Pharmacokinetics and pharmacodynamics of NTBC (2-(2-nitro-4-fluoromethylbenzoyl)-1,3-cyclohexanedione) and mesotrione, inhibitors of 4-hydroxyphenyl pyruvate dioxygenase (HPPD) following a single dose to healthy male volunteers. (33/410)

AIMS: NTBC (2-(2-nitro-4-fluoromethylbenzoyl)-1,3-cyclohexanedione) and mesotrione (2-(4-methylsulphonyl-2-nitrobenzoyl)-1,3-cyclohexanedione) are inhibitors of 4-hydroxyphenyl pyruvate dioxygenase (HPPD). NTBC has been successfully used as a treatment for hereditary tyrosinaemia type 1 (HT-1), while mesotrione has been developed as an herbicide. The pharmacokinetics of the two compounds were investigated in healthy male volunteers following single oral administration. The aim of the NTBC study was to assess the bioequivalence of two different formulations and to determine the extent of the induced tyrosinaemia. The mesotrione study was performed to determine the magnitude and duration of the effect on tyrosine catabolism. Additionally, the urinary excretion of unchanged mesotrione was measured to assess the importance of this route of clearance and to help develop a strategy for monitoring occupational exposure. METHODS: A total of 28 volunteers participated in two separate studies with the compounds. In the first study, the relative bioavailability of NTBC from liquid and capsule formulations was compared and the effect on plasma tyrosine concentrations measured. In the second study the pharmacokinetics of mesotrione were determined at three doses. Plasma tyrosine concentrations were monitored and the urinary excretion of mesotrione and tyrosine metabolites was measured. RESULTS: Both compounds were well tolerated at the dose levels studied. Peak plasma concentrations of NTBC were rapidly attained following a single oral dose of 1 mg x kg(-1) body weight of either formulation and the half-life in plasma was approximately 54 h. There were no statistical differences in mean (+/- s.d.) AUC(0,infinity) (capsule 602 +/- 154 vs solution 602 +/- 146 microg x ml(-1) h) or t1/2 (capsule 55 +/- 13 vs solution 54 +/- 8 h) and these parameters supported the bioequivalence of the two formulations. Mesotrione was also rapidly absorbed, with a significant proportion of the dose eliminated unchanged in urine. The plasma half-life was approximately 1 h and was independent of dose and AUC(0,infinity) and Cmax increased linearly with dose. Following administration of 1 mg NTBC x kg(-1) in either formulation, the concentrations of tyrosine in plasma increased to approximately 1100 nmol x ml(-1). Concentrations were still approximately 8 times those of background at 14 days after dosing, but had returned to background levels within 2 months of the second dose. Administration of mesotrione resulted in an increase in tyrosine concentrations which reached a maximum of approximately 300 nmol x ml(-1) following a dose of 4 mg x kg(-1) body weight. Concentrations returned to those of background within 2 days of dosing. Urinary excretion of tyrosine metabolites was increased during the 24 h immediately following a dose of 4 mg mesotrione x kg(-1), but returned to background levels during the following 24 h period. CONCLUSIONS: NTBC and mesotrione are both inhibitors of HPPD, although the magnitude and duration of their effect on tyrosine concentrations are very different. When normalized for dose, the extent of the induced tyrosinaemia after administration of NTBC and over the duration of these studies, was approximately 400 fold greater than that following administration of mesotrione. The persistent and significant effect on HPPD following administration of NTBC make it suitable for the treatment of patients with hereditary tyrosinaemia type 1 (HT-1), whilst the minimal and transient effects of mesotrione minimize the likelihood of a clinical effect in the event of systemic exposure occurring during occupational use.  (+info)

Two-dimensional IR spectroscopy can be designed to eliminate the diagonal peaks and expose only the crosspeaks needed for structure determination. (34/410)

