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S Toki

Publications and source records attributed to S Toki.

89 records · Page 5Linked to original sources
PubMed1977

Dehydrogenation of indanol by rabbit liver 3-hydroxyhexobarbital dehydrogenase.

1. Among the several enzyme activities in rabbit liver cytosol able to dehydrogenate 1-indanol, only the main activity was not separable from 3-hydroxyhexobarbital dehydrogenase during purification including polyacrylamide gel disc electrophoresis. 2. Results of mixed substrate method indicated that the same enzyme catalyses the dehydrogenation of 1-indanol and 3-hydroxyhexobarbital. The ratio between the two dehydrogenation activities was almost constant as the enzyme underwent thermal inactivation. The Ki values of p-chloromercuribenzoate, the Km values for NAD+, and the Km values for NADP+ were very similar for the two dehydrogenations. These results lead to the conclusion that the same enzyme catalyses the dehydrogenation of 3-hydroxyhexobarbital and 1-indanol. 3. 1-Tetralol, 1-acenaphthenol, 9-fluorenol, thiochroman-4-ol and 4-chromanol also served as substrate of the enzyme, but 2-indanol, 2-tetralol, and trans- and cis-indan-1,2-diol were not oxidized. 4. Reversibility of the reaction was also confirmed using 1-indanone as substrate.

Alcohol Oxidoreductases↗
PubMed1975

Guinea pig liver 3-hydroxyhexobarbital dehydrogenase. Purification and properties.

3-Hydroxyhexobarbital dehydrogenase, which catalyzes the reversible oxidation of 3-hydroxyhexobarbital to 3-oxohexobarbital, has been purified 470-fold from the soluble fraction of guinea pig liver with a yield of 47%. The specific activity of the purified enzyme is 9.4 units/mg of protein. Results of polyacrylamide gel disc electrophoresis and isoelectric focusing indicated that the purified enzyme preparation is a single and homogeneous protein. NADP+ served as preferred co-factor, but NAD+ is also utilized in the presence of phosphate ion. The guinea pig liver enzyme possessed a relatively narrow substrate specificity in comparison with the rabbit liver enzyme. It is very distinctive that guinea pig liver 3-hydroxyhexobarbital dehydrogenase catalyzes the dehydrogenation of 17beta-hydroxysteroids such as testosterone, 4-androstene-3beta,17beta-diol, 5alpha-androstane-3alpha,17beta-diol, 5alpha-androstane-3beta,17beta-diol, 5alpha-androstan-17beta-ol-3-one, and 5beta-androstane-3alpha,17beta-diol.

Adrenal Glands↗
PubMed1970

Metabolism of beta-ionone. Isolation, characterization and identification of the metabolitesin the urine of rabbits.

1. Rabbits dosed orally with beta-ionone excreted in the urine unchanged beta-ionone, 3-oxo-beta-ionone, 3-oxo-beta-ionol, dihydro-3-oxo-beta-ionol and 3-hydroxy-beta-ionol. 2. Excretion products were isolated as 2,4-dinitrophenylhydrazone derivatives (beta-ionone, 3-oxo-beta-ionone, 3-oxo-beta-ionol and dihydro-3-oxo-beta-ionol) and as p-nitrobenzoate derivatives (3-oxo-beta-ionol, dihydro-3-oxo-beta-ionol and 3-hydroxy-beta-ionol), which were characterized and identified by comparison with the synthetic authentic compounds. 3. The glucuronides of 3-oxo-beta-ionol and dihydro-3-oxo-beta-ionol were also detected in the urine. The latter compound was isolated as free glucuronide, sodium salt and 2,4-dinitrophenylhydrazone.

Alcohols↗
PubMedDate not supplied

In vivo formation of codeinone-glutathione adduct: isolation and identification of a new metabolite in the bile of codeine-treated guinea pig.

