京都大学 / 0048 / 新制・課程博士 / 博士(工学) / 甲第19752号 / 工博第4207号 / 新制||工||1649(附属図書館) / 32788 / 京都大学大学院工学研究科合成・生物化学専攻 / (主査)教授 濵地 格, 教授 梅田 眞郷, 教授 跡見 晴幸 / 学位規則第4条第1項該当 / Doctor of Philosophy (Engineering) / Kyoto University / DGAM

京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第20793号 / 医博第4293号 / 新制||医||1025(附属図書館) / 京都大学大学院医学研究科医学専攻 / (主査)教授 妻木 範行, 教授 鈴木 茂彦, 教授 瀬原 淳子 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

No description available.

No description available.

Die Fähigkeit sich an die Rotation der Erde und den daraus resultierenden Tag- und Nacht-Rhythmus anzupassen, basiert auf einer komplexen Regulation verschiedener physiologischer Prozesse. Auf molekularer Ebene liegt diesen Prozessen eine Orchestration von Uhr-Genen zugrunde – auch als innere Uhr bezeichnet – die einen aktivierenden bzw. reprimierenden Einfluss auf die Expression einer Vielzahl weiterer Gene hat. Ausgehend von dieser Regulation lassen sich auf unterschiedlichsten Ebenen tageszeitabhängige, wiederkehrende Rhythmen beobachten. Während diese wiederkehrenden Rhythmen auf einigen Ebenen bereits gut erforscht und beschrieben sind, gibt es weitere Ebenen wie den Metabolismus, über die das Wissen bisher noch begrenzt ist. So handelt es sich bei Drosophila beispielsweise um den Organismus, dessen innere Uhr auf molekularer Ebene wahrscheinlich mit am besten charakterisiert ist. Dennoch ist bisher nur wenig über Stoffklassen bekannt, deren Metabolismus durch die innere Uhr kontrolliert wird. Zwar konnte bereits gezeigt werden, dass sich eine gestörte innere Uhr auf die Anlage der Energiespeicher auswirkt, inwiefern dies allerdings einen Einfluss auf dem intermediären Stoffwechsel hat, blieb bisher weitgehend unerforscht. Auch die Frage, welche Metaboliten wiederkehrende, tageszeitabhängige Rhythmen aufweisen, wurde bisher nur für eine begrenzte Anzahl Metaboliten untersucht. Bei der hier durchgeführten Arbeit wurden deshalb zunächst die globalen Metabolit-Profile von Fliegen mit einer auf molekularer Ebene gestörten inneren Uhr (per01) mit Fliegen, die über eine funktionale Uhr verfügen (CantonS), zu zwei Zeitpunkten verglichen. Um die Anzahl der zeitgleich untersuchten Gewebe und somit die Komplexität der Probe zu reduzieren, wurden hierfür die Köpfe von den Körpern der Fliegen getrennt und separat analysiert. Beide Körperteile wurden sowohl auf kleine hydrophile als auch auf hydrophobe Metaboliten hin mittels UPLC-ESI-qTOF-MS untersucht. Die anschließend durchgeführte, statistische Analyse brachte hervor, dass sich Unterschiede zwischen den beiden Fliegenlinien besonders in den Spiegeln der essentiellen Aminosäuren, den Kynureninen, den Pterinaten sowie den Spiegeln der Glycero(phospho)lipiden und Fettsäureester zeigten. Bei den Lipiden zeigte sich, dass die Auswirkungen weniger ausgeprägt für die Anlage der Speicher- und Strukturlipide als für die Intermediate des Lipidabbaus, die Diacylglycerole (DAGs) sowie die Acylcarnitine (ACs), waren. Um zu bestätigen, dass die inneren Uhr tatsächlich einen regulatorischen Einfluss auf die ausgemachten Stoffwechselwege hat, wurden anschließend die Spiegel aller Mitglieder darauf hin untersucht, ob diese wiederkehrende, tageszeitabhängige Schwankungen aufweisen. Hierfür wurden Proben alle zwei Stunden über drei aufeinanderfolgende Tage genommen und analysiert, bevor mittels JTK_CYCLE eine statistische Analyse der Daten durchgeführt und die Metaboliten herausgefiltert wurden, die ein rhythmisches Verhalten bei einer Periodenlänge von 24h zeigten. Hierbei bestätigte sich, dass besonders die Mitglieder des intermediären Lipidmetablismus hiervon betroffen waren. So konnten zwar auch für einige Aminosäuren robuste Rhythmen ausgemacht werden, besonders ausgeprägt waren diese jedoch erneut bei den DAGs und den ACs. Die abschließende Untersuchung letzterer unter Freilaufbedingungen (DD) sowie in per01 brachte hervor, dass die ausgemachten Rhythmen unter diesen Bedingungen entweder nicht mehr detektiert werden konnten oder deutlich abgeschwächt vorlagen. Lediglich zwei kurzkettige