Stanley F. Osman

Partitioning of Intermediary Carbon Metabolism in Vesicular-Arbuscular Mycorrhizal Leek.

Vesicular-arbuscular mycorrhizal fungi are symbionts for a large variety of crop plants; however, the form in which they take up carbon from the host is not established. To trace the course of carbon metabolism, we have used nuclear magnetic resonance spectroscopy with [13C]glucose labeling in vivo and in extracts to examine leek (Allium porrum) roots colonized by Glomus etunicatum (and uncolonized controls) as well as germinating spores. These studies implicate glucose as a likely substrate for vesicular-arbuscular mycorrhizal fungi in the symbiotic state. Root feeding of 0.6 mM 1-[13C]glucose labeled only the fungal metabolites trehalose and glycogen. The time course of this labeling was dependent on the status of the host. Incubation with 50 mM 1-[13C]glucose caused labeling of sucrose (in addition to fungal metabolites) with twice as much labeling in uncolonized plants. There was no detectable scrambling of the label from C1 glucose to the C6 position of glucose moieties in trehalose or glycogen. Labeling of mannitol C1,6 in the colonized root tissue was much less than in axenically germinating spores. Thus, carbohydrate metabolism of host and fungus are significantly altered in the symbiotic state.

Synergistic interaction between potato glycoalkaloidsα-solanine andα-chaconine in relation to destabilization of cell membranes: Ecological implications

In studies of the lysis of rabbit erythrocytes, red beet cells, andPenicillium notatum protoplasts by the potato glycoalkaloids α-solanine and α-chaconine, the latter was consistently the more membrane-disruptive compound and erythrocytes the more susceptible cell type. A 1∶1 mixture of solanine and chaconine produced pronounced synergistic effects in all three test systems. In beet cells, such effects were apparent from an early stage of treatment and persisted over a period of several hours. With erythrocytes and fungal protoplasts, the synergism was maximal with mixtures containing approximately 70% chaconine, whereas with beet cells it peaked at approximately 40% chaconine. Synergistic interactions between solanine and chaconine also occurred with regard to cholesterol binding in vitro, with a maximum response corresponding to the 50% mixture. The implications of these findings for the nature and efficacy of chemical defense systems in plants are discussed.

Structural studies of a phosphocholine substituted β-(1,3); (1,6) macrocyclic glucan from Bradyrhizobium japonicum USDA 110

In our previous in vivo 31P study of intact nitrogen-fixing nodules (Rolin, D.B., Boswell, R.T., Sloger, C., Tu, S.I. and Pfeffer, P.E., 1989 Plant Physiol. 89, 1238-1246), we observed an unknown phosphodiester. The compound was also observed in the spectra of isolated bacteroids as well as extracts of the colonizing Bradyrhizobium japonicum USDA 110. In order to characterize the phosphodiester in the present study, we took advantage of the relatively hydrophobic nature of the material and purified it by elution from a C-18 silica reverse-phase chromatography column followed by final separation on an aminopropyl silica HPLC column. Structural characterization of this compound with a molecular weight of 2271 (FAB mass spectrometry), using 13C-1H and 31P-1H heteronuclear 2D COSY and double quantum 2D phase sensitive homonuclear 1H COSY NMR spectra, demonstrated that the molecule contained beta-(1,3); beta-(1,6); beta-(1,3,6) and beta-linked non-reducing terminal glucose units in the ratio of 5:6:1:1, respectively, as well as one C-6 substituted phosphocholine (PC) moiety associated with one group of (1,3) beta-glucose residues. Carbohydrate degradation analysis indicated that this material was a macrocyclic glucan, (absence of a reducing end group) with two separated units containing three consecutively linked beta-(1,3) glucose residues and 6 beta-(1,6) glucose residues. The sequences of beta-(1,3)-linked glucose units contained a single non-reducing, terminal, unsubstituted glucose linked at the C-6 position and a PC group attached primarily to an unsubstituted C-6 position of a beta-(1,3)-linked glucose.

Cutinase production byStreptomyces spp.

Forty-fiveStreptomyces strains, including representatives of the plant pathogensS. acidiscabies, S. scabies, andS. ipomoea, were screened for ability to produce enzymes (cutinases) capable of hydrolyzing the insoluble plant biopolyester cutin. Initially, all strains were tested for production of extracellular esterase in liquid shake (250 rpm) cultures at room temperature in defined (glycerol-asparagine) or complex (tryptone-yeast extract with or without addition of mannitol) broth media supplemented with either tomato or apple cutin. Esterase activity was determined by a spectrophotometric assay utilizing the model substratep-nitrophenyl butyrate. Of the five strains exhibiting highest esterase activity, four (S. acidiscabies ATCC 49003,S. “scabies” ATCC 15485 and IMRU 3018, andS. badius ATCC 19888) were confirmed to produce enzymes with cutin-degrading activity (cutinases). Confirmation of extracellular cutinase production was accomplished by use of a new high-performance liquid chromatography method for separation and quantification of released cutin monomers. Monomer identification was confirmed by GC/MS analyses. Cutinase production was induced 2- to 17-fold by inclusion of cutin in the media. To our knowledge this constitutes the first report of cutinase production byStreptomyces spp. other thanS. scabies.

