Stereochemical inversion of pyrrolizidine alkaloids byMechanitis polymnia (Lepidoptera: Nymphalidae: Ithomiinae): Specificity and evolutionary significance

Pyrrolizidine alkaloids (PAs), acquired by adults or larvae of Danainae and Ithomiinae butterflies and Arctiidae moths from plants, protect these lepidopterans against predators and are biosynthetic precursors of male sex pheromones. The investigation of PAs in many species of wild-caught adults of Ithomiinae showed lycopsamine (1) [(7R)-OH, (2S)-OH, (3S)-OH] as the main alkaloid. In incorporation experiments, PA-free (freshly emerged) adults of the ithomiineMechanitis polymnia were fed seven PAs: lycopsamine and four of its known natural stereoisomers—indicine (2) [(7R)-OH, (2R)-OH, (3S)-OH], intermedine (3) [(7R)-OH, (2S)-OH, (3R)-OH], rinderine (4) [(7S)-OH, (2S)-OH, (3R)-OH], and echinatine (5) [(7S)-OH, (2S)-OH, (3S)-OH], and two PAs without the 7-OH: supinine (6) [(2S)-OH, (3R)-OH] and amabiline (7) [(2S)-OH, (3S)-OH]. Males epimerized PAs 3, 4, and 5 mainly to lycopsamine (1). Females fed these same three PAs changed a smaller proportion to lycopsamine; their lesser capacity to modify PAs corresponds to their normal acquisition of already transformed PAs from males during mating rather than through visits of adults to plant sources of PAs. The alkaloids1 and2, both 7R and 3S, were incorporated without or with minimum change by males and females. Feeding experiments with6 and7 (males only) showed an inversion at the 3 center of6 and no change in7. The inversion from 7S to 7R (probably via oxyreduction) may be closely related to the evolution of acquisition of PAs by butterflies and moths. Two hypotheses are discussed: (1) The ancestral butterflies are probably adapted to tolerate, assimilate, and use (7R)-PAs (most common in plants; all widespread 1,2-unsaturated macrocyclic PA diesters show this configuration). The development of (7R)-PA receptors in the butterflies could lead to a specialization on this configuration in two ways: to help find PA plants and to utilize these components in sexual chemical communication. A later appearance of (7S)-PAs in plants could have selected an enzymatic system for the inversion of this chiral center in order to continue producing (7R)-PA-derived pheromones. (2) The inversion would be due to the evolution of a enzyme system specialized in the transport of (7R)-PAs to the integument; the failure of this system to carry (7S)-PAs led to an enzymatic system to invert them to transportable (7R)-PAs. In this case, the 7R configuration is an effect and not a cause of (7R)-PA-derived pheromones. In both hypotheses, the partial inversion of the 3-asymmetric center, when the butterfly was fed intermedine (3), rinderine (4), and supinine (6), could be fortuitous due to the conformation of the molecule and/or the enzymatic system.This paper is part of the doctoral thesis of J.R.T.     

Dual role of pyrrolizidine alkaloids in nectar

Pyrrolizidine alkaloids occur in several plant families, attracting ithomiine and danaine butterflies that specialize on the flowers. I show that pyrrolizidine alkaloids in artificial nectar also inhibit at least one butterfly,Agraulis vanillae (Nymphalidae: Heliconiinae), a more general forager. Inhibition was demonstrated in two ways: quantity of nectar consumed and number of artificial flowers visited. The amount of sucrose solution consumed by individual butterflies was measured using a microcapillary tube. Number of flower visits was determined using an array of artificial orange and yellow flowers.A. vanillae drank less sucrose solution with monocrotaline, a pyrrolizidine alkaloid, than without. When monocrotaline was placed into yellow flowers,A. vanillae learned to visit predominantly orange flowers. Evolutionarily, pyrrolizidine alkaloids in nectar may represent an adaptation to exclude butterflies. Ithomiines and danaines, seeking these compounds in larval food plants, were not excluded. Alternatively, ithomiines and danaines overcame the nectar defense. In either case, the plant effectively increased floral constancy by inhibiting generalist butterflies and attracting specialists.     