Codeinone-glutathione adduct (CO-GSH) in the bile of guinea pigs given a subcutaneous injection of codeine was isolated and identified. Synthesized authentic CO-GSH was characterized by the mass and nuclear magnetic resonance spectra and used as the standard sample. The metabolite was isolated by preparative high-performance liquid chromatography on a C18 column. The fractions containing the conjugated metabolite were purified using Sep-Pak C18 cartridges. For further purification of the metabolite CO-GSH, a Radial Pak CN column was used. Structure assignment of the metabolite was then performed by fast-atom-bombardment mass spectrometry and 500 MHz Fourier-transform-NMR spectrometric analysis and identified as S-[4,5-epoxy-3-methoxy-17-methyl-6-oxomorphinan-(8S)-yl] glutathione.

Animals↗
PubMedDate not supplied

In vivo formation of codeinone and morphinone from codeine. Isolation and identification from guinea pig bile.

Codeinone (CO) and morphinone (MO) were isolated and identified in the bile of guinea pigs given sc injections of codeine. Authentic CO was synthesized and characterized by the NMR and mass spectra of its 2-mercaptoethanol (ME) adduct. This material was then used as the standard to identify the CO-ME adduct in the bile of codeine-treated animals. The MO-ME adduct was also identified in the bile with authentic materials prepared earlier. The results of our investigations indicated that 10.5 and 2.7% dose of CO and MO, respectively, were produced for 6 hr after the codeine was given. The metabolites were separated by preparative HPLC on a reverse phase column packed with C18 gel using a 10 mM sodium phosphate buffer, pH 6.8/CH3CN, 1:1 (v/v) as an eluate. For the further purification of metabolites, we used another reverse phase column with the same mobile phase. A structural elucidation of the ME adduct of metabolites was then performed by fast atom bombardment mass spectroscopy and 400 MHz fourier transform-NMR spectrometric analysis, and identified as (8S)-(2-hydroxyethylthio)dihydrocodeinone and (8S)-(2-hydroxyethylthio)dihydromorphinone, respectively.

Animals↗
PubMedDate not supplied

Stimulation mechanism of guinea pig liver-mediated reduction of naloxone by morphine.

The mechanism of the stimulatory effect of morphine on the reduction of naloxone has been elucidated using guinea pig liver naloxone reductase that is identical with morphine 6-dehydrogenase. The reaction products were quantitated by means of HPLC. When naloxone was incubated with the enzyme in the presence of NAD(P)H at pH 7.4 or pH optima, the production of 6 alpha-naloxol increased according to the added amount of morphine. The stimulation was predominant with NADH at pH 7.4. Under these conditions, the production of morphinone also increased in proportion to the amount of morphine. The enzymatic reduction of naloxone proceeded even if NAD(P)H was replaced by NAD(P)+ and morphine. At a fairly low concentration of NADH (0.01 mM), the enzyme produced 6 alpha-naloxol (0.3 mM), exceeding the stoichiometric amount in the presence of 16 mM morphine. Although the Vmax values for naloxone was increased by the addition of morphine, the Km values for naloxone remained unaltered. Besides other substrates for guinea pig liver morphine 6-dehydrogenase such as codeine, normorphine and ethylmorphine also enhanced the reduction of naloxone. From these results we concluded that the stimulation of guinea pig liver-mediated reduction of naloxone by morphine is caused by the acceleration of the redox of pyridine nucleotides conducted by the enzyme. These phenomena were further supported by the experiments with the liver cytosol. In addition, we confirmed that, in the guinea pig, the biliary excretion of the metabolites, naloxol and naloxol-3-glucuronide, increased after sc injection of naloxone with morphine.

Alcohol Oxidoreductases↗
PubMedDate not supplied

(8S)-(glutathion-S-yl)dihydromorphinone, a novel metabolite of morphine from guinea pig bile.

A novel glutathione-conjugated metabolite of morphine has been isolated from the bile of guinea pigs given morphine. The metabolite was separated by preparative HPLC on a reverse phase column (YMC-GEL C18) using methanol/water (1:4, v/v) as eluate and purified by HPLC on another reverse phase column (mu-Bondapak phenyl) using water/acetonitrile/trimethylamine/acetic acid (150:3:2:1, v/v) as a mobile phase. The unambiguous structure assignment of the metabolite was performed by fast atom bombardment mass spectrometry and 400 MHz fourier transform NMR spectrometric analysis, and it was identified as (8S)-glutathion-S-yl)dihydromorphinone, in comparison with the synthetic morphinone-glutathione adduct.

