the transition to toroidal mode has been achieved, k| / 2 ɛnk< 0.1 [3]. We first excite the slab ETG mode [1] and gradually increase the magnetic curvature drive by increasing the magnetic mirror ratio, and observe an increase of the ETG mode amplitude up to 2 times and a small change in mode frequency. Alternatively, we can shorten the mirror cell length via moving the mirror coil to increase the bounce average curvature drive and the mode amplitude. [4pt] [1] X.Wei, V.Sokolov, and A.K. Sen, Phys. Plasmas 17, 042108 (2010).[0pt] [2] V.Sokolov, and A.K. Sen, Phys. Rev. Lett. 107, 155001 (2011).[0pt] [3] J.Y.Kim and W.Horton, Phys.Fluids B 3, 1167 (1991).

  4. Different electrostatic potentials define ETGE and DLG motifs as hinge and latch in oxidative stress response.

    PubMed

    Tong, Kit I; Padmanabhan, Balasundaram; Kobayashi, Akira; Shang, Chengwei; Hirotsu, Yosuke; Yokoyama, Shigeyuki; Yamamoto, Masayuki

    2007-11-01

    Nrf2 is the regulator of the oxidative/electrophilic stress response. Its turnover is maintained by Keap1-mediated proteasomal degradation via a two-site substrate recognition mechanism in which two Nrf2-Keap1 binding sites form a hinge and latch. The E3 ligase adaptor Keap1 recognizes Nrf2 through its conserved ETGE and DLG motifs. In this study, we examined how the ETGE and DLG motifs bind to Keap1 in a very similar fashion but with different binding affinities by comparing the crystal complex of a Keap1-DC domain-DLG peptide with that of a Keap1-DC domain-ETGE peptide. We found that these two motifs interact with the same basic surface of either Keap1-DC domain of the Keap1 homodimer. The DLG motif works to correctly position the lysines within the Nrf2 Neh2 domain for efficient ubiquitination. Together with the results from calorimetric and functional studies, we conclude that different electrostatic potentials primarily define the ETGE and DLG motifs as a hinge and latch that senses the oxidative/electrophilic stress.

  5. Lithocholate glucuronide is a cholestatic agent

    SciTech Connect

    Oelberg, D.G.; Chari, M.V.; Little, J.M.; Adcock, E.W.; Lester, R.

    1984-06-01

    Lithocholic acid and its taurine, glycine, and sulfate derivatives are potent cholestatic agents. (3 beta-/sup 3/H)lithocholate 3-O-beta-D-glucuronide was synthesized, and chemical and radiochemical purity were established. The aqueous solubility of lithocholate glucuronide was determined and found to be greater than that of lithocholic acid or several of its derivatives. In the range of concentrations examined, calcium ions precipitated lithocholate glucuronide stoichiometrically. The material was administered to rats prepared with an external biliary fistula. When 17-25 micrograms quantities were administered, 89.1 +/- 4.5% (mean +/- SEM) of the radiolabel was secreted in bile within the first 20 h after administration, the major fraction being secreted in less than 20 min. Four-fifths of the radiolabeled material in bile was the administered unaltered parent compound, while a minor fraction consisted of a more polar derivative(s). We showed that increasing biliary concentrations of more polar derivatives were observed with milligram doses of (3H)lithocholate glucuronide, and with time after the administration of these loading doses. Milligram doses of (3H)lithocholate glucuronide resulted in partial or complete cholestasis. When induced cholestasis was partial, secretion in bile remained the primary excretory route (82.5-105.6% recovery in bile), while, when complete cholestasis was induced, wide tissue distribution of radiolabel was observed. Cholestasis developed rapidly during infusion of (3H)lithocholate glucuronide. Bile flow was diminished within 10-20 min of the start of an infusion of 0.05 mumol, 100 g-1 body weight, minute-1, administered concomitantly with an equimolar infusion of taurocholate. The results establish that lithocholate glucuronide exerts cholestatic effects comparable to those exerted by unconjugated lithocholic acid.

  6. Ethyl ether

    Integrated Risk Information System (IRIS)

    Ethyl ether ; CASRN 60 - 29 - 7 Human health assessment information on a chemical substance is included in the IRIS database only after a comprehensive review of toxicity data , as outlined in the IRIS assessment development process . Sections I ( Health Hazard Assessments for Noncarcinogenic Effect

  7. Ethyl carbamate

    Integrated Risk Information System (IRIS)

    Ethyl carbamate ; CASRN 51 - 79 - 6 Human health assessment information on a chemical substance is included in the IRIS database only after a comprehensive review of toxicity data , as outlined in the IRIS assessment development process . Sections I ( Health Hazard Assessments for Noncarcinogenic Ef

  8. Ethyl acetate

    Integrated Risk Information System (IRIS)

    Ethyl acetate ; CASRN 141 - 78 - 6 Human health assessment information on a chemical substance is included in the IRIS database only after a comprehensive review of toxicity data , as outlined in the IRIS assessment development process . Sections I ( Health Hazard Assessments for Noncarcinogenic Eff

  9. Ethyl chloride

    Integrated Risk Information System (IRIS)

    Ethyl chloride ; CASRN 75 - 00 - 3 Human health assessment information on a chemical substance is included in the IRIS database only after a comprehensive review of toxicity data , as outlined in the IRIS assessment development process . Sections I ( Health Hazard Assessments for Noncarcinogenic Eff

  10. Glucuronides from metabolites to medicines: a survey of the in vivo generation, chemical synthesis and properties of glucuronides.

    PubMed

    Stachulski, Andrew V; Meng, Xiaoli

    2013-06-01

    Covering: 1998 to 2011. Previous review: Nat. Prod. Rep., 1998, 15, 173-186. The fourteen years that have passed since the previous review on this topic have seen a significant increase of interest in many aspects of glucuronide chemistry and biology. Glucuronides are the most important class of phase 2 xenobiotic metabolites and typically act in a detoxifying role. While this is generally true for O-alkyl and O-aryl glucuronides, a number of glucuronides are known to be pharmacologically active per se. Additionally the use of glucuronide prodrugs, notably to ameliorate the cytotoxicity of anticancer agents, has markedly increased. Whereas the previous review covered only the synthesis of O-glucuronides, we now include N-, S- and C-glucuronides also and discuss both synthetic and biological aspects. Synthetic methods for all classes of glucuronides are reviewed and updated, together with advances in the enzymatic synthesis of glucuronides and methods for their detection. Finally we discuss the biological reactivity of glucuronides where known, including the important morphine-6-glucuronide. A lively debate has continued for several years on whether O-acyl glucuronide metabolites of carboxylic acids are toxic, affecting both the safety assessment of well-used drugs and new drug development programmes. We summarise the current understanding, together with other known examples of interaction between glucuronides and macromolecules.

  11. In silico prediction of acyl glucuronide reactivity

    NASA Astrophysics Data System (ADS)

    Potter, Tim; Lewis, Richard; Luker, Tim; Bonnert, Roger; Bernstein, Michael A.; Birkinshaw, Timothy N.; Thom, Stephen; Wenlock, Mark; Paine, Stuart

    2011-11-01

    Drugs and drug candidates containing a carboxylic acid moiety, including many widely used non-steroidal anti-inflammatory drugs (NSAIDs) are often metabolized to form acyl glucuronides (AGs). NSAIDs such as Ibuprofen are amongst the most widely used drugs on the market, whereas similar carboxylic acid drugs such as Suprofen have been withdrawn due to adverse events. Although the link between these AG metabolites and toxicity is not proven, there is circumstantial literature evidence to suggest that more reactive acyl glucuronides may, in some cases, present a greater risk of exhibiting toxic effects. We wished therefore to rank the reactivity of potential new carboxylate-containing drug candidates, and performed kinetic studies on synthetic acyl glucuronides to benchmark our key compounds. Driven by the desire to quickly rank the reactivity of compounds without the need for lengthy synthesis of the acyl glucuronide, a correlation was established between the degradation half-life of the acyl glucuronide and the half life for the hydrolysis of the more readily available methyl ester derivative. This finding enabled a considerable broadening of chemical property space to be investigated. The need for kinetic measurements was subsequently eliminated altogether by correlating the methyl ester hydrolysis half-life with the predicted 13C NMR chemical shift of the carbonyl carbon together with readily available steric descriptors in a PLS model. This completely in silico prediction of acyl glucuronide reactivity is applicable within the earliest stages of drug design with low cost and acceptable accuracy to guide intelligent molecular design. This reactivity data will be useful alongside the more complex additional pharmacokinetic exposure and distribution data that is generated later in the drug discovery process for assessing the overall toxicological risk of acidic drugs.

  12. Determination of propofol glucuronide from hair sample by using mixed mode anion exchange cartridge and liquid chromatography tandem mass spectrometry.

    PubMed

    Kwak, Jae-Hwan; Kim, Hye Kyung; Choe, Sanggil; In, Sangwhan; Pyo, Jae Sung

    2016-03-15

    The main objective of this study was to develop and validate a simpler and less time consuming analytical method for determination of propofol glucuronide from hair sample, by using mixed mode anion exchange cartridge and liquid chromatography-tandem mass spectrometry (LC-MS/MS). The study uses propofol glucuronide, a major metabolite of propofol, as a marker for propofol abuse. The hair sample was digested in sodium hydroxide solution and loaded in mixed-mode anion cartridge for solid phase extraction. Water and ethyl acetate were used as washing solvents to remove interfering substances from the hair sample. Consequently, 2% formic acid in ethyl acetate was employed to elute propofol glucuronide from the sorbent of mixed-mode anion cartridge, and analyzed by LC-MS/MS. The method validation parameters such as selectivity, specificity, LOD, LLOQ, accuracy, precision, recovery, and matrix effect were also tested. The linearity of calibration curves showed good correlation, with correlation coefficient 0.998. The LOD and LLOQ of the propofol glucuronide were 0.2 pg/mg and 0.5 pg/mg, respectively. The intra and inter-day precision and accuracy were acceptable within 15%. The mean values of recovery and matrix effect were in the range of 91.7-98.7% and 87.5-90.3%, respectively, signifying that the sample preparation, washing and extraction procedure were efficient, and there was low significant hair matrix effect for the extraction of propofol glucuronide from hair sample on the mixed mode anion cartridge. To evaluate the suitability of method, the hair of propofol administered rat was successfully analyzed with this method.

  13. AGN feedback and star formation in ETGs: negative and positive feedback

    NASA Astrophysics Data System (ADS)

    Ciotti, Luca; Ostriker, Jeremiah P.; Novak, Greg; Negri, Andrea; Pellegrini, Silvia; Posacki, Silvia

    2015-08-01

    AGN feedback from supermassive black holes at the center of Early Type Galaxies is commonly invoked as the explanation for the quenching of star formation in these systems, that after this phase are considered “red and dead”. The situation is by far more complicated, due to the significant amount of mass injected in the galaxy by the evolving stellar population over cosmological times. In absence of feedback, this mass would lead to unobserved galactic cooling flows, and to central black holes two orders of magnitude more massive than observed. I will present the results of state-of-the-art hydrodynamical simulations with radiative transport and star formation of the “passive” evolution of ETGs, focusing in particular on highly structured spatial and temporal nature of the intermittent AGN feedback, that is not only negative (shutting down the cooling episodes of the ISM), but also positive, inducing star formation in the inner regions of the host galaxy.

  14. Acyl glucuronides: the good, the bad and the ugly.

    PubMed

    Regan, Sophie L; Maggs, James L; Hammond, Thomas G; Lambert, Craig; Williams, Dominic P; Park, B Kevin

    2010-10-01

    Acyl glucuronidation is the major metabolic conjugation reaction of most carboxylic acid drugs in mammals. The physiological consequences of this biotransformation have been investigated incompletely but include effects on drug metabolism, protein binding, distribution and clearance that impact upon pharmacological and toxicological outcomes. In marked contrast, the exceptional but widely disparate chemical reactivity of acyl glucuronides has attracted far greater attention. Specifically, the complex transacylation and glycation reactions with proteins have provoked much inconclusive debate over the safety of drugs metabolised to acyl glucuronides. It has been hypothesised that these covalent modifications could initiate idiosyncratic adverse drug reactions. However, despite a large body of in vitro data on the reactions of acyl glucuronides with protein, evidence for adduct formation from acyl glucuronides in vivo is limited and potentially ambiguous. The causal connection of protein adduction to adverse drug reactions remains uncertain. This review has assessed the intrinsic reactivity, metabolic stability and pharmacokinetic properties of acyl glucuronides in the context of physiological, pharmacological and toxicological perspectives. Although numerous experiments have characterised the reactions of acyl glucuronides with proteins, these might be attenuated substantially in vivo by rapid clearance of the conjugates. Consequently, to delineate a relationship between acyl glucuronide formation and toxicological phenomena, detailed pharmacokinetic analysis of systemic exposure to the acyl glucuronide should be undertaken adjacent to determining protein adduct concentrations in vivo. Further investigation is required to ascertain whether acyl glucuronide clearance is sufficient to prevent covalent modification of endogenous proteins and consequentially a potential immunological response.

  15. A highly toxic morphine-3-glucuronide derivative.

    PubMed

    Salvatella, Mariona; Arsequell, Gemma; Valencia, Gregorio; Rodríguez, Raquel E

    2004-02-23

    By the coupling of octylamine to the uronic acid function of morphine-3-glucuronide (M3G) a new glycoconjugate (morphine-3-octylglucuronamide, M3GOAM) was prepared. When assayed in both rats and mice up to ng/kg (i.p.) doses none of the animals survived. The aliphatic octyl chain may be the lethal factor since a closely related derivative (M3GNH2), was not toxic and showed similar opioid antagonist properties than naloxone.

  16. 21 CFR 584.200 - Ethyl alcohol containing ethyl acetate.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 6 2013-04-01 2013-04-01 false Ethyl alcohol containing ethyl acetate. 584.200... Ethyl alcohol containing ethyl acetate. The feed additive ethyl alcohol containing ethyl acetate meets the requirement of 27 CFR 21.62, being not less than 92.5 percent ethyl alcohol, each 100...

  17. 21 CFR 584.200 - Ethyl alcohol containing ethyl acetate.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 6 2014-04-01 2014-04-01 false Ethyl alcohol containing ethyl acetate. 584.200... Ethyl alcohol containing ethyl acetate. The feed additive ethyl alcohol containing ethyl acetate meets the requirement of 27 CFR 21.62, being not less than 92.5 percent ethyl alcohol, each 100...

  18. 21 CFR 584.200 - Ethyl alcohol containing ethyl acetate.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 6 2012-04-01 2012-04-01 false Ethyl alcohol containing ethyl acetate. 584.200... Ethyl alcohol containing ethyl acetate. The feed additive ethyl alcohol containing ethyl acetate meets the requirement of 27 CFR 21.62, being not less than 92.5 percent ethyl alcohol, each 100...

  19. 21 CFR 584.200 - Ethyl alcohol containing ethyl acetate.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 6 2011-04-01 2011-04-01 false Ethyl alcohol containing ethyl acetate. 584.200... Ethyl alcohol containing ethyl acetate. The feed additive ethyl alcohol containing ethyl acetate meets the requirement of 27 CFR 21.62, being not less than 92.5 percent ethyl alcohol, each 100...

  20. 21 CFR 584.200 - Ethyl alcohol containing ethyl acetate.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Ethyl alcohol containing ethyl acetate. 584.200... Ethyl alcohol containing ethyl acetate. The feed additive ethyl alcohol containing ethyl acetate meets the requirement of 27 CFR 21.62, being not less than 92.5 percent ethyl alcohol, each 100...

  1. IFU Spectroscopy of 10 ETG nuclei: Properties of the circumnuclear gas emission

    NASA Astrophysics Data System (ADS)

    Ricci, T. V.

    2015-09-01

    LINERs are galactic nuclei containing spectra with prominence of low-ionization lines. Several ionization sources are able to produce a LINER-like spectra (e.g. shocks, low-luminosity AGNs, pAGB stars). In this work, we will present an analysis performed on a sample of 10 massive ETGs with distances up to 30 Mpc. This sample was observed with the GMOS-IFU installed on the Gemini-South Telescope. The data cubes of the galaxies have a FOV of 3.5 - 5 arcsec2 and a spatial resolution of about 0.6 -- 1.0 arcsec. After subtracting the stellar component of the galaxies by means of spectral synthesis, we studied the emission lines all over the FOV of the data cubes. In galaxies where an AGN was clearly detected, we found ionized gaseous discs and also a low-velocity extended emission perpendicular to the gas discs. We concluded that only ionizing photons emerging from the AGNs are not enough to photoionize the gaseous discs of the galaxies. On the other hand, it seems to be responsible for the photoionization of the low-velocity extended gas perpendicular to the discs. We propose a scheme where some collimating agent, somehow aligned to the gaseous discs, may be present in some LINER-like AGNs in the local universe.

  2. Intestinal first-pass glucuronidation activities of selected dihydroxyflavones.

    PubMed

    Wong, Yin Cheong; Zhang, Li; Lin, Ge; Zuo, Zhong

    2009-01-21

    Flavonoids have low bioavailabilities due to extensive intestinal first-pass metabolisms, especially glucuronidation. The present study aimed to evaluate the intestinal glucuronidation of dihydroxyflavones and provide more information on their structure-activity relationships. Seven dihydroxyflavones, namely 3,7-, 5,7-, 6,7-, 7,8-, 2',7-, 3',7-, and 4',7-dihydroxyflavone and a monohydroxyflavone, 7-hydroxyflavone, were investigated by incubating each hydroxyflavone at various concentrations with either human jejunum microsome or rat intestinal microsome. Two mono-glucuronides were identified for each dihydroxyflavone. For human jejunum microsome, most of the studied dihydroxyflavones demonstrated greater glucuronidation activities than that of 7-hydroxyflavone except for 3,7-dihydroxyflavone and 4',7-dihydroxyflavone. 3',7-dihydroxyflavone had the greatest intrinsic clearance which was at least seven times greater than that of all other dihydroxyflavones. In addition, species difference in glucuronidation activity was observed with human jejunum microsome higher than rat intestinal microsome for all hydroxyflavones except for 3,7-dihydroxyflavone. The results further demonstrated that the hydroxyl group positions do affect the intestinal glucuronidation activity of hydroxyflavones. Increasing the number of hydroxyl groups on A- or B-ring (except for 4'-OH) would enhance the glucuronidation activity of flavones, whereas adding a 3-OH on C-ring might not. Furthermore, existence of hydroxyl group at 3' position may enhance the glucuronidation activity of flavonoids.

  3. Glucuronidation, a new metabolic pathway for pyrrolizidine alkaloids.

    PubMed

    He, Yu-Qi; Yang, Li; Liu, Hui-Xin; Zhang, Jiang-Wei; Liu, Yong; Fong, Alan; Xiong, Ai-Zhen; Lu, Yan-Liu; Yang, Ling; Wang, Chang-Hong; Wang, Zheng-Tao

    2010-03-15

    Pyrrolizidine alkaloids (PAs) possess significant hepatotoxicity to humans and animals after metabolic activation by liver P450 enzymes. Metabolism pathways of PAs have been studied for several decades, including metabolic activation, hydroxylation, N-oxidation, and hydrolysis. However, the glucuronidation of intact PAs has not been investigated, although glucuronidation plays an important role in the elimination and detoxication of xenobiotics. In this study, PAs glucuronidation was investigated, and three important points were found. First, we demonstrated that senecionine (SEN)-a representative hepatotoxic PA-could be conjugated by glucuronic acid via an N-glucuronidation reaction catalyzed by uridine diphosphate glucuronosyl transferase in human liver microsomes. Second, glucuronidation of SEN was catalyzed not only by human but also other animal species and showed significant species differences. Rabbits, cattle, sheep, pigs, and humans showed the significantly higher glucuronidation activity than mice, rats, dogs, and guinea pigs on SEN. Kinetics of SEN glucuronidation in humans, pigs, and rabbits followed the one-site binding model of the Michaelis-Menten equation, while cattle and sheep followed the two-sites binding model of the Michaelis-Menten equation. Third, besides SEN, other hepatotoxic PAs including monocrotaline, adonifoline, and isoline also underwent N-glucuronidation in humans and several animal species such as rabbits, cattle, sheep, and pigs.

  4. A specific immunoassay for the determination of morphine and its glucuronides in human blood.

    PubMed

    Beike, J; Blaschke, G; Mertz, A; Köhler, H; Brinkmann, B

    1998-01-01

    The development of specific antisera for immunochemical determination of morphine, morphine-3-glucuronide and morphine-6-glucuronide is described. Morphine was N-demethylated to normorphine and N-alkylated to give N-aminopropyl-normorphine as hapten for antisera against morphine. As haptens for antisera against morphine-3-glucuronide and morphine-6-glucuronide, N-aminopropyl-nor-morphine was glucuronidated in position 3 or 6 respectively. Each of these three haptens were coupled to BSA employing the glutaraldehyde method to obtain three different immunogens. Immunisation of rabbits with these conjugates gave anti-morphine, anti-morphine-3-glucuronide and anti-morphine-6-glucuronide antisera, which were tested in a competitive, heterogeneous radioimmunoassay. Tracers for this radioimmunoassay procedure were synthesised by substitution of morphine and morphine-6-glucuronide in position 2 with 125I and indirect iodination of the morphine-3-glucuronide hapten according to the method of Bolton and Hunter. The resulting antisera show very specific reactions with morphine, morphine-3-glucuronide and morphine-6-glucuronide. Cross reactivities of each antiserum with structurally related opiates and opioides are very low. The cross reactivities of the anti-morphine antiserum against morphine-3-glucuronide, morphine-6-glucuronide, codeine, codeine-6-glucuronide or dihydrocodeine were less than 0.3%, the anti-morphine-3-glucuronide antiserum against morphine, morphine-6-glucuronide, codeine, codeine-6-glucuronide or dihydrocodeine less than 0.1% and the anti-morphine-6-glucuronide antiserum against morphine, morphine-3-glucuronide, codeine or dihydrocodeine less than 0.1%, against codeine-6-glucuronide less than 2.3%. The determination of morphine, morphine-3-glucuronide and morphine-6-glucuronide in blood samples (limit of detection= 3, 1, 0.5 ng/g) of nine cases of fatal heroin overdose with this radioimmunoassay method and the comparison with a GC/MS method is described.

  5. Non-linear Dynamics in ETG Mode Saturation and Beam-Plasma Instabilities

    NASA Astrophysics Data System (ADS)

    Tokluoglu, Erinc K.

    Non-linear mechanisms arise frequently in plasmas and beam-plasma systems resulting in dynamics not predicted by linear theory. The non-linear mechanisms can influence the time evolution of plasma instabilities and can be used to describe their saturation. Furthermore time and space averaged non-linear fields generated by instabilities can lead to collisionless transport and plasma heating. In the case of beam-plasma systems counter-intuitive beam defocusing and scaling behavior which are interesting areas of study for both Low-Temperature and High Energy Density physics. The non-linear mode interactions in form of phase coupling can describe energy transfer to other modes and can be used to describe the saturation of plasma instabilities. In the first part of this thesis, a theoretical model was formulated to explain the saturation mechanism of Slab Electron Temperature Gradient (ETG) mode observed in the Columbia Linear Machine (CLM), based on experimental time-series data collected through probe diagnostics [1]. ETG modes are considered to be a major player in the unexplained high levels of electron transport observed in tokamak fusion experiments and the saturation mechanism of these modes is still an active area of investigation. The data in the frequency space indicated phase coupling between 3 modes, through a higher order spectral correlation coefficient known as bicoherence. The resulting model is similar to [2], which was a treatment for ITG modes observed in the CLM and correctly predicts the observed saturation level of the ETG turbulence. The scenario is further supported by the fact that the observed mode frequencies are in close alignment with those predicted theoretical dispersion relations. Non-linear effects arise frequently in beam-plasma systems and can be important for both low temperature plasma devices commonly used for material processing as well as High Energy Density applications relevant to inertial fusion. The non-linear time averaged

  6. Glucuronidation in the polar bear (Ursus maritimus).

    PubMed

    Sacco, James C; James, Margaret O

    2004-01-01

    Polar bears bioaccumulate lipophilic pollutants, including polychlorinated biphenyls (PCBs), into their bodies from their exclusive diet of marine organisms. Hydroxylated PCB metabolites (OH-PCBs) have been found in plasma, presumably due to CYP-dependent biotransformation of PCBs in liver. Little is known about the phase 2 metabolism of hydroxylated xenobiotics in polar bears. The objective of this study was to examine UDP-glucuronosyltransferase (UGT) activity with OH-PCBs and a hydroxylated polycyclic aromatic hydrocarbon, 3-hydroxy-benzo(a)pyrene (3-OH-BaP), in polar bear liver. Samples of frozen polar bear liver were used to prepare microsomes. UGT activity with 3-OH-BaP in Brij-treated microsomes, measured by a fluorescence assay, was readily measurable with protein concentrations in assay tubes of up to 10 g/ml, but dropped off very sharply at higher protein concentrations. The apparent Km for 3-OH-BaP was 1.71 +/- 0.04 microM, and Vmax 1.26 +/- 0.16 nmol/min/mg protein (mean +/- SD, n=3). UGT activities with a model tetrachloro-OH-PCB (4'-OH-CB72) and a model hexachloro-OH-PCB (4'-OH-CB159) were assayed with [14-C]-UDPGA and separation of the [14-C]-glucuronide by ion-pair extraction and thin-layer chromatography. [14-C]-glucuronide conjugates were readily formed by polar bear liver microsomes in the absence of added substrate, apparently from contaminants present in liver. This phenomenon was not observed using hepatic microsomes from laboratory-held catfish. Glucuronidation efficiency was much higher with 4'-OH-CB72 (Km 7.3 microM; Vmax 1.55 nmol/min/mg) than 4'-OH-CB159 (Km 16.1 microM; Vmax 0.46 nmol/min/mg). The identities of the aglycones present in polar bear liver are not known, but could include OH-PCBs or hydroxylated metabolites of other persistent organic pollutants. This study demonstrates that UGT with high activity for 3-OH-BaP and other substrates is present in polar bear liver.

  7. Structure- and isoform-specific glucuronidation of six curcumin analogs.

    PubMed

    Lu, Danyi; Liu, Hui; Ye, Wencai; Wang, Ying; Wu, Baojian

    2017-04-01

    1. In the present study, we aimed to characterize the glucuronidation of six curcumin analogs (i.e. RAO-3, RAO-8, RAO-9, RAO-18, RAO-19, and RAO-23) derived from galangal using human liver microsomes (HLM) and twelve expressed UGT enzymes. 2. Formation of glucuronide was confirmed using high-resolution mass spectrometry. Single glucuronide metabolite was generated from each of six curcumin analogs. The fragmentation patterns were analyzed and were found to differ significantly between alcoholic and phenolic glucuronides. 3. All six curcumin analogs except one (RAO-23) underwent significant glucuronidation in HLM and expressed UGT enzymes. In general, the methoxy group (close to the phenolic hydroxyl group) enhanced the glucuronidation liability of the curcumin analogs. 4. UGT1A9 and UGT2B7 were primarily responsible for the glucuronidation of two alcoholic analogs (RAO-3 and RAO-18). By contrast, UGT1A9 and four UGT2Bs (UGT2B4, 2B7, 2B15 and 2B17) played important roles in conjugating three phenolic analogs (RAO-8, RAO-9, and RAO-19). Interestingly, the conjugated double bonds system (in the aliphatic chain) was crucial to the substrate selectivity of gastrointestinal UGTs (i.e. UGT1A7, 1A8 and 1A10). 5. In conclusion, glucuronidation of six curcumin analogs from galangal were structure- and isoform-specific. The knowledge should be useful in identifying a curcumin analog with improved metabolic property.

  8. 3-O-Hydroxytyrosol glucuronide and 4-O-hydroxytyrosol glucuronide reduce endoplasmic reticulum stress in vitro.

    PubMed

    Giordano, Elena; Dangles, Olivier; Rakotomanomana, Njara; Baracchini, Silvia; Visioli, Francesco

    2015-10-01

    Endoplasmic reticulum (ER) stress is important for atherosclerosis development and is mediated by the unfolded protein response (UPR). In this work, we synthesized two among the most physiologically-prominent hydroxytyrosol HT hepatic metabolites, i.e. 3-O-HT glucuronide and 4-O-HT glucuronide and we tested their activities on ER stress (in human hepatocarcinoma HepG2 cells), to gain further insight into the cardiopreventive properties of HT, extra virgin olive oil, and the Mediterranean diet. We report that 3-O-HT glucuronide and 4-O-HT glucuronide inhibit tunicamycin-induced ER stress. As compared with the effects of the parent molecule, 3-O-HT glucuronide and 4-O-HT glucuronide at 10 μM and 25 μM alone induced a milder change in mRNA expression levels of both CCAAT-enhancer-binding protein homologous protein (CHOP) and glucose regulated protein GRP78 immunoglobulin heavy chain binding protein (BiP). In conclusion, we add further evidence to the hypothesis that the HT intake might be atheroprotective and reiterate the usefulness to preferably use high-quality, high-(poly)phenol extra virgin olive oil as a prominent condiment.

  9. Glucuronidation of active tamoxifen metabolites by the human UDP glucuronosyltransferases.

    PubMed

    Sun, Dongxiao; Sharma, Arun K; Dellinger, Ryan W; Blevins-Primeau, Andrea S; Balliet, Renee M; Chen, Gang; Boyiri, Telih; Amin, Shantu; Lazarus, Philip

    2007-11-01

    Tamoxifen (TAM) is an antiestrogen that has been widely used in the treatment and prevention of breast cancer in women. One of the major mechanisms of metabolism and elimination of TAM and its major active metabolites 4-hydroxytamoxifen (4-OH-TAM) and 4-OH-N-desmethyl-TAM (endoxifen; 4-hydroxy-N-desmethyl-tamoxifen) is via glucuronidation. Although limited studies have been performed characterizing the glucuronidation of 4-OH-TAM, no studies have been performed on endoxifen. In the present study, characterization of the glucuronidating activities of human UDP glucuronosyltransferases (UGTs) against isomers of 4-OH-TAM and endoxifen was performed. Using homogenates of individual UGT-overexpressing cell lines, UGTs 2B7 approximately 1A8 > UGT1A10 exhibited the highest overall O-glucuronidating activity against trans-4-OH-TAM as determined by Vmax/K(M), with the hepatic enzyme UGT2B7 exhibiting the highest binding affinity and lowest K(M) (3.7 microM). As determined by Vmax/K(M), UGT1A10 exhibited the highest overall O-glucuronidating activity against cis-4-OH-TAM, 10-fold higher than the next-most active UGTs 1A1 and 2B7, but with UGT1A7 exhibiting the lowest K(M). Although both N- and O-glucuronidation occurred for 4-OH-TAM in human liver microsomes, only O-glucuronidating activity was observed for endoxifen; no endoxifen-N-glucuronidation was observed for any UGT tested. UGTs 1A10 approximately 1A8 > UGT2B7 exhibited the highest overall glucuronidating activities as determined by Vmax/K(M) for trans-endoxifen, with the extrahepatic enzyme UGT1A10 exhibiting the highest binding affinity and lowest K(M) (39.9 microM). Similar to that observed for cis-4-OH-TAM, UGT1A10 also exhibited the highest activity for cis-endoxifen. These data suggest that several UGTs, including UGTs 1A10, 2B7, and 1A8 play an important role in the metabolism of 4-OH-TAM and endoxifen.

  10. Morphine, morphine-6-glucuronide and morphine-3-glucuronide pharmacokinetics in newborn infants receiving diamorphine infusions.

    PubMed

    Barrett, D A; Barker, D P; Rutter, N; Pawula, M; Shaw, P N

    1996-06-01

    1. The pharmacokinetics of morphine, morphine-6-glucuronide (M6G) and morphine-3-glucuronide (M3G) were studied in 19 ventilated newborn infants (24-41 weeks gestation) who were given a loading dose of 50 micrograms kg-1 or 200 micrograms kg-1 of diamorphine followed by an intravenous infusion of 15 micrograms kg-1 h-1 of diamorphine. Plasma concentrations of morphine, M3G and M6G were measured during the accrual to steady-state and at steady state of the diamorphine infusion. 2. Following both the 50 micrograms kg-1 or 200 micrograms kg-1 loading doses the mean steady-state plasma concentration (+/- s.d.) of morphine, M3G and M6G were 86 +/- 52 ng ml-1, 703 +/- 400 ng ml-1 and 48 +/- 28 ng ml-1 respectively and morphine clearance was found to be 4.6 +/- 3.2 ml min-1 kg-1. 3. M3G formation clearance was estimated to be 2.5 +/- 1.8 ml min-1 kg-1, and the formation clearance of M6G was estimated to be 0.46 +/- 0.32 ml min-1 kg-1. 4. M3G metabolite clearance was 0.46 +/- 0.60 ml min-1 kg-1, the elimination half-life was 11.1 +/- 11.3 h and the volume of distribution was 0.55 +/- 1.13 l kg-1. M6G metabolite clearance was 0.71 +/- 0.36 ml min-1 kg-1, the elimination half-life was 18.2 +/- 13.6 h and the volume of distribution was 1.03 +/- 0.88 l kg-1. 5. No significant effect of the loading dose (50 micrograms kg-1 or 200 micrograms kg-1) on the plasma morphine or metabolite concentrations or their derived pharmacokinetic parameters was found. 6. We were unable to identify correlations between gestational age of the infants and any of the determined pharmacokinetic parameters. 7. M3G: morphine and M6G: morphine steady-state plasma concentration ratios were 11.0 +/- 10.8 and 0.8 +/- 0.8, respectively. 8. The metabolism of morphine in neonates, in terms of the respective contributions of each glucuronide pathway, was similar to that in adults.

  11. Microvascular protective activity of flavonoid glucuronides fraction from Tulipa gesneriana.

    PubMed

    Budzianowski, J; Korzeniowska, K; Chmara, E; Mrozikiewicz, A

    1999-03-01

    A mixture of flavonoid glucuronides, consisting of 7-O-glucuronides of kaempferol and quercetin 3-O-rutinosides, 3-O-gentiobiosides and 3-O-glucosides, was isolated from the perianths of Tulipa gesneriana L. var. 'Paradae'. It showed protective activity against the increased (both chloroform and histamine) skin vascular permeability in rabbits. The protective effect, measured as the reduction in leakage of Evans blue, was 59.8% after peritoneal treatment at a dose of 25 mg/kg, while that of troxerutin was 45.5%.

  12. Diethylstilbestrol can effectively accelerate estradiol-17-O-glucuronidation, while potently inhibiting estradiol-3-O-glucuronidation

    SciTech Connect

    Zhu, Liangliang; Xiao, Ling; Xia, Yangliu; Zhou, Kun; Wang, Huili; Huang, Minyi; Ge, Guangbo; Wu, Yan; Wu, Ganlin; Yang, Ling

    2015-03-01

    This in vitro study investigates the effects of diethylstilbestrol (DES), a widely used toxic synthetic estrogen, on estradiol-3- and 17-O- (E2-3/17-O) glucuronidation, via culturing human liver microsomes (HLMs) or recombinant UDP-glucuronosyltransferases (UGTs) with DES and E2. DES can potently inhibit E2-3-O-glucuronidation in HLM, a probe reaction for UGT1A1. Kinetic assays indicate that the inhibition follows a competitive inhibition mechanism, with the Ki value of 2.1 ± 0.3 μM, which is less than the possible in vivo level. In contrast to the inhibition on E2-3-O-glucuronidation, the acceleration is observed on E2-17-O-glucuronidation in HLM, in which cholestatic E2-17-O-glucuronide is generated. In the presence of DES (0–6.25 μM), K{sub m} values for E2-17-O-glucuronidation are located in the range of 7.2–7.4 μM, while V{sub max} values range from 0.38 to 1.54 nmol/min/mg. The mechanism behind the activation in HLM is further demonstrated by the fact that DES can efficiently elevate the activity of UGT1A4 in catalyzing E2-17-O-glucuronidation. The presence of DES (2 μM) can elevate V{sub max} from 0.016 to 0.81 nmol/min/mg, while lifting K{sub m} in a much lesser extent from 4.4 to 11 μM. Activation of E2-17-O-glucuronidation is well described by a two binding site model, with K{sub A}, α, and β values of 0.077 ± 0.18 μM, 3.3 ± 1.1 and 104 ± 56, respectively. However, diverse effects of DES towards E2-3/17-O-glucuronidation are not observed in liver microsomes from several common experimental animals. In summary, this study issues new potential toxic mechanisms for DES: potently inhibiting the activity of UGT1A1 and powerfully accelerating the formation of cholestatic E2-17-O-glucuronide by UGT1A4. - Highlights: • E2-3-O-glucuronidation in HLM is inhibited when co-incubated with DES. • E2-17-O-glucuronidation in HLM is stimulated when co-incubated with DES. • Acceleration of E2-17-O-glucuronidationin in HLM by DES is via activating the

  13. Quantitation of Buprenorphine, Norbuprenorphine, Buprenorphine Glucuronide, Norbuprenorphine Glucuronide, and Naloxone in Urine by LC-MS/MS.

    PubMed

    Marin, Stephanie J; McMillin, Gwendolyn A

    2016-01-01

    Buprenorphine is an opioid drug that has been used to treat opioid dependence on an outpatient basis, and is also prescribed for managing moderate to severe pain. Some formulations of buprenorphine also contain naloxone to discourage misuse. The major metabolite of buprenorphine is norbuprenorphine. Both compounds are pharmacologically active and both are extensively metabolized to their glucuronide conjugates, which are also active metabolites. Direct quantitation of the glucuronide conjugates in conjunction with free buprenorphine, norbuprenorphine, and naloxone in urine can distinguish compliance with prescribed therapy from specimen adulteration intended to mimic compliance with prescribed buprenorphine. This chapter quantitates buprenorphine, norbuprenorphine, their glucuronide conjugates and naloxone directly in urine by liquid chromatography tandem mass spectrometry (LC-MS/MS). Urine is pretreated with formic acid and undergoes solid phase extraction (SPE) prior to analysis by LC-MS/MS.

  14. Fate of glucuronide conjugated estradiol in the environment

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The fate and transport of conjugated reproductive hormones, which are polar compared to parent hormones, are little understood. Laboratory bench-scale soil (Hamar; Sandy, mixed, frigid typic Endoaquolls) sorption studies were conducted using [14C] 17ß-estradiol-3-glucuronide for a range of concentra...

  15. Morphine, morphine-6-glucuronide and morphine-3-glucuronide pharmacokinetics in newborn infants receiving diamorphine infusions

    PubMed Central

    BARRETT, D. A.; BARKER, D. P.; RUTTER, N.; PAWULA, M.; SHAW, P. N.

    1996-01-01

    1The pharmacokinetics of morphine, morphine-6-glucuronide (M6G) and morphine-3-glucuronide (M3G) were studied in 19 ventilated newborn infants(24–41 weeks gestation) who were given a loading dose of 50 μg kg−1 or 200 μg kg−1 of diamorphine followed by an intravenous infusion of 15 μg kg−1 h−1 of diamorphine. Plasma concentrations of morphine, M3G and M6G were measured during the accrual to steady-state and at steady state of the diamorphine infusion. 2Following both the 50 μg kg−1 or 200 μg kg−1 loading doses the mean steady-state plasma concentration (±s.d.) of morphine, M3G and M6G were 86±52 ng ml−1, 703±400 ng ml−1 and 48±28 ng ml−1 respectively and morphine clearance was found to be 4.6±3.2 ml min−1 kg−1. 3M3G formation clearance was estimated to be 2.5±1.8 ml min−1 kg−1, and the formation clearance of M6G was estimated to be 0.46±0.32 ml min−1 kg−1. 4M3G metabolite clearance was 0.46±0.60 ml min−1 kg−1, the elimination half-life was 11.1±11.3 h and the volume of distribution was 0.55±1.13 l kg−1. M6G metabolite clearance was 0.71±0.36 ml min−1 kg−1, the elimination half-life was 18.2±13.6 h and the volume of distribution was 1.03±0.88 l kg−1. 5No significant effect of the loading dose (50 μg kg−1 or 200 μg kg−1) on the plasma morphine or metabolite concentrations or their derived pharmacokinetic parameters was found. 6We were unable to identify correlations between gestational age of the infants and any of the determined pharmacokinetic parameters. 7M3G:morphine and M6G:morphine steady-state plasma concentration ratios were 11.0±10.8 and 0.8±0.8, respectively. 8The metabolism of morphine in neonates, in terms of the respective contributions of each glucuronide pathway, was similar to that in adults. PMID:8799518

  16. Interindividual variability in nicotine metabolism: C-oxidation and glucuronidation.

    PubMed

    Nakajima, Miki; Yokoi, Tsuyoshi

    2005-08-01

    Nicotine has roles in the addiction to smoking, replacement therapy for smoking cessation, as a potential medication for several diseases such as Parkinson's disease, Alzheimer's disease, and ulcerative colitis. The absorbed nicotine is rapidly and extensively metabolized and eliminated to urine. A major pathway of nicotine metabolism is C-oxidation to cotinine, which is catalyzed by CYP2A6 in human livers. Cotinine is subsequently metabolized to trans-3'-hydroxycotinine by CYP2A6. Nicotine and cotinine are glucuronidated to N-glucuronides mainly by UGT1A4 and partly by UGT1A9. Trans-3'-hydroxycotinine is glucuronidated to O-glucuronide mainly by UGT2B7 and partly by UGT1A9. Approximately 90% of the total nicotine uptake is eliminated as these metabolites and nicotine itself. The nicotine metabolism is an important determinant of the clearance of nicotine. Recently, advances in the understanding of the interindividual variability in nicotine metabolism have been made. There are substantial data suggesting that the large interindividual differences in cotinine formation are associated with genetic polymorphisms of the CYP2A6 gene. Interethnic differences have also been observed in the cotinine formation and the allele frequencies of the CYP2A6 alleles. Since the genetic polymorphisms of the CYP2A6 gene have a major impact on nicotine clearance, its relationships with smoking behavior or the risk of lung cancer have been suggested. The metabolic pathways of the glucuronidation of nicotine, cotinine, and trans-3'-hydroxycotinine in humans would be one of the causal factors for the interindividual differences in nicotine metabolism. This review mainly summarizes recent results from our studies.

  17. Chlorimuron-ethyl

    Integrated Risk Information System (IRIS)

    Chlorimuron - ethyl ; CASRN 90982 - 32 - 4 Human health assessment information on a chemical substance is included in the IRIS database only after a comprehensive review of toxicity data , as outlined in the IRIS assessment development process . Sections I ( Health Hazard Assessments for Noncarcinog

  18. Methyl ethyl ketone (MEK)

    Integrated Risk Information System (IRIS)

    EPA 635 / R - 03 / 009 www.epa.gov / iris TOXICOLOGICAL REVIEW OF METHYL ETHYL KETONE ( CAS No . 78 - 93 - 3 ) In Support of Summary Information on the Integrated Risk Information System ( IRIS ) September 2003 U.S . Environmental Protection Agency Washington , DC DISCLAIMER This document has been r

  19. Ethyl alcohol production

    SciTech Connect

    Hofman, V.; Hauck, D.

    1980-11-01

    Recent price increases and temporary shortages of petroleum products have caused farmers to search for alternate sources of fuel. The production of ethyl alcohol from grain is described and the processes involved include saccharification, fermentation and distillation. The resulting stillage has potential as a livestock feed.

  20. Analysis of glucuronide and sulfate steroids in urine by ultra-high-performance supercritical-fluid chromatography hyphenated tandem mass spectrometry.

    PubMed

    Doué, Mickael; Dervilly-Pinel, Gaud; Pouponneau, Karinne; Monteau, Fabrice; Le Bizec, Bruno

    2015-06-01

    Profiling conjugated urinary steroids to detect anabolic-steroid misuse is recognized as an efficient analytical strategy in both chemical-food-safety and anti-doping fields. The relevance and robustness of such profiling rely on the analysis of glucuronide and sulfate steroids, which is expected to have properties including accuracy, specificity, sensitivity, and, if possible, rapidity. In this context, the ability of ultra-high-performance supercritical-fluid chromatography (UHPSFC) hyphenated tandem mass spectrometry (MS-MS) to provide reliable and accurate phase II analysis of steroids was assessed. Four stationary phases with sub-2 μm particles (BEH, BEH 2-ethyl-pyridine, HSS C18 SB, and CSH fluorophenyl) were screened for their capacity to separate several conjugated steroid isomers. Analytical conditions including stationary phase, modifier composition and percentage, back pressure, column temperature, and composition and flow rate of make-up solvent were investigated to improve the separation and/or the sensitivity. Thus, an analytical procedure enabling the analysis of eight glucuronide and 12 sulfate steroids by two different methods in 12 and 15 min, respectively, was optimized. The two procedures were evaluated, and UHPSFC-MS-MS analysis revealed its ability to provide sensitive (limits of quantification: 0.1 ng mL(-1) and 0.5 ng mL(-1) for sulfate and glucuronide steroids, respectively) and reliable quantitative performance (R(2) > 0.995, RSD < 20%, and bias < 30%) through the use of suitable labeled internal standards. Comparison with UHPLC-MS-MS was performed, and UHPSFC-MS-MS obtained better performance in terms of sensitivity. Finally, as a proof of concept, this so-called green technology was used in a chemical-food-safety context to profile steroid conjugates in urine samples from bovines treated with estradiol. Graphical Abstract Glucuronide and sulfate steroids analysis in urine by ultra-high performance supercritical fluid

  1. In Vitro Stability of Free and Glucuronidated Cannabinoids in Urine Following Controlled Smoked Cannabis

    PubMed Central

    Desrosiers, Nathalie A.; Lee, Dayong; Scheidweiler, Karl B.; Concheiro-Guisan, Marta; Gorelick, David A.; Huestis, Marilyn A.

    2014-01-01

    Analyte stability is an important factor in urine test interpretation, yet cannabinoid stability data are limited. A comprehensive study of Δ9-tetrahydrocannabinol (THC), 11-hydroxy-THC (11-OH-THC), 11-nor-9-carboxy-THC (THCCOOH), cannabidiol, cannabinol, THC-glucuronide, and THCCOOH-glucuronide stabilities in authentic urine was completed. Urine samples after ad libitum cannabis smoking were pooled to prepare low and high pools for each study participant; baseline concentrations were measured within 24h at room temperature (RT), 4°C and −20°C. Stability at RT, 4°C and −20°C was evaluated by Friedman tests for up to 1 year. THCCOOH, THC-glucuronide, and THCCOOH-glucuronide were quantified in baseline pools. RT THCCOOH baseline concentrations were significantly higher than −20°C, but not 4°C baseline concentrations. After 1 week at RT, THCCOOH increased, THCCOOH-glucuronide decreased, but THC-glucuronide was unchanged. In RT low pool, total THCCOOH (THCCOOH+THCCOOH-glucuronide) was significantly lower after 1 week. At 4°C, THCCOOH was stable 2 weeks, THCCOOH-glucuronide 1 month and THC-glucuronide for at least 6 months. THCCOOH was stable frozen for 1 year, but 6 months high pool results were significantly higher than baseline; THC-glucuronide and THCCOOH-glucuronide were stable for 6 months. Total THCCOOH was stable 6 months at 4°C, and frozen 6 months (low) and 1 year (high). THC, cannabidiol and cannabinol were never detected in urine; although not detected initially, 11-OH-THC was detected in 2 low and 3 high pools after one week at RT. Substantial THCCOOH-glucuronide deconjugation was observed at RT and 4°C. Analysis should be conducted within 3 months if non-hydrolyzed THCCOOH or THCCOOH-glucuronide quantification is required. PMID:24292435

  2. The effect of morbid obesity on morphine glucuronidation.

    PubMed

    Lloret-Linares, Celia; Luo, Huilong; Rouquette, Alexandra; Labat, Laurence; Poitou, Christine; Tordjman, Joan; Bouillot, Jean-Luc; Mouly, Stéphane; Scherrmann, Jean-Michel; Bergmann, Jean-François; Declèves, Xavier

    2017-04-01

    The purpose of the present work was to study the change in morphine metabolic ratio in obese subjects before and after Roux-en-Y Gastric Bypass (RYGB) and to identify clinical and/or biological factors associated with this change. The pharmacokinetics (PK) of oral morphine (30mg), morphine-3-glucuronide (M3G) and morphine-6-glucuronide (M6G) was performed in patients before (n=25; mean BMI=43.2 (35.4-61.9)kg/m(2)), 7-15days (n=16) and 6 months after RYGB (n=19; mean BMI=32.3 (25.4-46.0)kg/m(2)). Morphine Cmax and AUC0-inf were significantly increased and morphine Tmax significantly shortened at 6 months after RYGB compared with preoperative data, indicating an important increase in the rate and extent of morphine absorption. The morphine metabolic ratio 0-inf M3G+M6G/Morphine, decreased significantly from the preoperative to 6 months postoperative period with an average of -26% (range -74%; +21%; p=0.004), but not in the immediate post-operative period. The change in morphine metabolic ratio was associated with a change in BMI, fat mass in kg, and triglyceride levels (rho=0.5, p≤0.04). The degree of change in several markers of low-grade inflammation, or the level of liver steatosis and fibrosis before surgery, was not associated with the change in morphine metabolic ratios. Our findings indicate that RYGB-induced weight loss significantly decreases morphine metabolic ratio, arguing for an effect of morbid obesity on glucuronidation. With glucuronide exposure at 6 months similar to preoperative values, a higher morphine AUC0-inf should encourage reducing morphine dosage in patients undergoing RYGB and chronically receiving immediate-release oral morphine.

  3. Ethyl N-phenyloxamate.

    PubMed

    García-Báez E, Efrén V; Gómez-Castro, Carlos Z; Höpfl, Herbert; Martínez-Martínez, Francisco J; Padilla-Martínez, Itzia I

    2003-10-01

    The oxamate group in the title compound, C(10)H(11)NO(3), is almost coplanar with the phenyl ring because of intramolecular hydrogen-bonding interactions, and the structure can be described as an anilide single bonded to an ethyl carboxylate group. The supramolecular structure is achieved through intermolecular hard N-H...O and soft C-H...X (X = O and phenyl) hydrogen-bonding interactions.

  4. Enzyme-assisted synthesis and structural characterization of the 3-, 8-, and 15-glucuronides of deoxynivalenol.

    PubMed

    Uhlig, Silvio; Ivanova, Lada; Fæste, Christiane Kruse

    2013-02-27

    4-Deoxynivalenol is one of the most prevalent mycotoxins in grain-based food and feed products worldwide. Conjugation of deoxynivalenol to glucuronic acid and elimination via the urine appears to be the major metabolism pathway, although with differing efficiency in different species. In order to make pure deoxynivalenol glucuronides for analytical methodologies available we intended to enzymatically synthesize glucuronides of deoxynivalenol using rat and human liver microsomes supplemented with uridine 5'-diphosphoglucuronic acid and alamethicin as detergent. Three glucuronides were isolated and purified using solid-phase extraction of microsomal incubations and subsequent semipreparative hydrophilic interaction chromatography. NMR spectra were obtained for all three compounds from solutions in methanol, showing that deoxynivalenol 3-O-β-D-glucuronide and deoxynivalenol 15-O-β-D-glucuronide were the major products from incubations of deoxynivalenol with rat and human liver microsomes, respectively. The NMR spectra of a third glucuronide showed replacement of the C-8 carbonyl by a ketal carbon. This glucuronide was finally identified as deoxynivalenol 8-O-β-D-glucuronide. The present study provides unequivocal structural evidence for three glucuronides of deoxynivalenol formed by liver enzymes.

  5. Regioselective Glucuronidation of Diosmetin and Chrysoeriol by the Interplay of Glucuronidation and Transport in UGT1A9-Overexpressing HeLa Cells

    PubMed Central

    Zhao, Min; Chen, Qingwei; Wang, Liping; Jiang, Huangyu; Luo, Feifei; Zhu, Lijun; Lu, Linlin; Wang, Xinchun; Liu, Zhongqiu

    2016-01-01

    This study aimed to determine the reaction kinetics of the regioselective glucuronidation of diosmetin and chrysoeriol, two important methylated metabolites of luteolin, by human liver microsomes (HLMs) and uridine-5′-diphosphate glucuronosyltransferase (UGTs) enzymes. This study also investigated the effects of breast cancer resistance protein (BCRP) on the efflux of diosmetin and chrysoeriol glucuronides in HeLa cells overexpressing UGT1A9 (HeLa—UGT1A9). After incubation with HLMs in the presence of UDP-glucuronic acid, diosmetin and chrysoeriol gained two glucuronides each, and the OH—in each B ring of diosmetin and chrysoeriol was the preferable site for glucuronidation. Screening assays with 12 human expressed UGT enzymes and chemical-inhibition assays demonstrated that glucuronide formation was almost exclusively catalyzed by UGT1A1, UGT1A6, and UGT1A9. Importantly, in HeLa—UGT1A9, Ko143 significantly inhibited the efflux of diosmetin and chrysoeriol glucuronides and increased their intracellular levels in a dose-dependent manner. This observation suggested that BCRP-mediated excretion was the predominant pathway for diosmetin and chrysoeriol disposition. In conclusion, UGT1A1, UGT1A6, and UGT1A9 were the chief contributors to the regioselective glucuronidation of diosmetin and chrysoeriol in the liver. Moreover, cellular glucuronidation was significantly altered by inhibiting BCRP, revealing a notable interplay between glucuronidation and efflux transport. Diosmetin and chrysoeriol possibly have different effects on anti-cancer due to the difference of UGT isoforms in different cancer cells. PMID:27832172

  6. Separation of a BMS drug candidate and acyl glucuronide from seven glucuronide positional isomers in rat plasma via high-performance liquid chromatography with tandem mass spectrometric detection.

    PubMed

    Xue, Y-J; Simmons, Neal J; Liu, Jane; Unger, Steve E; Anderson, Danielle F; Jenkins, Rand G

    2006-01-01

    A high-performance liquid chromatography/tandem mass spectrometry (LC/MS/MS) method has been developed and validated for the determination of a BMS drug candidate and its acyl glucuronide (1-O-beta glucuronide) in rat plasma. A 50-microL aliquot of each plasma sample was fortified with acetonitrile containing the internal standard to precipitate proteins and extract the analytes of interest. After mixing and centrifugation, the supernatant from each sample was transferred to a 96-well plate and injected into an LC/MS/MS system. Chromatographic separation was achieved isocratically on a Phenomenex Luna C(18), 3 mm x 150 mm, 3 microm column. The mobile phase contained 0.075% formic acid in 70:30 (v/v) acetonitrile/water. Under the optimized chromatographic conditions, the BMS drug candidate and its acyl glucuronide were separated from its seven glucuronide positional isomers within 10 min. Resolution of the parent from all glucuronides and acyl glucuronide from its positional isomers was critical to avoid their interference with quantitation of parent or acyl glucuronide. Detection was by positive ion electrospray MS/MS on a Sciex API 4000. The standard curve, which ranged from 5 to 5000 ng/mL, was fitted to a 1/x(2) weighted quadratic regression model for both the BMS drug candidate and its acyl glucuronide. Whole blood and plasma stability experiments were conducted to establish the sample collection, storage, and processing conditions. The validation results demonstrated that this method was rugged and repeatable. The same methodology has also been used in mouse and human plasma for the determination of the BMS drug candidate and its acyl glucuronide.

  7. Organochlorines inhibit acetaminophen glucuronidation by redirecting UDP-glucuronic acid towards the D-glucuronate pathway

    SciTech Connect

    Chan, Tom S. Wilson, John X.; Selliah, Subajini; Bilodeau, Marc; Zwingmann, Claudia; Poon, Raymond; O'Brien, Peter J.

    2008-11-01

    Industry-derived organochlorines are persistent environmental pollutants that are a continuing health concern. The effects of these compounds on drug metabolism are not well understood. In the current study we present evidence that the inhibition of acetaminophen (APAP) glucuronidation by minute concentrations of organochlorines correlates well with their ability to stimulate the D-glucuronate pathway leading to ascorbate synthesis. A set of 6 arylated organochlorines, including 5 PCB (polychlorinated biphenyl) congeners, were assessed for their effects on APAP glucuronidation in isolated hepatocytes from male Sprague-Dawley rats. The capacity of each organochlorine to inhibit APAP glucuronidation was found to be directly proportional to its capacity to stimulate ascorbate synthesis. PCB153, PCB28 and bis-(4-chlorophenyl sulfone) (BCPS) in increasing order were the most effective organochlorines for inhibiting APAP glucuronidation and stimulating the D-glucuronate pathway. None of the 3 inhibitors of APAP glucuronidation were able to alter the expression of UGT1A6, UGT1A7 and UGT1A8 (the major isoforms responsible for APAP glucuronidation in the rat), however, their efficacy at inhibiting APAP glucuronidation was proportional to their capacity to deplete UDP-glucuronic acid (UDPGA). BCPS-mediated inhibition of APAP glucuronidation in isolated hepatocytes had non-competitive characteristics and was insensitive to the inactivation of cytochrome P450. The effective organochlorines were also able to selectively stimulate the hydrolysis of UDPGA to UDP and glucuronate in isolated microsomes, but could not inhibit APAP glucuronidation in microsomes when UDPGA was in excess. We conclude that organochlorines are able to inhibit APAP glucuronidation in hepatocytes by depleting UDPGA via redirecting UDPGA towards the D-glucuronate pathway. Because the inhibition is non-competitive, low concentrations of these compounds could have long term inhibitory effects on the

  8. Evidence for an UDP-glucuronic acid/phenol glucuronide antiport in rat liver microsomal vesicles.

    PubMed

    Bánhegyi, G; Braun, L; Marcolongo, P; Csala, M; Fulceri, R; Mandl, J; Benedetti, A

    1996-04-01

    The transport of glucuronides synthesized in the luminal compartment of the endoplasmic reticulum by UDP-glucuronosyltransferase isoenzymes was studied in rat liver microsomal vesicles. Microsomal vesicles were loaded with p-nitrophenol glucuronide (5 mM), phenolphthalein glucuronide or UDP-glucuronic acid, by a freeze-thawing method. In was shown that: (i) the loading procedure resulted in millimolar intravesicular concentrations of the different loading compounds; (ii) addition of UDP-glucuronic acid (5 mM) to the vesicles released both intravesicular glucuronides within 1 min; (iii) glucuronides stimulated the release of UDP-glucuronic acid from UDP acid-loaded microsomal vesicles; (iv) trans-stimulation of UDP-glucuronic acid entry by loading of microsomal vesicles with p-nitrophenol glucuronide, phenolphthalein glucuronide, UDP-glucuronic acid and UDP-N-acetyl-glucosamine almost completely abolished the latency of UDP-glucuronosyltransferase, although mannose 6-phosphatase latency remained unaltered; (v) the loading compounds by themselves did not stimulate UDP-glucuronosyltransferase activity. This study indicates that glucuronides synthesized in the lumen of endoplasmic reticulum can leave by an antiport, which concurrently transports USP-glucuronic acid into the lumen of the endoplasmic reticulum.

  9. UDP-glucuronosyltransferases 1A6 and 1A10 catalyze reduced menadione glucuronidation

    SciTech Connect

    Nishiyama, Takahito; Ohnuma, Tomokazu; Inoue, Yuu; Kishi, Takehiko; Ogura, Kenichiro; Hiratsuka, Akira

    2008-06-27

    Menadione (2-methyl-1,4-naphthoquine), also known as vitamin K3, has been widely used as a model compound in the field of oxidative stress-related research. The metabolism of menadione has been studied, and it is known that menadione undergoes a two-electron reduction by NAD(P)H:Quinone oxidoreductase 1 (NQO1) after which the reduced form of menadione (2-methyl-1,4-naphthalenediol, menadiol) is glucuronidated and excreted in urine. To investigate which human UDP-glucuronosyltransferase (UGT) isoforms participate in the glucuronidation of menadiol reduced by NQO1 from menadione, we first constructed heterologously expressed NQO1 in Sf9 cells and tested the menadiol glucuronidating activity of 16 human recombinant UGT isoforms. Of the 16 UGT isoforms, UGTs 1A6, 1A7, 1A8, 1A9, and 1A10 catalyzed menadiol glucuronidation, and, of these, UGTs 1A6 and 1A10 catalyzed menadiol glucuronidation at much higher rates than the other UGTs. Menadiol was regioselectively glucuronidated in the manner of 4-position > 1-position by UGTs 1A7, 1A8, 1A9, and 1A10. In contrast to these UGTs, only UGT1A6 exhibited 1-menadiol-preferential glucuronidating activity. The results suggest possible detoxification pathways for quinones via NQO1 reduction followed by UGT glucuronidation.

  10. Direct measurement of salicylphenolic glucuronide in human urine.

    PubMed

    Imhoff, D M; Reece, P A; Dimitriadis, E; Ward, A D; Bochner, F

    1986-01-01

    Indirect measurement of salicylphenolic glucuronide (SPG) has suggested that the formation of this metabolite from therapeutic doses of salicyclic acid (SA) is capacity-limited in humans. A direct high performance liquid chromatographic (HPLC) assay for SPG in human urine is described. SPG was prepared by a published method and purified by HPLC. On treatment with beta-glucuronidase, SPG yielded the expected amount of SA. Spectroscopic data, melting point, and optical rotation of the glucuronide and/or its triacetyl dimethyl ester derivative were consistent with the proposed structure. SPG was assayed using a 5-micron C18 column (temperature 55 degrees C) and fluorescence detection. A nonlinear gradient mobile phase at a flow rate of 2 ml/min was used, beginning with 100% 0.1 M pH 2.1 phosphate buffer and finishing with 84% buffer, 16% acetonitrile. Total run time was 25 min. Urine (10 microliter) was injected directly on the column, and quantitation was performed using urine standards. Within-run precision for SPG ranged from 1.2% at 150 mg/L to 2.4% at 5 mg/L. The limit of detection was less than 1 mg/L. A pilot study in two volunteers, each receiving a single 500-mg dose of sodium salicylate, was carried out to validate the usefulness of the assay.

  11. 21 CFR 184.1295 - Ethyl formate.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... Substances Affirmed as GRAS § 184.1295 Ethyl formate. (a) Ethyl formate (C3H6O2, CAS Reg. No. 109-94-4) is also referred to as ethyl methanoate. It is an ester of formic acid and is prepared by esterification of formic acid with ethyl alcohol or by distillation of ethyl acetate and formic acid in the...

  12. 21 CFR 184.1295 - Ethyl formate.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... Substances Affirmed as GRAS § 184.1295 Ethyl formate. (a) Ethyl formate (C3H6O2, CAS Reg. No. 109-94-4) is also referred to as ethyl methanoate. It is an ester of formic acid and is prepared by esterification of formic acid with ethyl alcohol or by distillation of ethyl acetate and formic acid in the...

  13. Trans-stilbene oxide administration increased hepatic glucuronidation of morphine but decreased biliary excretion of morphine glucuronide in rats

    SciTech Connect

    Fuhrman-Lane, C.; Fujimoto, J.M.

    1982-09-01

    The effect of the inducing agent trans-stilbene oxide (TSO) on the metabolism and biliary excretion of (/sup 14/C)morphine was studied in the isolated in situ perfused rat liver. After administration of morphine by intraportal injection or by the segmented retrograde intrabiliary injection technique, the TSO-treated group showed a marked decrease in the biliary recovery of morphine as its glucuronide conjugate (morphine-3-glucuronide (MG)). However, recovery of MG in the venous outflow of the single pass perfusate was greatly increased. These findings suggested that TSO treatment enhanced the formation of MG from morphine and changed the primary route of hepatic elimination of MG. TSO treatment also decreased the excretion of morphine (as MG) in the bile of anesthetized renal-ligated rats. This decreased biliary function required several days to develop and appeared closely associated with the inductive effect of TSO. After i.v. administration of (/sup 14/C)MG itself, biliary recovery was also markedly decreased in TSO-treated rats. It is postulated that the effect of the TSO treatment led to either a decrease in canalicular transport of MG into bile or an increase in the efficiency of transfer of MG to the blood at the sinusoidal side of the hepatocyte. Regardless of the mechanism, the results indicate the need to study compartmentalization of drug transport and metabolism functions.

  14. Rectal administration of nicomorphine in patients improves biological availability of morphine and its glucuronide conjugates.

    PubMed

    Koopman-Kimenai, P M; Vree, T B; Booij, L H; Dirksen, R

    1994-12-02

    The pharmacokinetics of 30 mg nicomorphine after rectal administration with a suppository are described in 8 patients under combined general and epidural anaesthesia. No nicomorphine or 6-mononicotinoylmorphine could be detected in the serum. Morphine appeared almost instantaneously with a lag-time of 8 min and had a final elimination half-life of 1.48 +/- 0.48 h. Morphine was metabolized to morphine-3-glucuronide and morphine-6-glucuronide. These glucuronide conjugates appeared after a lag-time of 12 min and the half-life of these two glucuronide conjugates was similar: about 2.8 h (P > 0.8). The glucuronide conjugate of 6-mononicotinoylmorphine was not detected. In the urine only morphine and its glucuronides were found. The renal clearance value for morphine was 162 ml.min-1 and for the glucuronides 81 ml.min-1. This study shows that administration of a suppository with 30 mg nicomorphine gives an excellent absolute bioavailability of morphine and its metabolites of 88%. The lipid-soluble prodrug nicomorphine is quickly absorbed and immediately hydrolysed to morphine.

