Metabolism and excretion of acetylchromenes by the migratory grasshopper

The extent of metabolism and excretion of three acetylchromenes (two toxic, one relatively nontoxic) were examined in adult migratory grasshoppers (Melanoplus sanguinipes) following topical administration. Both the total amount excreted (parent plus metabolites) and the proportion of parent compound in the excreta were inversely correlated with contact toxicity. Both toxic and nontoxic acetylchromenes are rapidly absorbed from the cuticle, with maximum excretion of parent and metabolite chromenes from 4 to 8 h posttreatment in each case. Much of the applied compounds (60–80%) apparently remains within the insect, and cannot be recovered by extraction of the insect. Metabolites formed result from simple oxidative and reductive transformations. For all of the compounds tested (including the allatocidin precocene II), the major mode of metabolism results from aliphatic hydroxylation of one of the geminal methyl groups on the chromene. No conjugated metabolites were found in the excreta.     

Infrared and Raman studies show that poly(dA).poly(dT) and d(AAAAATTTTT)2 exhibit a heteronomous conformation in films at 75% relative humidity and a B-type conformation at high humidities and in solution

The decadeoxynucleotide d(AAAAATTTTT)2 in duplex form and the double-helical polynucleotide poly(dA).poly(dT) have been studied by Raman and infrared (IR) spectroscopy under a variety of environmental conditions. The IR spectra have been taken of cast films and compared to the IR spectra of the alternating poly(dA-dT), which shows clear B-genus and A-genus vibrational spectra under conditions of high (greater than 92%) and low (75%) relative humidity (RH). From the IR data, it is shown that d-(AAAAATTTTT)2 and poly(dA).poly(dT) adopt a B-genus conformation in films with high water content. When the relative humidity of the film is decreased, the IR spectra reflect a gradual evolution of the geometry of both d(AAAAATTTTT)2 and poly(dA).poly(dT) into a form intermediate between the B genus and A genus, but the IR spectrum of a pure A genus has not been obtained. In these DNAs at 75% RH, the IR bands of adenosine have the same frequencies as those found in poly(dA-dT) at 75% RH where the local furanose conformation is C3' endo/anti, but the thymidine frequencies do not resemble those of poly(dA-dT) at 75% RH but rather those of poly(dA-dT) at high humidities. It is concluded that both poly(dA).poly(dT) and d(AAAAATTTTT)2 adopt a fully heteronomous duplex geometry in cast films at low humidity. For studies in aqueous solution the Raman effect was employed. As a model for the heteronomous conformation in solution, the duplex poly(rA).poly(dT) was used.(ABSTRACT TRUNCATED AT 250 WORDS)     

Relationships between structure, toxicity and genetic effects in Salmonella typhimurium and Saccharomyces cerevisiae for substituted aniline mustards

A series of 4-substituted aniline mustards of widely varying reactivities have been evaluated for their mutagenic effects in Salmonella typhimurium strains of varying uvrB gene and plasmid status, and for their ability to cause mitotic crossing-over in Saccharomyces cerevisiae. The 4-methyl aniline mustard N,N-bis(2-chloroethyl)-4-methylaniline and its corresponding half-mustard N-(2-chloroethyl)-4-methylaniline showed widely different effects in the various bacterial strains, with the half-mustard being much less toxic than the full mustard in the uvrB- strain TA100. However, in the uvrB+ strain TA1978+, possessing an intact excision repair system, both compounds were equally toxic and the full mustard was the more mutagenic. Both compounds were equally effective in promoting mitotic crossing-over in yeast. For a series of 4-substituted full mustards, the toxicity in S. typhimurium strain TA100 correlated with substituent electronic parameters in the same way as does mammalian cell toxicity, supporting the view that the primary mode of toxicity is via DNA cross-linking, even for unreactive analogues. However, there were no obvious correlations between substituent physiochemical properties and mutagenic potential in bacteria, suggesting that mutagenic events are subject to a variety of influences other than the reactivity of the mustard group. In contrast, the most chemically reactive compounds were the most toxic and most recombinogenic in yeast.     

Cloning of the egl gene of Pseudomonas solanacearum and analysis of its role in phytopathogenicity.