Animals↗
PubMedDate not supplied

Preparation of four optical isomers of hydroxylated hexobarbital and activities of 3-hydroxyhexobarbital dehydrogenase from guinea pig and rabbit liver.

The preparation of four optical isomers of 3'-hydroxyhexobarbital [5-(3'-hydroxy-1'-cyclohexen-1'-yl)-1,5-dimethylbarbituric acid] is described. The absolute configuration of the four isomers were assigned as (3'S, 5 R) and (3'R, 5R) for alpha- and beta-isomers from (R)-(--)-hexobarbital, respectively. Some of these isomeric 3'-hydroxyhexobarbitals, which are formed in the reaction of hexobarbital with liver microsomal mono-oxygenase, are oxidized to 3'-oxohexobarbital [5-(3'-oxo-1'-cyclohexen-1'-yl)-1,5-dimethylbarbituric acid] by dehydrogenase localized in the soluble fraction of liver homogenates. Comparison of activities among the four isomers for 3-hydroxyhexobarbital dehydrogenase was made by use of enzymes from guinea pig and rabbit liver. It became evident that either enzyme had a quite different activity towards each optical isomer, and the configuration of the 3'-position of hydroxyhexobarbital was an important factor affecting the reactivity of enzyme; isomers with 3'S-configuration were preferentially dehydrogenated to enantiomers with 3'R-configuration. Both enzymes had the highest activity towards (3'S, 5R)-3'-(--)-hydroxy-(--)-hexobarbital, irrespective of their having specificity towards different types of substrates.

Alcohol Oxidoreductases↗
PubMedDate not supplied

Stereoselective formation of glucuronides in metabolism of hexobarbital enantiomers in vivo: isolation and quantitation of glucuronides in rabbit urine.

An improved column-chromatographic method was described for isolation and purification on preparative scale of a glucuronide from urine of rabbits administered (RS)-hexobarbital. The analytically pure preparation obtained was found to be a mixture of two glucuronides of diastereomeric 3'-hydroxyhexobarbitals. Rates of hydrolysis of the glucuronides were dependent on the enzyme preparations used as well as on the configuration of the substrate. For quantitative determination, the glucuronides were hydrolyzed completely by beta-glucuronidases from either Escherichia coli or abalone entrails under the conditions used. In vivo studies on the metabolism of (R)-(-)- and (S)-(+)-hexobarbital in the rabbit showed that the glucuronides excreted in 24-hr urine accounted for about 30% of the dose of each enantiomer, and that conjugation of the hydroxy isomers with glucuronic acid was so stereoselective that the isomers with S-configuration at the 3'-position were preferentially conjugated. There were almost no differences in the urinary metabolite profile between normal an PB-treated rabbits; however, a noticeable change was observed in recovery of unchanged (S)-hexobarbital after phenobarbital treatment.

Animals↗
PubMedDate not supplied

Studies on the mechanism of covalent binding of morphine metabolites to proteins in mouse.

The disposition of N--14CH3-labeled and C-6--3H-labeled morphines in mouse tissue was determined over 48 hr after sc injection. 3H radioactivity in tissues decreased more rapidly than 14C radioactivity, and 14C activity at 48 hr after injection was 2 to 3 times greater than 3H activity in brain, blood and liver. Only 14C radioactivity in the brain and other tissues accumulated significantly by repeated co-administrations of N--14CH3- and C-6--3H-morphines. However, 3H radioactivity did not show much accumulation in brain. The ratio of 14C radioactivity irreversibly bound to macromolecules (insoluble in HCl-methanol) to the total radioactivity increased with time in the liver. [14C]Morphinone-cysteine conjugate was detected in proteolytic digests of mouse liver protein dosed with radiolabeled morphine. Morphinone-glutathione conjugate was also detected in an incubation mixture of morphine and cytosol fraction of mouse liver. These results seem to indicate that morphine is metabolized to morphinone, which then binds covalently to the thiol group of cysteine residues in protein.

Animals↗