The chemical basis of attraction of ithomiine butterflies to plants containing pyrrolizidine alkaloids

The plantHeliotropium indicum L. (Boraginaceae) contains pyrrolizidine alkaloids. When dried, it is a powerful attractant for male ithomiine and danaine butterflies, which congregate and feed at its dead shoots. The butterflies use alkaloids derived from the plants for the formation of chemicals with pheromone/allomone activity. Baiting with alkaloids and esterifying acids, which form a part of the alkaloid molecules, indicates that a volatile product derived from the esterifying acids attracts males to the plants, where intact alkaloids then act as phagostimulants.     

Lepidoptera and pyrrolizidine alkaloids Exemplification of complexity in chemical ecology

Pyrrolizidine alkaloids (PAs) are defensive secondary metabolites found in numerous plant groups. Various insects belonging to different orders have special requirements for these compounds and sequester them from such plants for their own defense and often as pheromone precursors. The fitness of these insects depends on PAs and, in some cases, PAs even act as regulators of androconial organ development. This article discusses selected behavioral, chemical, physiological, and phylogenetic aspects of insect-PA relationships, and raises questions about the complex interactions of the variety of PA-related adaptations as they occur among a diverse array of species. Although many superficial similarities are recognized, few generalizations can yet be drawn. However, insect-PA relationships not only exemplify basic features of chemical ecology but illustrate a multiplicity of aspects and adaptations, which we should expect to find in any thorough study of insect-plant relationship.Paper presented at the Symposium Sequestration of Natural Products by Insects during the 5th Annual Meeting of the International Society of Chemical Ecology, Athens, Georgia, June 24–27, 1988.     

Seducin,male sex pheromone of the cockroachNauphoeta cinerea: Isolation,identification, and bioassay

The male sex pheromone of the cockroachNauphoeta cinerea was isolated from adult sternal glands and identified by gas chromatography-mass spectrometry. This pheromone, which attracts females from a distance, is a multicomponent blend. It is composed principally of three compounds: 3-hydroxy-2-butanone (acetoin), 2-methylthiazolidine, and 4-ethyl-2-meth-oxyphenol. A mixture of synthetic compounds in a ratio of 441 (2700 ng) elicits maximal response and short latency of response by receptive females, similar to that induced by male sternal gland extracts. Tested separately, each compound induces a positive response, but the moving times spent by females are generally longer than with the mixture or extracts. These three volatile compounds also are found in small amounts (about 1/40 of sternal glands) in tergal gland secretions licked by females. Acetoin also is found in the sternal and tergal glands of two closely related speciesLeucophaea maderae andHenschoutedenia flexivitta. The biological relationship of each compound and pheromone blend to behavioral response is discussed.     

Importance of quinolizidine alkaloids in the relationship between larvae ofUresiphita reversalis (Lepidoptera: Pyralidae) and a host plant,Genista monspessulana

Larvae ofUresiphita reversalis feed almost exclusively on legumes in the tribe Genisteae, which characteristically contain a variety of quinolizidine alkaloids. The larvae are aposematic, and onGenista monspessulana, a major host in California, they feed on the youngest leaves, at the periphery of the plant. These leaves, which were preferred over older foliage in choice tests, contained four to five times the level of alkaloid found in older leaves. The major alkaloids detected in these plants were dehydroaphylline andN-methylcytisine, together accounting for 74% of the total. Preliminary analyses showed the alkaloid profile of exuviae from larvae feeding on these plants was very similar to that of the plants. Two alkaloids, sparteine and cytisine, which are known components of some hosts ofU. reversalis, were phagostimulants for fifth-instar larvae when added to sucrose-impregnated glass-fiber disks. In addition, when sparteine was added to foliage ofG. monspessulana, effectively doubling the percent dry weight of alkaloid, the growth rate of late-instar larvae was positively affected. Cytisine added to plants had no discernible effect on growth of larvae. Alkaloid levels in larvae and in their frass were proportional to levels in the plants on which they fed. Although the majority of alkaloid was excreted, that which was sequestered by the insect was found entirely in the integument, possibly confering some protection from predators.     