  15. Reversible binding of tolmetin, zomepirac, and their glucuronide conjugates to human serum albumin and plasma.

    PubMed

    Ojingwa, J C; Spahn-Langguth, H; Benet, L Z

    1994-02-01

    Acyl glucuronides of drugs and bilirubin have been shown in the past decade to be reactive metabolites undergoing acyl migration and irreversible binding. The latter reaction has been hypothesized to be facilitated by or to proceed through the formation of a reversible complex. Furthermore, it has been suggested that the decreased binding seen in patients with compromised excretory function may be due to competition by elevated plasma concentrations of the glucuronides. In these reversible binding studies, we characterized the extent and the "site" of binding of tolmetin, zomepirac, their glucuronides and isomeric conjugates. We also examined the displacement between the parent drugs and their glucuronide conjugates using a rapid ultrafiltration method. Tolmetin exhibited three classes of binding sites with a primary association constant of 1.7 x 10(6) M-1 (Kd1 = 0.60 microM). The primary association constant of zomepirac (1.16 x 10(6) M-1, Kd1 = 0.86 microM) is similar to that of tolmetin. The beta 1 and alpha/beta 3 glucuronides of both compounds bind to a lesser extent than their parent aglycones. The isomeric glucuronide conjugates of both compounds showed much stronger binding than the beta/1 conjugates. Of the four glucuronides investigated, tolmetin glucuronide-alpha/beta 3 isomer was bound by fatty acid free human serum albumin with the highest affinity (4.6 x 10(5) M-1, Kd = 2.22 microM). Protein binding of the parent drugs and conjugates were decreased significantly at pH 5.0. In displacement studies, except for salicylate and acetylsalicylate, drugs known to bind to Sites I and II as well as the digitoxin and tamoxifen binding sites had little inhibitory effect on the binding of tolmetin, zomepirac, and their glucuronide conjugates.

  16. Gas chromatographic-mass spectrometric assay for 6-hydroxymelatonin sulfate and 6-hydroxymelatonin glucuronide in urine

    SciTech Connect

    Francis, P.L.; Leone, A.M.; Young, I.M.; Stovell, P.; Silman, R.E.

    1987-04-01

    Circulating melatonin is hydroxylated to 6-hydroxymelatonin and excreted in urine as the sulfate and glucuronide conjugates. We extracted these two compounds from urine by using octadecylsilane-bonded silica cartridges to eliminate most of the urea and electrolytes, and silica cartridges to separate the sulfate and glucuronide conjugates. After hydrolyzing the separated conjugates enzymically, we determined the free hydroxymelatonin by gas chromatography-mass spectrometry. Though recoveries were low and variable, we were able to quantify the analyte in the original sample by adding deuterated sulfate and glucuronide conjugates to the urines before extraction.

  17. Glucuronidation and Sulfation Kinetics of Diflunisal in Man.

    NASA Astrophysics Data System (ADS)

    Loewen, Gordon Rapheal

    Diflunisal is a nonsteroidal anti-inflammatory drug used in the treatment of arthritis and musculoskeletal pain. Diflunisal exhibits concentration- and dose-dependent kinetics, the mechanism of which has not been determined. The purpose of this study was to determine the mechanism(s) responsible for non-linear disposition of diflunisal and to examine environmental factors which may affect the elimination of diflunisal. The metabolites of diflunisal, including a new metabolite, the sulphate conjugate, were purified by column and semi-preparative high pressure liquid chromatography. Assays for the quantitation of diflunisal and conjugates in urine and diflunisal in plasma were developed. Plasma protein binding of diflunisal in blank plasma and in plasma obtained following multiple doses of diflunisal was determined by equilibrium dialysis. Total body clearance of diflunisal decreased when dose increased from 100 to 750 mg. Total clearance increased when dose increased from 750 to 1000 mg. The percent of recovered dose eliminated as the acyl glucuronide decreased and the percent eliminated as the sulphate increased with increasing dose of diflunisal. Plasma protein binding of diflunisal was concentration dependent over a range of diflunisal plasma concentrations of 3 to 257 mug/ml. Total clearance, and to a lesser degree, unbound clearance of diflunisal were decreased following multiple dose administration of 250 and 500 mg diflunisal. Percent of recovered dose eliminated as the acyl glucuronide decreased and percent eliminated as the sulphate conjugate increased following multiple dosing. Plasma protein binding of diflunisal was similar in blank plasma and plasma obtained at steady state. Unbound clearance of diflunisal exceeded liver plasma flow. Frequency distributions of the elimination of the conjugates of diflunisal were normally distributed. Sex, smoking, and use of vitamins or oral contraceptives were identified as factors which may affect the elimination of

  18. Optimization to eliminate the interference of migration isomers for measuring 1-O-beta-acyl glucuronide without extensive chromatographic separation.

    PubMed

    Xue, Y-J; Akinsanya, J Billy; Raghavan, Nirmala; Zhang, Donglu

    2008-01-01

    A highly selected reaction monitoring (SRM) method has been investigated for the determination of muraglitazar 1-O-beta-acyl glucuronide in animal and human plasma without chromatographic separation of this naturally formed acyl glucuronide from its migration isomers. In the ion source or the collision cell, glucuronides are often prone to lose the dehydrated glucuronic acid (176 Da) and convert back into the parent drug (aglycone). The extent of loss of the glucuronide moiety can differ among glucuronides. For the naturally occurring muraglitazar 1-O-beta-acyl glucuronide, or its synthetic anomer 1-O-alpha-glucuronide, the loss of the glucuronide moiety was a major fragment ion. The loss of the glucuronide moiety was greater for the 1-O-beta-acyl glucuronide than the 1-O-alpha-anomer. In addition, the loss of the glucuronide moiety was insignificant (less than 0.01%) with the other glucuronide isomers (2-, 3- or 4-O, alpha or beta). Given the fact that the 1-O-alpha-anomer was a minor impurity in the muraglitazar 1-O-beta-acyl glucuronide reference standard, and not either a conversion product of 1-O-beta-acyl glucuronide or endogenously formed, the SRM transition corresponding to the loss of the glucuronide moiety was very specific for 1-O-beta-acyl glucuronide, and practically free from interference of the other isomers under optimized collision-cell conditions. As a result, extensive chromatographic separation of 1-O-beta-acyl glucuronide from its migration isomers was not required. The use of this specific SRM transition effectively reduced the separation time from 12.0 min of a long-column high-performance liquid chromatography (HPLC) method to 2.5 min by use of a shorter column. The standard curve performance and analysis results of 1-O-beta-acyl glucuronide incubation samples showed that the short-column method could produce equivalent results to the long-column method but with a 4.5-fold improvement in sample throughput. This approach may be useful for

  19. Mutual Regioselective Inhibition of Human UGT1A1-Mediated Glucuronidation of Four Flavonoids

    PubMed Central

    Ma, Guo; Wu, Baojian; Gao, Song; Yang, Zhen; Ma, Yong; Hu, Ming

    2013-01-01

    UDP-glucuronosyltransferase (UGT) 1A1-catalyzed glucuronidation is an important elimination pathway of flavonoids, and mutually inhibitory interactions may occur when two or more flavonoids are co-administered. Our recent research suggested that glucuronidation of flavonoids displayed distinct positional preferences, but whether this will lead to the mutually regioselective inhibition of UGT1A1-mediated glucuronidation of flavonoids is unknown. Therefore, we chose three monohydroxyflavone isomers 3-hydroxyflavone (3HF), 7-hydroxyflavone (7HF), 4′-hydroxyflavone (4′HF) and one trihydroxyflavone 3,7,4′-trihydroxyflavone (3,7,4′THF) as the model compounds to characterize the possible mutually regioselective inhibition of glucuronidation using expressed human UGT1A1. Apparent kinetic parameters [e.g., reaction velocity (V), Michaelis-Menten constant (Km), maximum rate of metabolism (Vmax), concentration at which inhibitor achieve 50% inhibition or IC50] and the Lineweaver-Burk plots were used to evaluate the apparent kinetic mechanisms of inhibition of glucuronidation. The results showed that UGT1A1-mediated glucuronidation of three monohydroxyflavones (i.e., 3HF, 7HF and 4′HF) and 3,7,4′THF was mutually inhibitory, and the mechanisms of inhibition appeared to be the mixed-typed inhibition. Specifically, the inhibitory effects displayed certain positional preference. Glucuronidation of 3HF was more easily inhibited by 3,7,4′THF than that of 7HF or 4′HF. Compared to 7-O-glucuronidation of 3,7,4′THF, 3-O-glucuronidation of 3,7,4′THF was more inhibited by 3HF and 4′HF, whereas glucuronidation at both 3-OH and 7-OH positions of 3,7,4′THF was more easily inhibited by 7HF than by 3HF and 4′HF. In conclusion, 3HF, 7HF, 4′HF and 3,7,4′THF were both substrates and inhibitors of UGT1A1, and they exhibited mutually regioselective inhibition of UGT1A1-mediated glucuronidation via a mixed-type inhibitory mechanism. PMID:23786524

  20. In vitro glucuronidation of the antibacterial triclocarban and its oxidative metabolites.

    PubMed

    Schebb, N H; Franze, B; Maul, R; Ranganathan, A; Hammock, B D

    2012-01-01

    Triclocarban (3,4,4'-trichlorocarbanilide; TCC) is widely used as an antibacterial in bar soaps. During use of these soaps, a significant portion of TCC is absorbed by humans. For the elimination from the body, glucuronidation plays a key role in both biliary and renal clearance. To investigate this metabolic pathway, we performed microsomal incubations of TCC and its hydroxylated metabolites 2'-OH-TCC, 3'-OH-TCC, and 6-OH-TCC. Using a new liquid chromatography-UV-mass spectrometry method, we could show a rapid glucuronidation for all OH-TCCs by the uridine-5'-diphosphate-glucuronosyltransferases (UGT) present in liver microsomes of humans (HLM), cynomolgus monkeys (CLM), rats (RLM), and mice (MLM). Among the tested human UGT isoforms, UGT1A7, UGT1A8, and UGT1A9 showed the highest activity for the conjugation of hydroxylated TCC metabolites followed by UGT1A1, UGT1A3, and UGT1A10. Due to this broad pattern of active UGTs, OH-TCCs can be efficiently glucuronidated in various tissues, as shown for microsomes from human kidney (HKM) and intestine (HIM). The major renal metabolites in humans, TCC-N-glucuronide and TCC-N'-glucuronide, were formed at very low conversion rates (<1%) by microsomal incubations. Low amounts of N-glucuronides were generated by HLM, HIM, and HKM, as well as by MLM and CLM, but not by RLM, according to the observed species specificity of this metabolic pathway. Among the human UGT isoforms, only UGT1A9 had activity for the N-glucuronidation of TCC. These results present an anomaly where in vivo the predominant urinary metabolites of TCC are N and N'-glucuronides, but these compounds are slowly produced in vitro.

  1. Glucuronidation in the chimpanzee (Pan troglodytes): studies with acetaminophen, oestradiol and morphine.

    PubMed

    Wong, H; Grace, J E; Wright, M R; Browning, M R; Grossman, S J; Bai, S A; Christ, D D

    2006-12-01

    The chimpanzee has recently been characterized as a surrogate for oxidative drug metabolism in humans and as a pharmacokinetic model for the selection of drug candidates. In the current study, the glucuronidation of acetaminophen, morphine and oestradiol was evaluated in the chimpanzee to extend the characterization of this important animal model. Following oral administration of acetaminophen (600 mg) to chimpanzees (n=2), pharmacokinetics were comparable with previously reported human values, namely mean oral clearance 0.91 vs. 0.62+/-0.05 l h-1 kg-1, apparent volume of distribution 2.29 vs. 1.65+/-0.25 l kg-1, and half-life 1.86 vs. 1.89+/-7h, for chimpanzee vs. human, respectively. Urinary excretions (percentage of dose) of acetaminophen, acetaminophen glucuronide and acetaminophen sulfate were also similar between chimpanzees and humans, namely 2.3 vs. 5.0, 63.1 vs. 54.7, and 25.0 vs. 32.3%, respectively. Acetaminophen, oestradiol and morphine glucuronide formation kinetics were investigated using chimpanzee (n=2) and pooled human liver microsomes (n=10). V(max) (app) and K(m)(app) (or S(50)(app)) for acetaminophen glucuronide, morphine 3- and 6-glucuronide, and oestradiol 3- and 17-glucuronide formation were comparable in both species. Eadie-Hofstee plots of oestradiol 3-glucuronide formation in chimpanzee microsomes were characteristic of autoactivation kinetics. Western immunoblot analysis of chimpanzee liver microsomes revealed a single immunoreactive band when probed with anti-human UGT1A1, anti-human UGT1A6, and anti-human UGT2B7. Taken collectively, these data demonstrate similar glucuronidation characteristics in chimpanzees and humans.

  2. Identification of the human UDP-glucuronosyltransferase isoforms involved in the glucuronidation of the phytochemical ferulic acid.

    PubMed

    Li, Xiaojun; Shang, Liang; Wu, Yaohua; Abbas, Suzanne; Li, Dong; Netter, Patrick; Ouzzine, Mohamed; Wang, Hui; Magdalou, Jacques

    2011-01-01

    Ferulic acid (FA), a member of the hydroxycinnamate family, is an abundant dietary antioxidant that may offer beneficial effects against cancer, cardiovascular disease, diabetes, osteoarthritis and Alzheimer's disease. In this study, evidence for sulfation and glucuronidation of FA was investigated upon incubation with human liver microsomes and cytosol. Two main glucuronides, M1 (ether O-glucuronide) and M2 (ester acylglucuronide), were formed with a similar affinity (apparent K(m) 3.53 and 5.15 mM, respectively). A phenol sulfoconjugate was also formed with a higher affinity (K(m) 0.53 mM). Identification of the UDP-glucuronosyltransferase (UGT) isoforms involved in FA glucuronidation was investigated with 12 human recombinant enzymes. FA was mainly glucuronidated by UGT1A isoforms and by UGT2B7. UGT1A4, 2B4, 2B15 and 2B17 failed to glucuronidate the substance. Examination of the kinetic constants revealed that FA was mainly glucuronidated by UGT1A1 at the two nucleophilic groups. UGT1A3 was able to glucuronidate these two positions with the same, but low, efficiency. UGT1A6 and 1A8 were involved in the formation of the ether glucuronide only, whereas UGT1A7, 1A10 and 2B7 preferentially glucuronidated the carboxyl group. Moreover, octyl gallate, a marker substrate of UGT1A1, competitively inhibited FA glucuronidation mediated by this isoform. Altogether, the results suggest that FA glucuronidation is primarily mediated by UGT1A1.

  3. In vitro glucuronidation kinetics of deoxynivalenol by human and animal microsomes and recombinant human UGT enzymes.

    PubMed

    Maul, Ronald; Warth, Benedikt; Schebb, Nils Helge; Krska, Rudolf; Koch, Matthias; Sulyok, Michael

    2015-06-01

    The mycotoxin deoxynivalenol (DON), formed by Fusarium species, is one of the most abundant mycotoxins contaminating food and feed worldwide. Upon ingestion, the majority of the toxin is excreted by humans and animal species as glucuronide conjugate. First in vitro data indicated that DON phase II metabolism is strongly species dependent. However, kinetic data on the in vitro metabolism as well as investigations on the specific enzymes responsible for DON glucuronidation in human are lacking. In the present study, the DON metabolism was investigated using human microsomal fractions and uridine-diphosphoglucuronyltransferases (UGTs) as well as liver microsomes from five animal species. Only two of the twelve tested human recombinant UGTs led to the formation of DON glucuronides with a different regiospecificity. UGT2B4 predominantly catalyzed the formation of DON-15-O-glucuronide (DON-15GlcA), while for UGT2B7 the DON-3-O-glucuronide (DON-3GlcA) metabolite prevailed. For human UGTs, liver, and intestinal microsomes, the glucuronidation activities were low. The estimated apparent intrinsic clearance (Clapp,int) for all human UGT as well as tissue homogenates was <1 mL/min mg protein. For the animal liver microsomes, moderate Clapp,int between 1.5 and 10 mL/min mg protein were calculated for carp, trout, and porcine liver. An elevated glucuronidation activity was detected for rat and bovine liver microsomes leading to Clapp,int between 20 and 80 mL/min mg protein. The obtained in vitro data points out that none of the animal models is suitable for estimating the human DON metabolism with respect to the metabolite pattern and formation rate.

  4. 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.

  5. Regioselective Glucuronidation of Flavonols by Six Human UGT1A Isoforms

    PubMed Central

    Wu, Baojian; Hu, Ming

    2012-01-01

    Purpose Flavonols, a class of polyphenols, show a variety of biological activities such as antioxidant and anticancer. However, rapid in vivo O-glucuronidation posed a challenge to develop them as therapeutic agents. The objective of this paper is to determine the regioselective glucuronidation of flavonols by UGT1A isoforms (i.e., UGT1A1, UGT1A3, UGT1A7, UGT1A8, UGT1A9 and UGT1A10). Methods The kinetics of UGT1A1-, 1A3- and 1A7~1A10-mediated metabolisms of four flavonols that contain 7-OH group were characterized and kinetic parameters (Km, Vmax and intrinsic clearance (CLint=Vmax/Km)) were determined. Results UGT1A1 and 1A3 regioselectively metabolized 7-OH, whereas UGT1A7~1A10 preferred to glucuronidate 3-OH group. UGT1A1 and UGT1A9 were the most efficient conjugating enzymes with Km of ≤1 µM and Vmax/Km of >3 ml/min/mg protein, resulting in a CLint value as high as 6 ml/min/mg protein. Additionally, the four flavonols generally strongly self-inhibited the UGT1A1-mediated glucuronidation, with Ks (substrate inhibition constant) of ≤ 5.4 µM. Conclusion UGT1A isoforms displayed distinct positional preferences between 3-OH and 7-OH in the glucuronidation of flavonols. The differentiated kinetics properties between 3-O- and 7-O- glucuronidation indicated that at least two distinct binding modes within the catalytic domain were responsible for the formation of these two glucuronide isomers. PMID:21472492

  6. The role of xenobiotic glucuronidating enzymes in drug resistance of tumour tissues and cells.

    PubMed

    Burchell, B; Baird, S; Coughtrie, M W

    1990-01-01

    Xenobiotic and endobiotic glucuronidation is regulated by many cellular features such as (a) access of substrates to a family of microsomal enzymes, the UDP-glucuronosyltransferases (UDPGTs) and (b) mechanisms of transport and excretion of glucuronides from the cell. We have isolated molecular biological probes identifying human UDPGTs to facilitate the examination of mechanisms regulating the functional expression of UDPGTs during natural development, in genetic diseases and in cancer cells. UDPGTs are encoded by a multigene family. Seven rat and four human UDPGT cDNAs have been cloned and expressed in cell culture to determine their substrate specificity and ability to glucuronidate xenobiotics. Analysis of the amino acid sequences of cloned UDPGTs has provided information about their method of synthesis and topological orientation with the lumen of the endoplasmic reticulum (ER). The location of hepatic UDPGT raises interesting questions about the role of possible transporters essential for functional glucuronidation. Measurement of the UDPGT activities and UDPGT mRNA in carcinoma tissue and in hepatic nodules from rats suggest that increases in the level of a phenol-UDPGT may contribute to drug resistance in cancer. Human UDPGTs stably expressed in cell lines have been used to study the glucuronidation of carcinogens and anticancer drugs. Human phenol UDPGT will catalyse the glucuronidation of carcinogens. The high levels of expression of human phenol UDPGT in tissue culture cells could also lead to an increased resistance to xenobiotic toxicity caused by mitoxantrone, mitomycin C, and adriamycin. This molecular cell biology approach is being used to further assess the glucuronidation and effective toxicity of anticancer drugs.

  7. Identification of a novel N-carbamoyl glucuronide: in vitro, in vivo, and mechanistic studies.

    PubMed

    Gunduz, Mithat; Argikar, Upendra A; Baeschlin, Daniel; Ferreira, Suzie; Hosagrahara, Vinayak; Harriman, Shawn

    2010-03-01

    1-[4-Aminomethyl-4-(3-chlorophenyl)-cyclohexyl]-tetrahydro-pyrimidin- 2-one, 1, was developed as an inhibitor of dipeptidyl peptidase-4 enzyme. Biotransformation studies with 1 revealed the presence of an N-carbamoyl glucuronide metabolite (M1) in rat bile and urine. N-Carbamoyl glucuronides are rarely observed, and little is understood regarding the mechanism of N-carbamoyl glucuronidation. The objectives of the current investigation were to elucidate the structure of the novel N-carbamoyl glucuronide, to investigate the mechanism of N-carbamoyl glucuronide formation in vitro using stable labeled CO(2), UDP glucuronosyltransferase (UGT) reaction phenotyping, and to assess whether M1 was formed to the same extent in vitro across species-mouse, rat, hamster, dog, monkey, and human. Structure elucidation was performed on a mass spectrometer with accurate mass measurement and MS(n) capabilities. (13)C-labeled carbon dioxide was used for identification of the mechanism of N-carbamoyl glucuronidation. Mechanistic studies with (13)C-labeled CO(2) in rat liver microsomes revealed that CO(2) from the bicarbonate buffer (in equilibrium with exogenous CO(2)) may be responsible for the formation of M1. M1 was formed in vitro in liver microsomes from multiple species, mainly rat and hamster, followed by similar formation in dog, monkey, mouse, and human. M1 could be detected in UGT1A1, UGT1A3, and UGT2B7 Supersomes in a CO(2)-rich environment. In conclusion, our study demonstrates that formation of M1 was observed in microsomal incubations across various species and strongly suggests incorporation of CO(2) from the bicarbonate buffer, in equilibrium with exogenous CO(2), into the carbamoyl moiety of the formed N-carbamoyl glucuronide.

  8. Direct radioimmunoassay of urinary estrogen and pregnanediol glucuronides during the menstrual cycle

    SciTech Connect

    Stanczyk, F.Z.; Miyakawa, I.; Goebelsmann, U.

    1980-06-15

    Assays measuring immunoreactive estrone glucuronide (E/sub 1/G), estradiol-3-glucuronide (E/sub 2/-3G), estradiol-17..beta..-glucuronide (E/sub 2/-17G), estriol-3-glucuronide (E/sub 3/-3G), estriol-16..cap alpha..-glucuronide (E/sub 3/-16G), and pregnanediol-3..cap alpha..-glucuronide (Pd-3G) directly in diluted urine were developed and validated. These estrogen and pregnanediol glucuronide fractions were measured in aliquots of 24-hour and overnight samples of urine collected daily from seven women for one menstrual cycle. Urinary hormone excretion was correlated with daily serum estradiol (E/sub 2/), progesterone (P), and lutenizing hormonee (LH) levels. A sharp midcycle LH peak preceded by a preovulatory rise in serum E/sub 2/ and followed by luteal phase serum P levels were noted in each of the seven apparently ovulatory cycles. Twenty-four-hour and overnight urinary excretion patterns of estrogen glucuronides were similar to those of serum E/sub 2/. Of the five estrogen glucuronide fractions tested, excretion of E/sub 2/-17G exhibited the earliest and steepest ascending slope of the preovulatory estrogen surge and correlated best with serum E/sub 2/ levels. Urinary excretion of E/sub 1/-G, E/sub 2/-3G, and E/sub 3/-16G also showed an early and steep preovulatory rise and preceded that of E/sub 3/-3G, whereas urinary excretion of E/sub 3/-3G exhibited the poorest correlation with serum E/sub 2/ concentrations. The urinary excretion of Pd-3G rose parallel to serum P levels and was markedly elevated 2 to 3 days after the midcycle LH peak in both 24-hour and overnight collections of urine. These results indicate that among the urinary estrogen conjugate fractions tested, E/sub 2/-17G is the one that most suitably predicts ovulation.

  9. Nitrosation Reactions of Ethyl Centralite

    DTIC Science & Technology

    1977-02-01

    up in ethyl alcohol, and the extract was treated with animai charcoal and filtered to give a yellow solution. Evaporation to half volume and dilution...stirrer, thermometer, and dropping funnel. Concentrated hydrochloric acid (25 ml) was slowly added while stirring, followed by a solution of a NaNO (6

  10. S-Ethyl dipropylthiocarbamate (EPTC)

    Integrated Risk Information System (IRIS)

    S - Ethyl dipropylthiocarbamate ( EPTC ) ; CASRN 759 - 94 - 4 Human health assessment information on a chemical substance is included in the IRIS database only after a comprehensive review of toxicity data , as outlined in the IRIS assessment development process . Sections I ( Health Hazard Assessme

  11. Detection of interstellar ethyl cyanide

    NASA Technical Reports Server (NTRS)

    Johnson, D. R.; Lovas, F. J.; Gottlieb, C. A.; Gottlieb, E. W.; Litvak, M. M.; Thaddeus, P.; Guelin, M.

    1977-01-01

    Twenty-four millimeter-wave emission lines of ethyl cyanide (CH3CH2CN) have been detected in the Orion Nebula (OMC-1) and seven in Sgr B2. To derive precise radial velocities from the astronomical data, a laboratory measurement of the rotational spectrum of ethyl cyanide has been made at frequencies above 41 GHz. In OMC-1, the rotational temperature of ethyl cyanide is 90 K (in good agreement with other molecules), the local-standard-of-rest radial velocity is 4.5 + or - 1.0 km/s (versus 8.5 km/s for most molecules), and the column density is 1.8 by 10 to the 14th power per sq cm (a surprisingly high figure for a complicated molecule). The high abundance of ethyl cyanide in the Orion Nebula suggests that ethane and perhaps larger saturated hydrocarbons may be common constituents of molecular clouds and have escaped detection only because they are nonpolar or only weakly polar.

  12. Separation and Purification of Two Flavone Glucuronides from Erigeron multiradiatus (Lindl.) Benth with Macroporous Resins

    PubMed Central

    Zhang, Zhi-feng; Liu, Yuan; Luo, Pei; Zhang, Hao

    2009-01-01

    Scutellarein-7-O-β-D-glucuronide (SG) and apigenin-7-O-β-D-glucuronide (AG) are two major bioactive constituents with known pharmacological effects in Erigeron multiradiatus. In this study, a simple method for preparative separation of the two flavone glucuronides was established with macroporous resins. The performance and adsorption characteristics of eight macroporous resins including AB-8, HPD100, HPD450, HPD600, D100, D101, D141, and D160 have been evaluated. The results confirmed that D141 resin offered the best adsorption and desorption capacities and the highest desorption ratio for the two glucuronides among the tested resins. Sorption isotherms were constructed for D141 resin under optimal ethanol conditions and fitted well to the Freundlich and Langmuir models (R2 > 0.95). Dynamic adsorption and desorption tests was performed on column packed with D141 resin. After one-run treatment with D141 resin, the two-constituent content in the final product was increased from 2.14% and 1.34% in the crude extract of Erigeron multiradiatus to 24.63% and 18.42% in the final products with the recoveries of 82.5% and 85.4%, respectively. The preparative separation of SG and AG can be easily and effectively achieved via adsorption and desorption on D141 resin, and the method developed can be referenced for large-scale separation and purification of flavone glucuronides from herbal raw materials. PMID:19918373

  13. Desorption chemical ionization and fast atom bombardment mass spectrometric studies of the glucuronide metabolites of doxylamine.

    PubMed

    Lay, J O; Korfmacher, W A; Miller, D W; Siitonen, P; Holder, C L; Gosnell, A B

    1986-11-01

    Three glucuronide metabolites of doxylamine succinate were collected in a single fraction using high-performance liquid chromatography (HPLC) from the urine of dosed male Fischer 344 rats. The metabolites were then separated using an additional HPLC step into fractions containing predominantly a single glucuronide metabolite. Analysis of the metabolites by methane and ammonia desorption chemical ionization, with and without derivatization, revealed fragment ions suggestive of a hydroxylated doxylamine moiety. Identification of the metabolites as glucuronides of doxylamine, desmethyldoxylamine and didesmethyldoxylamine was accomplished, based on determination of the molecular weight and exact mass of each metabolite using fast atom bombardment (FAB) ionization. This assignment was confirmed by the fragmentation observed in FAB mass spectrometric and tandem mass spectrometric experiments. Para-substitution of the glucuronide on the phenyl moiety was observed by 500-MHz nuclear magnetic resonance (NMR) spectrometry. A fraction containing all three glucuronide metabolites, after a single stage of HPLC separation, was also analysed by FAB mass spectrometry, and the proton- and potassium-containing quasimolecular ions for all three metabolites were observed.

  14. Isolation, purification, and structural characterization of flunixin glucuronide in the urine of greyhound dogs.

    PubMed

    Brady, T C; Kind, A J; Hyde, W H; Favrow, M; Hill, D W

    1998-04-01

    A urinary metabolite of flunixin in greyhound dogs was isolated and purified by a gradient-elution solid-phase extraction technique. The purified metabolite was shown to be hydrolyzed to free flunixin by strong base and by beta-glucuronidase, suggesting the presence of a C1-beta-glucuronide ester of flunixin. The metabolite was further characterized by positive-ion, tandem MS with electrospray ionization. Mass spectral data showed the presence of a protonated molecular ion (M+1) at m/z 473, which was consistent with the molecular weight of protonated flunixin glucuronide, and a product ion at m/z 297, which was consistent with the molecular weight of protonated flunixin. Collisionally induced dissociation of the m/z 297 product ion showed a fragmentation pattern consistent with that of standard flunixin. These data support the contention that this metabolite of flunixin in greyhound urine is the C1-beta-glucuronide of flunixin. Acyl glucuronide metabolites of some organic acid drugs have been shown to bind covalently to tissue proteins in vitro, in vivo, and ex vivo. The presence of this metabolite may, therefore, have pharmacokinetic and pharmacodynamic implications for flunixin in greyhound dogs, as well as in other animal species in which the acyl glucuronide of flunixin is a metabolite.

  15. New Flavonol Glucuronides from the Flower Buds of Syzygium aromaticum (Clove).

    PubMed

    Ryu, Byeol; Kim, Hye Mi; Lee, Jin Su; Lee, Chan Kyu; Sezirahiga, Jurdas; Woo, Jeong-Hwa; Choi, Jung-Hye; Jang, Dae Sik

    2016-04-20

    Repeated chromatography of the EtOAc-soluble fraction from the 70% EtOH extract of the flower buds of Syzygium aromaticum (clove) led to the isolation and characterization of four new flavonol glucuronides, rhamnetin-3-O-β-d-glucuronide (1), rhamnazin-3-O-β-d-glucuronide (2), rhamnazin-3-O-β-d-glucuronide-6″-methyl ester (3), and rhamnocitrin-3-O-β-d-glucuronide-6″-methyl ester (4), together with 15 flavonoids (5-19) having previously known chemical structures. The structures of the new compounds 1-4 were determined by interpretation of spectroscopic data, particularly by 1D- and 2D-NMR studies. Six flavonoids (6, 7, 9, 14, 18, and 19) were isolated from the flower buds of S. aromaticum for the first time in this study. The flavonoids were examined for their cytotoxicity against human ovarian cancer cells (A2780) using MTT assays. Among the isolates, pachypodol (19) showed the most potent cytotoxicity on A2780 cells with an IC50 value of 8.02 μM.

  16. The cynomolgus monkey (Macaca fascicularis) is the best animal model for the study of steroid glucuronidation.

    PubMed

    Barbier, Olivier; Bélanger, Alain

    2003-06-01

    Intense research efforts performed during the past decade clearly established the major role of glucuronidation and uridine-diphospho-glucuronosyltransferase (UGT) enzymes for steroid metabolism in humans. However, a clear understanding of the physiological importance of this metabolic process requires in vivo studies. Numerous evidences ascertain that simians are the most appropriate animal models for such studies. Indeed human and monkey have a similar pattern of steroidogenesis, unlike common laboratory mammals such as rat or mouse. Furthermore, human and monkey are unique in having high levels of circulating androsterone glucuronide and androstane-3alpha-diol glucuronide (3alpha-Diol-G). In addition, characterization of eight monkey UGT proteins demonstrated the similarity of their conjugation activity toward steroid hormones. Like human ones, monkey enzymes are expressed in steroid target tissues, where they preferentially glucuronidate androgen and estrogen metabolites. In monkey tissues, immunohistochemical studies demonstrated that UGT2B proteins are expressed in a cell-type specific manner in ovary and kidney, where they control androgens and aldosterone inactivation. These results identify the cynomolgus monkey as an appropriate animal model for the determination of cellular localization of UGT enzymes in steroid target tissues and for the identification of endogenous or exogenous stimuli affecting steroid glucuronidation.

  17. 49 CFR 173.322 - Ethyl chloride.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... 49 Transportation 2 2012-10-01 2012-10-01 false Ethyl chloride. 173.322 Section 173.322 Transportation Other Regulations Relating to Transportation PIPELINE AND HAZARDOUS MATERIALS SAFETY... SHIPMENTS AND PACKAGINGS Gases; Preparation and Packaging § 173.322 Ethyl chloride. Ethyl chloride must...

  18. 49 CFR 173.322 - Ethyl chloride.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 49 Transportation 2 2013-10-01 2013-10-01 false Ethyl chloride. 173.322 Section 173.322 Transportation Other Regulations Relating to Transportation PIPELINE AND HAZARDOUS MATERIALS SAFETY... SHIPMENTS AND PACKAGINGS Gases; Preparation and Packaging § 173.322 Ethyl chloride. Ethyl chloride must...

  19. 49 CFR 173.322 - Ethyl chloride.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 49 Transportation 2 2014-10-01 2014-10-01 false Ethyl chloride. 173.322 Section 173.322 Transportation Other Regulations Relating to Transportation PIPELINE AND HAZARDOUS MATERIALS SAFETY... SHIPMENTS AND PACKAGINGS Gases; Preparation and Packaging § 173.322 Ethyl chloride. Ethyl chloride must...

  20. 49 CFR 173.322 - Ethyl chloride.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 49 Transportation 2 2010-10-01 2010-10-01 false Ethyl chloride. 173.322 Section 173.322 Transportation Other Regulations Relating to Transportation PIPELINE AND HAZARDOUS MATERIALS SAFETY... SHIPMENTS AND PACKAGINGS Gases; Preparation and Packaging § 173.322 Ethyl chloride. Ethyl chloride must...

  1. 21 CFR 172.868 - Ethyl cellulose.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 3 2011-04-01 2011-04-01 false Ethyl cellulose. 172.868 Section 172.868 Food and... Multipurpose Additives § 172.868 Ethyl cellulose. The food additive ethyl cellulose may be safely used in food in accordance with the following prescribed conditions: (a) The food additive is a cellulose...

  2. 21 CFR 172.868 - Ethyl cellulose.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 3 2012-04-01 2012-04-01 false Ethyl cellulose. 172.868 Section 172.868 Food and... Multipurpose Additives § 172.868 Ethyl cellulose. The food additive ethyl cellulose may be safely used in food in accordance with the following prescribed conditions: (a) The food additive is a cellulose...

  3. 21 CFR 172.868 - Ethyl cellulose.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 3 2013-04-01 2013-04-01 false Ethyl cellulose. 172.868 Section 172.868 Food and... Multipurpose Additives § 172.868 Ethyl cellulose. The food additive ethyl cellulose may be safely used in food in accordance with the following prescribed conditions: (a) The food additive is a cellulose...

  4. 21 CFR 573.420 - Ethyl cellulose.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Ethyl cellulose. 573.420 Section 573.420 Food and... Listing § 573.420 Ethyl cellulose. The food additive ethyl cellulose may be safely used in animal feed in accordance with the following prescribed conditions: (a) The food additive is a cellulose ether...

  5. 21 CFR 172.868 - Ethyl cellulose.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Ethyl cellulose. 172.868 Section 172.868 Food and... Multipurpose Additives § 172.868 Ethyl cellulose. The food additive ethyl cellulose may be safely used in food in accordance with the following prescribed conditions: (a) The food additive is a cellulose...

  6. 21 CFR 184.1293 - Ethyl alcohol.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 3 2011-04-01 2011-04-01 false Ethyl alcohol. 184.1293 Section 184.1293 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD FOR HUMAN... Substances Affirmed as GRAS § 184.1293 Ethyl alcohol. (a) Ethyl alcohol (ethanol) is the chemical C2H5OH....

  7. 21 CFR 184.1293 - Ethyl alcohol.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 3 2013-04-01 2013-04-01 false Ethyl alcohol. 184.1293 Section 184.1293 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD FOR HUMAN... Substances Affirmed as GRAS § 184.1293 Ethyl alcohol. (a) Ethyl alcohol (ethanol) is the chemical C2H5OH....

  8. 21 CFR 184.1293 - Ethyl alcohol.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 3 2014-04-01 2014-04-01 false Ethyl alcohol. 184.1293 Section 184.1293 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) DIRECT FOOD....1293 Ethyl alcohol. (a) Ethyl alcohol (ethanol) is the chemical C2H5OH. (b) The ingredient meets...

  9. 21 CFR 184.1293 - Ethyl alcohol.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Ethyl alcohol. 184.1293 Section 184.1293 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD FOR HUMAN... Substances Affirmed as GRAS § 184.1293 Ethyl alcohol. (a) Ethyl alcohol (ethanol) is the chemical C2H5OH....

  10. 21 CFR 184.1293 - Ethyl alcohol.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 3 2012-04-01 2012-04-01 false Ethyl alcohol. 184.1293 Section 184.1293 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD FOR HUMAN... Substances Affirmed as GRAS § 184.1293 Ethyl alcohol. (a) Ethyl alcohol (ethanol) is the chemical C2H5OH....

  11. Investigation of the hepatic glucuronidation pattern of the Fusarium mycotoxin deoxynivalenol in various species.

    PubMed

    Maul, Ronald; Warth, Benedikt; Kant, Jill-Sandra; Schebb, Nils Helge; Krska, Rudolf; Koch, Matthias; Sulyok, Michael

    2012-12-17

    Deoxynivalenol (DON) is one of the most abundant mycotoxins contaminating food and feed worldwide. Upon absorption, the major portion of the toxin is excreted by humans and animal species as glucuronide. However, consistent in vitro data on DON glucuronidation are lacking. In the present study, the metabolism of DON was investigated using liver microsomes from humans and six different animal species. It was shown that all animal and human liver microsomes led to the formation of up to three different mono-O-glucuronides with significant interspecies differences. While the activity of human liver microsomes was low (0.8 to 2.2 pmol·min(-1)·mg(-1)), bovine liver and rat liver microsomes conjugated DON with activities of 525 pmol·min(-1)·mg(-1) and 80 pmol·min(-1)·mg(-1), respectively.

  12. Determination of salbutamol and salbutamol glucuronide in human urine by means of liquid chromatography-tandem mass spectrometry.

    PubMed

    Mareck, Ute; Guddat, Sven; Schwenke, Anne; Beuck, Simon; Geyer, Hans; Flenker, Ulrich; Elers, Jimmi; Backer, Vibeke; Thevis, Mario; Schänzer, Wilhelm

    2011-01-01

    The determination of salbutamol and its glucuronide in human urine following the inhalative and oral administration of therapeutic doses of salbutamol preparations was performed by means of direct urine injection utilizing liquid chromatography-tandem mass spectrometry (LC-MS/MS) and employing d(3)-salbutamol and d(3)-salbutamol glucuronide as internal standards. Unconjugated salbutamol was detected in all administration study urine samples. Salbutamol concentrations following inhalation were commonly (99%) below 1000 ng/ml whereas values after oral administration frequently (48%) exceeded this threshold. While salbutamol glucuronide was not detected in urine samples collected after inhalation of the drug, 26 out of 82 specimens obtained after oral application contained salbutamol glucuronide with a peak value of 63 ng/ml. The percentage of salbutamol glucuronide compared to unconjugated salbutamol was less than 3%. Authentic doping control urine samples indicating screening results for salbutamol less than 1000 ng/ml, showed salbutamol glucuronide concentrations between 2 and 6 ng/ml, whereas adverse analytical findings resulting from salbutamol levels higher than 1000 ng/ml, had salbutamol glucuronide values between 8 and 15 ng/ml. The approach enabled the rapid determination of salbutamol and its glucuronic acid conjugate in human urine and represents an alternative to existing procedures since time-consuming hydrolysis or derivatization steps were omitted. Moreover, the excretion of salbutamol glucuronide in human urine following the administration of salbutamol was proven.

  13. A novel method for the determination of the site of glucuronidation by ion mobility spectrometry-mass spectrometry.

    PubMed

    Shimizu, Atsushi; Ohe, Tomoyuki; Chiba, Masato

    2012-08-01

    Glucuronidation not only plays a detoxifying role in living body, but it also can complicate pharmacological and toxicological profiles of new drug candidates by forming active and reactive conjugated metabolites. The opportunity to elucidate structure of conjugated metabolites has increased in drug metabolism studies at the early development stage. General methodologies for the structure elucidation of glucuronide conjugate(s) include liquid chromatography-tandem mass spectrometry (LC-MS/MS) and NMR spectroscopy. In many cases, LC-MS/MS alone cannot unequivocally identify the site(s) of conjugation in isomeric glucuronidations. In the present study, we established a new strategy for the structure elucidation of glucuronide conjugates using ion mobility spectrometry (IMS)-mass spectrometry. Linear correlation between calculated collision cross-sections (CCS) and actual drift times from IMS was found for each set of parent compound (raloxifene, losartan, telmisartan, and estradiol) and the corresponding MS/MS product ions. Thus, obtained regression lines accurately and selectively projected the actual drift times of authentic standards of glucuronide conjugate based on the theoretical CCS values. The established method was used for the accurate assignment of predominant formation of phenolic glucuronide conjugate (SCH 60663) in the isomeric (phenolic and benzylic) glucuronidations of ezetimibe in the incubated sample with cryopreserved human hepatocytes. This application demonstrates the potential to facilitate the structure identification of glucuronide conjugates at the early development stage of new drug candidates.

  14. Structure-Dependent Deconjugation of Flavonoid Glucuronides by Human β-Glucuronidase - In Vitro and In Silico Analyses.

    PubMed

    Untergehrer, Monika; Bücherl, Daniel; Wittmann, Hans-Joachim; Strasser, Andrea; Heilmann, Jörg; Jürgenliemk, Guido

    2015-08-01

    Flavonoid glycosides are extensively metabolized to glucuronidated compounds after oral intake. Recently, a cleavage of quercetin glucuronides by β-glucuronidase has been found. To characterize the deglucuronidation reaction and its structural prerequisites among the flavonoid subtypes more precisely, four flavonol glucuronides with varying glucuronidation positions, five flavone 7-O-glucuronides with varying A- and B-ring substitution as well as one flavanone- and one isoflavone-7-O-glucuronide were analyzed in a human monocytic cell line. Investigation of the deglucuronidation rates by HPLC revealed a significant influence of the glucuronidation position on enzyme activity for flavonols. Across the flavonoid subtypes, the C-ring saturation also showed a significant influence on deglucuronidation, whereas A- and B-ring variations within the flavone-7-O-glucuronides did not affect the enzymes' activity. Results were compared to computational binding studies on human β-glucuronidase. Additionally, molecular modeling and dynamic studies were performed to obtain detailed insight into the binding and cleavage mode of the substrate at the active site of the human β-glucuronidase.

  15. Interpretation of the presence of 6-monoacetylmorphine in the absence of morphine-3-glucuronide in urine samples: evidence of heroin abuse.

    PubMed

    von Euler, Mia; Villén, Tomas; Svensson, Jan-Olof; Ståhle, Lars

    2003-10-01

    The presence of morphine in a urinary sample may be caused not only by intake of heroin but also by intake of poppy-seed-containing food shortly before urine sampling or intake of drugs containing morphine, ethyl morphine, or codeine. To facilitate the interpretation, the heroin-specific metabolite 6-monoacetylmorphine (6-MAM) can be analyzed along with morphine-3-glucuronide (M3G) in an LC-MS verification analysis. In sporadic samples positive in the immunologic opiate screening test, 6-MAM, but not M3G, was found. To systematically analyze the finding all specimens with positive 6-MAM and/or M3G found during a 1-year period were investigated (n = 1923). Of these, 423 were positive for 6-MAM. In 32 (7.6%) of the samples 6-MAM was detected while the M3G concentrations were below cutoff (300 ng/mL) and in some cases even below the limit of detection (15 ng/mL). The 32 samples with this excretion pattern came from 13 different individuals, all but one with previously known heroin abuse. Eleven urine samples, nine containing M3G and 6-MAM and two with only 6-MAM, were also analyzed for the presence of heroin. In six samples, including the two with only 6-MAM, heroin was detected. There are several plausible explanations for these findings. The intake may have taken place shortly before urine sampling. High concentrations of heroin and 6-MAM may inhibit UGT 2B7, the enzyme responsible for glucuronidation of morphine. The hydrolyzation of 6-MAM to morphine may be disturbed by either internal or external causes. To elucidate this, further studies are required. Nevertheless, our finding demonstrates that routine measurement of 6-MAM when verifying opioid-positive immunologic screening results facilitates interpretation of low concentrations of M3G in urine specimens.

  16. In vitro characterization of glucuronidation of vanillin: identification of human UDP-glucuronosyltransferases and species differences.

    PubMed

    Yu, Jian; Han, Jing-Chun; Hua, Li-Min; Gao, Ya-Jie

    2013-09-01

    Vanillin is a food flavoring agent widely utilized in foods, beverages, drugs, and perfumes and has been demonstrated to exhibit multiple pharmacological activities. Given the importance of glucuronidation in the metabolism of vanillin, the UDP-glucuronosyltransferase conjugation pathway of vanillin was investigated in this study. Vanillin glucuronide was identified by high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) and a hydrolysis reaction catalyzed by β-glucuronidase. The kinetic study showed that vanillin glucuronidation by HLMs and HIMs followed Michaelis-Menten kinetics and the kinetic parameters were as follows: 134.9 ± 13.5 μM and 81.3 ± 11.3 μM for K(m) of HLMs and HIMs, 63.8 ± 2.0 nmol/min/mg pro and 13.4 ±2.0 nmol/min/mg pro for Vmax of HLMs and HIMs. All UDP-glucuronosyltransferase (UGT) isoforms except UGT1A4, 1A9, and 2B7 showed the capability to glucuronidate vanillin, and UGT1A6 exerted the higher V(max)/K(m) values than other UGT isoforms for the glucuronidation of vanillin when assuming expression of isoforms is similar in recombinant UGTs. Kinetic analysis using liver microsomes from six studied speices indicated that vanillin had highest affinity for the monkey liver microsomes enzyme (K(m)  = 25.6 ± 3.2 μM) and the lowest affinity for the mice liver microsomes enzyme (K(m)  = 149.1 ± 18.4 μM), and intrinsic clearance was in the following order: monkey > dog > minipig > mice > rat ~ human. These data collectively provided important information for understanding glucuronidation of vanillin.

  17. Glucuronidation versus oxidation of the flavonoid galangin by human liver microsomes and hepatocytes.

    PubMed

    Otake, Yoko; Hsieh, Faye; Walle, Thomas

    2002-05-01

    In a previous study, we used human liver microsomes for the first time to study cytochrome P450 (P450)-mediated oxidation of the flavonoid galangin. The combination of CYP1A2 and CYP2C9 produced a V(max)/K(m) value of 13.6 +/- 1.1 microl/min/mg of protein. In the present extended study, we determined glucuronidation rates for galangin with the same microsomes. Two major and one minor glucuronide were identified by liquid chromatography/mass spectrometry. The V(max)/K(m) values for the two major glucuronides conjugated in the 7- and 3-positions were 155 +/- 30 and 427 +/- 26 microl/min/mg of protein, thus, exceeding that of oxidation by 11 and 31 times, respectively. This highly efficient glucuronidation appeared to be catalyzed mainly by the UDP-glucuronosyltransferase (UGT)1A9 isoform but also by UGT1A1 and UGT2B15. Sulfation of galangin by the human liver cytosol, mediated mainly but not exclusively by sulfotransferase (SULT) 1A1, also appeared to be efficient. These conclusions were strongly supported by experiments using the S9 fraction of the human liver, in which all three metabolic pathways could be directly compared. When galangin metabolism was examined in fresh plated hepatocytes from six donors, glucuronidation clearly predominated followed by sulfation. Oxidation occurred only to a minor extent in two of the donors. This study for the first time establishes that glucuronidation and sulfation of galangin, and maybe other flavonoids, are more efficient than P450-mediated oxidation, clearly being the metabolic pathways of choice in intact cells and therefore likely also in vivo.

  18. The relative protein abundance of UGT1A alternative splice variants as a key determinant of glucuronidation activity in vitro.

    PubMed

    Rouleau, Mélanie; Roberge, Joannie; Falardeau, Sarah-Ann; Villeneuve, Lyne; Guillemette, Chantal

    2013-04-01

    Alternative splicing (AS) is one of the most significant components of the functional complexity of human UDP-glucuronosyltransferase enzymes (UGTs), particularly for the UGT1A gene, which represents one of the best examples of a drug-metabolizing gene regulated by AS. Shorter UGT1A isoforms [isoform 2 (i2)] are deficient in glucuronic acid transferase activity but function as negative regulators of enzyme activity through protein-protein interaction. Their abundance, relative to active UGT1A enzymes, is expected to be a determinant of the global transferase activity of cells and tissues. Here we tested whether i2-mediated inhibition increases with greater abundance of the i2 protein relative to the isoform 1 (i1) enzyme, using the extrahepatic UGT1A7 as a model and a series of 23 human embryonic kidney 293 clonal cell lines expressing variable contents of i1 and i2 proteins. Upon normalization for i1, a significant reduction of 7-ethyl-10-hydroxycamptothecin glucuronide formation was observed for i1+i2 clones (mean of 53%) compared with the reference i1 cell line. In these clones, the i2 protein content varied greatly (38-263% relative to i1) and revealed two groups: 17 clones with i2 < i1 (60% ± 3%) and 6 clones with i2 ≥ i1 (153% ± 24%). The inhibition induced by i2 was more substantial for clones displaying i2 ≥ i1 (74.5%; P = 0.001) compared with those with i2 < i1 (45.5%). Coimmunoprecipitation supports a more substantial i1-i2 complex formation when i2 exceeds i1. We conclude that the relative abundance of regulatory i2 proteins has the potential to drastically alter the local drug metabolism in the cells, particularly when i2 surpasses the protein content of i1.

  19. Age-related increases in F344 rat intestine microsomal quercetin glucuronidation

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The objective of this study was to establish the extent age modifies intestinal quercetin glucuronidation capacity. Pooled microsomal fractions of three equidistant small intestine (SI) segments from 4, 12, 18, and 28 mo male F344 rats (n=8/group) were employed to model the enzyme kinetics of UDP-gl...

  20. DEVELOPMENT OF A CLASS-SELECTIVE ENZYME IMMUNOASSAY FOR URINARY PHENOLIC GLUCURONIDES. (R825433)

    EPA Science Inventory

    Class-selective immunoassays for the measurement of glucuronides in human urine can aid evaluation of human exposure to complex mixtures of xenobiotics. Therefore, an enzyme immunoassay (EIA) for the group-selective detection of phenolic

  1. Profiling serum bile acid glucuronides in humans: gender divergences, genetic determinants and response to fenofibrate

    PubMed Central

    Trottier, Jocelyn; Perreault, Martin; Rudkowska, Iwona; Levy, Cynthia; Dallaire-Theroux, Amélie; Verreault, Mélanie; Caron, Patrick; Staels, Bart; Vohl, Marie-Claude; Straka, Robert J.; Barbier, Olivier

    2014-01-01

    Glucuronidation, catalyzed by UDP-glucuronosyltransferase (UGT) enzymes detoxifies cholestatic bile acids (BAs). We aimed at i) characterizing the circulating BA-glucuronide (-G) pool composition in humans, ii) evaluating how sex and UGT polymorphisms influence this composition, and iii) analyzing the effects of lipid-lowering drug fenofibrate on the circulating BA-G profile in 300 volunteers and 5 cholestatic patients. Eleven BA-Gs were determined in pre- and post-fenofibrate samples. Men exhibited higher BA-G concentrations, and various genotype/BA-G associations were discovered in relevant UGT genes. The chenodeoxycholic acid-3G concentration was associated with the UGT2B7 802C>T polymorphism. Glucuronidation assays confirmed the predominant role of UGT2B7 and UGT1A4 in CDCA-3G formation. Fenofibrate exposure increased the serum levels of 5 BA-G species, including CDCA-3G, and up-regulated expression of UGT1A4, but not UGT2B7, in hepatic cells. This study demonstrates that fenofibrate stimulates BA glucuronidation in humans, and thus reduces bile acid toxicity in the liver. PMID:23756370

  2. Profiling serum bile acid glucuronides in humans: gender divergences, genetic determinants, and response to fenofibrate.

    PubMed

    Trottier, J; Perreault, M; Rudkowska, I; Levy, C; Dallaire-Theroux, A; Verreault, M; Caron, P; Staels, B; Vohl, M-C; Straka, R J; Barbier, O

    2013-10-01

    Glucuronidation, catalyzed by uridine 5'-diphospho-glucuronosyltransferase (UGT) enzymes, detoxifies cholestatic bile acids (BAs). We aimed to (i) characterize the circulating BA-glucuronide (BA-G) pool composition in humans, (ii) determine how sex and UGT polymorphisms influence this composition, and (iii) analyze the effects of the lipid-lowering drug fenofibrate on the circulating BA-G profile in 300 volunteers and 5 cholestatic patients. Eleven BA-Gs were determined in pre- and postfenofibrate samples. Men exhibited higher BA-G concentrations, and various genotype/BA-G associations were discovered in relevant UGT genes. The chenodeoxycholic acid-3G (CDCA-3G) concentration was associated with the UGT2B7 802C>T polymorphism. Glucuronidation assays confirmed the predominant role of UGT2B7 and UGT1A4 in CDCA-3G formation. Fenofibrate exposure increased the serum levels of five BA-G species, including CDCA-3G, and upregulated expression of UGT1A4, but not UGT2B7, in hepatic cells. This study demonstrated that fenofibrate stimulates BA glucuronidation in humans and thus reduces BA toxicity in the liver.

  3. Nitrosation of glycine ethyl ester and ethyl diazoacetate to give the alkylating agent and mutagen ethyl chloro(hydroximino)acetate.

    PubMed

    Zhou, Lin; Haorah, James; Chen, Sheng C; Wang, Xiaojie; Kolar, Carol; Lawson, Terence A; Mirvish, Sidney S

    2004-03-01

    Whereas nitrosation of secondary amines produces nitrosamines, amino acids with primary amino groups and glycine ethyl ester were reported to react with nitrite to give unidentified agents that alkylated 4-(p-nitrobenzyl)pyridine to produce purple dyes and be direct mutagens in the Ames test. We report here that treatment of glycine ethyl ester at 37 degrees C with excess nitrite acidified with HCl, followed by ether extraction, gave 30-40% yields of a product identified as ethyl chloro(hydroximino)acetate [ClC(=NOH)COOEt, ECHA] and a 9% yield of ethyl chloroacetate. The ECHA was identical to that synthesized by a known method from ethyl acetoacetate, strongly alkylated nitrobenzylpyridine, and may have arisen by N-nitrosation of glycine ethyl ester to give ethyl diazoacetate, which was C-nitrosated and reacted with chloride to give ECHA. Nitrosation of ethyl diazoacetate also yielded ECHA. Ethyl nitroacetate was not an intermediate as its nitrosation did not produce ECHA. ECHA reacted with aniline to give ethyl (hydroxamino)(phenylimino)acetate [PhN=C(NHOH)CO2Et]. This product was different from ethyl [(phenylamino)carbonyl]carbamate [PhNHC(=O)NHCO2Et], which was synthesized by reacting ethyl isocyanatoformate (OCN.CO2Et) with aniline. ECHA reacted with guanosine to give a derivative, which may have been a guanine-C(=NOH)CO2Et derivative. ECHA showed moderate toxicity and weak but significant mutagenicity without activation in Salmonella typhimurium TA-100 (mean, 1.31 x control value for 12-18 microg/plats) and for V79 mammalian cells (1.5-1.7 x control value for 60-100 microM). In conclusion, gastric nitrosation of glycine derivatives such as peptides with a N-terminal glycine might produce ECHA analogues that alkylate bases of gastric mucosal DNA and thereby initiate gastric cancer.

  4. Role of glucuronidation for hepatic detoxification and urinary elimination of toxic bile acids during biliary obstruction.

    PubMed

    Perreault, Martin; Białek, Andrzej; Trottier, Jocelyn; Verreault, Mélanie; Caron, Patrick; Milkiewicz, Piotr; Barbier, Olivier

    2013-01-01

    Biliary obstruction, a severe cholestatic condition, results in a huge accumulation of toxic bile acids (BA) in the liver. Glucuronidation, a conjugation reaction, is thought to protect the liver by both reducing hepatic BA toxicity and increasing their urinary elimination. The present study evaluates the contribution of each process in the overall BA detoxification by glucuronidation. Glucuronide (G), glycine, taurine conjugates, and unconjugated BAs were quantified in pre- and post-biliary stenting urine samples from 12 patients with biliary obstruction, using liquid chromatography-tandem mass spectrometry (LC-MS/MS). The same LC-MS/MS procedure was used to quantify intra- and extracellular BA-G in Hepatoma HepG2 cells. Bile acid-induced toxicity in HepG2 cells was evaluated using MTS reduction, caspase-3 and flow cytometry assays. When compared to post-treatment samples, pre-stenting urines were enriched in glucuronide-, taurine- and glycine-conjugated BAs. Biliary stenting increased the relative BA-G abundance in the urinary BA pool, and reduced the proportion of taurine- and glycine-conjugates. Lithocholic, deoxycholic and chenodeoxycholic acids were the most cytotoxic and pro-apoptotic/necrotic BAs for HepG2 cells. Other species, such as the cholic, hyocholic and hyodeoxycholic acids were nontoxic. All BA-G assayed were less toxic and displayed lower pro-apoptotic/necrotic effects than their unconjugated precursors, even if they were able to penetrate into HepG2 cells. Under severe cholestatic conditions, urinary excretion favors the elimination of amidated BAs, while glucuronidation allows the conversion of cytotoxic BAs into nontoxic derivatives.

  5. Role of Glucuronidation for Hepatic Detoxification and Urinary Elimination of Toxic Bile Acids during Biliary Obstruction

    PubMed Central

    Perreault, Martin; Białek, Andrzej; Trottier, Jocelyn; Verreault, Mélanie; Caron, Patrick; Milkiewicz, Piotr; Barbier, Olivier

    2013-01-01

    Biliary obstruction, a severe cholestatic condition, results in a huge accumulation of toxic bile acids (BA) in the liver. Glucuronidation, a conjugation reaction, is thought to protect the liver by both reducing hepatic BA toxicity and increasing their urinary elimination. The present study evaluates the contribution of each process in the overall BA detoxification by glucuronidation. Glucuronide (G), glycine, taurine conjugates, and unconjugated BAs were quantified in pre- and post-biliary stenting urine samples from 12 patients with biliary obstruction, using liquid chromatography-tandem mass spectrometry (LC-MS/MS). The same LC-MS/MS procedure was used to quantify intra- and extracellular BA-G in Hepatoma HepG2 cells. Bile acid-induced toxicity in HepG2 cells was evaluated using MTS reduction, caspase-3 and flow cytometry assays. When compared to post-treatment samples, pre-stenting urines were enriched in glucuronide-, taurine- and glycine-conjugated BAs. Biliary stenting increased the relative BA-G abundance in the urinary BA pool, and reduced the proportion of taurine- and glycine-conjugates. Lithocholic, deoxycholic and chenodeoxycholic acids were the most cytotoxic and pro-apoptotic/necrotic BAs for HepG2 cells. Other species, such as the cholic, hyocholic and hyodeoxycholic acids were nontoxic. All BA-G assayed were less toxic and displayed lower pro-apoptotic/necrotic effects than their unconjugated precursors, even if they were able to penetrate into HepG2 cells. Under severe cholestatic conditions, urinary excretion favors the elimination of amidated BAs, while glucuronidation allows the conversion of cytotoxic BAs into nontoxic derivatives. PMID:24244729

  6. Simultaneous determination of sulfation and glucuronidation of flavones in FVB mouse intestine in vitro and in vivo.

    PubMed

    Fan, Yanfang; Tang, Lan; Zhou, Juan; Feng, Qian; Xia, Bijun; Liu, Zhongqiu

    2013-04-01

    Glucuronidation and sulfation are the two major phase II metabolic pathways for flavones, natural compounds that hold great potential for improving human health. We investigated the positional preference for sulfation and glucuronidation of seven structurally similar flavones in vitro and in situ. An FVB mouse intestinal perfusion model was used in addition to three small intestine S9 fractions catalyzing sulfation only (Sult enzymes), glucuronidation only (Ugt enzymes) or both (Sult and Ugt enzymes). In both the single and co-reaction S9 systems, flavones containing 7-OH groups were conjugated only at 7-OH despite the presence of other hydroxyl groups, and 7-OH glucuronidation was faster than sulfation (P <0.05). The sulfation rate was enhanced in the Sult-Ugt co-reaction system, while glucuronidation was usually unchanged by the presence of Sult. In the intestinal perfusate, sulfation patterns were the same in the small intestine and colon, and the excretion rate of 7-O-sulfate was the fastest or second fastest. The excretion of 7-O-glucuronidates was faster in small intestine (P < 0.05) than in colon. The S9-mediated sulfation rates of the different flavones were significantly correlated with the excretion rates of the same flavones from perfused intestine. In conclusion, flavone glucuronidation and sulfation rates were sensitive to minor changes in molecular structure. In intestinal S9 fractions, both Ugts and Sults preferentially catalyzed reactions at 7-OH. The sulfation rate was significantly enhanced by simultaneous glucuronidation, but glucuronidation was unaltered by sulfation. Sulfation rates in mouse S9 fractions correlated with sulfation rates in perfused intestine.

  7. The Human UGT1A3 Enzyme Conjugates Norursodeoxycholic Acid into a C23-ester Glucuronide in the Liver*

    PubMed Central

    Trottier, Jocelyn; El Husseini, Diala; Perreault, Martin; Pâquet, Sophie; Caron, Patrick; Bourassa, Sylvie; Verreault, Mélanie; Inaba, Ted T.; Poirier, Guy G.; Bélanger, Alain; Guillemette, Chantal; Trauner, Michael; Barbier, Olivier

    2010-01-01

    Norursodeoxycholic acid (norUDCA) exhibits efficient anti-cholestatic properties in an animal model of sclerosing cholangitis. norUDCA is eliminated as a C23-ester glucuronide (norUDCA-23G) in humans. The present study aimed at identifying the human UDP-glucuronosyltransferase (UGT) enzyme(s) involved in hepatic norUDCA glucuronidation and at evaluating the consequences of single nucleotide polymorphisms in the coding region of UGT genes on norUDCA-23G formation. The effects of norUDCA on the formation of the cholestatic lithocholic acid-glucuronide derivative and of rifampicin on hepatic norUDCA glucuronidation were also explored. In vitro glucuronidation assays were performed with microsomes from human tissues (liver and intestine) and HEK293 cells expressing human UGT enzymes and variant allozymes. UGT1A3 was identified as the major hepatic UGT enzyme catalyzing the formation of norUDCA-23G. Correlation studies using samples from a human liver bank (n = 16) indicated that the level of UGT1A3 protein is a strong determinant of in vitro norUDCA glucuronidation. Analyses of the norUDCA-conjugating activity by 11 UGT1A3 variant allozymes identified three phenotypes with high, low, and intermediate capacity. norUDCA is also identified as a competitive inhibitor for the hepatic formation of the pro-cholestatic lithocholic acid-glucuronide derivative, whereas norUDCA glucuronidation is weakly stimulated by rifampicin. This study identifies human UGT1A3 as the major enzyme for the hepatic norUDCA glucuronidation and supports that some coding polymorphisms affecting the conjugating activity of UGT1A3 in vitro may alter the pharmacokinetic properties of norUDCA in cholestasis treatment. PMID:19889628

  8. Simultaneous Quantification of Buprenorphine, Norbuprenorphine, Buprenorphine-Glucuronide and Norbuprenorphine-Glucuronide in Human Umbilical Cord by Liquid Chromatography Tandem Mass Spectrometry

    PubMed Central

    Concheiro, Marta; Shakleya, Diaa M.; Huestis, Marilyn A.

    2009-01-01

    A LCMS method was developed and validated for the simultaneous determination of buprenorphine (BUP), norbuprenorphine (NBUP), buprenorphine glucuronide (BUP-Gluc) and norbuprenorphine glucuronide (NBUP-Gluc) in human umbilical cord. Quantification was achieved by selected ion monitoring of precursor ions m/z 468.4 for BUP; 414.3 for NBUP; 644.4 for BUP-Gluc and 590 for NBUP-Gluc. BUP and NBUP were identified by MS2, with m/z 396, 414 and 426 for BUP, and m/z 340, 364 and 382 for NBUP. Glucuronide conjugates were identified by MS3 with m/z 396 and 414 for BUP-Gluc and m/z 340 and 382 for NBUP-Gluc. The assay was linear 1–50 ng/g. Intra, inter-day and total assay imprecision (%RSD) were <14.5%, and analytical recovery ranged from 94.1% to 112.3% for all analytes. Extraction efficiencies were >66.3%, and process efficiency >73.4%. Matrix effect ranged, in absolute value, from 3.7% to 27.4% (CV<21.8%, n=8). The method was selective with no endogenous or exogenous interferences from 41 compounds evaluated. Sensitivity was high with limits of detection of 0.8 ng/g. In order to prove method applicability, an authentic umbilical cord obtained from an opioid-dependent pregnant woman receiving BUP pharmacotherapy was analyzed. Interestingly, BUP was not detected but concentrations of the other metabolites were NBUP-Gluc 13.4 ng/g, BUP-Gluc 3.5 ng/g and NBUP 1.2 ng/g. PMID:19406593

  9. Post-mortem levels and tissue distribution of codeine, codeine-6-glucuronide, norcodeine, morphine and morphine glucuronides in a series of codeine-related deaths.

    PubMed

    Frost, Joachim; Løkken, Trine Nordgård; Helland, Arne; Nordrum, Ivar Skjåk; Slørdal, Lars

    2016-05-01

    This article presents levels and tissue distribution of codeine, codeine-6-glucuronide (C6G), norcodeine, morphine and the morphine metabolites morphine-3-glucuronide (M3G) and morphine-6-glucuronide (M6G) in post-mortem blood (peripheral and heart blood), vitreous fluid, muscle, fat and brain tissue in a series of 23 codeine-related fatalities. CYP2D6 genotype is also determined and taken into account. Quantification of codeine, C6G, norcodeine, morphine, M3G and M6G was performed with a validated solid phase extraction LC-MS method. The series comprise 19 deaths (83%) attributed to mixed drug intoxication, 4 deaths (17%) attributed to other causes of death, and no cases of unambiguous monointoxication with codeine. The typical peripheral blood concentration pattern in individual cases was C6G≫codeine≫norcodeine>morphine, and M3G>M6G>morphine. In matrices other than blood, the concentration pattern was similar, although in a less systematic fashion. Measured concentrations were generally lower in matrices other than blood, especially in brain and fat, and in particular for the glucuronides (C6G, M3G and M6G) and, to some extent, morphine. In brain tissue, the presumed active moieties morphine and M6G were both below the LLOQ (0.0080mg/L and 0.058mg/L, respectively) in a majority of cases. In general, there was a large variability in both measured concentrations and calculated blood/tissue concentration ratios. There was also a large variability in calculated ratios of morphine to codeine, C6G to codeine and norcodeine to codeine in all matrices, and CYP2D6 genotype was not a reliable predictor of these ratios. The different blood/tissue concentration ratios showed no systematic relationship with the post-mortem interval. No coherent degradation or formation patterns for codeine, morphine, M3G and M6G were observed upon reanalysis in peripheral blood after storage.

