John Ansede

Publications (13) View all

• Article: Comparison of selenium and sulfur volatilization by dimethylsulfoniopropionate lyase (DMSP) in two marine bacteria and estuarine sediments

  John H Ansede, Duane C Yoch
  [show abstract] [hide abstract]
  ABSTRACT: The volatile forms of selenium emitted from soils and plants have been identified by others as dimethylselenide and dimethyldiselenide. Dimethylselenoniopropionate, a possible precursor of dimethylselenide, is the selenium analogue of the common marine osmoprotectant, dimethylsulfoniopropionate. Dimethylselenoniopropionate was organically synthesized from dimethylselenide and acrylate and used in a comparative study with dimethylsulfoniopropionate as a growth substrate, an inducer and substrate for dimethylsulfoniopropionate lyase in two marine bacterial isolates, Alcaligenes sp. strain M3A and Pseudomonas doudoroffii, and in salt marsh sediments and estuarine creek water samples. In P. doudoroffii, the rate of dimethylselenoniopropionate transport into cells was about half that observed with dimethylsulfoniopropionate; Alcaligenes does not take up these molecules. The rate and extent of induction of dimethylsulfoniopropionate lyase in P. doudoroffii at low concentrations of dimethylselenoniopropionate were similar to that of dimethylsulfoniopropionate. In Alcaligenes the low constitutive level of dimethylsulfoniopropionate lyase cleaved dimethylselenoniopropionate into dimethylselenide and acrylate, but induction of dimethylsulfoniopropionate lyase by dimethylselenoniopropionate did not follow. Dimethylsulfoniopropionate lyases from both strains, when induced by dimethylsulfoniopropionate, utilized dimethylselenoniopropionate as a substrate with emission of dimethylselenide. Rates of partially purified dimethylsulfoniopropionate lyase activity with dimethylselenoniopropionate were half those with dimethylsulfoniopropionate. Anoxic salt marsh sediments catalyzed dimethylsulfoniopropionate and dimethylselenoniopropionate degradation at equal rates. In sediments containing high organic matter, both dimethylsulfide and dimethylselenide served as substrates for methanogenesis. These data support the hypothesis that organic selenium molecules can be volatilized by the same biochemical pathways as organic sulfur molecules.
  FEMS Microbiology Ecology 01/2006; 23(4):315 - 324. · 3.41 Impact Factor
• Article: In vitro metabolism of an orally active O-methyl amidoxime prodrug for the treatment of CNS trypanosomiasis.

  J H Ansede, R D Voyksner, M A Ismail, D W Boykin, R R Tidwell, J E Hall
  [show abstract] [hide abstract]
  ABSTRACT: A new aza-analogue of furamidine, 6-[5-(4-amidinophenyl)-furan-2-yl]nicotinamidine (DB820), has potent in vitro antitrypanosomal activity; however, it suffers from poor oral activity because of its positively charged amidine groups. The dimethoxyamidine prodrug of DB820, N-methoxy-6-{5-[4-(N-methoxyamidino)phenyl]-furan-2-yl}-nicotinamidine (DB844), has potent oral activity in mouse models of both early-stage and CNS African trypanosomiasis. Metabolism of DB844 in human liver microsomes (HLM) was investigated using liquid chromatography-mass spectrometry (LC-MS/MS). The metabolism of DB844 in HLM was NADPH-dependent and resulted in the production of eight metabolites over a 90?min incubation. O-Demethylation and N-dehydroxylation reactions resulted in the metabolic conversion of DB844 to its active DB820 metabolite. Chromatographic conditions used for LC-MS analysis allowed for the separation and identification of all metabolites including positional isomers. Demethylation of either the phenyl or pyridine side of DB844 (DB844 m/z 366.2) resulted in the production of two metabolites (M1A, M1B), each with a molecular ion of m/z of 352.3 and MS(2) fragments of 288.1, 305.2, 321.2 and 335.2. However, the intensities of the MS(2) fragments were different among the two isomeric metabolites, and comparison to an authentic standard allowed for the structural determination of each metabolite. The isomeric metabolites M2A and M2B, resulting from amidoxime reductions of M1A and M1B, were also chromatographically separated and had distinguishable MS(2) profiles that allowed for their structural assignments when compared to an authentic standard. The di-amidoxime product resulting from O-demethylation of either side of DB844 was also identified as an abundant metabolite during microsomal incubations. The active antitrypanosomal metabolite, DB820, was the last metabolite to be formed and thus provides evidence that DB844 may effectively be metabolized to its active metabolite in vivo.
  Xenobiotica 04/2005; 35(3):211-26. · 1.79 Impact Factor
• Article: Nuclear magnetic resonance analysis of [1-13C]dimethylsulfoniopropionate (DMSP) and [1-13C]acrylate metabolism by a DMSP lyase-producing marine isolate of the alpha-subclass of Proteobacteria.