Behavioral responses of male turnip moths,Agrotis segetum,to sex pheromone in a flight tunnel and in the field

The response of individual male turnip mothsAgrotis segetum was observed in a sustained flight tunnel to a mixture of decyl acetate, (Z)-5-decenyl acetate, (Z)-7-dodecenyl acetate, and (Z)-9-tetradecenyl acetate in proportions similar to those found in gland extracts from virgin females (0.6: 1:5:2.5). Lures containing 3–30 g (Z)-5-decenyl acetate proved to be maximally attractive, with approximately 60% of the males completing all behavioral steps from activation to copulation efforts. A 300-g dosage caused significant arrestment of upwind flight. Peak response to synthetics, however, was significantly lower than to female glands. Omitting decyl acetate from the blend did not affect the activity, while omission of any of the three monounsaturated acetates caused a dramatic decrease in response. In the field maximum trap catches were achieved with 1- to 30-g lures. The subtractive assay carried out in the field confirmed the neutrality of decyl acetate and the importance of the three monoenes. Adding 1% of (Z)-8-dodecenyl acetate (earlier reported as an inhibitor) to the four-component mixture decreased the trap catch to about 50%, and increasing the amount of (Z)-8-dodecenyl acetate to 27% decreased the activity further to about 10%. (Z)-8-Dodecenyl acetate also decreased the number of successful flights in the flight tunnel.Schiff., Lepidoptera: Noctuidae.This study was made within the Swedish project Odour Signals for Control of Pest Insects.     

Isolation and tentative identification of lineatin,a pheromone from the frass ofTrypodendron lineatum (Coleoptera: Scolytidae)

An attractant compound was isolated from frass produced byTrypodendron lineatum female beetles boring in Douglas fir. The proposed structure is one of two isomeric tricyclic acetals, to which the trivial name lineatin is assigned.Research supported by: National Science Foundation (U.S.A.) Grant No. PCM 74-13643, National Research Council (Canada) Operating Grant No. A3881, Canada Department of Fisheries and Forestry Extramural Research Grant No. F-58, Canadian Forestry Service Science Subvention Grant, and the Council of Forest Industries of British Columbia.     

Pheromone biosynthetic pathways: Conversion of ipsdienone to (−)-ipsdienol,a mechanism for enantioselective reduction in the male bark beetle,Ips paraconfusus

The enantiomeric composition of the pheromone components (+)-ipsdienoI, e.e. 87.6%, and (–)-ipsenol, e.e. 93.8%, produced by the male bark beetleIps paraconfusus (Scolytidae) under natural conditions was determined by HPLC separation of their diastereomeric ester derivatives. Males confined in an atmosphere of ipsdienone produced (–)-ipsdienol, e.e. 28%, and (–)-ipsenol, e.e. 86%, indicating an enantiomeric selectivity in the conversion of the ketone to the alcohols. These findings demonstrate an enantioselective conversion mechanism in the biosynthetic pathway to the pheromones from myrcene, a host-plant terpene.     