  10. The development of a high-performance liquid chromatography-tandem mass spectrometric method for simultaneous quantification of morphine, morphine-3-β-glucuronide, morphine-6-β-glucuronide, hydromorphone, and normorphine in serum

    PubMed Central

    Sartori, David; Lewis, Tamorah; Breaud, Autumn; Clarke, William

    2015-01-01

    Objectives Development and validation of a selective, robust high-performance liquid chromatography-tandem mass spectrometric (HPLC/MS-MS) method for the quantification of morphine, morphine-3-β-glucuronide, morphine-6-β-glucuronide, hydromorphone, and normorphine in human serum. Design and methods Drug-free human serum samples spiked with morphine, morphine-3-β-glucuronide, morphine-6-β-glucuronide, hydromorphone, and normorphine were prepared by protein precipitation using methanol containing the internal standards. Samples were injected onto a Thermo Scientific AccuCore PFP column for chromatographic separation. Detection was achieved using a Thermo Scientific TSQ Vantage mass spectrometer. Assay validation followed the new Clinical and Laboratory Standards Institute (CLSI) C62-A guidelines. Results The analytical measuring range for all analytes was determined to be 5 to 1,000 ng/mL. Intra- and inter-assay precision for three quality control levels were ≤ 7.0% and ≤ 13.5%, respectively. Carryover, stability, linearity, matrix effects, extraction and processing efficiency and method comparison characteristics were acceptable relative to the CLSI C62 guidelines. Conclusion The validation of this HPLC-MS/MS method demonstrated a robust and rapid assay for the quantification of morphine, morphine-3-β-glucuronide, morphine-6-β-glucuronide, hydromorphone, and normorphine. PMID:26118474

  11. Characterization of raloxifene glucuronidation: potential role of UGT1A8 genotype on raloxifene metabolism in vivo.

    PubMed

    Sun, Dongxiao; Jones, Nathan R; Manni, Andrea; Lazarus, Philip

    2013-07-01

    Raloxifene is a second-generation selective estrogen receptor modulator used for the prevention and treatment of osteoporosis and the prevention of breast cancer in postmenopausal women. Raloxifene is extensively metabolized by glucuronidation to form raloxifene-6-glucuronide (ral-6-Gluc) and raloxifene-4'-glucuronide (ral-4'-Gluc). The goal of the present study was to determine whether functional polymorphisms in active UGTs could play a role in altered raloxifene glucuronidation in vivo. Using homogenates from HEK293 UGT-overexpressing cell lines, raloxifene was shown to be glucuronidated primarily by the hepatic UGTs 1A1 and 1A9 and the extra-hepatic UGTs 1A8 and 1A10; no detectable raloxifene glucuronidation activity was found for UGT2B enzymes. Functional UGT1A1 transcriptional promoter genotypes were significantly (Ptrend = 0.005) associated with ral-6-Gluc formation in human liver microsomes, and, consistent with the decreased raloxifene glucuronidation activities observed in vitro with cell lines overexpressing UGT1A8 variants, the UGT1A8*2 variant was significantly (P = 0.023) correlated with total raloxifene glucuronide formation in human jejunum homogenates. While ral-4'-Gluc exhibited 1:100th the anti-estrogenic activity of raloxifene itself as measured by binding to the estrogen receptor, raloxifene glucuronides comprised about 99% of the circulating raloxifene dose in raloxifene-treated subjects, with ral-4'-Gluc comprising ~70% of raloxifene glucuronides. Plasma ral-6-Gluc (Ptrend = 0.0025), ral-4'-Gluc (Ptrend = 0.001), and total raloxifene glucuronides (Ptrend = 0.001) were increased in raloxifene-treated subjects who were predicted slow metabolizers [UGT1A8 (*1/*3)] versus intermediate metabolizers [UGT1A8 (*1/*1) or UGT1A8 (*1/*2)] versus fast metabolizers [UGT1A8 (*2/*2). These data suggest that raloxifene metabolism may be dependent on UGT1A8 genotype and that UGT1A8 genotype may play an important role in overall response to raloxifene.

  12. 21 CFR 573.420 - Ethyl cellulose.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ..., FEEDS, AND RELATED PRODUCTS FOOD ADDITIVES PERMITTED IN FEED AND DRINKING WATER OF ANIMALS Food Additive Listing § 573.420 Ethyl cellulose. The food additive ethyl cellulose may be safely used in animal feed in accordance with the following prescribed conditions: (a) The food additive is a cellulose ether...

  13. 21 CFR 573.420 - Ethyl cellulose.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ..., FEEDS, AND RELATED PRODUCTS FOOD ADDITIVES PERMITTED IN FEED AND DRINKING WATER OF ANIMALS Food Additive Listing § 573.420 Ethyl cellulose. The food additive ethyl cellulose may be safely used in animal feed in accordance with the following prescribed conditions: (a) The food additive is a cellulose ether...

  14. 21 CFR 573.420 - Ethyl cellulose.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ..., FEEDS, AND RELATED PRODUCTS FOOD ADDITIVES PERMITTED IN FEED AND DRINKING WATER OF ANIMALS Food Additive Listing § 573.420 Ethyl cellulose. The food additive ethyl cellulose may be safely used in animal feed in accordance with the following prescribed conditions: (a) The food additive is a cellulose ether...

  15. 21 CFR 573.420 - Ethyl cellulose.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ..., FEEDS, AND RELATED PRODUCTS FOOD ADDITIVES PERMITTED IN FEED AND DRINKING WATER OF ANIMALS Food Additive Listing § 573.420 Ethyl cellulose. The food additive ethyl cellulose may be safely used in animal feed in accordance with the following prescribed conditions: (a) The food additive is a cellulose ether...

  16. 27 CFR 21.108 - Ethyl ether.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 27 Alcohol, Tobacco Products and Firearms 1 2010-04-01 2010-04-01 false Ethyl ether. 21.108 Section 21.108 Alcohol, Tobacco Products and Firearms ALCOHOL AND TOBACCO TAX AND TRADE BUREAU, DEPARTMENT....108 Ethyl ether. (a) Odor. Characteristic odor. (b) Specific gravity at 15.56 °/15.56 °C. Not...

  17. Soy isoflavone metabolism in cats compared with other species: Urinary metabolite concentrations and glucuronidation by liver microsomes

    PubMed Central

    Redmon, Joanna M.; Shrestha, Binu; Cerundolo, Rosario; Court, Michael H.

    2016-01-01

    Soybean is a common source of protein in many pet foods. Slow glucuronidation of soy-derived isoflavones in cats has been hypothesized to result in accumulation with adverse health consequences. Here we evaluated species’ differences in soy isoflavone glucuronidation using urine samples from cats and dogs fed a soy-based diet and liver microsomes from cats compared with microsomes from 12 other species.Significant concentrations of conjugated (but not unconjugated) genistein, daidzein, and glycitein, and the gut microbiome metabolites, dihydrogenistein and dihydrodaidzein were found in cat and dog urine samples. Substantial amounts of conjugated equol were also found in cat urine but not in dog urine.β-glucuronidase treatment showed that all these compounds were significantly glucuronidated in dog urine while only daidzein (11%) and glycitein (37%) showed any glucuronidation in cat urine suggesting that alternate metabolic pathways including sulfation predominate in cats.Glucuronidation rates of genistein, daidzein, and equol by cat livers were consistently ranked within the lowest three out of 13 species’ livers evaluated. Ferret and mongoose livers were also ranked in the lowest four species.Our results demonstrate that glucuronidation is a minor pathway for soy isoflavone metabolism in cats compared with most other species. PMID:26366946

  18. Soy isoflavone metabolism in cats compared with other species: urinary metabolite concentrations and glucuronidation by liver microsomes.

    PubMed

    Redmon, Joanna M; Shrestha, Binu; Cerundolo, Rosario; Court, Michael H

    2016-01-01

    1. Soybean is a common source of protein in many pet foods. Slow glucuronidation of soy-derived isoflavones in cats has been hypothesized to result in accumulation with adverse health consequences. Here, we evaluated species' differences in soy isoflavone glucuronidation using urine samples from cats and dogs fed a soy-based diet and liver microsomes from cats compared with microsomes from 12 other species. 2. Significant concentrations of conjugated (but not unconjugated) genistein, daidzein and glycitein, and the gut microbiome metabolites, dihydrogenistein and dihydrodaidzein, were found in cat and dog urine samples. Substantial amounts of conjugated equol were also found in cat urine but not in dog urine. 3. β-Glucuronidase treatment showed that all these compounds were significantly glucuronidated in dog urine while only daidzein (11%) and glycitein (37%) showed any glucuronidation in cat urine suggesting that alternate metabolic pathways including sulfation predominate in cats. 4. Glucuronidation rates of genistein, daidzein and equol by cat livers were consistently ranked within the lowest 3 out of 13 species' livers evaluated. Ferret and mongoose livers were also ranked in the lowest four species. 5. Our results demonstrate that glucuronidation is a minor pathway for soy isoflavone metabolism in cats compared with most other species.

  19. Glucuronide transport across the endoplasmic reticulum membrane is inhibited by epigallocatechin gallate and other green tea polyphenols.

    PubMed

    Révész, Katalin; Tütto, Anna; Margittai, Eva; Bánhegyi, Gábor; Magyar, Judit E; Mandl, József; Csala, Miklós

    2007-01-01

    Toxic endogenous or exogenous compounds can be inactivated by various conjugation reactions. Glucuronidation (i.e. conjugation with glucuronate) is especially important due to the large number of drugs and chemical carcinogens that are detoxified through this pathway. Stable and harmless glucuronides can be reactivated by enzymatic hydrolysis thus inhibitors of glucuronidase activity reduce the risk of chemical carcinogenesis. The aim of this study was to reveal whether this mechanism contributes to the anti-cancer effect of green tea flavanols, which has been shown in various animal models. Therefore, we investigated the effect of these polyphenols on deglucuronidation in rat liver microsomes and in Hepa 1c1c7 mouse hepatoma cells, using 4-methylumbelliferyl glucuronide as model substrate. Tea flavanols inhibited beta-glucuronidase in intact vesicles, where glucuronide transport across the microsomal membrane is rate-limiting, but were almost ineffective in permeabilized vesicles. Epigallocatechin gallate, the major green tea flavanol was shown to have a concentration-dependent inhibitory effect on both beta-glucuronidase activity and glucuronide transport in native vesicles. Epigallocatechin gallate also inhibited beta-glucuronidase activity in native Hepa 1c1c7 mouse hepatoma cells, while failed to affect the enzyme in alamethicin-permeabilized cells, where the endoplasmic membrane barrier was eliminated. Our findings indicate that tea flavanols inhibit deglucuronidation in the endoplasmic reticulum at the glucuronide transport stage. This phenomenon might potentially contribute to the cancer-preventing dietary or pharmacological effect attributed to these catechins.

  20. Hormonal monitoring of early pregnancy by a direct radioimmunoassay of steroid glucuronides in first morning urine

    SciTech Connect

    Mendizabal, A.F.; Quiroga, S.; Farinati, Z.; Lahoz, M.; Nagle, C.

    1984-11-01

    The usefulness of the direct 4-hour radioimmunoassay of estriol-16-glucuronide (E/sub 3/G) and pregnanediol-3-glucuronide (P/sub 2/G) in first morning urine (FMU) for establishing a prognosis of the early pregnancy outcome was evaluated in 106 patients that became pregnant. Microaliquots of FMU were serially assayed from day 3 of the conception cycle until day 80 of pregnancy. The E/sub 3/G and P/sub 2/G profiles of 19 pregnancies which terminated in spontaneous abortion with either a diagnosis of the blighted ovum syndrome (n = 11) or presumption of a corpus luteum/trophoblast failure (n = 8) have been compared with those of clinically normal pregnancies (n = 87). Normal pregnancies displayed typical patterns of E/sub 3/G and P/sub 2/G development, while variations were observed in abortive events that reflected changes of the fetoplacental unit.

  1. Synthesis and Evaluation of the Anti-Oxidant Capacity of Curcumin Glucuronides, the Major Curcumin Metabolites

    PubMed Central

    Choudhury, Ambar K.; Raja, Suganya; Mahapatra, Sanjata; Nagabhushanam, Kalyanam; Majeed, Muhammed

    2015-01-01

    Curcumin metabolites namely curcumin monoglucuronide and curcumin diglucuronide were synthesized using an alternative synthetic approach. The anti-oxidant potential of these curcumin glucuronides was compared with that of curcumin using DPPH scavenging method and Oxygen Radical Absorbance Capacity (ORAC) assay. The results show that curcumin monoglucuronide exhibits 10 fold less anti-oxidant activity (DPPH method) and the anti-oxidant capacity of curcumin diglucuronide is highly attenuated compared to the anti-oxidant activity of curcumin. PMID:26783957

  2. Reaction of primary and secondary amines to form carbamic acid glucuronides.

    PubMed

    Schaefer, William H

    2006-12-01

    Glucuronidation is an important mechanism used by mammalian systems to clear and eliminate both endogenous and foreign chemicals. Many functional groups are susceptible to conjugation with glucuronic acid, including hydroxyls, phenols, carboxyls, activated carbons, thiols, amines, and selenium. Primary and secondary amines can also react with carbon dioxide (CO(2)) via a reversible reaction to form a carbamic acid. The carbamic acid is also a substrate for glucuronidation and results in a stable carbamate glucuronide metabolite. The detection and characterization of these products has been facilitated greatly by the advent of soft ionization mass spectrometry techniques and high field NMR instrumentation. The formation of carbamate glucuronide metabolites has been described for numerous pharmaceuticals and they have been identified in all of the species commonly used in drug metabolism studies (rat, dog, mouse, rabbit, guinea pig, and human). There has been no obvious species specificity for their formation and no preference for 1 degrees or 2 degrees amines. Many biological reactions have also been described in the literature that involve the reaction of CO(2) with amino groups of biomolecules. For example, CO(2) generated from cellular respiration is expired in part through the reversible formation of a carbamate between CO(2) and the alpha-amino groups of the alpha- and beta-chains of hemoglobin. Also, carbamic acid products of several amines, such as beta-N-methylamino-L-alanine (BMAA), ethylenediamine, and L-cysteine have been implicated in toxicity. Studies suggested that a significant portion of amino-compounds in biological samples (that naturally contain CO(2)/bicarbonate) can be present as a carbamic acid.

  3. IN VITRO GLUCURONIDATION OF APREPITANT: A MODERATE INHIBITOR OF UGT2B7

    PubMed Central

    House, Larry; Ramirez, Jacqueline; Seminerio, Michael; Mirkov, Snezana; Ratain, Mark J.

    2016-01-01

    Aprepitant, an oral antiemetic, commonly used in the prevention of chemotherapy-induced nausea and vomiting, is primarily metabolized by CYP3A4. Aprepitant glucuronidation has yet to be evaluated in humans. The contribution of human UDP-glucuronosyltransferase (UGT) isoforms to the metabolism of aprepitant was investigated by performing kinetic studies, inhibition studies, and correlation analyses. In addition, aprepitant was evaluated as an inhibitor of UGTs.Glucuronidation of aprepitant was catalyzed by UGT1A4 (82%), UGT1A3 (12%), and UGT1A8 (6%) and Kms were 161.6 ± 15.6 µM, 69.4 ± 1.9 µM, and 197.1 ± 28.2 µM, respectively. Aprepitant glucuronidation was significantly correlated with both UGT1A4 substrates anastrazole and imipramine (rs = 0.77, P < 0.0001 for both substrates; n = 44), and with the UGT1A3 substrate thyroxine (rs = 0.58, P < 0.0001; n = 44).We found aprepitant to be a moderate inhibitor of UGT2B7 with a Ki of ~10 µM for 4-MU, morphine, and zidovudine. Our results suggest aprepitant can alter clearance of drugs primarily eliminated by UGT2B7. Given the likelihood for first-pass metabolism by intestinal UGT2B7, this is of particular concern for oral aprepitant co-administered with oral substrates of UGT2B7, such as zidovudine and morphine. PMID:26053558

  4. Preparation and separation of the glucuronide and sulfate conjugates of thyroxine and triiodothyronine

    SciTech Connect

    Hays, M.T.; Hsu, L.

    1987-01-01

    An enzymatic method for synthesis of labelled thyroxine glucuronide (T4G) and triiodothyronine glucuronide (T3G) from labelled thyroxine (T4) and triiodothyronine (T3) is presented. The synthetic glucuronides are completely digested by beta-glucuronidase, with recovery of the parent T4 or T3. They have distinctive elution patterns on HPLC and on Sephadex G25 chromatography, and can be clearly separated from T4 and T3 as well as from synthetic T4 sulfate (T4S) and T3 sulfate (T3S). On LH 20 chromatography, elution of T4G and T3G is intermediate between that of T4 and T3 and that of T4S and T3S. T3G can be well separated from other thyronines by HPLC alone, but T4G coelutes with rT3 on HPLC; these are then separated by adding a Sephadex G25 chromatography step. Biosynthetic /sup 131/I-T3G and /sup 125/I-T4G from the bile of a cat given /sup 131/I-T3 and /sup 125/I-T4 had similar HPLC chromatographic patterns to those of synthetic T3G and T4G. That the identified peaks from analysis of the bile were indeed T3G and T4G was confirmed by recovery of the parent T3 and T4 after beta-glucuronidase digestion.

  5. Extensive intestinal glucuronidation of raloxifene in vivo in pigs and impact for oral drug delivery.

    PubMed

    Thörn, Helena Anna; Yasin, Mohammed; Dickinson, Paul Alfred; Lennernäs, Hans

    2012-09-01

    In this study an advanced multisampling site pig model, with simultaneous venous blood sampling pre- and post liver, was applied to quantify the role of the intestine in relation to the liver in first-pass glucuronidation of raloxifene in vivo. The pharmacokinetic of raloxifene (a BCS/BDDCS class II compound) in humans is characterized by extensive metabolism (>90%) and the major metabolite is the 4'-β-glucuronide (R-4-G). Following intra-jejunal (i.j.) single dose administration in pigs raloxifene was metabolized in the gut (E(G)) during first-pass to more than 70% and a high concentration (AUC(0-6 h) ratio R-4-G/raloxifene >100) of R-4-G was reached in the portal vein. The hepatic extraction (E(H)) of raloxifene was ~50% and as in humans the bioavailability become low (~7%) in pigs. Interestingly the E(H) of raloxifene and R-4-G was time-dependent after i.j. administration. It is clear that the gut was the dominating organ in first-pass extraction of raloxifene in vivo in pigs. The quantification in this study support earlier human data and emphasize that intestinal glucuronidation should be considered early in the pharmaceutical development.

  6. Regioselective Sulfation and Glucuronidation of Phenolics: Insights into the Structural Basis of Conjugation

    PubMed Central

    Wu, Baojian; Basu, Sumit; Meng, Shengnan; Wang, Xiaoqiang; Zhang, Shuxing; Hu, Ming

    2012-01-01

    The phase II metabolism sulfation and glucuronidation, mediated by sulfotransferases (SULTs) and UDP-glucuronosyltransferases (UGTs) respectively, are significant metabolic pathways for numerous endo- and xenobiotics. Understanding of SULT/UGT substrate specificity (including regioselectivity (i.e., position preference)) is of great importance in predicting contribution of sulfation/glucuronidation to drug and metabolite disposition in vivo. This review summarizes regioselective sulfation and glucuronidation of phenolic compounds with multiple hydroxyl (OH) groups as the potential conjugation sites. The strict regioselective patterns were highlighted for several SULT and UGT isoforms towards flavonoids, a large class of natural polyphenols. To seek for a molecular-level explanation, the enzyme structures (i.e., SULT crystal structures and homology-based UGT structure models) combined with molecular docking was employed. In particular, the structural bases for regioselective metabolism of flavonoids by SULT1A3 and UGT1A1 were discussed. It was concluded that the regioselective nature of these phase II enzymes was determined by the size and shape of binding pocket. While the molecular structures of the enzymes can be used to explain regioselective metabolism regarding the binding property, predicting the turnover at different positions remains a particularly difficult task. PMID:21933112

  7. Cryptotanshinone and dihydrotanshinone I exhibit strong inhibition towards human liver microsome (HLM)-catalyzed propofol glucuronidation.

    PubMed

    Cong, Ming; Hu, Cui-Min; Cao, Yun-Feng; Fang, Zhong-Ze; Tang, Shu-Hong; Wang, Jia-Rui; Luo, Jun-Sheng

    2013-03-01

    Danshen is one of the most famous herbs in the world, and more and more danshen-prescribed drugs interactions have been reported in recent years. Evaluation of inhibition potential of danshen's major ingredients towards UDP-glucuronosyltransferases (UGTs) will be helpful for understanding detailed mechanisms for danshen-drugs interaction. Therefore, the aim of the present study is to investigate the inhibitory situation of cryptotanshinone and dihydrotanshinone I towards UGT enzyme-catalyzed propofol glucuronidation. In vitro the human liver microsome (HLM) incubation system was used, and the results showed that cryptotanshinone and dihydrotanshinone I exhibited dose-dependent inhibition towards HLM-catalyzed propofol glucuronidation. Dixon plot and Lineweaver-Burk plot showed that the inhibition type was best fit to competitive inhibition type for both cryptotanshinone and dihydrotanshinone I. The second plot using the slopes from the Lineweaver-Burk plot versus the concentrations of cryptotanshinone or dihydrotanshinone I was employed to calculate the inhibition parameters (Ki) to be 0.4 and 1.7μM, respectively. Using the reported maximum plasma concentration (Cmax), the altered in vivo exposure of propofol increased by 10% and 8.2% for the co-administration of dihydrotanshinone I and cryptotanshinone, respectively. All these results indicated the possible danshen-propofol interaction due to the inhibition of dihydrotanshinone I and cryptotanshinone towards the glucuronidation reaction of propofol.

  8. Pharmacokinetics of epidurally administered nicomorphine with its metabolites and glucuronide conjugates in patients undergoing pulmonary surgery during combined epidural local anaesthetic block and general anaesthesia.

    PubMed

    Koopman-Kimenai, P M; Vree, T B; Booij, L H; Hasenbos, M A

    1995-08-01

    After epidural administration of 15 mg 3, 6-dinicotinoylmorphine (nicomorphine) in 10 patients undergoing pulmonary surgery, the parent compound was quickly metabolized into the metabolites 6-mononicotinoylmorphine and morphine. The mean apparent half-lives (+/- SD) of elimination were 10 min (0.165 h +/- 0.053 h) for 3,6-dinicotinoylmorphine and 1.77 h +/- 1.23 h for 6-mononicotinoylmorphine. Morphine is subsequently metabolized into morphine-3-glucuronide and morphine-6-glucuronide. The apparent half-lives of morphine, morphine-3-glucuronide, and morphine-6-glucuronide are similar: 3.63 h +/- 1.63 h, 4.10 h +/- 0.57 h, and 4.20 h +/- 1.64 h respectively. The possible glucuronide conjugate of 6-mononicotinoylmorphine was not detected. The prodrug 3,6-dinicotinoylmorphine was biotransformed into three active compounds: 6-mononicotinoylmorphine, morphine, and morphine-6-glucuronide.

  9. Studies to further investigate the inhibition of human liver microsomal CYP2C8 by the acyl-β-glucuronide of gemfibrozil.

    PubMed

    Jenkins, S M; Zvyaga, T; Johnson, S R; Hurley, J; Wagner, A; Burrell, R; Turley, W; Leet, J E; Philip, T; Rodrigues, A D

    2011-12-01

    In previous studies, gemfibrozil acyl-β-glucuronide, but not gemfibrozil, was found to be a mechanism-based inhibitor of cytochrome P450 2C8. To better understand whether this inhibition is specific for gemfibrozil acyl-β-glucuronide or whether other glucuronide conjugates are potential substrates for inhibition of this enzyme, we evaluated several pharmaceutical compounds (as their acyl glucuronides) as direct-acting and metabolism-dependent inhibitors of CYP2C8 in human liver microsomes. Of 11 compounds that were evaluated as their acyl glucuronide conjugates, only gemfibrozil acyl-β-glucuronide exhibited mechanism-based inhibition, indicating that CYP2C8 mechanism-based inhibition is very specific to certain glucuronide conjugates. Structural analogs of gemfibrozil were synthesized, and their glucuronide conjugates were prepared to further examine the mechanism of inhibition. When the aromatic methyl groups on the gemfibrozil moiety were substituted with trifluoromethyls, the resulting glucuronide conjugate was a weaker inhibitor of CYP2C8 and mechanism-based inhibition was abolished. However, the glucuronide conjugates of monomethyl gemfibrozil analogs were mechanism-based inhibitors of CYP2C8, although not as potent as gemfibrozil acyl-β-glucuronide itself. The ortho-monomethyl analog was a more potent inhibitor than the meta-monomethyl analog, indicating that CYP2C8 favors the ortho position for oxidation and potential inhibition. Molecular modeling of gemfibrozil acyl-β-glucuronide in the CYP2C8 active site is consistent with the ortho-methyl position being the favored site of covalent attachment to the heme. Moreover, hydrogen bonding to four residues (Ser100, Ser103, Gln214, and Asn217) is implicated.

  10. Simultaneous Quantification of Free and Glucuronidated Cannabinoids in Human Urine by Liquid Chromatography-Tandem Mass Spectrometry

    PubMed Central

    Scheidweiler, Karl B.; Desrosiers, Nathalie A.; Huestis, Marilyn A.

    2012-01-01

    Background Cannabis is the most commonly abused drug of abuse and is commonly quantified during urine drug testing. We conducted a controlled drug administration studies investigating efficacy of urinary cannabinoid glucuronide metabolites for documenting recency of cannabis intake and for determining stability of urinary cannabinoids. Methods A liquid chromatography tandem mass spectrometry method was developed and validated quantifying Δ9-tetrahydrocannabinol (THC), 11-hydroxy-THC (11-OH-THC), 11-nor-9-carboxy-THC (THCCOOH), cannabidiol, cannabinol, THC-glucuronide and THCCOOH-glucuronide in 0.5 ml human urine via supported-liquid extraction. Chromatography was performed on an Ultra Biphenyl column with a gradient of 10 mmol/l ammonium acetate, pH 6.15 and 15% methanol in acetonitrile at 0. 4ml/min. Analytes were monitored by positive and negative mode electrospray ionization and multiple reaction monitoring mass spectrometry. Results Linear ranges were 0.5–50 ng/ml for THC-glucuronide, 1–100 ng/ml for THCCOOH, 11-OH-THC and cannabidiol, 2–100 ng/ml for THC and cannabinol, and 5–500 ng/ml for THCCOOH-glucuronide (R2>0.99). Mean extraction efficiencies were 34–73% with analytical recovery (bias) 80.5–118.0% and total imprecision 3.0–10.2% coefficient of variation. Conclusion This method simultaneously quantifies urinary cannabinoids and phase II glucuronide metabolites, and enables evaluation of urinary cannabinoid glucuronides for documenting recency of cannabis intake and cannabinoid stability. The assay is applicable for routine urine cannabinoid testing. PMID:22771478

  11. Quaternary ammonium-linked glucuronidation of tamoxifen by human liver microsomes and UDP-glucuronosyltransferase 1A4.

    PubMed

    Kaku, Teppei; Ogura, Kenichiro; Nishiyama, Takahito; Ohnuma, Tomokazu; Muro, Kei; Hiratsuka, Akira

    2004-06-01

    Tamoxifen (TAM), a nonsteroidal antiestrogen, is the most widely used drug for chemotherapy of hormone-dependent breast cancer in women. In the present study, we found a new potential metabolic pathway of TAM via N-linked glucuronic acid conjugation for excretion in humans. TAM N(+)-glucuronide was isolated from a reaction mixture consisting of TAM and human liver microsomes fortified with UDP-glucuronic acid (UDPGA) and identified with a synthetic specimen by high-performance liquid chromatography-electrospray ionization-mass spectrometry. However, no TAM-glucuronidating activity was detected in microsomes from rat, mouse, monkey, dog, and guinea pig livers. A strong correlation (r(2) =0.92 ) was observed between N-glucuronidating activities toward TAM and trifluoperazine, a probe substrate for human UDP-glucuronosyltransferase (UGT) 1A4, in human liver microsomes from eight donors (five females, three males). However, no correlation ( (r(2) =0.02 )) was observed in the activities between 7-hydroxy-4-(trifluoromethyl)coumarin and TAM. Only UGT1A4 catalyzed the N-linked glucuronidation of TAM among recombinant UGTs (UGT1A1, UGT1A3, UGT1A4, UGT1A6, UGT1A9, UGT2B4, UGT2B7, UGT2B15, and UGT2B17) expressed in insect cells. Apparent K(m) values for TAM N-glucuronidation by human liver microsomes and recombinant UGT1A4 were 35.8 and 32.4 microM, respectively. These results strongly suggested that UGT1A4 could play a role in metabolism and excretion of TAM without Phase I metabolism in human liver. TAM N(+)-glucuronide still had binding affinity similar to TAM itself for human estrogen receptors, ERalpha and ERbeta, suggesting that TAM N(+)-glucuronide might contribute to the biological activity of TAM in vivo.

  12. Process for the preparation of ethyl benzene

    DOEpatents

    Smith, L.A. Jr.; Arganbright, R.P.; Hearn, D.

    1995-12-19

    Ethyl benzene is produced in a catalyst bed under 0.25 to 50 atmospheres of pressure and at temperatures in the range of 50 C to 300 C, using as the catalyst a mole sieve characterized as acidic by feeding ethylene to the catalyst bed while benzene is conveniently added through the reflux to result in a molar excess present in the reactor to that required to react with ethylene, thereby reacting substantially all of the ethylene and recovering benzene as the principal overhead and ethyl benzene and diethyl benzene in the bottoms. The bottoms are fractionated, the ethyl benzene recovered and the bottoms are contacted with benzene in the liquid phase in a fixed bed straight pass reactor under conditions to transalkylate the benzene thereby converting most of the diethyl benzene to ethyl benzene which is again separated and recovered. 2 figs.

  13. Process for the preparation of ethyl benzene

    DOEpatents

    Smith, Jr., Lawrence A.; Arganbright, Robert P.; Hearn, Dennis

    1995-01-01

    Ethyl benzene is produced in a catalyst bed under 0.25 to 50 atmospheres of pressure and at temperatures in the range of 50.degree. C. to 300.degree. C., using as the catalyst a mole sieve characterized as acidic by feeding ethylene to the catalyst bed while benzene is conveniently added through the reflux to result in a molar excess present in the reactor to that required to react with ethylene, thereby reacting substantially all of the ethylene and recovering benzene as the principal overhead and ethyl benzene and diethyl benzene in the bottoms. The bottoms are fractionated, the ethyl benzene recovered and the bottoms are contacted with benzene in the liquid phase in a fixed bed straight pass reactor under conditions to transalkylate the benzene thereby converting most of the diethyl benzene to ethyl benzene which is again separated and recovered.

  14. Methyl Ethyl Ketoxime; Final Test Rule

    EPA Pesticide Factsheets

    EPA is issuing this final test rule under section 4 of the Toxic Substances Control Act (TSCA), requiring manufacturers and processors of methyl ethyl ketoxime (MEKO, CAS No. 96-29-7) to perform testing for health effects.

  15. Day-to-day variations during clinical drug monitoring of morphine, morphine-3-glucuronide and morphine-6-glucuronide serum concentrations in cancer patients. A prospective observational study

    PubMed Central

    Klepstad, Pål; Hilton, Priscilla; Moen, Jorunn; Kaasa, Stein; Borchgrevink, Petter C; Zahlsen, Kolbjørn; Dale, Ola

    2004-01-01

    Background The feasibility of drug monitoring of serum concentrations of morphine, morphine-6-glucuronide (M6G) and morphine-3-glucuronide (M3G) during chronic morphine therapy is not established. One important factor relevant to drug monitoring is to what extent morphine, M6G and M3G serum concentrations fluctuate during stable morphine treatment. Methods We included twenty-nine patients admitted to a palliative care unit receiving oral morphine (n = 19) or continuous subcutaneous (sc) morphine infusions (n = 10). Serum concentrations of morphine, M6G and M3G were obtained at the same time on four consecutive days. If readmitted, the patients were followed for another trial period. Day-to-day variations in serum concentrations and ratios were determined by estimating the percent coefficient of variation (CV = (mean/SD) ×100). Results The patients' median morphine doses were 90 (range; 20–1460) mg/24 h and 135 (range; 30–440) mg/24 h during oral and sc administration, respectively. Intraindividual fluctuations of serum concentrations estimated by median coefficients of day-to-day variation were in the oral group for morphine 46%, for M6G 25% and for M3G 18%. The median coefficients of variation were lower in patients receiving continuous sc morphine infusions (morphine 10%, M6G 13%, M3G 9%). Conclusion These findings indicate that serum concentrations of morphine and morphine metabolites fluctuate. The fluctuations found in our study are not explained by changes in morphine doses, administration of other drugs or by time for collection of blood samples. As expected the day-to-day variation was lower in patients receiving continuous sc morphine infusions compared with patients receiving oral morphine. PMID:15461818

  16. Methylation, Glucuronidation, and Sulfonation of Daphnetin in Human Hepatic Preparations In Vitro: Metabolic Profiling, Pathway Comparison, and Bioactivity Analysis.

    PubMed

    Liang, Si-Cheng; Xia, Yang-Liu; Hou, Jie; Ge, Guang-Bo; Zhang, Jiang-Wei; He, Yu-Qi; Wang, Jia-Yue; Qi, Xiao-Yi; Yang, Ling

    2016-02-01

    Our previous study demonstrated that daphnetin is subject to glucuronidation in vitro. However, daphnetin metabolism is still poorly documented. This study aimed to investigate daphnetin metabolism and its consequent effect on the bioactivity. Metabolic profiles obtained by human liver S9 fractions and human hepatocytes showed that daphnetin was metabolized by glucuronidation, sulfonation, and methylation to form 6 conjugates which were synthesized and identified as 7-O-glucuronide, 8-O-glucuronide, 7-O-sulfate and 8-O-sulfate, 8-O-methylate, and 7-O-suflo-8-O-methylate. Regioselective 8-O-methylation of daphnetin was investigated using in silico docking calculations, and the results suggested that a close proximity (2.03 Å) of 8-OH to the critical residue Lysine 144 might be the responsible mechanism. Compared with glucuronidation and sulfonation pathways, the methylation of daphnetin had a high clearance rate (470 μL/min/mg) in human liver S9 fractions and contributed to a large amount (37.3%) of the methyl-derived metabolites in human hepatocyte. Reaction phenotyping studies showed the major role of SULT1A1, -1A2, and -1A3 in daphnetin sulfonation, and soluble COMT in daphnetin 8-O-methylation. Of the metabolites, only 8-O-methyldaphnetin exhibited an inhibitory activity on lymphocyte proliferation comparable to that of daphnetin. In conclusion, methylation is a crucial pathway for daphnetin clearance and might be involved in pharmacologic actions of daphnetin in humans.

  17. Comparison of enzymically glucuronidated flavonoids with flavonoid aglycones in an in vitro cellular model of oxidative stress protection.

    PubMed

    Stevenson, David E; Cooney, Janine M; Jensen, Dwayne J; Wibisono, Reginald; Adaim, Aselle; Skinner, Margot A; Zhang, Jingli

    2008-01-01

    This study modeled, in vitro, the potential effect of conjugative (phase II) metabolism on the cytoprotective capacity of fruit flavonoids against oxidative stress. Flavonoid aglycones were compared with their corresponding isomeric mixtures of glucuronides for their ability to enhance the survival of cultured human Jurkat T and neuroblastoma cells stressed with hydrogen peroxide. Various polyphenolic compounds were tested as substrates in vitro for an ovine liver glucuronyl transferase preparation. Flavonoids and their glycoside derivatives were found to be good substrates, whereas phenolic acids were either poor or nonsubstrates. Five common flavonoids were glucuronidated to prepare mixtures for bioassay testing. Glucuronidation generally weakened the cytoprotective capacities of flavonoids (in the presence of H(2)O(2)), but some compounds were weakened much more than others. The concentration that halved cell death was well below 0.5 microM for most flavonoids tested, but glucuronidation increased median effective concentration values to a range of 1-16 microM. This compares with the generally accepted physiological range (0.1-10 microM) for circulating dietary polyphenolics detected in the body. Therefore, some flavonoids may retain a reduced cytoprotective capacity in vitro, after glucuronidation, whereas others may be effectively inactivated.

  18. Structures of (-)-epicatechin glucuronide identified from plasma and urine after oral ingestion of (-)-epicatechin: differences between human and rat.

    PubMed

    Natsume, Midori; Osakabe, Naomi; Oyama, Makoto; Sasaki, Motoko; Baba, Seigo; Nakamura, Yoshimasa; Osawa, Toshihiko; Terao, Junji

    2003-04-01

    (-)-epicatechin is one of the most potent antioxidants present in the human diet. Particularly high levels are found in black tea, apples, and chocolate. High intake of catechins has been associated with reduced risk of cardiovascular diseases. There have been several reports concerning the bioavailability of catechins, however, the chemical structure of (-)-epicatechin metabolites in blood, tissues, and urine remains unclear. In the present study, we purified and elucidated the chemical structure of (-)-epicatechin metabolites in human and rat urine after oral administration. Three metabolites were purified from human urine including (-)-epicatechin-3'-O-glucuronide, 4'-O-methyl-(-)-epicatechin-3'-O-glucuronide, and 4'-O-methyl-(-)-epicatechin-5 or 7-O-glucuronide, according to 1H- and 13C-NMR, HMBC, and LC-MS analyses. The metabolites purified from rat urine were 3'-O-methyl-(-)-epicatechin, (-)-epicatechin-7-O-glucuronide, and 3'-O-methyl-(-)-epicatechin-7-O-glucuronide. These compounds were also detected in the blood of humans and rats by LC-MS. The presence of these metabolites in blood and urine suggests that catechins are metabolized and circulated in the body after administration of catechin-containing foods.

  19. 40 CFR 721.10595 - Octadecen-1-aminium, N-ethyl-N,N-dimethy-, ethyl sulfate (1:1).

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 32 2013-07-01 2013-07-01 false Octadecen-1-aminium, N-ethyl-N,N... Significant New Uses for Specific Chemical Substances § 721.10595 Octadecen-1-aminium, N-ethyl-N,N-dimethy... chemical substance identified as octadecen-1-aminium, N-ethyl-N,N-dimethy-, ethyl sulfate (1:1) (PMN...

  20. [Glucuronidation of antitumour therapeutics--detoxification, mechanism of resistance or prodrug formation?].

    PubMed

    Mróz, Anna; Mazerska, Zofia

    2015-12-31

    The physiological role of phase I and II of xenobiotic biotransformations is their detoxification and better excretion outside the organism. UDP-glucuronosyltransferases (UGTs) being the enzymes of phase II metabolism catalyse the conjugation of glucuronic acid to the lipophilic substrate by its specific nucleophilic group. UGT isoenzymes of various substrate specificities and different expression profiles in selected tissues belong to the large UGT superfamily. Usually, glucuronidation is the detoxification process, but sometimes (morphine, tamoxifen) glucuronides express biological activity higher than or comparable to the native compound. The level of UGT gene expression is individual for patients, because of their genetic status as well as epigenetic conditions. Also, xenobiotics are able to modulate UGT level and gene expression by the interaction with nuclear receptors. Moreover, one can find a lower level of UGT in the tumour compared to normal tissue, which results in the protection against deactivation of the drug and in the promotion of its selective activity in tumor tissue. On the other hand, UGT activity is considered as the possible cause of resistance to chemotherapy. Metabolism by hepatic and intestinal UGT isoenzymes is responsible for the "first-pass effect", whereas acquired resistance consists in the induction of UGT gene expression by the chemotherapeutic or its metabolite. Moreover, UGT induction can be associated with the induction of membrane transporters, particularly proteins of the ABC family, responsible for drug excretion outside the cell. The above resistance effects can be fortified by the overexpression of selected UGT isoenzymes sometimes observed in specific types of tumours. It is also considered that many advanced tumours are characterized by a higher level of β-glucuronidase. This enzyme has a chance to be the molecular target of directed antitumour therapy, as it catalyses β-glucuronide hydrolysis, leading to active aglycones.

  1. Interactions between morphine and the morphine-glucuronides measured by conditioned place preference and locomotor activity.

    PubMed

    Vindenes, Vigdis; Ripel, Ase; Handal, Marte; Boix, Fernando; Mørland, Jørg

    2009-07-01

    After intake of heroin or morphine, active metabolites are formed in the body. The two most important morphine metabolites are morphine-6-glucuronide (M6G) and morphine-3-glucuronide (M3G). M6G and M3G are present for longer time periods and in higher concentrations than the parent drug, but their potential contribution to reward and to development of dependence and addiction is not clear. We tested the effects of morphine and M6G separately (doses of 10, 20, 30 and 50 micromol/kg), administered together, and also in combination with with 200 microm l/kg M3G in male C57BL/6J-Bom mice. M3G in doses of 50, 100, 200, 300 and 400 micromol/kg were also tested alone. We evaluated the rewarding effects in a conditioning place preference (CPP) model and the psychomotor stimulating effects by recording locomotor activity. Mice were subjected to three consecutive conditioning days with drugs or saline before testing. Changes in locomotor activity from conditioning day one to day three were also compared to the expression of CPP on the test day. This study revealed that coadministration of morphine and M6G induced CPP of similar magnitude to the sum of equimolar doses of these compounds alone, and different ratios of the two drugs did not affect the results. M3G did not cause CPP and reduced the CPP induced by both morphine and M6G when coadministered with these drugs. Morphine induced locomotor activity was reduced by coadministration of M3G, but this was not seen when M3G was co-injected with M6G. The changes in locomotor activity during the conditioning periods did not correlated with the expression of CPP. This study revealed that the morphine-glucuronides in different and complex ways can influence the pharmacological effects of psychomotor activation and reward observed after intake of morphine.

  2. Glucuronidated Quercetin Lowers Blood Pressure in Spontaneously Hypertensive Rats via Deconjugation

    PubMed Central

    Galindo, Pilar; Rodriguez-Gómez, Isabel; González-Manzano, Susana; Dueñas, Montserrat; Jiménez, Rosario; Menéndez, Carmen; Vargas, Félix; Tamargo, Juan; Santos-Buelga, Celestino; Pérez-Vizcaíno, Francisco; Duarte, Juan

    2012-01-01

    Background Chronic oral quercetin reduces blood pressure and restores endothelial dysfunction in hypertensive animals. However, quercetin (aglycone) is usually not present in plasma, because it is rapidly metabolized into conjugated, mostly inactive, metabolites. The aim of the study is to analyze whether deconjugation of these metabolites is involved in the blood pressure lowering effect of quercetin. Methodology/Principal Findings We have analyzed the effects on blood pressure and vascular function in vitro of the conjugated metabolites of quercetin (quercetin-3-glucuronide, Q3GA; isorhamnetin-3-glucuronide, I3GA; and quercetin-3′-sulfate, Q3'S) in spontaneously hypertensive rats (SHR). Q3GA and I3GA (1 mg/kg i.v.), but not Q3'S, progressively reduced mean blood pressure (MBP), measured in conscious SHR. The hypotensive effect of Q3GA was abolished in SHR treated with the specific inhibitor of β-glucuronidase, saccharic acid 1,4-lactone (SAL, 10 mg/ml). In mesenteric arteries, unlike quercetin, Q3GA had no inhibitory effect in the contractile response to phenylephrine after 30 min of incubation. However, after 1 hour of incubation Q3GA strongly reduced this contractile response and this effect was prevented by SAL. Oral administration of quercetin (10 mg/Kg) induced a progressive decrease in MBP, which was also suppressed by SAL. Conclusions Conjugated metabolites are involved in the in vivo antihypertensive effect of quercetin, acting as molecules for the plasmatic transport of quercetin to the target tissues. Quercetin released from its glucuronidated metabolites could be responsible for its vasorelaxant and hypotensive effect. PMID:22427863

  3. Milk Thistle Constituents Inhibit Raloxifene Intestinal Glucuronidation: A Potential Clinically Relevant Natural Product-Drug Interaction.

    PubMed

    Gufford, Brandon T; Chen, Gang; Vergara, Ana G; Lazarus, Philip; Oberlies, Nicholas H; Paine, Mary F

    2015-09-01

    Women at high risk of developing breast cancer are prescribed selective estrogen response modulators, including raloxifene, as chemoprevention. Patients often seek complementary and alternative treatment modalities, including herbal products, to supplement prescribed medications. Milk thistle preparations, including silibinin and silymarin, are top-selling herbal products that may be consumed by women taking raloxifene, which undergoes extensive first-pass glucuronidation in the intestine. Key constituents in milk thistle, flavonolignans, were previously shown to be potent inhibitors of intestinal UDP-glucuronosyl transferases (UGTs), with IC50s ≤ 10 μM. Taken together, milk thistle preparations may perpetrate unwanted interactions with raloxifene. The objective of this work was to evaluate the inhibitory effects of individual milk thistle constituents on the intestinal glucuronidation of raloxifene using human intestinal microsomes and human embryonic kidney cell lysates overexpressing UGT1A1, UGT1A8, and UGT1A10, isoforms highly expressed in the intestine that are critical to raloxifene clearance. The flavonolignans silybin A and silybin B were potent inhibitors of both raloxifene 4'- and 6-glucuronidation in all enzyme systems. The Kis (human intestinal microsomes, 27-66 µM; UGT1A1, 3.2-8.3 µM; UGT1A8, 19-73 µM; and UGT1A10, 65-120 µM) encompassed reported intestinal tissue concentrations (20-310 µM), prompting prediction of clinical interaction risk using a mechanistic static model. Silibinin and silymarin were predicted to increase raloxifene systemic exposure by 4- to 5-fold, indicating high interaction risk that merits further evaluation. This systematic investigation of the potential interaction between a widely used herbal product and chemopreventive agent underscores the importance of understanding natural product-drug interactions in the context of cancer prevention.

  4. In Vitro Glucuronidation of Fenofibric Acid by Human UDP-Glucuronosyltransferases and Liver Microsomes

    PubMed Central

    Tojcic, Jelena; Benoit-Biancamano, Marie-Odile; Court, Michael H.; Straka, Robert J.; Caron, Patrick

    2009-01-01

    Fenofibric acid (FA), the active moiety of fenofibrate, is an agonist of the peroxisome proliferator-activated nuclear receptor α that modulates triglyceride and cholesterol profiles. Lipid response to fenofibrate and FA serum concentrations is highly variable. Although FA is reported to be almost exclusively inactivated by UDP-glucuronosyltransferases (UGTs) into FA-glucuronide (FA-G), the contribution of UGT isoenzymes has never been systematically assessed. Heterologously expressed human UGT1A and UGT2B and their coding variants were tested for FA glucuronidation using liquid chromatography/mass spectrometry. Recombinant UGT2B7 presented the highest Vmax/Km value (2.10 μl/min/mg), 16-fold higher than the activity of other reactive UGTs, namely, UGT1A3, UGT1A6, and UGT1A9 (0.13, 0.09, and 0.02 μl/min/mg, respectively). UGT2B7.1 (His268) and UGT2B7.2 (Tyr268) enzyme activity was similar, whereas UGT1A3.2 (R11A47), UGT1A3.3 (Trp11), and UGT1A9.3 (Thr33) showed 61 to 96% reduced Vmax/Km values compared with the respective (1) reference proteins. FA-G formation by a human liver bank (n = 48) varied by 10-fold, but the rate of formation was not associated with common genetic variations in UGT1A3, UGT1A6, UGT1A9, and UGT2B7. Correlation with activities for the probe substrates zidovudine (UGT2B7; r2 = 0.75), mycophenolic acid (UGT1A9; r2 = 0.42), fulvestrant (UGT1A3; r2 = 0.36), but not serotonin (UGT1A6; r2 = 0.06) indicated a primary role for UGT2B7 and lesser roles of UGT1A9 and UGT1A3 in hepatic FA glucuronidation. This was confirmed by a strong correlation of FA-G formation with UGT2B7 protein content and inhibition by fluconazole, a known UGT2B7 selective inhibitor. Additional studies are required to identify genetic factors contributing to the observed FA glucuronidation variability. PMID:19661212

  5. Two new glucuronide saponins, Achyranthosides G and H, from Achyranthes fauriei root.

    PubMed

    Ando, Hidehiro; Fukumura, Motonori; Hori, Yumiko; Hirai, Yasuaki; Toriizuka, Kazuo; Kuchino, Yoshiyuki; Ida, Yoshiteru

    2008-01-01

    Two oleanolic acid saponins named achyranthosides G (1) and H (2) were newly isolated from Achyranthes fauriei root as methyl esters in addition to methyl esters of achyranthosides A - F and five oleanolic acid glucuronides (chikusetsusaponins IVa, V, 28-deglucosyl chikusetsusaponin V, pseudoginsenoside RT(1), and oleanolic acid 3-O-beta-D-glucuronopyranoside) as well as oleanolic acid 28-O-beta-D-glucopyranoside, beta-ecdysterone, and polypodine B. Their structures were characterized as follows on the basis of the chemical and spectroscopic evidences.

  6. Neuroexcitatory effects of morphine and hydromorphone: evidence implicating the 3-glucuronide metabolites.

    PubMed

    Smith, M T

    2000-07-01

    1. Morphine is recommended by the World Health Organization as the drug of choice for the management of moderate to severe cancer pain. 2. Education of health professionals in the past decade has resulted in a large increase in the prescribing of opioids, such as morphine, and in the magnitude of the doses administered, resulting in an improvement in the quality of pain relief available for many cancer patients. 3. However, the reported incidence of neuroexcitatory side effects (allodynia, myoclonus, seizures) in patients administered large doses of systemic morphine or its structural analogue, hydromorphone (HMOR), has also increased. 4. Clinically, increasing the magnitude of the morphine or HMOR dose administered to patients already exhibiting neuroexcitatory opioid related side effects, results in an exacerbation rather than an attenuation of the excitatory behaviours. 5. In contrast, cessation of the opioid or rotation to a structurally dissimilar opioid (e.g. from morphine/HMOR to methadone or fentanyl), usually results in a restoration of analgesia and resolution of the neuroexcitatory opioid side effects over a period of hours to days. 6. To explain the clinical success of 'opioid rotation', it is essential to understand the in vivo metabolic fate of morphine and HMOR. 7. Following systemic administration, morphine and HMOR are metabolized primarily to the corresponding 3-glucuronide metabolites, morphine-3-glucuronide (M3G) and hydromorphone-3-glucuronide (H3G), which are not only devoid of analgesic activity but evoke a range of dose-dependent excitatory behaviours, including allodynia, myoclonus and seizures, following intracerebroventricular (i.c.v.) administration to rats. 8. Several studies have shown that, following chronic oral or subcutaneous morphine administration to patients with cancer pain, the cerebrospinal fluid (CSF) concentrations of M3G exceed those of morphine and morphine-6-glucuronide (analgesically active morphine metabolite) by

  7. 2D QSAR Study for Gemfibrozil Glucuronide as the Mechanism-based Inhibitor of CYP2C8

    PubMed Central

    Taxak, N.; Bharatam, P. V.

    2013-01-01

    Mechanism-based inhibition of cytochrome P450 involves the bioactivation of the drug to a reactive metabolite, which leads to cytochrome inhibition via various mechanisms. This is generally seen in the Phase I of drug metabolism. However, gemfibrozil (hypolipidemic drug) leads to mechanism-based inhibition after generating glucuronide conjugate (gemfibrozil acyl-β-glucuronide) in the Phase II metabolism reaction. The mechanism involves the covalent binding of the benzyl radical (generated from the oxidation of aromatic methyl group in conjugate) to the heme of CYP2C8. This article deals with the development of a 2D QSAR model based on the inhibitory potential of gemfibrozil, its analogues and corresponding glucuronide conjugates in inhibiting the CYP2C8-catalysed amodiaquine N-deethylation. The 2D QSAR model was developed using multiple linear regression analysis in Accelrys Discovery Studio 2.5 and helps in identifying the descriptors, which are actually contributing to the inhibitory potency of the molecules studied. The built model was further validated using leave one out method. The best quantitative structure activity relationship model was selected having a correlation coefficient (r) of 0.814 and cross-validated correlation coefficient (q2) of 0.799. 2D QSAR revealed the importance of volume descriptor (Mor15v), shape descriptor (SP09) and 3D matrix-based descriptor (SpMax_RG) in defining the activity for this series of molecules. It was observed that volume and 3D matrix-based descriptors were crucial in imparting higher potency to gemfibrozil glucuronide conjugate, as compared with other molecules. The results obtained from the present study may be useful in predicting the inhibitory potential (IC50 for CYP2C8 inhibition) of the glucuronide conjugates of new molecules and compare with the standard gemfibrozil acyl-β-glucuronide (in terms of pIC50 values) in early stages of drug discovery and development. PMID:24591743

  8. Identification of Recent Cannabis Use: Whole-Blood and Plasma Free and Glucuronidated Cannabinoid Pharmacokinetics following Controlled Smoked Cannabis Administration

    PubMed Central

    Schwope, David M.; Karschner, Erin L.; Gorelick, David A.; Huestis, Marilyn A.

    2013-01-01

    BACKGROUND Δ9-Tetrahydrocannabinol (THC) is the most frequently observed illicit drug in investigations of accidents and driving under the influence of drugs. THC-glucuronide has been suggested as a marker of recent cannabis use, but there are no blood data following controlled THC administration to test this hypothesis. Furthermore, there are no studies directly examining whole-blood cannabinoid pharmacokinetics, although this matrix is often the only available specimen. METHODS Participants (9 men, 1 woman) resided on a closed research unit and smoked one 6.8% THC cannabis cigarette ad libitum. We quantified THC, 11-hydroxy-THC (11-OH-THC), 11-nor-9-carboxy-THC (THCCOOH), cannabidiol (CBD), cannabinol (CBN), THC-glucuronide and THCCOOH-glucuronide directly in whole blood and plasma by liquid chromatography/ tandem mass spectrometry within 24 h of collection to obviate stability issues. RESULTS Median whole blood (plasma) observed maximum concentrations (Cmax) were 50 (76), 6.4 (10), 41 (67), 1.3 (2.0), 2.4 (3.6), 89 (190), and 0.7 (1.4) μg/L 0.25 h after starting smoking for THC, 11-OH-THC, THCCOOH, CBD, CBN, and THCCOOH-glucuronide, respectively, and 0.5 h for THC-glucuronide. At observed Cmax, whole-blood (plasma) detection rates were 60% (80%), 80% (90%), and 50% (80%) for CBD, CBN, and THC-glucuronide, respectively. CBD and CBN were not detectable after 1 h in either matrix (LOQ 1.0 μg/L). CONCLUSIONS Human whole-blood cannabinoid data following cannabis smoking will assist whole blood and plasma cannabinoid interpretation, while furthering identification of recent cannabis intake. PMID:21836075

  9. Simultaneous determination of THC-COOH and THC-COOH-glucuronide in urine samples by LC/MS/MS.

    PubMed

    Weinmann, W; Vogt, S; Goerke, R; Müller, C; Bromberger, A

    2000-09-11

    A fast method using liquid-liquid extraction and HPLC/tandem-mass spectrometry (LC/MS/MS) was developed for the simultaneous detection of 11-Nor-Delta(9)-tetrahydrocannabinol-9-carboxylic acid beta-glucuronide (THC-COOH-glucuronide) and 11-Nor-Delta(9)-tetrahydrocannabinol-9-carboxylic acid (THC-COOH) in urine samples. This highly specific method, which combines chromatographic separation and MS/MS analysis, can be used for the confirmation of positive immunoassay results even without hydrolysis of the sample or derivatisation of extracts. Liquid-liquid extraction was optimised: with ethylacetate/diethylether (1:1, v/v) THC-COOH-glucuronide and THC-COOH could be extracted in one step. Molecular ions of the glucuronide (MH(+), m/z 521) and THC-COOH (MH(+), m/z 345) were generated using a PE/SCIEX turboionspray source in positive ionisation mode; specific fragmentation was performed in the collision cell of an API 365 triple-quadrupole mass spectrometer and yielded major fragments at m/z 345 (for THC-COOH-glucuronide) and m/z 327 as well as m/z 299 for both cannabinoids. Chromatographic separation was performed using a reversed-phase C8 column and gradient elution with 0.1% formic acid/1 mM ammonium formate and acetonitrile/0.1% formic acid. Retention times were 22.2 min for the glucuronide and 26.8 min for THC-COOH. After enzymatic hydrolysis of urine samples with beta-glucuronidase/arylsulfatase (37 degrees C, 5 h), THC-COOH-glucuronide was no longer detectable by LC/MS/MS in urine samples. However, the THC-COOH concentration was increased. For quantitation of THC-COOH, THC-COOH-D(3) was added to the urine samples as internal standard prior to analysis. From the difference of THC-COOH in the native urine and urine after enzymatic hydrolysis, molar concentration ratios of THC-COOH-glucuronide/THC-COOH in urine samples of cannabis users were determined and found to be between 1.3 and 4.5.

  10. Identification of Human UDP-Glucuronosyltransferase 1A4 as the Major Isozyme Responsible for the Glucuronidation of 20(S)-Protopanaxadiol in Human Liver Microsomes

    PubMed Central

    Li, Jia; He, Chunyong; Fang, Lianxiang; Yang, Li; Wang, Zhengtao

    2016-01-01

    20(S)-protopanaxadiol (PPD), one of the representative aglycones of ginsenosides, has a broad spectrum of pharmacological activities. Although phase I metabolism has been investigated extensively, information regarding phase II metabolism of this compound remains to be elucidated. Here, a glucuronidated metabolite of PPD in human liver microsomes (HLMs) and rat liver microsomes (RLMs) was unambiguously identified as PPD-3-O-β-d-glucuronide by nuclear magnetic resonance spectroscopy and high resolution mass spectrometry. The chemical inhibition and recombinant human UDP-Glucuronosyltransferase (UGT) isoforms assay showed that the PPD glucuronidation was mainly catalyzed by UGT1A4 in HLM, whereas UGT1A3 showed weak catalytic activity. In conclusion, PPD-3-O-β-d-glucuronide was first identified as the principal glucuronidation metabolite of PPD in HLMs, which was catalyzed by UGT1A4. PMID:27005621

  11. Identification of a new metabolite of GHB: gamma-hydroxybutyric acid glucuronide.

    PubMed

    Petersen, Ida Nymann; Tortzen, Christian; Kristensen, Jesper Langgaard; Pedersen, Daniel Sejer; Breindahl, Torben

    2013-06-01

    Gamma-hydroxybutyric acid (GHB) is an important analyte in clinical and forensic toxicology with a narrow detection window of 3-6 h. In the search of improved detection methods, the existence in vivo of a glucuronated GHB metabolite (GHB-GLUC) was hypothesized. Chemically pure standards of GHB-GLUC and a deuterated analogue for chromatography were synthesized. Liquid chromatography and tandem mass spectrometry were used for targeted analysis in anonymous clinical urine samples (n = 50). GHB-GLUC was found in concentrations ranging from 0.11 to 5.0 µg/mL (mean: 1.3 ± 1.2 µg/mL). Thus far, this is the first report of a GHB glucuronide detected in biological samples. Given that glucuronides generally have longer half-life values than their corresponding free drugs, GHB-GLUC should theoretically be a biomarker of GHB intoxication. It is also proposed that the hitherto unexplained reports of elevated GHB concentrations in some biological samples, which has caused the setting of a relatively high cutoff value (10 µg/mL), represent total GHB measurements (sum of free GHB and actively chemically hydrolyzed GHB-GLUC). To address these challenges, the present study must be followed by comprehensive pharmacokinetic and stability studies after the controlled administration of GHB.

  12. Isolation and determination of benzo(a)pyrene glucuronide and sulfate conjugates in soybean leaves

    SciTech Connect

    Negishi, T.; Nakano, M.; Kobayashi, S.; Kim, C.H.

    1987-08-01

    BaP is metabolized in mammalian systems by the mixed function oxidase system of liver microsomes. This system catalyzes the oxidation of BaP via epoxide intermediate to phenol, diol and quinone metabolites. One of these 7,8-dihydroxy-9,10-epoxy-7,8,9,10-tetrahydro-BaP is thought to act as the ultimate carcinogen by binding covalently to cellular DNA. It is also known that Cunninghamella elegans oxidized BaP to its phenol, diol and quinone metabolites. In addition, the alcohols were detected as glucuronide and sulfate conjugates. These metabolites are remarkably similar to those observed in higher organisms. On the other hand, some investigators have demonstrated that plants take up BaP and anthracene from soil or culture medium containing these compounds. This paper reports the finding that soybeans grown in BaP polluted soil take it up and metabolize to its phenol, diol and the glucuronide and sulfate conjugates of the alcohols.

  13. 99mTc-d-penicillamine-glucuronide: synthesis, radiolabeling, in vitro and in vivo evaluation.

    PubMed

    Teksöz, Serap; Içhedef, Ciğdem Acar; Ozyüncü, Seniha; Müftüler, Fazilet Zümrüt Biber; Unak, Perihan; Medine, Ilker Emin; Ertay, Türkan; Eren, Mine Şencan

    2011-10-01

    The current study was aimed at synthesizing a glucuronide derivative of D-penicillamine (D-PA) to be used for imaging purposes. First of all, D-PA-glucuronide (D-PA-Glu) was synthesized by experimental treatments starting with uridine 5'-diphospho-glucuronosyltransferase enzyme rich microsome preparate. Then, the synthesized compound was labeled with technetium ((99m)Tc) by using a reduction method with stannous chloride. Quality controls were performed by using high-performance liquid chromatography and thin-layer radio chromatography (TLRC). Radiolabeling yield of (99m)Tc-D-PA-Glu was more than 98% according to TLRC results. In vitro evaluations of radiolabeled complexes were investigated on PC-3 human prostate cancer cells. (99m)Tc-D-PA-Glu exhibited more accumulation on PC-3 cells versus (99m)Tc-D-PA at 240 minutes. In order to determine its radiopharmaceutical potential, biodistribution studies were carried out in male Albino Wistar rats. The biodistribution results of (99m)Tc-D-PA-Glu, showed the highest uptake in prostate at 120 minutes postinjection with the main excretion route being through kidneys and bladder. (99m)Tc-D-PA-Glu and (99m)Tc-D-PA have exhibited different biodistribution results.

  14. Identification of flunixin glucuronide and depletion of flunixin and its marker residue in bovine milk.

    PubMed

    Jedziniak, P; Olejnik, M; Szprengier-Juszkiewicz, T; Smulski, S; Kaczmarowski, M; Żmudzki, J

    2013-12-01

    Residues of flunixin [and its marker residue 5-hydroxyflunixin (5OHFLU)] were determined in milk from cows that intravenously received therapeutic doses of the drug. The samples were collected during each milking (every 12 h) for six consecutive days, and concentrations of flunixin and its metabolites were determined by the method with and without enzymatic hydrolysis (beta-glucuronidase). The highest flunixin concentration in milk was observed 12 h after dosing (2.4 ± 1.42 μg/kg, mean ± SD). Flunixin concentrations in the samples determined with enzymatic hydrolysis were significantly higher (P < 0.05), which suggests the transfer of flunixin glucuronide to the milk. Additionally, unambiguous identification of flunixin glucuronide in the bovine milk was performed with linear ion-trap mass spectrometry. The 5OHFLU concentrations analyzed without enzymatic hydrolysis (22.3 ± 16.04 μg/kg) were similar to this obtained with enzymatic hydrolysis. Flunixin and 5OHFLU concentrations dropped below the limits of detection at 48 h after last dosing.

  15. A newborn screening method for cerebrotendinous xanthomatosis using bile alcohol glucuronides and metabolite ratios.

    PubMed

    Vaz, Frédéric M; Bootsma, Albert H; Kulik, Willem; Verrips, Aad; Wevers, Ron A; Schielen, Peter C; DeBarber, Andrea E; Huidekoper, Hidde H

    2017-03-17

    Cerebrotendinous xanthomatosis (CTX) is a treatable neurodegenerative metabolic disorder of bile acid synthesis where symptoms can be prevented if treatment with chenodeoxycholic acid supplementation is initiated early in life, making CTX an excellent candidate for newborn screening. We developed a new dried blood spot screening assay for this disorder based on different ratios between the accumulating cholestanetetrol glucuronide (tetrol) and specific bile acids/bile acid intermediates, without the need for derivatization. A quarter-inch dried blood spot punch was extracted with methanol, internal standards were added and after concentration the extract was injected into the tandem mass spectrometer using a 2 minute flow injection analysis where specific transitions were measured for cholestanetetrol glucuronide, tauro-chenodeoxycholic acid (t-CDCA) and tauro-trihydroxycholestanoic acid (t-THCA). A proof of principle experiment was performed using 216 Guthrie cards from healthy term/preterm newborns, CTX patients and Zellweger patients. Using two calculated biomarkers, tetrol/t-CDCA and t-THCA/tetrol, this straightforward method achieved an excellent separation between dried blood spots of CTX patients and those of controls, Zellweger patients and newborns with cholestasis. The results of this small pilot study indicate that the tetrol/t-CDCA ratio is an excellent derived biomarker for CTX that has the potential to be used in neonatal screening programs.

