Purification, characterization, and partial structure of D factor from Polysphondylium violaceum.

The A component of D factor (DfA) was overproduced during development of wild type Polyspondylium violaceum strain China after starvation in liquid medium. Crude DfA excreted by strain China was partially purified by ultrafiltration using Amicon YM10 and YM2 filters with DfA extracted from the filtrate by absorption onto a preparative grade C-18 resin. The concentrated material was further purified on a C-18 analytical column using both acetonitrile:water and methanol:water gradients. This highly purified fraction was a single component with a final specific activity of greater than 10(6) units per mg dry weight. Purified DfA is red having a broad visible absorbance at 500 nm and a ultraviolet (uv) absorbance at 290-300 nm. The red chromophore is sensitive to pH and to oxidation-reduction. 1H and 13C nmr studies with purified DfA indicate that it is a C11 compound with both polar and non-polar regions. The non-polar region has been identified as a hexanone and is the same as the side chain of DIF from Dictyostelium discoideum. Purified DfA has been used in studies with the D factor non-producing mutant, tsg-119 cyc-1 aggA586 (A586), to show that neither production of glorin nor chemotactic sensitivity to glorin are affected by D factor. However, founder cells develop in A586 mutant populations only after addition of D factor. These data suggest that DfA may be necessary for induction of aggregate formation by aggregation-competent amoebae.

The disposition of quinfamide in the rat.

The disposition of quinfamide 1-(dichloroacetyl)-6-(2-furoyloxy)-1, 2, 3, 4-tetrahydroquinoline, an enteric anti-amoebic agent, was studied in the rat. A peak blood level equivalent to 2.3 micrograms/ml of quinfamide was observed at 7 hr following a 20 mg/kg oral dose. Urinary recovery of radioactivity was much higher (84%) following intravenous than oral (48%) administration. Drug levels, in all of the tissues examined. were low. The major pathways of quinfamide metabolism in the rat involve hydrolysis of one or both ester groups, acetylation of the de-acylated product to 1-acetyl-1, 2, 3, 4-tetrahydro-6-quinolinol, oxidation of this to the 1-glycolyl metabolite, and aromatization to 6-hydroxyquinoline.

Metabolism of amrinone in animals.

The biotransformation of 14C-amrinone was studied in rats, dogs, and monkeys by automated gradient high-performance liquid chromatography. The major pathways of metabolism elucidated are: A) glucuronidation at the primary amino nitrogen atom and/or the enolized oxygen atom of the pyridone ring; B) addition of glutathione at the pyridone 2-position and ultimate hydrolysis of this compound to the 2-S-cysteinyl metabolite; C) formation of the primary amino N-acetyl derivative and subsequent oxidation of this to the corresponding N-glycolate. In each species studied, urine was the primary route of elimination and unchanged amrinone was the major urinary excretion product, the other known pathways being: rat, A and C; dog, A and B; monkey, A.

Metabolism of arildone, an antiviral agent, in laboratory animals.

After arildone administration, four compounds were identified in the excreta of laboratory animals: unchanged drug, arildone; the O-desmethyl metabolite, 4-[6-(2-chloro-4-hydroxy)phenoxy]hexyl-3,5-heptanedione; the sulfate ester of 2-chloro-4-methoxyphenol; and a labile conjugate of chlorohydroquinone, tentatively characterized as the sulfate ester. The concentrations of each of these were determined in the urine and/or plasma of rats, dogs, and mice after administration of 14C-arildone.

Disposition of trilostane in the rat and monkey.

The metabolism of trilostane, a novel inhibitor of adrenal steroidogenesis, was studied in the rat and monkey. In the rat, a peak blood level, equivalent to 2 microgram/ml of trilostane, was observed following a 25 mg/kg oral dose; excretion was mainly via the feces. In the monkey, the peak plasma level, equivalent to 15 microgram/ml, was observed 2 hr after a 20 mg/kg oral dose; elimination of radioactivity was predominantly in the urine. The five major metabolites of trilostane in monkey urine have been isolated and partially characterized. The primary metabolic pathways involved hydroxylation and glucuronide formation.

Analysis of cyclazocine in plasma.

The analysis of plasma cyclazocine by two methods is described. The radioimmunoassay employed a 125I-labeled radioligand, rabbit antiserum, and sepration of bound from free cyclazocine with a second antibody. The radioimmunoassay was specific for cyclazocine and had a detection limit of approximately 20 pg/ml. The GLC method employed a mass spectrometer as the detector and had a detection limit of approximately 109 pg/ml. Both techniques had acceptable accuracy and precision when used to quantitate cyclazocine in dog and human plasma. The methods were used successfully to quantitate cyclazocine from beagle hounds receiving 0.5 mg of 3H-cylazocine/kg iv. The decline in plasma cyclazocine fitted a two-compartment body model with a mean plasma clearance rate of 39.2 liters/hr.