The egl gene of Pseudomonas solanacearum was cloned on a cosmid and expressed in Escherichia coli. Restriction endonuclease mapping, transposon mutagenesis, and subclone analysis showed that the egl gene was located on a 2.7-kilobase XhoI-SalI P. solanacearum DNA fragment. Immunoabsorption experiments and sodium dodecyl sulfate-polyacrylamide gel electrophoretic analysis showed that the egl gene encodes the 43-kilodalton endoglucanase that is the major excreted endoglucanase of P. solanacearum. In E. coli, the egl gene appeared to be expressed from its own promoter, but its product was restricted to the cytoplasm. The cloned egl gene was mutagenized with Tn5 and used to specifically mutate the chromosomal egl gene of P. solanacearum by site-directed mutagenesis. The resultant mutant was identical to the wild-type strain in production of extracellular polysaccharide and extracellular polygalacturonase as well as several other excreted proteins but produced at least 200-fold less endoglucanase. This mutant strain was significantly less virulent on tomato than the wild-type strain in plant bioassay experiments. Virulence of the endoglucanase-deficient strain was restored to near wild-type levels by complementation in trans with the cloned egl gene, indicating that the egl gene is important but not absolutely required for pathogenesis.     

Possible mode of action of a photosynthetic inhibitor produced byPandorina morum

The mode of action of the photosynthetic inhibitor produced byPandorina morum was examined by exposingVolvox globator and isolated spinach chloroplasts to a partially purified inhibitor preparation. Oxygen evolution ofVolvox, whole chloroplasts, and broken chloroplasts (-Calvin cycle) was reduced indicating that the substances inhibit the light reactions of photosynthesis. Oxygen evolution studies of other Volvocaceae confirmed the observation thatPandorina morum is not significantly influenced by its own inhibitor. Molecular weight approximation by gel filtration established that the inhibitor has a low, molecular weight (probably below 100 mw).     

Methylmercury-Induced Elevations in Intrasynaptosomal Zinc Concentrations: An 19F-NMR Study

Abstract: Methylmercury (MeHg) increases the concentration of intracellular Ca²⁺ ([Ca²⁺]_i) and another endogenous polyvalent cation in both synaptosomes and NG108-15 cells. In synaptosomes, the elevation in [Ca²⁺]_i was strictly dependent on extracellular Ca²⁺ (Ca²⁺_e); similarly, in NG108-15 cells, a component of the elevations in [Ca²⁺]_i was Ca²⁺_e dependent. The MeHg-induced elevations in endogenous polyvalent cation concentration were independent of Ca²⁺_e in synaptosomes and NG108-15 cells. The pattern of alterations in fura-2 fluorescence suggested the endogenous polyvalent cation may be Zn²⁺. Using ¹⁹F-NMR spectroscopy of rat cortical synaptosomes loaded with the fluorinated chelator 1,2-bis(2-amino-5-fluorophenoxy)ethane-N,N,N′,N′-tetraacetic acid (5F-BAPTA), we have determined unambiguously that MeHg increases the free intrasynaptosomal Zn²⁺ concentration ([Zn²⁺]_i). In buffer containing 200 µM EGTA to prevent the Ca²⁺_e-dependent elevations in [Ca²⁺]_i, the [Zn²⁺]_i was 1.37 ± 0.20 nM; following a 40-min exposure to MeHg-free buffer [Zn²⁺]_i was 1.88 ± 0.53 nM. Treatment of synaptosomes for 40 min with 125 µM MeHg yielded [Zn²⁺]_i of 2.69 ± 0.55 nM, whereas 250 µM MeHg significantly elevated [Zn²⁺]_i to 3.99 ± 0.68 nM. No Zn²⁺ peak was observed in synaptosomes treated with the cell-permeant heavy metal chelator N,N,N′,N′-tetrakis(2-pyridylmethyl)ethylenediamine (TPEN, 100 µM) following 250 µM MeHg exposure. [Ca²⁺]_i in buffer containing 200 µM EGTA was 338 ± 26 nM and was 370 ± 64 nM following an additional 40-min exposure to MeHg-free buffer. [Ca²⁺]_i was 498 ± 28 or 492 ± 53 nM during a 40-min exposure to 125 or 250 µM MeHg, respectively. None of the values of [Ca²⁺]_i differed significantly from either pretreatment levels or buffer-treated controls.     

Detection and separation of the anthrapyrazole CI-941 and its metabolites in serum and urine by high-performance liquid chromatography

A high-performance liquid chromatographic method using ion-pairing chromatography on reversed-phase C₁₈ material with a mobile phase of acetonitrile—water (19:81, v/v) containing 5 mM 1-pentanesulfonic acid was developed for the detection and separation of the anthrapyrazole CI-941 (I) and its metabolites. After sample clean-up with solid-phase extraction, I and its metabolites were measurable at a wavelength of 491 nm. A detection limit of 5 ng/ml was achievable for I. The dicarboxylic acid derivative and the isomers of the monocarboxylic acid derivative could be separated. Application of the method to a human pharmacokinetic study showed two and four metabolites of I in serum and urine respectively.