  16. Synthesis and Antitumor Properties of BQC-Glucuronide, a Camptothecin Prodrug for Selective Tumor Activation.

    PubMed

    Prijovich, Zeljko M; Burnouf, Pierre-Alain; Chou, Hua-Cheng; Huang, Ping-Ting; Chen, Kai-Chuan; Cheng, Tian-Lu; Leu, Yu-Lin; Roffler, Steve R

    2016-04-04

    Major limitations of camptothecin anticancer drugs (toxicity, nonselectivity, water insolubility, inactivation by human serum albumin) may be improved by creating glucuronide prodrugs that rely on beta-glucuronidase for their activation. We found that the camptothecin derivative 5,6-dihydro-4H-benzo[de]quinoline-camptothecin (BQC) displays greater cytotoxicity against cancer cells than the clinically used camptothecin derivatives SN-38 and topotecan even in the presence of human serum albumin. We synthesized the prodrug BQC-glucuronide (BQC-G), which was 4000 times more water soluble and 20-40 times less cytotoxic than BQC. Importantly, even in the presence of human serum albumin, BQC-G was efficiently hydrolyzed by beta-glucuronidase and produced greater cytotoxicity (IC50 = 13 nM) than camptothecin, 9-aminocamptothecin, SN-38, or topotecan (IC50 > 3000, 1370, 48, and 28 nM, respectively). BQC-G treatment of mice bearing human colon cancer xenografts with naturally or artificially elevated beta-glucuronidase activity produced significant antitumor activity, showing that BQC-G is a potent prodrug suitable for selective intratumoral drug activation.

  17. 40 CFR 721.10244 - Phosphonic acid, P-[2-[bis(2-hydroxyethyl)amino]ethyl]-, 2-[bis(2- chloroethoxy)phosphinyl]ethyl...

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 32 2012-07-01 2012-07-01 false Phosphonic acid, P- ethyl]-, 2- ethyl... New Uses for Specific Chemical Substances § 721.10244 Phosphonic acid, P- ethyl]-, 2- ethyl 2... substance identified as phosphonic acid, P- ethyl]-, 2- ethyl 2-chloroethyl ester (PMN P-09-195; CAS...

  18. 40 CFR 721.10244 - Phosphonic acid, P-[2-[bis(2-hydroxyethyl)amino]ethyl]-, 2-[bis(2- chloroethoxy)phosphinyl]ethyl...

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 31 2014-07-01 2014-07-01 false Phosphonic acid, P- ethyl]-, 2- ethyl... New Uses for Specific Chemical Substances § 721.10244 Phosphonic acid, P- ethyl]-, 2- ethyl 2... substance identified as phosphonic acid, P- ethyl]-, 2- ethyl 2-chloroethyl ester (PMN P-09-195; CAS...

  19. 40 CFR 721.10244 - Phosphonic acid, P-[2-[bis(2-hydroxyethyl)amino]ethyl]-, 2-[bis(2- chloroethoxy)phosphinyl]ethyl...

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 32 2013-07-01 2013-07-01 false Phosphonic acid, P- ethyl]-, 2- ethyl... New Uses for Specific Chemical Substances § 721.10244 Phosphonic acid, P- ethyl]-, 2- ethyl 2... substance identified as phosphonic acid, P- ethyl]-, 2- ethyl 2-chloroethyl ester (PMN P-09-195; CAS...

  20. Efflux Transport Characterization of Resveratrol Glucuronides in UDP-Glucuronosyltransferase 1A1 Transfected HeLa Cells: Application of a Cellular Pharmacokinetic Model to Decipher the Contribution of Multidrug Resistance-Associated Protein 4.

    PubMed

    Wang, Shuai; Li, Feng; Quan, Enxi; Dong, Dong; Wu, Baojian

    2016-04-01

    Resveratrol undergoes extensive metabolism to form biologically active glucuronides in humans. However, the transport mechanisms for resveratrol glucuronides are not fully established. Here, we aimed to characterize the efflux transport of resveratrol glucuronides using UGT1A1-overexpressing HeLa cells (HeLa1A1 cells), and to determine the contribution of multidrug resistance-associated protein (MRP) 4 to cellular excretion of the glucuronides. Two glucuronide isomers [i.e., resveratrol 3-O-glucuronide (R3G) and resveratrol 4'-O-glucuronide (R4'G)] were excreted into the extracellular compartment after incubation of resveratrol (1-100 μM) with HeLa1A1 cells. The excretion rate was linearly related to the level of intracellular glucuronide, indicating that glucuronide efflux was a nonsaturable process. MK-571 (a dual inhibitor of UGT1A1 and MRPs) significantly decreased the excretion rates of R3G and R4'G while increasing their intracellular levels. Likewise, short-hairpin RNA (shRNA)-mediated silencing of MRP4 caused a significant reduction in glucuronide excretion but an elevation in glucuronide accumulation. Furthermore, β-glucuronidase expressed in the cells catalyzed the hydrolysis of the glucuronides back to the parent compound. A cellular pharmacokinetic model integrating resveratrol transport/metabolism with glucuronide hydrolysis/excretion was well fitted to the experimental data, allowing derivation of the efflux rate constant values in the absence or presence of shRNA targeting MRP4. It was found that a large percentage of glucuronide excretion (43%-46%) was attributed to MRP4. In conclusion, MRP4 participated in cellular excretion of R3G and R4'G. Integration of mechanistic pharmacokinetic modeling with transporter knockdown was a useful method to derive the contribution percentage of an exporter to overall glucuronide excretion.

  1. High-Throughput LC-MS/MS Method for Direct Quantification of Glucuronidated, Sulfated, and Free Enterolactone in Human Plasma.

    PubMed

    Nørskov, Natalja P; Kyrø, Cecilie; Olsen, Anja; Tjønneland, Anne; Knudsen, Knud Erik Bach

    2016-03-04

    Sulfation and glucuronidation constitute a major pathway in humans and may play an important role in biological activity of metabolites including the enterolignan, enterolactone. Because the aromatic structure of enterolactone has similarities to steroid metabolites, it was hypothesized that enterolactone may protect against hormone-dependent cancers. This led to numerous epidemiological studies. In this context, there has been a demand for rapid, sensitive, high-throughput methods to measure enterolactone in biofluids. Different methods have been developed using GC-MS, HPLC, LC-MS/MS and a fluoroimmunoassay; however, most of these methods measure the total concentration of enterolactone, without any specification of its conjugation pattern. Here for the first time we present a high-throughput LC-MS/MS method to quantify enterolactone in its intact form as glucuronide, sulfate, and free enterolactone. The method has shown good accuracy and precision at low concentration and very high sensitivity, with LLOQ for enterolactone sulfate at 16 pM, enterolactone glucuronide at 26 pM, and free enterolactone at 86 pM. The short run time of 2.6 min combined with simple sample clean up and high sensitivity make this method attractive for the high-throughput of samples needed for epidemiological studies. Finally, we have adapted the new method to quantify enterolactone and its conjugates in 3956 plasma samples from an epidemiological study. We found enterolactone glucuronide to be the major conjugation form and that conjugation pattern was similar between men and women.

  2. N(+)-glucuronidation of aliphatic tertiary amines, a general phenomenon in the metabolism of H1-antihistamines in humans.

    PubMed

    Luo, H; Hawes, E M; McKay, G; Korchinski, E D; Midha, K K

    1991-10-01

    1. Representative drugs of the various structural classes of H1 antihistamines were chosen for study. The drugs chosen (class name in parentheses) were chlorpheniramine maleate and pheniramine maleate (alkylamines), diphenhydramine hydrochloride and doxylamine succinate (ethanolamines), pyrilamine maleate and tripelennamine hydrochloride (ethylenediamines), promethazine hydrochloride (phenothiazine), cyclizine lactate (piperazine) and terfenadine (miscellaneous). In each case oral dose(s) were administered over no more than 6 h to two healthy volunteers and the total urine collected for 36 h. 2. Metabolites from urine were separated by h.p.l.c. and individually collected prior to mass spectrometric analysis in the fast atom bombardment mode. The structure of each metabolite identified as a quaternary ammonium-linked glucuronide metabolite was confirmed by direct comparison of its mass spectrum and chromatographic behaviour with that of a synthetic authentic compound. 3. For eight of the nine drugs studied, metabolism by the N(+)-glucuronidation pathway was observed in each of the volunteers. Terfenadine was the exception. 4. The amount of each N(+)-glucuronide in the urine was estimated by h.p.l.c. analysis. The mean proportion of dose excreted as the metabolite was 14.3%, 6.5% and 4.0% for cyclizine, tripelennamine and diphenhydramine, respectively. Promethazine was the only case where the N(+)-glucuronide accounted for less than 1.0% of the administered dose in both volunteers examined.

  3. Human and Rat ABC Transporter Efflux of Bisphenol A and Bisphenol A Glucuronide: Interspecies Comparison and Implications for Pharmacokinetic Assessment

    EPA Science Inventory

    Significant interspecies differences exist between human and rodent with respect to absorption, distribution, and excretion of bisphenol A (BPA) and its primary metabolite, BPA-glucuronide (BPA-G). ATP-Binding Cassette (ABC) transporter enzymes play important roles in these physi...

  4. 21 CFR 173.228 - Ethyl acetate.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 3 2014-04-01 2014-04-01 false Ethyl acetate. 173.228 Section 173.228 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) SECONDARY DIRECT FOOD ADDITIVES PERMITTED IN FOOD FOR HUMAN CONSUMPTION Solvents, Lubricants, Release Agents and...

  5. Ethyl p-nitrophenyl phenylphosphorothioate (EPN)

    Integrated Risk Information System (IRIS)

    Ethyl p - nitrophenyl phenylphosphorothioate ( EPN ) ; CASRN 2104 - 64 - 5 Human health assessment information on a chemical substance is included in the IRIS database only after a comprehensive review of toxicity data , as outlined in the IRIS assessment development process . Sections I ( Health Ha

  6. Manufacturing Ethyl Acetate From Fermentation Ethanol

    NASA Technical Reports Server (NTRS)

    Rohatgi, Naresh K.; Ingham, John D.

    1991-01-01

    Conceptual process uses dilute product of fermentation instead of concentrated ethanol. Low-concentration ethanol, extracted by vacuum from fermentation tank, and acetic acid constitutes feedstock for catalytic reaction. Product of reaction goes through steps that increases ethyl acetate content to 93 percent by weight. To conserve energy, heat exchangers recycle waste heat to preheat process streams at various points.

  7. Raloxifene glucuronidation in liver and intestinal microsomes of humans and monkeys: contribution of UGT1A1, UGT1A8 and UGT1A9.

    PubMed

    Kishi, Naoki; Takasuka, Akane; Kokawa, Yuki; Isobe, Takashi; Taguchi, Maho; Shigeyama, Masato; Murata, Mikio; Suno, Manabu; Hanioka, Nobumitsu

    2016-01-01

    1. Raloxifene is an antiestrogen that has been marketed for the treatment of osteoporosis, and is metabolized into 6- and 4'-glucuronides by UDP-glucuronosyltransferase (UGT) enzymes. In this study, the in vitro glucuronidation of raloxifene in humans and monkeys was examined using liver and intestinal microsomes and recombinant UGT enzymes (UGT1A1, UGT1A8 and UGT1A9). 2. Although the K(m) and CL(int) values for the 6-glucuronidation of liver and intestinal microsomes were similar between humans and monkeys, and species differences in Vmax values (liver microsomes, humans > monkeys; intestinal microsomes, humans < monkeys) were observed, no significant differences were noted in the K(m) or S50, Vmax and CL(int) or CLmax values for the 4'-glucuronidation of liver and intestinal microsomes between humans and monkeys. 3. The activities of 6-glucuronidation in recombinant UGT enzymes were UGT1A1 > UGT1A8 >UGT1A9 for humans, and UGT1A8 > UGT1A1 > UGT1A9 for monkeys. The activities of 4'-glucuronidation were UGT1A8 > UGT1A1 > UGT1A9 in humans and monkeys. 4. These results demonstrated that the profiles for the hepatic and intestinal glucuronidation of raloxifene by microsomes were moderately different between humans and monkeys.

  8. β-Lactam analogues of combretastatin A-4 prevent metabolic inactivation by glucuronidation in chemoresistant HT-29 colon cancer cells.

    PubMed

    Malebari, Azizah M; Greene, Lisa M; Nathwani, Seema M; Fayne, Darren; O'Boyle, Niamh M; Wang, Shu; Twamley, Brendan; Zisterer, Daniela M; Meegan, Mary J

    2017-04-21

    Glucuronidation by uridine 5-diphosphoglucuronosyl transferase enzymes (UGTs) is a cause of intrinsic drug resistance in cancer cells. Glucuronidation of combretastatin A-4 (CA-4) was previously identified as a mechanism of resistance in hepatocellular cancer cells. Herein, we propose chemical manipulation of β-lactam bridged analogues of Combretastatin A-4 as a novel means of overcoming drug resistance associated with glucuronidation due to the expression of UGTs in the CA-4 resistant human colon cancer HT-29 cells. The alkene bridge of CA-4 is replaced with a β-lactam ring to circumvent potential isomerisation while the potential sites of glucuronate conjugation are deleted in the novel 3-substituted-1,4-diaryl-2-azetidinone analogues of CA-4. We hypothesise that glucuronidation of CA-4 is the mechanism of drug resistance in HT-29 cells. Ring B thioether containing 2-azetidinone analogues of CA-4 such as 4-(4-(methylthio)phenyl)-3-phenyl-1-(3,4,5-trimethoxyphenyl)azetidin-2-one (27) and 3-hydroxy-4-(4-(methylthio)phenyl)-1-(3,4,5-trimethoxyphenyl)azetidin-2-one (45) were identified as the most potent inhibitors of tumour cell growth, independent of UGT status, displaying antiproliferative activity in the low nanomolar range. These compounds also disrupted the microtubular structure in MCF-7 and HT-29 cells, and caused G2/M arrest and apoptosis. Taken together, these findings highlight the potential of chemical manipulation as a means of overcoming glucuronidation attributed drug resistance in CA-4 resistant human colon cancer HT-29 cells, allowing the development of therapeutically superior analogues.

  9. Analysis of R- and S-Hydroxywarfarin Glucuronidation Catalyzed by Human Liver Microsomes and Recombinant UDP-Glucuronosyltransferases

    PubMed Central

    Bratton, Stacie M.; Mosher, Carrie M.; Khallouki, Farid; Finel, Moshe; Court, Michael H.; Moran, Jeffery H.

    2012-01-01

    Coumadin (R-, S-warfarin) is a challenging drug to accurately dose, both initially and for maintenance, because of its narrow therapeutic range and wide interpatient variability and is typically administered as a racemic (Rac) mixture, which complicates the biotransformation pathways. The goal of the current work was to identify the human UDP-glucuronosyltransferases (UGTs) involved in the glucuronidation of the separated R- and S-enantiomers of 6-, 7-, and 8-hydroxywarfarin and the possible interactions between these enantiomers. The kinetic and inhibition constants for human recombinant 1A family UGTs toward these separated enantiomers have been assessed using high-performance liquid chromatography (HPLC)-UV-visible analysis, and product confirmations have been made using HPLC-mass spectrometry/mass spectrometry. We found that separated R- and S-enantiomers of 6-, 7-, and 8-hydroxywarfarin demonstrate significantly different glucuronidation kinetics and can be mutually inhibitory. In some cases significant substrate inhibition was observed, as shown by Km, Vmax, and Ki, comparisons. In particular, UGT1A1 and extrahepatic UGT1A10 have significantly higher capacities than other isoforms for S-7-hydroxywarfarin and R-7-hydroxywarfarin glucuronidation, respectively. Activity data generated using a set of well characterized human liver microsomes supported the recombinant enzyme data, suggesting an important (although not exclusive) role for UGT1A1 in glucuronidation of the main warfarin metabolites, including Rac-6- and 7-hydroxywarfarin and their R- and S-enantiomers in the liver. This is the first demonstration that the R- and S-enantiomers of hydroxywarfarins are glucuronidated, with significantly different enzymatic affinity and capacity, and supports the importance of UGT1A1 as the major hepatic isoform involved. PMID:21972237

  10. Bioavailability of the glucuronide and sulfate conjugates of genistein and daidzein in breast cancer resistance protein 1 knockout mice.

    PubMed

    Álvarez, Ana I; Vallejo, Fernando; Barrera, Borja; Merino, Gracia; Prieto, Julio G; Tomás-Barberán, Francisco; Espín, Juan C

    2011-11-01

    The dietary polyphenols genistein and daidzein are potent effectors of biological processes. The plasma profile of both isoflavones is governed by the presence of phase II conjugates, mainly glucuronides and sulfates. Breast cancer resistance protein (ABCG2/BCRP) interacts with genistein and daidzein, which are among the natural substrates of the transporter and competitively inhibit ABCG2-mediated drug efflux. ABCG2/BCRP can also transport glucuronide and sulfate conjugates. In this study, we analyzed the plasma levels of aglycones and derived conjugated metabolites, glucuronides, and sulfates, after intragastric administration of these isoflavones to wild-type and Bcrp1(-/-) knockout mice. The results show that overall plasmatic profile is mainly governed by sulfate and glucuronide derivatives, the concentration of which was significantly increased (7- to 10-fold) in Bcrp1(-/-) mice. The total AUC h nM (0-180 min), as the sum of aglycones, glucuronides, and sulfates, was 901 ± 207 in wild-type mice versus 4988 ± 508 in Bcrp1(-/-) mice after genistein administration (50 mg/kg b.wt.); 584.3 ± 90 in wild-type mice versus 4012 ± 612 in Bcrp1(-/-) after daidzein administration (50 mg/kg); and 926 ± 140 in wild-type mice versus 5174 ± 696 in Bcrp1(-/-) after genistein+daidzein administration (25 + 25 mg/kg). Therefore, our results indicate a direct and conclusive Bcrp1 efflux action on phase II metabolites of these isoflavones in vivo and suggest a possible novel concept for ABCG2/BCRP as part of metabolism-driven efflux transport of these conjugates.

  11. A novel method for the quantitation of gingerol glucuronides in human plasma or urine based on stable isotope dilution assays.

    PubMed

    Schoenknecht, Carola; Andersen, Gaby; Schieberle, Peter

    2016-11-15

    The bio-active compounds of ginger (Zingiber officinale Roscoe), the gingerols, are gaining considerable attention due to their numerous beneficial health effects. In order to elucidate the physiological relevance of the ascribed effects their bioavailability has to be determined taking their metabolization into account. To quantitate in vivo generated [6]-, [8]- and [10]-gingerol glucuronides in human plasma and urine after ginger tea consumption, a simultaneous and direct liquid chromatography-tandem mass spectrometry method based on stable isotope dilution assays was established and validated. The respective references as well as the isotopically labeled substances were synthesized and characterized by mass spectrometry and NMR. Selective isolation of gingerol glucuronides from human plasma and urine by a mixed-phase anion-exchange SPE method led to recovery rates between 80.8 and 98.2%. LC-MS/MS analyses in selected reaction monitoring modus enabled a highly sensitive quantitation of gingerol glucuronides with LoQs between 3.9-9.8nmol/L in plasma and 39.3-161.1nmol/L in urine. The method precision in plasma and urine varied in the range±15%, whereas the intra-day accuracy in plasma and urine showed values between 78 and 122%. The developed method was then applied to a pilot study in which two volunteers consumed one liter ginger tea. Pharmacokinetic parameters like the maximum concentration (cmax), the time to reach cmax (tmax), area under the curve (AUC), elimination rate constant (kel) and elimination half-life (t1/2) were calculated from the concentration-time curve of each gingerol glucuronide. The obtained results will enable more detailed investigation of gingerol glucuronides as bioactives in their physiologically relevant concentrations.

  12. 21 CFR 172.872 - Methyl ethyl cellulose.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 3 2014-04-01 2014-04-01 false Methyl ethyl cellulose. 172.872 Section 172.872... Methyl ethyl cellulose. The food additive methyl ethyl cellulose may be safely used in food in accordance with the following prescribed conditions. (a) The additive is a cellulose ether having the...

  13. 40 CFR 180.441 - Quizalofop ethyl; tolerances for residues.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... residues of the herbicide quizalofop ethyl, including its metabolites and degradates, in or on the....05 (2) Tolerances are established for residues of the herbicide quizalofop ethyl, including its... with regional registration are established for residues of the herbicide quizalofop ethyl,...

  14. 40 CFR 180.441 - Quizalofop ethyl; tolerances for residues.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... combined residues of the herbicide quizalofop (2- propanoic acid) and quizalofop ethyl (ethyl-2- propanoate...) Tolerances are established for the combined residues of the herbicide quizalofop (2- propanoic acid... herbicide quizalofop-p ethyl ester , and its acid metabolite quizalofop-p , and the S enantiomers of...

  15. 40 CFR 180.441 - Quizalofop ethyl; tolerances for residues.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... residues of the herbicide quizalofop ethyl, including its metabolites and degradates, in or on the....05 (2) Tolerances are established for residues of the herbicide quizalofop ethyl, including its... with regional registration are established for residues of the herbicide quizalofop ethyl,...

  16. 40 CFR 180.441 - Quizalofop ethyl; tolerances for residues.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... residues of the herbicide quizalofop ethyl, including its metabolites and degradates, in or on the....05 (2) Tolerances are established for residues of the herbicide quizalofop ethyl, including its... with regional registration are established for residues of the herbicide quizalofop ethyl,...

  17. 21 CFR 177.1320 - Ethylene-ethyl acrylate copolymers.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 3 2011-04-01 2011-04-01 false Ethylene-ethyl acrylate copolymers. 177.1320... Basic Components of Single and Repeated Use Food Contact Surfaces § 177.1320 Ethylene-ethyl acrylate copolymers. Ethylene-ethyl acrylate copolymers may be safely used to produce packaging materials,...

  18. 21 CFR 177.1320 - Ethylene-ethyl acrylate copolymers.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 3 2012-04-01 2012-04-01 false Ethylene-ethyl acrylate copolymers. 177.1320... Basic Components of Single and Repeated Use Food Contact Surfaces § 177.1320 Ethylene-ethyl acrylate copolymers. Ethylene-ethyl acrylate copolymers may be safely used to produce packaging materials,...

  19. 21 CFR 177.1320 - Ethylene-ethyl acrylate copolymers.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 3 2013-04-01 2013-04-01 false Ethylene-ethyl acrylate copolymers. 177.1320... Basic Components of Single and Repeated Use Food Contact Surfaces § 177.1320 Ethylene-ethyl acrylate copolymers. Ethylene-ethyl acrylate copolymers may be safely used to produce packaging materials,...

  20. Synthesis of a precursor for the preparation of 9 alpha,11 alpha-tritiated 5 alpha-androstane-3 alpha,17 beta-diol 17-glucuronide

    SciTech Connect

    Rao, P.N.; Damodaran, K.M.

    1984-03-01

    Starting from 11 beta-hydroxytestosterone, the synthesis of a strategic precursor, C-9 (11) unsaturated 5 alpha-androstane-3 alpha, 17 beta-diol 17-glucuronide (9a), for the preparation of 9 alpha,11 alpha-tritiated 5 alpha-androstane-3 alpha, 17 beta-diol 17-glucuronide has been achieved. The authors optimized the reaction conditions for catalytic reduction employing hydrogen and subsequent base hydrolysis followed by purification on Amberlite XAD-2 resin to obtain the saturated 5 alpha-androstane-3 alpha, 17 beta-diol 17-glucuronide.

  1. Puerarin-7-O-glucuronide, a water-soluble puerarin metabolite, prevents angiotensin II-induced cardiomyocyte hypertrophy by reducing oxidative stress.

    PubMed

    Hou, Ning; Cai, Bin; Ou, Cai-Wen; Zhang, Zhen-Hui; Liu, Xia-Wen; Yuan, Mu; Zhao, Gan-Jian; Liu, Shi-Ming; Xiong, Long-Gen; Luo, Jian-Dong; Luo, Cheng-Feng; Chen, Min-Sheng

    2017-02-16

    This study aimed to investigate the anti-oxidant and anti-hypertrophic effects of puerarin-7-O-glucuronide, a water-soluble puerarin metabolite. The anti-oxidant effects of puerarin-7-O-glucuronide were assessed by measurement of intracellular superoxide levels, total superoxide dismutase (SOD) activity, total anti-oxidant capacity, and glutathione (GSH)/glutathione disulfide (GSSG) ratio in cultured neonatal rat cardiomyocytes (NRCMs) stimulated with the xanthine oxidase (XO)/xanthine (X) system or angiotensin II. The activity of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase and expression of NADPH oxidase subunits p22(phox) and p47(phox) were determined. The anti-hypertrophic effects of puerarin-7-O-glucuronide in angiotensin II-challenged NRCMs were characterized by changes in cell morphology and expression of hypertrophic genes. In the pharmacokinetic study, the plasma concentration of puerarin-7-O-glucuronide was determined by rapid resolution-liquid chromatography-tandem mass spectrometry (RR-LC-MS/MS). Puerarin-7-O-glucuronide prevented XO/X-induced increase in intracellular superoxide production and decreases in total SOD activity, GSH/GSSG ratio, and total anti-oxidant capacity. Puerarin-7-O-glucuronide also reversed angiotensin II-induced increases in intracellular superoxide production and NADPH oxidase activity and decreases in total SOD activity. These anti-oxidant effects of puerarin-7-O-glucuronide were accompanied by downregulation of p22(phox) and p47(phox). Furthermore, puerarin-7-O-glucuronide prevented angiotensin II-induced increases in cell surface area and perimeter, as well as changes in Nppa, Myh7, and Myh6. In the pharmacokinetic study, puerarin-7-O-glucuronide was cleared with a half-life of 0.94 h after intravenous administration. Puerarin could be detected in rat plasma, albeit in low concentration, as early as 5 min after intravenous administration of puerarin-7-O-glucuronide. These anti-oxidant and anti

  2. Identifying and applying a highly selective probe to simultaneously determine the O-glucuronidation activity of human UGT1A3 and UGT1A4

    PubMed Central

    Jiang, Li; Liang, Si-Cheng; Wang, Chao; Ge, Guang-Bo; Huo, Xiao-Kui; Qi, Xiao-Yi; Deng, Sa; Liu, Ke-Xin; Ma, Xiao-Chi

    2015-01-01

    Glucuronidation mediated by uridine 5′-diphospho (UDP)-glucuronosyltransferase is an important detoxification pathway. However, identifying a selective probe of UDP- glucuronosyltransferase is complicated because of the significant overlapping substrate specificity displayed by the enzyme. In this paper, desacetylcinobufagin (DACB) 3-O- and 16-O-glucuronidation were found to be isoform-specific probe reactions for UGT1A4 and UGT1A3, respectively. DACB was well characterized as a probe for simultaneously determining the catalytic activities of O-glucuronidation mediated by UGT1A3 and UGT1A4 from various enzyme sources, through a sensitive analysis method. PMID:25884245

  3. Skin of the male African catfish, Clarias gariepinus: a source of steroid glucuronides

    SciTech Connect

    Ali, S.A.; Schoonen, W.G.; Lambert, J.G.; Van den Hurk, R.; Van Oordt, P.G.

    1987-06-01

    Steroid metabolism in the skin of mature male African catfish, Clarias gariepinus, reared in the laboratory, was studied in vitro by tissue incubations with (/sup 3/H)pregnenolone, (/sup 3/H)dehydroepiandrosterone, (/sup 3/H)17 alpha-hydroxyprogesterone, (/sup 3/H)androstenedione, (/sup 14/C)11 beta-hydroxyandrostenedione, and (/sup 3/H)testosterone as precursors. While pregnenolone was not converted to any other steroid, dehydroepiandrosterone was transformed mainly to 5-androstene-3 beta, 17 beta-diol. The products of 17 alpha-hydroxyprogesterone incubations were 5 beta-pregnane-3 alpha,17 alpha-diol-20-one, 5 beta-pregnane-3 alpha,17 alpha, 20 beta-triol, and 5 beta-pregnan-17 alpha-o1-3,20-dione. The major steroids of androstenedione incubations were etiocholanolone, testosterone, and androsterone. Testosterone was converted mainly to etiocholanolone and androstenedione, and only small quantities of 11 beta-hydroxytestosterone, 11-ketotestosterone, and 11-ketoandrostenedione were the metabolites found in 11 beta-hydroxyandrostenedione incubation. These results demonstrated the presence of the enzymes 5 alpha- and 5 beta-reductases and 3 alpha-, 11 beta-, 17 beta-, and 20 beta-hydroxysteroid dehydrogenases in the skin. From enzymehistochemical results it appeared that the steroid conversions take place in the epithelial cells. Moreover, the presence of UDP-glucose dehydrogenase, an enzyme involved in the synthesis of glucuronic acid, in these cells indicates the possibility of steroid glucuronide formation. Indeed significant amounts of water-soluble steroid conjugates, particularly 5 beta-dihydrotestosterone- and testosterone-glucuronide, were found in the incubations with androstenedione and testosterone, indicating the presence of the UDP-glucuronosyl transferase in the catfish skin.

  4. Simultaneous modelling of the Michaelis-Menten kinetics of paracetamol sulphation and glucuronidation.

    PubMed

    Reith, David; Medlicott, Natalie J; Kumara De Silva, Rohana; Yang, Lin; Hickling, Jeremy; Zacharias, Mathew

    2009-01-01

    1. The aim of the present study was to perform an in vivo estimation of the Michaelis-Menten constants of the major metabolic pathways of paracetamol (APAP). 2. A two-occasion, single-dose cross-over trial was performed using 60 and 90 mg/kg doses of APAP in healthy patients undergoing third molar dental extraction. Plasma samples were collected over 24 h and urine was collected for 8 h after dosing. Twenty patients were enrolled in the study and complete data for plasma and urine were available for both doses for 13 volunteers who were included in the analysis; seven of the volunteers were men, the median age (range) was 22 years (19-31) and the median weight (range) was 68 kg (50-86). 3. The mean (95% CI) k(m) for APAP glucuronidation was 6.89 mmol/L (3.57-10.22) and the V(max) was 0.97 mmol/h per kg (0.65-1.28). The k(m) for APAP sulphation was 0.097 mmol/L (0.041-0.152) and the V(max) was 0.011 mmol/h per kg (0.009-0.013). For the combined excretion of APAP-cysteine and APAP-mercapturate, the k(m) was 0.303 mmol/L (0.131-0.475) and the V(max) was 0.004 mmol/h per kg (0.002-0.005). 4. The estimates for in vivo Michaelis-Menten constants for APAP glucuronidation and sulphation were in the order of those reported previously using in vitro methods.

  5. Inhibitory effect of ciprofloxacin on β-glucuronidase-mediated deconjugation of mycophenolic acid glucuronide.

    PubMed

    Kodawara, Takaaki; Masuda, Satohiro; Yano, Yoshitaka; Matsubara, Kazuo; Nakamura, Toshiaki; Masada, Mikio

    2014-07-01

    The interaction between mycophenolate (MPA) and quinolone antibiotics such as ciprofloxacin is considered to reduce the enterohepatic recycling of MPA, which is biotransformed in the intestine from MPA glucuronide (MPAG) conjugate excreted via the biliary system; however, the molecular mechanism underlying this biotransformation of MPA is still unclear. In this study, an in vitro system was established to evaluate β-glucuronidase-mediated deconjugation and to examine the influence of ciprofloxacin on the enzymatic deconjugation of MPAG and MPA resynthesis. Resynthesis of MPA via deconjugation of MPAG increased in a time-dependent manner from 5 to 60 min in the presence of β-glucuronidase. Ciprofloxacin and phenolphthalein-β-d-glucuronide (PhePG), a typical β-glucuronidase substrate, significantly decreased the production of MPA from MPAG in the β-glucuronidase-mediated deconjugation system. In addition, enoxacin significantly inhibited the production of MPA from MPAG, while levofloxacin and ofloxacin had no inhibitory effect on MPA synthesis. Pharmacokinetic analysis revealed that ciprofloxacin showed a dose-dependent inhibitory effect on MPA production from MPAG via β-glucuronidase with a half-maximal inhibitory concentration (IC50 ) value of 30.4 µm. While PhePG inhibited the β-glucuronidase-mediated production of MPA from MPAG in a competitive manner, ciprofloxacin inhibited MPA synthesis via noncompetitive inhibition. These findings suggest that the reduction in the serum MPA concentration during the co-administration of ciprofloxacin is at least in part due to the decreased enterohepatic circulation of MPA because of noncompetitive inhibition of deconjugation of MPAG by intestinal β-glucuronidase.

  6. 40 CFR 180.595 - Flufenpyr-ethyl; tolerances for residues.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... residues of the herbicide, flufenpyr-ethyl; acetic acid, -phenoxy]-ethyl ester], in or on the following...) Tolerances are established for residues of the herbicide flufenpyr-ethyl; acetic acid, -phenoxy]-ethyl...

  7. 40 CFR 180.595 - Flufenpyr-ethyl; tolerances for residues.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... residues of the herbicide, flufenpyr-ethyl; acetic acid, -phenoxy]-ethyl ester], in or on the following...) Tolerances are established for residues of the herbicide flufenpyr-ethyl; acetic acid, -phenoxy]-ethyl...

  8. 40 CFR 180.595 - Flufenpyr-ethyl; tolerances for residues.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... residues of the herbicide, flufenpyr-ethyl; acetic acid, -phenoxy]-ethyl ester], in or on the following...) Tolerances are established for residues of the herbicide flufenpyr-ethyl; acetic acid, -phenoxy]-ethyl...

  9. 40 CFR 180.595 - Flufenpyr-ethyl; tolerances for residues.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... residues of the herbicide, flufenpyr-ethyl; acetic acid, -phenoxy]-ethyl ester], in or on the following...) Tolerances are established for residues of the herbicide flufenpyr-ethyl; acetic acid, -phenoxy]-ethyl...

  10. 40 CFR 180.595 - Flufenpyr-ethyl; tolerances for residues.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... residues of the herbicide, flufenpyr-ethyl; acetic acid, -phenoxy]-ethyl ester], in or on the following...) Tolerances are established for residues of the herbicide flufenpyr-ethyl; acetic acid, -phenoxy]-ethyl...

  11. Synthesis of Ethyl Salicylate Using Household Chemicals

    NASA Astrophysics Data System (ADS)

    Solomon, Sally; Hur, Chinhyu; Lee, Alan; Smith, Kurt

    1996-02-01

    Ethyl salicylate is synthesized, isolated, and characterized in a three-step process using simple equipment and household chemicals. First, acetylsalicylic acid is extracted from aspirin tablets with isopropyl alcohol, then hydrolyzed to salicylic acid with muriatic acid, and finally, the salicylic acid is esterified using ethanol and a boric acid catalyst. The experiment can be directed towards high school or university level students who have sufficient background in organic chemistry to recognize the structures and reactions that are involved.

  12. Production of ethyl alcohol from bananas

    SciTech Connect

    Jones, R.L.; Towns, T.

    1983-12-01

    The production of ethyl alcohol from waste bananas presents many special problems. During cooking, matting of the latex fibers from the banana peel recongeal when cooled and left untreated. This problem has been addressed by Alfaro by the use of CaC1/sub 2/. Separation of solids prior to distillation of the mashes in an economical fashion and use of the by product are also of concern to banana processors.

  13. Estimation of measurement uncertainty for the quantification of 11-nor-delta 9-tetrahydrocannabinol-9-carboxylic acid and its glucuronide in urine using liquid chromatography-tandem mass spectrometry.

    PubMed

    Kim, Jin Young; Kwon, Woonyong; Kim, Hee Seung; Suh, Sungill; In, Moon Kyo

    2014-04-01

    Recently, the estimation of the measurement uncertainty has become a significant issue in the quality control of forensic drug testing. In the present study, the uncertainty of the measurement was calculated for the quantification of 11-nor-delta 9-tetrahydrocannabinol-9-carboxylic acid (THC-COOH) and its glucuronide conjugate (THC-COOH-glu) in urine using liquid chromatography-tandem mass spectrometry. The procedure was based on liquid-liquid extraction of a volume of urine (800 µL) with ethyl acetate. The sources of uncertainty were identified and classified into four major categories as follows: standard preparation, calibration curve, method precision and bias. The overall contribution of combined standard uncertainty on THC-COOH increased in the order of standard preparation (0.9%), method precision (10.4%), calibration curve (30.3%) and bias (58.4%) and, while calibration curve (53.0%) and bias (40.4%) gave the bigger contributions to the combined standard uncertainty for THC-COOH-glu than method precision and standard preparation, which accounted for 6.3 and 0.3%, respectively. The reliability of a measurement was expressed by stating the expanded uncertainty of the measurement result at 95% confidence level. The concentrations of THC-COOH and THC-COOH-glu in the urine sample with their expanded uncertainties were 10.20 ± 1.14 ng/mL and 25.42 ± 5.01 ng/mL, respectively.

  14. 40 CFR 721.3152 - Ethanaminium, N-ethyl-2-hydroxy-N,N-bis(2-hydroxyethyl)-, diester with C12-18 fatty acids, ethyl...

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ...-bis(2-hydroxyethyl)-, diester with C12-18 fatty acids, ethyl sulfates (salts). 721.3152 Section 721... Ethanaminium, N-ethyl-2-hydroxy-N,N-bis(2-hydroxyethyl)-, diester with C12-18 fatty acids, ethyl sulfates... ethanaminium, N-ethyl-2-hydroxy-N,N-bis(2-hydroxyethyl)-, diester with C12-18 fatty acids, ethyl...

  15. 40 CFR 721.3152 - Ethanaminium, N-ethyl-2-hydroxy-N,N-bis(2-hydroxyethyl)-, diester with C12-18 fatty acids, ethyl...

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ...-bis(2-hydroxyethyl)-, diester with C12-18 fatty acids, ethyl sulfates (salts). 721.3152 Section 721... Ethanaminium, N-ethyl-2-hydroxy-N,N-bis(2-hydroxyethyl)-, diester with C12-18 fatty acids, ethyl sulfates... ethanaminium, N-ethyl-2-hydroxy-N,N-bis(2-hydroxyethyl)-, diester with C12-18 fatty acids, ethyl...

  16. 40 CFR 721.3152 - Ethanaminium, N-ethyl-2-hydroxy-N,N-bis(2-hydroxyethyl)-, diester with C12-18 fatty acids, ethyl...

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ...-bis(2-hydroxyethyl)-, diester with C12-18 fatty acids, ethyl sulfates (salts). 721.3152 Section 721... Ethanaminium, N-ethyl-2-hydroxy-N,N-bis(2-hydroxyethyl)-, diester with C12-18 fatty acids, ethyl sulfates... ethanaminium, N-ethyl-2-hydroxy-N,N-bis(2-hydroxyethyl)-, diester with C12-18 fatty acids, ethyl...

  17. 40 CFR 721.3152 - Ethanaminium, N-ethyl-2-hydroxy-N,N-bis(2-hydroxyethyl)-, diester with C12-18 fatty acids, ethyl...

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ...-bis(2-hydroxyethyl)-, diester with C12-18 fatty acids, ethyl sulfates (salts). 721.3152 Section 721... Ethanaminium, N-ethyl-2-hydroxy-N,N-bis(2-hydroxyethyl)-, diester with C12-18 fatty acids, ethyl sulfates... ethanaminium, N-ethyl-2-hydroxy-N,N-bis(2-hydroxyethyl)-, diester with C12-18 fatty acids, ethyl...

  18. Chemical and thermochemical aspects of the ozonolysis of ethyl oleate: decomposition enthalpy of ethyl oleate ozonide.

    PubMed

    Cataldo, Franco

    2013-01-01

    Neat ethyl oleate was ozonized in a bubble reactor and the progress of the ozonolysis was followed by infrared (FT-IR) spectroscopy and by the differential scanning calorimetry (DSC). The ozonolysis was conducted till a molar ratio O3/C=C≈1 when the exothermal reaction spontaneously went to completion. A specific thermochemical calculation on ethyl oleate ozonation has been made to determine the theoretical heat of the ozonization reaction using the group increment approach. A linear relationship was found both in the integrated absorptivity of the ozonide infrared band at 1110 cm(-1) and the ozonolysis time as well as the thermal decomposition enthalpy of the ozonides and peroxides formed as a result of the ozonation. The DSC decomposition temperature of ozonated ethyl oleate occurs with an exothermal peak at about 150-155 °C with a decomposition enthalpy of 243.0 kJ/mol at molar ratio O3/C=C≈1. It is shown that the decomposition enthalpy of ozonized ethyl oleate is a constant value (≈243 kJ/mol) at any stage of the O3/C=C once an adequate normalization of the decomposition enthalpy for the amount of the adsorbed ozone is taken into consideration. The decomposition enthalpy of ozonized ethyl oleate was also calculated using a simplified thermochemical model, obtaining a result in reasonable agreement with the experimental value.

  19. Glucuronidation of Dihydrotestosterone and trans-Androsterone by Recombinant UDP-Glucuronosyltransferase (UGT) 1A4: Evidence for Multiple UGT1A4 Aglycone Binding Sites

    PubMed Central

    Zhou, Jin; Tracy, Timothy S.

    2010-01-01

    UDP-glucuronosyltransferase (UGT) 1A4-catalyzed glucuronidation is an important drug elimination pathway. Although atypical kinetic profiles (nonhyperbolic, non-Michaelis-Menten) of UGT1A4-catalyzed glucuronidation have been reported occasionally, systematic kinetic studies to explore the existence of multiple aglycone binding sites in UGT1A4 have not been conducted. To this end, two positional isomers, dihydrotestosterone (DHT) and trans-androsterone (t-AND), were used as probe substrates, and their glucuronidation kinetics with HEK293-expressed UGT1A4 were evaluated both alone and in the presence of a UGT1A4 substrate [tamoxifen (TAM) or lamotrigine (LTG)]. Coincubation with TAM, a high-affinity UGT1A4 substrate, resulted in a concentration-dependent activation/inhibition effect on DHT and t-AND glucuronidation, whereas LTG, a low-affinity UGT1A4 substrate, noncompetitively inhibited both processes. The glucuronidation kinetics of TAM were then evaluated both alone and in the presence of different concentrations of DHT or t-AND. TAM displayed substrate inhibition kinetics, suggesting that TAM may have two binding sites in UGT1A4. However, the substrate inhibition kinetic profile of TAM became more hyperbolic as the DHT or t-AND concentration was increased. Various two-site kinetic models adequately explained the interactions between TAM and DHT or TAM and t-AND. In addition, the effect of TAM on LTG glucuronidation was evaluated. In contrast to the mixed effect of TAM on DHT and t-AND glucuronidation, TAM inhibited LTG glucuronidation. Our results suggest that multiple aglycone binding sites exist within UGT1A4, which may result in atypical kinetics (both homotropic and heterotropic) in a substrate-dependent fashion. PMID:20007295

  20. Glucuronidation of dihydrotestosterone and trans-androsterone by recombinant UDP-glucuronosyltransferase (UGT) 1A4: evidence for multiple UGT1A4 aglycone binding sites.

    PubMed

    Zhou, Jin; Tracy, Timothy S; Remmel, Rory P

    2010-03-01

    UDP-glucuronosyltransferase (UGT) 1A4-catalyzed glucuronidation is an important drug elimination pathway. Although atypical kinetic profiles (nonhyperbolic, non-Michaelis-Menten) of UGT1A4-catalyzed glucuronidation have been reported occasionally, systematic kinetic studies to explore the existence of multiple aglycone binding sites in UGT1A4 have not been conducted. To this end, two positional isomers, dihydrotestosterone (DHT) and trans-androsterone (t-AND), were used as probe substrates, and their glucuronidation kinetics with HEK293-expressed UGT1A4 were evaluated both alone and in the presence of a UGT1A4 substrate [tamoxifen (TAM) or lamotrigine (LTG)]. Coincubation with TAM, a high-affinity UGT1A4 substrate, resulted in a concentration-dependent activation/inhibition effect on DHT and t-AND glucuronidation, whereas LTG, a low-affinity UGT1A4 substrate, noncompetitively inhibited both processes. The glucuronidation kinetics of TAM were then evaluated both alone and in the presence of different concentrations of DHT or t-AND. TAM displayed substrate inhibition kinetics, suggesting that TAM may have two binding sites in UGT1A4. However, the substrate inhibition kinetic profile of TAM became more hyperbolic as the DHT or t-AND concentration was increased. Various two-site kinetic models adequately explained the interactions between TAM and DHT or TAM and t-AND. In addition, the effect of TAM on LTG glucuronidation was evaluated. In contrast to the mixed effect of TAM on DHT and t-AND glucuronidation, TAM inhibited LTG glucuronidation. Our results suggest that multiple aglycone binding sites exist within UGT1A4, which may result in atypical kinetics (both homotropic and heterotropic) in a substrate-dependent fashion.

  1. Theoretical study of the decomposition of ethyl and ethyl 3-phenyl glycidate.

    PubMed

    Josa, Daniela; Peña-Gallego, Angeles; Rodríguez-Otero, Jesús; Cabaleiro-Lago, Enrique M

    2013-01-01

    The mechanism of the decomposition of ethyl and ethyl 3-phenyl glycidate in gas phase was studied by density functional theory (DFT) and MP2 methods. A proposed mechanism for the reaction indicates that the ethyl side of the ester is eliminated as ethylene through a concerted six-membered cyclic transition state, and the unstable intermediate glycidic acid decarboxylates rapidly to give the corresponding aldehyde. Two possible pathways for glycidic acid decarboxylation were studied: one via a five-membered cyclic transition state, and the other via a four-membered cyclic transition state. The results of the calculations indicate that the decarboxylation reaction occurs via a mechanism with five-membered cyclic transition state.

  2. Identification of Flavone Glucuronide Isomers by Metal Complexation and Tandem Mass Spectrometry: Regioselectivity of UDP-Glucuronosyltransferase Isozymes in the Biotransformation of Flavones

    PubMed Central

    Robotham, Scott A.; Brodbelt, Jennifer S.

    2013-01-01

    Flavone Glucuronide isomers of five flavones (chrysin, apigenin, luteolin, baicalein, and scutellarein) were differentiated by collision induced dissociation (CID) of [Co(II) (flavone-H) (4,7-diphenyl-1,10-phenanthroline)2]+ complexes. The complexes were generated via post-column addition of a metal/ligand solution after separation of the glucuronide products generated upon incubation of each flavone with an array of UDP-glucuronosyl-transferase (UGT) isozymes. Elucidation of the glucuronide isomers allowed a systematic investigation of the regioselectivity of twelve human UDP-glucuronosyl-transferase (UGT) isozymes, including eight UGT1A and four UGT2B isozymes. Glucuronidation of the 7-OH position was the preferred site for all the flavones except for luteolin, which possessed adjacent hydroxyl groups on the B ring. For all flavones and UGT isozymes, glucuronidation of the 5-OH position was never observed. As confirmed by the metal complexation/MS/MS strategy, glucuronidation of the 6-OH position only occurred for baicalein and scutellarein when incubated with three of the UGT isozymes. PMID:23362992

  3. Structure-activity relationship (SAR): effort towards blocking N-glucuronidation of indazoles (PF-03376056) by human UGT1A enzymes.

    PubMed

    Rose, Kelly; Yang, Young-Sun; Sciotti, Richard; Cai, Hongliang

    2009-01-01

    GyrATPase is a cellular enzyme that has been used as an antibacterial target for treatment of nosocomial and community acquired bacterial infections. The leading chemical series targeted at inhibiting this enzyme, indazoles, were rapidly cleared in rats (CL > 70 mL/min/kg). The predominant metabolite identified in both urine and bile samples from a bile duct-cannulated study corresponded to direct glucuronidation of the parent compound and was excreted rapidly. Subsequently, a carefully designed analog was used to pinpoint the site of glucuronidation (N-glucuronidation) by incubation with rat hepatocytes and followed by mass spectrometry analysis. Reaction mapping with an array of recombinant UGT isozymes revealed that N-glucuronidation was predominantly catalyzed by the UGT1A family of enzymes. Based on the results, the following approaches were considered to reduce or eliminate glucuronidation: 1) adding sterically hindered substitutions on the phenyl ring of the indazole core; 2) changing the electron distribution by substituting with electron-donating or -withdrawing groups; 3) replacing the site of glucuronidation. The resulted compounds were evaluated in vitro in rat hepatocytes to assess their metabolic stabilities followed by in vivo efficacy studies in the murine peritonitis sepsis model (at 50 mg/kg) for selected compounds.

  4. Simultaneous determination of mycophenolic acid and its glucuronide and glycoside derivatives in canine and feline plasma by UHPLC-UV.

    PubMed

    Rivera, Sol Maiam; Hwang, Julianne K; Slovak, Jeniffer E; Court, Michael H; Villarino, Nicolas F

    2017-02-01

    Cats and dogs can suffer from multiple autoimmune diseases. Mycophenolic acid (MPA) is a potentially useful immunosuppressant drug in cats and dogs, because of its well-documented efficacy in controlling autoimmune disease in humans. However, the pharmacokinetics and pharmacodynamics in these species remain to be determined. We have developed and validated a sensitive, precise, accurate and reproducible method that provides consistent quantification of MPA and its major derivatives, MPA phenol glucoside and MPA phenol glucuronide, in canine and feline plasma using ultra-high-pressure liquid chromatography coupled to an ultraviolet detector. The main advantages of this novel method include a small sample volume, easy sample preparation, a short chromatographic analysis time and the option to select either phenolphthalein β-d-glucuronide or mycophenolic acid carboxybutoxy ether as internal standard. Results of validation indicate that this analytical method is suitable to study the plasma disposition of MPA and its derivatives in dogs and cats.

  5. Identification of brain-targeted bioactive dietary quercetin-3-O-glucuronide as a novel intervention for Alzheimer's disease.

    PubMed

    Ho, Lap; Ferruzzi, Mario G; Janle, Elsa M; Wang, Jun; Gong, Bing; Chen, Tzu-Ying; Lobo, Jessica; Cooper, Bruce; Wu, Qing Li; Talcott, Stephen T; Percival, Susan S; Simon, James E; Pasinetti, Giulio Maria

    2013-02-01

    Epidemiological and preclinical studies indicate that polyphenol intake from moderate consumption of red wines may lower the relative risk for developing Alzheimer's disease (AD) dementia. There is limited information regarding the specific biological activities and cellular and molecular mechanisms by which wine polyphenolic components might modulate AD. We assessed accumulations of polyphenols in the rat brain following oral dosage with a Cabernet Sauvignon red wine and tested brain-targeted polyphenols for potential beneficial AD disease-modifying activities. We identified accumulations of select polyphenolic metabolites in the brain. We demonstrated that, in comparison to vehicle-control treatment, one of the brain-targeted polyphenol metabolites, quercetin-3-O-glucuronide, significantly reduced the generation of β-amyloid (Aβ) peptides by primary neuron cultures generated from the Tg2576 AD mouse model. Another brain-targeted metabolite, malvidin-3-O-glucoside, had no detectable effect on Aβ generation. Moreover, in an in vitro analysis using the photo-induced cross-linking of unmodified proteins (PICUP) technique, we found that quercetin-3-O-glucuronide is also capable of interfering with the initial protein-protein interaction of Aβ(1-40) and Aβ(1-42) that is necessary for the formation of neurotoxic oligomeric Aβ species. Lastly, we found that quercetin-3-O-glucuronide treatment, compared to vehicle-control treatment, significantly improved AD-type deficits in hippocampal formation basal synaptic transmission and long-term potentiation, possibly through mechanisms involving the activation of the c-Jun N-terminal kinases and the mitogen-activated protein kinase signaling pathways. Brain-targeted quercetin-3-O-glucuronide may simultaneously modulate multiple independent AD disease-modifying mechanisms and, as such, may contribute to the benefits of dietary supplementation with red wines as an effective intervention for AD.

  6. 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.

  7. β-Glucuronidase activity and mitochondrial dysfunction: the sites where flavonoid glucuronides act as anti-inflammatory agents.

    PubMed

    Kawai, Yoshichika

    2014-05-01

    Epidemiological and experimental studies suggest that the consumption of flavonoid-rich diets decreases the risk of various chronic diseases such as cardiovascular diseases. Although studies on the bioavailability of flavonoids have been well-characterized, the tissue and cellular localizations underlying their biological mechanisms are largely unknown. The development and application of novel monoclonal antibodies revealed that macrophages could be the major target of dietary flavonoids in vivo. Using macrophage-like cell lines in vitro, we examined the molecular basis of the interaction between the macrophages and flavonoids, especially the glucuronide metabolites. We have found that extracellular β-glucuronidase secreted from macrophages is essential for the bioactivation of the glucuronide conjugates into the aglycone, and that the enzymatic activity, which requires an acidic pH, is promoted by the increased secretion of lactate in response to the mitochondrial dysfunction. This review describes our recent findings indicating the molecular mechanisms responsible for the anti-inflammatory activity of dietary flavonoids within the inflammation sites. We propose that the extracellular activity of β-glucuronidase associated with the status of the mitochondrial function in the target cells might be important biomarkers for the specific sites where the glucuronides of dietary flavonoids can act as anti-atherosclerotic and anti-inflammatory agents in vivo.

  8. Measurement of direct ethanol metabolites in a case of a former driving under the influence (DUI) of alcohol offender, now claiming abstinence.

    PubMed

    Wurst, Friedrich M; Yegles, Michel; Alling, Christer; Aradottir, Steina; Dierkes, Jutta; Wiesbeck, Gerhard A; Halter, Claudia C; Pragst, Fritz; Auwaerter, Volker

    2008-05-01

    A 37-year-old female subject had been convicted of driving under the influence of alcohol, and 19 months later, claimed abstinence after supervised disulfiram treatment. Our aim was to elucidate the value of direct ethanol metabolites as measures of abstinence. Ethyl glucuronide (EtG) and fatty acid ethyl esters (FAEE) in hair, phosphatidylethanol in whole blood and EtG and ethyl sulphate in urine were measured. The results were compared with self-report of alcohol consumption and traditional blood biomarkers for chronically elevated alcohol consumption as carbohydrate deficient transferrin (CDT), gamma glutamyl transpeptidase, mean corpuscular erythrocyte volume, aspartate aminotransferase and alanine aminotransferase. EtG was found in distal parts of hair only, whereas the proximal parts were negative. Furthermore, FAEE concentrations were found in the typical distribution over the hair length and showed values typical for either moderate social drinking or abstinence. CDT was above cut-off in 9 out of 16 analyses with a decreasing tendency and the lowest values in the last 2 months before the end of sampling. The data suggest that in addition to traditional markers, a combination of direct ethanol metabolites can be useful in the expert assessment of judging driving ability. A careful individual interpretation of the results for the different markers, however, is an absolute necessity.

  9. Biomarkers for the Detection of Prenatal Alcohol Exposure: A Review.

    PubMed

    Bager, Heidi; Christensen, Lars Porskjaer; Husby, Steffen; Bjerregaard, Lene

    2017-02-01

    Alcohol exposure during pregnancy can cause adverse effects to the fetus, because it interferes with fetal development, leading to later physical and mental impairment. The most common clinical tool to determine fetal alcohol exposure is maternal self-reporting. However, a more objective and useful method is based on the use of biomarkers in biological specimens alone or in combination with maternal self-reporting. This review reports on clinically relevant biomarkers for detection of prenatal alcohol exposure (PAE). A systematic search was performed to ensure a proper overview in existing literature. Studies were selected to give an overview on clinically relevant neonatal and maternal biomarkers. The direct biomarkers fatty acid ethyl esters (FAEEs), ethyl glucuronide (EtG), ethyl sulfate, and phosphatidylethanol (PEth) were found to be the most appropriate biomarkers in relation to detection of PAE. To review each biomarker in a clinical context, we have compared the advantages and disadvantages of each biomarker, in relation to its window of detectability, ease of collection, and the ease and cost of analysis of each biomarker. The biomarkers PEth, FAEEs, and EtG were found to be applicable for detection of even low levels of alcohol exposure. Meconium is an accessible matrix for determination of FAEEs and EtG, and blood an accessible matrix for determination of PEth.

  10. [Direct metabolites of ethanol as biological markers of alcohol use: basic aspects and applications].

    PubMed

    Thon, N; Weinmann, W; Yegles, M; Preuss, U; Wurst, F M

    2013-09-01

    In addition to self reports and questionnaires, biomarkers are of relevance in the diagnosis of and therapy for alcohol use disorders. Traditional biomarkers such as gamma-glutamyl transpeptidase or mean corpuscular volume are indirect biomarkers and are subject to the influence of age, gender and non-alcohol related diseases, among others. Direct metabolites of ethanol such as ethyl glucuronide (EtG), ethyl sulphate (EtS) and phosphatidylethanol (PEth) are direct metabolites of ethanol, that are positive after intake of ethyl alcohol. They represent useful diagnostic tools for identifying alcohol use even more accurately than traditional biomarkers. Each of these drinking indicators remains positive in serum and urine for a characteristic time spectrum after the cessation of ethanol intake - EtG and EtS in urine up to 7 days, EtG in hair for months after ethanol has left the body. Applications include clinical routine use, emergency room settings, proof of abstinence in alcohol rehabilitation programmes, driving under influence offenders, workplace testing, assessment of alcohol intake in the context of liver transplantation and foetal alcohol syndrome. Due to their properties, they open up new perspectives for prevention, interdisciplinary cooperation, diagnosis of and therapy for alcohol-related problems.

  11. Glucuronidation of fenamates: kinetic studies using human kidney cortical microsomes and recombinant UDP-glucuronosyltransferase (UGT) 1A9 and 2B7.

    PubMed

    Gaganis, Paraskevi; Miners, John O; Knights, Kathleen M

    2007-05-15

    Mefenamic acid, a non-steroidal anti-inflammatory drug (NSAID), is used commonly to treat menorrhagia. This study investigated the glucuronidation kinetics of flufenamic, mefenamic and niflumic acid using human kidney cortical microsomes (HKCM) and recombinant UGT1A9 and UGT2B7. Using HKCM Michaelis-Menten (MM) kinetics were observed for mefenamic (K(m)(app) 23 microM) and niflumic acid (K(m)(app) 123 microM) glucuronidation, while flufenamic acid exhibited non-hyperbolic (atypical) glucuronidation kinetics. Notably, the intrinsic renal clearance of mefenamic acid (CL(int) 17+/-5.5 microL/minmg protein) was fifteen fold higher than that of niflumic acid (CL(int) 1.1+/-0.8 microL/minmg protein). These data suggest that renal glucuronidation of mefenamic acid may result in high intrarenal exposure to mefenamic acyl-glucuronide and subsequent binding to renal proteins. Diverse kinetics were observed for fenamate glucuronidation by UGT2B7 and UGT1A9. Using UGT2B7 MM kinetics were observed for flufenamic (K(m)(app) 48 microM) and niflumic acid (K(m)(app) 135 microM) glucuronidation and atypical kinetics with mefenamic acid. Similarity in K(m)(app) between HKCM and UGT2B7 suggests that UGT2B7 may be the predominant renal UGT isoform catalysing niflumic acid glucuronidation. In contrast, UGT1A9 glucuronidation kinetics were characterised by negative cooperativity with mefenamic (S(50) 449 microM, h 0.4) and niflumic acid (S(50) 7344 microM, h 0.4) while atypical kinetics were observed with flufenamic acid. Additionally, potent inhibition of the renal glucuronidation of the UGT substrate 'probe' 4-methylumbelliferone by flufenamic, mefenamic and niflumic acid was observed. These data suggest that inhibitory metabolic interactions may occur between fenamates and other substrates metabolised by UGT2B7 and UGT1A9 in human kidney.

  12. Quantitative Determination of Common Urinary Odorants and Their Glucuronide Conjugates in Human Urine

    PubMed Central

    Wagenstaller, Maria; Buettner, Andrea

    2013-01-01

    Our previous study on the identification of common odorants and their conjugates in human urine demonstrated that this substance fraction is a little-understood but nonetheless a promising medium for analysis and diagnostics in this easily accessible physiological medium. Smell as an indicator for diseases, or volatile excretion in the course of dietary processes bares high potential for a series of physiological insights. Still, little is known today about the quantitative composition of odorous or volatile targets, as well as their non-volatile conjugates, both with regard to their common occurrence in urine of healthy subjects, as well as in that of individuals suffering from diseases or other physiological misbalancing. Accordingly, the aim of our study was to develop a highly sensitive and selective approach to determine the common quantitative composition of selected odorant markers in healthy human subjects, as well as their corresponding glucuronide conjugates. We used one- and two-dimensional high resolution gas chromatography-mass spectrometry in combination with stable isotope dilution assays to quantify commonly occurring and potent odorants in human urine. The studies were carried out on both native urine and on urine that had been treated by glucuronidase assays, with analysis of the liberated odor-active compounds using the same techniques. Analytical data are discussed with regard to their potential translation as future diagnostic tool. PMID:24958143

  13. Temporal indication of cannabis use by means of THC glucuronide determination.

    PubMed

    Mareck, Ute; Haenelt, Nadine; Geyer, Hans; Guddat, Sven; Kamber, Matthias; Brenneisen, Rudolf; Thevis, Mario; Schänzer, Wilhelm

    2009-11-01

    According to the regulations of the World Anti-Doping Agency (WADA), the use of cannabinoids is forbidden in competition. In doping controls, the detection of cannabinoid misuse is based on the analysis of the non-psychoactive metabolite 11-nor-9-carboxy-delta-9-tetrahydrocannabinol (carboxy-THC). The determination of values greater than 15 ng/mL in urine represents an adverse analytical finding; however, no accurate prediction of the time of application is possible as the half-life of carboxy-THC ranges between three and four days. Consequently the detection of carboxy-THC in doping control urine samples collected in competition might also result from cannabis use in out-of-competition periods. The analysis of the glucuronide of the pharmacologically active delta 9-tetrahydrocannabinol (THC-gluc) may represent a complementary indicator for the detection of cannabis misuse in competition.An assay for the determination of THC-gluc in human urine was established. The sample preparation consisted of liquid-liquid extraction of urine specimens, and extracts were analysed by liquid chromatography/tandem mass spectrometry (LC-MS/MS). Authentic doping-control urine samples as well as specimens obtained from a controlled smoking study were analysed and assay characteristics such as specificity, detection limit (0.1 ng/mL), precision (>90%), recovery ( approximately 80%), and extraction efficiency (90%) were determined.

  14. Morphine-3-D glucuronide stability in postmortem specimens exposed to bacterial enzymatic hydrolysis.

    PubMed

    Carroll, F T; Marraccini, J V; Lewis, S; Wright, W

    2000-12-01

    Medical examiners frequently rely on the finding of free morphine present in postmortem specimens to assist in certifying deaths associated with narcotics. In vitro hydrolysis of morphine-3-D glucuronide (M3DG) to free morphine was studied using variable specimen pH, initial degree of specimen putrefaction, storage temperature and time, and the effectiveness of sodium fluoride (NaF) preservation. Reagent M3DG was added to opiate-free fresh blood and urine and to autopsy-derived blood specimens. Reagent bovine glucuronidase was also added to certain specimens. Freshly collected and refrigerated NaF-preserved blood produced minimal free morphine, whereas four of five autopsy blood specimens produced free morphine from M3DG. Increased storage time, temperature, and initial degree of putrefaction resulted in greater free morphine generation despite the absence of viable bacteria. Hydrolysis occurring during specimen storage can generate free morphine from M3DG and may result in erroneous conclusions in certifying narcotic deaths.

  15. Quercetin-3-O-glucuronide induces ABCA1 expression by LXRα activation in murine macrophages

    SciTech Connect

    Ohara, Kazuaki; Wakabayashi, Hideyuki; Taniguchi, Yoshimasa; Shindo, Kazutoshi; Yajima, Hiroaki; Yoshida, Aruto

    2013-11-29

    Highlights: •The major circulating quercetin metabolite (Q3GA) activated LXRα. •Q3GA induced ABCA1 via LXRα activation in macrophages. •Nelumbo nucifera leaf extracts contained quercetin glycosides. •N. nucifera leaf extract feeding elevated HDLC in mice. -- Abstract: Reverse cholesterol transport (RCT) removes excess cholesterol from macrophages to prevent atherosclerosis. ATP-binding cassette, subfamily A, member 1 (ABCA1) is a crucial cholesterol transporter involved in RCT to produce high density lipoprotein-cholesterol (HDLC), and is transcriptionally regulated by liver X receptor alpha (LXRα), a nuclear receptor. Quercetin is a widely distributed flavonoid in edible plants which prevented atherosclerosis in an animal model. We found that quercetin-3-O-glucuronide (Q3GA), a major quercetin metabolite after absorption from the digestive tract, enhanced ABCA1 expression, in vitro, via LXRα in macrophages. In addition, leaf extracts of a traditional Asian edible plant, Nelumbo nucifera (NNE), which contained abundant amounts of quercetin glycosides, significantly elevated plasma HDLC in mice. We are the first to present experimental evidence that Q3GA induced ABCA1 in macrophages, and to provide an alternative explanation to previous studies on arteriosclerosis prevention by quercetin.

  16. Nasal administration of morphine-6-glucuronide in sheep--a pharmacokinetic study.

    PubMed

    Illum, L; Davis, S S; Pawula, M; Fisher, A N; Barrett, D A; Farraj, N F; Shaw, P N

    1996-11-01

    The pharmacokinetics of morphine-6-glucuronide (M6G) after both intravenous dosing and nasal administration were studied in sheep. The nasal formulation consisted of M6G in combination with an absorption promoting delivery system in the form of chitosan. The mean half-life of M6G after intravenous administration was 51.0 +/- 8.2 min and that after intranasal dosing was 45.0 +/- 5.5 min. M6G clearance and volume of distribution were 5.4 +/- 1.5 mL min-1 kg-1 and 0.4 +/- 0.1 L kg-1 respectively. The plasma profile after nasal administration demonstrated rapid absorption of M6G. The bioavailability of M6G in the chitosan formulation was found to be 31.4%. These results suggest that M6G administered in combination with the chitosan delivery system may be considered as a suitable non-parenteral means of administering this analgesic.

