Les cyanobactéries ont développé des mécanismes de photo-protection pour se prémunir des dommages causés par un excès de lumière. L’un d’eux repose sur l’activité de l’Orange Caroténoïde Protéine (OCP), protéine soluble qui attache un kéto-caroténoïde, l’hydroxy-echinenone. Sous illumination, l’OCP se photo-convertit en forme active et interagit avec les phycobilisomes pour dissiper l’énergie collectée sous forme de chaleur. En conséquence, l’énergie d’excitation reçue par les centres réactionnels et la fluorescence du complexe photosynthétique diminuent. L’OCP a aussi la faculté de neutraliser l’oxygène singulet pour lutter contre la photo-oxydation. J’ai développé un système d’expression hétérologue pour reconstituer la voie de biosynthèse de cette protéine dans E.coli. Ce système permet l’obtention d’une grande quantité d’OCP liant son caroténoïde in vivo. Grâce à ce système robuste et rapide, les OCPs de trois cyanobactéries : Synechocystis, Arthrospira et Anabaena ont été produites, liant différents caroténoïdes. Toutes les OCPs recombinantes sont photo-actives et capables de quencher la fluorescence des phycobilisomes in vitro. Elles possèdent toutes la faculté de neutraliser l'oxygène singulet quel que soit le caroténoïde lié. Ce système d'expression hétérologue nous a permis d’élucider les déterminants structurels impliqués dans la photo-activation et la structure de la forme active de l’OCP. Il constitue une avancée fondamentale dans l'étude des protéines à caroténoïde et dans la production d'antioxydants solubles qui présentent un grand intérêt pour l’industrie de la santé. / Cyanobacteria have developed some photo-protective mechanisms to protect themselves from stress caused by excess light. One of them relies on the activity of the soluble Orange Carotenoid Protein (OCP) that binds a keto-carotenoid, the hydroxyechinenone. Under illumination, the OCP gets photo-converted to an active form and can interact with phycobilisomes to dissipate the collected energy as heat. Consequently, the excitation energy arriving at the photosynthetic reaction centers and the phycobilisome fluorescence emission decrease. The OCP can also quench the singlet oxygen to fight against photo-oxidation. I developed a heterologous expression system in which the biosynthetic pathway of the OCP is built in E.coli. The expression system allows the production of a large amount of OCP binding its carotenoid in vivo. Thanks to this robust and fast expression system, OCPs from three different cyanobacteria: Synechocystis, Anabaena and Arthrospira were produced, binding different carotenoids. All recombinant OCPs are photoactive and able to induce a large phycobilisome fluorescence quenching. Moreover, they all have the ability to quench the singlet oxygen, whatever the bound carotenoid. This heterologous expression system allowed us to elucidate the structural determinants involved in the photo-activation and structure of the active form of the OCP. This work represents a fundamental advance in the study of caroteno-proteins and in the production of others soluble antioxidants that are of great interest to the health industry.