Glycoalkaloids and resistance to the Colorado potato beetle inSolanum chacoense Bitter

Resistance of 20 F2Solarium chacoense Bitter clones to the Colorado potato beetle,Leptinotarsa decemlineata (Say), was measured in a field test. Levels of total glycoalkaloids (TGA) and composición of the glycoalkaloid mixtures in foliage of the clones were also determined. Clones with either commersonine or dehydrocommersonine as the major foliar glycoalkaloid were significantly more resistant (lower damage ratings, fewer larvae and adult insects) than clones with solanine and chaconine. Damage ratings were negatively correlated (r = -0.67, p = 0.01) with foliar TGA levels. The results indicate that the types of glycoalkaloids present in the foliage ofS. chacoense may be as important as the level of TGA in limiting damage and numbers of insects.ResumenResistencia de 20 clones F2 deSolarium chacoense Bitter, al “Colorado potato beetle”,Leptinotarsa decemlineata (Say), se evaluó en una prueba de campo. Asimismo, se determinó niveles de glicoalcaloides totales (TGA) y composición de mezclas de glicoalcaloides en el follaje. Clones con cualquiera de los dos commersonine o dehydrocommersonine como el mayor componente de glicoalcaloide en el follaje fueron significativamente mas resistentes (lecturas bajas de daños, pocas larvas e insectos adultos) que clones con solanine y chaconine. Las lecturas de daño mostraron correlación negativa (r = -0.67, p = 0.01) con los niveles de glicoalcaloides totales en el follaje (GA). Los resultados indicaron que los tipos de glicoalcaloides presentes en el follaje deS. chacoense puede ser tan importante como el nivel de glicoalcaloides totales en la limitación del daño y número de los insectos.

Potato glycoalkaloids: Increases and variations of ratios in aged slices over prolonged storage

Four commercial cultivars of potatoes were maintained under normal storage conditions at 44 F for 34 weeks. Except for a final 10 week interval tubers were withdrawn at 6 week intervals. After slicing, a portion of the slices was immediately analyzed for total glycoalkaloid content. The remaining slices were aged for four days in the dark at room temperature, then similarly analyzed.The total glycoalkaloid content of the aged slices increased dramatically on aging. This increase on aging of slices reached a maximum early in storage then decreased gradually over the storage period. In determining the individual glycoalkaloids, α-solanine and α-chaconine both increased in these slices, but the greatest increase was in the former. Appearing solely in the aged slices of the Kennebec variety, α-and β-solamarine appeared early in the storage period and gradually decreased over the storage period. Analyses of the unaged slices indicated that the glycoalkaloid content and composition of the potato tubers was little affected by storage. Aging of potato sprouts did not change their glycoalkaloid content.ResumenSe mantuvieron cuatro cultivares comerciales de papa bajo condiciones normales de almacenamiento a 44°F por 34 semanas. Con excepción del intervalo final de 10 semanas, los tubérculos se sacaron a intervalos de 6 semanas. Después de obtener discos de los tubérculos, una porción de los discos fue inmediatamente analizada para determinar el contenido de glicoalcaloides totales. El resto de los discos se envejecieron en la oscuridad, a temperatura de cuarto por cuatro días, luego se analizaron en forma similar.El contenido de glicoalcaloides totales aumentó dramáticamente en los discos envejecidos. Este aumento con la edad de los discos, alcanzó un máximo al inicio del almacenamiento, luego disminuyó gradualmente conforme avanzó el período de almacenamiento. En la determination de los glicoalcaloides individuales, α-solanina y α-chaconina aumentaron en los discos, pero el aumento más grande fue con el prirnero. Apareciendo solos en los discos envejecidos de la variedad Kennebec, α-y β-solamarina apareció al inicio del período de almacenamiento y gradualmente disminuyó durante el período de almacenamiento. Los análisis de los discos sin almacenamiento indicaron que el contenido de glicoalcaloides y la composición de los mismos en los tubéculos de papa fueron muy poco afectados por el almacenamiento. El envejecimiento de los brotes de papa no cambió su contenido de glicoalcaloides.

Commersonine, a new glycoalkaloid from two Solanum species

Abstract Selected plant introduction lines of S. chacoense and S. commersonii contain two major glycoalkaloids, demissine and a new compound called commersonine. In contrast, other plant introduction lines of S. chacoense contain only solanine and chaconine as the major glycoalkaloids. The isolation and characterization of the new glycoalkaloid is described.

Isolubimin: A possible precursor of lubimin in infected potato slices

Abstract Potato slices treated with spirovetiva-1(10),11-diene-2-one yield lubimin and rishitin within 24 hr. A vetispirane which has not been detected in fungally infected potatoes was also isolated. This compound, isolubimin, appears to be an intermediate in the conversion of the above spirovetivone to lubimin.

Separation of lactose, lactobionic acid and lactobionolactone by high-performance liquid chromatography☆

Abstract Complete separation of the three title compounds has been achieved for the first time on either a β-cyclodextrinor an aminopropyl-silica gel-bonded-phase HPLC column eluted with acetonitrile-aqueous buffer mixtures. An HPLC system using either of these stationary phases and a refractive index detector directly quantified lactose, lactobionic acid and lactobionolactone at levels between 0.3 and 40 μg of analyte per injection. Simple separations of lactose and lactobionic acid were also readily accomplished on a calcium-form cation-exchange type column eluted with 1.2 m M CaSO 4 solution. These methods are useful for monitoring the chemical or enzymatic conversion of lactose into the valuable derivatives, lactobionic acid and lactobionolactone.

Isoflavone glucoside stress metabolites of soybean leaves

Abstract The isoflavone glucosides daidzin, genistin and ononin, the isoflavones daidzein and formononetin, and glyceollins I-III accumulated in soybean leaves inoculated with phytopathogenic bacteria. Treatment of leaves with sodium iodoacetate or yeast extract also led to isoflavonoid accumulation. Various other stress-inducing treatments were not effective. Bacterially-induced accumulation of isoflavone glucosides and the occurrence of ononin and formononetin in soybean are reported for the first time.