Molecular recognition has become a very important field of research in chemistry during the last decades. This<br />chemical phenomenon is responsible for all processes occurring in biology and asymmetric synthesis is based<br />upon the capability of molecules or substrates to recognise each other in a selective manner. In this thesis, the<br />design, preparation and evaluation of a series of new synthetic receptors has been described. The importance of<br />regioselectivity and stereoselectivity in molecular recognition has also been underlined with two different<br />biological examples.<br />The capability of host molecules, derived from (+)-tartaric acid, to accommodate various guests in a selective<br />manner was demonstrated using 1H-NMR spectroscopy (paper I). These host molecules, known as TADDOLs,<br />enantioselectively recognised the valuable chiral alcohols glycidol and menthol. Macromolecular receptors, i.e.<br />molecularly imprinted polymers (MIPs), were also prepared in order to catalyse the aldol reaction between either<br />(R)- or (S)-camphor and benzaldehyde (paper II). With the help of analytical methods, it was demonstrated that<br />the MIPs interacted in a selective manner with the enantiomers of camphor. Moreover, these MIPs enhanced<br />significantly the rate of the aldol condensation mentioned above.<br />Regarding biological systems, various regioisomeric analogues of benzoic acid have been tested as antifeedants<br />against the pine weevil Hylobius abietis (paper III and IV). The regioisomers studied displayed very different<br />antifeedant activities. The significance of stereoisomerism on pheromone function has been shown in the<br />preparation of lures for the control of the insect pest Argyrotaenia sphaleropa (paper V). It was demonstrated that<br />male leafrollers could be caught by a lure containing components of the female sex pheromone gland.

<p>The Autumn gum moth, <em>Mnesampela privata </em>(Lepidoptera: Geometridae) is an endemic Australian moth whose larvae feed upon species of <em>Eucalyptus.</em> The moths favorite host plants are <em>E. globulus </em>and<em> E. nitens</em> which are the most important species used in commercial plantations of the Australian pulpwood industry. The autumn gum moth has become one of the most significant outbreak insects of eucalyptus plantations throughout Australia. As a consequence great financial losses to the forest industry occur. Today insecticides such as pyrethroids are used for control of eucalyptus defoliators as <em>M. privata</em>.</p><p>The carrot psyllid, <em>Trioza apicalis </em>(Homoptera: Psylloidea), is one of the major pests of carrot (<em>Daucus carota</em>) in northern Europe. The psyllid causes curling of the carrot leafs and reduction of plant growth. Today the carrot crops are protected with the pyrethroid insecticide cypermethrin, which is toxic to aquatic organisms and is, from 2010, prohibited for use in Sweden by the Swedish Chemicals Inspectorate.</p><p>An alternative to insecticides is to protect the seedlings with semiochemicals, a chemical substance or mixture of them that carries a message. This thesis describes the identification and the syntheses of semiochemicals from the above mentioned insect species.</p><p>From analysis of abdominal tip extracts of <em>M. privata</em> females from Tasmania a blend of (3<em>Z</em>,6<em>Z</em>,9<em>Z</em>)-3,6,9-nonadecatriene and (3<em>Z,6Z,9Z</em>)-3,6,9-heneicosatriene was identified as the sex pheromone of this species. The identification of the C₁₉- and C₂₁-trienes was confirmed by synthesis.</p><p>In the analysis of carrot leaf extracts we found a compound, α-<em>cis</em>-bergamotene, that induces antennal response in the carrot psyllid. This is just the beginning of the studies of trying to manipulate this psyllid with semiochemicals instead of insecticides.</p>

