Doctor of Philosophy / Department of Biology / John M. Blair / Juniperus virginiana, an important woody plant invader in the U.S. Central Plains, has increased considerably in density and cover in large areas previously dominated by tallgrass prairie. Change in the phenology and nitrogen use efficiency of the dominant plant communities as J. virginiana replaces native prairies may lead to increased plant productivity and biomass accumulation, but may also alter the microclimate and litter quality that affect soil microbial communities responsible for key soil processes. I have focused my investigations on changes in key soil processes that could lead to differences in soil N availability, as well as changes in ecosystem C and N pools and fluxes as J. virginiana expands into native grasslands. Juniperus virginiana forest soils exhibit greater cumulative annual net N mineralization (11.52 ± 0.38 µg N g¯1 soil y¯1) compared to prairie soils (7.90 ± 0.26 µg N g¯1 soil y¯1) (F = 60.67, P = 0.016), yet slightly reduced potential soil C flux. Examination of internal soil N cycling revealed that both J. virginiana and prairie soils minimize potential soil N losses, by rapid microbial immobilization of inorganic N, and constraining nitrification via substrate limitation or environmental constraints. Leaf-level photosynthetic nitrogen use efficiency (NUE) was over a magnitude higher in the dominant grass, Andropogon gerardii, but high annual ecosystem-level NUE and greater soil N availability may contribute to the higher productivity and rapid accrual of C in newly established J. virginiana forests. Increased plant productivity and elimination of fire in J. virginiana forests have allowed at least 80,000 kg ha-1 increase in ecosystem C storage in about half a century. Soil organic C, an important long-term sink, has also increased significantly in J. virginiana forests, with approximately 34% replacement of C4 grass-derived soil C with new C from trees in the A-horizon. The observed high productivity of J. virginiana and increased N availability necessary to support continued plant biomass accumulation are possible because of substantial (~ 44%) increase in ecosystem N in measured pools, which is a likely a result of reduced volatilization of N from biomass burning, possible increased exogenous N inputs, and/or N translocation from deeper soil horizons. Reduced fire return intervals in prairie provide an opportunity for J. virginiana to establish and facilitate N accrual, which may allow this species to accelerate is own establishment through creating conditions of increased N availability and efficient utilization of N.

<p>Karakterizacija metanolnih ekstrakata i etarskih ulja iglica i &scaron;i&scaron;arki osam vrsta samoniklih kleka (rod Juniperus L.) obuhvatala je fitohemijski skrining i ispitivanje antioksidantne, antiinflamatorne i antimikrobne aktivnosti. Primenom LC-MS/MS i GC-MS tehnike detektovan i odreĎen je sadržaj fenolnih i terpenoidnih sekundarnih biomolekula. Antioksidantna aktivnost ekstrakata i etarskih ulja (sposobnost neutralizacije slobodnih radikala, redukcioni potencijal i inhibicija lipidne peroksidacije) ispitana je primenom spektrofotometrijskih metoda. U cilju odreĎivanja antiinflamatornog potencijala, primenjena je ex vivo metoda za praćenje aktivnosti trombocitne ciklooksigenaze-1 i 12-lipoksigenaze. Antimikrobna aktivnost etarskih ulja odreĎena je na &scaron;est odabranih bakterijskih sojeva. Ispitani ekstrakti i etarska ulja pokazali su značajnu biolo&scaron;ku aktivnost.</p> / <p> Characterization of methanol extracts and essential oils of eight Juniperus L. species included phytochemical screening and evaluation of antioxidant, anti-inflammatory and antimicrobial activity. The presence and content of phenolics and terpenoids was confirmed by LC-MS/MS and GC-MS technique. Antioxidant activity of extracts and essential oils (radical scavenger capacity, reduction potential and inhibition of lipid peroxidation) was examined by spectrophotometric methods. With the intention to evaluate anti-inflammatory activity, an ex vivo method was applied to measure activity of platelet cyclooxygenase-1 and 12-lipoxygenase. Antimicrobial activity was evaluated according to six bacterial strains. Examined extracts and essential oils showed noticeable biological activity.</p>