  17. Molecular Pathways: GLI1-Induced Drug Glucuronidation in Resistant Cancer Cells.

    PubMed

    Zahreddine, Hiba Ahmad; Borden, Katherine L B

    2015-05-15

    Drug resistance remains a major impediment in the development of durable cancer therapies. Studies in acute myelogenous leukemia (AML) patients revealed a new form of multidrug resistance. Here, increased glioma-associated protein GLI1 leads to elevation of the UDP-glucuronosyl transferase (UGT) enzymes. UGTs add glucuronic acid to xenobiotics and metabolites. Traditionally, the loss of these enzymes is thought to contribute to cancer as a result of impaired clearance of environmental carcinogens. However, we demonstrate that overexpression of UGTs can contribute to oncogenesis by promoting drug resistance. Indeed, UGT levels in AML patients treated with ribavirin and/or cytarabine were elevated at relapse relative to diagnosis. This was reversed by GLI1 inhibition, suggesting a clinically relevant strategy to overcome drug resistance. Further, overexpression of UGTs can also lead to drug resistance in other cancers, such as certain Hsp90 inhibitors and vorinostat in colorectal and chronic lymphoblastic leukemia, respectively. Not all drugs are targets of glucuronidation, suggesting that UGT status could be relevant to treatment choice. Here, we describe several facets of UGT biology and how these could be exploited clinically. These studies demonstrate how drugs in cancer cells can be metabolized differentially than their normal counterparts. In summary, we describe a new form of drug resistance relevant to a variety of cancer contexts.

  18. UHPLC-MS/MS quantification of buprenorphine, norbuprenorphine, methadone, and glucuronide conjugates in umbilical cord plasma.

    PubMed

    Kyle, Amy Redmond; Carmical, Jennifer; Shah, Darshan; Pryor, Jason; Brown, Stacy

    2015-10-01

    Opioid use during pregnancy can result in the newborn being physically dependent on the substance, thus experiencing drug withdrawal, termed neonatal abstinence syndrome (NAS). Buprenorphine and methadone are two drugs used to treat opioid withdrawal and are approved for use in pregnancy. Quantification of these compounds in umbilical cord plasma would help assess in utero exposure of neonates in cases of buprenorphine or methadone use during pregnancy. An LC-MS/MS method using solid-phase extraction sample preparation was developed and validated for the simultaneous quantification of methadone, buprenorphine, norbuprenorphine, and glucuronide metabolites in umbilical cord plasma. The average accuracy (percentage error) and precision (relative standard deviation) were <15% for each validated concentration. Our data establishes a 2 week maximum freezer storage window in order to achieve the most accurate cord plasma concentrations of these analytes. Additionally, we found that the umbilical cord tissue analysis was less sensitive compared with analysis with umbilical cord blood plasma, indicating that this may be a more appropriate matrix for determination of buprenorphine and metabolite concentrations. This method was successfully applied to the analysis of cord blood from women with known buprenorphine or methadone use during pregnancy.

  19. 21 CFR 172.872 - Methyl ethyl cellulose.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 3 2012-04-01 2012-04-01 false Methyl ethyl cellulose. 172.872 Section 172.872... CONSUMPTION Multipurpose Additives § 172.872 Methyl ethyl cellulose. The food additive methyl ethyl cellulose... a cellulose ether having the general formula [C6H(10 -x-y)O5(CH3)x(C2H5)y]n, where x is the...

  20. 21 CFR 172.872 - Methyl ethyl cellulose.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 3 2011-04-01 2011-04-01 false Methyl ethyl cellulose. 172.872 Section 172.872... CONSUMPTION Multipurpose Additives § 172.872 Methyl ethyl cellulose. The food additive methyl ethyl cellulose... a cellulose ether having the general formula [C6H(10 -x-y)O5(CH3)x(C2H5)y]n, where x is the...

  1. 21 CFR 172.872 - Methyl ethyl cellulose.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 3 2013-04-01 2013-04-01 false Methyl ethyl cellulose. 172.872 Section 172.872... CONSUMPTION Multipurpose Additives § 172.872 Methyl ethyl cellulose. The food additive methyl ethyl cellulose... a cellulose ether having the general formula [C6H(10 -x-y)O5(CH3)x(C2H5)y]n, where x is the...

  2. 21 CFR 172.872 - Methyl ethyl cellulose.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Methyl ethyl cellulose. 172.872 Section 172.872... CONSUMPTION Multipurpose Additives § 172.872 Methyl ethyl cellulose. The food additive methyl ethyl cellulose... a cellulose ether having the general formula [C6H(10 -x-y)O5(CH3)x(C2H5)y]n, where x is the...

  3. IRIS Toxicological Review of Ethyl Tertiary Butyl Ether (Etbe) ...

    EPA Pesticide Factsheets

    EPA is conducting a peer review and public comment of the scientific basis supporting the human health hazard and dose-response assessment of ethyl tertiary butyl ether (ETBE) that when finalized will appear on the Integrated Risk Information System (IRIS) database. The draft Toxicological Review of Ethyl Tertiary Butyl Ether provides scientific support and rationale for the hazard and dose-response assessment pertaining to chronic exposure to ethyl tertiary butyl ether.

  4. Going the distance with ethyl alcohol

    SciTech Connect

    Hairston, D.W.

    1995-12-01

    If all had gone according to plan, ethyl alcohol would be in the driver`s seat now, cruising down the highway and getting ready to speed into high gear. Instead, this renewable fuel, chemical reagent and solvent is navigating a complex obstacle course, watching warily for sharp turns and mixed signals. Globally, the supply and demand for all grades of ethyl alcohol is awry. Production of industrial-grade material is running at full throttle and prices are going up. Much of the upheaval over ethanol can be traced to the US Environmental Protection Agency and the renewable oxygenate standard (ROS) of the Clean Air Act. Under ROS, 15% of oxygenates used in gasoline sold this year was to be derived from a renewable source. Next month, that percentage was to have been doubled to 30%. Enticed by projections of upwards of 2 billion gal/yr of fermentation alcohol to comply with ROS, producers rushed to expand capacity. But to the producers` dismay, EPA was forced to backpedal on ROS. When representatives of the petroleum industry filed suit and won a stay, EPA rescinded its ROS regulation and ethanol producers were left in the lurch. High prices for corn is also putting the squeeze on inventories of industrial alcohol. Synthetic ethanol production, from ethylene for example, is booming, however. This paper discusses the ethanol market factors.

  5. Morphine glucuronidation and glucosidation represent complementary metabolic pathways that are both catalyzed by UDP-glucuronosyltransferase 2B7: kinetic, inhibition, and molecular modeling studies.

    PubMed

    Chau, Nuy; Elliot, David J; Lewis, Benjamin C; Burns, Kushari; Johnston, Martin R; Mackenzie, Peter I; Miners, John O

    2014-04-01

    Morphine 3-β-D-glucuronide (M3G) and morphine 6-β-D-glucuronide (M6G) are the major metabolites of morphine in humans. More recently, morphine-3-β-d-glucoside (M-3-glucoside) was identified in the urine of patients treated with morphine. Kinetic and inhibition studies using human liver microsomes (HLM) and recombinant UGTs as enzyme sources along with molecular modeling were used here to characterize the relationship between morphine glucuronidation and glucosidation. The M3G to M6G intrinsic clearance (C(Lint)) ratio (∼5.5) from HLM supplemented with UDP-glucuronic acid (UDP-GlcUA) alone was consistent with the relative formation of these metabolites in humans. The mean C(Lint) values observed for M-3-glucoside by incubations of HLM with UDP-glucose (UDP-Glc) as cofactor were approximately twice those for M6G formation. However, although the M3G-to-M6G C(Lint) ratio remained close to 5.5 when human liver microsomal kinetic studies were performed in the presence of a 1:1 mixture of cofactors, the mean C(Lint) value for M-3-glucoside formation was less than that of M6G. Studies with UGT enzyme-selective inhibitors and recombinant UGT enzymes, along with effects of BSA on morphine glycosidation kinetics, were consistent with a major role of UGT2B7 in both morphine glucuronidation and glucosidation. Molecular modeling identified key amino acids involved in the binding of UDP-GlcUA and UDP-Glc to UGT2B7. Mutagenesis of these residues abolished morphine glucuronidation and glucosidation. Overall, the data indicate that morphine glucuronidation and glucosidation occur as complementary metabolic pathways catalyzed by a common enzyme (UGT2B7). Glucuronidation is the dominant metabolic pathway because the binding affinity of UDP-GlcUA to UGT2B7 is higher than that of UDP-Glc.

  6. Effect of MRP2 and MRP3 Polymorphisms on Anastrozole Glucuronidation and MRP2 and MRP3 Gene Expression in Normal Liver Samples

    PubMed Central

    Edavana, Vineetha Koroth; Penney, Rosalind B; Yao- Borengasser, Aiwei; Starlard-Davenport, Athena; Dhakal, Ishwori B; Kadlubar, Susan

    2015-01-01

    Anastrozole is an aromatase inhibitor (AI) used as adjuvant therapy for breast cancer. Anastrozole is subject to direct glucuronidation catalyzed by UDP-glucuronosyltransferase1A4 (UGT1A4). Interindividual variability in anastrozole glucuronidation may be affected by UGT1A4 SNPs. Interplay between drug metabolizing genes such as UGT1A4 and transporter genes may also be affected by genetic variability. Thus, we hypothesize that genetic variability in MRPs could influence anastrozole glucuronidation. The correlation between UGT1A4 and MRP2 or MRP3 transporter gene expressions and the correlation between MRP2 or MRP3 mRNA and anastrozole glucuronidation were analyzed in normal human liver samples. MRP2 and MRP3 mRNA levels were significantly correlated with UGT1A4 mRNA, with anastrozole glucuronidation and with each other (p<0.05). The data also demonstrated that MRP2 SNPs are positively correlated with MRP2 mRNA expression, while there was no association between MRP3 SNPs from this study and MRP3 expression. Significant correlations (p<0.05) between certain MRP2 SNPs (3972C>T, 2366C>T and −24C>T) and anastrozole glucuronidation were observed. There were no observed correlations between MRP3 SNPs and anastrozole glucuronidation. MRP2 polymorphisms have been identified as playing a role in the disposition of other drugs, and the data presented here indicate for the first time that MRP2 SNPs could influence anastrozole metabolism and contribute to interindividual variation in treatment responses. PMID:26985457

  7. Ground-state stability and rotational activation parameters for individual rotamers of (R,S,S,R)-(N,N'-dimethyl-2,3-diaminobutane)PtG(2) complexes (G = 9-EtG, 3'-GMP, and 5'-GMP).

    PubMed

    Colonna, Gianpiero; Di Masi, Nicola G; Marzilli, Luigi G; Natile, Giovanni

    2003-02-24

    Rate constants for guanine rotation about the Pt-N7 bond in (R,S,S,R)-Me(2)DABPtG(2) complexes (Me(2)DAB = N,N'-dimethyl-2,3-diaminobutane; G = 9-EtG, 3'-GMP, and 5'-GMP) were evaluated from line-shape analysis of H8 resonances. Three diastereomers, two in head-to-tail (DeltaHT and LambdaHT) and one in head-to-head (HH) conformations, exist in equilibrium in solution. The two guanines are equivalent in DeltaHT and LambdaHT conformers and nonequivalent in the HH form; therefore, four rate constants (k(Delta)(HT), k(Lambda)(HT), k(HH)()s, and k(HH)()d; sub-subscripts s and d stand for H8-shielded and -deshielded guanine, respectively) were evaluated. Activation parameters (DeltaH and DeltaS) were evaluated from the rate constant dependence on temperature. High values of DeltaH (78-93 kJ mol(-)(1)) and DeltaS (51-71 J K(-)(1) mol(-)(1)) were found for G rotation in the preferred DeltaHT rotamer having the six-membered ring of each guanine more canted toward the cis-G and a favorable dipole-dipole internucleotide interaction. Lower values of DeltaH() (64-76 kJ mol(-)(1)) and very small values of DeltaS() (-7-11 J K(-)(1) mol(-)(1)) were found for G rotation in the less favorable LambdaHT rotamer, indicating that the ground-state entropy of this rotamer is close to that of the activated complex and the ground-state enthalpy closer to that of the activated complex than for the DeltaHT rotamer. For the two guanines in the HH rotamer there is no large difference in activation parameters. In general DeltaH falls in the range 66-84 kJ mol(-)(1) (rather close to the values for the LambdaHT rotamer) and DeltaS in the range 14-41 J K(-)(1) mol(-)(1). The equilibrium constant between HT and HH rotamers was also evaluated together with the corresponding thermodynamic parameters (DeltaH and DeltaS). It is found that the low enthalpy is the major stabilizing factor for DeltaHT as compared to HH, while the entropy factor would favor the latter rotamer. In contrast the greater

  8. The effect of the use of mouthwash on ethylglucuronide concentrations in urine.

    PubMed

    Costantino, Anthony; Digregorio, E John; Korn, Warren; Spayd, Stephanie; Rieders, Frederic

    2006-01-01

    Two studies were performed to evaluate the effect of alcohol containing mouthwash on the appearance of ethyl glucuronide (EtG) in urine. In the first study, 9 volunteers were given a 4-oz bottle of mouthwash, which contained 12% ethanol. They gargled with all 4 oz. of the mouthwash at intervals over a 15-min period. All urine samples were collected over the next 24 h. Of 39 provided urine samples, there were 20 > 50 ng/mL, 12 > 100 ng/mL, 5 > 200 ng/mL, 3 > 250 ng/mL, and 1 > 300 ng/mL. The peak concentrations were all within 12 h after the exposure. In the second study, 11 participants gargled 3 times daily for 5 days. The first morning void was collected. Sixteen of the 55 submitted samples contained EtG concentrations of greater than 50 ng/mL. All of them were less than 120 ng/mL. These studies show that incidental exposure to mouthwash containing 12% ethanol, when gargling according to the manufacturer's instructions, can result in urinary EtG values greater than 50 ng/mL. All specimens were negative for ethanol. The limits of detection and quantitation for the EtG testing were 50 ng/mL.

  9. Quaternary ammonium-linked glucuronidation of trans-4-hydroxytamoxifen, an active metabolite of tamoxifen, by human liver microsomes and UDP-glucuronosyltransferase 1A4.

    PubMed

    Ogura, Kenichiro; Ishikawa, Yuko; Kaku, Teppei; Nishiyama, Takahito; Ohnuma, Tomokazu; Muro, Kei; Hiratsuka, Akira

    2006-04-28

    Tamoxifen (TAM), a nonsteroidal antiestrogen, is the most widely used drug for chemotherapy of hormone-dependent breast cancer in women. Trans-4-hydroxy-TAM (trans-4-HO-TAM), one of the TAM metabolites in humans, has been considered to be an active metabolite of TAM because of its higher affinity toward estrogen receptors (ERs) than the parent drug and other side-chain metabolites. In the present study, we found a new potential metabolic pathway of trans-4-HO-TAM and its geometrical isomer, cis-4-HO-TAM, via N-linked glucuronic acid conjugation for excretion in humans. N+-Glucuronides of 4-HO-TAM isomers were isolated along with O-glucuronides from a reaction mixture consisting of trans- or cis-4-HO-TAM and human liver microsomes fortified with UDP-glucuronic acid and identified with their respective synthetic specimens by high performance liquid chromatography-electrospray ionization time-of-flight mass spectrometry. Although N- and O-glucuronidating activities of human liver microsomes toward trans-4-HO-TAM were nearly comparable, O-glucuronidation was predominant for cis-4-HO-TAM conjugation. Only UGT1A4 catalyzed the N-linked glucuronidation of 4-HO-TAM among recombinant human UGT isoforms (UGT1A1, UGT1A3, UGT1A4, UGT1A6, UGT1A7, UGT1A8, UGT1A9, UGT1A10, UGT2B4, UGT2B7, UGT2B15, and UGT2B17) expressed in insect cells. In contrast, all UGT isoforms, except for UGT1A3 and UGT1A4, catalyzed O-glucuronidation of 4-HO-TAM. Although O-glucuronidation of 4-HO-TAM greatly decreased binding affinity for human ERs, 4-HO-TAM N+-glucuronide still had binding affinity similar to 4-HO-TAM itself, suggesting that N+-glucuronide might contribute to the biological activity of TAM in vivo.

  10. Disruption of thyroid hormone homeostasis in Ugt1a-deficient Gunn rats by microsomal enzyme inducers is not due to enhanced thyroxine glucuronidation

    SciTech Connect

    Richardson, Terrilyn A.; Klaassen, Curtis D.

    2010-10-01

    Microsomal enzyme inducers (MEI) that increase UDP-glucuronosyltransferases (UGTs) are thought to increase glucuronidation of thyroxine (T{sub 4}), thus reducing serum T{sub 4}, and subsequently increasing thyroid stimulating hormone (TSH). Ugt1a1 and Ugt1a6 mediate T{sub 4} glucuronidation. Therefore, this experiment determined the involvement of Ugt1a enzymes in increased T{sub 4} glucuronidation, decreased serum T{sub 4}, and increased TSH after MEI treatment. Male Wistar and Ugt1a-deficient Wistar (Gunn) rats were fed a control diet or diet containing pregnenolone-16{alpha}-carbonitrile (PCN; 800 ppm), 3-methylcholanthrene (3-MC; 200 ppm), or Aroclor 1254 (PCB; 100 ppm) for 7 days. Serum T{sub 4}, triiodothyronine (T{sub 3}), and TSH concentrations, hepatic T{sub 4}/T{sub 3} glucuronidation, and thyroid histology and follicular cell proliferation were investigated. PCN, 3-MC, and PCB treatments decreased serum T{sub 4}, whereas serum T{sub 3} was maintained in both Gunn and Wistar rats (except for PCB treatment). TSH was increased in Wistar and Gunn rats after PCN (130 and 277%) or PCB treatment (72 and 60%). T{sub 4} glucuronidation in Wistar rats was increased after PCN (298%), 3-MC (85%), and PCB (450%), but was extremely low in Gunn rats, and unchanged after MEI. T{sub 3} glucuronidation was increased after PCN (121%) or PCB (58%) in Wistar rats, but only PCN increased T{sub 3} glucuronidation in Gunn rats (43%). PCN treatment induced thyroid morphological changes and increased follicular cell proliferation in both strains. These data demonstrate that T{sub 4} glucuronidation cannot be increased in Ugt1a-deficient Gunn rats. Thus, the decrease in serum T{sub 4}, increase in TSH, and increase in thyroid cell proliferation after MEI are not dependent on increased T{sub 4} glucuronidation, and cannot be attributed to Ugt1a enzymes.

  11. Determination of glucuronide conjugates of hydroxyl triphenyl phosphate (OH-TPHP) metabolites in human urine and its use as a biomarker of TPHP exposure.

    PubMed

    Su, Guanyong; Letcher, Robert J; Yu, Hongxia; Gooden, David M; Stapleton, Heather M

    2016-04-01

    In vitro studies using avian hepatocytes or human liver microsomes suggest that hydroxylation is an important pathway in the metabolism of triphenyl phosphate (TPHP), a chemical used as a flame retardant and plasticizer. TPHP metabolism can lead to the formation of para(p)- and meta(m)-hydroxyl-(OH-)TPHP products as well as their glucuronide conjugates. To determine whether the TPHP hydroxylation and depuration pathway also occurs in vivo in humans, the present study developed a sensitive method for quantification of p- and m-OH-TPHP glucuronides in human urine samples. In n = 1 pooled urine sample and n = 12 individual urine samples collected from four human volunteers from Ottawa (ON, Canada), p- and m-OH-TPHP glucuronides were detectable in 13 and 9 of the 13 analyzed samples and at concentrations ranging from glucuronide and diphenyl phosphate concentrations (DPHP, a known dealkylated metabolite of TPHP). To our knowledge, this is the first report demonstrating that TPHP hydroxylation and conjugation occurs in vivo in humans, and further suggests that p-OH-TPHP glucuronide can be used as a specific biomarker of TPHP exposure in humans.

  12. Species- and gender-dependent differences in the glucuronidation of a flavonoid glucoside and its aglycone determined using expressed UGT enzymes and microsomes.

    PubMed

    Dai, Peimin; Luo, Feifei; Wang, Ying; Jiang, Huangyu; Wang, Liping; Zhang, Guiyu; Zhu, Lijun; Hu, Ming; Wang, Xinchun; Lu, Linlin; Liu, Zhongqiu

    2015-12-01

    Flavonoids occur naturally as glucosides and aglycones. Their common phenolic hydroxyl groups may trigger extensive UDP-glucuronosyltransferase (UGT)- catalysed metabolism. Unlike aglycones, glucosides contain glucose moieties. However, the influence of these glucose moieties on glucuronidation of glucosides and aglycones remains unclear. In this study, the flavonoid glucoside tilianin and its aglycone acacetin were used as model compounds. The glucuronidation characteristics and enzyme kinetics of tilianin and acacetin were compared using human UGT isoforms, liver microsomes and intestinal microsomes obtained from different animal species. Tilianin and acacetin were metabolized into different glucuronides, with UGT1A8 produced as the main isoform. Assessment of enzyme kinetics in UGT1A8, human liver microsomes and human intestinal microsomes revealed that compared with tilianin, acacetin displayed lower Km (0.6-, 0.7- and 0.6-fold, respectively), higher Vmax (20-, 60- and 230-fold, respectively) and higher clearance (30-, 80- and 300-fold, respectively). Furthermore, glucuronidation of acacetin and tilianin showed significant species- and gender-dependent differences. In conclusion, glucuronidation of flavonoid aglycones is faster than that of glucosides in the intestine and the liver. Understanding the metabolism and species- and gender-dependent differences between glucosides and aglycones is crucial for the development of drugs from flavonoids.

  13. Quercetin-3-O-glucuronide promotes the proliferation and migration of neural stem cells.

    PubMed

    Baral, Samrat; Pariyar, Ramesh; Kim, Jaehyo; Lee, Ho-Sub; Seo, Jungwon

    2017-04-01

    Quercetin is a bioactive compound exerting therapeutic effects on in vivo animal models of neurodegeneration or neurotoxicity. However, the narrow therapeutic dose-range of quercetin has been a point of concern since previous studies have demonstrated that quercetin induces cytotoxicity in vitro. Quercetin is metabolized to quercetin glucuronates such as quercetin-3-O-glucuronide (Q3GA), primarily detected in the plasma and the brain. Here, we examined whether and how quercetin or Q3GA regulates neural stem cells (NSCs) in vivo and in vitro. Immunohistochemistry showed that oral administration of quercetin increased nestin-, DCX-, BrdU/DCX-, and BrdU/NeuN-positive cells in the dentate gyrus of mice. However, quercetin decreased the viability of human embryonic NSCs in culture, accompanied by decreased Akt phosphorylation and increased cleavage of caspase-3 and PARP. In contrast, Q3GA increased BrdU-positive cell proliferation, Akt phosphorylation, and cyclin D1 expression. PI3K/Akt inhibitor LY294002 reversed Q3GA-induced Akt phosphorylation and cyclin D1 expression, thereby reducing Q3GA-induced proliferation. Furthermore, Q3GA increased the protein secretion of BDNF and its blockade using anti-BDNF antibody reversed Q3GA-induced proliferation. Under differentiation state, Q3GA promotes NSC migration, along with increased mRNA expression of CXCR4. Moreover, Q3GA significantly reversed scopolamine-induced reduction of Akt phosphorylation in the mouse hippocampus and ameliorated scopolamine-induced memory impairments. Our results demonstrate that quercetin and its metabolite Q3GA control NSC viability in a converse manner through contrary regulation of Akt, accounting for the conflicting effects of quercetin in vivo and in vitro. This study provides a novel mechanism for the positive effects of Q3GA on neurogenesis and suggests its therapeutic potential in neurodegenerative diseases.

  14. KEY COMPARISON: Final report on CCQM-K69 key comparison: Testosterone glucuronide in human urine

    NASA Astrophysics Data System (ADS)

    Liu, Fong-Ha; Mackay, Lindsey; Murby, John

    2010-01-01

    The CCQM-K69 key comparison of testosterone glucuronide in human urine was organized under the auspices of the CCQM Organic Analysis Working Group (OAWG). The National Measurement Institute Australia (NMIA) acted as the coordinating laboratory for the comparison. The samples distributed for the key comparison were prepared at NMIA with funding from the World Anti-Doping Agency (WADA). WADA granted the approval for this material to be used for the intercomparison provided the distribution and handling of the material were strictly controlled. Three national metrology institutes (NMIs)/designated institutes (DIs) developed reference methods and submitted data for the key comparison along with two other laboratories who participated in the parallel pilot study. A good selection of analytical methods and sample workup procedures was displayed in the results submitted considering the complexities of the matrix involved. The comparability of measurement results was successfully demonstrated by the participating NMIs. Only the key comparison data were used to estimate the key comparison reference value (KCRV), using the arithmetic mean approach. The reported expanded uncertainties for results ranged from 3.7% to 6.7% at the 95% level of confidence and all results agreed within the expanded uncertainty of the KCRV. Main text. To reach the main text of this paper, click on Final Report. Note that this text is that which appears in Appendix B of the BIPM key comparison database kcdb.bipm.org/. The final report has been peer-reviewed and approved for publication by the CCQM, according to the provisions of the CIPM Mutual Recognition Arrangement (MRA).

  15. Bisphenol A glucuronide deconjugation is a determining factor of fetal exposure to bisphenol A.

    PubMed

    Gauderat, Glenn; Picard-Hagen, Nicole; Toutain, Pierre-Louis; Corbel, Tanguy; Viguié, Catherine; Puel, Sylvie; Lacroix, Marlène Z; Mindeguia, Pierre; Bousquet-Melou, Alain; Gayrard, Véronique

    2016-01-01

    Previous studies in experimental animals have shown that maternal exposure to bisphenol A (BPA) during late pregnancy leads to high plasma concentrations of BPA glucuronide (BPAG) in fetus compared to mother due to the inability of BPAG to cross the placental barrier. A recent in vitro study has reported that BPAG can exert adipogenic effect underlining the need for characterization of the fetal disposition of BPAG. Experiments were conducted in chronically catheterized fetal sheep to determine the contribution of BPAG hydrolysis to BPA to the elimination of BPAG from the fetal compartment and its resulting effect on the overall fetal exposure to free BPA. Serial sampling of fetal arterial blood, amniotic fluid, maternal venous blood and urine was performed following separate single doses of BPA and BPAG administered intravenously to eight fetal/maternal pairs after cesarean section, and repeated BPAG doses given to two fetal sheep. On average 67% of the BPA entering the fetal circulation was rapidly eliminated through fetal to maternal clearance, with a very short half-life (20 min), while the remaining fraction (24%) was glucuronoconjugated. BPA conjugation-deconjugation cycling was responsible for a 43% increase of the overall fetal exposure to free BPA. A very specific pattern of fetal exposure to free BPA was observed due to its highly increased persistence with a hydrolysis-dependent plasma terminal free BPA half-life of several tens of hours. These findings suggest that although the high fetal to maternal clearance of free BPA protects the fetus from transient increases in free BPA plasma concentrations associated with maternal BPA intake, low but sustained basal free BPA concentrations are maintained in the fetus through BPA conjugation-deconjugation cycling. The potential health implications of these low but sustained basal concentrations of free BPA in fetal plasma should be addressed especially when considering time-dependent effects.

  16. [Concentration of cotinine and trans-3'-hydroxycotinine and its glucuronides in urine of smoking pregnant women].

    PubMed

    Kedziora, Hanna; Florek, Ewa; Piekoszewski, Wojciech; Breborowicz, Grzegorz H; Anholcer, Andrzej; Gomółka, Ewa; Kulza, Maksymilian; Stanek, Artur

    2007-01-01

    To assess the exposure to tobacco smoke constituents, the biomarkers are used. In most studies conducted among active smokers, the cotinine is usually utilised as such a biomarker. The aim of this study was to estimate an application of determining two nicotine metabolites, cotinine and trans-3'-hydroxycotinine (3-HC) in order to assess cigarette smoking among pregnant women. There were 25 patients (15 smokers, 10 non-smokers) admitted to Delivery Ward in Gynecology and Obstetrics Clinical Hospital of the University of Medical Sciences in Poznan. The free and total form of cotinine and trans-3'-hydroxycotinine in urine were determined by the means of high performance liquid chromatography with spectrometry detection. The results of this studies indicated that in urine both cotinine and trans-3'-hydroxycotinine are conjugated with glucoronide acid in the high degree. The linear correlation between free and conjugated form of cotinine and whole cotinine concentration indicates the possibility of application of all three of them as biomarkers of tobacco smoke exposure. The high conjugation of trans-3'-hydroxycotinine with glucuronic acid (over 80 %), and high correlation between glucuronide and total trans-3'-hydroxycotinine concentration proves necessity of having the urine samples hydrolysed or determining the 3'-hydroxycotinine glucoronide, when application of 3-HC as a biomarker of tobacco smoke exposure is concerned. Our studies confirm other authors' observations that division of 3'-hydroxycotinine and cotinine concentrations might be used as a predictor of differences in cotinine metabolite ratio (polimorphism). The studies shown that using analytical equipment, nowadays available in the most laboratories (HPLC), it is possible to determined two biomarkers which are very useful and give various information, no matter whether they are conjugated with glucuronic acid, or not.

  17. Morphine-6-glucuronide: analgesic effects and receptor binding profile in rats

    SciTech Connect

    Abbott, F.V.; Palmour, R.M.

    1988-01-01

    The antinociceptive effects of morphine-6-glucuronide (M6G) were examined in two animal models of pain, the tail immersion test (reflex withdrawal to noxious heat) and the formalin test (behavioral response to minor tissue injury). In the tail immersion test, M6G produced and increase in withdrawal latency that rose rapidly between 0.01 and 0.025 ug ICV or 1 and 2 mg/kg SC. A further increase occurred at doses greater than 0.2 ug ICV or 4 mg/kg SC and was associated with marked catelepsy and cyanosis. Naloxone, 0.1 mg/kg SC, shifted the lower component of the dose-effect relation by a factor of 24. In the formalin test, 0.01 ug M6G ICV produced hyperalgesia, while between 0.05 and 0.2 ug ICV, antinociception increased rapidly without toxicity. The dose effect relations for hyperalgesia and antinociception were shifted to the right by factors of 20- and 3-fold, respectively. By comparison, ICV morphine was 60 (formalin test) to 145-200 (tail immersion test) times less potent than M6G. At sub-nanomolar concentrations, M6G enhanced the binding of (/sup 3/H)-etorphine, (/sup 3/H)-dihydromorphine and (/sup 3/H)-naloxone to rat brain membrane receptors by 20-40%. At higher concentrations, M6G displaced each ligand from binding sites, with K/sub i/ values of about 30 nM, as compared to morphine K/sub i/ values of about 3 nM.

  18. Postmortem redistribution of the heroin metabolites morphine and morphine-3-glucuronide in rabbits over 24 h.

    PubMed

    Maskell, Peter D; Albeishy, Mohammed; De Paoli, Giorgia; Wilson, Nathan E; Seetohul, L Nitin

    2016-03-01

    The interpretation of postmortem drug levels is complicated by changes in drug blood levels in the postmortem period, a phenomena known as postmortem drug redistribution. We investigated the postmortem redistribution of the heroin metabolites morphine and morphine-3-glucuronide in a rabbit model. Heroin (1 mg/kg) was injected into anesthetised rabbit; after 1 h, an auricular vein blood sample was taken and the rabbit was euthanised. Following death rabbits were placed in a supine position at room temperature and divided into three groups namely (1) immediate autopsy, (2) autopsy after 30 minutes and (3) autopsy 24 h after death. Various samples which included femoral blood, cardiac blood, lung, liver, kidney, vitreous humour, subcutaneous and abdominal fat, liver, bone marrow and skeletal muscle were taken. The samples were analysed with a validated LC-MS/MS method. It was observed that within minutes there was a significant increase in free morphine postmortem femoral blood concentration compared to the antemortem sample (0.01 ± 0.01 to 0.05 ± 0.02 mg/L).Various other changes in free morphine and metabolite concentrations were observed during the course of the experiment in various tissues. Principal component analysis was used to investigate possible correlations between free morphine in the various samples. Some correlations were observed but gave poor predictions (>20 % error) when back calculating. The results suggest that rabbits are a good model for further studies of postmortem redistribution but that further study and understanding of the phenomena is required before accurate predictions of the blood concentration at the time of death are possible.

  19. Elucidation of the Mechanisms through Which the Reactive Metabolite Diclofenac Acyl Glucuronide Can Mediate Toxicity

    PubMed Central

    Scialis, Renato J.

    2016-01-01

    We have previously reported that mice lacking the efflux transporter Mrp3 had significant intestinal injury after toxic diclofenac (DCF) challenge, and proposed that diclofenac acyl glucuronide (DCF-AG), as a substrate of Mrp3, played a part in mediating injury. Since both humans and mice express the uptake transporter OATP2B1 in the intestines, OATP2B1 was characterized for DCF-AG uptake. In vitro assays using human embryonic kidney (HEK)-OATP2B1 cells demonstrated that DCF-AG was a substrate with a maximal velocity (Vmax) and Km of 17.6 ± 1.5 pmol/min per milligram and 14.3 ± 0.1 μM, respectively. Another key finding from our in vitro assays was that DCF-AG was more cytotoxic compared with DCF, and toxicity occurred within 1–3 hours of exposure. We also report that 1 mM DCF-AG caused a 6-fold increase in reactive oxygen species (ROS) by 3 hours. Investigation of oxidative stress through inhibition of superoxide dismutase (SOD) revealed that DCF-AG had 100% inhibition of SOD at the highest tested dose of 1 mM. The SOD and ROS results strongly suggest DCF-AG induced oxidative stress in vitro. Lastly, DCF-AG was screened for pharmacologic activity against COX-1 and COX-2 and was found to have IC50 values of 0.620 ± 0.105 and 2.91 ± 0.36 μM, respectively, which represents a novel finding. Since cyclooxygenase (COX) inhibition can lead to intestinal ulceration, it is plausible that DCF-AG can also contribute to enteropathy via COX inhibition. Taken in context, the work presented herein demonstrated the multifactorial pathways by which DCF-AG can act as a direct contributor to toxicity following DCF administration. PMID:26869668

  20. Weeding the Astrophysical Garden: Ethyl Cyanide

    NASA Astrophysics Data System (ADS)

    De Lucia, F. C.; Fortman, S. M.; Medvedev, I. R.; Neese, C. F.

    2009-12-01

    It is well known that many, if not most, of the unidentified features in astrophysical spectra arise from relatively low lying excited vibrational and torsional states of a relatively small number of molecular species— the astrophysical weeds. It is also well known that the traditional quantum mechanical assignment and fitting of these excited state spectra is a formidable task, one that is made harder by the expected perturbations and interactions among these states. We have previously proposed an alternative fitting and analysis approach based on experimental, intensity calibrated spectra taken at many temperatures. In this paper we discuss the implementation of this approach and provide details in the context of one of these weeds, ethyl cyanide.

  1. Cognitive effects of creatine ethyl ester supplementation.

    PubMed

    Ling, Jonathan; Kritikos, Minos; Tiplady, Brian

    2009-12-01

    Supplementation with creatine-based substances as a means of enhancing athletic performance has become widespread. Until recently, however, the effects of creatine supplementation on cognitive performance has been given little attention. This study used a new form of creatine--creatine ethyl ester--to investigate whether supplementation would improve performance in five cognitive tasks, using a double-blind, placebo-controlled study. Creatine dosing led to an improvement over the placebo condition on several measures. Although creatine seems to facilitate cognition on some tasks, these results require replication using objective measures of compliance. The improvement is discussed in the context of research examining the influence of brain energy capacity on cognitive performance.

  2. The gelation of oil using ethyl cellulose.

    PubMed

    Davidovich-Pinhas, M; Barbut, S; Marangoni, A G

    2015-03-06

    The characterization of the thermo-gelation mechanism and properties of ethyl cellulose/canola oil oleogels was performed using rheology and thermal analysis. Thermal analysis detected no evidence for thermal transitions contributed to secondary conformational changes, suggesting a gelation mechanism that does not involve secondary ordered structure formation. Rheological analysis demonstrated a relationship between the polymer molecular weight and the final gel strength, the cross-over behavior as well as the gel point temperature. Increasing polymer molecular weight led to an increase in final gel strength, the modulus at cross-over, and the gel point temperature. Cooling/heating rates affect gel modulus only for the low molecular weight samples. A decrease in gel strength with increasing cooling rate was detected. The cross-over temperature was not affected by the cooling/heating rates. Cooling rate also affected the gelation setting time where slow cooling rates produced a stable gel faster.

  3. IRIS TOXICOLOGICAL REVIEW OF METHYL ETHYL KETONE (2003 Final)

    EPA Science Inventory

    EPA is announcing the release of the final report, "Toxicological Review of Methyl Ethyl Ketone: in support of the Integrated Risk Information System (IRIS)". The updated Summary for Methyl Ethyl Ketone and accompanying Quickview have also been added to the IRIS Database.

  4. IRIS Toxicological Review of Methyl Ethyl Ketone (2003 Final)

    EPA Science Inventory

    EPA announced the release of the final report, Toxicological Review of Methyl Ethyl Ketone: in support of the Integrated Risk Information System (IRIS). The updated Summary for Methyl Ethyl Ketone and accompanying toxicological review have been added to the IRIS Database....

  5. Evaluation of 3,3′,4′-trihydroxyflavone and 3,6,4′-trihydroxyflavone (4′-O-glucuronidation) as the in vitro functional markers for hepatic UGT1A1

    PubMed Central

    Wu, Baojian; Zhang, Shuxing; Hu, Ming

    2012-01-01

    Identifying UDP-glucuronosyltransferase (UGT)-selective probes (substrates that are primarily glucuronidated by a single isoform) is complicated by the enzymes' large overlapping substrate specificity. Here, regioselective glucuronidation of two flavonoids 3,3′,4′-trihydroxyflavone (33′4′THF) and 3,6,4′-trihydroxyflavone (364′THF) is used to probe the activities of hepatic UGT1A1. The glucuronidation kinetics of 33′4′THF and 364′THF was determined using 12 recombinant human UGT isoforms and pooled human liver microsomes (pHLM). The individual contribution of main UGT isoforms to the metabolism of the two flavonoids in pHLM was estimated using the relative activity factor approach. UGT1A1 activity correlation analyses using flavonoids-4′-O-glucuronidation vs. β-estradiol-3-glucuronidation (a well-recognized marker for UGT1A1) or vs. SN-38 glucuronidation were performed using a bank of HLMs (n=12) including three UGT1A1-genotyped HLMs (i.e., UGT1A1*1*1, UGT1A1*1*28 and UGT1A1*28*28). The results showed that UGT1A1 and 1A9, followed by 1A7, were the main isoforms for glucuronidating the two flavonoids, where UGT1A1 accounted for 92 ± 7 % and 91 ± 10 % of 4′-O-glucuronidation of 33′4′THF and 364′THF, respectively, and UGT1A9 accounted for most of the 3-O-glucuronidation. Highly significant correlations (R2 > 0.944, p < 0.0001) between the rates of flavonoids 4′-O-glucuronidation and that of estradiol-3-glucuronidation or SN-38 glucuronidation were observed across 12 HLMs. In conclusion, UGT1A1-mediated 4′-O-glucuronidation of 33′4′THF and 364′THF were highly correlated with glucuronidation of estradiol (3-OH) and SN-38. This study demonstrated for the first time that regioselective glucuronidation of flavonoids can be applied to probe hepatic UGT1A1 activity in vitro. PMID:21985641

  6. Photoisomerization of ethyl ferulate: A solution phase transient absorption study

    NASA Astrophysics Data System (ADS)

    Horbury, Michael D.; Baker, Lewis A.; Rodrigues, Natércia D. N.; Quan, Wen-Dong; Stavros, Vasilios G.

    2017-04-01

    Ethyl ferulate (ethyl 4-hydroxy-3-methoxycinnamate) is currently used as a sunscreening agent in commercial sunscreen blends. Recent time-resolved gas-phase measurements have demonstrated that it possesses long-lived (>ns) electronic excited states, counterintuitive to what one might anticipate for an effective sunscreening agent. In the present work, the photodynamics of ethyl ferulate in cyclohexane, are explored using time-resolved transient electronic absorption spectroscopy, upon photoexcitation to the 11ππ∗ and 21ππ∗ states. We demonstrate that ethyl ferulate undergoes efficient non-radiative decay to repopulate the electronic ground state, mediated by trans-cis isomerization. These results strongly suggest that even mild perturbations induced by a non-polar solvent, as may be found in a closer-to-market sunscreen blend, may contribute to our understanding of ethyl ferulate's role as a sunscreening agent.

  7. [Effect of dimethicone on pharmacokinetics and pharmacodynamics of ethyl biscoumacetate].

    PubMed

    Copie, X; Pinquier, J L; Letrait, M; Paltiat, M H; Pello, J Y; Rey, E; Chanteclair, G; de Lauture, D; Olive, G; Strauch, G

    1993-01-01

    The influence of dimeticone (Gel de Polysilane Midy) on the pharmacokinetics and pharmacodynamics of oral ethyl biscoumacetate was studied in 6 healthy volunteers in a randomised single dose, two-way cross-over study. Each volunteer received at one week interval a single dose (300 mg) of ethyl biscoumacetate, either alone or with dimeticone. Ethyl biscoumacetate levels were measured in plasma for 24 hours. Pharmacodynamic parameters were measured for 96 hours. Ethyl biscoumacetate peak concentration was significantly higher when administered with dimeticone (40.3 +/- 25.3 mg/l vs 31.0 +/- 25.7 mg/l; p = 0.031), without significant change in the area under curve. Other pharmacokinetic and pharmacodynamic parameters did not differ significantly. The slight increase of the ethyl biscoumacetate bioavailability with dimeticone in repeated dosing might have pharmacodynamic consequence; a clinical trial should address this question.

  8. Spectroscopy reveals that ethyl esters interact with proteins in wine.

    PubMed

    Di Gaspero, Mattia; Ruzza, Paolo; Hussain, Rohanah; Vincenzi, Simone; Biondi, Barbara; Gazzola, Diana; Siligardi, Giuliano; Curioni, Andrea

    2017-02-15

    Impairment of wine aroma after vinification is frequently associated to bentonite treatments and this can be the result of protein removal, as recently demonstrated for ethyl esters. To evaluate the existence of an interaction between wine proteins and ethyl esters, the effects induced by these fermentative aroma compounds on the secondary structure and stability of VVTL1, a Thaumatin-like protein purified from wine, was analyzed by Synchrotron Radiation Circular Dichroism (SRCD) spectroscopy. The secondary structure of wine VVTL1 was not strongly affected by the presence of selected ethyl esters. In contrast, VVTL1 stability was slightly increased by the addition of ethyl-octanoate, -decanoate and -dodecanoate, but decreased by ethyl-hexanoate. This indicates the existence of an interaction between VVTL1 and at least some aroma compounds produced during fermentation. The data suggest that proteins removal from wine by bentonite can result in indirect removal of at least some aroma compounds associated with them.

  9. Parameters Affecting Ethyl Ester Production by Saccharomyces cerevisiae during Fermentation▿

    PubMed Central

    Saerens, S. M. G.; Delvaux, F.; Verstrepen, K. J.; Van Dijck, P.; Thevelein, J. M.; Delvaux, F. R.

    2008-01-01

    Volatile esters are responsible for the fruity character of fermented beverages and thus constitute a vital group of aromatic compounds in beer and wine. Many fermentation parameters are known to affect volatile ester production. In order to obtain insight into the production of ethyl esters during fermentation, we investigated the influence of several fermentation variables. A higher level of unsaturated fatty acids in the fermentation medium resulted in a general decrease in ethyl ester production. On the other hand, a higher fermentation temperature resulted in greater ethyl octanoate and decanoate production, while a higher carbon or nitrogen content of the fermentation medium resulted in only moderate changes in ethyl ester production. Analysis of the expression of the ethyl ester biosynthesis genes EEB1 and EHT1 after addition of medium-chain fatty acid precursors suggested that the expression level is not the limiting factor for ethyl ester production, as opposed to acetate ester production. Together with the previous demonstration that provision of medium-chain fatty acids, which are the substrates for ethyl ester formation, to the fermentation medium causes a strong increase in the formation of the corresponding ethyl esters, this result further supports the hypothesis that precursor availability has an important role in ethyl ester production. We concluded that, at least in our fermentation conditions and with our yeast strain, the fatty acid precursor level rather than the activity of the biosynthetic enzymes is the major limiting factor for ethyl ester production. The expression level and activity of the fatty acid biosynthetic enzymes therefore appear to be prime targets for flavor modification by alteration of process parameters or through strain selection. PMID:17993562

  10. High-performance liquid chromatographic assay for N-glucuronidation of nicotine and cotinine in human liver microsomes.

    PubMed

    Nakajima, Miki; Kwon, Jun-Tack; Tanaka, Eriko; Yokoi, Tsuyoshi

    2002-03-01

    A method for the determination of N-glucuronidation of nicotine and cotinine in human liver microsomes by high-performance liquid chromatography was developed. Nicotine or cotinine was incubated with human liver microsomes and UDP-glucuronic acid in a 200-microl incubation mixture. The nicotine N-glucuronide (Nic-glu) and cotinine N-glucuronide (Cot-glu) formed were analyzed by ion-pair chromatography with a C-18 column. The sensitivity of quantification at 260 nm absorption was improved by using a noise-base clean Uni-3, and the limit of quantification was 10 pmol/200 microl mixture for both Nic-glu and Cot-glu. Linear standard curves were obtained within the concentration ranges 25-1000 pmol/200 microl mixture for Nic-glu and 100-5000 pmol/200 microl mixture for Cot-glu. The intraassay precision and accuracy were < or =11.1% coefficient of variation (CV) and 97.5-106.6% for Nic-glu and < or =4.6% CV and 96.7-100.4% for Cot-glu. The interassay precision and accuracy were < or =7.2% CV and 98.2-106.1% for Nic-glu and < or =4.6% CV and 96.8-99.3% for Cot-glu. This is the first report of the in vitro determination of Nic-glu and Cot-glu in human liver microsomes. Furthermore, this highly sensitive HPLC method can be used for the determination of Nic-glu and Cot-glu in biological specimens in vivo.

  11. Simultaneous determination of mycophenolic acid and its glucuronides in human plasma using isocratic ion pair high-performance liquid chromatography.

    PubMed

    Mino, Yasuaki; Naito, Takafumi; Matsushita, Tomomi; Kagawa, Yoshiyuki; Kawakami, Junichi

    2008-02-13

    Therapeutic drug monitoring (TDM) of mycophenolic acid (MPA) following administration of mycophenolate mofetil (MMF) or the enteric-coated sodium salt of MPA formulations, seems beneficial because of the large intra- and inter-individual variability in MPA pharmacokinetics. MPA is an active component from these oral formulations and are further metabolised to inactive phenolic glucuronide (MPAG) and active acyl glucuronide (AcMPAG). This study aims to determine simultaneously these three metabolites of MMF using isocratic ion pair HPLC and to evaluate the short-term stability of AcMPAG in human plasma. Samples were prepared using solid phase extraction. Chromatographic separation was achieved over an RP column (TSKgel ODS-80Ts, 150 mm x 4.6 mm i.d., 5 microm particle size) with acetonitrile and 30 mM tetra-n-butylammonium bromide containing 5 mM ammonium acetate at pH 9.0 (33/67, v/v) as the mobile phase. The flow rate of the mobile phase was 1ml/min, and the wavelength of determination by UV detection was 250 nm (run time, 16 min). Calibration curves for MPA, MPAG and AcMPAG in human plasma were linear over a concentration range of 0.05-50, 0.1-400 and 0.08-8 microg/ml, respectively. Intra- and inter-assay R.S.D. were<6.5%. Extraction efficiencies were more than 85% for all analytes. Since AcMPAG was unstable in human plasma, plasma acidification was needed for the quantification of AcMPAG. Large interindividual variability was observed in the AcMPAG pharmacokinetics in the early period after renal transplantation. In conclusion, a simple, accurate and reproducible HPLC method to measure simultaneously these three MMF metabolites has been established. The method will be helpful in evaluating pharmacokinetics of MPA and its glucuronides.

  12. Alcohol Consumption during Pregnancy: Analysis of Two Direct Metabolites of Ethanol in Meconium

    PubMed Central

    Sanvisens, Arantza; Robert, Neus; Hernández, José María; Zuluaga, Paola; Farré, Magí; Coroleu, Wifredo; Serra, Montserrat; Tor, Jordi; Muga, Robert

    2016-01-01

    Alcohol consumption in young women is a widespread habit that may continue during pregnancy and induce alterations in the fetus. We aimed to characterize prevalence of alcohol consumption in parturient women and to assess fetal ethanol exposure in their newborns by analyzing two direct metabolites of ethanol in meconium. This is a cross-sectional study performed in September 2011 and March 2012 in a series of women admitted to an obstetric unit following childbirth. During admission, socio-demographic and substance use (alcohol, tobacco, cannabis, cocaine, and opiates) during pregnancy were assessed using a structured questionnaire and clinical charts. We also recorded the characteristics of pregnancy, childbirth, and neonates. The meconium analysis was performed by liquid chromatography—tandem mass spectrometry (LC-MS/MS) to detect the presence of ethyl glucuronide (EtG) and ethyl sulfate (EtS). Fifty-one parturient and 52 neonates were included and 48 meconium samples were suitable for EtG and EtS detection. The median age of women was 30 years (interquartile range (IQR): 26–34 years); EtG was present in all meconium samples and median concentration of EtG was 67.9 ng/g (IQR: 36.0–110.6 ng/g). With respect to EtS, it was undetectable (<0.01 ng/g) in the majority of samples (79.1%). Only three (6%) women reported alcohol consumption during pregnancy in face-to-face interviews. However, prevalence of fetal exposure to alcohol through the detection of EtG and EtS was 4.2% and 16.7%, respectively. Prevention of alcohol consumption during pregnancy and the detection of substance use with markers of fetal exposure are essential components of maternal and child health. PMID:27011168

  13. Impact of hair-care products on FAEE hair concentrations in substance abuse monitoring.

    PubMed

    Gareri, Joey; Appenzeller, Brice; Walasek, Paula; Koren, Gideon

    2011-04-01

    Previous studies have indicated that the use of high-ethanol-content (>65%) hair-care products may elevate fatty acid ethyl ester (FAEE) concentrations in hair. In this case series, nine individuals were identified by FAEE analysis to be chronic alcohol abusers in the context of child-welfare substance abuse monitoring. Based on patient claims of moderate or no alcohol consumption, the presence of ethanol in the patients' hair-care regimens was investigated. Samples were additionally tested for the presence of ethyl glucuronide (EtG). From a total of nine patients, 12 hair samples were submitted for analysis. Patient histories were obtained as well as Material Safety Data Sheets (MSDS) listing hair-care product ethanol content. Hair samples were pre-washed to remove external contamination and analyzed for FAEE and EtG by GC-MS. According to the Society of Hair Testing consensus guidelines, FAEE levels exceeding 0.50 ng/mg and/or EtG levels exceeding 30 pg/mg indicate chronic excessive alcohol consumption. Upon initial analysis, the nine samples exhibited positive FAEE findings ranging from 0.496 to 4.984 ng/mg. MSDS review revealed the presence of ethanol from 10% to 95% by volume in at least one hair-care product used by each individual. Results of the EtG analysis ranged from 1.9 to 23.5 pg/mg. These findings indicate that regular use of products with ethanol content as low as 10% can impact FAEE results. EtG analysis should be used to confirm FAEE findings and appears to be unaffected by hair-care products, likely due to alternative mechanisms of incorporation.

  14. Alcohol Consumption during Pregnancy: Analysis of Two Direct Metabolites of Ethanol in Meconium.

    PubMed

    Sanvisens, Arantza; Robert, Neus; Hernández, José María; Zuluaga, Paola; Farré, Magí; Coroleu, Wifredo; Serra, Montserrat; Tor, Jordi; Muga, Robert

    2016-03-22

    Alcohol consumption in young women is a widespread habit that may continue during pregnancy and induce alterations in the fetus. We aimed to characterize prevalence of alcohol consumption in parturient women and to assess fetal ethanol exposure in their newborns by analyzing two direct metabolites of ethanol in meconium. This is a cross-sectional study performed in September 2011 and March 2012 in a series of women admitted to an obstetric unit following childbirth. During admission, socio-demographic and substance use (alcohol, tobacco, cannabis, cocaine, and opiates) during pregnancy were assessed using a structured questionnaire and clinical charts. We also recorded the characteristics of pregnancy, childbirth, and neonates. The meconium analysis was performed by liquid chromatography-tandem mass spectrometry (LC-MS/MS) to detect the presence of ethyl glucuronide (EtG) and ethyl sulfate (EtS). Fifty-one parturient and 52 neonates were included and 48 meconium samples were suitable for EtG and EtS detection. The median age of women was 30 years (interquartile range (IQR): 26-34 years); EtG was present in all meconium samples and median concentration of EtG was 67.9 ng/g (IQR: 36.0-110.6 ng/g). With respect to EtS, it was undetectable (<0.01 ng/g) in the majority of samples (79.1%). Only three (6%) women reported alcohol consumption during pregnancy in face-to-face interviews. However, prevalence of fetal exposure to alcohol through the detection of EtG and EtS was 4.2% and 16.7%, respectively. Prevention of alcohol consumption during pregnancy and the detection of substance use with markers of fetal exposure are essential components of maternal and child health.

  15. Determination of sulfates and glucuronides of endogenic steroids in biofluids by high-performance liquid chromatography/orbitrap mass spectrometry

    NASA Astrophysics Data System (ADS)

    Semenistaya, E. N.; Virus, E. D.; Rodchenkov, G. M.

    2009-04-01

    the possibility of selective determination of testosterone and epitestosterone glucuronides in urine by high-performance liquid chromatography/high-resolution mass spectrometry using solid phase microextraction on a meps cartridge was studied. the effect of the biological matrix on the spectra of conjugated steroids can be taken into account by using the spectra of conjugates recorded for urine samples after hydrolysis as reference spectra. the conditions of fragmentation in the ion source were optimized for separate analytes. this method was used for analyzing real samples with different testosterone/epitestosterone ratios. variations in conjugate contents and qualitative changes in the steroid profile of endogenic compounds were observed.

  16. Human UDP-Glucuronosyltransferase 1A1 is the Primary Enzyme Responsible for the N-glucuronidation of N-hydroxy-PhIP in vitro

    SciTech Connect

    Malfatti, M A; Felton, J S

    2004-04-06

    UDP-Glucuronosyltransferase 1A proteins (UGT1A) catalyze the glucuronidation of many endogenous and xenobiotic compounds including heterocyclic amines and their hydroxylated metabolites (the main topic of this study). Studies have shown that in humans UGT1A mediated glucuronidation is an important pathway in the detoxification of food-borne carcinogenic heterocyclic amines. The biotransformation of 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), the most mass abundant heterocyclic amine found in cooked meats, is highly dependent on cytochrome P4501A2 hydroxylation followed by UGT catalyzed glucuronidation of the N-hydroxy-PhIP reactive intermediate. To determine which UGT1A proteins are involved in the glucuronidation of N-hydroxy-PhIP, microsomal preparations from baculovirus infected insect cells that express all of the known functional human UGT1A isozymes (UGT1A1, -1A3, -1A4, -1A6, -1A7, -1A8, -1A9, -1A10) were exposed to N-hydroxy-PhIP and the reaction products were isolated by HPLC. All UGT1A proteins except UGT1A6 showed some degree of activity towards N-hydroxy-PhIP. The formation of both N-hydroxy-PhIP-N{sup 2}-glucuronide and N-hydroxy-PhIP-N3-glucuronide was both time and substrate concentration dependent in all the microsomal incubations that showed appreciable activity. UGT1A1 was the most efficient in converting N-hydroxy-PhIP to both conjugates producing 5 times more of the N{sup 2}-conjugate than UGT1A4, the next active UGT, and 286 times more than UGT1A7, the least active UGT. With an apparent Km of 52 {micro}M and a K{sub cat} of 114 min-1, UGT1A1 was also the most catalytically efficient in forming N-hydroxy-PhIP-N{sup 2}-glucuronide. Catalytic constants for UGT1A4, UGT1A8 and UGT1A9 were 52 min-1, 35 min{sup -1} and 3.7 min{sup -1}, respectively. The catalytic efficiency for N-hydroxy-PhIP-N3-glucuronide formation was 8, 10, and 6 times lower for UGT1A1, -1A4, and -1A8, respectively, when compared to the k{sub cat} values for N

  17. A modern approach to the synthesis of 2-(4-chlorophenyl)[2-(14)C]thiazol-4-ylacetic acid ([(14)C] fenclozic acid) and its acyl glucuronide metabolite.

    PubMed

    Killick, David A; Bushby, Nick

    2013-01-01

    An updated approach to the 1960s synthesis of [(14)C] fenclozic acid from labelled potassium cyanide is presented. By employing modern synthetic methodology and purification techniques, many of the inherent hazards in the original synthesis are avoided or significantly reduced. The concomitant labelled stereoselective synthesis of the key acyl glucuronide metabolite (the 1-β-O-acyl glucuronide) is also described.

  18. Evaluation of pharmaceutical excipients as cosolvents in 4-methyl umbelliferone glucuronidation in human liver microsomes: applications for compounds with low solubility.

    PubMed

    Argikar, Upendra A; Liang, Guiqing; Bushee, Jennifer L; Hosagrahara, Vinayak P; Lee, Wendy

    2011-01-01

    Standard incubation procedures for carrying out microsomal assays involve the use of less than 1% w/v organic solvents to minimize the potential inhibitory effects of organic solvents on metabolic activity. This presents a practical limitation for poorly soluble xenobiotics, which cannot be incubated at concentrations high enough to obtain a V(max), and therefore subsequent values for K(m) and Cl(int) cannot be calculated. Our goal was to study the application of a variety of pharmaceutical excipients to aid the solubilization of compounds in vitro in glucuronidation incubations, without affecting the reaction kinetics. In vitro glucuronidation incubations were carried out in human liver microsomes with 4-methylumbelliferone (4-MU) and the kinetics of 4-MU glucuronidation in the presence of excipients were compared to that in control incubations without any excipients. In addition, IC(75) values were calculated for each excipient. We observed that HPBCD (Hydroxypropyl-β-cyclodextrin) may be employed in in vitro glucuronidation incubations up to 0.5% w/v without affecting the Cl(int) of 4-MU. Although NMP (N-methyl-2-pyrrolidone) and DMA (N,N-dimethylacetamide); showed low IC(75) values approximately 0.1% w/v each, neither excipients altered the Cl(int) of 4-MUG (4-methylumbelliferyl-β-D-glucuronide) formation. Our studies point toward possible applications of pharmaceutical excipients to carry out in vitro glucuronidation of substrates with poor aqueous solubility, in order to estimate Cl(int) and subsequently scaled organ clearance values.

  19. Stereoselective pharmacokinetics of ketoprofen and ketoprofen glucuronide in end-stage renal disease: evidence for a ‘futile cycle’ of elimination

    PubMed Central

    Grubb, N G; Rudy, D W; Brater, D C; Hall, S D

    1999-01-01

    Aims To assess if futile cycling of ketoprofen occurs in patients with decreased renal function. Methods Ketoprofen was administered to six haemodialysis-dependent patients with end-stage renal disease as single (50 mg) or multiple doses (50 mg three times daily, for 7 days). Plasma and dialysate concentrations of the unconjugated and glucuronidated R- and S-enantiomers of ketoprofen were determined using h.p.l.c. following the single and multiple dosing. Results The oral clearance was decreased and terminal elimination half-lives of R- and S-ketoprofen and the corresponding acyl glucuronides were increased in functionally anephric patients compared with healthy subjects. In contrast with the R-isomers, S-ketoprofen and S-ketoprofen glucuronide exhibited an unexpected accumulation (2.7–3.8 fold) after repeated dosing achieving S:R ratios of 3.3±1.7 and 11.2±5.3, respectively. The plasma dialysis clearances for R- and S-ketoprofen glucuronides were 49.4±19.8 and 39.0±15.9 ml min−1, respectively, and 10.8±17.6 and 13.3±23.5 ml min−1 for unconjugated R- and S-ketoprofen. Conclusions The selective accumulation of S-ketoprofen and its acyl glucuronide are consistent with amplification of chiral inversion subsequent to futile cycling between R-ketoprofen and R-ketoprofen glucuronide. Severe renal insufficiency, and possibly more modest decrements, results in a disproportionate increase in systemic exposure to the S-enantiomer which inhibits both pathologic and homeostatic prostaglandin synthesis. PMID:10583018

  20. Identification of anovulation and transient luteal function using a urinary pregnanediol-3-glucuronide ratio algorithm.

    PubMed Central

    Kassam, A; Overstreet, J W; Snow-Harter, C; De Souza, M J; Gold, E B; Lasley, B L

    1996-01-01

    The sensitivity and specificity of a urinary pregnanediol-3-glucuronide (PdG) ratio algorithm to identify anovulatory cycles was studied prospectively in two independent populations of women. Urinary hormone data from the first group was used to develop the algorithm, and data from the second group was used for its validation. PdG ratios were calculated by a cycles method in which daily PdG concentrations indexed by creatinine (CR) from cycle day 11 onward were divided by a baseline PdG (average PdG/Cr concentration for cycle days 6-10). In the interval method, daily PdG/CR concentrations from day 1 onward were divided by baseline PdG (lowest 5-day average of PdG/CR values throughout the collection period). Evaluation of the first study population (n = 6) resulted in cycles with PdG ratios > or = 3 for > or = 3 consecutive days being classified as ovulatory; otherwise they were anovulatory. The sensitivity and specificity of the PdG ratio algorithm to identify anovulatory cycles in the second population were 75% and 89.5%, respectively, for all cycles (n = 88); 50% and 88.3% for first cycles (n = 40) using the cycles method; 75% and 92.2%, respectively, for all cycles (n = 89); and 50% and 94.1% for first cycles (n = 40) using the interval method. The "gold standard" for anovulation was weekly serum samples < or = 2 ng/ml progesterone. The sensitivity values for all cycles and for the first cycle using both methods were underestimated because of apparent misclassification of cycles using serum progesterone due to infrequent blood collection. Blood collection more than once a week would have greatly improved the sensitivity and modestly improved the specificity of the algorithm. The PdG ratio algorithm provides an efficient approach for screening urine samples collected in epidemiologic studies of reproductive health in women. Images Figure 1. A Figure 1. B Figure 1. C Figure 2. A Figure 2. B PMID:8732951

  1. Ethyl Esterification for MALDI-MS Analysis of Protein Glycosylation.

    PubMed

    Reiding, Karli R; Lonardi, Emanuela; Hipgrave Ederveen, Agnes L; Wuhrer, Manfred

    2016-01-01

    Ethyl esterification is a technique for the chemical modification of sialylated glycans, leading to enhanced stability when performing matrix-assisted laser desorption/ionization (MALDI)-mass spectrometry (MS), as well as allowing the efficient detection of both sialylated and non-sialylated glycans in positive ion mode. In addition, the method shows specific reaction products for α2,3- and α2,6-linked sialic acids, leading to an MS distinguishable mass difference. Here, we describe the ethyl esterification protocol for 96 glycan samples, including enzymatic N-glycan release, the aforementioned ethyl esterification, glycan enrichment, MALDI target preparation, and the MS(/MS) measurement.

  2. On the cause of low thermal stability of ethyl halodiazoacetates

    PubMed Central

    Mortén, Magnus; Hennum, Martin

    2016-01-01

    Summary Rates for the thermal decomposition of ethyl halodiazoacetates (halo = Cl, Br, I) have been obtained, and reported herein are their half-lives. The experimental results are supported by DFT calculations, and we provide a possible explanation for the reduced thermal stability of ethyl halodiazoacetates compared to ethyl diazoacetate and for the relative decomposition rates between the chloro, bromo and iodo analogs. We have also briefly studied the thermal, non-catalytic cyclopropanation of styrenes and compared the results to the analogous Rh(II)-catalyzed reactions. PMID:27559411

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

    PubMed Central

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

    2015-01-01

    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. PMID:26255293

  4. Determination of free and glucuronidated kaempferol in rat plasma by LC-MS/MS: application to pharmacokinetic study.

    PubMed

    Zhang, Wei-Dong; Wang, Xiao-Juan; Zhou, Si-Yuan; Gu, Yi; Wang, Rong; Zhang, Tao-Li; Gan, Hong-Quan

    2010-08-01

    Flavanoid kaempferol is mainly present as glucuronides and sulfates in rat plasma, and small amounts of the intact aglycone are also detected. In the this study, a rapid, specific and sensitive liquid chromatography-electrospray ionization-tandem mass spectrometry method (HPLC-MS/MS) was developed and validated for determination of kaempferol and its major metabolite glucuronidated kaempferol in rat plasma. A liquid-liquid extraction with acetic ether was involved for the extraction of kaempferol and internal standard. Analytes were separated on a C18 column (150 mm x 2.1 mm, 4.5 microm, Waters Corp.) with isocratic elution at a flow-rate of 0.3 ml min(-1). The mobile phase was consisted of 0.5% formic acid and acetonitrile (50:50, v/v). The Quattro Premier HPLC-MS/MS was operated under the multiple reaction-monitoring mode (MRM) using the electrospray ionization technique. The method was validated according to the FDA guidelines for validation of bioanalytical method. The validated method was successfully applied to the study of the pharmacokinetics in rats after oral administration of kaempferol with different doses.

  5. Is urine an alternative to cosmetically treated hair for the detection of drugs and alcohol?

    PubMed

    Agius, Ronald; Dufaux, Bertin; Kahl, Hans-Gerhard; Nadulski, Thomas

    2014-06-01

    This study attempts to assess the utility of the urine matrix as an alternative to cosmetically treated hair for the detection of drugs and alcohol for driving licence re-granting in 1026 cosmetically treated hair samples and 33 262 urine routine samples. No significant difference was observed between the percentage positive samples in cosmetically treated hair to those in urine at both the 95% and 99% significance level for amphetamines, cocaine, opiates, benzodiazepines, and methadone. Significant difference was found between the positivity rates of cannabinoids in cosmetically treated hair and that in urine indicating urine to be a better alternative to the use of the hair matrix even when cosmetically treated. The opposite was observed for the alcohol consumption marker ethyl glucuronide (EtG) for which the positivity rate in cosmetically treated hair was twice that in urine samples. Particularly for alcohol abstinence monitoring, as for the rehabilitative driving licence re-granting medical and psychological assessment (MPA) programme in Germany, it seems that ethyl glucuronide (EtG) in hair presents a much better alternative than urine testing, even when cosmetically treated hair is analyzed. Moreover, segmentation is an additional advantage of hair testing which can provide additional useful information.