Identification of a Triterpenoid Saponin from a Crucifer, Barbarea vulgaris, as a Feeding Deterrent to the Diamondback Moth, Plutella xylostella

Larvae of the diamondback moth, Plutella xylostella, a crucifer specialist, refuse to feed on a crucifer, Barbarea vulgaris, because of the presence of a feeding deterrent, which is extractable with chloroform. We isolated a feeding deterrent from B. vulgaris leaves, by successive fractionations with silica-gel, ODS, i.e., C₁₈ reversed phase, and Sephadex LH-20 column chromatographies, and ODS-HPLC, guided by a bioassay for feeding deterrent activity. The structure of the compound was determined to be a monodesmosidic triterpenoid saponin, 3-O-[O--D-glucopyranosyl-(14)--D-glucopyranosyl]-hederagenin, based on FAB-MS, ¹H- and ¹³C-NMR spectra, and hydrolysis experiments. When the compound was applied to cabbage leaf disks at greater than 0.18 g/mm², consumption of the disks by third instars was less than 11% of control disks treated with the solvent alone. Furthermore, all first instars died on the disks treated with the same concentrations. Because the concentration of the compound in the fresh leaves of B. vulgaris was comparable to the effective dose in the cabbage leaf disk tested, we conclude that the unacceptability of B. vulgaris to P. xylostella larvae is primarily due to this saponin.     

Female sex pheromone of the melonworm,Diaphania hyalinata (Lepidoptera: Pyralidae), and analysis of male responses to pheromone in a flight tunnel

Ten C₁₆ chain-length compounds were identified from heptane extracts of ovipositors of female melonworm,Diaphania hyalinata (L.). The major constituents of the extracts were (E)-11-hexadecenal and (E,E)-10,12-hexadecadienal [(E,E)-10,12–16:Ald] and the alcohols and acetates of these olefins were found in trace amounts (<2%). Extracts also contained traces of (E,Z)- and (Z,Z)-10,12-16:Ald, hexadecanal, and 1-hexadecanol. Analysis of the behavioral responses of males to synthetic mixtures of these compounds and responses to ovipositor extracts in a flight tunnel showed that a synthetic mixture of the 10 compounds elicited a behavioral repertoire from males that was indistinguishable from that elicited by ovipositor extract. Flight tunnel studies also indicated that six of the 10 compounds probably represent the essential components of the female's sex pheromone.Supported in part by USDA Cooperative Agreement No. 58-32U4-1-299. Scientific Article No. A-4073, Contribution No. 7058 of the Maryland Agricultural Experiment Station. Mention of a commercial product in this paper does not constitute a recommendation by the U.S. Department of Agriculture.     

Studies related to the structure and reactivity of coals : 18. The chemical structures of products from reactions of a suite of higher rank Australian coals

The structure of the oils, asphaltenes and residues obtained by the thermal reactions of a suite of Australian higher rank coals under hydrogen or nitrogen have been studied by chemical and spectroscopic methods. The host-guest model that has been used to describe the structure of Australian brown coals cannot be applied directly to the higher rank coals. Evidence is provided that suggests that a modified version of the model may be of use in describing the structure of some subbituminous coals. The methodology has proved to be useful in the understanding of structural features of coals which are often not rank dependent, e.g. Callide coal (ABL2), a subbituminous coal, has been shown to have characteristics of both very high and also low rank coals.     

从兴趣出发,以OBE理念打造充满魅力的微生物学课堂教学

针对微生物学课程,我们基于OBE理念,大力推行启发式和联想式教学,加强师生互动,营造轻松活泼、充满魅力的课堂氛围,引导学生从“要我学”的被动学习向“我要学、我爱学”的主动学习和自主学习转变。教学改革的实施大大提高了学生对微生物学课程和生物专业的兴趣,教学效果得到了专家和学生的高度认可。     

Bioaccumulation of lanthanum,uranium and thorium,and use of a model system to develop a method for the biologically-mediated removal of plutonium from solution

