Cytochrome P-450-catalysed monoterpenoid oxidation in catmint (Nepeta racemosa) and avocado (Persea americana); evidence for related enzymes with different activities.

A cytochrome P-450 present in ripening avocado (Persea americana) fruit mesocarp (CYP71A1) had previously been shown to metabolize the monoterpenoids nerol and geraniol (Hallahan et al. (1992) Plant Physiol. 98, 1290-1297). Using DNA encoding CYP71A1 as a hybridization probe, we have shown by Southern analysis that a related gene is present in the catmint, Nepeta racemosa. RNA blot analysis, together with Western analysis of catmint leaf polypeptides using avocado cyt P-450 antiserum, showed that a closely related gene is expressed in catmint leaves. Cytochrome P-450 in catmint microsomes catalysed the specific hydroxylation of nerol and geraniol at C-10, whereas avocado CYP71A1, in either avocado microsomes or heterologously expressed in yeast, catalysed 2,3- or 6,7-epoxidation of these substrates. These results suggest that orthologous genes of the CYP71 family are expressed in these two plant species, but catalyse dissimilar reactions with monoterpenoid substrates.

Glucosinolate Biosynthesis (Further Characterization of the Aldoxime-Forming Microsomal Monooxygenases in Oilseed Rape Leaves).

The initial steps in glucosinolate biosynthesis are thought to proceed from amino acids, via N-hydroxy amino acids, to aldoximes. We showed previously that microsomes from green leaves of oilseed rape (Brassica napus cv Bienvenu) contain two distinct monooxygenases that catalyze the conversion of homophenylalanine and dihomomethionine to their respective aldoximes. Further characterization of these enzymes has now demonstrated that the latter enzyme catalyzes the NADPH-dependent oxidative decarboxylation of two higher homologs of methionine, in addition to dihomomethionine. No activity was found for either enzyme with L-methionine, DL-homomethionine, L-phenylalanine, L-tyrosine, or L-tryptophan. Both of these rape monooxygenase activities are dependent on O2, not requiring any other O2 species or radical. The presence of an unoxidized sulfur atom and its relative position in the side chain of the aliphatic substrates are important for binding to the active site of the methionine-homolog enzyme. Neither enzyme has any characteristics of a cytochrome P450-type enzyme, and antiserum raised against cytochrome P450 reductase did not significantly inhibit monooxygenase activity.

Cigarette smoking and theophylline metabolism: effects of cimetidine.

The inhibition of theophylline metabolism by cimetidine was investigated in young male cigarette smokers (greater than 20 cigarettes/day) and nonsmokers by stable isotope methodology. Subjects received oral theophylline (510 mg/day) for 14 days and cimetidine (1200 mg/day) over days 1 to 7 or 8 to 14. On days 7 and 14, a tracer dose (10 mg) of stable isotope-labeled theophylline was injected intravenously with the oral dose of theophylline. Serial plasma samples were then obtained for 24 hours and both molecular forms of theophylline were assayed by mass spectrometry after purification by HPLC. Theophylline bioavailability, volume of distribution, and protein binding were of the same order in both groups and were not affected by cimetidine. Although the basal theophylline elimination rate constant was 46% greater and clearance was 54% greater in smokers than in nonsmokers, the proportionate changes in steady-state plasma concentrations, t1/2, and clearance due to cimetidine were much the same in both groups. Plasma thiocyanate concentrations were higher in smokers than in nonsmokers and were related to theophylline clearance. Our findings indicate that cimetidine inhibits theophylline metabolism to a similar extent in both smokers and nonsmokers. Determination of plasma thiocyanate levels may be valuable in the prediction of theophylline clearance.

Loprazolam. A preliminary review of its pharmacodynamic and pharmacokinetic properties and therapeutic efficacy in insomnia.

Loprazolam is a 1,4-benzodiazepine with hypnotic properties, advocated for use in acute or chronic insomnia. As loprazolam has a half-life of 7 to 8 hours in healthy adults it may have advantages over longer-acting hypnotics, particularly when residual sedative effects on the day after ingestion are undesirable, although at doses greater than 1 mg residual sedation may occur. In addition, it may reduce daytime anxiety, following a hypnotic dose the night before, more effectively than the short-acting drug, triazolam. In short term comparative studies loprazolam was clearly superior to placebo, and was at least as effective as triazolam, flurazepam, nitrazepam, flunitrazepam or temazepam in hastening sleep onset, reducing nocturnal awakenings and increasing total sleep duration and quality. In the small number of patients with chronic insomnia who have received extended treatment with loprazolam, no evidence of tolerance has occurred, although rebound insomnia was evident 3 days after drug withdrawal in several studies. Thus, with its 'intermediate' elimination half-life, loprazolam would appear to have some potential advantages over both long- and short-acting hypnotics in selected patients, although further studies are needed to fully elucidate its place in therapy.

Ciclopirox olamine 1% cream. A preliminary review of its antimicrobial activity and therapeutic use.