The power of two-dimensional (2D) IR spectroscopy as a structural method with unprecedented time resolution is greatly improved by the introduction of IR polarization conditions that completely eliminate diagonal peaks from the spectra and leave only the crosspeaks needed for structure determination. This approach represents a key step forward in the applications of 2D IR to proteins, peptides, and other complex molecules where crosspeaks are often obscured by diagonal peaks. The technique is verified on the model compound 1,3-cyclohexanedione and subsequently used to clarify the distribution of structures that the acetylproline-NH(2) dipeptide adopts in chloroform. In both cases, crosspeaks are revealed that were not observed before, which, in the case of the dipeptide, has led to additional information about the structure of the amino group end of the peptide.  (+info)

The role of glutamine 114 in old yellow enzyme. (35/410)

Glutamine 114 of OYE1 is a well conserved residue in the active site of the Old Yellow Enzyme family. It forms hydrogen bonds to the O2 and N3 of the flavoprotein prosthetic group, FMN. Glutamine 114 was mutated to asparagine, introducing an R-group that is one methylene group shorter. The resultant enzyme was characterized to determine the effect of the mutation on the mechanistic behavior of the enzyme, and the crystal structure was solved to determine the effect of the mutation on the structure of the protein. The Q114N mutation results in little change in the protein structure, moving the amide group of residue 114 out of H-bonding distance, allowing repositioning of the FMN prosthetic group to form new interactions that replace the lost H-bonds. The mutation decreases the ability to bind ligands, as all dissociation constants for substituted phenols are larger than for the wild type enzyme. The rate constant for the reductive half-reaction with beta-NADPH is slightly greater, whereas that for the oxidative half-reaction with 2-cyclohexenone is smaller than for the wild type enzyme. Oxidation with molecular oxygen is biphasic and involves formation and reaction with O(2), a phenomenon that is more pronounced with this mutation than with wild type enzyme. When superoxide dismutase is added to the reaction, we observe a single-phase reaction typical of the wild type enzyme. Turnover reactions using beta-NADPH with 2-cyclohexenone and molecular oxygen were studied to further characterize the mutant enzyme.  (+info)

Sorbicillinol, a key intermediate of bisorbicillinoid biosynthesis in Trichoderma sp. USF-2690. (36/410)

In the course of our screening program for free radical scavengers from Trichoderma sp. USF-2690, we found an unidentified metabolite (1) that appeared by the method used for HPLC analysis. Metabolite 1 gradually decreased with the production of bisorbicillinoids and was easily missed during the general isolation procedure. The LC-ESI-MS (negative) analysis for 1 gave m/z 247 as the (M-1)- ion peak. The hydrolysis of synthetic 6-O-acetylsorbicillinol (+/- -2) by 0.05 M KOH and acetylation of product 1 in an aqueous solution indicated that the structure of 1 was (6S)-4-(2,4-hexadienoyl)-3,6-dihydroxy-2,6-dimethyl-2,4-cyclohexadien-1-one, designated sorbicillinol, a quinol that has been postulated to be important in bisorbicillinoid biosynthesis.  (+info)

Regioselective nucleophilic addition of methoxybenzene derivatives to the beta-carbon of p-benzoquinone mono O,S-acetal. (37/410)

Regioselective nucleophilic addition of electron rich aromatics to the beta-position of acetal carbon of p-benzoquinone mono O,S-acetal was achieved by modifying the acetal moiety.  (+info)

Kinetic and structural basis of reactivity of pentaerythritol tetranitrate reductase with NADPH, 2-cyclohexenone, nitroesters, and nitroaromatic explosives. (38/410)