<p>U radu su prikazani eksperimentalni rezultati natkritične ekstrakcije etarskog ulja bobica<br />kleke (<em>Juniperus communis</em> L.) i ulja iz semena uljane tikve golice (<em>Cucurbita pepo</em> L.). Ispitan je uticaj pritiska, temperature, stepena usitnjenosti čestica i protoka natkritičnog<br />ugljendioksida na promenu prinosa ulja sa vremenom. U cilju poređenja, usitnjeno seme uljane tikve ekstrahovano je i u ekstraktoru većih dimenzija, NOVA-SWISS, High<br />pressure extraction plant, kao i heksanom i petroletrom u ekstraktoru tipa Sokslet.<br />Praćena je i promena kvaliteta ekstrakata sa vremenom: u etarskom ulju kleke, GC-FID i GC-MS metodama, određen je relativni sadržaj 50 terpenskih jedinjenja i sve komponente ulja su svrstane u 5 osnovnih grupa (monoterpene, seskviterpene, oksidovane monoterpene, oksidovane seskviterpene i ostale komponente). U tikvinom ulju ekstrahovanom natkritičnim ugljendioksidom određen je masnokiselinski sastav GC-MS analizom, sadržaj tokoferola HPLC analizom, sterola i skvalena GC-MS metodom. Određeni su uslovi koji favorizuju ekstrakciju ispitanih jedinjenja za obe sirovine. Dat je dateljan prikaz matematičkih modela koji se koriste za opisivanje natkritične ekstrakcije etarskih ulja i masnih ulja, počev&scaron;i od najop&scaron;tijeg modela koji uključuje diferencijalne bilanse mase za rastvorak u masi natkritičnog fluida, u fluidu unutar pora čestica usitnjenog matrijala i u čvrstoj fazi, koji se uvođenjem određenih pretpostavki pojednostavljuje i svodi na modele koji su izabrani da budu ispitani u okviru ovoga rada. Ispitani su modeli kreireni po analogiji sa hlađenjem vrele kugle u masi fluida, tj. modeli tipa jedne sfere i to: Model jedne sfere-1 (MJS-1), koji pored uticaja koeficijenta efektivne difuzije ulja u materijalu na brzinu prenosa mase uzima u obzir uticaj koeficijenta prenosa mase kroz film natkritičnog fluida oko čestice, pri čemu je njegova vrednost procenjena preko postojećih korelacija; MJS-1 (2 par), u kome je spolja&scaron;nji koeficijent prenosa mase uzet kao drugi prilagodljiv parametar modela; MJS-2, gde je koeficijent efektivne difuzije jedini prilagođeni parametar, Model karakterističnog vremena i pro&scaron;ireni model klipnog toka koji je predložila Sovov&aacute;. Za modelovanje natkritične ekstrakcije ulja semena tikve kori&scaron;ćen je i kombinovani model Honga i sar. Softverskim paketima Mathcad 2001 Professional i Solver dodatka unutar Microsoft Excel 2003, određeni su parametri ispitanih modela u cilju najboljeg slaganja modela sa ekperimentalnim podacima. Za obe ispitane sirovine, među ispitanim modelima, izabrani su modeli koji najbolje opisuju njihovu ekstrakciju natkritičnim ugljendioksidom. Pro&scaron;ireni model klipnog toka koji je predložila Sovov&aacute; pokazao se podjednako dobrim za modelovanje natkritične ekstrakcije obe sirovine i ne&scaron;to bolji od ostalih primenjenih modela.</p> / <p>This study provides results of supercritical carbon dioxide (SCCO₂) extraction of juniper berries (<em>Juniperus communis</em> L.) and pumpkin seeds (<em>Cucurbita pepo</em> L. convar.&nbsp;<em>citrullina</em>) in a laboratorysupercritical fluid extraction apparatus. The influence<br />of pressure, temperature, particle size and carbon dioxide flow on the extraction kinetics of pumpkin seed oil and juniper berry essential oil was studied. Ground pumpkin seeds were also extracted with supercritical carbon dioxide in NOVA-SWISS, High Pressure Extraction Plant, and with hexane and petroleum ether in a laboratory Soxhlet extractor. This work was also aimed to investigate the evolution of the composition of juniper fruit supercritical CO₂ extracts with time, at different extraction pressures and to emphasize the most favorable condition for the extraction of different terpene hydrocarbon groups, reporting the qualitative differences among extracts collected during successive extraction time periods. Juniper berry extracts were analyzed by capillary gas chromatography, using flame ionization (GC-FID) and mass spectrometric detection (GC-MS). More than 200 constituents were detected in the extracts and the contents of 50 compounds were reported in the work. Dependence of the percentage yields of monoterpene, sesquiterpene, oxygenated monoterpene and oxygenated sesquiterpene hydrocarbon groups on extraction time was investigated and conditions that favored the yielding of each terpene groups were emphasized. GC-MS analysis of FAME, prepared by transesterification of pumpkin seed oil with KOH in methanol, was performed. Fatty acid compositions of supercritical CO₂ pumpkin seed extract fractions collected in successive time intervals over the course of the extraction were determined. The same fractions were analyzed by high pressure liquid chromatography (HPLC), using diode-array detector (DAD) in order to determine a- and g-tocopherol contents. Sterol and squalene contents were determined by GC-MS analysis, as well. Conditions that favored the yielding of tocopherols, squalene and sterols were emphasized. A general mass transfer model and its simlifications were analysed. Extraction curves were evaluated by &ldquo;hot sphere&rdquo; mathematical models SSM-1 (Single Sphere Model 1 &ndash; in which the external mass transfer coefficient also influences the extraction profile and film mass transfer coefficients were estimated by the correlations), SSM-1 (2 par) (film mass transfer coefficient is used as the second adjustable parameter), SSM-2 (only effective diffusivity influence is considered), Characteristic time model and by the extended Lack&rsquo;s plug-flow model given by Sovov&aacute;. A combined model of Hong et al. was also fitted to the experimental data for pumpkin seed oil SCCO_2&nbsp;extractions. Relative merits of the models are demonstrated. Good agreement between the extended Lack&rsquo;s plug-flow model and the experimental measurements was obtained.</p>