La tomate est le deuxième légume le plus consommé dans le monde et la plus grande partie est consommée sous forme transformée. La tomate d’industrie est typiquement une culture exigeante en eau. La production se situe majoritairement en zone méditerranéenne, confrontée à des périodes de sécheresse de plus en plus fréquentes, et contrainte, dans le contexte de changement climatique, à réduire le prélèvement des ressources en eau pour son agriculture. L’eau est un facteur majeur qui impacte le rendement et la qualité des tomates et il y a donc un lien direct entre l’amont agricole (production), et l’aval industriel (transformation). Bien que des travaux visant à mieux comprendre les liens pré-post-récolte existent, mieux adapter la production face aux contraintes climatiques en évolution reste un challenge pour la filière, et c’est dans ce cadre que ce situe ce travail de thèse. Dans ce contexte, l’objectif de la thèse était d’analyser au cours de la chaine de production et de transformation, l’impact de facteurs agronomiques et technologiques sur la qualité des purées de tomate, et d’identifier au cours de cette chaine des leviers pour améliorer la qualité gustative, nutritionnelle et environnementale du produit fini. Etant donné cet objectif, ce travail a été construit pour analyser 1) quels sont les principaux facteurs agronomiques et technologiques responsables de la variabilité de la qualité des fruits frais et transformés au sein des zones de production françaises de tomates d’industrie ; 2) quels sont les effets de la variété, des apports en eau, du stade de développement, et du procédé sur la qualité des fruits frais et leur aptitude à la transformation ; 3) Quels sont les processus responsables des modifications des propriétés rhéologiques des purées. Nos résultats ont montré des différences de qualité des fruits et des produits finis entre les deux grandes régions de production françaises de tomates d’industrie en 2015. Le choix du cultivar et des pratiques, comme la densité de plantation ou la durée du cycle de production, étaient responsables de ces différences. En l’absence de déficit hydrique, les différences climatiques (amplitudes thermiques et cumul de précipitations) n’ont pas semblé être un facteur majeur de variation. L’exploration des liens pré-post-récoltes a ensuite confirmé que les critères à la récolte couramment utilisés (rendement et °Brix) renseignaient peu sur l’aptitude à la transformation des tomates. De plus, nous avons montré que diminuer l’irrigation jusqu’à ne remplacer que 60 % de l’évapotranspiration des plantes n’a que très peu affecté le rendement en fruit frais, et a augmenté le rendement en matière sèche jusqu’à +27%, ce qui est positif pour l’usine. L’efficience de l’utilisation de l’eau d’irrigation a ainsi été augmentée d’en moyenne 20% sur tout le cycle. La composition de la matière sèche des fruits à la récolte n’a pas été significativement affectée par le déficit hydrique en 2016. En revanche, lorsque ces mêmes fruits ont été transformés selon des procédés industriels, le déficit hydrique a amélioré la viscosité de la purée sans affecter sa couleur. Il a également limité la perte de viscosité obtenue lorsque le procédé possède une phase de macération à basse température (Cold Break (CB)). Ce résultat a suggéré que la réactivité enzymatique des tissus a été réduite. En revanche, un stress hydrique plus drastique en 2017 a induit des fortes différences de teneur en matière sèche. Comme les purée sont concentrée à la même teneur en SSC, alors les purée WD, où moins d’eau a été éliminée, montre une viscosité plus faible. La maitrise de la matière sèche n’est pas le seul facteur affectant la rhéologie des purées. C’est un phénomène complexe dépendant de plusieurs facteurs. / Tomato is the second most consumed vegetable in the world and most of it is consumed in processed form. The industry tomato is typically a demanding crop in water. Production is mainly in the Mediterranean zone, facing increasingly frequent periods of drought, and constrained, in the context of climate change, to reduce the withdrawal of water resources for its agriculture. Water is a major factor affecting the yield and quality of tomatoes, so there is a direct link between upstream agriculture (production) and industrial downstream (processing). Although works to better understand pre-post-harvest links exists, better adapting production to changing climatic constraints remains a challenge for the industry, and it is in this context that this thesis work is located. In this context, the objective of the thesis was to analyze during the chain of production and processing, the impact of agronomic and technological factors on the quality of tomato purées, and to identify during this chain levers to improve the taste, nutritional and environmental of the finished product. Given this objective, this work was constructed to analyze 1) what are the main agronomic and technological factors responsible for the variability of the quality of fresh and processed fruit within the French production areas of industrial tomatoes; 2) what are the effects of variety, water supply, stage of development and process on the quality of fresh fruits and their suitability for processing; 3) what are the processes responsible for the changes in the rheological properties of the purees. Our results showed differences in the quality of fruit and processed products between the two major French industrial tomato production regions in 2015. The choice of cultivar and practices, such as planting density or production cycle time, were responsible for these differences. In the absence of a water deficit, climatic differences (thermal amplitudes and cumulative precipitation) did not seem to be a major factor of variation. Exploration of pre-post-harvest links then confirmed that the commonly used harvest criteria (yield and °Brix) provided little information on the processing ability of tomatoes. In addition, we have shown that decreasing irrigation to replace only 60% of plant evapotranspiration has had little effect on the yield of fresh fruit, and has increased dry matter yield to + 27%, which is positive for the plant. The efficiency of the use of irrigation water has thus been increased by an average of 20% over the entire cycle. The composition of the dry matter of the fruits at harvest was not significantly affected by the water deficit in 2016. On the other hand, when these same fruits were transformed according to industrial processes, the water deficit improved the viscosity of the purees without affecting its color. It also limited the loss of viscosity obtained when the process has a low temperature maceration phase (Cold Break (CB)). This result suggested that the enzymatic reactivity of the tissues was reduced. On the other hand, more drastic water stress in 2017 led to significant differences in dry matter content. As the purees are concentrated to the same SSC content, then the WD purees, which less water has been removed, shows a lower viscosity. The mastery of the dry matter is not the only factor affecting the rheology of purees. It is a complex phenomenon depending on several factors. Indeed, when the differences in viscosity are induced by the cooking method, it is mainly the viscosity of the serum (liquid part of the mash) that is involved. When the viscosity variations are induced by the use of Variety with a high potential for viscosity (Ketchup tomato), it is the size and shape of the particles and their aggregation potential that are involved.