  6. Ethyl pyruvate: a novel treatment for sepsis.

    PubMed

    Fink, Mitchell P

    2007-01-01

    Ethyl pyruvate (EP), a simple aliphatic ester derived from pyruvic acid, improves survival and ameliorates organ system dysfunction in mice with peritonitis induced by caecal ligation and perforation, even when treatment is started as late as 12-24 hours after the onset of sepsis. In studies using lipopolysaccharide-stimulated RAW 264.7 murine macrophage like cells, EP inhibits activation of the pro-inflammatory transcription factor, NF-kappaB, and down regulates secretion of a number of pro-inflammatory cytokines, such as tumour necrosis factor (TNF). In this reductionist in vitro system, EP also blocks secretion of the late-appearing pro inflammatory cytokine-like molecule, high mobility group box 1 (HMGB1). In murine models of endotoxaemia or sepsis, treatment with EP decreases circulating levels of TNF and HMGB1. While the molecular events responsible for the salutary effects of EP remain to be elucidated, one mechanism may involve covalent modification of a critical thiol residue in the p65 component of NF-kappaB. EP warrants evaluation as a therapeutic agent for the treatment of sepsis in humans.

  7. Residual behavior of quizalofop ethyl on onion (Allium cepa L.).

    PubMed

    Sahoo, S K; Mandal, Kousik; Singh, Gurmail; Kumar, Rajinder; Chahil, G S; Battu, R S; Singh, Balwinder

    2013-02-01

    Quizalofop ethyl, a phenoxy propionate herbicide, is used for postemergence control of annual and perennial grass weeds in broad-leaved crops in India. The experiments were designed to study the dissipation kinetics of quizalofop ethyl on onion for two seasons. A simple, rapid, and sensitive method for estimation of quizalofop ethyl residues in onion and soil was developed and validated. The recoveries of quizalofop ethyl residues from onion and soil at different spiking level range from 84.81 to 92.68 %. The limit of quantification of this method was found to be 0.01 μg g(-1). The risk assessment through consumption of the onion in comparison to its acceptable daily intake which is an important parameter for the safety of the consumer was also evaluated. Standardized methodology supported by recovery studies was adopted to estimate residues of quizalofop ethyl on onion and soil. The average initial deposits of quizalofop ethyl on onion were observed to be 0.25 and 0.33 mg kg(-1), following single application of the herbicide at 50 g active ingredient (a.i.) ha(-1) during 2009 and 2010, respectively. The half-life values (T (1/2)) of quizalofop ethyl on onion crop were worked out to be 0.85 and 0.79 days, respectively, during 2009 and 2010. At harvest time, the residues of quizalofop ethyl on onion and soil were found to be below the determination limit of 0.01 mg kg(-1) following single application of the herbicide at 50 and 100 g a.i. ha(-1) for both the periods.

  8. Atmospheric Oxidation Mechanisms for Diethyl Ether and its Oxidation Products, Ethyl Formate and Ethyl Acetate.

    NASA Astrophysics Data System (ADS)

    Orlando, J. J.; Tyndall, G. S.

    2006-12-01

    Carbon-containing compounds are present in the earth's atmosphere as the result of emissions from natural and anthropogenic sources. Their oxidation in the atmosphere, initiated by such oxidants as OH, ozone, and nitrate radicals, leads to potentially harmful secondary pollutants such as ozone, carbonyl species, organic acids and aerosols. Ethers and esters are two classes of compounds that contribute to the complex array of organic compounds found in anthropogenically-influenced air. Additional ester is present as a result of the oxidation of the ethers. In this paper, the oxidation of diethyl ether and its two main oxidation products, ethyl formate and ethyl acetate, are studied over ranges of temperature, oxygen partial pressure, and NOx concentration, using an environmental chamber / FTIR absorption technique. Major end-products (the esters from diethyl ether; organic acids and anhydrides from the esters) are quantified, and these data are interpreted in terms of the chemistry of the various alkoxy and peroxy radicals generated. Emphasis is placed on the effects of chemical activation on the behavior of the alkoxy radicals, as well as on a novel peroxy radical rearrangement that may contribute to the observed products of ether oxidation under some conditions. Finally, the data are used, in conjunction with data on similar species, to provide a general representation of ether and ester oxidation in the atmosphere.

  9. Toxicity Studies of Ethyl Maltol and Iron Complexes in Mice.

    PubMed

    Li, Zhen; Lu, Jieli; Wu, Chonghui; Pang, Quanhai; Zhu, Zhiwei; Nan, Ruipeng; Du, Ruochen; Chen, Jia

    2017-01-01

    Ethyl maltol and iron complexes are products of ethyl maltol and the iron found in the cooking pots used to prepare the Chinese dish, hot-pot. Because their safety is undocumented, the toxicity study of ethyl maltol and iron complexes was conducted in male and female Kunming (KM) mice. The animal study was designed based on the preliminary study conducted to determine the median lethal dose (LD50). The doses used in the study were 0, 1/81, 1/27, 1/9, and 1/3 of the LD50 (mg kg body weight (BW)(-1) day(-1)) dissolved in the water. The oral LD50 of the ethyl maltol and iron complexes was determined to be 743.88 mg kg BW(-1) in mice. The ethyl maltol and iron complexes targeted the endocrine organs including the liver and kidneys following the 90 D oral exposure. Based on the haematological data, the lowest-observed-adverse-effect level (LOAEL) of the ethyl maltol and iron complexes was determined to be 1/81 LD50 (9.18 mg kg BW(-1) day(-1)) in both male and female mice. Therefore, we suggest that alternative strategies for preparing the hot-pot, including the use of non-Fe-based cookware, need to be developed and encouraged to avoid the formation of the potentially toxic complexes.

  10. Toxicity Studies of Ethyl Maltol and Iron Complexes in Mice

    PubMed Central

    Li, Zhen; Lu, Jieli; Wu, Chonghui; Zhu, Zhiwei; Nan, Ruipeng; Du, Ruochen; Chen, Jia

    2017-01-01

    Ethyl maltol and iron complexes are products of ethyl maltol and the iron found in the cooking pots used to prepare the Chinese dish, hot-pot. Because their safety is undocumented, the toxicity study of ethyl maltol and iron complexes was conducted in male and female Kunming (KM) mice. The animal study was designed based on the preliminary study conducted to determine the median lethal dose (LD50). The doses used in the study were 0, 1/81, 1/27, 1/9, and 1/3 of the LD50 (mg kg body weight (BW)−1 day−1) dissolved in the water. The oral LD50 of the ethyl maltol and iron complexes was determined to be 743.88 mg kg BW−1 in mice. The ethyl maltol and iron complexes targeted the endocrine organs including the liver and kidneys following the 90 D oral exposure. Based on the haematological data, the lowest-observed-adverse-effect level (LOAEL) of the ethyl maltol and iron complexes was determined to be 1/81 LD50 (9.18 mg kg BW−1 day−1) in both male and female mice. Therefore, we suggest that alternative strategies for preparing the hot-pot, including the use of non-Fe-based cookware, need to be developed and encouraged to avoid the formation of the potentially toxic complexes. PMID:28197411

  11. Androgen glucuronides analysis by liquid chromatography tandem-mass spectrometry: could it raise new perspectives in the diagnostic field of hormone-dependent malignancies?

    PubMed

    Kalogera, Eleni; Pistos, Constantinos; Provatopoulou, Xeni; Athanaselis, Sotirios; Spiliopoulou, Chara; Gounaris, Antonia

    2013-12-01

    Breast and prostate constitute organs of intense steroidogenic activity. Clinical and epidemiologic data provide strong evidence on the influence of androgens and estrogens on the risk of typical hormone-dependent malignancies, like breast and prostate cancer. Recent studies have focused on the role of androgen metabolites in regulating androgen concentrations in hormone-sensitive tissues. Steroid glucuronidation has been suggested to have a prominent role in controlling the levels and the biological activity of unconjugated androgens. It is well-established that serum levels of androgen glucuronides reflect androgen metabolism in androgen-sensitive tissues. Quantitative analysis of androgen metabolites in blood specimens is the only minimally invasive approach permitting an accurate estimate of the total pool of androgens. During the past years, androgen glucuronides analysis most often involved radioimmunoassays (RIA) or direct immunoassays, both methods bearing serious limitations. However, recent impressive technical advances in mass spectrometry, and particularly in high performance liquid chromatography coupled with mass spectrometry (LC-MS/MS), have overcome these drawbacks enabling the simultaneous, quantitative analysis of multiple steroids even at low concentrations. Blood androgen profiling by LC-MS/MS, a robust and reliable technique of high selectivity, sensitivity, specificity, precision and accuracy emerges as a promising new approach in the study of human pathology. The present review offers a contemporary insight in androgen glucuronides profiling through the application of LC-MS/MS, highlighting new perspectives in the study of steroids and their implication in hormone-dependent malignancies.

  12. Inhibition of glucuronidation and oxidative metabolism of buprenorphine using GRAS compounds or dietary constituents/supplements: in vitro proof of concept.

    PubMed

    Maharao, Neha V; Joshi, Anand A; Gerk, Phillip M

    2017-03-01

    The present study investigated the potential of generally recognized as safe (GRAS) compounds or dietary substances to inhibit the presystemic metabolism of buprenorphine and to increase its oral bioavailability. Using IVIVE, buprenorphine extraction ratios in intestine and liver were predicted as 96% and 71%, respectively. In addition, the relative fraction of buprenorphine metabolized by oxidation and glucuronidation in these two organs was estimated using pooled human intestinal and liver microsomes. In both organs, oxidation appeared to be the major metabolic pathway with a 6 and 4 fold higher intrinsic clearance than glucuronidation in intestine and liver, respectively. The oral bioavailability of buprenorphine was predicted to be 1.16%. Inhibition of 75% and 50% of intestinal and hepatic presystemic metabolism would result in an Foral of 49%, which is comparable to the bioavailability of sublingual buprenorphine. In human liver microsomes, chrysin, curcumin, ginger extract, hesperitin, magnolol, quercetin and silybin inhibited ≥50% glucuronidation, whereas chrysin, curcumin, ginger extract, 6-gingerol, pterostilbene, resveratrol and silybin exhibited ≥30% inhibition of oxidation. In human intestinal microsomes, curcumin, ginger extract, α-mangostin, quercetin and silybin inhibited ≥50% glucuronidation while chrysin, ginger extract, α-mangostin, pterostilbene and resveratrol exhibited ≥30% inhibition of oxidation. These results demonstrate the feasibility of our proposed approach of using GRAS or dietary compounds to inhibit the presystemic metabolism of buprenorphine and thus improve its oral bioavailability. An oral buprenorphine formulation containing these inhibitors or their combinations has promising potential to replace sublingual buprenorphine. Copyright © 2016 John Wiley & Sons, Ltd.

  13. Challenges in the indirect quantitation of acyl-glucuronide metabolites of a cardiovascular drug from complex biological mixtures in the absence of reference standards.

    PubMed

    Srinivasan, Karthik; Nouri, Parya; Kavetskaia, Olga

    2010-07-01

    This paper describes the quantitation of acyl-glucuronide metabolites (M26 and M5) of a cardiovascular-drug (torcetrapib) from monkey urine, in the absence of their reference standards. LC/MS/MS assays for M1 and M4 (aglycones of M26 and M5, respectively) were characterized from normal and base-treated urine, as their respective reference standards were available. The in vivo study samples containing M26 and M5 were treated with 1 n sodium hydroxide to hydrolyze them to their respective aglycones. The study samples were assayed for M1 and M4 before and after alkaline hydrolysis and the difference in the concentrations provided an estimate of the urinary levels of M26 and M5. Prior to the main sample analysis, conditions for alkaline hydrolysis of the glucuronides were optimized by incubating pooled study samples. During incubations, a prolonged increase in M4 levels over time was observed, which is inconsistent with the base-hydrolysis of an acyl-glucuronide (expected to hydrolyze rapidly). Possible interference of the metabolite M9 (an ether-glucuronide metabolite isobaric to M4) was investigated to explain this observation using chromatographic and wet-chemistry approaches. The strategies adopted herein established that the LC/MS/MS assay and our approach were reliable. The metabolite exposure was then correlated to toxicological observations to gain initial insights into the physiological role of these metabolites.

  14. Tentative Structural Assignment of a Glucuronide Metabolite of Methyltestosterone in Tilapia Bile by Liquid Chromatography-Quadrupole-Time-of-Flight Mass Spectrometry.

    PubMed

    Nishshanka, Upul; Chu, Pak-Sin; Evans, Eric; Reimschuessel, Renate; Hasbrouck, Nicholas; Amarasinghe, Kande; Jayasuriya, Hiranthi

    2015-06-24

    Methyltestosterone (MT), a strong androgenic steroid, is not approved for use in fish aquaculture in the United States. It is used in the U.S. under an investigational new animal drug exemption (INAD) only during the early life stages of fish. There is a possibility that farmers feed fish with MT to enhance production for economic gains. Therefore, there is a need to develop methods for the detection of MT and its metabolite residues in fish tissue for monitoring purposes. Previously, our laboratory developed a liquid chromatography-quadrupole time-of-flight (LC-QTOF) method for characterization of 17-O-glucuronide metabolite (MT-glu) in bile of tilapia dosed with MT. The system used was an Agilent 6530 Q-TOF equipped with electrospray jet stream technology, operating in positive ion mode. Retrospective analysis of the data generated in that experiment by a feature-finding algorithm, combined with a search against an in-house library of possible MT-metabolites, resulted in the discovery of a major glucuronide metabolite of MT in the bile extracts. Preliminary data indicate it to be a glucuronide of a hydroxylated MT (OHMT-glu) which persists in tilapia bile for at least 2 weeks after dosing. We present the tentative structural assignment of the OHMT-glu in tilapia bile and time course of development. This glucuronide can serve as a marker to monitor illegal use of MT in tilapia culture.

  15. 19 CFR 10.99 - Importation of ethyl alcohol for nonbeverage purposes.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 19 Customs Duties 1 2013-04-01 2013-04-01 false Importation of ethyl alcohol for nonbeverage... Provisions Ethyl Alcohol § 10.99 Importation of ethyl alcohol for nonbeverage purposes. (a) If claim is made... of ethyl alcohol of an alcoholic strength by volume of 80 percent volume or higher under...

  16. 19 CFR 10.99 - Importation of ethyl alcohol for nonbeverage purposes.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 19 Customs Duties 1 2012-04-01 2012-04-01 false Importation of ethyl alcohol for nonbeverage... Provisions Ethyl Alcohol § 10.99 Importation of ethyl alcohol for nonbeverage purposes. (a) If claim is made... of ethyl alcohol of an alcoholic strength by volume of 80 percent volume or higher under...

  17. 19 CFR 10.99 - Importation of ethyl alcohol for nonbeverage purposes.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 19 Customs Duties 1 2010-04-01 2010-04-01 false Importation of ethyl alcohol for nonbeverage... Provisions Ethyl Alcohol § 10.99 Importation of ethyl alcohol for nonbeverage purposes. (a) If claim is made... of ethyl alcohol of an alcoholic strength by volume of 80 percent volume or higher under...

  18. Characterization of niflumic acid as a selective inhibitor of human liver microsomal UDP-glucuronosyltransferase 1A9: application to the reaction phenotyping of acetaminophen glucuronidation.

    PubMed

    Miners, John O; Bowalgaha, Kushari; Elliot, David J; Baranczewski, Pawel; Knights, Kathleen M

    2011-04-01

    Enzyme selective inhibitors represent the most valuable experimental tool for reaction phenotyping. However, only a limited number of UDP-glucuronosyltransferase (UGT) enzyme-selective inhibitors have been identified to date. This study characterized the UGT enzyme selectivity of niflumic acid (NFA). It was demonstrated that 2.5 μM NFA is a highly selective inhibitor of recombinant and human liver microsomal UGT1A9 activity. Higher NFA concentrations (50-100 μM) inhibited UGT1A1 and UGT2B15 but had little effect on the activities of UGT1A3, UGT1A4, UGT1A6, UGT2B4, UGT2B7, and UGT2B17. NFA inhibited 4-methylumbelliferone and propofol (PRO) glucuronidation by recombinant UGT1A9 and PRO glucuronidation by human liver microsomes (HLM) according to a mixed (competitive-noncompetitive) mechanism, with K(i) values ranging from 0.10 to 0.40 μM. Likewise, NFA was a mixed or noncompetitive inhibitor of recombinant and human liver microsomal UGT1A1 (K(i) range 14-18 μM), whereas competitive inhibition (K(i) 62 μM) was observed with UGT2B15. NFA was subsequently applied to the reaction phenotyping of human liver microsomal acetaminophen (APAP) glucuronidation. Consistent with previous reports, APAP was glucuronidated by recombinant UGT1A1, UGT1A6, UGT1A9, and UGT2B15. NFA concentrations in the range of 2.5 to 100 μM inhibited APAP glucuronidation by UGT1A1, UGT1A9, and UGT2B15 but not by UGT1A6. The mean V(max) for APAP glucuronidation by HLM was reduced by 20, 35, and 40%, respectively, in the presence of 2.5, 50, and 100 μM NFA. Mean K(m) values decreased in parallel with V(max), although the magnitude of the decrease was smaller. Taken together, the NFA inhibition data suggest that UGT1A6 is the major enzyme involved in APAP glucuronidation.

  19. Cremophor EL-based nanoemulsion enhances transcellular permeation of emodin through glucuronidation reduction in UGT1A1-overexpressing MDCKII cells.

    PubMed

    Zhang, Tianpeng; Dong, Dong; Lu, Danyi; Wang, Shuai; Wu, Baojian

    2016-03-30

    Oral emodin, a natural anthraquinone and active component of many herbal medicines, is poorly bioavailable because of extensive first-pass glucuronidation. Here we aimed to prepare emodin nanoemulsion (EMO-NE) containing cremophor EL, and to assess its potential for enhancing transcellular absorption of emodin using UGT1A1-overexpressing MDCKII cells (or MDCK1A1 cells). EMO-NE was prepared using a modified emulsification technique and subsequently characterized by particle size, morphology, stability, and drug release. MDCKII cells were stably transfected with UGT1A1 using the lentiviral transfection approach. Emodin transport and metabolism were evaluated in Transwell-cultured MDCK1A1 cells after apical dosing of EMO-NE or control solution. The obtained EMO-NE (116 ± 6.5 nm) was spherical and stable for at least 2 months. Emodin release in vitro was a passive diffusion-driven process. EMO-NE administration increased the apparent permeability of emodin by a 2.3-fold (p<0.001) compared to the pure emodin solution (1.2 × 10(-5) cm/s vs 5.3 × 10(-6) cm/s). Further, both apical and basolateral excretion of emodin glucuronide (EMO-G) were significantly decreased (≥56.5%, p<0.001) in EMO-NE group. This was accompanied by a marked reduction (57.4%, p<0.001) in total emodin glucuronidation. It was found that the reduced glucuronidation was due to inhibition of cellular metabolism by cremophor EL. Cremophor EL inhibited UGT1A1-mediated glucuronidation of emodin using the mixed-type inhibition mechanism. In conclusion, cremophor EL-based nanoemulsion greatly enhanced transcellular permeation of emodin through inhibition of UGT metabolism. This cremophor EL-based nanoformulation may be a promising strategy to improve the oral bioavailability of emodin.

  20. Investigation of a recently detected 11-nor-9-carboxy-Δ(9)-tetrahydrocannabinol isomer: Studies on the degradation of 11-nor-9-carboxy-Δ(9)-tetrahydrocannabinol glucuronide.

    PubMed

    Hanisch, Stephanie; Paulke, Alexander; Toennes, Stefan W

    2016-09-10

    An isomer of the tetrahydrocannabinol (THC) metabolite 11-nor-9-carboxy-Δ(9)-THC (THCCOOH) had been detected in blood of cannabis users. The present study was initiated to elucidate whether the labile metabolite THCCOOH-glucuronide could be the precursor. THCCOOH-glucuronide was incubated in human serum and albumin (HSA) solution at various temperatures (-18, 4.5, 22 and 37°C) and pH values (pH 7.4 and 8.3) for seven days in the presence or absence of the esterase inhibitor sodium fluoride. Analysis of incubation samples was performed using LC-MS/MS. Marked degradation of THCCOOH-glucuronide was observed at 37°C. It was found that not only THCCOOH, but also the isomer is a degradation product of THCCOOH-glucuronide and its in-vivo production is assumed. Degradation to THCCOOH and the isomer occurred at alkaline pH, in the presence of fluoride-sensitive esterases and of HSA alone. To inhibit isomer formation during sample storage, refrigeration and controlling of the pH are recommended. However, THCCOOH and the isomer exhibit similar properties during incubations in serum, but differ in their interaction with HSA. The present study confirmed the nature of the isomer as degradation product of the abundant THC metabolite THCCOOH-glucuronide. Serum albumin and esterases are obviously involved. The isomer is formed not only during storage, but also under physiological conditions, suggesting that it can be considered an in-vivo metabolite. However, the chemical structure of the isomer remains unknown and further research is necessary.

  1. Pharmacokinetics and elucidation of the rates and routes of N-glucuronidation of PF-592379, an oral dopamine 3 agonist in rat, dog, and human.

    PubMed

    Attkins, Neil; Betts, Alison; Hepworth, David; Heatherington, Anne C

    2010-11-01

    PF-592379 is a potent, selective agonist of the dopamine 3 receptor, for the treatment of male erectile dysfunction and female sexual dysfunction. In vivo, PF-592379 has low-moderate clearance relative to liver blood flow of 6.3 and 8.5 ml/min/kg in dog and 44.8 and 58.2 ml/min/kg in rat. It has high permeability in Caco-2 cells and was completely absorbed in rat and dog pharmacokinetic studies with an oral bioavailability of 28% in both rats and 61 and 87% in the dogs. These data are consistent with the physicochemical properties of PF-592379, which indicate complete absorption by the transcellular route. Elimination of PF-592379 was predominantly metabolic in nature. In vitro routes of metabolism studies indicate that metabolism in the rat is a combination of P450 mechanisms and N-glucuronidation, whereas in dog and human, N-glucuronidation is the major route. NMR analysis indicates that N-glucuronidation is non-quaternary in nature and occurs on both the pyridyl amine and ring nitrogen. Rates of clearance via N-glucuronidation were predicted to be low in humans compared with acyl or phenolic glucuronidation. PF-592379 was predicted to have complete absorption from the gastrointestinal tract and an oral bioavailability of >60% in the clinic. Clinical data verified that PF-592379 is a low clearance compound in human, with a mean oral clearance of 6.5 ml/min/kg following a 200 mg oral dose. PF-592379 has ideal pharmacokinetic properties for an oral D3 agonist, intended for on demand dosing.

  2. The Impact of Glucuronidation on the Bioactivation and DNA Adduction of the Cooked-Food Carcinogen 2-Amino-1-methyl-6-phenylimidazo[4,5-b] pyridine in vivo

    SciTech Connect

    Malfatti, M A; Ubick, E A; Felton, J S

    2005-03-31

    UDP-glucuronosyltransferases (UGTs) catalyze the glucuronidation of many different chemicals. Glucuronidation is especially important for detoxifying reactive intermediates from metabolic reactions, which otherwise can be biotransformed into highly reactive cytotoxic or carcinogenic species. Detoxification of certain food-borne carcinogenic heterocyclic amines (HAs) is highly dependent on UGT1A-mediated glucuronidation. 2-Amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), the most mass abundant carcinogenic HA found in well-done cooked meat, is extensively glucuronidated by UGT1A proteins. In humans, CYP1A2 catalyzed N-hydroxylation and subsequent UGT1A-mediated glucuronidation is a dominant pathway in the metabolism of PhIP. Therefore, changes in glucuronidation rates could significantly alter PhIP metabolism. To determine the importance of UGT1A-mediated glucuronidation in the biotransformation of PhIP, UGT1A proficient Wistar and UGT1A deficient Gunn rats were exposed to a single 100 {micro}g/kg oral dose of [{sup 14}C]-PhIP. Urine was collected over 24 h and the PhIP urinary metabolite profiles were compared between the two strains. After the 24 h exposure, livers and colon were removed and analyzed for DNA adduct formation by accelerator mass spectrometry. Wistar rats produced several PhIP and N-hydroxy-PhIP glucuronides that accounted for {approx}25% of the total amount of recovered urinary metabolites. In the Gunn rats, PhIP and N-hydroxy-PhIP glucuronides were reduced by 68-92%, compared to the Wistar rats, and comprised only 4% of the total amount of recovered urinary metabolites. PhIP-DNA adduct analysis from the Gunn rats revealed a correlation between reduced PhIP and N-hydroxy-PhIP glucuronide levels in the urine and increased hepatic DNA adducts, compared to the Wistar rats. These results indicate that UGT1A-mediated glucuronidation of PhIP and N-hydroxy-PhIP is an important pathway for PhIP detoxification. Failure to form glucuronide conjugates

  3. Differential effects of microsomal enzyme inducers on in vitro thyroxine (T(4)) and triiodothyronine (T(3)) glucuronidation.

    PubMed

    Hood, A; Klaassen, C D

    2000-05-01

    Microsomal enzyme inducers that increase UDP-glucuronosyltransferase (UDP-GT) activity are suspected to affect the thyroid gland by increasing the glucuronidation of T(4), which reduces serum thyroxine (T(4)). In response to reduced serum T(4), serum thyroid-stimulating hormone (TSH) increases. However, not all microsomal enzyme inducers that reduce serum T(4) produce an increase in serum TSH. We have shown that serum TSH is increased the most in rats treated with the microsomal enzyme inducers phenobarbital (PB) or pregnenolone-16alpha-carbonitrile (PCN), whereas TSH is affected less in rats treated with 3-methylcholanthrene (3MC) and Aroclor 1254 (PCB). It is unclear why serum TSH is differentially affected by various microsomal enzyme inducers. We propose that the glucuronidation of T(3) might be the reason serum TSH is increased by some microsomal enzyme inducers but not by others. Male Sprague-Dawley rats were fed either a basal diet or a diet containing PB (at 300, 600, 1200, or 2400 ppm), PCN (at 200, 400, 800, or 1600 ppm), 3MC (at 50, 100, 200, or 400 ppm), or PCB (at 25, 50, 100, or 200 ppm) for 7 days; and T(4) and T(3) UDP-GT activities were then determined. T(4) UDP-GT activity was increased in rats treated with PB (120%), PCN (250 to 400%), 3MC (400 to 600%), or PCB (300 to 430%). In contrast, T(3) UDP-GT activity was increased in rats treated with PB (90%) or PCN (120 to 200%), whereas 3MC and PCB treatments did not have an appreciable effect. In conclusion, differential effects on T(3) glucuronosyltransferase activity were found in rats treated with microsomal enzyme inducers.

  4. Determining the partial photoionization cross-sections of ethyl radicals.

    PubMed

    FitzPatrick, B L; Maienschein-Cline, M; Butler, L J; Lee, S-H; Lin, J J

    2007-12-13

    Using a crossed laser-molecular beam scattering apparatus, these experiments photodissociate ethyl chloride at 193 nm and detect the Cl and ethyl products, resolved by their center-of-mass recoil velocities, with vacuum ultraviolet photoionization. The data determine the relative partial cross-sections for the photoionization of ethyl radicals to form C2H5+, C2H4+, and C2H3+ at 12.1 and 13.8 eV. The data also determine the internal energy distribution of the ethyl radical prior to photoionization, so we can assess the internal energy dependence of the photoionization cross-sections. The results show that the C2H4++H and C2H3++H2 dissociative photoionization cross-sections strongly depend on the photoionization energy. Calibrating the ethyl radical partial photoionization cross-sections relative to the bandwidth-averaged photoionization cross-section of Cl atoms near 13.8 eV allows us to use these data in conjunction with literature estimates of the Cl atom photoionization cross-sections to put the present bandwidth-averaged cross-sections on an absolute scale. The resulting bandwidth-averaged cross-section for the photoionization of ethyl radicals to C2H5+ near 13.8 eV is 8+/-2 Mb. Comparison of our 12.1 eV data with high-resolution ethyl radical photoionization spectra allows us to roughly put the high-resolution spectrum on the same absolute scale. Thus, one obtains the photoionization cross-section of ethyl radicals to C2H5+ from threshold to 12.1 eV. The data show that the onset of the C2H4++H dissociative photoionization channel is above 12.1 eV; this result offers a simple way to determine whether the signal observed in photoionization experiments on complex mixtures is due to ethyl radicals. We discuss an application of the results for resolving the product branching in the O+allyl bimolecular reaction.

  5. Chemical fingerprint and metabolic profile analysis of ethyl acetate fraction of Gastrodia elata by ultra performance liquid chromatography/quadrupole-time of flight mass spectrometry.

    PubMed

    Tang, Chunlan; Wang, Li; Liu, Xinxin; Cheng, Mengchun; Xiao, Hongbin

    2016-02-01

    The chemical fingerprint and metabolic profile of traditional Chinese medicine is very complicated and has been a great challenge. In the present study, chemical fingerprint of ethyl acetate fraction of Gastrodia elata (EtAcGE) and metabolic profile of rat plasma sample after intragastric administration of EtAcGE (2.5g/kg) were investigated using ultra-high performance liquid chromatography coupled with quadrupole-time of flight mass spectrometry (UPLC/Q-TOF MS). A total of 38 chemical constituents of EtAcGE were identified by comparing their retention time, accurate molecular mass and characteristic fragment ions with those of references, or tentatively characterized by comparing molecular formula, fragment ions with that of known compound or information available in literature. And 40 compounds were detected in dosed rat plasma sample, including 16 prototypes and 24 metabolites underwent metabolic process of glucuronidation, glucosylation, sulfation, methylation, hydroxylation, dehydrogenation or mixed modes. The metabolic "soft spots" was hydroxyl or carboxy group. This is the first research for chemical fingerprint and metabolic profile of EtAcGE, which lay a foundation for the further investigation of EtAcGE.

  6. Utility of coloured hair for the detection of drugs and alcohol.

    PubMed

    Agius, Ronald

    2014-06-01

    The aim of this paper is to assess the utility of coloured hair for the detection of drugs and alcohol in a large statistically significant population. The positivity rate, the 1st, 5th, 50th, 95th, and 99th percentiles of five amphetamines, cannabinoids, cocaine, four opiates, methadone, buprenorphine, seven benzodiazepines, and ethyl glucuronide (EtG) in 9488 non-treated and 1026 cosmetically treated (dyed or bleached) authentic hair samples was compared. Analytical methods used were accredited for forensic purposes at the cut-offs defined by the German driving licence re-granting medical and psychological assessment (MPA) guidelines. Considering only the drug classes for which at least 10 positive samples were detected, the positivity rate in non-treated hair was highest for alcohol (4.50%; measured using EtG at concentrations ≥ 7 pg/mg hair), followed by THC (2.00%), cocaine (1.75%), and amphetamine (0.59%). While the 1st to 99th percentile range was significantly lower for drugs in treated, compared to non-treated hair, no significant change was observed for EtG. Additionally, no significant difference in the positivity rate was observed between treated hair and non-treated hair for both drugs and EtG. This study is the first attempt to evaluate the influence of cosmetic treatment, mainly dying, on the positivity rate for both drugs and EtG in hair samples submitted routinely for abstinence testing and the first to indicate that dyed and eventually bleached hair is not necessarily useless in detecting drugs and/or alcohol consumption, thus making coloured hair analysis still useful, often being the only possibility to prove such misuse.

  7. Health and Environmental Effects Profile for ethyl methacrylate

    SciTech Connect

    Not Available

    1986-06-01

    The Health and Environmental Effects Profile for ethyl methacrylate was prepared to support listings of hazardous constituents of a wide range of waste streams under Section 3001 of the Resource Conservation and Recovery Act (RCRA) and to provide health-related limits for emergency actions under Section 101 of the Comprehensive Environmental Response, Compensation and Liability Act (CERCLA). Both published literature and information obtained from Agency program office files were evaluated as they pertained to potential human health, aquatic life and environmental effects. Quantitative estimates are presented provided sufficient data are available. Ethyl methacrylate has been determined to be a systemic toxicant. An acceptable daily intake (ADI) for ethyl methacrylate is 0.086 mg/kg/day for oral exposure.

  8. Fragrance material review on ethyl phenyl carbinyl acetate.

    PubMed

    McGinty, D; Letizia, C S; Api, A M

    2012-09-01

    A toxicologic and dermatologic review of ethyl phenyl carbinyl acetate when used as a fragrance ingredient is presented. Ethyl phenyl carbinyl acetate is a member of the fragrance structural group Aryl Alkyl Alcohol Simple Acid Esters (AAASAE). The AAASAE fragrance ingredients are prepared by reacting an aryl alkyl alcohol with a simple carboxylic acid (a chain of 1-4 carbons) to generate formate, acetate, propionate, butyrate, isobutyrate and carbonate esters. This review contains a detailed summary of all available toxicology and dermatology papers that are related to this individual fragrance ingredient and is not intended as a stand-alone document. Available data for ethyl phenyl carbinyl acetate were evaluated, then summarized, and includes: physical properties; acute toxicity; skin irritation; and skin sensitization data. A safety assessment of the entire AAASAE will be published simultaneously with this document; please refer to Belsito et al. (2012) for an overall assessment of the safe use of this material and all AAASAE in fragrances.

  9. Fragrance material review on 2-(p-tolyloxy)ethyl acetate.

    PubMed

    McGinty, D; Letizia, C S; Api, A M

    2012-09-01

    A toxicologic and dermatologic review of 2-(p-tolyloxy)ethyl acetate when used as a fragrance ingredient is presented. 2-(p-tolyloxy)ethyl acetate is a member of the fragrance structural group aryl alkyl alcohol simple acid esters (AAASAE). The AAASAE fragrance ingredients are prepared by reacting an aryl alkyl alcohol with a simple carboxylic acid (a chain of 1-4 carbons) to generate formate, acetate, propionate, butyrate, isobutyrate and carbonate esters. This review contains a detailed summary of all available toxicology and dermatology papers that are related to this individual fragrance ingredient and is not intended as a stand-alone document. Available data for 2-(p-tolyloxy)ethyl acetate were evaluated, then summarized, and includes physical properties data. A safety assessment of the entire AAASAE will be published simultaneously with this document. Please refer to Belsito et al. (2012) for an overall assessment of the safe use of this material and all AAASAE in fragrances.

  10. NEW GROUND-STATE MEASUREMENTS OF ETHYL CYANIDE

    SciTech Connect

    Brauer, Carolyn S.; Pearson, John C.; Drouin, Brian J.; Yu, Shanshan

    2009-09-01

    The spectrum of ethyl cyanide, or propionitrile (CH{sub 3}CH{sub 2}CN), has been repeatedly observed in the interstellar medium with large column densities and surprisingly high temperatures in hot core sources. The construction of new, more sensitive, observatories accessing higher frequencies such as Herschel, ALMA, and SOFIA have made it important to extend the laboratory data for ethyl cyanide to coincide with the capabilities of the new instruments. We report extensions of the laboratory measurements of the rotational spectrum of ethyl cyanide in its ground vibrational state to 1.6 THz. A global analysis of the ground state, which includes all of the previous data and 3356 newly assigned transitions, has been fitted to within experimental error to J = 132, K = 36, using both Watson A-reduced and Watson S-reduced Hamiltonians.

  11. IRIS Toxicological Review of Ethyl Tertiary Butyl Ether (ETBE) ...

    EPA Pesticide Factsheets

    The IRIS Toxicological Review of Ethyl Tertiary Butyl Ether (ETBE) was released for external peer review in April 2017. EPA’s Science Advisory Board’s (SAB) Chemical Assessment Advisory Committee (CAAC) will conduct a peer review of the scientific basis supporting the ETBE assessment and release a final report of their review. Information regarding the peer review can be found on the SAB website. EPA is conducting an Integrated Risk Information System (IRIS) health assessment for Ethyl Tertiary Butyl Ether (ETBE). The outcome of this project is a Toxicological Review and IRIS Summary for ETBE that will be entered into the IRIS database.

  12. Synthesis and Characterization of New Optically Active Poly (ethyl L-lysinamide)s and Poly (ethyl L-lysinimide)s

    PubMed Central

    Zahmatkesh, Saeed; Vakili, Mohammad Reza

    2010-01-01

    Ethyl L-lysine dihydrochloride was reacted with three different dianhydrides to yield the poly (ethyl L-lysinimide)s (PI1−3); it was also reacted with two different diacyl chlorides to yield the poly (ethyl L-lysinamide)s (PA4-5). The resulting polymers have inherent viscosities in the range of 0.15 to 0.42 dL g−1. These polymers are prepared from an inexpensive starting material and are optically active, potentially ion exchangeable, semicrystalline, thermally stable, and soluble in polar aprotic solvents such as DMF, DMSO, NMP, DMAc, and sulfuric acid. All of the above polymers were fully characterized by FT-IR and 1H NMR spectroscopy, elemental analysis, WAX diffraction, TGA, inherent viscosity measurement, and specific rotation. PMID:22331998

  13. 5α-Androst-16-en-3α-ol β-D-glucuronide, precursor of 5α-androst-16-en-3α-ol in human sweat.

    PubMed

    Starkenmann, Christian; Mayenzet, Fabienne; Brauchli, Robert; Troccaz, Myriam

    2013-12-01

    5α-Androst-16-en-3α-ol (α-androstenol) is an important contributor to human axilla sweat odor. It is assumed that α-andostenol is excreted from the apocrine glands via a H2 O-soluble conjugate, and this precursor was formally characterized in this study for the first time in human sweat. The possible H2 O-soluble precursors, sulfate and glucuronide derivatives, were synthesized as analytical standards, i.e., α-androstenol, β-androstenol sulfates, 5α-androsta-5,16-dien-3β-ol (β-androstadienol) sulfate, α-androstenol β-glucuronide, α-androstenol α-glucuronide, β-androstadienol β-glucuronide, and α-androstenol β-glucuronide furanose. The occurrence of α-androstenol β-glucuronide was established by ultra performance liquid chromatography (UPLC)/MS (heated electrospray ionization (HESI)) in negative-ion mode in pooled human sweat, containing eccrine and apocrine secretions and collected from 25 female and 24 male underarms. Its concentration was of 79 ng/ml in female secretions and 241 ng/ml in male secretions. The release of α-androstenol was observed after incubation of the sterile human sweat or α-androstenol β-glucuronide with a commercial glucuronidase enzyme, the urine-isolated bacteria Streptococcus agalactiae, and the skin bacteria Staphylococcus warneri DSM 20316, Staphylococcus haemolyticus DSM 20263, and Propionibacterium acnes ATCC 6919, reported to have β-glucuronidase activities. We demonstrated that if α- and β-androstenols and androstadienol sulfates were present in human sweat, their concentrations would be too low to be considered as potential precursors of malodors; therefore, the H2 O-soluble precursor of α-androstenol in apocrine secretion should be a β-glucuronide.

  14. Decreased Expression of Multidrug Resistance-Associated Protein 4 (MRP4/ABCC4) Leads to Reduced Glucuronidation of Flavonoids in UGT1A1-Overexpressing HeLa Cells: The Role of Futile Recycling.

    PubMed

    Sun, Hua; Zhou, Xiaotong; Zhang, Xingwang; Wu, Baojian

    2015-07-08

    In this study, the role of futile recycling (or deglucuronidation) in the disposition of two flavonoids (i.e., genistein and apigenin) was explored using UGT1A1-overexpressing HeLa cells (or HeLa1A1 cells). Glucuronidation of the flavonoids by HeLa1A1 cell lysate followed the substrate inhibition kinetics (Vmax = 0.10 nmol/min/mg, Km = 0.54 μM, and Ksi = 2.0 μM for genistein; Vmax = 0.19 nmol/min/mg, Km = 0.56 μM, and Ksi = 3.7 μM for apigenin). Glucuronide was efficiently generated and excreted after incubation of the cells with the aglycone (at doses of 1.25-20 nmol). The excretion rates were 0.40-0.69 and 0.84-1.1 nmol/min/mg protein for genistein glucuronide (GG) and apigenin glucuronide (AG), respectively. Furthermore, glucuronide excretion and total glucuronidation were significantly reduced in MRP4 knocked-down as compared to control cells. The alterations were well characterized by a two-compartment pharmacokinetic model incorporating the process of futile recycling (defined by a first-order rate constant, Kde). The derived Kde values were 15 and 25 h(-1) for GG and AG, respectively. This was well consistent with the in vitro observation that AG was subjected to more efficient futile recycling compared to GG. In conclusion, futile recycling was involved in cellular glucuronidation, accounting for transporter-dependent glucuronidation of flavonoids.

  15. Kinetic modeling of the interactions between 4-methylumbelliferone, 1-naphthol, and zidovudine glucuronidation by udp-glucuronosyltransferase 2B7 (UGT2B7) provides evidence for multiple substrate binding and effector sites.

    PubMed

    Uchaipichat, Verawan; Galetin, Aleksandra; Houston, J Brian; Mackenzie, Peter I; Williams, J Andrew; Miners, John O

    2008-10-01

    Interactions between the UGT2B7-catalyzed glucuronidation of zidovudine (AZT), 4-methylumbelliferone (4MU), and 1-naphthol (1NP) were analyzed using multisite and empirical kinetic models to explore the existence of multiple substrate and effector binding sites within this important drug metabolizing enzyme. 4MU and 1NP glucuronidation by UGT2B7 exhibit sigmoidal kinetics characteristic of homotropic cooperativity (autoactivation), which may be modeled assuming the existence of two equivalent, interacting substrate binding sites. In contrast, UGT2B7-catalyzed AZT glucuronidation follows hyperbolic (Michaelis-Menten) kinetics. Although 4MU and 1NP decreased the binding affinity of AZT, the kinetics of AZT glucuronidation changed from hyperbolic to sigmoidal in the presence of both modifiers. Data were well described by a generic two-substrate binding site model in which there is no interaction between the sites in the absence of 4MU or 1NP, but heterotropic cooperativity results from the binding of modifier. Inhibition of 4MU and 1NP glucuronidation by AZT and interactions between 4MU and 1NP required more complex three-site models, where the modifier acts via a distinct effector site to alter either substrate binding affinity or Vmax without affecting the homotropic cooperativity characteristic of 4MU and 1NP glucuronidation. It is noteworthy that 1NP inhibited 4MU glucuronidation, whereas 4MU activated 1NP glucuronidation. The results are consistent with the existence of two "catalytic" sites for each substrate within the UGT2B7 active site, along with multiple effector sites. The multiplicity of binding and effector sites results in complex kinetic interactions between UGT2B7 substrates, which potentially complicates inhibition screening studies.

  16. Enantiomer selective glucuronidation of the non-steroidal pure anti-androgen bicalutamide by human liver and kidney: role of the human UDP-glucuronosyltransferase (UGT)1A9 enzyme.

    PubMed

    Grosse, Laurent; Campeau, Anne-Sophie; Caron, Sarah; Morin, Frédéric-Alexandre; Meunier, Kim; Trottier, Jocelyn; Caron, Patrick; Verreault, Mélanie; Barbier, Olivier

    2013-08-01

    Bicalutamide (Casodex(®) ) is a non-steroidal pure anti-androgen used in the treatment of localized prostate cancer. It is a racemate drug, and its activity resides in the (R)-enantiomer, with little in the (S)-enantiomer. A major metabolic pathway for bicalutamide is glucuronidation catalysed by UDP-glucuronosyltransferase (UGT) enzymes. While (S)bicalutamide is directly glucuronidated, (R)bicalutamide requires hydroxylation prior to glucuronidation. The contribution of human tissues and UGT isoforms in the metabolism of these enantiomers has not been extensively investigated. In this study, both (R) and/or (S)bicalutamide were converted into glucuronide (-G) derivatives after incubation of pure and racemic solutions with microsomal extracts from human liver and kidney. Intestinal microsomes exhibited only low reactivity with these substrates. Km values of liver and kidney samples for (S)bicalutamide glucuronidation were similar, and lower than values obtained with the (R)-enantiomer. Among the 16 human UGTs tested, UGT1A8 and UGT1A9 were able to form both (S) and (R)bicalutamide-G from pure or racemic substrates. UGT2B7 was also able to form (R)bicalutamide-G. Kinetic parameters of the recombinant UGT2B7, UGT1A8 and UGT1A9 enzymes support a predominant role of the UGT1A9 isoform in bicalutamide metabolism. Accordingly, (S)bicalutamide inhibited the ability of human liver and kidney microsomes to glucuronidate the UGT1A9 probe substrate, propofol. In conclusion, the present study provides the first comprehensive analysis of in vitro bicalutamide glucuronidation by human tissues and UGTs and identifies UGT1A9 as a major contributor for (R) and (S) glucuronidation in the human liver and kidney.

  17. Enantiomer selective glucuronidation of the non-steroidal pure anti-androgen bicalutamide by human liver and kidney: role of the human UDP-glucuronosyltransferase (UGT)1A9 enzyme

    PubMed Central

    Grosse, Laurent; Campeau, Anne-Sophie; Caron, Sarah; Morin, Frédéric-Alexandre; Meunier, Kim; Trottier, Jocelyn; Caron, Patrick; Verreault, Mélanie; Barbier, Olivier

    2013-01-01

    Bicalutamide (Casodex®) is a non-steroidal pure anti-androgen used in the treatment of localized prostate cancer. It is a racemate drug and its activity resides in the (R)-enantiomer, with little in the (S)-enantiomer. A major metabolic pathway for bicalutamide is glucuronidation catalyzed by UDP-glucuronosyltransferase (UGT) enzymes. While (S)bicalutamide is directly glucuronidated, (R)bicalutamide requires hydroxylation prior to glucuronidation. The contribution of human tissues and UGT isoforms in the metabolism of these enantiomers has not been extensively investigated. In this study, both (R) and/or (S)bicalutamide were converted into glucuronide (-G) derivatives following incubation of pure and racemic solutions with microsomal extracts from human liver and kidney. Intestinal microsomes exhibited only low reactivity with these substrates. Km values of liver and kidney samples for (S)bicalutamide glucuronidation were similar, and lower than values obtained with the (R)-enantiomer. Among the 16 human UGTs tested, UGT1A8 and UGT1A9 were able to form both (S) and (R)bicalutamide-G from pure or racemic substrates. UGT2B7 was also able to form (R)bicalutamide-G. Kinetic parameters of the recombinant UGT2B7, UGT1A8 and UGT1A9 enzymes support a predominant role of the UGT1A9 isoform in bicalutamide metabolism. Accordingly, (S)bicalutamide inhibited the ability of human liver and kidney microsomes to glucuronidate the UGT1A9 probe substrate, propofol. In conclusion, the present study provides the first comprehensive analysis of in vitro bicalutamide glucuronidation by human tissues and UGTs, and identifies UGT1A9 as a major contributor for (R) and (S) glucuronidation in the human liver and kidney. PMID:23527766

  18. Modulation of Strawberry/Cranberry Phenolic Compounds Glucuronidation by Co-Supplementation with Onion: Characterization of Phenolic Metabolites in Rat Plasma Using an Optimized μSPE-UHPLC-MS/MS Method.

    PubMed

    Dudonné, Stéphanie; Dubé, Pascal; Pilon, Geneviève; Marette, André; Jacques, Hélène; Weisnagel, John; Desjardins, Yves

    2014-03-26

    Plant phenolic compounds are suggested to exert pharmacological activities in regards to obesity and type-2 diabetes, but their mode of action is poorly understood due to a lack of information about their bioavailability. This work aimed to study the bioavailability of GlucoPhenol phenolic compounds, a strawberry-cranberry extracts blend, by characterizing plasma phenolic profile in obese rats. A comparison was performed by co-supplementation with an onion extract. Using an optimized μSPE-UHPLC-MS/MS method, 21 phenolic metabolites were characterized, mostly conjugated metabolites and microbial degradation products of the native phenolic compounds. Their kinetic profiles revealed either an intestinal or hepatic formation. Among identified metabolites, isorhamnetin glucuronide sulfate was found in greater amount in plasma. Three glucuronidated conjugates of strawberry-cranberry phenolic compounds, p-hydroxybenzoic acid glucuronide, catechins glucuronide, and methyl catechins glucuronide were found in higher quantities when GlucoPhenol was ingested together with onion extract (+252%, +279%, and +118% respectively), suggesting a possible induction of glucuronidation processes by quercetin. This work allowed the characterization of actual phenolic metabolites generated in vivo following a phenolic intake, the analysis of their kinetics and suggested a possible synergistic activity of phenolic compounds for improving bioavailability.

  19. An in vitro experiment on the interaction of charcoal or wheat bran with 11-nor-9-carboxy-Δ(9)-tetrahydrocannabinol and its glucuronide.

    PubMed

    Skopp, Gisela; Mikus, Gerd

    2013-11-01

    The rather long yet variable terminal half-lives and detection times since last use of urinary cannabinoids may partly be attributed to their enterohepatic circulation which generally can be interrupted or restricted by chemical adsorbents. Therefore, an in vitro experiment was performed to study the adsorption/binding of 11-nor-9-carboxy-Δ9-tetrahydrocannabinol (THC-COOH) and its glucuronide to activated charcoal and wheat bran; remaining concentrations were determined by liquid chromatography/tandem mass spectrometry. Adsorption/binding of 1,000 ng/mL of free or conjugated THC-COOH was complete using as little as 5 mg of charcoal whereas adsorption/binding to wheat bran increased with increasing amounts. Taking of remedies affecting enterohepatic recycling of THC-COOH and its glucuronide may challenge interpretation of cannabinoid concentrations used to detect or assess frequency of drug use or the time since last drug consumption.

  20. Development of a fast screening and confirmatory method by liquid chromatography-quadrupole-time-of-flight mass spectrometry for glucuronide-conjugated methyltestosterone metabolite in tilapia.

    PubMed

    Amarasinghe, Kande; Chu, Pak-Sin; Evans, Eric; Reimschuessel, Renate; Hasbrouck, Nicholas; Jayasuriya, Hiranthi

    2012-05-23

    This paper describes the development of a fast method to screen and confirm methyltestosterone 17-O-glucuronide (MT-glu) in tilapia bile. The method consists of solid-phase extraction (SPE) followed by high-performance liquid chromatography-mass spectrometry. The system used was an Agilent 6530 Q-TOF with an Agilent Jet stream electrospray ionization interface. The glucuronide detected in the bile was characterized as MT-glu by comparison with a chemically synthesized standard. MT-glu was detected in bile for up to 7 days after dosing. Semiquantification was done with matrix-matched calibration curves, because MT-glu showed signal suppression due to matrix effects. This method provides a suitable tool to monitor the illegal use of methyltestosterone in tilapia culture.

  1. Quantification of cannabinoids and their free and glucuronide metabolites in whole blood by disposable pipette extraction and liquid chromatography-tandem mass spectrometry.

    PubMed

    Scheidweiler, Karl B; Newmeyer, Matthew N; Barnes, Allan J; Huestis, Marilyn A

    2016-07-01

    Identifying recent cannabis intake is confounded by prolonged cannabinoid excretion in chronic frequent cannabis users. We previously observed detection times ≤2.1h for cannabidiol (CBD) and cannabinol (CBN) and Δ(9)-tetrahydrocannabinol (THC)-glucuronide in whole blood after smoking, suggesting their applicability for identifying recent intake. However, whole blood collection may not occur for up to 4h during driving under the influence of drugs investigations, making a recent-use marker with a 6-8h detection window helpful for improving whole blood cannabinoid interpretation. Other minor cannabinoids cannabigerol (CBG), Δ9-tetrahydrocannabivarin (THCV), and its metabolite 11-nor-9-carboxy-THCV (THCVCOOH) might also be useful. We developed and validated a sensitive and specific liquid chromatography-tandem mass spectrometry method for quantification of THC, its phase I and glucuronide phase II metabolites, and 5 five minor cannabinoids. Cannabinoids were extracted from 200μL whole blood via disposable pipette extraction, separated on a C18 column, and detected via electrospray ionization in negative mode with scheduled multiple reaction mass spectrometric monitoring. Linear ranges were 0.5-100μg/L for THC and 11-nor-9-carboxy-THC (THCCOOH); 0.5-50μg/L for 11-hydroxy-THC (11-OH-THC), CBD, CBN, and THC-glucuronide; 1-50μg/L for CBG, THCV, and THCVCOOH; and 5-500μg/L for THCCOOH-glucuronide. Inter-day accuracy and precision at low, mid and high quality control (QC) concentrations were 95.1-113% and 2.4-8.5%, respectively (n=25). Extraction recoveries and matrix effects at low and high QC concentrations were 54.0-84.4% and -25.8-30.6%, respectively. By simultaneously monitoring multiple cannabinoids and metabolites, identification of recent cannabis administration or discrimination between licit medicinal and illicit recreational cannabis use can be improved.

  2. Interaction of hesperetin glucuronide conjugates with human BCRP, MRP2 and MRP3 as detected in membrane vesicles of overexpressing baculovirus-infected Sf9 cells.

    PubMed

    Brand, Walter; Oosterhuis, Berend; Krajcsi, Peter; Barron, Denis; Dionisi, Fabiola; van Bladeren, Peter J; Rietjens, Ivonne M C M; Williamson, Gary

    2011-12-01

    The citrus flavonoid hesperetin (4'-methoxy-3',5,7-trihydroxyflavanone) is the aglycone of hesperidin, the major flavonoid present in sweet oranges. Hesperetin 7-O-glucuronide (H7G) and hesperetin 3'-O-glucuronide (H3'G) are the two most abundant metabolites of hesperetin in vivo. In this study, their interaction with specific ABC transporters, believed to play a role in the disposition and bioavailability of hesperetin, was studied using Sf9 membranes from cells overexpressing human BCRP (ABCG2), MRP2 (ABCC2) and MRP3 (ABCC3). Both H7G and H3'G were tested for their potential to activate and inhibit ATPase activity, and to inhibit vesicular transport by these transporters. Both H7G and H3'G demonstrated interaction with all tested ABC transporters, especially with BCRP and MRP3. An interesting difference between H7G and H3'G was seen with respect to the interaction with BCRP: H7G stimulated the ATPase activity of BCRP up to 76% of the maximal effect generated by the reference activator sulfasalazine, with an EC(50) of 0.45 µM, suggesting that H7G is a high affinity substrate of BCRP, whereas H3'G did not stimulate BCRP ATPase activity. Only moderate inhibition of BCRP ATPase activity at high H3'G concentrations was observed. This study provides information on the potential of hesperetin glucuronide conjugates to act as specific ABC transporter substrates or inhibitors and indicates that regio-specific glucuronidation could affect the disposition of hesperetin.

  3. Human UDP-glucuronosyltransferase (UGT) 2B10 in drug N-glucuronidation: substrate screening and comparison with UGT1A3 and UGT1A4.

    PubMed

    Kato, Yukiko; Izukawa, Takeshi; Oda, Shingo; Fukami, Tatsuki; Finel, Moshe; Yokoi, Tsuyoshi; Nakajima, Miki

    2013-07-01

    Recent observations revealed that human UDP-glucuronosyltransferase (UGT) 2B10 catalyzes N-glucuronidation of amine-containing compounds. Knowledge of the substrate specificity and clinical significance of UGT2B10 is still limited. The purpose of this study was to expand the knowledge of UGT2B10 substrates and to evaluate its significance in drug clearance. Using recombinant UGT2B10, we found that it catalyzes the N-glucuronidation of amitriptyline, imipramine, ketotifen, pizotifen, olanzapine, diphenhydramine, tamoxifen, ketoconazole, and midazolam. These are drugs that were previously reported to be substrates for UGT1A4 or UGT1A3, and that contain in their structure either tertiary aliphatic amines, cyclic amines, or an imidazole group. UGT2B10 was inactive in the glucuronidation of desipramine, nortriptyline, carbamazepine, and afloqualone. This group of drugs contains secondary or primary amines, and these results suggest that UGT2B10 preferably conjugates tertiary amines. This preference is partial because UGT2B10 did not conjugate the tertiary cyclic amine in trifluoperazine. Kinetic analyses revealed that the affinity and clearance of UGT2B10 for amitriptyline, imipramine, and diphenhydramine are significantly higher than the corresponding values of UGT1A4 and UGT1A3, although the Vmax values of UGT1A4 toward these drugs are considerably higher. These findings suggest that UGT2B10 plays a major role in the N-glucuronidation of these drugs at therapeutic concentrations. These results are also supported by inhibition studies with nicotine and hecogenin. In conclusion, this study expands the understanding of the substrate specificity of UGT2B10, highlighting its preference for tertiary amines with higher affinities and clearance values than those of UGT1A4 and UGT1A3.

  4. Differences between human and rat intestinal and hepatic bisphenol A glucuronidation and the influence of alamethicin on in vitro kinetic measurements.

    PubMed

    Mazur, Christopher S; Kenneke, John F; Hess-Wilson, Janet K; Lipscomb, John C

    2010-12-01

    The extent to which membrane-disrupting agents, such as alamethicin, may alter cofactor transport and influence in vitro kinetic measurements of glucuronidation is a major concern regarding the characterization and extrapolation of inter- and intraspecies pharmacokinetics of bisphenol A (BPA). An additional concern is the omission of a BPA intestinal metabolism component in current pharmacokinetic models used to assess oral exposure. In this study, BPA glucuronidation in native hepatic microsomes from female rat and female human liver displayed higher V(max) values than that in males. In the presence of alamethicin, all hepatic V(max) values increased; however, this increase was disproportionately greater in males and gender differences were no longer observed. Female rats exhibited a much higher K(m) than all other species and genders; the addition of alamethicin had little influence on K(m) values for any of the test systems. The dissimilar K(m) measured for female rat suggests that different UDP-glucuronosyltransferase (UGT) enzyme(s) are involved in BPA glucuronidation. The presence of different UGTs in female rat was confirmed using Hill coefficients measured from diclofenac-mediated chemical inhibition assays within hepatic microsomes and purified human UGT2B7 and UGT2B15. Mixed-gender human intestinal microsomes showed little BPA glucuronidation reactivity compared with those from male rat intestine. Male rat intestinal microsomes in the presence of alamethicin exhibited a V(max) that was nearly 30-fold higher than that for mixed human microsomes. The species and gender metabolic differences we observed between rat and human liver and intestine provide key information for delineating BPA pharmacokinetics needed for human health risk assessment.

  5. [What ethanol metabolites as biological markers tell us about alcohol use].

    PubMed

    Wurst, Friedrich Martin; Thon, Natasha; Weinmann, Wolfgang; Yegles, Michel; Preuss, Ulrich

    2014-01-01

    Alcohol and tobacco related disorders are the two leading and most expensive causes of illness in central Europe. In addition to self reports and questionnaires, biomarkers are of relevance in diagnosis and therapy of alcohol use disorders. Traditional biomarkers such as gamma glutamyl transpeptidase or mean corpuscular volume are indirect biomarkers and are subject to influence of age, gender and non alcohol related diseases, among others.Direct ethanol metabolites such as ethyl glucuronide (EtG), ethyl sulphate (EtS) and phosphatidylethanol (PEth) are direct metabolites of ethanol, that are positive after intake of ethyl alcohol. They represent useful diagnostic tools for identifying alcohol use even more accurately than traditional biomarkers. Each of these drinking indicators remains positive in serum and urine for a characteristic time spectrum after the cessation of ethanol intake--EtG and EtS in urine up to 7 days, EtG in hair for months after ethanol has left the body. Applications include clinical routine use, emergency room settings, proof of abstinence in alcohol rehabilitation programs, driving under influence offenders, workplace testing, assessment of alcohol intake in the context of liver transplantation and fetal alcohol syndrome.

  6. 77 FR 41346 - Trinexapac-ethyl; Proposed Pesticide Tolerance

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-07-13

    ... AGENCY 40 CFR Part 180 Trinexapac-ethyl; Proposed Pesticide Tolerance AGENCY: Environmental Protection... Division (7505P), Office of Pesticide Programs, Environmental Protection Agency, 1200 Pennsylvania Ave. NW... affected by this action if you are an agricultural producer, food manufacturer, or pesticide...

  7. 21 CFR 177.1320 - Ethylene-ethyl acrylate copolymers.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... blended with polyethylene or with one or more olefin copolymers complying with § 177.1520 or with a mixture of polyethylene and one or more olefin copolymers, in such proportions that the ethyl acrylate... prescribed in paragraph (c)(2) of this section, when tested by the methods prescribed for polyethylene...

  8. 77 FR 60917 - Trinexapac-ethyl; Pesticide Tolerances

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-10-05

    ....662 be amended by establishing tolerances for trinexapac-ethyl in or on barley, bran at 2.5 ppm... barley, bran at 2.5 ppm; sugarcane, molasses at 2.5 ppm; and wheat, bran at 6.0 ppm. In addition, as... commodities: ``Barley, bran'', ``Sugarcane, molasses'', and ``Wheat, bran''. 0 ii. Removing the entry...

  9. Synthesis of Ethyl Nalidixate: A Medicinal Chemistry Experiment

    ERIC Educational Resources Information Center

    Leslie, Ray; Leeb, Elaine; Smith, Robert B.

    2012-01-01

    A series of laboratory experiments that complement a medicinal chemistry lecture course in drug design and development have been developed. The synthesis of ethyl nalidixate covers three separate experimental procedures, all of which can be completed in three, standard three-hour lab classes and incorporate aspects of green chemistry such as…

  10. Reactivity of 2-ethyl-1-hexanol in the atmosphere.

    PubMed

    Gallego-Iniesta García, María Paz; Moreno Sanroma, Alberto; Martín Porrero, María Pilar; Tapia Valle, Araceli; Cabañas Galán, Beatriz; Salgado Muñoz, María Sagrario

    2010-04-07

    Rate coefficients at room temperature for the reaction of 2-ethyl-1-hexanol with OH and NO(3) radicals and with Cl atoms have been determined in a 150 L PTFE chamber using GC-FID/SPME and FTIR as detection systems. The rate coefficients k (in units of cm(3) molecule(-1) s(-1)) obtained were: (1.13 +/- 0.31) 10(-11) for the OH reaction, (2.93 +/- 0.92) 10(-15) for the NO(3) reaction and (1.88 +/- 0.25) 10(-10) for the Cl reaction. Despite the high concentrations of 2-ethyl-1-hexanol, especially in indoor air, this is the first kinetic study carried out to date for these reactions. The results are consistent with the expected reactivity given the chemical structure of 2-ethyl-1-hexanol. Calculated atmospheric lifetimes reveal that the dominant loss process for 2-ethyl-1-hexanol is clearly the daytime reaction with the hydroxyl radical.

  11. Dissociation of the Ethyl Radical: An Exercise in Computational Chemistry

    ERIC Educational Resources Information Center

    Nassabeh, Nahal; Tran, Mark; Fleming, Patrick E.

    2014-01-01

    A set of exercises for use in a typical physical chemistry laboratory course are described, modeling the unimolecular dissociation of the ethyl radical to form ethylene and atomic hydrogen. Students analyze the computational results both qualitatively and quantitatively. Qualitative structural changes are compared to approximate predicted values…

  12. Kinetics of Ethyl Acetate Synthesis Catalyzed by Acidic Resins

    ERIC Educational Resources Information Center

    Antunes, Bruno M.; Cardoso, Simao P.; Silva, Carlos M.; Portugal, Ines

    2011-01-01

    A low-cost experiment to carry out the second-order reversible reaction of acetic acid esterification with ethanol to produce ethyl acetate is presented to illustrate concepts of kinetics and reactor modeling. The reaction is performed in a batch reactor, and the acetic acid concentration is measured by acid-base titration versus time. The…

  13. Pregnane × Receptor (PXR) expression in colorectal cancer cells restricts irinotecan chemosensitivity through enhanced SN-38 glucuronidation

    PubMed Central

    2010-01-01

    Background Clinical efficacy of chemotherapy in colorectal cancer is subjected to broad inter-individual variations leading to the inability to predict outcome and toxicity. The topoisomerase I inhibitor irinotecan (CPT-11) is worldwide approved for the treatment of metastatic colorectal cancer and undergoes extensive peripheral and tumoral metabolism. PXR is a xenoreceptor activated by many drugs and environmental compounds regulating the expression of drug metabolism and transport genes in detoxification organs such as liver and gastrointestinal tract. Considering the metabolic pathway of irinotecan and the tissue distribution of Pregnane × Receptor (PXR), we hypothesized that PXR could play a key role in colon cancer cell response to irinotecan. Results PXR mRNA expression was quantified by RT-quantitative PCR in a panel of 14 colon tumor samples and their matched normal tissues. PXR expression was modulated in human colorectal cancer cells LS174T, SW480 and SW620 by transfection and siRNA strategies. Cellular response to irinotecan and its active metabolic SN38 was assessed by cell viability assays, HPLC metabolic profiles and mRNA quantification of PXR target genes. We showed that PXR was strongly expressed in colon tumor samples and displayed a great variability of expression. Expression of hPXR in human colorectal cancer cells led to a marked chemoresistance to the active metabolite SN38 correlated with PXR expression level. Metabolic profiles of SN38 showed a strong enhancement of SN38 glucuronidation to the inactive SN38G metabolite in PXR-expressing cells, correlated with an increase of UDPglucuronosyl transferases UGT1A1, UGT1A9 and UGT1A10 mRNAs. Inhibition of PXR expression by lentivirus-mediated shRNA, led to SN38 chemoresistance reversion concomitantly to a decrease of UGT1A1 expression and SN38 glucuronidation. Similarly, PXR mRNA expression levels correlated to UGT1A subfamily expression in human colon tumor biopsies. Conclusion Our results

  14. Beta-glucuronidase is not required for transfer of [3H]-estrone-[14C]glucuronide across guinea pig fetal membranes.

    PubMed

    Goldhawk, D E; Hobkirk, R

    1998-07-01

    To understand the means whereby a charged, estrogen conjugate may be transferred across guinea pig amnion and chorion, the permeability to [3H]estrone-[14C]glucuronide was examined at 45 days and near term. No evidence of deconjugation was obtained in either early or late amnion, despite significantly greater transfer near term. Early amnion was virtually impermeable, regardless of ATP depletion. In contrast, early chorion transferred estrone-glucuronide without any requirement for deconjugation or ATP. No effect of tissue orientation was observed in amnion; whereas, incubations from maternal to fetal side of late chorion exhibited beta-glucuronidase activity. Inhibition of the latter demonstrated that hydrolysis was concomitant with but not required for transport. [3H]Estrone produced by deconjugation was enzymatically reduced after pubic symphysis relaxation, although beta-glucuronidase activity began prior to this stage. Transport across late fetal membranes was not saturable and chorion incubations from maternal to fetal side demonstrated a lower transport capacity. In either tissue orientation, late chorion displayed a lower rate of transfer than amnion. These results indicate that fetal membranes possess distinct abilities for transferring intact estrone-glucuronide, depending on stage of development and tissue orientation. The passive nature of transport and its dependence on structural characteristics is consistent with possible regulation of tight junctions.