Plasma pentazocine radioimmunoassay.

A sensitive and specific radioimmunoassay of dog and human plasma pentazocine is described. Rabbit antiserum and the second antibody method separated bound from free pentazocine. The radioimmunoassay employed an 125I-labeled radioligand and required extraction from the sample prior to quantitation. The method had a detection limit of approximately 200 pg/assay tube (1 ng/ml). The assay was used successfully to measure pentazocine in the plasma of beagle hounds given 0.3 mg of pentazocine/kg iv. The decline in plasma levels fitted a two-compartment body model with a 100-min mean overall half-life and a 3.2-liters/hr mean plasma clearance rate.

Absorption and disposition of N-(1, 1-dimethylethyl)-N'-[2-(4-pyridinyl)-4-pyrimidinyl]urea in rats and dogs.

N-(1, 1-Dimethylethyl)-N'-[2-(4-pyridinyl)-4-pyrimidinyl]urea, Win 40, 882, was eliminated from the blood stream of dogs by two apparent first-order processes with alpha- and beta-phase half-lives of 0.2 hr and 1.4 hr, respectively. Radioactivity of the administered dose was excreted by rats in the feces and via the kidneys; about 40-45% of the dose was recovered in the feces, with the remainder in the urine, over a six day period. One of the terminal methyl groups of the tert-butyl moiety of Win 40,882 is sequentially oxidized by the rat to the alcohol, aldehyde and carboxylic acid. In addition, some of the aldehyde was further metabolized to generate two different monohydroxylated aldehydic metabolites; these hydroxyaldehydes accounted for less than 10% of the dose administered. An unusual metabolic pattern was noted in the excretion of Win 40,882. Over 30% of the urinary metabolites contained the carboxaldehyde function; only8% of the urinary radioactivity was represented by the further oxidation of the aldehyde group to generate the carboxylic acid.

Radioimmunoassay for danazol in human and monkey plasma.

A sensitive method is described for the radioimmunoassay of danazol in monkey and human plasma. Antiserum was developed in rabbits, and a second antibody was used to separate bound from free danazol. The radioimmunoassay was specific for danazol, and the limit of detection ranged from 1.4 to 2.8 ng/ml. Exogeneous danazol could be quantitated accurately in both monkey and human plasma. The radioimmunoassay results agreed with values obtained by inverse isotope dilution after intravenous administration of 14C-danazol to monkeys. The assay was used successfully to measure danazol in plasma from human volunteers receiving 200 mg of danazol.

Absorption, excretion and disposition of cyclindole in laboratory animals and human volunteers.

Radioactivity from orally administered single doses of cyclindole-14C was excreted primarily in the urine of the rat (99%/48 hr), monkey (80%/120 hr), and dog (70%/144 hr). Approximately 38-58% of a daily dose of cyclindole was recovered from 24-hr human urine, as determined by gas chromatography. Substantial amounts of unchanged drug were voided by dogs. Cyclindole was metabolized primarily by N-demethylation and/or hydroxylation in the 7-position. Hydroxylation at the 6-carbon atom was of minor importance in humans only; none of the animal species excreted the 6-hydroxy metabolite. Dogs were capable of N-demethylation, but no metabolites oxidized at the 6- or 7-carbon atoms were detected in dog urine.

Experimental antiulcer drugs. 1. Indole-1-alkanamides and pyrrole-1-alkanamides.

The synthesis and gastric antisecretory activity of a series of indole-1-alkanamides and pyrrole-1-alkanamides are presented. A marked elevation of the pH of the gastric secretions of the rat was observed after oral administration of 100 mg/kg of 2,3-dimethylindole-1-acetamide (2), -1-propionamide (8), and -1-butyramide (13). Replacement of either methyl group by a hydrogen atom or an ethyl radical resulted in greatly diminished activity. In the acetamide and propionamide series the 3-hydroxymethyl-2-methyl (14 and 15) derivatives exhibited activity but only when administered by the subcutaneous route. 2,3-Dimethylindole (18) was active and 2,3,4,5-tetramethylpyrrole-1-acetamide was moderately active. A number of the activ compounds were tested in the mouse mydriasis test for anticholinergic activity and found to be inactive. They were also found to be inactive in blocking histamine-induced acid secretion in the dog.

A gas chromatography-mass fragmentographic method for the assay of ajmaline and its monochloroacetyl ester.

A gas chromatograph-mass fragmentography method for simultaneous assay of 17-monochloroacetyl-ajmaline (MCAA) and its hydrolysis product, ajmaline, is described. Recovery of both compounds from whole blood averaged 71%. About 5% of MCAA was hydrolyzed during the assay procedure. The method was accurate and precise to within a few percent. It was suitable for assays of blood levels in the dog after an i.v. dose of 2 mg/kg or an oral dose of 5 mg/kg.