Sexual selection can influence the mating system of an organism through multiple mechanisms. These mechanisms result in variation in reproductive success among individuals, and include scramble competition, endurance rivalries, contests, mate choice and cryptic choice, and sperm competition. Understanding the mating system of a species requires the identification of which processes are occurring, and to what degree. In this thesis, I explored the influence of mate choice mechanisms on the mating system of the terrestrial red-legged salamander, Plethodon shermani. I also documented the potential for post-copulatory processes (such as sperm competition and cryptic choice) to influence mating system dynamics. The evolution of mate choice requires (among other factors) variation in the reproductive value of potential mates. This variation is made apparent to choosy individuals through cues. Most animals use multiples cues incorporating many modalities to assess the reproductive quality of potential mates. In Chapter 2, I tested the contribution of two cues (chemical and visual) to mate choice by female P. shermani. I found that a male visual cue ("foot-dancing") increased courtship success. In contrast, delivery of non-volatile pheromones during courtship did not influence courtship success in the laboratory setting, but did affect the duration of one of the courtship stages. In Chapter 4, I identified a tactile cue that was significantly correlated with male reproductive success. Thus, P. shermani females could use at least three modalities to assess the reproductive quality of potential mates. Mate choice can also evolve in males. In Chapter 3, I tested this possibility in P. shermani. I found that males vary the reproductive effort they invest in a particular courtship based on the reproductive value of their partner, indicating male mate choice is occurring. A male invested most when paired with a female with large, well developed ova, and invested less with females that were non-gravid or had small ova. In addition to documenting male mate choice, I showed that the male visual display ("foot-dancing") that affected female mate choice was correlated with male condition, implying foot-dancing may be an honest indicator of male quality. Finally, in Chapter 5, I explored the potential for post-copulatory processes to influence the P. shermani mating system. The opportunity for sperm from multiple males to overlap in the female reproductive tract (i.e., the opportunity for females to mate multiply) is necessary for post-copulatory processes such as cryptic choice and sperm competition. The capacity for long-term sperm storage by females can increase the likelihood that this overlap in sperm from multiple males will occur. I found that females can store viable sperm for at least 9 months and in some cases beyond oviposition. In addition, I documented one female with sperm in her sperm storage organ from a mating that occurred 17 months earlier. Such lengthy sperm storage allows the possibility of sperm from one breeding season to interact with sperm from a subsequent season. Thus, the potential for post-copulatory sexual selection within this salamander system is high. / Graduation date: 2012

The Autumn gum moth, Mnesampela privata (Lepidoptera: Geometridae) is an endemic Australian moth whose larvae feed upon species of Eucalyptus. The moths favorite host plants are E. globulus and E. nitens which are the most important species used in commercial plantations of the Australian pulpwood industry. The autumn gum moth has become one of the most significant outbreak insects of eucalyptus plantations throughout Australia. As a consequence great financial losses to the forest industry occur. Today insecticides such as pyrethroids are used for control of eucalyptus defoliators as M. privata. The carrot psyllid, Trioza apicalis (Homoptera: Psylloidea), is one of the major pests of carrot (Daucus carota) in northern Europe. The psyllid causes curling of the carrot leafs and reduction of plant growth. Today the carrot crops are protected with the pyrethroid insecticide cypermethrin, which is toxic to aquatic organisms and is, from 2010, prohibited for use in Sweden by the Swedish Chemicals Inspectorate. An alternative to insecticides is to protect the seedlings with semiochemicals, a chemical substance or mixture of them that carries a message. This thesis describes the identification and the syntheses of semiochemicals from the above mentioned insect species. From analysis of abdominal tip extracts of M. privata females from Tasmania a blend of (3Z,6Z,9Z)-3,6,9-nonadecatriene and (3Z,6Z,9Z)-3,6,9-heneicosatriene was identified as the sex pheromone of this species. The identification of the C19- and C21-trienes was confirmed by synthesis. In the analysis of carrot leaf extracts we found a compound, α-cis-bergamotene, that induces antennal response in the carrot psyllid. This is just the beginning of the studies of trying to manipulate this psyllid with semiochemicals instead of insecticides.

