Stereoselective and regioselective hydroxylation of warfarin and selective O-dealkylation of phenoxazone ethers in human placenta

The oxidative metabolism of warfarin and a series of phenoxazone ethers was studied in two groups of human placentas which exhibited high or low levels of aryl hydrocarbon hydroxylase (AHH). Warfarin metabolism was stereoselective (mean R/S = 2.48) for the R-enantiomer and regioselective for the 6- and 8- positions in the high AHH group whereas warfarin metabolism in the low AHH group displayed no significant overall stereoselectivity (mean R/S = 1.24) and was regioselective for the 7- position. The high AHH group metabolized the methyl, ethyl, propyl and butyl ethers of phenoxazone rapidly, while the low AHH group catalyzed their biotransformation at very low or negligible rates. Neither group detectably metabolized phenoxazone or pentyloxyphenoxazone whereas both groups metabolized benzyloxyphenoxazone at low but similar rates. Rates of warfarin R-6 and R-8 hydroxylation were highly correlated with metabolism of benzo(alpha)pyrene (r = 0.99) and the C1-C4 phenoxazone ethers (r greater than 0.87), but poorly correlated with metabolism of benzyloxyphenoxazone (r less than 0.50). These data support the use of warfarin and the phenoxazone ethers as sensitive biochemical probes for P-450 isozymes in human extrahepatic tissues. They indicate the presence of a multiplicity of xenobiotic metabolizing P-450's in placental tissue which has not been exposed to inducing agents that elevate AHH.     

Theoretical studies on the mechanism of conversion of androgens to estrogens by aromatase.

Semiempirical molecular orbital calculations (AM1) were used to model several possible reaction mechanisms for the third oxidation of the aromatase-catalyzed conversion of androgens to estrogens. The reaction mechanisms considered are based on the assumption that the third oxidation is initiated by 1 beta-hydrogen atom abstraction. Homolytic cleavage of the C10-C19 bond was modeled for both the 3-keto and 2-en-3-ol forms of the androgen 1-radicals. The addition of a protein nucleophile to the 19-oxo intermediate was also considered, and -OCH3, -SCH3, and -NHCH3 were used to represent the Ser, Cys, and Lys adducts. The transition states were estimated and optimized from the reaction coordinates obtained by constraining and increasing the C10-C19 bond lengths. The enthalpies of activation range from 14 to 21 kcal and are approximately 2 kcal lower for cleavage of the enol form. Given the tendency for AM1 to overestimate activation energies, all reactions may be energetically accessible. Other reactions modeled include a homolytic cleavage reaction from a thioether radical cation and the direct additions of oxygen radical compounds to the carbonyl of the 1-radical-2-en-3-ol-19-oxo androgen. A mechanism is proposed in which the 19-oxo intermediate is subject to initial nucleophilic attack by the protein. Since rotation of the 19-carbonyl can bring the oxygen within 2.1 A of the 2 beta-hydrogen, the formation of a tetrahedral intermediate can occur with concomitant removal of the 2 beta-proton. Enolization activates the C1-position for hydrogen atom abstraction, since the resulting radical is resonance stabilized.(ABSTRACT TRUNCATED AT 250 WORDS)     

NAG infection of grass frog (Rana temporaria) tadpoles]

Rana temporaria tadpoles (2250) were contaminated with three types of NAG vibrio cultures. Clinical, bacteriological and morphological examinations showed the larvae to be suffering from an acute infection during the first 2 days after the contamination. Then the vibrios persisted in the tadpole organism for a long time and were excreted in the feces.     

Nutrition of a Hemoflagellate (Leishmania tarentolae) Having an Interchangeable Requirement for Choline or Pyridoxal.

SUMMARY. Leptomonads of Leishmania tarentolae were grown continuously in a defined medium containing: inorganic salts', glucose, hemin, 17 amino acids, purines and pyrimidines, and a mixture of vitamins of the B group. In this medium the population of organisms reached about 20 to 50 million per ml. alter 1 week at 27°C. Only slightly better growth occurred in a partially defined medium containing bovine plasma fraction V. In earlier experiments, however, omission of the plasma fraction resulted in decreased growth, and under these circumstances cholesterol or lecithin had growth-stimulating effects. In later experiments in the fully-defined medium no effect of these lipids could be found. The leptomonads were shown to require at least the following substances: inorganic salts; a source of purines and pyrimidines; tryptophan and the nine other amino acids essential for the growth of rats, glutamic acid, tyrosine, proline, serine, one or more of the group alanine, glycine and aspartic acid; folic acid, biotin, pantothenic acid, nicotinamide, riboflavm, thiamine, and either pyridoxine plus choline or pyridoxal or pyridoxamine. Choline at 2 × 10⁻⁵ m gave optimal growth in the presence of pyridoxine at 1 × 10⁻⁵ m. In a medium with a suboptimal concentration of choline (0.4 × 10⁻⁵ m) the leptomonads grew through nine transfers but they were mostly somewhat rounded and aflagellate.     

NAG-infection in grass frogs (Rana temporaria) subjected to hypothermia]

Rana temporaria kept under hypothermic conditions approaching anabiosis were inoculated with NAG-vibrios and examined clinically, bacteriologically, histologically, and electron microscopically. Oral inoculation of hypothermic frogs with NAG-vibrios resulted in 18 to 24 hours in the development of acute NAG-infection resembling the cholera-like syndrome, and characterized by general intoxication and local enteropathogenic effects. NAG-vibrios persisted in the frog gastrointestinal tract for a long time after the cessation of the acute period of the disease.     

A stable isotope assay for phenprocoumon and its metabolites

A gas chromatographic/mass spectrometric assay for quantifying phenprocoumon and its 4'-, 6-, 7- and 8-hydroxy metabolites in microsomal preparations is described. This assay which uses deuterium-labeled analogs of the phenprocoumon metabolites as internal standards has a lower limit of quantitation of 20 ng ml-1. Diazomethane is used to derivatize both metabolites and parent compound yielding along with the expected 4-methoxy derivative a minor amount of the 2-methoxychromone. Resolution of the methylated metabolites is accomplished by capillary gas chromatography.