The growth rings of eastern redcedar (Juniperus virginiana L.) often contain a high frequency of false intra-annual growth bands, which complicates the dendrochronology of this species. However, exactly dated false rings replicated among many trees can reflect major weather changes during the growing season. Sixty-one trees from two sites (Oklahoma and Kansas) were dated and used to compile replicated chronologies of false rings at both locations extending from AD 1700–2000. False-ring events during the modern instrumental era were compared with the daily weather data from nearby stations. Significant false-ring events occurred at both locations during years that experienced a dramatic late-growing season weather reversal, when an extended period of high temperatures and drought was followed by prolonged cool and wet conditions. Synoptic weather maps for these events indicate that all ten replicated false-ring events in the instrumental era occurred during the highly unseasonable penetration of a cold front into the region. However, none of the significant false-ring events occurred in the same year at both sites. These separate false-ring chronologies indicate that there may be phenological differences in the timing of radial growth in redcedar between Kansas and Oklahoma, and that the weather conditions responsible for false-ring formation often occur at the mesoscale and do not often impact central Kansas and northcentral Oklahoma simultaneously.

Ce travail de thèse, financé par l'Agence de Développement Economique de la Corse de la Collectivité Territoriale de Corse, a pour objectif la caractérisation de diverses plantes poussant à l'état spontané en Corse (genévriers, pin maritime) à travers la composition de leurs huiles essentielles et résines. Pour cela, différentes techniques chromatographiques et spectroscopiques ont été mises œuvre, incluant la RMN 13C des mélanges, méthode mise au point et développée au laboratoire. Ainsi, nous avons montré différents aspects de l'application de la méthode d'analyse par RMN 13C à travers i) l'étude d'huiles essentielles du genre Cymbopogon, ii) l'étude de la variabilité chimique des huiles essentielles d'hybrides entre la clémentine commune et la mandarine Willow Leaf et iii) la mise au point d'une méthode de quantification du dipropylene glycol. Ensuite, pour chacune des espèces du genre Juniperus (J. thurifera, J. oxycedrus ssp. macrocarpa, J. communis var. communis et J. communis ssp. alpina), nous avons réalisé un échantillonnage aussi important que possible en fonction de la disponibilité de la plante. Les huiles essentielles, préparées par hydrodistillation, ont été analysées par combinaison de différentes techniques (CC, CPG(Ir), CPG-SM, RMN 13C). Les résultats des analyses ont été soumis à un traitement statistique des données (ACP, k-means) quand le nombre d'échantillons était suffisant. Ainsi nous avons montré, dans deux cas, une variabilité chimique intraspécifique. Les échantillons de J. communis var. communis et de J. communis ssp. alpina se répartissent en deux groupes selon, dans le premier cas, leur teneur en -pinène et en sabinène et dans le second cas, leur teneur en limonène et en sabinène. Pour ce qui est des deux autres espèces, J. thurifera et J. oxycedrus ssp. macrocarpa, leurs huiles essentielles présentent comme composé majoritaire, respectivement le limonène et par l'-pinène, avec toutefois quelques compositions atypiques. Enfin, nous avons caractérisé l'huile essentielle de pin maritime (Pinus pinaster Ait.) de Corse, par l'analyse de 46 échantillons provenant d'aiguilles récoltées dans les différents massifs forestiers de l'île. Nous observons 3 types de compositions, dominées respectivement par des monoterpènes (-pinène), des sesquiterpènes (germacrène D) ou par des diterpènes (abiétadiène), sans qu'il soit possible pour autant de dégager des groupes bien définis. Nous avons également décrit la composition chimique de la fraction acide de 23 échantillons d'oléorésines de pin maritime, provenant de trois forêts de Corse. Sur l'ensemble de l'échantillonnage, nous avons identifié 11 acides résiniques. L'analyse statistique (ACP, K-means) de ces résultats suggère l'existence de deux groupes, sur la base des teneurs des acides lévopimarique et déhydroabiétique. Par ailleurs, le pinorésinol, produit à forte valeur ajoutée de la famille des lignanes, a été mis en évidence, pour la première fois à notre connaissance, dans des résines de pins.