  15. Determination of Serotonin and Dopamine Metabolites in Human Brain Microdialysis and Cerebrospinal Fluid Samples by UPLC-MS/MS: Discovery of Intact Glucuronide and Sulfate Conjugates

    PubMed Central

    Suominen, Tina; Uutela, Päivi; Ketola, Raimo A.; Bergquist, Jonas; Hillered, Lars; Finel, Moshe; Zhang, Hongbo; Laakso, Aki; Kostiainen, Risto

    2013-01-01

    An UPLC-MS/MS method was developed for the determination of serotonin (5-HT), dopamine (DA), their phase I metabolites 5-HIAA, DOPAC and HVA, and their sulfate and glucuronide conjugates in human brain microdialysis samples obtained from two patients with acute brain injuries, ventricular cerebrospinal fluid (CSF) samples obtained from four patients with obstructive hydrocephalus, and a lumbar CSF sample pooled mainly from patients undergoing spinal anesthesia in preparation for orthopedic surgery. The method was validated by determining the limits of detection and quantification, linearity, repeatability and specificity. The direct method enabled the analysis of the intact phase II metabolites of 5-HT and DA, without hydrolysis of the conjugates. The method also enabled the analysis of the regioisomers of the conjugates, and several intact glucuronide and sulfate conjugates were identified and quantified for the first time in the human brain microdialysis and CSF samples. We were able to show the presence of 5-HIAA sulfate, and that dopamine-3-O-sulfate predominates over dopamine-4-O-sulfate in the human brain. The quantitative results suggest that sulfonation is a more important phase II metabolism pathway than glucuronidation in the human brain. PMID:23826355

  16. Isolation and characterization of a β-glucuronide of hydroxylated SARM S1 produced using a combination of biotransformation and chemical oxidation.

    PubMed

    Rydevik, Axel; Lagojda, Andreas; Thevis, Mario; Bondesson, Ulf; Hedeland, Mikael

    2014-09-01

    In this study, using mass spectrometry and nuclear magnetic resonance (NMR) spectroscopy, it has been confirmed that biotransformation with the fungus Cunninghamella elegans combined with chemical oxidation with the free radical tetramethylpiperidinyl-1-oxy (TEMPO) can produce drug glucuronides of β-configuration. Glucuronic acid conjugates are a common type of metabolites formed by the human body. The detection of such conjugates in doping control and other kinds of forensic analysis would be beneficial owing to a decrease in analysis time as hydrolysis can be omitted. However the commercial availability of reference standards for drug glucuronides is poor. The selective androgen receptor modulator (SARM) SARM S1 was incubated with the fungus C. elegans. The sample was treated with the free radical TEMPO oxidizing agent and was thereafter purified by SPE. A glucuronic acid conjugate was isolated using a fraction collector connected to an ultra high performance liquid chromatographic (UHPLC) system. The isolated compound was characterized by NMR spectroscopy and mass spectrometry and its structure was confirmed as a glucuronic acid β-conjugate of hydroxylated SARM S1 bearing the glucuronide moiety on carbon C-10.

  17. Quantification of phenolic acids and their methylates, glucuronides, sulfates and lactones metabolites in human plasma by LC-MS/MS after oral ingestion of soluble coffee.

    PubMed

    Marmet, Cynthia; Actis-Goretta, Lucas; Renouf, Mathieu; Giuffrida, Francesca

    2014-01-01

    Chlorogenic acids and derivatives like phenolic acids are potentially bioactive phenolics, which are commonly found in many foods. Once absorbed, chlorogenic and phenolic acids are highly metabolized by the intestine and the liver, producing glucuronidated and/or sulphated compounds. These metabolites were analyzed in human plasma using a validated liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) method. After protein precipitation, phenolic acids and their metabolites were extracted by using ethanol and chromatographic separation was achieved by reversed-phase using an Acquity UPLC BEH C18 column combined with a gradient elution system using 1% acetic acid aqueous solution and 1% acetic acid with 100% acetonitrile. The method was able to quantify 56 different compounds including 24 phenolic acids, 4 lactones, 15 sulfates and 13 glucuronides metabolites between 5 and 1000nM in plasma for most of them, except for m-dihydrocoumaric acid, 5-ferulloylquinic-glucuronide, 4-methoxycinnamic acid, 3-phenylpropionic acid, 3-(4-methoxyphenyl)propionic acid (25 to 1000nM) and p-dihydrocoumaric acid (50-1000nM). Values of repeatability and intermediate reproducibility were below 15% of deviation in general, and maximum 20% for the lowest concentrations. The validated method was successfully applied to quantify phenolic acids and their metabolites in plasma obtained after oral ingestion of soluble coffee. In conclusion, the developed and validated method is proved to be very sensitive, accurate and precise for the quantification of these possible dietary phenols.

  18. Accurate Prediction of Glucuronidation of Structurally Diverse Phenolics by Human UGT1A9 Using Combined Experimental and In Silico Approaches

    PubMed Central

    Wu, Baojian; Wang, Xiaoqiang; Zhang, Shuxing; Hu, Ming

    2012-01-01

    Purpose The catalytic selectivity of human UGT1A9, an important membrane-bound enzyme catalyzing glucuronidation of xenobiotics were determined experimentally using 145 phenolics, and analyzed by 3D-QSAR methods. Methods The catalytic efficiency of UGT1A9 was determined by kinetic profiling. Quantitative structure activity relationships were analyzed using the CoMFA and CoMSIA techniques. Molecular alignment of the substrate structures was made by superimposing the glucuronidation site and its adjacent aromatic ring to achieve maximal steric overlap. For a substrate with multiple active glucuronidation sites, each site was considered as a separate substrate. Results The 3D-QSAR analyses produced statistically reliable models with good predictive power (CoMFA: q2 = 0.548, r2= 0.949, r2pred = 0.775; CoMSIA: q2 = 0.579, r2= 0.876, r2pred = 0.700). The contour coefficient maps were applied to elucidate structural features among substrates that are responsible for the selectivity differences. Furthermore, the contour coefficient maps were overlaid in the catalytic pocket of a homology model of UGT1A9; this enabled us to identify the UGT1A9 catalytic pocket with a high degree of confidence. Conclusion The CoMFA/CoMSIA models can predict the substrate selectivity and in vitro clearance of UGT1A9. Our findings also provide a possible molecular basis for understanding UGT1A9 functions and its substrate selectivity. PMID:22302521

  19. 40 CFR 180.430 - Fenoxaprop-ethyl; tolerances for residues.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... residues of the herbicide fenoxaprop-ethyl, including its metabolites and degradates, in or on the...-limited tolerances are established for residues of the herbicide fenoxaprop-ethyl, including...

  20. 40 CFR 180.430 - Fenoxaprop-ethyl; tolerances for residues.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... residues of the herbicide fenoxaprop-ethyl, including its metabolites and degradates, in or on the...-limited tolerances are established for residues of the herbicide fenoxaprop-ethyl, including...

  1. 40 CFR 180.430 - Fenoxaprop-ethyl; tolerances for residues.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... combined residues of the herbicide fenoxaprop-ethyl phenoxy]propanoate] and its metabolites (6-chloro-2... tolerances are established for combined residues of the herbicide fenoxaprop-ethyl, phenoxy]propanoic...

  2. 40 CFR 180.430 - Fenoxaprop-ethyl; tolerances for residues.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... residues of the herbicide fenoxaprop-ethyl, including its metabolites and degradates, in or on the...-limited tolerances are established for residues of the herbicide fenoxaprop-ethyl, including...

  3. Validation and application of a method for the determination of buprenorphine, norbuprenorphine, and their glucuronide conjugates in human meconium.

    PubMed

    Kacinko, Sherri L; Shakleya, Diaa M; Huestis, Marilyn A

    2008-01-01

    A novel liquid chromatography tandem mass spectrometry method for quantification of buprenorphine, norbuprenorphine, and glucuronidated conjugates was developed and validated. Analytes were extracted from meconium using buffer, concentrated by solid-phase extraction and quantified within 13.5 min. In order to determine free and total concentrations, specimens were analyzed with and without enzyme hydrolysis. Calibration was achieved by linear regression with a 1/x weighting factor and deuterated internal standards. All analytes were linear from 20 to 2000 ng/g with a correlation of determination of >0.98. Accuracy was >or=85.7% with intra-assay and interassay imprecisionor=85.0%. There was suppression of ionization by the polar matrix; however, this did not interfere with sensitivity or analyte quantification due to inclusion of deuterated internal standards. Analytes were stable on the autosampler, at room temperature, at 4 degrees C, and when exposed to three freeze/thaw cycles. This sensitive and specific method can be used to monitor in utero buprenorphine exposure and to evaluate correlations, if any, between buprenorphine exposure and neonatal outcomes.

  4. Bidirectional placental transfer of Bisphenol A and its main metabolite, Bisphenol A-Glucuronide, in the isolated perfused human placenta.

    PubMed

    Corbel, T; Gayrard, V; Puel, S; Lacroix, M Z; Berrebi, A; Gil, S; Viguié, C; Toutain, P-L; Picard-Hagen, N

    2014-08-01

    The widespread human exposure to Bisphenol A (BPA), an endocrine disruptor interfering with developmental processes, raises the question of the risk for human health of BPA fetal exposure. In humans, highly variable BPA concentrations have been reported in the feto-placental compartment. However the human fetal exposure to BPA still remains unclear. The aim of the study was to characterize placental exchanges of BPA and its main metabolite, Bisphenol A-Glucuronide (BPA-G) using the non-recirculating dual human placental perfusion. This high placental bidirectional permeability to the lipid soluble BPA strongly suggests a transport by passive diffusion in both materno-to-fetal and feto-to-maternal direction, leading to a calculated ratio between fetal and maternal free BPA concentrations of about 1. In contrast, BPA-G has limited placental permeability, particularly in the materno-to-fetal direction. Thus the fetal exposure to BPA conjugates could be explained mainly by its limited capacity to extrude BPA-G.

  5. Interaction of the electrophilic ketoprofenyl-glucuronide and ketoprofenyl-coenzyme A conjugates with cytosolic glutathione S-transferases.

    PubMed

    Osbild, Sandra; Bour, Jérome; Maunit, Benoît; Guillaume, Cécile; Asensio, Carine; Muller, Jean-François; Netter, Patrick; Kirsch, Glbert; Bagrel, Denyse; Lapicque, Françoise; Battaglia, Eric

    2008-02-01

    Carboxylic acid-containing drugs are metabolized mainly through the formation of glucuronide and coenzyme A esters. These conjugates have been suspected to be responsible for the toxicity of several nonsteroidal anti-inflammatory drugs because of the reactivity of the electrophilic ester bond. In the present study we investigated the reactivity of ketoprofenyl-acylglucuronide (KPF-OG) and ketoprofenyl-acyl-coenzyme A (KPF-SCoA) toward cytosolic rat liver glutathione S-transferases (GST). We observed that KPF-SCoA, but not KPF-OG inhibited the conjugation of 1-chloro-2,4-dinitrobenzene and 4-nitroquinoline N-oxide catalyzed by both purified cytosolic rat liver GST and GST from FAO and H5-6 rat hepatoma cell lines. Photoaffinity labeling with KPF-SCoA suggested that the binding of this metabolite may overlap the binding site of 4-methylumbelliferone sulfate. Furthermore, high-performance liquid chromatography and mass spectrometry analysis showed that both hydrolysis and transacylation reactions were observed in the presence of GST and glutathione. The formation of ketoprofenyl-S-acyl-glutathione could be kinetically characterized (apparent K(m) = 196.0 +/- 70.6 microM). It is concluded that KPF-SCoA is both a GST inhibitor and a substrate of a GST-dependent transacylation reaction. The reactivity and inhibitory potency of thioester CoA derivatives toward GST may have potential implications on the reported in vivo toxicity of some carboxylic acid-containing drugs.

  6. 40 CFR 721.4090 - Ethanaminium, N-[bis(diethylamino)-methylene]-N-ethyl-, bromide.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 30 2010-07-01 2010-07-01 false Ethanaminium, N- -N-ethyl-, bromide... Substances § 721.4090 Ethanaminium, N- -N-ethyl-, bromide. (a) Chemical substance and significant new uses subject to reporting. (1) The chemical substance identified as ethanaminium, N- -N-ethyl-, bromide (PMN...

  7. 40 CFR 721.4090 - Ethanaminium, N-[bis(diethylamino)-methylene]-N-ethyl-, bromide.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 32 2012-07-01 2012-07-01 false Ethanaminium, N- -N-ethyl-, bromide... Substances § 721.4090 Ethanaminium, N- -N-ethyl-, bromide. (a) Chemical substance and significant new uses subject to reporting. (1) The chemical substance identified as ethanaminium, N- -N-ethyl-, bromide (PMN...

  8. 40 CFR 721.4090 - Ethanaminium, N-[bis(diethylamino)-methylene]-N-ethyl-, bromide.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 31 2014-07-01 2014-07-01 false Ethanaminium, N- -N-ethyl-, bromide... Substances § 721.4090 Ethanaminium, N- -N-ethyl-, bromide. (a) Chemical substance and significant new uses subject to reporting. (1) The chemical substance identified as ethanaminium, N- -N-ethyl-, bromide (PMN...

  9. 40 CFR 721.4090 - Ethanaminium, N-[bis(diethylamino)-methylene]-N-ethyl-, bromide.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 32 2013-07-01 2013-07-01 false Ethanaminium, N- -N-ethyl-, bromide... Substances § 721.4090 Ethanaminium, N- -N-ethyl-, bromide. (a) Chemical substance and significant new uses subject to reporting. (1) The chemical substance identified as ethanaminium, N- -N-ethyl-, bromide (PMN...

  10. 40 CFR 721.4250 - Hexanoic acid, 2-ethyl-, ethenyl ester.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 31 2011-07-01 2011-07-01 false Hexanoic acid, 2-ethyl-, ethenyl ester... Substances § 721.4250 Hexanoic acid, 2-ethyl-, ethenyl ester. (a) Chemical substance and significant new uses subject to reporting. (1) The chemical substance identified as hexanoic acid, 2-ethyl-, ethenyl ester...

  11. 40 CFR 721.4250 - Hexanoic acid, 2-ethyl-, ethenyl ester.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 30 2010-07-01 2010-07-01 false Hexanoic acid, 2-ethyl-, ethenyl ester... Substances § 721.4250 Hexanoic acid, 2-ethyl-, ethenyl ester. (a) Chemical substance and significant new uses subject to reporting. (1) The chemical substance identified as hexanoic acid, 2-ethyl-, ethenyl ester...

  12. 40 CFR 721.7290 - Propanoic acid, 2-(trimethoxysilyl)-, ethyl ester.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ...)-, ethyl ester. 721.7290 Section 721.7290 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY... Specific Chemical Substances § 721.7290 Propanoic acid, 2-(trimethoxysilyl)-, ethyl ester. (a) Chemical... acid, 2-(trimethoxysilyl)-, ethyl ester (PMN P-01-22; CAS No. 137787-41-8) is subject to...

  13. 40 CFR 721.10064 - 2-Propenoic acid, 2-[2-(ethenyloxy)ethoxy]ethyl ester.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 31 2011-07-01 2011-07-01 false 2-Propenoic acid, 2- ethyl ester. 721... Substances § 721.10064 2-Propenoic acid, 2- ethyl ester. (a) Chemical substance and significant new uses subject to reporting. (1) The chemical substance identified as 2-propenoic acid, 2- ethyl ester (PMN...

  14. 40 CFR 721.7290 - Propanoic acid, 2-(trimethoxysilyl)-, ethyl ester.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ...)-, ethyl ester. 721.7290 Section 721.7290 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY... Specific Chemical Substances § 721.7290 Propanoic acid, 2-(trimethoxysilyl)-, ethyl ester. (a) Chemical... acid, 2-(trimethoxysilyl)-, ethyl ester (PMN P-01-22; CAS No. 137787-41-8) is subject to...

  15. 40 CFR 721.10064 - 2-Propenoic acid, 2-[2-(ethenyloxy)ethoxy]ethyl ester.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 30 2010-07-01 2010-07-01 false 2-Propenoic acid, 2- ethyl ester. 721... Substances § 721.10064 2-Propenoic acid, 2- ethyl ester. (a) Chemical substance and significant new uses subject to reporting. (1) The chemical substance identified as 2-propenoic acid, 2- ethyl ester (PMN...

  16. 40 CFR 721.10064 - 2-Propenoic acid, 2-[2-(ethenyloxy)ethoxy]ethyl ester.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 31 2014-07-01 2014-07-01 false 2-Propenoic acid, 2- ethyl ester. 721... Substances § 721.10064 2-Propenoic acid, 2- ethyl ester. (a) Chemical substance and significant new uses subject to reporting. (1) The chemical substance identified as 2-propenoic acid, 2- ethyl ester (PMN...

  17. 40 CFR 721.4250 - Hexanoic acid, 2-ethyl-, ethenyl ester.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 32 2013-07-01 2013-07-01 false Hexanoic acid, 2-ethyl-, ethenyl ester... Substances § 721.4250 Hexanoic acid, 2-ethyl-, ethenyl ester. (a) Chemical substance and significant new uses subject to reporting. (1) The chemical substance identified as hexanoic acid, 2-ethyl-, ethenyl ester...

  18. 40 CFR 721.10300 - Benzeneacetic acid, .alpha.-chloro-.alpha.-phenyl-, ethyl ester.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ...-.alpha.-phenyl-, ethyl ester. 721.10300 Section 721.10300 Protection of Environment ENVIRONMENTAL....-phenyl-, ethyl ester. (a) Chemical substance and significant new uses subject to reporting. (1) The chemical substance identified as benzeneacetic acid, .alpha.-chloro-.alpha.-phenyl-, ethyl ester (PMN...

  19. 40 CFR 721.10300 - Benzeneacetic acid, .alpha.-chloro-.alpha.-phenyl-, ethyl ester.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ...-.alpha.-phenyl-, ethyl ester. 721.10300 Section 721.10300 Protection of Environment ENVIRONMENTAL....-phenyl-, ethyl ester. (a) Chemical substance and significant new uses subject to reporting. (1) The chemical substance identified as benzeneacetic acid, .alpha.-chloro-.alpha.-phenyl-, ethyl ester (PMN...

  20. 40 CFR 721.10064 - 2-Propenoic acid, 2-[2-(ethenyloxy)ethoxy]ethyl ester.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 32 2013-07-01 2013-07-01 false 2-Propenoic acid, 2- ethyl ester. 721... Substances § 721.10064 2-Propenoic acid, 2- ethyl ester. (a) Chemical substance and significant new uses subject to reporting. (1) The chemical substance identified as 2-propenoic acid, 2- ethyl ester (PMN...

  1. 40 CFR 721.10365 - Butanoic acid, 3-mercapto-2-methyl-, ethyl ester.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ...-, ethyl ester. 721.10365 Section 721.10365 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY... Specific Chemical Substances § 721.10365 Butanoic acid, 3-mercapto-2-methyl-, ethyl ester. (a) Chemical... acid, 3-mercapto-2-methyl-, ethyl ester (PMN P-10-56; CAS No. 888021-82-7) is subject to...

  2. 40 CFR 721.4250 - Hexanoic acid, 2-ethyl-, ethenyl ester.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 31 2014-07-01 2014-07-01 false Hexanoic acid, 2-ethyl-, ethenyl ester... Substances § 721.4250 Hexanoic acid, 2-ethyl-, ethenyl ester. (a) Chemical substance and significant new uses subject to reporting. (1) The chemical substance identified as hexanoic acid, 2-ethyl-, ethenyl ester...

  3. 40 CFR 721.10300 - Benzeneacetic acid, .alpha.-chloro-.alpha.-phenyl-, ethyl ester.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ...-.alpha.-phenyl-, ethyl ester. 721.10300 Section 721.10300 Protection of Environment ENVIRONMENTAL....-phenyl-, ethyl ester. (a) Chemical substance and significant new uses subject to reporting. (1) The chemical substance identified as benzeneacetic acid, .alpha.-chloro-.alpha.-phenyl-, ethyl ester (PMN...

  4. 40 CFR 721.7290 - Propanoic acid, 2-(trimethoxysilyl)-, ethyl ester.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ...)-, ethyl ester. 721.7290 Section 721.7290 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY... Specific Chemical Substances § 721.7290 Propanoic acid, 2-(trimethoxysilyl)-, ethyl ester. (a) Chemical... acid, 2-(trimethoxysilyl)-, ethyl ester (PMN P-01-22; CAS No. 137787-41-8) is subject to...

  5. 40 CFR 721.4250 - Hexanoic acid, 2-ethyl-, ethenyl ester.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 32 2012-07-01 2012-07-01 false Hexanoic acid, 2-ethyl-, ethenyl ester... Substances § 721.4250 Hexanoic acid, 2-ethyl-, ethenyl ester. (a) Chemical substance and significant new uses subject to reporting. (1) The chemical substance identified as hexanoic acid, 2-ethyl-, ethenyl ester...

  6. 40 CFR 721.7290 - Propanoic acid, 2-(trimethoxysilyl)-, ethyl ester.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ...)-, ethyl ester. 721.7290 Section 721.7290 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY... Specific Chemical Substances § 721.7290 Propanoic acid, 2-(trimethoxysilyl)-, ethyl ester. (a) Chemical... acid, 2-(trimethoxysilyl)-, ethyl ester (PMN P-01-22; CAS No. 137787-41-8) is subject to...

  7. 40 CFR 721.10365 - Butanoic acid, 3-mercapto-2-methyl-, ethyl ester.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ...-, ethyl ester. 721.10365 Section 721.10365 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY... Specific Chemical Substances § 721.10365 Butanoic acid, 3-mercapto-2-methyl-, ethyl ester. (a) Chemical... acid, 3-mercapto-2-methyl-, ethyl ester (PMN P-10-56; CAS No. 888021-82-7) is subject to...

  8. 40 CFR 721.10064 - 2-Propenoic acid, 2-[2-(ethenyloxy)ethoxy]ethyl ester.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 32 2012-07-01 2012-07-01 false 2-Propenoic acid, 2- ethyl ester. 721... Substances § 721.10064 2-Propenoic acid, 2- ethyl ester. (a) Chemical substance and significant new uses subject to reporting. (1) The chemical substance identified as 2-propenoic acid, 2- ethyl ester (PMN...

  9. 40 CFR 721.10365 - Butanoic acid, 3-mercapto-2-methyl-, ethyl ester.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ...-, ethyl ester. 721.10365 Section 721.10365 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY... Specific Chemical Substances § 721.10365 Butanoic acid, 3-mercapto-2-methyl-, ethyl ester. (a) Chemical... acid, 3-mercapto-2-methyl-, ethyl ester (PMN P-10-56; CAS No. 888021-82-7) is subject to...

  10. 40 CFR 721.7290 - Propanoic acid, 2-(trimethoxysilyl)-, ethyl ester.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ...)-, ethyl ester. 721.7290 Section 721.7290 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY... Specific Chemical Substances § 721.7290 Propanoic acid, 2-(trimethoxysilyl)-, ethyl ester. (a) Chemical... acid, 2-(trimethoxysilyl)-, ethyl ester (PMN P-01-22; CAS No. 137787-41-8) is subject to...

  11. 40 CFR 180.430 - Fenoxaprop-ethyl; tolerances for residues.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... residues of the herbicide fenoxaprop-ethyl, including its metabolites and degradates, in or on the...-limited tolerances are established for residues of the herbicide fenoxaprop-ethyl, including its... established for residues of the herbicide fenoxaprop-ethyl, including its metabolites and degradates, in or...

  12. 76 FR 82320 - Ethyl Alcohol for Fuel Use: Determination of the Base Quantity of Imports

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-12-30

    ... COMMISSION Ethyl Alcohol for Fuel Use: Determination of the Base Quantity of Imports AGENCY: United States.... domestic market for fuel ethyl alcohol during the 12-month period ending on the preceding September 30. This determination is to be used to establish the ``base quantity'' of imports of fuel ethyl...

  13. 78 FR 9938 - Ethyl Alcohol for Fuel Use: Determination of the Base Quantity of Imports

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-02-12

    ... COMMISSION Ethyl Alcohol for Fuel Use: Determination of the Base Quantity of Imports AGENCY: United States... is equal to 7 percent of the U.S. domestic market for fuel ethyl alcohol during the 12-month period...'' of imports of fuel ethyl alcohol, and the Commission transmitted it determinations to the...

  14. 40 CFR 721.9514 - Ethyl silicate, reaction products with modified alkoxysilane salt (generic).

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 32 2012-07-01 2012-07-01 false Ethyl silicate, reaction products with... Significant New Uses for Specific Chemical Substances § 721.9514 Ethyl silicate, reaction products with.... (1) The chemical substance identified generically as Ethyl silicate, reaction products with...

  15. 40 CFR 721.9514 - Ethyl silicate, reaction products with modified alkoxysilane salt (generic).

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 31 2014-07-01 2014-07-01 false Ethyl silicate, reaction products with... Significant New Uses for Specific Chemical Substances § 721.9514 Ethyl silicate, reaction products with.... (1) The chemical substance identified generically as Ethyl silicate, reaction products with...

  16. 40 CFR 721.9514 - Ethyl silicate, reaction products with modified alkoxysilane salt (generic).

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 32 2013-07-01 2013-07-01 false Ethyl silicate, reaction products with... Significant New Uses for Specific Chemical Substances § 721.9514 Ethyl silicate, reaction products with.... (1) The chemical substance identified generically as Ethyl silicate, reaction products with...

  17. 40 CFR 721.9514 - Ethyl silicate, reaction products with modified alkoxysilane salt (generic).

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 31 2011-07-01 2011-07-01 false Ethyl silicate, reaction products with... Significant New Uses for Specific Chemical Substances § 721.9514 Ethyl silicate, reaction products with.... (1) The chemical substance identified generically as Ethyl silicate, reaction products with...

  18. 40 CFR 721.9514 - Ethyl silicate, reaction products with modified alkoxysilane salt (generic).

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 30 2010-07-01 2010-07-01 false Ethyl silicate, reaction products with... Significant New Uses for Specific Chemical Substances § 721.9514 Ethyl silicate, reaction products with.... (1) The chemical substance identified generically as Ethyl silicate, reaction products with...

  19. 40 CFR 721.4090 - Ethanaminium, N-[bis(diethylamino)-methylene]-N-ethyl-, bromide.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 31 2011-07-01 2011-07-01 false Ethanaminium, N- -N-ethyl-, bromide... Substances § 721.4090 Ethanaminium, N- -N-ethyl-, bromide. (a) Chemical substance and significant new uses subject to reporting. (1) The chemical substance identified as ethanaminium, N- -N-ethyl-, bromide (PMN...

  20. Homo- and hetero-dimerization of human UDP-glucuronosyltransferase 2B7 (UGT2B7) wild type and its allelic variants affect zidovudine glucuronidation activity.

    PubMed

    Yuan, Lingmin; Qian, Sainan; Xiao, Yongsheng; Sun, Hongying; Zeng, Su

    2015-05-01

    Most human UDP-glucuronosyltransferase (UGT; EC 2.4.1.17) genes contain non-synonymous single nucleotide polymorphisms (nsSNPs) which cause amino acid substitutions. Allelic variants caused by nsSNPs may exhibit absent or reduced enzyme activity. UGT2B7 is one of the most important UGTs that glucuronidates abundant endobiotics and xenobiotics, such as estriol, morphine, and anticancer drugs. Three nsSNPs, UGT2B7*71S (211G>T), UGT2B7*2 (802C>T) and UGT2B7*5 (1192G>A) are observed in the UGT2B7 gene, and they code for allozymes UGT2B7*71S (A71S), UGT2B7*2 (H268Y), and UGT2B7*5 (D398N). UGT2B7 has been observed to form oligomers that affect its enzymatic activity and in this study, we investigated protein-protein interactions among UGT2B7 allozymes wild type (WT), A71S, H268Y and D398N, by performing a systematic quantitative fluorescence resonance energy transfer (FRET) analysis in combination with co-immunoprecipitation assay. Quantitative FRET analysis revealed that UGT2B7 allozymes formed homo- and hetero-dimers and showed distinct features in donor-acceptor distances. Both codon 71 and codon 268 in the N-terminal domain were involved in the dimeric interaction. Co-immunoprecipitation experiments also proved that UGT2B7 allozymes formed stable dimers. The glucuronidation activities of homo- and hetero-dimers were further tested with zidovudine as the substrate. An increase in activity was observed when WT hetero-dimerized with A71S compared with homo-dimers, while both H268Y and D398N impaired the activity of WT and A71S by forming hetero-dimers. In addition, zidovudine glucuronidation activity is associated with FRET distance. These findings provide insights into the consequences of amino acid substitution in UGT2B7 on zidovudine glucuronidation and the association between protein-protein interaction and glucuronidation activity.

  1. [A study on the mechanism of reductive alkylation for preparing 3-(beta-hydroxy-ethyl-sulfonyl) N-ethyl aniline with HPLC/MS].

    PubMed

    Zhang, R; Wu, Z W; Lin, L S; Yang, H Y

    2000-11-01

    Hydrogenating 3-(beta-hydroxy-ethyl-sulfonyl)-aniline and acetaldehyde in the presence of Raney Nickel as a catalyst, 3-(beta-hydroxy-ethyl-sulfonyl)-N-ethyl-aniline was obtained with 98% conversion and 95% monoalkylation selectivity under optimum conditions. By using high performance liquid chromatography/mass selective detection technique to characterize the structures of the products, the mechanism of reductive alkylation is proposed. From the intermediates determined, it is shown that the reaction mechanism would go via an unstable N-alpha-hydroxyethylaniline derivative and Schiff base stage. After hydrogenation of Schiff base, finally the product 3-(beta-hydroxyethyl-sulfonyl)-N-ethyl aniline was formed.

  2. Elevated plasma creatinine due to creatine ethyl ester use.

    PubMed

    Velema, M S; de Ronde, W

    2011-02-01

    Creatine is a nutritional supplement widely used in sport, physical fitness training and bodybuilding. It is claimed to enhance performance. We describe a case in which serum creatinine is elevated due to the use of creatine ethyl esther. One week after withdrawal, the plasma creatinine had normalised. There are two types of creatine products available: creatine ethyl esther (CEE) and creatine monohydrate (CM). Plasma creatinine is not elevated in all creatine-using subjects. CEE , but not CM, is converted into creatinine in the gastrointestinal tract. As a result the use of CEE may be associated with elevated plasma creatinine levels. Since plasma creatinine is a widely used marker for renal function, the use of CEE may lead to a false assumption of renal failure.

  3. Fragrance material review on 2-ethyl-1-butanol.

    PubMed

    McGinty, D; Letizia, C S; Api, A M

    2010-07-01

    A toxicologic and dermatologic review of 2-ethyl-1-butanol when used as a fragrance ingredient is presented. 2-Ethyl-1-butanol is a member of the fragrance structural group branched chain saturated alcohols. The common characteristic structural elements of the alcohols with saturated branched chain are one hydroxyl group per molecule, and a C(4)-C(12) carbon chain with one or several methyl side chains. This review contains a detailed summary of all available toxicology and dermatology papers that are related to this individual fragrance ingredient and is not intended as a stand-alone document. A safety assessment of the entire branched chain saturated alcohol group will be published simultaneously with this document; please refer to Belsito et al. (2010) for an overall assessment of the safe use of this material and all other branched chain saturated alcohols in fragrances.

  4. Identification of an antioxidant, ethyl protocatechuate, in peanut seed testa.

    PubMed

    Huang, Shiow Chyn; Yen, Gow-Chin; Chang, Lee-Wen; Yen, Wen-Jye; Duh, Pin-Der

    2003-04-09

    The antioxidant activity and identification of the antioxidant component of peanut seed testa were investigated. The antioxidant activity of peanut seed testa was studied in the linoleic acid model system by using the ferric thiocyanate method. Among the five organic solvent extracts, the ethanolic extracts of peanut seed testa (EEPST) produced higher yields and stronger antioxidant activity than other organic solvent extracts. EEPST was separated into 17 fractions on silica gel column chromatography. Fraction 17, which showed the largest yield and significant antioxidant activity, was separated by thin-layer chromatography. Four major antioxidative subfractions were present. Subfraction 17-2 was found to be effective in preventing oxidation of linoleic acid. This subfraction was further fractionated and isolated and characterized by UV, MS, IR, and (1)H NMR techniques. The active compound was identified as ethyl protocatechuate (3,4-dihydroxybenzoic acid ethyl ester).

  5. Fragrance material review on 2-ethyl-1-hexanol.

    PubMed

    McGinty, D; Scognamiglio, J; Letizia, C S; Api, A M

    2010-07-01

    A summary of the safety data available for 2-ethyl-1-hexanol when used as a fragrance ingredient is presented. 2-Ethyl-1-hexanol is a member of the fragrance structural group branched chain saturated alcohols in which the common characteristic structural element is one hydroxyl group per molecule, and a C(4) to C(12) carbon chain with one or several methyl side chains. This review contains a detailed summary of all available toxicology and dermatology papers that are related to this individual fragrance ingredient and is not intended as a stand-alone document. A safety assessment of the entire branched chain saturated alcohol group will be published simultaneously with this document; please refer to Belsito et al. (2010) for an overall assessment of the safe use of this material and all other branched chain saturated alcohols in fragrances.

  6. Highly efficient palladium-catalyzed hydrostannation of ethyl ethynyl ether.

    PubMed

    Andrews, Ian P; Kwon, Ohyun

    2008-12-08

    The palladium-catalyzed hydrostannation of acetylenes is widely exploited in organic synthesis as a means of forming vinyl stannanes for use in palladium-catalyzed cross-coupling reactions. Application of this methodology to ethyl ethynyl ether results in an enol ether that is challenging to isolate from the crude reaction mixture because of incompatibility with typical silica gel chromatography. Reported here is a highly efficient procedure for the palladium-catalyzed hydrostannation of ethyl ethynyl ether using 0.1% palladium(0) catalyst and 1.0 equiv of tributyltin hydride. The product obtained is a mixture of regioisomers that can be carried forward with exclusive reaction of the beta-isomer. This method is highly reproducible; relative to previously reported procedures, it is more economical and involves a more facile purification procedure.

  7. SPECTROSCOPIC CHARACTERIZATION AND DETECTION OF ETHYL MERCAPTAN IN ORION

    SciTech Connect

    Kolesniková, L.; Alonso, J. L.; Daly, A. M.; Tercero, B.; Cernicharo, J.; Gordon, B. P.; Shipman, S. T. E-mail: jlalonso@qf.uva.es E-mail: terceromb@cab.inta-csic.es E-mail: brittany.gordon@ncf.edu

    2014-03-20

    New laboratory data of ethyl mercaptan, CH{sub 3}CH{sub 2}SH, in the millimeter- and submillimeter-wave domains (up to 880 GHz) provided very precise values of the spectroscopic constants that allowed the detection of gauche-CH{sub 3}CH{sub 2}SH toward Orion KL. This identification is supported by 77 unblended or slightly blended lines plus no missing transitions in the range 80-280 GHz. A detection of methyl mercaptan, CH{sub 3}SH, in the spectral survey of Orion KL is reported as well. Our column density results indicate that methyl mercaptan is ≅ 5 times more abundant than ethyl mercaptan in the hot core of Orion KL.

  8. Purification and Characterization of Methyl Phthalyl Ethyl Glycolate (MPEG)

    DTIC Science & Technology

    2014-11-21

    plasticizer, HES 5808 propellant, purification, 1H NMR, 13C NMR, IR, UV- Vis, bp, HRMS, GC, GC-MS, EA, KF, flash column chromatography , dimethylformamide...We report the purification of methyl phthalyl ethyl glycolate (MPEG) by flash column chromatography (ca. grams), allowing us to establish an...analysis Et3N Triethylamine Et2O Diethyl ether IR Infrared spectroscopy GC Capillary gas chromatography GC-MS Capillary gas chromatography mass

  9. Bioequivalence Demonstration for Ω-3 Acid Ethyl Ester Formulations: Rationale for Modification of Current Guidance.

    PubMed

    Maki, Kevin C; Johns, Colleen; Harris, William S; Puder, Mark; Freedman, Steven D; Thorsteinsson, Thorsteinn; Daak, Ahmed; Rabinowicz, Adrian L; Sancilio, Frederick D

    2017-02-08

    The US Food and Drug Administration (FDA) draft guidance for establishing bioequivalence (BE) of ω-3 acid ethyl esters (containing both eicosapentaenoic acid [EPA] and docosahexaenoic acid [DHA] as ethyl esters), used to treat severe hypertriglyceridemia, recommends the conduct of 2 studies: one with participants in the fasting state and one with participants in the fed state. For the fasting study, the primary measures of BE are baseline-adjusted EPA and DHA levels in total plasma lipids. For the fed study, the primary measures of BE are EPA and DHA ethyl esters in plasma. This guidance differs from that established for icosapent ethyl (EPA ethyl esters) in which the primary measure of BE is baseline-adjusted total EPA in plasma lipids for both the fasting and fed states. The FDA guidance for ω-3 acid ethyl esters is not supported by their physiologic characteristics and triglyceride-lowering mechanisms because EPA and DHA ethyl esters are best characterized as pro-drugs. This article presents an argument for amending the FDA draft guidance for ω-3 acid ethyl esters to use baseline-adjusted EPA and DHA in total plasma lipids as the primary measures of BE for both fasting and fed conditions. This change would harmonize the approaches for demonstration of BE for ω-3 acid ethyl esters and icosapent ethyl (EPA ethyl esters) products for future development programs and is the most physiologically rational approach to BE testing.

  10. Sensory reception of the primer pheromone ethyl oleate

    NASA Astrophysics Data System (ADS)

    Muenz, Thomas S.; Maisonnasse, Alban; Plettner, Erika; Le Conte, Yves; Rössler, Wolfgang

    2012-05-01

    Social work force distribution in honeybee colonies critically depends on subtle adjustments of an age-related polyethism. Pheromones play a crucial role in adjusting physiological and behavioral maturation of nurse bees to foragers. In addition to primer effects of brood pheromone and queen mandibular pheromone—both were shown to influence onset of foraging—direct worker-worker interactions influence adult behavioral maturation. These interactions were narrowed down to the primer pheromone ethyl oleate, which is present at high concentrations in foragers, almost absent in young bees and was shown to delay the onset of foraging. Based on chemical analyses, physiological recordings from the antenna (electroantennograms) and the antennal lobe (calcium imaging), and behavioral assays (associative conditioning of the proboscis extension response), we present evidence that ethyl oleate is most abundant on the cuticle, received by olfactory receptors on the antenna, processed in glomeruli of the antennal lobe, and learned in olfactory centers of the brain. The results are highly suggestive that the primer pheromone ethyl oleate is transmitted and perceived between individuals via olfaction at close range.

  11. Separation optimization for the recovery of phenyl ethyl alcohol.

    PubMed

    Priddy, S A; Hanley, T R; Effler, W T

    1999-01-01

    Phenyl ethyl alcohol is a compound that occurs naturally in flower petals and in many common beverages, such as beer. Desire for the floral, rose-like notes imparted by phenyl ethyl alcohol has created a unique niche for this chemical in flavor and fragrance industries. Phenyl ethyl alcohol can be produced by Saccharomyces cerevisiae via bioconversion. Often this method of production results in extremely low yields, thus placing a great deal of importance on recovery and purification of the valuable metabolite. To determine the best method for recovering the chemical, a primary recovery step and a secondary recovery step were developed. The primary recovery step consisted of comparing dead-end filtration with crossflow ultrafiltration. Crossflow ultrafiltration was ultimately selected to filter the fermentation broth because of its high flow rates and low affinity for the product. The secondary recovery step consisted of a comparison of liquid- liquid extraction and hydrophobic resin recovery. The hydrophobic resin was selected because of its higher rate of recovery and a higher purity than the liquid-liquid extraction, the current practice of Brown-Forman.

  12. Photodissociation dynamics of ethyl ethynyl ether: A ketenyl radical precursor

    NASA Astrophysics Data System (ADS)

    Krisch, Maria; Miller, Johanna; Butler, Laurie; Su, Hongmei; Bersohn, Richard; Shu, Jinian

    2006-03-01

    We investigate the photodissociation dynamics of ethyl ethynyl ether at 193.3 nm with crossed laser-molecular beam photofragment translational spectroscopy and laser-induced fluorescence. We establish ethyl ethynyl ether as the first clean precursor to the ketenyl radical, a key species in combustion reactions. One major bond fission channel was observed for the system, cleavage along the HCCO-C2H5 bond, leading to ground state C2H5 (ethyl) radicals and HCCO (ketenyl) radical products in two distinct electronic states. We observed neither cleavage of the other C-O bond nor molecular elimination to form C2H4 + CH2CO (ketene). Ketenyl radicals formed in the higher recoil kinetic energy channel could be either X(^2A") or Ã(^2A') state ketenyl radical. We assign the lower recoil kinetic energy channel to the spin forbidden ã(^4A") state of the ketenyl radical, reached through intersystem crossing. Laser-induced fluorescence from the ketenyl radical peaks after a 20 μs delay, indicating that it is formed with a significant amount of internal energy and subsequently relaxes to the lowest vibrational level of the ground electronic state, a result consistent with the product assignment.

  13. IRIS Toxicological Review of Ethyl Tertiary Butyl Ether (Etbe) ...

    EPA Pesticide Factsheets

    In September 2016, EPA released the draft IRIS Toxicological Review of Ethyl Tertiary Butyl Ether (ETBE) for public comment and discussion. The draft assessment was reviewed internally by EPA and by other federal agencies and White House Offices before public release. Consistent with the May 2009 IRIS assessment development process, all written comments on IRIS assessments submitted by other federal agencies and White House Offices are made publicly available. Accordingly, interagency comments and the interagency science consultation materials provided to other agencies, including interagency review drafts of the IRIS Toxicological Review of Ethyl Tertiary Butyl Ether are posted on this site. EPA is undertaking an new health assessment for ethyl tertiary butyl ether (ETBE) for the Integrated Risk Information System (IRIS). The outcome of this project will be a Toxicological Review and IRIS Summary of ETBE that will be entered on the IRIS database. IRIS is an EPA database containing Agency scientific positions on potential adverse human health effects that may result from chronic (or lifetime) exposure to chemicals in the environment. IRIS contains chemical-specific summaries of qualitative and quantitative health information in support of two steps of the risk assessment process, i.e., hazard identification and dose-response evaluation. IRIS assessments are used nationally and internationally in combination with specific situational exposure assessment infor

  14. Ethyl brings MMT to market with aggressive sales efforts

    SciTech Connect

    1996-01-01

    Wasting no time in seizing a market opportunity that has been on hold for nearly two decades, Ethyl Corp. recently delivered MMT to refiners across the US immediately after the US EPA decided on Dec. 4, 1995, not to appeal court decisions giving Ethyl the right to market the gasoline additive in conventional gasoline. Among the refiners reportedly taking deliveries of MMT were Texaco and Marathon, though spokepersons from each company would not comment. With the last immediate roadblock to the sale of MMT in the US removed, Ethyl anticipates achieving a market penetration of 8 million-10 million lb annually. With MMT allowed in conventional gasoline at 1/32 gram per gallon, 10 million lb would represent blending the octane enhancer into 948,000 b/d of gasoline. Blending MMT into nearly a million b/d of gasoline would represent a market share of about 20% of the eligible gasoline-i.e., the 5.5 million b/d that is not RFG. That 20% figure also corresponds to the approximate share of the gasoline market controlled by independent refiners.

  15. Contribution of citrulline to the formation of ethyl carbamate during Chinese rice wine production.

    PubMed

    Wang, Peihong; Sun, Junyong; Li, Xiaomin; Wu, Dianhui; Li, Tong; Lu, Jian; Chen, Jian; Xie, Guangfa

    2014-04-01

    Ethyl carbamate is a well-known carcinogen and widely occurs in Chinese rice wine. To provide more clues to minimise ethyl carbamate accumulation, the levels of possible precursors of ethyl carbamate in Chinese rice wine were investigated by HPLC. Studies of the possible precursors of ethyl carbamate in Chinese raw rice wine with various additives and treatments indicated that significant amounts of urea can account for ethyl carbamate formation. It was also recognised that citrulline is another important precursor that significantly affects ethyl carbamate production during the boiling procedure used in the Chinese rice wine manufacturing process. Besides urea and citrulline, arginine was also found to be an indirect ethyl carbamate precursor due to its ability to form urea and citrulline by microorganism metabolism.

  16. Estimating Driver Risk Using Alcohol Biomarkers, Interlock BAC Tests and Psychometric Assessments: Initial Descriptives

    PubMed Central

    Marques, Paul; Tippetts, Scott; Allen, John; Javors, Martin; Alling, Christer; Yegles, Michel; Pragst, Fritz; Wurst, Friedrich

    2009-01-01

    Aim To identify alcohol biomarker and psychometric measures that relate to drivers’ blood alcohol concentration (BAC) patterns from ignition interlock devices (IIDs). Design, Setting, Participants, Measurements In Alberta, Canada, 534 drivers, convicted of driving under the influence of alcohol (DUI), installed IIDs and agreed to participate in a research study. IID BAC tests are an established proxy for predicting future DUI convictions. Three risk groups were defined by rates of failed BAC tests. Program entry and followup blood samples (n=302, 171) were used to measure phosphatidyl ethanol (PETH), carbohydrate deficient transferrin (%CDT), gamma glutamyltransferase (GGT) and other biomarkers. Program entry urine (n=130) was analyzed for ethyl glucuronide (ETG) and ethyl sulfate (ETS). Entry hair samples were tested for fatty acid ethyl esters (FAEE) (n=92) and ETG (n=146). Psychometric measures included the DSM-4 Diagnostic Interview Schedule Alcohol Module, Alcohol Use Disorders Identification Test (AUDIT), the Timeline Followback (TLFB), the Drinker Inventory of Consequences (DRINC), and the Temptation and Restraint Inventory (TRI). Findings Except for FAEE, all alcohol biomarkers were significantly related to the interlock BAC test profiles; higher marker levels predicted higher rates of interlock BAC test failures. PETH, the strongest with an overall ANOVA F ratio of 35.5, had significant correlations with all nine of the other alcohol biomarkers and with 16 of 19 psychometric variables. Urine ETG and ETS were strongly correlated with the IID BAC tests. Conclusions The findings suggest several alcohol biomarkers and assessments could play an important role in the prediction and control of driver alcohol risk when relicensing. PMID:19922520

  17. Production of ethyl (R)-2-hydroxy-4-phenylbutanoate via reduction of ethyl 2-oxo-4-phenylbutanoate in an interface bioreactor.

    PubMed

    Oda, S; Inada, Y; Kobayashi, A; Ohta, H

    1998-09-01

    Ethyl (R)-2-hydroxy-4-phenylbutanoate [(R)-EHPB], a useful intermediate for the synthesis of various anti-hypertension drugs, was produced via microbial reduction of ethyl 2-oxo-4-phenylbutanoate [EOPB] in an interface bioreactor. Rhodotorula minuta IFO 0920 and Candida holmii KPY 12402 were selected as the best type culture and isolated yeasts, respectively. The highest enantiomeric excess of (R)-EHPB produced by R. minuta and C. holmii were 95 and 94%, respectively. C. holmii was used for the reduction of EOPB in a pad-packed interface bioreactor (inner volume, 3 liter). After incubation for 4 days, 4.4 g of (R)-EHPB was obtained via extraction with methanol followed by column chromatography. The overall yield, chemical purity, and enantiomeric excess of (R)-EHPB were 58%, 99.1%, and 90%, respectively.

  18. The TLR4-Active Morphine Metabolite Morphine-3-Glucuronide Does Not Elicit Macrophage Classical Activation In Vitro

    PubMed Central

    Khabbazi, Samira; Xie, Nan; Pu, Wenjun; Goumon, Yannick; Parat, Marie-Odile

    2016-01-01

    Macrophages are abundant in the tumor microenvironment where they adopt a pro-tumor phenotype following alternative polarization induced by paracrine factors from cancer and stromal cells. In contrast, classically activated macrophages have tumoricidal activities, such that the polarization of tumor-associated macrophages has become a novel therapeutic target. Toll-like receptor 4 engagement promotes classical activation of macrophages, and recent literature suggests TLR4 agonism to prevent metastasis and promote survival in experimental metastasis models. A growing number of studies indicate that TLR4 can respond to opioids, including the opioid receptor-inactive morphine metabolite morphine-3-glucuronide (M3G). We measured the activation of TLR4 in a reporter cell line exogenously expressing TLR4 and TLR4 co-receptors, and confirmed that M3G weakly but significantly activates TLR4. We hypothesized that M3G would promote the expression of classical activation signature genes in macrophages in vitro. We exposed mouse and human macrophage cell lines to M3G or the TLR4 activator lipopolysaccharide (LPS), alone or in combination with interferon gamma (IFN-γ). The classical macrophage activation markers tested were iNOS, CD86, IL-6, or TNF-α in RAW 264.7 cells and IL-6, IL-12, IL-23, TNF-α, CXCL10, and CXCL11 in THP1 cells. Our results show that despite exhibiting TLR4-activation ability, M3G does not elicit the expression of classical activation markers in LPS-responsive macrophages. PMID:27909407

  19. The TLR4-Active Morphine Metabolite Morphine-3-Glucuronide Does Not Elicit Macrophage Classical Activation In Vitro.

    PubMed

    Khabbazi, Samira; Xie, Nan; Pu, Wenjun; Goumon, Yannick; Parat, Marie-Odile

    2016-01-01

    Macrophages are abundant in the tumor microenvironment where they adopt a pro-tumor phenotype following alternative polarization induced by paracrine factors from cancer and stromal cells. In contrast, classically activated macrophages have tumoricidal activities, such that the polarization of tumor-associated macrophages has become a novel therapeutic target. Toll-like receptor 4 engagement promotes classical activation of macrophages, and recent literature suggests TLR4 agonism to prevent metastasis and promote survival in experimental metastasis models. A growing number of studies indicate that TLR4 can respond to opioids, including the opioid receptor-inactive morphine metabolite morphine-3-glucuronide (M3G). We measured the activation of TLR4 in a reporter cell line exogenously expressing TLR4 and TLR4 co-receptors, and confirmed that M3G weakly but significantly activates TLR4. We hypothesized that M3G would promote the expression of classical activation signature genes in macrophages in vitro. We exposed mouse and human macrophage cell lines to M3G or the TLR4 activator lipopolysaccharide (LPS), alone or in combination with interferon gamma (IFN-γ). The classical macrophage activation markers tested were iNOS, CD86, IL-6, or TNF-α in RAW 264.7 cells and IL-6, IL-12, IL-23, TNF-α, CXCL10, and CXCL11 in THP1 cells. Our results show that despite exhibiting TLR4-activation ability, M3G does not elicit the expression of classical activation markers in LPS-responsive macrophages.

  20. Quantitative determination of free and total bisphenol A in human urine using labeled BPA glucuronide and isotope dilution mass spectrometry.

    PubMed

    Kubwabo, Cariton; Kosarac, Ivana; Lalonde, Kaela; Foster, Warren G

    2014-07-01

    Bisphenol A (BPA) is a widely used industrial chemical in the manufacturing of polycarbonate plastic bottles, food and beverage can linings, thermal receipts, and dental sealants. Animal and human studies suggest that BPA may disrupt normal hormonal function and hence, potentially, have negative effects on the human health. While total BPA is frequently reported, it is recognized that free BPA is the biologically active form and is rarely reported in the literature. The objective of this study was to develop a sensitive and improved method for the measurement of free and total BPA in human urine. Use of a labeled conjugated BPA (bisphenol A-d6 β-D-glucuronide) allowed for the optimization of the enzymatic reaction and permitted an accurate determination of the conjugated BPA concentration in urine samples. In addition, a (13)C12-BPA internal standard was used to account for the analytical recoveries and performance of the isotope dilution method. Solid-phase extraction (SPE) combined with derivatization and analysis using a triple quadrupole GC-EI/MS/MS system achieved very low method detection limit of 0.027 ng/mL. BPA concentrations were measured in urine samples collected during the second and third trimesters of pregnancy in 36 Canadian women. Total maternal BPA concentrations in urine samples ranged from not detected to 9.40 ng/mL (median, 1.21 ng/mL), and free BPA concentrations ranged from not detected to 0.950 ng/mL (median, 0.185 ng/mL). Eighty-six percent of the women had detectable levels of conjugated BPA, whereas only 22 % had detectable levels of free BPA in their urine. BPA levels measured in this study agreed well with data reported internationally.

  1. Predictability of idiosyncratic drug toxicity risk for carboxylic acid-containing drugs based on the chemical stability of acyl glucuronide.

    PubMed

    Sawamura, Ryoko; Okudaira, Noriko; Watanabe, Kengo; Murai, Takahiro; Kobayashi, Yoshimasa; Tachibana, Masaya; Ohnuki, Takashi; Masuda, Kayoko; Honma, Hidehito; Kurihara, Atsushi; Okazaki, Osamu

    2010-10-01

    Acyl glucuronides (AGs) formed from carboxylic acid-containing drugs have been considered to be a cause of idiosyncratic drug toxicity (IDT). Chemical stability of AGs is supposed to relate to their reactivity. In this study, the half-lives of 21 AGs of carboxylic drugs in potassium phosphate buffer (KPB), human serum albumin (HSA) solution, and human fresh plasma were analyzed in relation to the IDT risk derived from these drugs. The carboxylic drugs were classified into three safety categories of "safe," "warning," and "withdrawn" in terms of their IDT risk. As for the results, the half-lives of AGs in KPB correlated with the IDT risk better than those in HSA solution or in human fresh plasma with regard to the separation of the safe drugs from the warning drugs or the withdrawn drugs. In KPB, whereas the half-lives in the safe category were 7.2 h or longer, those in the withdrawn category were 1.7 h or shorter. The classification value of the half-life in KPB, which separated the safe drugs from the withdrawn drugs was calculated to be 3.6 h by regression analysis. In conclusion, this is the first report that clearly shows the relationship between the IDT risk and chemical stability of AGs in several in vitro systems. The KPB system was considered to be the best for evaluating the stability of AGs, and the classification value of the half-life in KPB serves as a useful key predictor for the IDT risk.

  2. Elevated concentrations of morphine 6-beta-D-glucuronide in brain extracellular fluid despite low blood–brain barrier permeability

    PubMed Central

    Stain-Texier, Frédérique; Boschi, Gabrielle; Sandouk, Pierre; Scherrmann, Jean-Michel

    1999-01-01

    This study was done to find out how morphine 6-beta-D-glucuronide (M6G) induces more potent central analgesia than morphine, despite its poor blood–brain barrier (BBB) permeability. The brain uptake and disposition of these compounds were investigated in plasma and in various brain compartments: extracellular fluid (ECF), intracellular space (ICS) and cerebrospinal fluid (CSF). Morphine or M6G was given to rats at 10 mg kg−1 s.c. Transcortical microdialysis was used to assess their distributions in the brain ECF. Conventional tissue homogenization was used to determine the distribution in the cortex and whole brain. These two procedures were combined to estimate drug distribution in the brain ICS. The blood and CSF pharmacokinetics were also determined. Plasma concentration data for M6G were much higher than those of morphine, with Cmax and AUC 4–5 times more higher, Tmax shorter, and VZf−1 (volume of distribution) and CL f−1 (clearance) 4–6 times lower. The concentrations of the compounds in various brain compartments also differed: AUC values for M6G were lower than those of morphine in tissue and CSF and higher in brain ECF. AUC values in brain show that morphine levels were four times higher in ICS than in ECF, whereas M6G levels were 125 higher in ECF than in ICS. Morphine entered brain cells, whereas M6G was almost exclusively extracellular. This high extracellular concentration, coupled with extremely slow diffusion into the CSF, indicates that M6G was predominantly trapped in the extracellular fluid and therefore durably available to bind at opioid receptors. PMID:10556926

  3. Glucuronidation does not suppress the estrogenic activity of quercetin in yeast and human breast cancer cell model systems.

    PubMed

    Ruotolo, Roberta; Calani, Luca; Brighenti, Furio; Crozier, Alan; Ottonello, Simone; Del Rio, Daniele

    2014-10-01

    Several plant-derived molecules, referred to as phytoestrogens, are thought to mimic the actions of endogenous estrogens. Among these, quercetin, one of the most widespread flavonoids in the plant kingdom, has been reported as estrogenic in some occasions. However, quercetin occurs in substantial amounts as glycosides such as quercetin-3-O-glucoside (isoquercitrin) and quercetin-3-O-rutinoside (rutin) in dietary sources. It is now well established that quercetin undergoes substantial phase II metabolism after ingestion by humans, with plasma metabolites after a normal dietary intake rarely exceeding nmol/L concentrations. Therefore, attributing phytoestrogenic activity to flavonoids without taking into account the fact that it is their phase II metabolites that enter the circulatory system, will almost certainly lead to misleading conclusions. With the aim of clarifying the above issue, the goal of the present study was to determine if plant-associated quercetin glycosides and human phase II quercetin metabolites, actually found in human biological fluids after intake of quercetin containing foods, are capable of interacting with the estrogen receptors (ER). To this end, we used a yeast-based two-hybrid system and an estrogen response element-luciferase reporter assay in an ER-positive human cell line (MCF-7) to probe the ER interaction capacities of quercetin and its derivatives. Our results show that quercetin-3-O-glucuronide, one of the main human phase II metabolites produced after intake of dietary quercetin, displays ERα- and ERβ-dependent estrogenic activity, the functional consequences of which might be related to the protective activity of diets rich in quercetin glycosides.

  4. Application of microdialysis to evaluate the efflux transport of estradiol 17-beta glucuronide across the rat blood-retinal barrier.

    PubMed

    Katayama, Kazunori; Ohshima, Yuki; Tomi, Masatoshi; Hosoya, Ken-ichi

    2006-09-30

    The purpose of the present study was to evaluate vitreous humor/retina-to-blood efflux transport in rats and determine the efflux transport of estradiol 17-beta glucuronide (E17betaG) across the blood-retinal barrier (BRB) by the use of microdialysis. [(3)H]E17betaG and [(14)C]D-mannitol, which were used as a model compound for amphipathic organic anions and a bulk flow marker, respectively, were injected into the vitreous humor of rat eye, and a microdialysis probe was placed in the vitreous humor. [(3)H]E17betaG and [(14)C]D-mannitol were bi-exponentially eliminated from the vitreous humor after vitreous bolus injection. The elimination rate constant of [(3)H]E17betaG during the terminal phase was 1.9-fold greater than that of [(14)C]D-mannitol and reduced the level of [(14)C]D-mannitol in the retinal presence of 0.3 mM E17betaG, suggesting that [(3)H]E17betaG is transported via a carrier-mediated efflux transport process across the BRB. The efflux transport of [(3)H]E17betaG was significantly inhibited by organic anions, such as probenecid, sulfobromophthalein, digoxin, and dehydroepiandrosterone sulfate, whereas it was not inhibited by p-aminohippuric acid. In conclusion, the efflux transport of [(3)H]E17betaG across the rat BRB was evaluated by microdialysis and its inhibition by organic anions suggests organic anion transporting polypeptide 1a4-mediated E17betaG efflux transport at the BRB.

  5. Development of an enzyme immunoassay for urinary pregnanediol-3-glucuronide in a female giant panda (Ailuropoda melanoleuca).

    PubMed

    Hama, Natsuki; Kanemitsu, Hideyasu; Tanikawa, Michiyo; Shibaya, Masami; Sakamoto, Kensuke; Oyama, Yujiro; Acosta, Tomas J; Ishikawa, Osamu; Pengyan, Wang; Okuda, Kiyoshi

    2009-07-01

    In order to enable monitoring of the reproductive status of the female giant panda after observation of estrus behavior, we developed an enzyme immunoassay (EIA) system for urinary pregnanediol-3-glucuronide (PdG), a progesterone metabolite, using commercial reagents and examined the changes in the urinary concentration of PdG in a female giant panda that showed pseudopregnancy and suspicious pseudopregnancy in 6 consecutive years. The developed EIA system had good reproducibility (intra- and interassay CVs 6.1% and 16.3%, respectively), good parallelism between the standard curve and the dose response curve of serial diluted samples and positive correlation (r=0.836) with the data for PdG in the same samples measured by gas chromatography. Urinary PdG in the female panda showed two phases of increase. The first elevation was observed immediately after estrus with the levels of PdG below 100 ng/Crmg, while the second phase was characterized by a drastic elevation above 100 ng/Crmg until the level began to decrease at the end of pseudopregnancy or suspicious pseudopregnancy. The length of the second phase had wider range than that of the first phase. In the present study, a new EIA assay system for urinary PdG in the female giant panda was developed, and we found that the length of the second phase is unstable in the pseudopregnant and suspicious pseudopregnant giant panda, in contrast with the unstable length of the first phase caused by delayed implantation in the pregnant giant panda.

  6. Effects of Andrographis paniculata and Orthosiphon stamineus extracts on the glucuronidation of 4-methylumbelliferone in human UGT isoforms.

    PubMed

    Ismail, Sabariah; Hanapi, Nur Aziah; Ab Halim, Mohd Rohaimi; Uchaipichat, Verawan; Mackenzie, Peter I

    2010-05-14

    The effects of Andrographis paniculata and Orthosiphon stamineus extracts on the in vitro glucuronidation of 4-methylumbelliferone (4MU) by recombinant human UGTs, UGT1A1, UGT1A3, UGT1A6, UGT1A7, UGT1A8, UGT1A10, UGT2B7 and UGT2B15 were determined. The potential inhibitory effects of both of the extracts on the activity of each of the UGT isoforms were investigated using 4MU as the substrate. Incubations contained UDP-glucuronic acid (UDPGA) as the cofactor, MgCl(2), cell lysate of respective isoform, and 4MU at the approximate apparent K(m) or S(50) value of each isoform. Final concentrations of Andrographis paniculata and Orthosiphon stamineus extracts used were 0.025, 0.25, 2.5, 25 and 50 microg/mL and 0.01, 0.10, 1.0, 10 and 50 microg/mL respectively. Both extracts variably inhibited the activity of most of the isoforms in a concentration dependent manner. Andrographis paniculata extract was the better inhibitor of all the isoforms studied (IC(50) 1.70 microg/mL for UGT1A3, 2.57 microg/mL for UGT1A8, 2.82 microg/mL for UGT2B7, 5.00 micorg/mL for UGT1A1, 5.66 microg/mL for UGT1A6, 9.88 microg/mL for UGT1A7 and 15.66 microg/mL for UGT1A10). Both extracts showed less than 70% inhibition of UGT2B15, so the IC(50) values were >50 microg/mL. The inhibition of human UGTs by Andrographis paniculata and Orthosiphon stamineus extracts in vitro suggests a potential for drug-herbal extract interactions in the therapeutic setting.

  7. Isoliquiritigenin showed strong inhibitory effects towards multiple UDP-glucuronosyltransferase (UGT) isoform-catalyzed 4-methylumbelliferone (4-MU) glucuronidation.

    PubMed

    Lu, Hang; Fang, Zhong-Ze; Cao, Yun-Feng; Hu, Cui-Min; Hong, Mo; Sun, Xiao-Yu; Li, Hua; Liu, Yan; Fu, Xiaoguang; Sun, Hongzhi

    2013-01-01

    Isoliquiritigenin, a herbal ingredient with chalcone structure, has been speculated to be able to inhibit one of the most drug-metabolizing enzymes (DMEs) UDP-glucuronosyltransferase (UGT). Therefore, the aim of the present study was to investigate the inhibition of isoliquiritigenin towards important UGT isoforms in the liver and intestine, including UGT1A1, 1A3, 1A6, 1A7, 1A8, 1A9 and 1A10. The recombinant UGT-catalyzed 4-methylumbelliferone (4-MU) glucuronidation was used as probe reactions. The results showed that 100μM of isoliquiritigenin inhibited the activity of UGT1A1, UGT1A3, UGT1A6, UGT1A7, UGT1A8, UGT1A9, and UGT1A10 by 95.2%, 76.1%, 78.9%, 87.2%, 67.2%, 94.8%, and 91.7%, respectively. The data fitting using Dixon plot and Lineweaver-Burk plot showed that the inhibition of UGT1A1, UGT1A9 and UGT1A10 by isoliquiritigenin was all best fit to the competitive inhibition, and the second plot using the slopes from the Lineweaver-Burk plot versus isoliquiritigenin concentrations was used to calculate the inhibition kinetic parameter (K(i)) to be 0.7μM, 0.3μM, and 18.3μM for UGT1A1, UGT1A9, and UGT1A10, respectively. All these results indicated the risk of clinical application of isoliquiritigenin on the drug-drug interaction and other possible diseases induced by the inhibition of isoliquiritigenin towards these UGT isoforms.

  8. Glucuronidation and sulfonation, in vitro, of the major endocrine-active metabolites of methoxychlor in the channel catfish, Ictalurus punctatus, and induction following treatment with 3-methylcholanthrene

    PubMed Central

    James, Margaret O.; Stuchal, Leah D.; Nyagode, Beatrice A.