Ciclopirox olamine is a substituted pyridone antimycotic, unrelated to the imidazole derivatives, with activity against a broad spectrum of dermatophytes, yeasts, actinomycetes, molds, other fungi, and a variety of Gram-positive and Gram-negative bacteria. The efficacy of ciclopirox olamine cream has been demonstrated in open and placebo-controlled studies in patients with superficial dermatophyte or yeast infections, and in double-blind comparative trials in patients with dermatomycoses, topical ciclopirox olamine was comparable to or better than clotrimazole in efficacy and caused a similar number of side effects. Ciclopirox olamine penetrates through fingernails and in preliminary studies has been successfully used in onychomycoses. However, further studies are needed to establish the role of ciclopirox in the treatment of onychomycoses and dermatomycoses relative to that of the more recently introduced antigungal agents.

Amoxapine: a review of its pharmacology and efficacy in depressed states.

Amoxapine is an N-demethylated dibenzoxazepine closely related in the neuroleptic loxapine. Its tricyclic structure appears to give it antidepressant properties resembling imipramine and amitriptyline. In uncontrolled trials it was shown to have antidepressant activity in usual doses up to 200 to 400mg daily. In placebo and double-blind controlled studies comparing amoxapine with the standard tricyclic antidepressants imipramine and amitriptyline, it was shown to be comparable in efficacy with a possibly somewhat faster onset of improvement of selected symptoms of depression in some studies. Because of the small study groups and lack of placebo control, many reports do not show statistically significant differences of treatment over standard drugs. To date there have been no studies comparing amoxapine with electroconvulsive therapy. Side effects were qualitatively similar to standard drugs with a suggestion that in standard doses or overdose myocardial effects are mild. However, the final place of amoxapine in the therapy of depressed states is still to be decided.

Alprazolam: a review of its pharmacodynamic properties and efficacy in the treatment of anxiety and depression.

Alprazolam is a triazolobenzodiazepine which is related to diazepam and other 1,4-benzodiazepines, and has a similar pharmacological profile. Relative to the newer benzodiazepines, alprazolam has an intermediate half-life of 10 to 12 hours in healthy young subjects. In placebo-controlled and double-blind comparative trials in patients with anxiety, alprazolam was of comparable efficacy to diazepam and generally caused a lower incidence of drowsiness. Alprazolam has antidepressant activity and has been shown to be similar in efficacy to imipramine in the treatment of unipolar depression. Thus, alprazolam may be particularly useful in patients with mixed anxiety/depression. However, its general acceptance as an antidepressant awaits further studies.

In vitro metabolism of 1-phenyl-2-(n-propylamino) propane (N-propylamphetamine) by rat liver homogenates.

In vitro incubation of (+/-)-N-(n-propyl)amphetamine (NPA) with the 12000 g supernatant fraction of rat liver hmogenate resulted in the formation of two N-oxygenated products identified as N-hydroxy-1-phenyl-2(n-propylamino)propane and N-[(1-methyl-2-phenyl) ethyl]-1-propanamine N-oxide by g.l.c., g.l.c.-m.s. and t.l.c. Amphetamine, phenylacetone,...

The induction of somatic mutations in mouse embryos by benzo(a)pyrene.

With the object of establishing a test for specific locus mutations in the somatic cells of mice, the effect of benzo(a)pyrene on the developing pigment cells of mouse embryos was investigated. Embryos were heterozygous for genes affecting coat-colour so that mutations could be scored as spots on the adult mouse. If the pregnant female was treated, with 2 mg/day orally, before day 7 or after day 10 of pregnancy no mutations were seen in the offspring. Treatment at any time from day 7 to day 10 was effective in producing mutations with no day giving a peak response. The effect of treatment on consecutive days appears to be additive. It is suggested that further chemicals be investigated lest differing rates of uptake and metabolism should alter the period in which treatment should be given in order to detect a mutagenic effect.

Urinary excretion of phenolic metabolites of N-(n-propyl)amphetamine in man.

In vivo metabolism of (+)-N-(n-propyl)amphetamine (NPA) in man (2 subjects) produced, among other products, two conjugated phenolic metabolites which are identified as 1-(4-hydroxyphenyl)-2-(n-propylamino)propane and 1-(4-hydroxy-3-methoxyphenyl)-2-n-propylamino)propane. In a 24 hr urine sample (pH uncontrolled), 13.4% of the NPA administered was excreted as the former, and 3.5% as the latter metabolite.

"Self-feeding" strain of Salmonella typhimurium with a mutation in the trpB gene and nutritional requirements of trpA gene mutants.

An indole-requiring (Ind(-)) mutant of Salmonella typhimurium, isolated from a culture of a leaky trpA mutant, was genetically analyzed by P22-mediated transduction. The mutation site giving the Ind(-) phenotype was shown to be in trpB, the second gene of the trp operon. A second mutation at this site resulted in change of nutritional requirement from indole to anthranilic acid (Anth(-)). This phenotype is normally associated with mutations in the first trp gene, trpA. However, the Anth(-) mutant also excreted anthranilic acid and showed "self-feeding" on unsupplemented media. Of two possible explanations for this aberrant phenotype, the first, that the trpB mutations may be in the "unusual" region, was dismissed on genetic evidence and on the biochemical evidence that an active anthranilate synthetase (AS) is produced. The alternative explanation, that the affected enzymatic activity, phosphoribosyl transferase, is unstable in vivo, but its AS component 2 activity is stable, is considered more probable.