  J H Ansede, P J Pellechia, D C Yoch
  [show abstract] [hide abstract]
  ABSTRACT: The prominence of the alpha-subclass of Proteobacteria in the marine bacterioplankton community and their role in dimethylsulfide (DMS) production has prompted a detailed examination of dimethylsulfoniopropionate (DMSP) metabolism in a representative isolate of this phylotype, strain LFR. [1-(13)C]DMSP was synthesized, and its metabolism and that of its cleavage product, [1-(13)C]acrylate, were studied using nuclear magnetic resonance (NMR) spectroscopy. [1-(13)C]DMSP additions resulted in the intracellular accumulation and then disappearance of both [1-(13)C]DMSP and [1-(13)C]beta-hydroxypropionate ([1-(13)C]beta-HP), a degradation product. Acrylate, the immediate product of DMSP cleavage, apparently did not accumulate to high enough levels to be detected, suggesting that it was rapidly beta-hydroxylated upon formation. When [1-(13)C]acrylate was added to cell suspensions of strain LFR it was metabolized to [1-(13)C]beta-HP extracellularly, where it first accumulated and was then taken up in the cytosol where it subsequently disappeared, indicating that it was directly decarboxylated. These results were interpreted to mean that DMSP was taken up and metabolized by an intracellular DMSP lyase and acrylase, while added acrylate was beta-hydroxylated on (or near) the cell surface to beta-HP, which accumulated briefly and was then taken up by cells. Growth on acrylate (versus that on glucose) stimulated the rate of acrylate metabolism eightfold, indicating that it acted as an inducer of acrylase activity. DMSP, acrylate, and beta-HP all induced DMSP lyase activity. A putative model is presented that best fits the experimental data regarding the pathway of DMSP and acrylate metabolism in the alpha-proteobacterium, strain LFR.
  Applied and Environmental Microbiology 08/2001; 67(7):3134-9. · 3.83 Impact Factor
• Source

  Available from: ncbi.nlm.nih.gov

  Article: Phylogenetic analysis of culturable dimethyl sulfide-producing bacteria from a spartina-dominated salt marsh and estuarine water.

  J H Ansede, R Friedman, D C Yoch
  [show abstract] [hide abstract]
  ABSTRACT: Dimethylsulfoniopropionate (DMSP), an abundant osmoprotectant found in marine algae and salt marsh cordgrass, can be metabolized to dimethyl sulfide (DMS) and acrylate by microbes having the enzyme DMSP lyase. A suite of DMS-producing bacteria isolated from a salt marsh and adjacent estuarine water on DMSP agar plates differed markedly from the pelagic strains currently in culture. While many of the salt marsh and estuarine isolates produced DMS and methanethiol from methionine and dimethyl sulfoxide, none appeared to be capable of producing both methanethiol and DMS from DMSP. DMSP, and its degradation products acrylate and beta-hydroxypropionate but not methyl-3-mecaptopropionate or 3-mercaptopropionate, served as a carbon source for the growth of all the alpha- and beta- but only some of the gamma-proteobacterium isolates. Phylogenetic analysis of 16S rRNA gene sequences showed that all of the isolates were in the group Proteobacteria, with most of them belonging to the alpha and gamma subclasses. Only one isolate was identified as a beta-proteobacterium, and it had >98% 16S rRNA sequence homology with a terrestrial species of Alcaligenes faecalis. Although bacterial population analysis based on culturability has its limitations, bacteria from the alpha and gamma subclasses of the Proteobacteria were the dominant DMS producers isolated from salt marsh sediments and estuaries, with the gamma subclass representing 80% of the isolates. The alpha-proteobacterium isolates were all in the Roseobacter subgroup, while many of the gamma-proteobacteria were closely related to the pseudomonads; others were phylogenetically related to Marinomonas, Psychrobacter, or Vibrio species. These data suggest that DMSP cleavage to DMS and acrylate is a characteristic widely distributed among different phylotypes in the salt marsh-estuarine ecosystem.
  Applied and Environmental Microbiology 04/2001; 67(3):1210-7. · 3.83 Impact Factor
• Article: An in vitro assay to assess transporter-based cholestatic hepatotoxicity using sandwich-cultured rat hepatocytes.

  John H Ansede, William R Smith, Cassandra H Perry, Robert L St Claire, Kenneth R Brouwer
  [show abstract] [hide abstract]
  ABSTRACT: Drug-induced cholestasis can result from the inhibition of biliary efflux of bile acids in the liver. Drugs may inhibit the hepatic uptake and/or the biliary efflux of bile acids resulting in an increase in serum concentrations. However, it is the intracellular concentration of bile acids that results in hepatotoxicity, and thus serum concentrations may not necessarily be an appropriate indicator of hepatotoxicity. In this study, sandwich-cultured rat hepatocytes were used as an in vitro model to assess the cholestatic potential of drugs using deuterium-labeled sodium taurocholate (d(8)-TCA) as a probe for bile acid transport. Eight drugs were tested as putative inhibitors of d(8)-TCA uptake and efflux. The hepatobiliary disposition of d(8)-TCA in the absence and presence of drugs was measured by using liquid chromatography/tandem mass spectrometry, and the accumulation (hepatocytes and hepatocytes plus bile), biliary excretion index (BEI), and in vitro biliary clearance (Cl(biliary)) were reported. Compounds were classified based on inhibition of uptake, efflux, or a combination of both processes. Cyclosporine A and glyburide showed a decrease in total (hepatocytes plus bile) accumulation, an increase in intracellular (hepatocytes only) accumulation, and a decrease in BEI and Cl(biliary) of d(8)-TCA, suggesting that efflux was primarily affected. Erythromycin estolate, troglitazone, and bosentan resulted in a decrease in accumulation (total and intracellular), BEI, and Cl(biliary) of d(8)-TCA, suggesting that uptake was primarily affected. Determination of a compound's relative effect on bile acid uptake, efflux, and direct determination of alterations in intracellular amounts of bile acids may provide useful mechanistic information on compounds that cause increases in serum bile acids.
  Drug metabolism and disposition: the biological fate of chemicals 11/2009; 38(2):276-80. · 3.74 Impact Factor