    2008-01-01

    The organochlorine pesticide, methoxychlor (MXC), is metabolized in animals to phenolic mono- and bis-demethylated metabolites (OH-MXC and HPTE respectively) that interact with estrogen receptors and may be endocrine disruptors. The phase II detoxication of these compounds will influence the duration of action of the estrogenic metabolites, but has not been investigated extensively. In this study, the glucuronidation and sulfonation of OH-MXC and HPTE were investigated in subcellular fractions of liver and intestine from untreated, MXC-treated and 3-methylcholanthrene (3-MC)-treated channel catfish, Ictalurus punctatus. MXC-treated fish were given i.p. injections of 2 mg MXC/kg daily for 6 days and sacrificed 24 hr after the last dose. The 3-MC treatment was a single 10 mg/kg i.p. dose 5 days prior to sacrifice. In hepatic microsomes from control fish, the Vmax value (mean ± S.D., n=4) for glucuronidation of OH-MXC was 270 ± 50 pmol/min/mg protein, higher than found for HPTE (110 ± 20 pmol/min/mg protein). For each substrate, the Vmax values observed in intestinal microsomes were approximately twice those found in the liver. The Km values for OH-MXC and HPTE glucuronidation in control liver were not significantly different and were 0.32 ± 0.04 mM for OH-MXC and 0.26 ± 0.06 mM for HPTE. The Km for the co-substrate, UDPGA, was higher in liver (0.28 ± 0.09 mM) than intestine (0.04 ± 0.02 mM). Treatment with 3-MC but not MXC increased the Vmax for glucuronidation in liver and intestine. Glucuronidation was a more efficient pathway than sulfonation for both substrates, in both tissues. The Vmax values for sulfonation of OH-MXC and HPTE respectively in liver cytosol were 7 ± 3 and 17 ± 4 pmol/min/mg protein and in intestinal cytosol were 13 ± 3 and 30 ± 5 pmol/min/mg protein. Treatment with 3-MC but not MXC increased rates of sulfonation of OH-MXC and HPTE and the model substrate, 3-hydroxy-benzo(a)pyrene in both intestine and liver. Comparison of the kinetics

  9. 21 CFR 172.225 - Methyl and ethyl esters of fatty acids produced from edible fats and oils.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Methyl and ethyl esters of fatty acids produced... Methyl and ethyl esters of fatty acids produced from edible fats and oils. Methyl esters and ethyl esters... following prescribed conditions: (a) The additive consists of a mixture of either methyl or ethyl esters...

  10. 21 CFR 172.225 - Methyl and ethyl esters of fatty acids produced from edible fats and oils.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 3 2012-04-01 2012-04-01 false Methyl and ethyl esters of fatty acids produced... Methyl and ethyl esters of fatty acids produced from edible fats and oils. Methyl esters and ethyl esters... following prescribed conditions: (a) The additive consists of a mixture of either methyl or ethyl esters...

  11. 21 CFR 172.225 - Methyl and ethyl esters of fatty acids produced from edible fats and oils.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 3 2013-04-01 2013-04-01 false Methyl and ethyl esters of fatty acids produced... Methyl and ethyl esters of fatty acids produced from edible fats and oils. Methyl esters and ethyl esters... following prescribed conditions: (a) The additive consists of a mixture of either methyl or ethyl esters...

  12. The Human UDP-glucuronosyltransferase UGT2A1 and UGT2A2 enzymes are highly active in bile acid glucuronidation.

    PubMed

    Perreault, Martin; Gauthier-Landry, Louis; Trottier, Jocelyn; Verreault, Mélanie; Caron, Patrick; Finel, Moshe; Barbier, Olivier

    2013-09-01

    Bile acids (BA) are essential modulators of lipid, glucose, and cholesterol homeostasis, but they exert cytotoxic effects in the cholestatic liver. Glucuronidation, catalyzed by the UDP-glucuronosyltransferase (UGT) enzymes is a pharmacologically relevant BA detoxification process. The present study characterized the BA-conjugating activity of the little-studied human UGTs of subfamily 2A: UGT2A1, 2A2, and 2A3. Recombinant UGT2As, expressed in baculovirus-infected insect cells, were assayed for the glucuronidation of six major bile acids: chenodeoxycholic acid (CDCA), cholic acid (CA), lithocholic acid (LCA), deoxycholic acid (DCA), hyocholic acid (HCA) and hyodeoxycholic acid (HDCA). UGT2A3 exhibited detectable but very low activity with all the tested BA substrates. UGT2A1 was highly efficient in forming LCA-3 and LCA-24G, CDCA-24, DCA-24, HCA-24, and HDCA-24G, whereas UGT2A2 was the most active enzyme for CA-24G and CDCA-24G formation and also was able to generate HDCA-6G, HDCA-24G, LCA-24G, and HCA-24G. The Km values of UGT2A1 varied between 102.2 ± 14.3 µM and 2.4 ± 1.2 mM. With the exception of CA-24G, a low affinity substrate for UGT2A2, all the Km values for UGT2A2 were in the 100 to 400 µM range. We demonstrate the high reactivity of the human UGT2A1 and UGT2A2 for bile acid glucuronidation. The physiologic importance of these reactions to BA disposition remains, however, to be clarified in vivo.

  13. The human UDP-glucuronosyltransferase UGT2A1 and UGT2A2 enzymes are highly active in bile acid glucuronidation

    PubMed Central

    Perreault, Martin; Gauthier-Landry, Louis; Trottier, Jocelyn; Verreault, Mélanie; Caron, Patrick; Finel, Moshe; Barbier, Olivier

    2013-01-01

    Bile acids (BA) are essential modulators of lipid, glucose and cholesterol homeostasis, but exert cytotoxic effects in the cholestatic liver. Glucuronidation, catalyzed by the UDP-glucuronosyltransferase (UGT) enzymes is a pharmacologically-relevant BA detoxification process. The present study aimed at characterizing the BA-conjugating activity of the little-studied human UGTs of subfamily 2A, UGT2A1, 2A2 and 2A3. Recombinant UGT2As, expressed in baculovirus-infected insect cells, were assayed for the glucuronidation of 6 major bile acids, chenodeoxycholic (CDCA), cholic (CA), lithocholic (LCA), deoxycholic (DCA), hyocholic (HCA) and hyodeoxycholic (HDCA) acids. UGT2A3 exhibited detectable, but very low, activity with all the tested BAs substrates. UGT2A1 was highly efficient in forming LCA-3 and -24G, CDCA-24, DCA-24, HCA-24 and HDCA-24G, while UGT2A2 was the most active enzyme for CA-24G and CDCA-24G formation, and was also able to generate HDCA-6G, HDCA-24G, LCA-24G and HCA-24G. The Km values of UGT2A1 varied between 102.2 ± 14.3 μM and 2.4 ± 1.2 mM. With the exception of CA-24G, a low affinity substrate for UGT2A2, all the Km values for UGT2A2 were in the 100 to 400 μM range. In conclusion, the present study demonstrates the high reactivity of the human UGT2A1 and UGT2A2 for bile acid glucuronidation. The physiological importance of these reactions to BA disposition remains, however, to be clarified in vivo. PMID:23756265

  14. Gas chromatography-mass spectrometry of ethyl palmitate calibration and resolution with ethyl oleate as biomarker ethanol sub acute in urine application study

    NASA Astrophysics Data System (ADS)

    Suaniti, Ni Made; Manurung, Manuntun

    2016-03-01

    Gas Chromatography-Mass Spectrometry is used to separate two and more compounds and identify fragment ion specific of biomarker ethanol such as palmitic acid ethyl ester (PAEE), as one of the fatty acid ethyl esters as early detection through conyugated reaction. This study aims to calibrate ethyl palmitate and develop analysis with oleate acid. This methode can be used analysis ethanol and its chemistry biomarker in ethanol sub-acute consumption as analytical forensic toxicology. The result show that ethanol level in urine rats Wistar were 9.21 and decreased 6.59 ppm after 48 hours consumption. Calibration curve of ethyl palmitate was y = 0.2035 x + 1.0465 and R2 = 0.9886. Resolution between ethyl palmitate and oleate were >1.5 as good separation with fragment ion specific was 88 and the retention time was 18 minutes.

  15. PET imaging of β-glucuronidase activity by an activity-based 124I-trapping probe for the personalized glucuronide prodrug targeted therapy.

    PubMed

    Su, Yu-Cheng; Cheng, Ta-Chun; Leu, Yu-Ling; Roffler, Steve R; Wang, Jaw-Yuan; Chuang, Chih-Hung; Kao, Chien-Han; Chen, Kai-Chuan; Wang, Hsin-Ell; Cheng, Tian-Lu

    2014-12-01

    Beta-glucuronidase (βG) is a potential biomarker for cancer diagnosis and prodrug therapy. The ability to image βG activity in patients would assist in personalized glucuronide prodrug cancer therapy. However, whole-body imaging of βG activity for medical usage is not yet available. Here, we developed a radioactive βG activity-based trapping probe for positron emission tomography (PET). We generated a (124)I-tyramine-conjugated difluoromethylphenol beta-glucuronide probe (TrapG) to form (124)I-TrapG that could be selectively activated by βG for subsequent attachment of (124)I-tyramine to nucleophilic moieties near βG-expressing sites. We estimated the specificity of a fluorescent FITC-TrapG, the cytotoxicity of tyramine-TrapG, and the serum half-life of (124)I-TrapG. βG targeting of (124)I-TrapG in vivo was examined by micro-PET. The biodistribution of (131)I-TrapG was investigated in different organs. Finally, we imaged the endogenous βG activity and assessed its correlation with therapeutic efficacy of 9-aminocamptothecin glucuronide (9ACG) prodrug in native tumors. FITC-TrapG showed specific trapping at βG-expressing CT26 (CT26/mβG) cells but not in CT26 cells. The native TrapG probe possessed low cytotoxicity. (124)I-TrapG preferentially accumulated in CT26/mβG but not CT26 cells. Meanwhile, micro-PET and whole-body autoradiography results demonstrated that (124)I-TrapG signals in CT26/mβG tumors were 141.4-fold greater than in CT26 tumors. Importantly, Colo205 xenografts in nude mice that express elevated endogenous βG can be monitored by using infrared glucuronide trapping probes (NIR-TrapG) and suppressed by 9ACG prodrug treatment. (124)I-TrapG exhibited low cytotoxicity allowing long-term monitoring of βG activity in vivo to aid in the optimization of prodrug targeted therapy.

  16. Determination of resveratrol and its sulfate and glucuronide metabolites in plasma by LC-MS/MS and their pharmacokinetics in dogs

    PubMed Central

    Muzzio, Miguel; Huang, Zhihua; Hu, Shu-Chieh; Johnson, William D.; McCormick, David L.; Kapetanovic, Izet M.

    2011-01-01

    An analytical approach for the determination of trans-resveratrol (3,5,4′-trihydroxy-trans-stilbene) and its glucuronide and sulfate conjugates in dog plasma by LC-MS/MS (without enzymatic hydrolysis of the conjugates) was validated to support pre-clinical toxicological and pharmacological studies. The approach required two independent sample extractions and consequent instrument runs. Samples for resveratrol determination were prepared by protein precipitation with acetonitrile; acetonitrile-methanol was used instead for resveratrol metabolites. Chromatographic separation was performed using a C18 column (30 × 2.0 mm) at a flow rate of 0.25 mL/min. For resveratrol the mobile phase consisted of A: 5 mM ammonium acetate in water-isopropanol (98:2, v/v) and B: methanol-isopropanol (98:2, v/v) and for metabolites the mobile phase was modified as follows: A: 0.1% (v/v) formic acid in water and B: 0.1% (v/v) formic acid in acetonitrile. Total run time was 12 min for each run with retention times of about 4-5 min for all analytes. A turbo ion spray source was used operating in negative mode for resveratrol and resveratrol sulfate and in positive mode for resveratrol glucuronide. Calibration curves were linear from 5 to 1000 ng/mL for resveratrol and its glucuronide, and 10 to 2000 ng/mL for resveratrol sulfate. Linearity was assessed using the internal standard method for resveratrol and the external standard method for the metabolites. Method accuracy was 90 to 112% of the true value for all analytes with precision of 9 %RSD or less for all validation experiments. The validated method was applied to a preclinical toxicology study in dogs after oral administration (200 to 1200 mg/kg) of the agent. Peak plasma resveratrol concentration (Cmax) for most animals was observed within 1 to 5 h of dosing, with group mean values in the 1.7 to 9.9 μg/mL (7.5 to 43 μM) range. Area under the plasma concentration-time curve (AUC) mean values for resveratrol ranged from 3.6 to 44

  17. Determination of resveratrol and its sulfate and glucuronide metabolites in plasma by LC-MS/MS and their pharmacokinetics in dogs.

    PubMed

    Muzzio, Miguel; Huang, Zhihua; Hu, Shu-Chieh; Johnson, William D; McCormick, David L; Kapetanovic, Izet M

    2012-02-05

    An analytical approach for the determination of trans-resveratrol (3,5,4'-trihydroxy-trans-stilbene) and its glucuronide and sulfate conjugates in dog plasma by LC-MS/MS (without enzymatic hydrolysis of the conjugates) was validated to support pre-clinical toxicological and pharmacological studies. The approach required two independent sample extractions and consequent instrument runs. Samples for resveratrol determination were prepared by protein precipitation with acetonitrile; acetonitrile-methanol was used instead for resveratrol metabolites. Chromatographic separation was performed using a C18 column (30 mm × 2.0 mm) at a flow rate of 0.25 mL/min. For resveratrol the mobile phase consisted of A: 5mM ammonium acetate in water-isopropanol (98:2, v/v) and B: methanol-isopropanol (98:2, v/v) and for metabolites the mobile phase was modified as follows: A: 0.1% (v/v) formic acid in water and B: 0.1% (v/v) formic acid in acetonitrile. Total run time was 12 min for each run with retention times of about 4-5 min for all analytes. A turbo ion spray source was used operating in negative mode for resveratrol and resveratrol sulfate and in positive mode for resveratrol glucuronide. Calibration curves were linear from 5 to 1000 ng/mL for resveratrol and its glucuronide, and 10-2000 ng/mL for resveratrol sulfate. Linearity was assessed using the internal standard method for resveratrol and the external standard method for the metabolites. Method accuracy was 90-112% of the true value for all analytes with precision of 9% RSD or less for all validation experiments. The validated method was applied to a preclinical toxicology study in dogs after oral administration (200-1200 mg/kg) of the agent. Peak plasma resveratrol concentration (C(max)) for most animals was observed within 1-5 h of dosing, with group mean values in the 1.7-9.9 μg/mL (7.5-43 μM) range. Area under the plasma concentration-time curve (AUC) mean values for resveratrol ranged from 3.6 to 44 h μg/mL for

  18. Pyrrolo[2,1-c][1,4]benzodiazepine-beta-glucuronide prodrugs with a potential for selective therapy of solid tumors by PMT and ADEPT strategies.

    PubMed

    Kamal, Ahmed; Tekumalla, Venkatesh; Raju, P; Naidu, V G M; Diwan, Prakash V; Sistla, Ramakrishna

    2008-07-01

    Pyrrolo[2,1-c][1,4]benzodiazepine-beta-glucuronide prodrugs 15a-b, with a potential for selective therapy of solid tumors by PMT and ADEPT have been designed, synthesized and evaluated for selective cytotoxicity in the presence of the enzyme beta-glucuronidase. The prodrugs have been found to possess reduced cytotoxicity compared to the parent moieties, and are excellent substrates for the enzyme, exhibiting cytotoxicity selectively in the presence of the enzyme. Enhanced water solubility and improved stability are the other important outcomes upon modifying these molecules as their prodrugs.

  19. Pallidol hexa­acetate ethyl acetate monosolvate

    PubMed Central

    Mao, Qinyong; Taylor, Dennis K.; Ng, Seik Weng; Tiekink, Edward R. T.

    2013-01-01

    The entire mol­ecule of pallidol hexa­acetate {systematic name: (±)-(4bR,5R,9bR,10R)-5,10-bis­[4-(acet­yloxy)phen­yl]-4b,5,9b,10-tetra­hydro­indeno­[2,1-a]indene-1,3,6,8-tetrayl tetra­acetate} is completed by the application of twofold rotational symmetry in the title ethyl acetate solvate, C40H34O12·C4H8O2. The ethyl acetate mol­ecule was highly disordered and was treated with the SQUEEZE routine [Spek (2009 ▶). Acta Cryst. D65, 148–155]; the crystallographic data take into account the presence of the solvent. In pallidol hexa­acetate, the dihedral angle between the fused five-membered rings (r.m.s. deviation = 0.100 Å) is 54.73 (6)°, indicating a significant fold in the mol­ecule. Significant twists between residues are also evident as seen in the dihedral angle of 80.70 (5)° between the five-membered ring and the pendent benzene ring to which it is attached. Similarly, the acetate residues are twisted with respect to the benzene ring to which they are attached [C—O(carb­oxy)—C—C torsion angles = −70.24 (14), −114.43 (10) and −72.54 (13)°]. In the crystal, a three-dimensional architecture is sustained by C—H⋯O inter­actions which encompass channels in which the disordered ethyl acetate mol­ecules reside. PMID:24046702

  20. Short synthesis of ethyl 3-(3-aminophenyl)propanoate.

    PubMed

    Nagel, Ulrike; Radau, Gregor; Link, Andreas

    2011-12-01

    A short and effective synthesis of ethyl 3-(3-aminophenyl)propanoate is presented, employing a tandem Knoevenagel condensation/alkylidene reduction of 3-nitrobenzaldehyde with Meldrum's acid in TEAF (triethylammonium formate) followed by reduction of the intermediate 3-(3-nitrophenyl)propanoic acid by stannous chloride in ethanol. The use of stannous chloride as the reducing agent in ethanol enabled the simultaneous esterification of the carboxylic acid. Thus, stannous chloride was acting simultaneously as a Lewis acid, which activates the carboxylic acid towards a nucleophilic attack by ethanol.

  1. Cytotoxic constituents of ethyl acetate fraction from Dianthus superbus.

    PubMed

    Ding, Chengli; Zhang, Wu; Li, Jie; Lei, Jiachuan; Yu, Jianqing

    2013-01-01

    The ethyl acetate fraction (EE-DS) from Dianthus superbus was found to possess the cytotoxic activity against cancer cells in previous study. To investigate cytotoxic constituents, the bioassay-guided isolation of compounds from EE-DS was performed. Two dianthramides (1 and 2), three flavonoids (3-5), two coumarins (6 and 7) and three other compounds (8-10) were obtained. Structures of isolated compounds were identified by spectroscopic analysis. Cytotoxicity of the compounds against HepG2 cells was evaluated. Compound 1 showed the strongest cytotoxicity, compounds 10, 4, 3 and 5 had moderate cytotoxicity.

  2. Micellar phase boundaries under the influence of ethyl alcohol

    PubMed Central

    Bergeron, Denis E.

    2016-01-01

    The Compton spectrum quenching technique is used to monitor the effect of ethyl alcohol (EtOH) additions on phase boundaries in two systems. In toluenic solutions of the nonionic surfactant, Triton X-100, EtOH shifts the boundary separating the first clear phase from the first turbid phase to higher water:surfactant ratios. In a commonly used scintillant, Ultima Gold AB, the critical micelle concentration is not shifted. The molecular interactions behind the observations and implications for liquid scintillation counting are discussed. PMID:26585642

  3. Ethyl Pyruvate Combats Human Leukemia Cells but Spares Normal Blood Cells

    PubMed Central

    Kurz, Susanne; Bigl, Marina; Buchold, Martin; Thieme, Rene; Wichmann, Gunnar; Dehghani, Faramarz

    2016-01-01

    Ethyl pyruvate, a known ROS scavenger and anti-inflammatory drug was found to combat leukemia cells. Tumor cell killing was achieved by concerted action of necrosis/apoptosis induction, ATP depletion, and inhibition of glycolytic and para-glycolytic enzymes. Ethyl lactate was less harmful to leukemia cells but was found to arrest cell cycle in the G0/G1 phase. Both, ethyl pyruvate and ethyl lactate were identified as new inhibitors of GSK-3β. Despite the strong effect of ethyl pyruvate on leukemia cells, human cognate blood cells were only marginally affected. The data were compiled by immune blotting, flow cytometry, enzyme activity assay and gene array analysis. Our results inform new mechanisms of ethyl pyruvate-induced cell death, offering thereby a new treatment regime with a high therapeutic window for leukemic tumors. PMID:27579985

  4. Pharmacokinetic comparison between quercetin and quercetin 3-O-β-glucuronide in rats by UHPLC-MS/MS

    NASA Astrophysics Data System (ADS)

    Yang, Le-Le; Xiao, Na; Li, Xiao-Wei; Fan, Yong; Alolga, Raphael N.; Sun, Xiao-Yue; Wang, Shi-Lei; Li, Ping; Qi, Lian-Wen

    2016-10-01

    Quercetin is a natural flavonoid widely distributed in human diet and functional foods. Quercetin 3-O-β-glucuronide (Q3G) is present in wine and some medicinal plants. Quercetin and Q3G may be metabolized from each other in vivo. While quercetin has been the subject of many studies, the pharmacokinetic profiles of quercetin and Q3G (in animals) have not yet been compared. Herein, we prepared a column-based method for rapid isolation of Q3G from Nelumbo nucifera. Then, we developed an UHPLC-MS/MS method to compare the pharmacokinetics of quercetin and Q3G. Our results showed that the plasma concentration-time curves of quercetin and Q3G show two maxima (Tmax1 ≈ 0.75 h, Tmax2 ≈ 5 h). After oral administration of 100 mg/kg quercetin or 100 mg/kg Q3G in rats, predominantly Q3G was detected in plasma with AUC at 39529.2 ± 6108.2 mg·h·L‑1 or 24625.1 ± 1563.8 mg·h·L‑1, 18-fold higher than quercetin with AUC at 1583.9 ± 583.3 mg·h·L‑1 or 1394.6 ± 868.1 mg·h·L‑1, respectively. After intravenous injection of 10 mg/kg in rats, Q3G showed extensive tissue uptake in kidney (409.2 ± 118.4 ng/g), liver (166.1 ± 52.9 ng/g), heart (97.7 ± 22.6 ng/g), and brain (5.8 ± 1.2 ng/g). In conclusion, we have shown that Q3G is a major active component in plasma and tissue for oral administration of quercetin or Q3G.

  5. Pharmacokinetic comparison between quercetin and quercetin 3-O-β-glucuronide in rats by UHPLC-MS/MS

    PubMed Central

    Yang, Le-Le; Xiao, Na; Li, Xiao-Wei; Fan, Yong; Alolga, Raphael N.; Sun, Xiao-Yue; Wang, Shi-Lei; Li, Ping; Qi, Lian-Wen

    2016-01-01

    Quercetin is a natural flavonoid widely distributed in human diet and functional foods. Quercetin 3-O-β-glucuronide (Q3G) is present in wine and some medicinal plants. Quercetin and Q3G may be metabolized from each other in vivo. While quercetin has been the subject of many studies, the pharmacokinetic profiles of quercetin and Q3G (in animals) have not yet been compared. Herein, we prepared a column-based method for rapid isolation of Q3G from Nelumbo nucifera. Then, we developed an UHPLC-MS/MS method to compare the pharmacokinetics of quercetin and Q3G. Our results showed that the plasma concentration-time curves of quercetin and Q3G show two maxima (Tmax1 ≈ 0.75 h, Tmax2 ≈ 5 h). After oral administration of 100 mg/kg quercetin or 100 mg/kg Q3G in rats, predominantly Q3G was detected in plasma with AUC at 39529.2 ± 6108.2 mg·h·L−1 or 24625.1 ± 1563.8 mg·h·L−1, 18-fold higher than quercetin with AUC at 1583.9 ± 583.3 mg·h·L−1 or 1394.6 ± 868.1 mg·h·L−1, respectively. After intravenous injection of 10 mg/kg in rats, Q3G showed extensive tissue uptake in kidney (409.2 ± 118.4 ng/g), liver (166.1 ± 52.9 ng/g), heart (97.7 ± 22.6 ng/g), and brain (5.8 ± 1.2 ng/g). In conclusion, we have shown that Q3G is a major active component in plasma and tissue for oral administration of quercetin or Q3G. PMID:27775094

  6. [Progesterone and pregnanediol-glucuronid concentrations in saliva, milk and urine of female alpacas and their application in pregnancy diagnosis].

    PubMed

    Volkery, Janine; Wittek, Thomas; Sobiraj, Axel; Gottschalk, Jutta; Einspanier, Almuth

    2010-01-01

    The objective of the present study was the measurement of the pregnancy associated hormones progesterone (P4) and pregnanediol-glucuronide (PdG) in saliva, milk and urine of alpacas and their potential use in pregnancy diagnosis. Sample of blood, saliva, milk and urine were obtained from 36 female alpacas before mating and throughout the pregnancy. Concentrations of P4 and PdG were determined using an enzyme immunoassay (EIA). Pregnancy was checked by ultrasonography at any sampling time. The milk samples were also tested using a commercial on-farm progesterone kit which was designed for dairy cattle. EIA-Concentrations of P4 in blood, milk and urine and urine PdG concentrations were significantly higher in pregnant than in not pregnant alpacas. There was no difference in concentrations of P4 or PdG in saliva. The accuracy of the progesterone kit was 90% for diagnosis of pregnancy and 69% for non-pregnancy. However, 70% of the false positive results also showed relatively high P4 milk concentrations in the EIA. Values of P4 in blood and PdG in urine are comparable to previous reports in alpacas and therefore can be confirmed as an indicator for pregnancy. Saliva seems unsuitable in pregnancy diagnosis in alpacas, whereas milk seems to be an adequate alternative. The use of milk and urine would simplify the pregnancy diagnosis in alpacas since in contrast to the current methods (e. g. blood progesterone) the owners can take the samples. The avoidance of blood sampling results in a considerable stress reduction for the animals. P4 measurement in milk and PdG measurement in urine are good alternatives in pregnancy diagnosis during the first month of pregnancy, when a trans-abdominal ultrasonographic examination is not yet reliable. However, since high values of P4 and PdG only show the presence of active luteal tissue and therefore are indirect markers of pregnancy the diagnosis should be confirmed using ultrasound later in pregnancy.

  7. Dietary Dihydromethysticin Increases Glucuronidation of 4-(Methylnitrosamino)-1-(3-Pyridyl)-1-Butanol in A/J Mice, Potentially Enhancing Its Detoxification

    PubMed Central

    Narayanapillai, Sreekanth C.; von Weymarn, Linda B.; Carmella, Steven G.; Leitzman, Pablo; Paladino, Jordan; Upadhyaya, Pramod; Hecht, Stephen S.; Murphy, Sharon E.

    2016-01-01

    Effective chemopreventive agents are needed against lung cancer, the leading cause of cancer death. Results from our previous work showed that dietary dihydromethysticin (DHM) effectively blocked initiation of lung tumorigenesis by 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) in A/J mice, and it preferentially reduced 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL)-derived DNA adducts in lung. This study explored the mechanism(s) responsible for DHM’s differential effects on NNK/NNAL-derived DNA damage by quantifying their metabolites in A/J mice. The results showed that dietary DHM had no effect on NNK or NNAL abundance in vivo, indicating that DHM does not affect NNAL formation from NNK. DHM had a minimal effect on cytochrome P450 2A5 (CYP2A5, which catalyzes NNK and NNAL bioactivation in A/J mouse lung), suggesting that it does not inhibit NNAL bioactivation. Dietary DHM significantly increased O-glucuronidated NNAL (NNAL-O-gluc) in A/J mice. Lung and liver microsomes from dietary DHM-treated mice showed enhanced activities for NNAL O-glucuronidation. These results overall support the notion that dietary DHM treatment increases NNAL detoxification, potentially accounting for its chemopreventive efficacy against NNK-induced lung tumorigenesis in A/J mice. The ratio of urinary NNAL-O-gluc and free NNAL may serve as a biomarker to facilitate the clinical evaluation of DHM-based lung cancer chemopreventive agents. PMID:26744252

  8. Dietary Dihydromethysticin Increases Glucuronidation of 4-(Methylnitrosamino)-1-(3-Pyridyl)-1-Butanol in A/J Mice, Potentially Enhancing Its Detoxification.

    PubMed

    Narayanapillai, Sreekanth C; von Weymarn, Linda B; Carmella, Steven G; Leitzman, Pablo; Paladino, Jordan; Upadhyaya, Pramod; Hecht, Stephen S; Murphy, Sharon E; Xing, Chengguo

    2016-03-01

    Effective chemopreventive agents are needed against lung cancer, the leading cause of cancer death. Results from our previous work showed that dietary dihydromethysticin (DHM) effectively blocked initiation of lung tumorigenesis by 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) in A/J mice, and it preferentially reduced 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL)-derived DNA adducts in lung. This study explored the mechanism(s) responsible for DHM's differential effects on NNK/NNAL-derived DNA damage by quantifying their metabolites in A/J mice. The results showed that dietary DHM had no effect on NNK or NNAL abundance in vivo, indicating that DHM does not affect NNAL formation from NNK. DHM had a minimal effect on cytochrome P450 2A5 (CYP2A5, which catalyzes NNK and NNAL bioactivation in A/J mouse lung), suggesting that it does not inhibit NNAL bioactivation. Dietary DHM significantly increased O-glucuronidated NNAL (NNAL-O-gluc) in A/J mice. Lung and liver microsomes from dietary DHM-treated mice showed enhanced activities for NNAL O-glucuronidation. These results overall support the notion that dietary DHM treatment increases NNAL detoxification, potentially accounting for its chemopreventive efficacy against NNK-induced lung tumorigenesis in A/J mice. The ratio of urinary NNAL-O-gluc and free NNAL may serve as a biomarker to facilitate the clinical evaluation of DHM-based lung cancer chemopreventive agents.

  9. Acitretin exhibits inhibitory effects towards UDP-glucuronosyltransferase (UGT)1A9-mediated 4-methylumbelliferone (4-MU) and propofol glucuronidation reaction.

    PubMed

    Yu, Ming-Lian; Yang, Yue; Wang, Cong-Min; Zhang, Meng-Meng; Bai, Miao-Chun; Guo, Yue-Ling

    2013-06-01

    The present study aimed to evaluate the potential risk of drug-drug interactions associated with acitretin which is a drug for therapy of psoriasis approved by the Food and Drug Administration (FDA). The initial screening of acitretin's inhibition towards 4-methylumbelliferone (4-MU) glucuronidation catalyzed by important UDP-glucuronosyltransferase (UGT) isoforms in the liver showed that UGT1A9 activity was strongly inhibited by acitretin with other UGT isoforms negligibly influenced. The inhibition type is best fit to competitive inhibition, and the inhibition kinetic parameter (K(i)) was determined to be 3.5 microM. The inhibition behaviour of acitretin towards UGT1A9 activity did not exhibit probe substrate-dependent behaviour when selecting human liver microsomes (HLMs)-catalyzed propofol-O-glucuronidation as probe reaction of UGT1A9. The same inhibition type and similar inhibition parameters (K(i) = 3.2 microM) were obtained. Using the maximum plasma exposure dose of acitretin (C(max)), the C(max)/K(i) values were calculated to be 0.23 and 0.25 when selecting 4-MU and propofol as probe substrates, respectively. All these results indicate a potential clinical drug-drug interaction between acitretin and 4-MU or propofol.

  10. Determination of opiates in whole blood and vitreous humor: a study of the matrix effect and an experimental design to optimize conditions for the enzymatic hydrolysis of glucuronides.

    PubMed

    Sanches, Livia Rentas; Seulin, Saskia Carolina; Leyton, Vilma; Paranhos, Beatriz Aparecida Passos Bismara; Pasqualucci, Carlos Augusto; Muñoz, Daniel Romero; Osselton, Michael David; Yonamine, Mauricio

    2012-04-01

    Undoubtedly, whole blood and vitreous humor have been biological samples of great importance in forensic toxicology. The determination of opiates and their metabolites has been essential for better interpretation of toxicological findings. This report describes the application of experimental design and response surface methodology to optimize conditions for enzymatic hydrolysis of morphine-3-glucuronide and morphine-6-glucuronide. The analytes (free morphine, 6-acetylmorphine and codeine) were extracted from the samples using solid-phase extraction on mixed-mode cartridges, followed by derivatization to their trimethylsilyl derivatives. The extracts were analysed by gas chromatography-mass spectrometry with electron ionization and full scan mode. The method was validated for both specimens (whole blood and vitreous humor). A significant matrix effect was found by applying the F-test. Different recovery values were also found (82% on average for whole blood and 100% on average for vitreous humor). The calibration curves were linear for all analytes in the concentration range of 10-1,500 ng/mL. The limits of detection ranged from 2.0 to 5.0 ng/mL. The method was applied to a case in which a victim presented with a previous history of opiate use.

  11. Synthesis and application of resorufin β-D-glucuronide, a low-cost chromogenic substrate for detecting Escherichia coli in drinking water.

    PubMed

    Magro, Germinal; Bain, Robert E S; Woodall, Claire A; Matthews, Robert L; Gundry, Stephen W; Davis, Anthony P

    2014-08-19

    The development of low-cost tests for Escherichia coli is hampered by the expense and limited choice of enzyme substrates. Most chromogenic substrates are required in costly amounts, while fluorogenic substrates require an additional apparatus (e.g., an ultraviolet lamp) to be detected. Herein, we propose an alternative chromogenic substrate, resorufin β-d-glucuronide (REG), which is exceptionally sensitive and may be employed in very small amounts. We show that REG can be produced similarly to other simple glucuronides and should therefore be no more expensive. The compound is used by both healthy and injured E. coli, resulting in a pronounced color change from orange to a bright pink. Because the released dye (resorufin) has a high extinction coefficient, substantially lower amounts are needed than for commercially available substrates. The potential of this substrate is demonstrated by a presence/absence test requiring just 0.1 mg of REG/100 mL of water sample, one hundredth of the quantity needed for common chromogenic substrates, with an estimated bulk cost of ≤0.1 U.S. cents/test. REG shows promise as a chromogenic substrate for E. coli detection and should be considered in the development of new water tests, especially for low-income settings.

  12. Inter-isoform Hetero-dimerization of Human UDP-Glucuronosyltransferases (UGTs) 1A1, 1A9, and 2B7 and Impacts on Glucuronidation Activity

    PubMed Central

    Yuan, Ling-Min; Gao, Zhang-Zhao; Sun, Hong-Ying; Qian, Sai-Nan; Xiao, Yong-Sheng; Sun, Lian-Li; Zeng, Su

    2016-01-01

    Human UDP-glucuronosyltransferases (UGTs) play a pivotal role in phase II metabolism by catalyzing the glucuronidation of endobiotics and xenobiotics. The catalytic activities of UGTs are highly impacted by both genetic polymorphisms and oligomerization. The present study aimed to assess the inter-isoform hetero-dimerization of UGT1A1, 1A9, and 2B7, including the wild type (1A1*1, 1A9*1, and 2B7*1) and the naturally occurring (1A1*1b, 1A9*2/*3/*5, and 2B7*71S/*2/*5) variants. The related enzymes were double expressed in Bac-to-Bac systems. The fluorescence resonance energy transfer (FRET) technique and co-immunoprecipitation (Co-IP) revealed stable hetero-dimerization of UGT1A1, 1A9, and 2B7 allozymes. Variable FRET efficiencies and donor-acceptor distances suggested that genetic polymorphisms resulted in altered affinities to the target protein. In addition, the metabolic activities of UGTs were differentially altered upon hetero-dimerization via double expression systems. Moreover, protein interactions also changed the regioselectivity of UGT1A9 for querectin glucuronidation. These findings provide in-depth understanding of human UGT dimerization as well as clues for complicated UGT dependent metabolism in humans. PMID:27857056

  13. 40 CFR 180.662 - Trinexapac-ethyl; tolerances for residues.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ...) PESTICIDE PROGRAMS TOLERANCES AND EXEMPTIONS FOR PESTICIDE CHEMICAL RESIDUES IN FOOD Specific Tolerances... residues of the plant growth inhibitor, trinexapac-ethyl, including its metabolites and degradates, in...

  14. Genotoxicity and cytotoxicity assessment of new ethyl-carbamates with ixodicidal activity.

    PubMed

    Prado-Ochoa, María Guadalupe; Muñoz-Guzmán, Marco Antonio; Vázquez-Valadez, Víctor Hugo; Velázquez-Sánchez, Ana María; Salazar, Ana María; Ramírez-Noguera, Patricia; Angeles, Enrique; Alba-Hurtado, Fernando

    2016-09-01

    The mammalian erythrocyte micronucleus test was used on the peripheral blood of Wistar rats exposed to two new ethyl-carbamates: ethyl-4-bromophenyl-carbamate (LQM 919) and ethyl-4-chlorophenyl-carbamate (LQM 996) to analyze their genotoxic potential. The mitotic index and cell proliferation kinetics in human lymphocyte cultures in the presence of these ethyl-carbamates were used to evaluate cytotoxicity and cytostaticity respectively. Exposure to greater acute doses (300mg/kg) and to all of the subchronic doses (12.5, 25 and 50mg/kg daily for 90 days) of these ethyl-carbamates induced an increased frequency (p<0.05) of micro-nucleated polychromatic erythrocytes (MN-PCE) compared with rats not exposed to the ethyl-carbamates. Increases in MN-PCE was higher in males than in females exposed to LQM 996 50mg/Kg (p<0.05). All observed changes in rats return 21days after suspending ethyl-carbamate exposure. The highest concentration (0.3mM) of both ethyl-carbamates in lymphocyte cultures increased the percentage of cells in first division metaphase and decreased the percentage of cells in third division metaphase, indicating an increase in cell cycle length or a possible cell cycle arrest in metaphase (cytostatic effect). The results of this study show that the evaluated ethyl-carbamates may induce genotoxic damage in rats and alterations in the human lymphocyte cell cycle.

  15. Dissociative photoionization of ethyl acrylate: Theoretical and experimental insights

    NASA Astrophysics Data System (ADS)

    Song, Yanlin; Chen, Jun; Ding, Mengmeng; Wei, Bin; Cao, Maoqi; Shan, Xiaobin; Zhao, Yujie; Huang, Chaoqun; Sheng, Liusi; Liu, Fuyi

    2015-08-01

    The photoionization and dissociation of ethyl acrylate have been investigated by time-of-flight mass spectrometer with tunable vacuum ultraviolet (VUV) source in the range of 9.0-20.0 eV. The photoionization mass spectrum (PIMS) for ethyl acrylate and photoionization efficiency (PIE) curves for its major fragment ions: C5H7O2+, C4H5O2+, C3H5O2+, C3H4O+, C3H3O+, C2H5O+, C2H3O+, C2H5+ and C2H4+ have been obtained. The formation channels of main fragments are predicted by Gaussian 09 program at G3B3 level and examined via their dissociation energies from experimental results. Based on our analysis, nine main dissociative photoionization channels are proposed: C5H7O2+ + H, C4H5O2+ + CH3, C3H5O2+ + C2H3, C3H4O+ + C2H4O, C3H3O+ + C2H5O, C2H5O+ + C3H3O, C2H3O+ + C3H5O, C2H5+ + C3H3O2, C2H4+ + C3H4O2, respectively. The results of this work lead to a better understanding of photochemistry in the environment.

  16. Growth of glycine ethyl ester hydrochloride and its characterizations

    NASA Astrophysics Data System (ADS)

    Venkatesan, G.; Pari, S.

    2016-11-01

    Single crystal of glycine ethyl ester hydrochloride by slow evaporation method is reported. The grown crystal characterized by single crystal X-ray diffraction, FT-IR, UV-Vis-NIR and fluorescence spectroscopy. It is established that the crystal falls under the monoclinic system and space group P21/c with the cell parameters as: a=8.565 Å, b=12.943 Å, c=6.272 Å, α=γ=90°, β=103.630º. UV-Vis-NIR spectrum shows indirect allowed transition with a band gap of 5.21 eV and other optical properties are measured. The crystal is also shown to have a high transmittance in the visible region. The third order nonlinear property and optical limiting have been investigated using Z-Scan technique. Complex impedance spectrum measured at the dc conductivity. Dependence of dielectric constant, dielectric loss and ac conductivity on frequency at different temperature of applied ac field is analyzed. The mechanical behavior has been assessed by Vickers microhardness indenter. The thermal behavior of glycine ethyl ester hydrochloride was analyzed using TG/DTA thermal curves. From the thermal study, the material was found to possess thermal stability up to 174 °C. The predicted NLO properties, UV-Vis transmittance and Z-scan studies indicate that is an attractive material for photonics optical limiting applications.

  17. Transesterification process to manufacture ethyl ester of rape oil

    SciTech Connect

    Korus, R.A.; Hoffman, D.S.; Bam, N.; Peterson, C.L.; Drown, D.C.

    1993-12-31

    A process for the production of the ethyl ester of winter rape [EEWR] for use as a biodiesel fuel has been studied. The essential part of the process is the transesterification of rape oil with ethanol, in the presence of a catalyst, to yield the ethyl ester of rape oil as a product and glycerin as a by-product. Experiments have been performed to determine the optimum conditions for the preparation of EEWR. The process variables were: (1) temperature, (2) catalyst, (3) rate of agitation, (4) water content of the alcohol used, and (5) the amount of excess alcohol used. The optimum conditions were: (1) room temperature, (2) 0.5% sodium methoxide or 1% potassium hydroxide catalyst by weight of rapeseed oil, (3) extremely vigorous agitation with some splashing during the initial phase of the reaction and agitation was not necessary after the reaction mixture became homogeneous, (4) absolute ethanol was necessary for high conversion, and (5) 50% excess ethanol with NaOCH{sub 3} or 100% excess with KOH gave a maximum conversion. Viscosity, cloud point and pour point of the EEWR were measured. A preliminary break-even cost for the commercial production of EEWR was found to be $0.55/liter [$2.08/US gallon].

  18. IRIS Toxicological Review of Ethyl Tertiary Butyl Ether (Etbe) ...

    EPA Pesticide Factsheets

    In August 2013, EPA released the draft literature searches and associated search strategies, evidence tables, and exposure response arrays for ETBE to obtain input from stakeholders and the public prior to developing the draft IRIS assessment. Specifically, EPA was interested in comments on the following: Draft literature search strategies The approach for identifying studies The screening process for selecting pertinent studies The resulting list of pertinent studies Preliminary evidence tables The process for selecting studies to include in evidence tables The quality of the studies in the evidence tables The literature search strategy, which describes the processes for identifying scientific literature, contains the studies that EPA considered and selected to include in the evidence tables. The preliminary evidence tables and exposure-response arrays present the key study data in a standardized format. The evidence tables summarize the available critical scientific literature. The exposure-response figures provide a graphical representation of the responses at different levels of exposure for each study in the evidence table. The draft Toxicological Review of Ethyl Tertiary Butyl Ether provides scientific support and rationale for the hazard and dose-response assessment pertaining to chronic exposure to ethyl tertiary butyl ether.

  19. Favoured conformations of methyl isopropyl, ethyl isopropyl, methyl tert-butyl, and ethyl tert-butyl 2-(triphenylphosphoranylidene)malonate.

    PubMed

    Castañeda, Fernando; Silva, Paul; Bunton, Clifford A; Garland, María Teresa; Baggio, Ricardo

    2008-07-01

    The conformations of organic compounds determined in the solid state are important because they can be compared with those in solution and/or from theoretical calculations. In this work, the crystal and molecular structures of four closely related diesters, namely methyl isopropyl 2-(triphenylphosphoranylidene)malonate, C(25)H(25)O(4)P, ethyl isopropyl 2-(triphenylphosphoranylidene)malonate, C(26)H(27)O(4)P, methyl tert-butyl 2-(triphenylphosphoranylidene)malonate, C(26)H(27)O(4)P, and ethyl tert-butyl 2-(triphenylphosphoranylidene)malonate, C(27)H(29)O(4)P, have been analysed as a preliminary step for such comparative studies. As a result of extensive electronic delocalization, as well as intra- and intermolecular interactions, a remarkably similar pattern of preferred conformations in the crystal structures results, viz. a syn-anti conformation of the acyl groups with respect to the P atom, with the bulkier alkoxy groups oriented towards the P atom. The crystal structures are controlled by nonconventional hydrogen-bonding and intramolecular interactions between cationoid P and acyl and alkoxy O atoms in syn positions.

  20. Determination of the main hydrolysis product of O-ethyl S-2-diisopropylaminoethyl methylphosphonothiolate, ethyl methylphosphonic acid, in human serum.

    PubMed

    Katagi, M; Nishikawa, M; Tatsuno, M; Tsuchihashi, H

    1997-02-21

    For the unequivocal proof of the use of a nerve agent O-ethyl S-2-diisopropylaminoethyl methylphosphonothiolate (VX), a rapid, accurate and sensitive method which allows us to identify its main hydrolysis product ethyl methylphosphonic acid (EMPA) in human serum was explored by GC-MS. GC-MS analysis was performed after solvent extraction with acetonitrile in acidic conditions from the serum sample, which was previously deproteinized by micro-ultrafiltration, and subsequent tert.-butyldimethylsilyl derivatization with N-methyl-N-(tert.-butyldimethylsilyl)trifluoroacetamide (MTBSTFA) with 1% tert.-butyldimethylsilyl chloride (t-BDMSC). Linear calibration curves were obtained in the concentration range from 50 to 500 ng/ml for EMPA in the full-scan EI mode and from 5 to 50 ng/ml for EMPA in the SIM EI mode. The relative standard deviation obtained at a sample concentration of 50 ng/ml was 8.4% in the full-scan mode and 7.3% in the SIM mode. Upon applying the full-scan EI and CI mode, 40 ng/ml and 80 ng/ml were the detection limits. Using the SIM-EI mode, in which the ion at m/z 153 was chosen, the limit was 3 ng/ml.

  1. Evaluation of direct and indirect ethanol biomarkers using a likelihood ratio approach to identify chronic alcohol abusers for forensic purposes.

    PubMed

    Alladio, Eugenio; Martyna, Agnieszka; Salomone, Alberto; Pirro, Valentina; Vincenti, Marco; Zadora, Grzegorz

    2017-02-01

    The detection of direct ethanol metabolites, such as ethyl glucuronide (EtG) and fatty acid ethyl esters (FAEEs), in scalp hair is considered the optimal strategy to effectively recognize chronic alcohol misuses by means of specific cut-offs suggested by the Society of Hair Testing. However, several factors (e.g. hair treatments) may alter the correlation between alcohol intake and biomarkers concentrations, possibly introducing bias in the interpretative process and conclusions. 125 subjects with various drinking habits were subjected to blood and hair sampling to determine indirect (e.g. CDT) and direct alcohol biomarkers. The overall data were investigated using several multivariate statistical methods. A likelihood ratio (LR) approach was used for the first time to provide predictive models for the diagnosis of alcohol abuse, based on different combinations of direct and indirect alcohol biomarkers. LR strategies provide a more robust outcome than the plain comparison with cut-off values, where tiny changes in the analytical results can lead to dramatic divergence in the way they are interpreted. An LR model combining EtG and FAEEs hair concentrations proved to discriminate non-chronic from chronic consumers with ideal correct classification rates, whereas the contribution of indirect biomarkers proved to be negligible. Optimal results were observed using a novel approach that associates LR methods with multivariate statistics. In particular, the combination of LR approach with either Principal Component Analysis (PCA) or Linear Discriminant Analysis (LDA) proved successful in discriminating chronic from non-chronic alcohol drinkers. These LR models were subsequently tested on an independent dataset of 43 individuals, which confirmed their high efficiency. These models proved to be less prone to bias than EtG and FAEEs independently considered. In conclusion, LR models may represent an efficient strategy to sustain the diagnosis of chronic alcohol consumption

  2. Reactions of ethyl diazoacetate catalyzed by methylrhenium trioxide

    SciTech Connect

    Zhu, Z.; Espenson, H.

    1995-11-03

    Methylrhenium trioxide (CH{sub 3}ReO{sub 3} or MTO) has found wise use in catalysis, including the epoxidation and metathesis of olefins, aldehyde olefination, and oxygen transfer. Extensive reports have now appeared in the area of MTO-catalyzed substrate oxidations with hydrogen peroxide. Certain catalytic applications of MTO for organic reactions that do not utilize peroxide have now been realized. In particular, a catalytic amount of MTO with ethyl diazoacetate (EDA) will convert aromatic imines to aziridines and convert aldehydes and ketones to epoxides. The aziridine preparation proceeds in high yields under anaerobic conditions more conveniently than with existing methods. Compounds with a three-membered heterocyclic ring can be obtained with the EDA/MTO catalytic system. Aromatic imines undergo cycloaddition reactions to give aziridines under mild conditions.

  3. Interaction of Ethyl Alcohol Vapor with Sulfuric Acid Solutions

    NASA Technical Reports Server (NTRS)

    Leu, Ming-Taun

    2006-01-01

    We investigated the uptake of ethyl alcohol (ethanol) vapor by sulfuric acid solutions over the range approx.40 to approx.80 wt % H2SO4 and temperatures of 193-273 K. Laboratory studies used a fast flow-tube reactor coupled to an electron-impact ionization mass spectrometer for detection of ethanol and reaction products. The uptake coefficients ((gamma)) were measured and found to vary from 0.019 to 0.072, depending upon the acid composition and temperature. At concentrations greater than approx.70 wt % and in dilute solutions colder than 220 K, the values approached approx.0.07. We also determined the effective solubility constant of ethanol in approx.40 wt % H2SO4 in the temperature range 203-223 K. The potential implications to the budget of ethanol in the global troposphere are briefly discussed.

  4. Hydroxide as general base in the saponification of ethyl acetate.

    PubMed

    Mata-Segreda, Julio F

    2002-03-13

    The second-order rate constant for the saponification of ethyl acetate at 30.0 degrees C in H(2)O/D(2)O mixtures of deuterium atom fraction n (a proton inventory experiment) obeys the relation k(2)(n) = 0.122 s(-1) M(-1) (1 - n + 1.2n) (1 - n + 0.48n)/(1 - n + 1.4n) (1 - n + 0.68n)(3). This result is interpreted as a process where formation of the tetrahedral intermediate is the rate-determining step and the transition-state complex is formed via nucleophilic interaction of a water molecule with general-base assistance from hydroxide ion, opposite to the direct nucleophilic collision commonly accepted. This mechanistic picture agrees with previous heavy-atom kinetic isotope effect data of Marlier on the alkaline hydrolysis of methyl formate.

  5. High-temperature unimolecular decomposition of ethyl propionate

    NASA Astrophysics Data System (ADS)

    Giri, Binod Raj; AlAbbad, Mohammed; Farooq, Aamir

    2016-11-01

    This work reports rate coefficients of the thermal unimolecular decomposition reaction of ethyl propionate (EP) behind reflected shock waves over the temperature range of 976-1300 K and pressures of 825-1875 Torr. The reaction progress was monitored by detecting C2H4 near 10.532 μm using CO2 gas laser absorption. In addition, G3//MP2/aug-cc-pVDZ and master equation calculations were performed to assess the pressure- and temperature-dependence of the reaction. Our calculations revealed that C2H4 elimination occurs via a six-centered retro-ene transition state. Our measured rate data are close to the high-pressure limit and showed no discernable temperature fall off.

  6. Tension of Ethyl Alcohol and Hexadecane by Shock Waves

    NASA Astrophysics Data System (ADS)

    Utkin, A. V.; Sosikov, V. A.; Fortov, V. E.

    2006-07-01

    The influences of strain rate and shock wave amplitude on the negative pressure in ethyl alcohol, and hexadecane have been investigated. The method of spall strength measurements was applied and wave profiles were registered by laser interferometer VISAR. Unlike other liquids the process of destruction in methyl alcohol and hexadecane are double staged. At the first stage formation of cavities starts and there is a kinked at free velocity profile was observed. At the second stage the cavity grow rate increases and the spall pulse occurs. The dependence of negative pressure from the strain rate was instigated. The value of the negative pressure correspondent to the kinked at free velocity profile was practically constant and equal to 14MPa for methyl alcohol, and the maximal strength value may be much higher and equal to about 50MPa. Theory of homogeneous bubble nucleation was used to explain the experimental results.

  7. IRIS Toxicological Review of Ethyl Tertiary Butyl Ether (Etbe) ...

    EPA Pesticide Factsheets

    In September 2016, the U.S. Environmental Protection Agency's (USEPA) released the draft Integrated Risk Information System (IRIS) Toxicological Review of Ethyl Tertiary Butyl Ether (ETBE). Consistent with the 2013 IRIS Enhancements, draft IRIS assessments are released prior to external peer review for a 60-day public comment period and discussed at an upcoming public science meeting. Accordingly, the toxicological review, supplementary information, and other materials pertaining to this draft assessment are posted on this site. This material is being released for public viewing and comment prior to a public meeting, providing an opportunity for the IRIS Program to engage in early discussions with stakeholders and the public on data that may be used to identify adverse health effects and characterize exposure-response relationships.

  8. 40 CFR 721.1085 - Benzenamine,4,4′-methylenebis[N-ethyl-N-methyl-.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 30 2010-07-01 2010-07-01 false Benzenamine,4,4â²-methylenebis[N-ethyl-N-methyl-. 721.1085 Section 721.1085 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY... Specific Chemical Substances § 721.1085 Benzenamine,4,4′-methylenebis[N-ethyl-N-methyl-. (a)...

  9. 40 CFR 721.1085 - Benzenamine,4,4′-methylenebis[N-ethyl-N-methyl-.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 32 2013-07-01 2013-07-01 false Benzenamine,4,4â²-methylenebis[N-ethyl-N-methyl-. 721.1085 Section 721.1085 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY... Specific Chemical Substances § 721.1085 Benzenamine,4,4′-methylenebis[N-ethyl-N-methyl-. (a)...

  10. 40 CFR 721.1085 - Benzenamine,4,4′-methylenebis[N-ethyl-N-methyl-.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 31 2014-07-01 2014-07-01 false Benzenamine,4,4â²-methylenebis[N-ethyl-N-methyl-. 721.1085 Section 721.1085 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY... Specific Chemical Substances § 721.1085 Benzenamine,4,4′-methylenebis[N-ethyl-N-methyl-. (a)...

  11. 40 CFR 721.1085 - Benzenamine,4,4′-methylenebis[N-ethyl-N-methyl-.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 32 2012-07-01 2012-07-01 false Benzenamine,4,4â²-methylenebis[N-ethyl-N-methyl-. 721.1085 Section 721.1085 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY... Specific Chemical Substances § 721.1085 Benzenamine,4,4′-methylenebis[N-ethyl-N-methyl-. (a)...

  12. Controlled Degradation of Poly(Ethyl Cyanoacrylate-Co-Methyl Methacrylate)(PECA-Co-PMMA) Copolymers

    Technology Transfer Automated Retrieval System (TEKTRAN)

    This paper describes a method for modifying poly(ethyl cyanoacrylate) in order to control the degradation and the stability as well as the glass transition temperatures. Copolymers of poly(ethyl cyanoacrylate-co-methyl methacrylate) (PECA-co-PMMA) with various compositions were synthesized by free ...

  13. 40 CFR 721.1085 - Benzenamine,4,4′-methylenebis[N-ethyl-N-methyl-.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 31 2011-07-01 2011-07-01 false Benzenamine,4,4â²-methylenebis[N-ethyl-N-methyl-. 721.1085 Section 721.1085 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY... Specific Chemical Substances § 721.1085 Benzenamine,4,4′-methylenebis[N-ethyl-N-methyl-. (a)...

  14. Polychlorinated biphenyl congeners that increase the glucuronidation and biliary excretion of thyroxine are distinct from the congeners that enhance the serum disappearance of thyroxine.

    PubMed

    Martin, L A; Wilson, D T; Reuhl, K R; Gallo, M A; Klaassen, C D

    2012-03-01

    Polychlorinated biphenyl (PCB) congeners differentially reduce serum thyroxine (T(4)) in rats, but little is known about their ability to affect biliary excretion of T(4). Thus, male Sprague-Dawley rats were orally administered Aroclor-1254, Aroclor-1242 (32 mg/kg per day), PCB-95, PCB-99, PCB-118 (16 mg/kg per day), PCB-126 (40 μg/kg per day), 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) (3.9 μg/kg per day), or corn oil for 7 days. Twenty-four hours after the last dose, [(125)I]T(4) was administered intravenously, and blood, bile, and urine samples were collected for quantifying [(125)I]T(4) and in bile [(125)I]T(4) metabolites. Serum T(4) concentrations were reduced by all treatments, but dramatic reductions occurred in response to Aroclor-1254, PCB-99 [phenobarbital (PB)-type congener], and PCB-118 (mixed-type congener). None of the treatments increased urinary excretion of [(125)I]T(4). Aroclor-1254, PCB-118, TCDD, and PCB-126 (TCDD-type congener) increased biliary excretion of T(4)-glucuronide by 850, 756, 710, and 573%, respectively, corresponding to marked induction of hepatic UDP-glucuronosyltransferase (UGT) activity toward T(4). PCB-95 and PCB-99 did not induce UGT activity; therefore, the increased biliary excretion of T(4)-glucuronide was related to the affinity of congeners for the aryl hydrocarbon receptor. The disappearance of [(125)I]T(4) from serum was rapid (within 15-min) and was increased by Aroclor-1254, PCB-99 and PCB-118. Thus, reductions in serum T(4) in response to PCBs did not always correspond with UGT activity toward T(4) or with increased biliary excretion of T(4)-glucuronide. The rapid disappearance of [(125)I]T(4) from the serum of rats treated with PB-like PCBs suggests that increased tissue uptake of T(4) is an additional mechanism by which PCBs may reduce serum T(4).

  15. Glucuronidation of 4-tert-octylphenol in humans, monkeys, rats, and mice: an in vitro analysis using liver and intestine microsomes.

    PubMed

    Hanioka, Nobumitsu; Isobe, Takashi; Ohkawara, Susumu; Tanaka-Kagawa, Toshiko; Jinno, Hideto

    2017-03-01

    4-tert-Octylphenol (4-tOP) is an endocrine-disrupting chemical. It is mainly metabolized into glucuronide by UDP-glucuronosyltransferase (UGT) enzymes in mammals. In the present study, the glucuronidation of 4-tOP in humans, monkeys, rats, and mice was examined in an in vitro system using microsomal fractions. The kinetics of 4-tOP glucuronidation by liver microsomes followed the Michaelis-Menten model for humans and monkeys, and the biphasic model for rats and mice. The K m, V max, and CL int values of human liver microsomes were 0.343 µM, 11.6 nmol/min/mg protein, and 33.8 mL/min/mg protein, respectively. The kinetics of intestine microsomes followed the Michaelis-Menten model for humans, monkeys, and rats, and the biphasic model for mice. The K m, V max, and CL int values of human intestine microsomes were 0.743 µM, 0.571 nmol/min/mg protein, and 0.770 mL/min/mg protein, respectively. The CL int values estimated by Eadie-Hofstee plots were in the order of mice (high-affinity phase) (3.0) > humans (1.0) ≥ monkeys (0.9) > rats (high-affinity phase) (0.4) for liver microsomes, and monkeys (10) > mice (high-affinity phase) (5.6) > rats (1.4) > humans (1.0) for intestine microsomes. The percentages of the CL int values of intestine microsomes to liver microsomes were in the order of monkeys (27 %) > rats (high-affinity phase in liver microsomes) (7.9 %) > mice (high-affinity phase in liver and intestine microsomes) (4.2 %) > humans (2.3 %). These results suggest that the metabolic abilities of UGT enzymes expressed in the liver and intestine toward 4-tOP markedly differ among species and imply that species differences are strongly associated with the toxicities of alkylphenols.

  16. Biomonitoring of N-ethyl-2-pyrrolidone in automobile varnishers.

    PubMed

    Koslitz, Stephan; Meier, Swetlana; Schindler, Birgit Karin; Weiss, Tobias; Koch, Holger Martin; Brüning, Thomas; Käfferlein, Heiko Udo

    2014-12-01

    N-alkyl-2-pyrrolidones are important organic solvents for varnishes in industry. This study investigates exposure to N-ethyl-2-pyrrolidone (NEP) in varnishing of hard plastic components in an automobile plant. Two specific biomarkers of exposure, 5-hydroxy-N-ethyl-2-pyrrolidone (5-HNEP) and 2-hydroxy-N-ethylsuccinimide (2-HESI), were analyzed in urine samples of 14 workers. For this purpose, pre-shift, post-shift and next day pre-shift urine samples were collected midweek. Twelve workers performed regular work tasks (loading, wiping and packing), whereas two workers performed special work tasks including cleaning the sprayer system with organic solvents containing N-alkyl-2-pyrrolidones. Spot urine samples of nine non-exposed persons of the same plant served as controls. Median post-shift urinary levels of workers with regular work tasks (5-HNEP: 0.15 mg/L; 2-HESI: 0.19 mg/L) were ∼5-fold higher compared to the controls (0.03 mg/L each). Continuously increasing metabolite levels, from pre-shift via post-shift to pre-shift samples of the following day, were observed in particular for the two workers with the special working tasks. Maximum levels were 31.01 mg/L (5-HNEP) and 8.45 mg/L (2-HESI). No clear trend was evident for workers with regular working tasks. In summary, we were able to show that workers can be exposed to NEP during varnishing tasks in the automobile industry.

  17. Searching for trans ethyl methyl ether in Orion KL(.)

    PubMed

    Tercero, B; Cernicharo, J; López, A; Brouillet, N; Kolesniková, L; Motiyenko, R A; Margulès, L; Alonso, J L; Guillemin, J-C

    2015-10-01

    We report on the tentative detection of trans ethyl methyl ether (tEME), t-CH3CH2OCH3, through the identification of a large number of rotational lines from each one of the spin states of the molecule towards Orion KL. We also search for gauche-trans-n-propanol, Gt-n-CH3CH2CH2OH, an isomer of tEME in the same source. We have identified lines of both species in the IRAM 30 m line survey and in the ALMA Science Verification data. We have obtained ALMA maps to establish the spatial distribution of these species. Whereas tEME mainly arises from the compact ridge component of Orion, Gt-n-propanol appears at the emission peak of ethanol (south hot core). The derived column densities of these species at the location of their emission peaks are ≤(4.0 ± 0.8) × 10(15) cm(-2) and ≤(1.0 ± 0.2)× 10(15) cm(-2) for tEME and Gt-n-propanol, respectively. The rotational temperature is ~100 K for both molecules. We also provide maps of CH3OCOH, CH3CH2OCOH, CH3OCH3, CH3OH, and CH3CH2OH to compare the distribution of these organic saturated O-bearing species containing methyl and ethyl groups in this region. Abundance ratios of related species and upper limits to the abundances of non-detected ethers are provided. We derive an abundance ratio N(CH3OCH3)/N(tEME) ≥ 150 in the compact ridge of Orion.

  18. Elucidation of the metabolites of the novel psychoactive substance 4-methyl-N-ethyl-cathinone (4-MEC) in human urine and pooled liver microsomes by GC-MS and LC-HR-MS/MS techniques and of its detectability by GC-MS or LC-MS(n) standard screening approaches.

    PubMed

    Helfer, Andreas G; Turcant, Alain; Boels, David; Ferec, Séverine; Lelièvre, Bénédicte; Welter, Jessica; Meyer, Markus R; Maurer, Hans H

    2015-05-01

    4-methyl-N-ethcathinone (4-MEC), the N-ethyl homologue of mephedrone, is a novel psychoactive substance of the beta-keto amphetamine (cathinone) group. The aim of the present work was to study the phase I and phase II metabolism of 4-MEC in human urine as well as in pooled human liver microsome (pHLM) incubations. The urine samples were worked up with and without enzymatic cleavage, the pHLM incubations by simple deproteinization. The metabolites were separated and identified by gas chromatography-mass spectrometry (GC-MS) and liquid chromatography-high resolution-tandem mass spectrometry (LC-HR-MS/MS). Based on the metabolites identified in urine and/or pHLM, the following metabolic pathways could be proposed: reduction of the keto group, N-deethylation, hydroxylation of the 4-methyl group followed by further oxidation to the corresponding 4-carboxy metabolite, and combinations of these steps. Glucuronidation could only be observed for the hydroxy metabolite. These pathways were similar to those described for the N-methyl homologue mephedrone and other related drugs. In pHLM, all phase I metabolites with the exception of the N-deethyl-dihydro isomers and the 4-carboxy-dihydro metabolite could be confirmed. Glucuronides could not be formed under the applied conditions. Although the taken dose was not clear, an intake of 4-MEC should be detectable in urine by the GC-MS and LC-MS(n) standard urine screening approaches at least after overdose.

  19. High-yield synthesis of bioactive ethyl cinnamate by enzymatic esterification of cinnamic acid.

    PubMed

    Wang, Yun; Zhang, Dong-Hao; Zhang, Jiang-Yan; Chen, Na; Zhi, Gao-Ying

    2016-01-01

    In this paper, Lipozyme TLIM-catalyzed synthesis of ethyl cinnamate through esterification of cinnamic acid with ethanol was studied. In order to increase the yield of ethyl cinnamate, several media, including acetone, isooctane, DMSO and solvent-free medium, were investigated in this reaction. The reaction showed a high yield by using isooctane as reaction medium, which was found to be much higher than the yields reported previously. Furthermore, several parameters such as shaking rate, water activity, reaction temperature, substrate molar ratio and enzyme loading had important influences on this reaction. For instance, when temperature increased from 10 to 50 °C, the initial reaction rate increased by 18 times and the yield of ethyl cinnamate increased by 6.2 times. Under the optimum conditions, lipase-catalyzed synthesis of ethyl cinnamate gave a maximum yield of 99%, which was of general interest for developing industrial processes for the preparation of ethyl cinnamate.

  20. Fungal degradation of an acetolactate synthase (ALS) inhibitor pyrazosulfuron-ethyl in soil.

    PubMed

    Sondhia, Shobha; Waseem, Uzma; Varma, R K

    2013-11-01

    Owing to reported phytotoxicity of some sulfonylurea class of herbicides in number of sensitive crops and higher persistence in soil, present study was conducted to isolate and identify pyrazosulfuron-ethyl degrading fungi from soil of rice field. Penicillium chrysogenum and Aspergillus niger, were isolated and identified from rhizospere soil of rice field, as potent pyrazosulfuron-ethyl degrading fungi. Degradation of pyrazosulfuron-ethyl by P. chrysogenum and A. niger, yielded transformation products/metabolites which were identified and characterized by LC/MS/MS. The rate of dissipation of pyrazosulfuron-ethyl was found higher in soil of rice field and soil inoculated with P. chrysogenum. This showed important route of degradation of pyrazosulfuron-ethyl by microbes apart from chemical degradation.