The effects of cypermethrin, copper and glyphosate on the endocrine system and subsequent response to female pheromones were investigated in mature male brown trout (Salmo trutta) parr.  Responses measured were the amount of strippable milt, blood plasma levels of both an androgen (11-ketotestosterone (11-KT)) and a progestin (17α,20β-dihydroxy-4-pregnen-3-one (17,20b-P)), and behavioural changes. This was done in a two phased investigation where parr were exposed to one of the following via ambient water: 1) 0.1 or 1.0 μg L-1 cypermethrin, 2) 10 or 100 μg L-1 copper (Cu2+), or 3) 150 μg L-1 glyphosate for a 96 hour period.  Phase one was a priming experiment exposing parr to a treatment followed by priming with PGF2α or ovarian fluid (OVF). Atlantic salmon (Salmo salar) parr were, also exposed to glyphosate during phase I. The second phase was centered on behavioural observations.  Exposed parr were placed in a 35,000 L stream aquarium together with two ovulated females and four anadromous males. After the experiments a blood sample was taken, milt volumes measured and testes weighed.  The plasma was analyzed for 11-KT and 17,20b-P concentrations using radioimmunoassay (RIA). Results from phase I-priming: 1.0 μg L-1 cypermethrin exposure lowered 17,20b-P and 11-KT; Copper exposure lowered milt volumes; glyphosate exposure lowered 11-KT in salmon and raised 17,20b-P in trout.  Results from phase II-behaviour: 1.0 μg L-1 cypermethrin exposure lowered 11-KT, milt and spawning behaviour; copper exposure lowered spawning behaviour and raised 11-KT; Glyphosate exposure lowered 11KT; continuous cypermethrin exposure raised 17,20b-P, 11-KT and gave a tendency towards increased aggression. It is concluded that low concentration exposure to the compounds examined can induce negative effects on male salmonid endocrine systems, either through a disruption in the olfactory system or through a direct effect. / Extern doktorand från Södertörns högskola

The past few decades have witnessed a revolution in recombinant DNA and nucleic acid sequencing technologies. Recently however, technologies capable of massively high-throughout, genome-wide data collection, combined with computational and statistical tools for data mining, integration and modeling have enabled the construction of predictive networks that capture cellular regulatory states, paving the way for ‘Systems biology’. Consequently, protein interactions can be captured in the context of a cellular interaction network and emergent ‘system’ properties arrived at, that may not have been possible by conventional biology. The ability to generate data from multiple, non-redundant experimental sources is one of the important facets to systems biology. Towards this end, we have established a novel platform called ‘spotted cell microarrays’ for conducting image-based genetic screens. We have subsequently used spotted cell microarrays for studying multidimensional phenotypes in yeast under different regulatory states. In particular, we studied the response to mating pheromone using a cell microarray comprised of the yeast non-essential deletion library and analyzed morphology changes to identify novel genes that were involved in mating. An important aspect of the mating response pathway is large-scale spatiotemporal changes to the proteome, an aspect of proteomics, still largely obscure. In our next study, we used an imaging screen and a computational approach to predict and validate the complement of proteins that polarize and change localization towards the mating projection tip. By adopting such hybrid approaches, we have been able to, not only study proteins involved in specific pathways, but also their behavior in a systemic context, leading to a broader comprehension of cell function. Lastly, we have performed a novel metabolic starvation-based screen using the GFP-tagged collection to study proteome dynamics in response to nutrient limitation and are currently in the process of rationalizing our observations through follow-up experiments. We believe this study to have implications in evolutionarily conserved cellular mechanisms such as protein turnover, quiescence and aging. Our technique has therefore been applied towards addressing several interesting aspects of yeast cellular physiology and behavior and is now being extended to mammalian cells. / text

The freshwater prawn, Macrobrachium rosenbergii (de Man, 1879) is an important aquaculture species but one that has the disadvantage of heterogeneous individual growth (HIG) according to different morphotypes. Chemical cues, especially, pheromones, are one of the most important communication types between individual prawns, along with visual and tactile methods. Testing pheromones, whilst restricting other cues, may therefore lead to a better understanding of the influence of these communicatory compounds on the prawn reproductive process. The three principle objectives of this study were therefore: 1) To examine the effect of moult stage and morphotype on pheromone-induced sexual behaviour 2) To examine the role of pheromone / urine concentrations on sexual attraction behaviour 3) To describe the functional morphology of the antennal gland and examine its possible role in pheromone production and release. Identical bioassay tanks were designed and constructed to study the reproductive