Biotransformation of tri-substituted methoxyamphetamines by Cunninghamella echinulata.

1. Four trimethoxyamphetamine analogues were incubated with the filamentous fungus Cunninghamella echinulata. 2. 2,4,5-Trimethoxyamphetamine and 2,5-dimethoxy-4-ethoxyamphetamine were poorly metabolized by C. echinulata ATCC 9244 and C. echinulata var. elegans ATCC 9245. 2,5-Dimethoxy-4-(n)-propoxyamphetamine was mainly metabolized through N-acetylation and O-dealkylation with minor amounts of several aliphatic hydroxylation metabolites formed. 2,5-Dimethoxy-4-methylthioamphetamine was extensively metabolized to the corresponding sulphoxide. 3. 2,5-Dimethoxy-4-methylthioamphetamine metabolism was inhibited by ethanol and quinidine. Sparteine did not inhibit the formation of the sulphoxide and may have shunted the substrate through alternate metabolic pathways. 4. Incubation conditions can affect the rate and extent of fungal biotransformation of 2,5-dimethoxy-4-methylthioamphetamine, and influence dextrose utilization, ammonia formation and pH.

Discriminative stimulus properties of substituted amphetamine derivatives.

Animals were trained to discriminate amphetamine (1 mg/kg) from saline in a fixed-ratio (FR 10), food-reinforced paradigm. Amphetamine-appropriate responding was engendered by the training dose, and by 3 mg/kg, while at lower doses there was a progressive decrease in the extent of responding on the drug-appropriate lever. The following three novel amphetamine derivatives were tested for their ability to produce amphetamine-appropriate responding: 2,5-dimethoxy-4-ethoxy-amphetamine (DMEA); 2,5-dimethoxy-4-methylthio-amphetamine (DMMTA), and 2,4,5-trimethoxy-amphetamine (TMA). DMEA produced only minimal (< 20%) amphetamine-appropriate responding over a dose range of 0.1-10 mg/kg. Substantial decreases in response rate limited testing of the other amphetamines to a dose maximum of 3 mg/kg, but over the range of 0.1-3.0 mg/kg there was little evidence for generalization. At 3 mg/kg of either DMMTA or TMA, only 2 of 10 animals completed at least one uninterrupted FR 10 on either lever, and with either compound only 1 of these 2 animals responded more than 50% on the drug-appropriate lever. Of the three compounds tested, DMMTA had the greatest response rate-decreasing effect.

The reinforcing and discriminative stimulus properties of para-ethoxy- and para-methoxyamphetamine.

The purpose of this study was two-fold: 1) to assess the degree to which para-methoxyamphetamine and para-ethoxyamphetamine maintain self-administration behavior, and 2) to determine the similarity or difference between these drugs and amphetamine in drug discrimination tests. Animals were trained to self-administer 0.3 mg/kg/infusion cocaine on a fixed-ratio 5 (FR5) schedule of reinforcement. Substitution of para-ethoxyamphetamine (PEA), para-methoxyamphetamine (PMA), or saline produced similar results; in all cases responding decreased substantially. A separate group of animals was trained to discriminate amphetamine (1 mg/kg) from saline in a fixed-ratio (FR10), food-reinforced paradigm. PEA and PMA produced only limited responding on the amphetamine-appropriate lever (maximum of approximately 30%). Both PMA and PEA had effects on response rate which were similar to those of amphetamine, although PMA had slightly greater rate-decreasing effects than the other two compounds. These data suggest that neither PMA nor PEA are reinforcing in rats, and do not possess amphetamine-like discriminative properties.

Effects of chronic treatment of rats with "designer" amphetamines on brain regional monoamines.

(+)-Amphetamine and two structurally related analogues, 4-methoxyamphetamine and a recent "designer drug," 4-ethoxyamphetamine, were given to rats via subcutaneous osmotic minipumps for 1-14 days. Regional brain levels of the drugs as well as monoamine neurotransmitters and some of their major acidic metabolites were determined. Amphetamine produced depletions of dopamine in the striatum after at least 3 days of treatment but not in the nucleus accumbens of olfactory tubercle, even after 14 days of treatment. In contrast, the two ring-substituted amphetamine analogues increased levels of the monoamines and decreased levels of their acid metabolites. These data indicate that the two ring-substituted amphetamine analogues, at least one of which is a potent hallucinogen, have potent monoamine oxidase inhibition properties that are sustained during chronic treatment. Furthermore, these two compounds do not share amphetamine's regionally selective neurotoxic effects on dopamine-releasing terminals, even though brain and striatal drug levels are the same or higher than those of amphetamine.