Anthropogenic activities are responsible for increases in atmospheric CO₂ and climate change. These increases are partly counterbalanced by natural processes, such as carbon uptake in land surfaces. These processes are themselves subject to climate change, creating a coupled carbon-climate system. I investigated the carbon sink that woody encroachment represents, using a Central Texas savanna as study site, and studied how climatic factors influence this carbon sink. Woody plant encroachment, a worldwide structural change in grassland and savanna ecosystems, alters many ecosystem properties, but the net effect on the carbon balance is uncertain. Woody encroachment represents one of the key uncertainties in the US carbon balance, and demands a more detailed understanding. To come to a process-based understanding of the encroachment effect on carbon dynamics, I analyzed patterns of carbon exchange using eddy-covariance technology. I expected the imbalance between carbon uptake and release processes associated with the encroaching trees specifically, to be responsible for the carbon sink. I also expected that the sink would vary in time, due to strong links between carbon fluxes and soil water in this semi-arid ecosystem. I further studied the ecophysiology of the dominant species, as well as soil respiration processes under different vegetation types, and scaled these findings in space and time. I found that the ecosystem was a significant carbon sink of 405 g C m⁻² yr⁻¹. The encroaching trees increased photosynthesis by 180% and decreased soil respiration by 14%, compared to the grassland, resulting in a strong carbon sink due to the encroachment process. The encroaching process also altered carbon dynamics in relation to climatic drivers. The evergreen species Ashe juniper effectively lengthened the growing season and widened the temperature range over which the ecosystem acts as a carbon sink. The drought resistance of the encroaching trees reduced the sensitivity of this savanna to drought. I conclude that encroachment in Central Texas savannas increased the carbon sink strength by increasing the carbon inputs into the ecosystem. Woody encroachment also reduced the sensitivity to climatic drivers. These two effects constitute a direct effect, as well as a negative feedback to the coupled carbon-climate system. / text

Woody plant encroachment, that is, a substantial increase in the abundance of woody plants in a grassland or savanna, occurs in many parts of the world. It often has large effects on plant and animal populations and communities and on ecosystem properties and processes. However, little is known about the dynamics of woody plant encroachment and how these are affected by soils, by topography, and by the spatial pattern of the vegetation. Encroachment in turn can affect the spatial pattern of the vegetation. Using data from historical aerial photographs, I measured changes in woody plant cover and constructed, parameterized and compared a set of dynamic models of woody plant encroachment in central Texas savannas. These models predicted final woody cover from initial woody cover and the initial spatial configuration of woody plants. Then I incorporated soil and topography into these models to determine their effects. Finally, I examined the effects of encroachment on the spatial pattern of the vegetation. Incorporating negative density dependence in our models improved their fit, demonstrating that encroachment is density-dependent. A function that predicted the formation of new woody patches from a density-independent seed supply also improved the models' performance. The improvement in the models that resulted from incorporating the total length of woody-herbaceous edges confirmed that encroachment in this system occurs in part by the outward expansion of woody patches. The spatial pattern of the vegetation changed during woody plant encroachment. Spatial pattern (measured as degree of fragmentation) often had a non-linear relationship with cover. Furthermore, the spatial heterogeneity in fragmentation, that is, plot-to-plot variation in the degree of fragmentation, also changed during encroachment. Topography and soil type had, in general, little effect the dynamics of woody plant encroachment. Therefore, a relatively simple model of woody plant encroachment provided good predictions of woody cover at the end of the time periods. Other systems experiencing woody plant encroachment, forest succession, or invasion by non-native plants could be modeled using the same approach. / text