behaviour of prawns. Experiments were set up to examine responses to pheromone release by live prawns over 30 minutes and behavioural response observations were made with the aid of a Closed-Circuit Videotape System (CCVS). Results were statistically analysed using a repeated measures general linear model (GLM). Three trials were designed to test the effect of moult stage of both males and females and male morphotypes on sexual attraction behavioural responses. Twelve prawns were used for each trial and each prawn was used five times (1 no-pheromone control and 4 for experimental tests). The first trial studied the effect of female moult stages (pre-, inter and newly-moulted) on sexual attraction behaviour of blue claw (BC) male. Results of this trial showed that newly-moulted females spent significantly (p<0.05) less time approaching the BC male than the pre- and inter-moult females. The second trial studied the effect of male moult stage (pre-, inter and newly-moulted) on sexual attraction to receptive females. Results showed that the time taken by the inter-moult males was (p<0.05) less than the pre- and newly-moulted males in approaching the newly-moulted female. The third trial tested the effect of male morphotypes (small male, SM, orange claw, OC and dominant blue claw, BC) on sexual attraction behaviour towards newly-moulted females. Results showed that the BC male was significantly more attractive (p<0.05) than other morphotypes to newly-moulted females and that the OC male was the least attractive. The role of moulting stage for both male and female prawns on reproductive response behaviour was investigated. Because BC males responded significantly faster towards newly-moulted female more than to either pre-or inter-moult females, results of the first trial suggest that BC males are able to use different chemical cues to gather information about a conspecific’s gender and can differentiate female’s moult stages. Since BC males responded significantly faster towards newly-moulted females more than to either pre-or inter-moult females, this suggests that females at this particular stage released a distinct sexual pheromone or concentration of pheromone that differed from those pheromones released by both pre- and inter-moult females. In contrast, newly-moulted females prefer the inter-moult BC males which indicate that females have an ability to distinguish the moult status of BC males. Furthermore, it indicates that pheromone characteristics change with the moult status of BC males. Also, newly-moulted females are most likely to be avoiding the potential costs of mate guarding with soft shell BC males. Results obtained from the third trial suggested that a newly-moulted female can discriminate male morphotypes (SM, OC and BC) from their pheromone cues. This indicates that male morphotypes release pheromones which differ from each other in some way. Newly-moulted females responded positively to both SM and BC males with different levels of attraction with the greatest attraction to BC males to BC males suggesting that pheromone released from the BC male may carry information relating to dominance status. Urine is believed to be one of the main carriers of pheromone and is usually released from the antennal gland. Different urine concentrations (0.1, 1.0, 2.0, 3.0, 5.0 and 10µl l-1) of collected urine from BC males were used to test the sexual attraction behaviour of receptive newly-moulted females. Also, the attractant capability of fresh urine following exposure to different temperature regimes (cooled at 4ºC, frozen at -70ºC and heated at 70ºC) was tested. Since newly-moulted female M. rosenbergii were attracted to BC male urine, this indicates the existence of sex pheromone in the fresh urine. Also, it was found that the sexual response of females to fresh urine of BC males was directly proportional to urine concentration with faster responses observed with increasing urine concentrations. At the three fresh urine concentrations 0.1 µl l-1, 1.0 µl l-1 and 2.0 µl l-1, statistical analysis indicated no significant difference (p>0.05) between these three concentrations while a significant (P<0.05) response was to concentrations more than 3.0 µl l-1. This may indicate that these three concentrations were not sufficient to elicit attraction behaviour in newly-moulted females. A concentration of 3.0 µl l-1 of fresh urine is suggested to be a sufficient concentration to elicit a significant sexual attraction under laboratory conditions. Response of newly-moulted female prawns to the various temperature treatments tested declined in response to nominally increasingly degradative treatments. Also, statistical analysis showed that temperature treatment and concentration added both had a significant effect on the response of females. The greatest degradation of urine attractiveness was found with the 70ºC heat treatment. It can be concluded that the pheromone components of prawn urine are friable when exposed to high temperatures. Using light and transmission electron microscopes, ultrastructural observation of the antennal gland (AG) of M. rosenbergii suggests that it has four distinct regions, the coelomosac, the nephridial tubules, the labyrinth and the bladder. Morphological and functional descriptions of each of these regions were compared with those of other aquatic Crustacea.