Removal of La³⁺, UO₂²⁺ and Th⁴⁺ from aqueous solution by a Citrobacter sp. was dependent on phosphatase-mediated phosphate release and the residence time in a plug-flow reactor (PFR) containing polyacrylamide gel-immobilized cells. In a stirred tank reactor (STR) lanthanum phosphate accumulated on the biomass rapidly, in preference to uranium or thorium phosphates. Thorium removal was not affected by the presence of uranium but was promoted in the presence of lanthanum. Analysis of the accumulated polycrystalline material by X-ray powder diffraction (XRD) analysis and proton induced X-ray emission (PIXE) suggested the formation of a mixed crystal of lanthanum and thorium phosphate. La³⁺, UO₂²⁺ and Th⁴⁺ are analogues of the corresponding species of Pu³⁺, PuO₂²⁺ and Pu⁴⁺. The La/U/Th model system was used to identify some potential problems in the bioremediation of wastes containing plutonium and to develop a method for the biologically-mediated removal of plutonium from solution, in a test solution of ²³⁹Pu ‘spiked’ with a ²⁴¹Pu tracer. © 1998 SCI.     

Switch from single pot to multiphase high shear wet granulation process, influence of the volume of granulation liquid in a pilot scale study

Among the high shear wet granulation equipments used in the pharmaceutical industry, two configurations are current: single pot process for which blending, granulation and drying are performed in the same apparatus and multiphase process that usually associates a mixer granulator and a fluid bed dryer. Pharmaceutical formulations are often developed with regard to a specific industrial apparatus, but production imperatives may require a switch to another type of equipment. In this work, granulation process switch was investigated at pilot scale on a first intention excipient formulation and with two drug substances chosen as model drugs on the basis of their different water solubility. Each one was tested at two concentrations, 1 and 25%. The volume of granulation liquid was first fixed at the same level whatever the granulation equipment and the formulation. In the second part of the study, the effect of the volume of granulation was highlighted. Regardless of the formulation tested, single pot granules, compared to multiphase one, had improved flow properties, compressibility and tablet cohesion but higher sticking phenomenon was observed when tableting. In the second part, the effect of an adjustment of the volume of granulation liquid for horizontal transposition between high shear granulation processes was discussed.     

Deposition of C60, C70 and C84 fullerene molecules, in benzene via a change of the fluid state, from a gas–liquid two phase region to the critical point

We dissolve C₆₀, C₇₀ or C₈₄ molecules in benzene and change the fluid state from a gas–liquid two-phase region (25.0 °C) to the critical point (289.0 °C) and from the critical point to the original state (25.0 °C) along the gas–liquid coexistence curve. We find that particle-like and whisker-like nano/micro clusters, which are composed of C₆₀ molecules, deposit on the surface of a silicon substrate placed vertically in C₆₀/benzene solution during the temperature change, whereas no appreciable clusters are detected on the silicon substrate in either C₇₀/benzene or C₈₄/benzene solutions. The clusters, in which fcc lattice structures are formed by C₆₀ molecules, remain stable in the solution. The present result suggests that C₆₀ molecules can be separated and extracted from a mixture of C₆₀, C₇₀ and C₈₄ molecules dissolved in benzene.     

Supported photocatalysis as a pre‐treatment prior to biological degradation for the removal of some dyes from aqueous solutions; Acid Red 183, Biebrich Scarlet,Methyl Red Sodium Salt,Orange II

BACKGROUND: The aim of this study was to assess the feasibility of coupling photocatalysis and a biological treatment for the removal of azo dyes from aqueous effluents. Biological processes do not always appear relevant for dyes removal, owing to the low or total absence of biodegradability of this class of pollutants. RESULTS: During photocatalysis pre‐treatment, a decrease in the chemical oxygen demand (COD) indicated oxidation of the target compound and thus a change in the chemical structure; better biodegradability or less toxicity could then be expected. However, the concomitant decrease in dissolved organic carbon (DOC), characteristic of a high mineralization yield, led to nearly constant COD:DOC ratios, which was unfavorable for an increase in biodegradability. It was confirmed by the low values found for the ratios biological oxygen demand (BOD₅) to COD, which remained in the range 0.09–0.19, namely below 0.4 after photocatalytic reaction. Moreover, toxicity increased or remained at a high level after irradiation of the azo dyes for 3 h, and decreased only for Orange II, from toxic (EC₅₀ = 53%) to moderately toxic (EC₅₀ = 76%). CONCLUSION: An integrated process involving photocataysis and biological treatment to treat azo dyes appeared unsuitable under the conditions tested and may only be considered for Orange II among the four dyes tested. Copyright © 2010 Society of Chemical Industry     

