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Glucuronidation of deoxynivalenol (DON) by different animal species: identification of iso-DON glucuronides and iso-deepoxy-DON glucuronides as novel DON metabolites in pigs, rats, mice, and cows.

ABSTRACT: The Fusarium mycotoxin deoxynivalenol (DON) is a frequent contaminant of cereal-based food and feed. Mammals metabolize DON by conjugation to glucuronic acid (GlcAc), the extent and regioselectivity of which is species-dependent. So far, only DON-3-glucuronide (DON-3-GlcAc) and DON-15-GlcAc have been unequivocally identified as mammalian DON glucuronides, and DON-7-GlcAc has been proposed as further DON metabolite. In the present work, qualitative HPLC-MS/MS analysis of urine samples of animals treated with DON (rats: 2 mg/kg bw, single bolus, gavage; mice: 1 mg/kg bw, single i.p. injection; pigs: 74 µg/kg bw, single bolus, gavage; cows: 5.2 mg DON/kg dry mass, oral for 13 weeks) revealed additional DON and deepoxy-DON (DOM) glucuronides. To elucidate their structures, DON and DOM were incubated with human (HLM) and rat liver microsomes (RLM). Besides the expected DON/DOM-3- and 15-GlcAc, minor amounts of four DON- and four DOM glucuronides were formed. Isolation and enzymatic hydrolysis of four of these compounds yielded iso-DON and iso-DOM, the identities of which were eventually confirmed by NMR. Incubation of iso-DON and iso-DOM with RLM and HLM yielded two main glucuronides for each parent compound, which were isolated and identified as iso-DON/DOM-3-GlcAc and iso-DON/DOM-8-GlcAc by NMR. Iso-DON-3-GlcAc, most likely misidentified as DON-7-GlcAc in the literature, proved to be a major DON metabolite in rats and a minor metabolite in pigs. In addition, iso-DON-8-GlcAc turned out to be one of the major DON metabolites in mice. DOM-3-GlcAc was the dominant DON metabolite in urine of cows and an important DON metabolite in rat urine. Iso-DOM-3-GlcAc was detected in urine of DON-treated rats and cows. Finally, DON-8,15-hemiketal-8-glucuronide, a previously described by-product of DON-3-GlcAc production by RLM, was identified in urine of DON-exposed mice and rats. The discovery of several novel DON-derived glucuronides in animal urine requires adaptation of the currently used methods for DON-biomarker analysis.

SUBMITTER: Schwartz-Zimmermann HE

PROVIDER: S-EPMC5719127 | biostudies-literature | 2017 Dec

REPOSITORIES: biostudies-literature

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Glucuronidation of deoxynivalenol (DON) by different animal species: identification of iso-DON glucuronides and iso-deepoxy-DON glucuronides as novel DON metabolites in pigs, rats, mice, and cows.

Schwartz-Zimmermann Heidi E HE Hametner Christian C Nagl Veronika V Fiby Iris I Macheiner Lukas L Winkler Janine J Dänicke Sven S Clark Erica E Pestka James J JJ Berthiller Franz F

Archives of toxicology 20170621 12

The Fusarium mycotoxin deoxynivalenol (DON) is a frequent contaminant of cereal-based food and feed. Mammals metabolize DON by conjugation to glucuronic acid (GlcAc), the extent and regioselectivity of which is species-dependent. So far, only DON-3-glucuronide (DON-3-GlcAc) and DON-15-GlcAc have been unequivocally identified as mammalian DON glucuronides, and DON-7-GlcAc has been proposed as further DON metabolite. In the present work, qualitative HPLC-MS/MS analysis of urine samples of animals ...[more]

PMID: 28638985

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Project description:BackgroundFusarium infection with concurrent production of deoxynivalenol (DON) causes an increasing safety concern with feed worldwide. This study was conducted to determine the effects of varying levels of DON in diets on growth performance, serum biochemical profile, jejunal morphology, and the differential expression of nutrients transporter genes in growing pigs.ResultsA total of twenty-four 60-day-old healthy growing pigs (initial body weight = 16.3 ± 1.5 kg SE) were individually housed and randomly assigned to receive one of four diets containing 0, 3, 6 or 12 mg DON/kg feed for 21 days. Differences were observed between control and the 12 mg/kg DON treatment group with regards to average daily gain (ADG), although the value for average daily feed intake (ADFI) in the 3 mg/kg DON treatment group was slightly higher than that in control (P<0.01). The relative liver weight in the 12 mg/kg DON treatment group was significantly greater than that in the control (P<0.01), but there were no significant differences in other organs. With regard to serum biochemistry, the values of blood urea nitrogen (BUN), alkaline phosphatase (ALP), alanine aminotransferase (ALT) and aspartate amino transferase (AST) in the 3 treatment groups were higher than those in the control, and the serum concentrations of L-valine, glycine, L-serine, and L-glutamine were significantly reduced in the 3 treatment groups, especially in the 12 mg/kg DON group (P<0.01). Serum total superoxide dismutase (T-SOD), glutathione peroxidase (GSH-Px) were markedly decreased after exposure to DON contaminated feeds (P<0.01). The villi height was markedly decreased and the lymphocyte cell numbers markedly increased in the 3 DON contaminated feeds (P<0.01). The mRNA expression levels of excitatory amino acid transporter-3 (EAAC-3), sodium-glucose transporter-1 (SGLT-1), dipeptide transporter-1 (PepT-1), cationic amino acid transporter-1 (CAT-1) and y(+)L-type amino acid transporter-1 (LAT-1) in control were slightly or markedly higher than those in the 3 DON treatment groups.ConclusionsThese results showed that feeds containing DON cause a wide range of effects in a dose-dependent manner. Such effects includes weight loss, live injury and oxidation stress, and malabsorption of nutrients as a result of selective regulation of nutrient transporter genes such as EAAC-3, SGLT-1, PepT-1, CAT-1 and LAT-1.