ACs zeigten auch unter DD-Bedingungen statistisch signifikante Rhythmen in ihren Spiegeln. Dies spricht dafür, dass neben der Regulation durch die innere Uhr weitere Faktoren, wie beispielsweise das Licht, eine entscheidende Rolle zu spielen scheinen. / The ability to adapt to the rotation of the earth and to the resulting day and night rhythm is based on a complex regulation of various endogenous processes. At the molecular level, these processes are based on an orchestration of clock genes - also known as the endogenous clock – which have an activating or repressing influence on the expression of diverse clock-controlled genes. Based on this regulation, recurring rhythms depending on the time of day can be observed at various levels, ranging from gene expression to behavior. While these recurring rhythms have been well characterized on certain output levels, little is known, however, on other levels like their influence on metabolism. Drosophila for example, is an organism whose endogenous clock is probably best characterized at the molecular level. However, little is known about substance classes and metabolic pathways that are controlled by the endogenous clock. It has already been shown that an impaired endogenous clock affects energy storage, but how an impaired clock influences intermediary lipid metabolism remains still unknown. Additionally, little is known on metabolites or metabolite classes, that display recurring, time-dependent rhythms. So far this has only been studied for certain metabolites or metabolite classes. Therefore, we compared global metabolite profiles at two timepoints between flies with an impaired endogenous clock (per01) and WT flies, possessing a functional clock (CantonS). In order to reduce the number of different tissues studied at once and thus the complexity of the sample, fly heads were separated from fly bodies and analyzed separately. In both body parts levels of small hydrophilic and hydrophobic metabolites were studied using UPLC-ESI-qTOF-MS. The subsequent statistical analysis revealed differences between the two fly lines, associated with the metabolism of essential amino acids, kynurenines, pterinates, glycero(phospho)lipids and fatty acid esters. Closer inspection of the lipid classes being affected revealed, that the effects were less pronounced for the formation of storage- and structural- lipids, compared to the intermediates of lipid degradation, the diacylglycerols (DAGs) and acylcarnitines (ACs). In order to confirm that the endogenous clock has indeed a regulatory influence on these metabolic pathways, the levels of all members were studied in a time-course experiment to determine, whether they display recurring, time-of-day-dependent fluctuations. For this purpose, samples were taken every two hours for three consecutive days, with heads and bodies analyzed separately, before a statistical analysis was carried out using JTK_CYCLE. The results were then filtered for those metabolites that showed a rhythmic behavior with a period length of 24 hours. The results confirmed that members of the intermediary lipid metabolism were particularly affected. Although robust rhythms could be detected for some amino acids, multiple DAG and AC species showed even more pronounced rhythms. The subsequent analysis of the latter under freerunning conditions (DD) and in per01 showed that the identified rhythms either diminished completely under these conditions or were significantly weakened. Only two short-chain ACs showed statistically significant rhythms in their levels under DD conditions. This suggests that in addition to regulation by the internal clock, other factors, such as light, seem to play a crucial role.

No description available.

京都大学 / 0048 / 新制・課程博士 / 博士(農学) / 甲第22499号 / 農博第2403号 / 新制||農||1077(附属図書館) / 学位論文||R2||N5279(農学部図書室) / 京都大学大学院農学研究科応用生物科学専攻 / (主査)教授 佐藤 健司, 教授 菅原 達也, 准教授 豊原 治彦 / 学位規則第4条第1項該当 / Doctor of Agricultural Science / Kyoto University / DFAM

Mémoire numérisé par la Direction des bibliothèques de l'Université de Montréal.