Pacific lamprey (Entosphenus tridentatus) are experiencing population declines throughout their range. Xenobiotics could be an important risk factor for lamprey populations. Our goal was to establish if common herbicides, as used in forest management, could affect reproductive fitness. We determined that atrazine was a likely compound of greatest concern to lamprey populations. Using an odorant response behavioral assay we were able to demonstrate that environmentally relevant concentrations of atrazine caused a depressed response to adult lamprey holding tank effluent, likely pheromones. Atrazine also depressed their activity level; the number of times they crossed into the effluent arm after being treated with atrazine was significantly lower than controls. In addition, activity level post exposure to atrazine differed between adult life history stages, something which was not significantly different during control trials. Using an odorant detection assay, based on evaluating ventilation rate, we were able to show that environmentally relevant concentrations of atrazine caused a significant increase in ventilatory response to a repulsive odorant, a conspecific necromone. Through the detection study we also showed that lamprey,exposed to atrazine, had a slight increase in ventilatory response to odor from adult lamprey. If we are concerned about the decline in Pacific lamprey populations, then we should logically be concerned with their exposure to atrazine in the environment. / Graduation date: 2012

Made available in DSpace on 2015-03-26T13:30:40Z (GMT). No. of bitstreams: 1 texto completo.pdf: 4221966 bytes, checksum: 09f2c3231a81c4d2d8a60ef298168caa (MD5) Previous issue date: 2005-02-25 / Conselho Nacional de Desenvolvimento Científico e Tecnológico / Adults of Triatoma brasiliensis (Reduviidae) show two pairs of exocrine glands: the metasternal and the Brindley glands. The first one is located in the ventral part of the metathorax, while Brindleys glands are found in the lateral part of the superior metathorax. In both glands, the secretory cells correspond to type III, according to the classification of Noirot & Quennedey (1991). Metasternal glands present tricoid sensilla next to their external opening. In the cytoplasm of the secretory cells of these glands granules strongly stained were usually found, suggesting that these glands produce different substances than those of Brindleys glands. The substances found in the secretion of the Brindleys glands of T.brasiliensis were identified for the first time, as a mixture of five fatty acids: acetic; propanoic; isobutyric; butyric and 2-methyl-butyric acids. Isobutiric acid was found to be the most abundant compound and no qualitative or quantitative differences were observed in relation to gland contents for both sexes. Our results demonstrated that the metasternal glands of T. brasiliensis are also involved in the synthesis of volatile substances. These glands produce basically ketones and alcohols, 3-pentanone was the major constituent of the mixture found in these glands for both sexes. After their mechanical disturbance, adults of T. brasiliensis emitted a mixture of 10 different compounds consisting of ketones, alchools and fatty acids, including substances from both metasternal and Brindleys glands,. Finally, we analyzed the sexual behavior of T. brasiliensis: the copula last, in average, 6 ± 1 min and the number of male attempts to copulate varied according to the insect age. Females of T. brasiliensis showed three types of rejection in response to male attempt to copulate: flattening, abdominal movements and evasion. The analysis of the female behavior revealed that the number of rejections in response to male attempts stayed the same during the pre-feeding period. After feeding a gradual decrease in the number of rejections was observed, diminishing until reaching zero rejection at approximately the day 8 post-feeding. We suggest a behavioral mechanism in the female, after feeding, that may trigger the acceptance of the male attempt to copulate in T. brasiliensis. The relevance of the results for understanding the behavioral repertoire of the specie, particularly in the sexual context, is discussed. / As glândulas exócrinas de T. brasiliensis foram analisadas do ponto de vista morfológico e histológico no presente trabalho. Insetos adultos de Triatoma brasiliensis apresentam dois pares de glândulas exócrinas produtoras de voláteis: as glândulas metasternais, localizadas no metatoráx ventral e as glândulas de