The objective of the study was to investigate the optimum conditions for in vitro regeneration of common juniper (Juniperus communis L.) by using indirect organogenesis approach. Throughout the study / callus induction, organogenesis, improved organogenesis and root induction experiments were performed sequentially. It was found that explant position, genotype, gender, treatments and sampling time had significant effects on callus induction rate in common juniper. The results of treatments indicated that IBA (indole-3-butyric acid) at concentration range 0.5-4.0 mg/l combined with MS medium supplemented with 0.1 mg/l BAP (benzylaminopurine), 3 % sucrose and 0.7% agar was the best one among the treatments to induce callus formation from common juniper explants collected as Spring buds. Also, a two-month culture was adequate period for the callus induction of common juniper regardless of position, before transferring the explants into organogenesis media. After a two-month culture in callus induction media, explants were transferred to organogenesis treatments in order to investigate adventitious bud development from callus tissues. There were significant differences among genotypes, treatments and explant-sampling times in initiation of organ development in common juniper. Additionally, it was found that excluding the auxin components while maintaining 1.0-2.0 mg/l BAP concentration in culture media, as refreshing after a month, stimulated the formation and development of adventitious buds and shoots. Among the treatments tested, it was found that 1.0 mg/l BAP plus 0.5 mg/l 2,4-D was the optimum culture media with adventitious bud formation capacity of 37.5% was though ageing of callus significantly affected the frequency of adventitious bud formation. Finally, rooting experiments were performed to investigate rooting efficiency of adventitious shoots. In the adventitious rooting experiments, no rooting was observed in any of the treatments used with common juniper explants. Although whole plantlet development from callus tissues could not be achieved as indirect organogenesis, the results of the study could aid to future studies dealing in vitro regeneration and production of secondary chemicals from common juniper.

Common juniper (Juniperus communis subs. Communis) is a shrub or a tree original in Czech Republic which was widespread there in the past. It grows either in lowlands or in mountains and it is undemanding to the soil. Nowadays, there is a lower number of it because of economic transformation in landscape. In consequence of this, it is on the list of endangered species of Czech Republic. The aim of this project was to chart the current state of population of common juniper in region "Mšály" (in The Protected Landscape Area of Třeboňsko). 144 living specimens were founded there. Dendrometry, mensuration of the specimens by GPS and drawing the new maps was realised. Another aim was comparison of the new data with the results of the research in 2004 2007. It was ascertained, that the observed population has a decreasing tendency in it's quantity. The average height of a specimen in the population has grew from 0,8 m to 4,9 m since 2007.

MADS-box genes are important regulators of reproductive development in seed plants, including both flowering plants and conifers. In this thesis the evolution of the AGAMOUS subfamily of MADS-box genes, and what the ancestral function of this group of genes might have been in the early seed plants about 300 million years ago, was addressed by the discovery of two novel conifer genes, both basal to all previously known AGAMOUS subfamily genes. DAL20, the most basal of these genes, was exclusively expressed in roots, unlike all previously known AGAMOUS subfamily genes. I also studied the evolutionary mechanisms leading to functional diversification of duplicated genes in two different subfamilies of MADS-box genes; the AGAMOUS and AGL6 subfamilies. Focus was on studying changes in gene expression pattern, representing changes in the transcriptional regulation between the genes, and on comparing the functional properties of the gene products, representing changes in the protein-coding sequence between the genes. Duplicated genes in the AGL6 subfamily were found to have evolved by both mechanisms. In the AGAMOUS subfamily I found duplicated spruce genes; DAL2 and DAL20, that appear to have functionally diversified mainly by changes in the transcriptional regulation. Conifer AGAMOUS subfamily genes were also used in a comparative developmental-genetics approach to evaluate hypotheses, based on the morphology of fossil and extant conifer seed cones, on the identity of the female reproductive organ, the ovuliferous scale, and the evolution of seed cone morphology in the conifer families Pinaceae, Taxodiaceae and Cupressaceae. Seed cones in these families have been hypothesized to have homologous ovule-bearing organs, but I found substantial differences in the expression patterns of orthologous AGAMOUS subfamily genes in seed cones of these families that are not compatible with this hypothesis, indicating that the evolutionary history of conifer seed cones is more diverse than previously thought.