Gene mapping in marsupials and monotremes. I. The chromosomes of rodent-marsupial (Macropus) cell hybrids, and gene assignments to the X chromosome of the grey kangaroo.

Somatic cell genetic mapping of marsupial and monotreme species will greatly extend the power of comparative gene mapping to detect ancient mammalian gene arrangements. The use of eutherian-marsupial cell hybrids for such mapping is complicated by the frequent retention of deleted and rearranged marsupial chromosomes. We used staining techniques, involving the fluorochromes Hoechst 33258 and chromomycin A3, to facilitate rapid and unequivocal identification of marsupial chromosomes and chromosome segments and to make chromosome assignment and regional localization of marsupial genes possible. Chromosome segregation in rodent-macropod hybrids was consistent with preferential loss of the marsupial complement. The extent of loss was very variable. Some hybrids retained 30% of the marsupial complement; some retained small centric fragments; and some, no cytologically identifiable marsupial material. We examined the chromosomes and gene products of a number of rodent-grey kangaroo Macropus giganteus hybrids, and have assigned the genes Pgk-A (phosphoglycerate kinase-A), Hpt (Hypoxanthine-phosphoribosyl transferase), and Gpd (Glucose-6-phosphate dehydrogenase) to the long arm of the kangaroo X chromosome, and provisionally established the gene order Pgk-A--Hpt--Gpd.

Cimetidine inhibits the in vitro N-demethylation of methadone.

The effect of cimetidine on the metabolism of methadone was investigated in hepatic microsomes isolated from male Sprague-Dawley rats. N-demethylation of methadone was determined by formation of formaldehyde. Cimetidine inhibited methadone N-demethylation in a noncompetitive manner with an IC50 of 5.3 X 10(-4) M. This observation is consistent with previous reports of microsomal enzyme inhibition by cimetidine and suggests that caution must be exercised when methadone and cimetidine are co-administered to patients in order to avoid excessive sedation or respiratory depression.

The aphid sex pheromone cyclopentanoids: synthesis in the elucidation of structure and biosynthetic pathways.

Identification of a range of aphid sex pheromones as comprising the cyclopentanoids (4aS,7S,7aR)-nepetalactone, (1R,4aS,7S,7aR)-nepetalactol and the (1S)- and (1R,4aR,7S,7aS)-nepetalactols required samples authenticated by 1H and 13C NMR. These and related compounds were provided by small scale synthesis and extraction from plants in the genus Nepeta (Lamiaceae). The subsequent discovery that the synthetic sex pheromones could attract males, and also parasitic wasps that attack aphids, has created a need for large scale syntheses of the cyclopentanoids. This is afforded by cyclisation of the 8-oxo-1-enamine of citronellal as originally developed by Schreiber and co-workers (1986). Investigation into the biosynthesis of the cyclopentanoids by plants for exploiting aphid sex pheromones in crop protection by means of molecular biology required synthesis of putative biosynthetic intermediates, some with radioactive isotopic labelling, particularly 8-oxidised monoterpene alcohols and aldehydes.

9-Methylgermacrene-B; proposed structure for novel homosesquiterpene from the sex pheromone glands ofLutzomyia longipalpis (Diptera: Psychodidae) from Lapinha, Brazil.

The principal volatile component (99 + %) of the sex pheromone glands ofLutzomyia longipalpis from Lapinha, Brazil, has been isolated and characterized as a novel homosesquiterpene with the specific structure proposed as 9-methylgermacrene-B, (E,E)-8-(1-methylethylidenyl)-1,5,10-trimethyl-1,5-cyclodecadiene.

3-Methyl-α-himachalene: Proposed structure for novel homosesquiterpene sex pheromone ofLutzomyia longipalpis (diptera: Psychodidae) from Jacobina, Brazil.

The principal behaviorally active volatile component (ca. 90% +) of the sex pheromone glands ofLutzomyia longipalpis from Jacobina. Brazil, has been isolated and characterized as a novel homosesquiterpene with the structure 3-methyl-α-himachalene (C16H26). A minor component (ca. 10%) of the gland extract has also been identified as the sesquiterpeneα-himachalene (C15H24). This work confirms that there are at least 3 members of theL. longipalpis species complex.

In vivo metabolism of N-alkylamphetamines in the rat--the effect of N-alkyl chain length on oxidation of the aromatic ring.

In vivo metabolism of N-alkylated amphetamines in the rat produced 4-hydroxy- and 4-hydroxy-3-methoxy-metabolites. The relative quantities of the phenolicmetabolites formed were dependent upon the length of the N-alkyl substituent.