Brindley, situadas no metatórax lateral superior. Nossos resultados descrevem pela primeira vez detalhes histológicos e morfológicos destas glândulas em Triatoma brasiliensis. Em ambas glândulas metasternais e de Brindley as células secretoras seriam do tipo III, segundo classificação proposta por Noirot & Quennedey (1991). A glândula metasternal apresenta sensilas tricóides na região próxima à sua abertura externa. No citoplasma de suas células secretoras foram encontrados grânulos fortemente corados, sugerindo que esta glândula produz substâncias diferentes daquelas produzidas pelas glândulas de Brindley, que apresentam vacúolos não corados, indicando ausência de grânulos. Uma mistura de cinco ácidos graxos foi identificada: ácido acético; ácido propanóico; ácido isobutírico; ácido butírico e ácido 2-metil-butírico. O componente mais abundante foi o ácido isobutírico e nenhuma diferença qualitativa ou quantitativa entre os sexos foi observada em relação ao seu conteúdo. Nossos resultados também demonstram que as glândulas metasternais de T. brasiliensis estão envolvidas na síntese de substâncias voláteis. Estas glândulas produzem basicamente álcoois e cetonas, sendo que o principal constituinte encontrado nas glândulas de ambos os sexos foi a 3-pentanona. Durante a perturbação mecânica, adultos de T. brasiliensis liberam uma mistura de 10 componentes diferentes, constituída por ácidos, cetonas e álcoois, incluindo substâncias provenientes das glândulas metasternais e de Brindley. Paralelamente, foi analisando o comportamento sexual de T. brasiliensis. Observamos que a cópula tem uma duração média de 6 +ou- 7 min. Fêmeas desta espécie apresentaram somente três tipos de comportamento de rejeição frente ás tentativas de cópula dos machos como o achatamento corporal, os movimentos abdominais e a evasão. O número de tentativas de cópula realizadas pelos machos variou em função da idade. A análise do comportamento das fêmeas revelou que o número de rejeições em resposta às tentativas dos machos se manteve igual durante o período pré-alimentação, não sendo afetado pela sua idade. No período pós-alimentação foi observada uma mudança gradual na porcentagem de tentativas de cópula rejeitadas pelas fêmeas que, diminuiu gradativamente até atingir 0 %. Sugere-se que algum mecanismo comportamental desencadeado na fêmea dias após alimentação esteja determinando a ausência de rejeições e conseqüente aceitação da cópula em T. brasiliensis. A importância destes resultados com relação ao comportamento destes insetos, e particularmente no contexto sexual, é discutida.

O objetivo da pnmeIra parte do presente projeto é a síntese total da Isocicutoxina (esquema 1) e de alguns feromânios sexuais de insetos (esquema 2), utilizando algumas das reações já estudadas sistematicamente no nosso laboratório. (Ver PDF) As etapas chaves para síntese da Isocicutoxina foram uma reação de acoplamento entre um diino e um telureto vinílico, seguida por urna reação de Wittig. As etapas chaves para a síntese dos feromânios sexuais foram urna reação de acoplamento cruzado de um telureto vinílico com um organocrupato, seguida por urna reação tipo SN2 com um reagente de Grignard. O objetivo da segunda parte do presente projeto é a revisão bibliográfica e a sistematização de dados de RMN de 1H, 13C e 125Te d.e compostos contendo átomos de telúrio. Os ligantes contendo telúrio tiveram seus valores de RMN sistematizados e seus valores de coeficientes de blindagem efetivos analisados (esquema 3). (Ver PDF) Através de um software especializado, um banco de dados foi criado e possibilitou a simulação de espectros de RMN de 13C e 125Te. / The main goal of the first part of this project is the total synthesis of isocicutoxin (scheme 1) and some insects\' sexual pheromones by means of some reactions already studied by our group (scheme 2). (See PDF) The key steps for the synthesis of isocicutoxin were a coupling reaction between a diyne and a vinylic telluride, followed by a Wittig reaction. The key steps for the synthesis ofthe sexual pheromones were a cross coupling reaction between a vinylic telluride and an organocuprate, followed by a SN2 reaction with a Grignard reagent. The mam goal of the second part of the project lS the bibliographic reVleW and the systematization of 1H, 13C and 125Te data of tellurium compounds. The ligands containing tellurium had their NMR and their effective shielding values analyzed (Scheme 3). Using specialized software, a database was developed, which allowed the prediction of NMR 13C and 125Te spectra.