Date palm (Phoenix dactylifera L.) is a fruit-bearing tree with numerous potential sustainable applications. Since ancient times, it has been considered a stable, secure, and sustainable food. This work provides comprehensive metabolic profiling of both parts, flesh and seed, of four P. dactylifera cultivars; Ajwa, Anbara, Sukkari, and Degelt Nour, which originated from two countries, Saudi Arabia, Tunisia. The analysis performed using mass spectrometry-untargeted metabolomics approaches, included a combination of LC-MS and GC-MS coupled to multivariate statistical analysis to reveal the differences in metabolite compositions among date varieties. The LC-MS seed results showed several classes of metabolites that belong to the flavonoids, phenolic acids, and amino acids derivatives, including citric acid, malic acid, lactic acid, hydroxyadipic acid, caffeic acid, which were at high concentrations in AJS followed by DNS and ARS. The LC-MS flesh analysis displayed that DNF had a high level of Isoquercitrin (flavonoid compound) and sinapic acid, and AJF was high concentrations level in hydroxyadipic acid and ascorbic acid. GC-MS concluded that seed samples of four date varieties are richer in metabolites classes than the flesh samples. The metabolites contributed to the seed metabolite compositions included several classes of amino acids, hydrocinnamic acids (caffeic, ferulic and sinapic acids), antioxidant phenolics, and long-chain fatty acids. The PCA and its loading analysis demonstrated the discriminating metabolites that were responsible for date varieties segregation, as HCA displayed the metabolic patterns and groups of metabolites that drive the clustering of the date samples, two groups of dates clustered together (AR and AJ) and (SR and DN). These clusterings are based on the similarities and differences observed in the metabolite compositions of the studied date samples that could be explained by differences in various metabolic responses and the environmental conditions, genotypes and geographical regions. This extensive date palm information would increase the potential of date fruits and seeds as low-cost sources of natural diet that may provide nutritious and bioactive components in the food and pharmaceutical fields to produce value-added products.

Maize (Zea mays L.) is a global food staple and is at risk from infection by the pathogenic fungus Aspergillus flavus L. The ubiquitous, soil-borne fungus causes ear rot of maize and produces the carcinogenic secondary metabolite known as aflatoxin. Aflatoxin B1 is the most potent carcinogenic mycotoxin known, causing hepatocellular carcinoma, along with many other serious health problems such as immunosuppression. Previous studies have shown that maize cob tissue plays an essential role in both facilitating and limiting the spread of the fungal pathogen A. flavus, however, little attention in the literature has been given to the cob. To date, there have not been any studies published describing the metabolome of maize cob tissue. This study assessed three different methods for disruption of maize cob tissue and investigated the global metabolome of maize cob of two resistant (Mp313E and Mp420) and two susceptible (B73 and SC212m) genotypes via liquid chromatography-mass spectrometry. Three treatments (control, water-inoculated, and fungus-inoculated) and three-time points (3, 9, 15 days after inoculation) were included in the experimental design. For the first time in maize cob, 69 metabolites were identified via the mzCloud online database. Out of them, 28 metabolites showed statistically significant differences in abundance across the treatments. Twenty-two metabolites were identified via Fragment Ion Search, of which 14 were statistically significant differences in abundance across the treatments. The majority of the metabolites identified where from the phenylpropanoid, linoleic acid, and terpenoid biosynthesis pathways. Thousands of unknown ions were detected, and for 521 compounds the formula could be derived, based on accurate monoisotopic masses. In the targeted metabolomics analysis, the MS3 spectral tree was obtained for zealexin B1 for the first time via a highly induced sample and was subsequently used to identify zealexin B1 in maize cobs (Va35). To date, this work is the sole metabolomic profiling study of maize cob tissue, and it provides insight into constitutive and induced molecular antifungal defenses of resistant and susceptible genotypes. A list of significant fungal-induced metabolites related to the maize-A. flavus defense response was compiled for further targeted metabolomic identification.