The reaction of pentaerythritol tetranitrate reductase with reducing and oxidizing substrates has been studied by stopped-flow spectrophotometry, redox potentiometry, and X-ray crystallography. We show in the reductive half-reaction of pentaerythritol tetranitrate (PETN) reductase that NADPH binds to form an enzyme-NADPH charge transfer intermediate prior to hydride transfer from the nicotinamide coenzyme to FMN. In the oxidative half-reaction, the two-electron-reduced enzyme reacts with several substrates including nitroester explosives (glycerol trinitrate and PETN), nitroaromatic explosives (trinitrotoluene (TNT) and picric acid), and alpha,beta-unsaturated carbonyl compounds (2-cyclohexenone). Oxidation of the flavin by the nitroaromatic substrate TNT is kinetically indistinguishable from formation of its hydride-Meisenheimer complex, consistent with a mechanism involving direct nucleophilic attack by hydride from the flavin N5 atom at the electron-deficient aromatic nucleus of the substrate. The crystal structures of complexes of the oxidized enzyme bound to picric acid and TNT are consistent with direct hydride transfer from the reduced flavin to nitroaromatic substrates. The mode of binding the inhibitor 2,4-dinitrophenol (2,4-DNP) is similar to that observed with picric acid and TNT. In this position, however, the aromatic nucleus is not activated for hydride transfer from the flavin N5 atom, thus accounting for the lack of reactivity with 2,4-DNP. Our work with PETN reductase establishes further a close relationship to the Old Yellow Enzyme family of proteins but at the same time highlights important differences compared with the reactivity of Old Yellow Enzyme. Our studies provide a structural and mechanistic rationale for the ability of PETN reductase to react with the nitroaromatic explosive compounds TNT and picric acid and for the inhibition of enzyme activity with 2,4-DNP.  (+info)

Inhibition of tumor necrosis factor-alpha -induced nuclear translocation and activation of NF-kappa B by dehydroxymethylepoxyquinomicin. (39/410)

We previously designed and synthesized an NF-kappaB inhibitor, dehydroxymethylepoxyquinomicin (DHMEQ), that showed anti-inflammatory activity in vivo. In the present study we looked into its mechanism of inhibition. DHMEQ inhibited tumor necrosis factor-alpha (TNF-alpha)- and 12-O-tetradecanoylphorbol-13-acetate-induced transcriptional activity of NF-kappaB in human T cell leukemia Jurkat cells. It also inhibited the TNF-alpha-induced DNA binding of nuclear NF-kappaB but not the phosphorylation and degradation of IkappaB. Moreover, DHMEQ inhibited the TNF-alpha-induced nuclear accumulation of p65, a component of NF-kappaB. It also inhibited TNF-alpha-induced nuclear transport of green fluorescent protein-tagged p65. On the other hand, DHMEQ did not inhibit the nuclear transport of Smad2 and large T antigen. Also, it did not inhibit TNF-alpha-induced activation of JNK but synergistically induced apoptosis with TNF-alpha in Jurkat cells. Taken together, these data indicate that DHMEQ is a unique inhibitor of NF-kappaB acting at the level of nuclear translocation.  (+info)

Effect of P-glycoprotein on the pharmacokinetics and tissue distribution of enaminone anticonvulsants: analysis by population and physiological approaches. (40/410)

Multidrug resistance (MDR), mediated by P-glycoprotein (Pgp) has been identified as altering the disposition of structurally diverse compounds. Previous in vitro studies in bovine brain microvascular endothelial cells and MCF/Adr [Adriamycin (doxorubicin)-resistant human breast cancer] cells displayed that the transport of enaminone anticonvulsants was influenced by Pgp. Therefore the objectives of this study was to further evaluate the influence of Pgp on the pharmacokinetics and tissue distribution of the enaminone analogs. mdr1ab (+/+) and mdr1ab (-/-) male mice (20 +/- 5 g) were administered DM5 (methyl 4-[(4'-chlorophenyl)amino]-6-methyl-2-oxo-3-cyclohexene-1-carboxylate) or DM44 (12.5 mg/kg, i.v.). Cohorts (n = 3) were sacrificed over a 12-h period, and samples were analyzed by a validated UV-high performance liquid chromatography assay method. Population analysis was used to estimate pharmacokinetic parameters and partition coefficients were determined for tissues. The clearance (0.51 versus 0.33 l/h/kg) and V(d) (1.25 versus 0.93 l/kg) of DM5 were found to be higher (p < 0.05), however the area under the curve (26.1 versus 38.2 microg/ml. h) was lower (p < 0.05) in mdr1a/1b (-/-) versus mdr1a/1b (+/+) mice, respectively. Similar findings were observed for DM44. Tissues known to express Pgp such as the heart, liver, lung, and brain displayed 2-fold or higher tissue levels in mdr1a/1b (-/-) versus mdr1a/1b (+/+) mice. These results strongly suggest that Pgp may influence enaminone tissue distribution and pharmacokinetics and may play a significant role in the effective treatment of epilepsy with these analogs.  (+info)