"Designer" amphetamines: effects on behavior and monoamines with or without reserpine and/or alpha-methyl-para-tyrosine pretreatment.

Rats were given single injections of vehicle or one of three doses of (+)-amphetamine (AM), 4-methoxyamphetamine (MA) or 4-ethoxyamphetamine (EA) after pretreatment with vehicle or reserpine, and vehicle or alpha-methyl-para-tyrosine (AMPT). EA is a "designer" drug that was recently seized from an illicit laboratory in Canada. Locomotion of the rats was recorded after treatment with the drugs, and whole brain levels of the drugs as well as monoamine neurotransmitters and their major acidic metabolites were then determined. Neither of the ring-substituted AM analogues influenced locomotion. AM induced locomotion in a dose-dependent manner, and this effect was blocked by AMPT but potentiated by reserpine. Brain concentrations of EA were lower than those of the other two drugs. The brain levels of monoamines and their metabolites indicate that AM releases a newly synthesized pool of dopamine which is transferred to vesicles after re-uptake. A very low dose of AM, but not higher doses, was found to elevate serotonin (5-hydroxytryptamine: 5-HT) levels independently of effects on catecholamines. Both MA and EA affected monoamine metabolites in a manner consistent with actions as reversible inhibitors of monoamine oxidase-an effect which has been previously demonstrated to be true for MA. Both drugs increased 5-HT levels at a very low dose, as did AM, but also increased noradrenaline levels at this dose. It is concluded that EA is not a psychomotor stimulant, but is similar in many of its effects to MA, a potent hallucinogen.

Biotransformation of 2-, 3-, and 4-methoxy-amphetamines by Cunninghamella echinulata.

1. Three methoxyamphetamine analogues have been incubated with Cunninghamella echinulata under different environmental and nutrient conditions. 2. The biotransformation of 4-methoxyamphetamine was inhibited by cobalt; the carbon source and other trace metals had no effect. The rate of biotransformation of 4-methoxyamphetamine and formation of 4-hydroxyamphetamine was greater in cultures incubated on 30 degrees angle brackets rather than flat. 3. Carbon monoxide, ethanol and quinidine had a significant effect on methoxyamphetamine metabolism. 4. Metabolism was influenced by the position of the methoxy side-chain and substrate concentration. In day 7 samples the relative order for biotransformation was 3- greater than 4- greater than 2-methoxyamphetamine. 5. O-Demethylation was the major metabolic route in the biotransformation of 4-methoxyamphetamine but occurred to a lesser extent with 3-methoxyamphetamine, and was only a trace pathway with 2-methoxyamphetamine. N-acetylation was a trace pathway.

Stability of methadone in four vehicles for oral administration.

The stability of methadone in vehicles commonly used for oral administration was determined. Solutions of methadone were prepared in (1) orange-flavored Tang, (2) grape-flavored Kool-Aid, (3) apple juice, (4) grape-flavored Crystal Light, and (5) grape-flavored Crystal Light plus 0.1% sodium benzoate. For each of the first four preparations listed, two solutions were formulated at each methadone hydrochloride concentration of 0.2, 0.8, and 1.5 mg/mL; one set of three concentrations of each solution was stored in a refrigerator at 5 degrees C for up to 55 days, and the other set was stored unprotected from light at 20-25 degrees C for up to 17 days. Only three Crystal Light plus sodium benzoate solutions were prepared at the same methadone concentrations and stored at 20-25 degrees C for up to 29 days. Methadone concentrations were measured by high-performance liquid chromatography. Methadone was stable (loss of potency, less than 5%) for up to 17, 11, 9, and 8 days when stored at 20-25 degrees C in Kool-Aid, Tang, apple juice, and Crystal Light, respectively, and for up to 29 days when stored at 20-25 degrees C in Crystal Light plus sodium benzoate. Methadone was stable for up to 55, 49, 47, and 34 days when stored at 5 degrees C in Kool-Aid, Tang, apple juice, and Crystal Light, respectively. All the solutions that did not contain sodium benzoate and were stored at room temperature displayed unacceptable microbial growth after approximately two weeks. No significant loss of methadone potency occurred in any of the vehicles for oral administration during the study.