Project description:The prevalence of deoxynivalenol (DON) is a concern for swine producers, and although there has been extensive research into the effects of DON in pigs, focus has been in young pigs and/or in short-term studies. The objective of the study was to determine the effect of long-term exposure to DON-contaminated diets in finisher pigs. A total of 200 pigs (76.6 ± 3.9 kg initial weight) were group housed (five pigs per pen; n = 10 pens/treatment) in a 6-wk trial. Pigs were fed a wheat-barley-soybean meal-based control (CONT) diet with no DON or the basal diet in which clean wheat was replaced by DON-contaminated wheat and wheat screenings to provide DON content of 1, 3, or 5 ppm (DON1, DON3, and DON5, respectively). Individual BW and pen feed intake were recorded weekly to calculate average daily gain (ADG), average daily feed intake (ADFI), and gain to feed ratio (G:F). Blood was collected on days 0, 14, and 43 and analyzed for indicators of liver and kidney health. Nitrogen (N)-balance was conducted immediately following the growth performance period to determine the effect of DON on nutrient utilization. Blood and urine samples collected during N balance were analyzed for DON content. Feeding DON reduced (P < 0.05) ADFI and ADG from days 0 to 28 compared with CONT, after which there was no effect of diet on ADFI and ADG. The G:F was lower (P < 0.05) in DON5 fed pigs compared with all treatments during days 0 to 7; however, no treatment effects on G:F was observed from days 8 to 42. Nitrogen retention was lower (P < 0.05) in DON3 and DON5 compared with DON1-fed pigs. Nitrogen retention efficiency was higher (P < 0.05) in DON1 compared with DON3 and DON5 and protein deposition for DON1 pigs was higher (P < 0.05) than all treatments. There were no treatment effects on indicators of liver and kidney health. As dietary DON intake increased, concentration of DON in blood and urine increased. Overall, although there was an initial decrease in ADG and ADFI in pigs receiving diets containing >1 ppm DON, pig performance recovered after a period of time, whereas nutrient utilization continued to be affected after recovery of performance. Moreover, the lack of DON on G:F indicates that the negative effects of DON on growth performance are largely due to reduced feed intake. Overall, although pigs maybe capable of adapting to intake of DON-contaminated diets, their final body weight will be reduced when fed diets containing >1 ppm DON.

Project description:Bupropion is a widely used antidepressant and smoking cessation aid in addition to being one of two US Food and Drug Administration-recommended probe substrates for evaluation of cytochrome P450 2B6 activity. Racemic bupropion undergoes oxidative and reductive metabolism, producing a complex profile of pharmacologically active metabolites with relatively little known about the mechanisms underlying their elimination. A liquid chromatography-tandem mass spectrometry assay was developed to simultaneously separate and detect glucuronide metabolites of (R,R)- and (S,S)-hydroxybupropion, (R,R)- and (S,S)-hydrobupropion (threo) and (S,R)- and (R,S)-hydrobupropion (erythro), in human urine and liver subcellular fractions to begin exploring mechanisms underlying enantioselective metabolism and elimination of bupropion metabolites. Human liver microsomal data revealed marked glucuronidation stereoselectivity [Cl(int), 11.4 versus 4.3 µl/min per milligram for the formation of (R,R)- and (S,S)-hydroxybupropion glucuronide; and Cl(max), 7.7 versus 1.1 µl/min per milligram for the formation of (R,R)- and (S,S)-hydrobupropion glucuronide], in concurrence with observed enantioselective urinary elimination of bupropion glucuronide conjugates. Approximately 10% of the administered bupropion dose was recovered in the urine as metabolites with glucuronide metabolites, accounting for approximately 40%, 15%, and 7% of the total excreted hydroxybupropion, erythro-hydrobupropion, and threo-hydrobupropion, respectively. Elimination pathways were further characterized using an expressed UDP-glucuronosyl transferase (UGT) panel with bupropion enantiomers (both individual and racemic) as substrates. UGT2B7 catalyzed the stereoselective formation of glucuronides of hydroxybupropion, (S,S)-hydrobupropion, (S,R)- and (R,S)-hydrobupropion; UGT1A9 catalyzed the formation of (R,R)-hydrobupropion glucuronide. These data systematically describe the metabolic pathways underlying bupropion metabolite disposition and significantly expand our knowledge of potential contributors to the interindividual and intraindividual variability in therapeutic and toxic effects of bupropion in humans.

Dataset Information

Glucuronidation of deoxynivalenol (DON) by different animal species: identification of iso-DON glucuronides and iso-deepoxy-DON glucuronides as novel DON metabolites in pigs, rats, mice, and cows.

Publications

Glucuronidation of deoxynivalenol (DON) by different animal species: identification of iso-DON glucuronides and iso-deepoxy-DON glucuronides as novel DON metabolites in pigs, rats, mice, and cows.

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