Interception of the enzymatic conversion of farnesyl diphosphate to 5-epi-aristolochene by using a fluoro substrate analogue: 1-fluorogermacrene A from (2E,6Z)-6-fluorofarnesyl diphosphate

Tobacco 5-epi-aristolochene synthase (TEAS) catalyzes the Mg(II)-dependent cyclizations and rearrangements of (E,E)-farnesyl diphosphate (PP) to the bicyclic sesquiterpene hydrocarbon via a tightly bound (+)-germacrene A as a deprotonated intermediate. With the native enzyme, only a few percent of the putative germacrene A intermediate is released from the active site during the catalytic cycle. 6-Fluorofarnesyl PP was designed and synthesized with the aim of arresting the cyclization-rearrangement mechanism en route to 5-epi-aristolochene. Indeed, incubation of (2E,6Z)-6-fluorofarnesyl PP with recombinant TEAS afforded (-)-1-fluorogermacrene A as the sole product in 58% yield. Steady-state kinetic experiments with farnesyl PP and the 6-fluoro analogue showed that the overall catalytic efficiencies (k(cat)/K(m)) are essentially the same for both substrates. 1-Fluorogermacrene A was characterized by chromatographic properties (TLC, GC), MS, optical rotation, UV, IR and (1)H NMR data, and by heat-induced Cope rearrangement to (+)-1-fluoro-beta-elemene. (1)H NMR spectra at room temperature revealed that this (E,E)-configured fluorocyclodecadiene exists in solution as a 7:3 mixture of UU and UD conformers. 1-Fluorogermacrene A underwent trifluoroacetic acid-catalyzed cyclization to give three 1alpha-fluoroselinene isomers at a rate estimated to be about 1000 times slower than that of the similar cyclization of (+)-germacrene A to the parent selinenes.     

Analysis and Manipulation of the Structure of Odor Plumes from a Piezo-Electric Release System and Measurements of Upwind Flight of Male Almond Moths, <Emphasis Type="Italic">Cadra cautella</Emphasis>, to Pheromone Plumes

We investigated the plume structure of a piezo-electric sprayer system, set up to release ethanol in a wind tunnel, using a fast response mini-photoionizaton detector. We recorded the plume structure of four different piezo-sprayer configurations: the sprayer alone; with a 1.6-mm steel mesh shield; with a 3.2-mm steel mesh shield; and with a 5 cm circular upwind baffle. We measured a 12 × 12-mm core at the center of the plume, and both a horizontal and vertical cross-section of the plume, all at 100-, 200-, and 400-mm downwind of the odor source. Significant differences in plume structure were found among all configurations in terms of conditional relative mean concentration, intermittency, ratio of peak concentration to conditional mean concentration, and cross-sectional area of the plume. We then measured the flight responses of the almond moth, Cadra cautella, to odor plumes generated with the sprayer alone, and with the upwind baffle piezo-sprayer configuration, releasing a 13:1 ratio of (9Z,12E)-tetradecadienyl acetate and (Z)-9-tetradecenyl acetate diluted in ethanol at release rates of 1, 10, 100, and 1,000 pg/min. For each configuration, differences in pheromone release rate resulted in significant differences in the proportions of moths performing oriented flight and landing behaviors. Additionally, there were apparent differences in the moths’ behaviors between the two sprayer configurations, although this requires confirmation with further experiments. This study provides evidence that both pheromone concentration and plume structure affect moth orientation behavior and demonstrates that care is needed when setting up experiments that use a piezo-electric release system to ensure the optimal conditions for behavioral observations.