Fungal metabolism of 4-substituted amphetamines.

1. rac.-4-Ethoxyamphetamine was incubated with 14 different yeast and fungal microorganisms. 4-Hydroxyamphetamine was the major metabolite; traces of N-acetyl-4-ethoxyamphetamine were also detected. 2. The major fungal (Cunninghamella) metabolite of 4-propoxyamphetamine and 4-benzyloxyamphetamine was 4-hydroxyamphetamine. The major metabolites of 4-methoxyamphetamine were N-acetyl-4-methoxyamphetamine and 4-hydroxyamphetamine. 3. Acetoin derivatives were formed when alkoxyamphetamine substrates were incubated in the presence of various fungi and yeasts. 4. The findings indicate that Cunninghamella echinulata may be a useful microbial model for drug disposition and interaction studies.

Antinociceptive activity of propionyl esters of morphine: a reevaluation.

The antinociceptive activity of the propionyl homologues of 3-O- and 6-O-acetyl- and 3,6-O-diacetylmorphine was re-investigated using materials of unequivocally established structure. Testing was in male Wistar rats at 60 min following subcutaneous administration by the rat tail-flick method. Results indicate that the antinociceptive activity of 3-O-propionylmorphine was similar to that of 3-O-acetylmorphine. 6-O-Propionylmorphine and 3,6-O-dipropionylmorphine had similar antinociceptive activity and, like 6-O-acetylmorphine, 6-O-propionylmorphine may be the pharmacologically active principle responsible for the antinociceptive activity of its disubstituted homologue.

Metabolic disposition of pyrazinamide in the rat: identification of a novel in vivo metabolite common to both rat and human.

Only limited studies have been reported on the disposition and pharmacokinetics of pyrazinamide (PZA) in both animals and humans. The metabolism of PZA has never been completely elucidated, consequently the metabolites of PZA, pyrazinoic acid (PA), 5-hydroxypyrazinoic acid (5-HOPA), and 5-hydroxypyrazinamide (5-HOPZA) were characterized and the disposition of PZA was examined following administration of 150 mg kg-1 of 14C-PZA to male Wistar rats. Comparable t1/2 for total radiolabel 14C (1.45 +/- 0.06 h) and PZA (1.39 +/- 0.04 h) in the blood compartment were observed. Cumulative 48 h excretion in urine and faeces accounted for 82.6 +/- 3.2 per cent and 11.0 +/- 1.3 per cent, respectively, of the dose administered. In the 0-6 h urine collections PA, 5-HOPA, 5-HOPZA, and PZA, respectively, accounted for 25.4 +/- 1.7, 17.7 +/- 1.2, 11.6 +/- 0.8, and 2.7 +/- 0.2 per cent of the administered dose. In the 6-12 h urine samples the proportions of PA and 5-HOPA increased statistically over the 0-6 h excretion whereas 5-HOPZA decreased. Administration of PZA to humans indicated 5-HOPZA was a major urinary metabolite in human. These data suggested that direct hydroxylation of PZA was an alternative pathway in the oxidation of PZA of importance to both human and rat.

Validation of liquid chromatographic method for assay of calcitriol and alfacalcidol in capsule formulations.

The validation of a liquid chromatographic procedure suitable for the determination of calcitriol and alfacalcidol in their respective formulations labeled to contain at least 0.25 micrograms drug per unit is described. The capsule content is diluted and chromatographed in 15-20 min on silica columns (5 micron) with a mobile phase of hexane-tetrahydrofuran-methylene dichloride-isopropanol (72 + 12 + 12 + 4, v/v) with detection at 254 nm. The calibration curve is linear. Recoveries of "spikes" averaged 101% with a standard deviation of 2%. Precision was better than 1.5%.

Quantitative analysis of ethynodiol diacetate and ethinyl estradiol/mestranol in oral contraceptive tablets by high-performance liquid chromatography.

A procedure is described for the assay of ethynodiol diacetate and ethinyl estradiol/mestranol by HPLC using two UV detectors at 210 and 280 nm. The system was acetonitrile 38% (v/v) in water as mobile phase on a 250 x 3.2-mm i.d. RP-2 column, with butylated hydroxytoluene as the internal standard. There was greater than 99% recovery from synthetic preparations and the coefficient of variation was greater than 2.0% for formulations.