Global energy demands have escalated over the past few decades, creating a necessity for alternative energy sources. Solar technologies inspired by the primary solar energy storing process known on earth, photosynthesis, have subsequently gained popularity. The natural photosynthetic apparatus comprises a network of membrane-bound pigment-protein complexes, with the main plant light-harvesting complex (LHCII) consisting of chlorophyll (Chl) and carotenoid (Car) pigments. Electronic excitation energy transfer (ET) of the harvested energy takes place amongst these pigments on ultrafast timescales. This energy is funnelled towards a photosynthetic reaction centre where charge separation is achieved, creating a Biobattery, which powers the subsequent manufacture of energy-rich chemical compounds for photosynthetic activity. Transient absorption pump-probe spectroscopy has proven to be a useful technique for monitoring the evolution of the excited state dynamics, such as electronic transitions and excitation ET amongst Car and Chl pigments of LHCII trimers isolated from spinach leaves. This method was utilized to probe samples excited under four different conditions: at pump excitation wavelengths (𝜆𝑒𝑥) of 489 nm (preferentially exciting Cars Lutein1 and Neoxanthin) and 506 nm (targeting Cars Lutein2 and Violaxanthin), each with an intensity of either 800 nJ/pulse (relatively high) or 500 nJ/pulse (comparatively low). A global analysis was applied to each dataset using the robust, open-source Glotaran software, from which three kinetic decay lifetimes for the various processes were extracted. General spectral observations encompassed a negative pump ground state bleach (GSB) at each 𝜆𝑒𝑥; negative Chl b and Chl a GSBs, superimposed with negative stimulated emission (SE) signals; and a positive excited state absorption (ESA) band. The first lifetime of a few picoseconds corresponded mainly to Car-S2 depopulation, resulting either from energy relaxation towards Car-S1, or ET to Chls. Small, but distinct Chl b signals of less than 3 mOD were also detected on this timescale. The second lifetime, which is between 10 and 12 ps, was characteristic to the Lutein Car-S1 lifetime, mainly depicting Car-S1 ET to Chl a. The third lifetime, which extended from ~200 ps to the nanosecond timescale, was attributed to Chl a fluorescence. The 𝜆𝑒𝑥 of 489 nm directly excites the Chl Soret region, whilst excitation at 506 nm shows a pump intensity-dependence. Laser pulse photon density values were ~1014 photons·cm-2·pulse-1 for these datasets. Singlet-singlet annihilation calculations performed on the samples excited at 506 nm provided low annihilation probabilities of 9.0% and 11.5% for a low and high pump intensity, respectively, limiting the possibility of sample photobleaching. Optimization and redevelopment of the experimental setup significantly improved both the data quality and various recorded parameters, concluding that pump-probe spectroscopy was successful on the prepared LHCII trimers. Results acquired and calculations performed correlated with literature, where minimal changes were noticed in the timescales and ET pathways. The robustness of plant systems was confirmed through both excitation-wavelength and intensity dependence. This work paves the way for advanced studies on the role Cars play in non-photochemical quenching (NPQ), a self-protection mechanism of plants against over-illumination; and for the tailoring of artificial light-harvesting antennas based on research conducted on their natural counterparts. / Globale energievereistes het oor die afgelope paar dekades toegeneem, wat die ontwikkeling van alternatiewe energiebronne noodsaaklik maak. Sontegnologieë, geïnspireer deur die primêre sonenergiebergingsproses op aarde, fotosintese, het daarom gewild geword. Die natuurlike fotosintetiese apparaat bestaan uit 'n netwerk van membraangebonde pigment-proteïenkomplekse, met die hoof ligversamelingskompleks in plante (LHCII) wat bestaan uit chlorofil- (Chl) en karotenoïed- (Car) pigmente. Die energie wat deur die pigmente geabsorbeer word, word tussen elektroniese opgewekte toestande op verskillende pigmente op ultravinnige tydskale oorgedra. Hierdie energie word na ʼn fotosintetiese reaksiesentrum gekanaliseer, waar 'n ladingskeiding geïnduseer word en 'n Biobattery sodoende geskep word. Die energie wat in dié battery gestoor is, word gebruik om energieryke chemiese verbindings te vervaardig — wat as brandstof vir die plant dien om sy lewensfunksies te verrig. Tydopgeloste-absorpsie-pomp-tasting-spektroskopie is 'n nuttige tegniek om die dinamika tussen opgewekte toestande te volg. ‘n Voorbeeld van sulke dinamika is die elektroniese opwekking en energie-oordrag tussen die Car- en Chl-pigmente van geïsoleerde LHCII-trimere in spinasieblare. Hierdie metode is gebruik om monsters onder vier verskillende toestande te ondersoek by pompgolflengtes (𝜆𝑒𝑥) van 489 nm (waar hoofsaaklik die Cars Luteïne1 en Neoksantine opgewek word) en 506 nm (vir Cars Luteïne2 en Violaksantine), en pompenergieë van ‘n relatief hoë 800 nJ/puls, of 500 nJ/puls vir elke golflengte. / Dissertation (MSc)--University of Pretoria, 2017. / National Research Foundation (NRF) / Physics / MSc / Unrestricted

Fresh fruits and vegetables (FFVs) are the most frequently wasted foods because of their perishability and handling requirements. However, there is a lack of information on how much each step of the supply chain impacts FFVs quality, particularly on tomatoes, and what measures need to be taken for an immediate and effective impact on waste reduction. There is also no information on how the supply chain affects the proteome of the tomato and what proteins are differentially regulated by the most impactful steps of the supply chain. The objectives of the work presented on this thesis were to evaluate the decline in the overall quality and quantify tomato waste at each step of the supply chain, from the farm to consumer; and to determine what proteins are impacted by the decline in quality that is associated with temperature abuse. To determine overall quality and tomato waste, light-red tomatoes were exposed to an optimum temperature (13 °C) and eighteen different time-temperature scenarios, normally encountered during supply chain, and sensory and physicochemical attributes measured at each step. To determine the impact of chilling and non-chilling temperatures normally encountered during tomato supply chain, on the proteome, light-red tomatoes were exposed to an optimum temperature (13 °C) and to two time and temperature supply chain scenarios (2 °C and 25 °C) that showed the most negative impact on tomato overall quality, and physicochemical and proteomic attributes were measured at each step. For the first tomato harvest, the steps with the highest impact on quality and waste were shipping to distribution center (DC; 20°C), cooling at the grower (25°C) and storing at the consumer (4°C). For the second tomato harvest, shipping to the store (2°C), cooling at the farm (10°C) and displaying at the store (20°C) negatively impacted quality. High temperatures during cooling, shipping and store display impacted sensory quality and resulted in increased weight loss, and decreased sugar, carotenoids, and ascorbic acid contents. Although low temperatures during shipping, cooling and consumer did not impact tomato sensory quality, they contributed to a decline in sugar, carotenoids and ascorbic acid contents. Overall, the most impactful steps on tomato quality and waste, regardless of the temperature, were shipping to DC, cooling, shipping to stores, displaying at the store, and consumer storage. Analysis of the differentially expressed proteins in the tomato showed that metabolic proteins were greatly impacted by temperature abuses such as phosphomannomutase, heme oxygenase 1, and MAP kinase; and that proteins regulating cellular membrane integrity such as vacuolar protein sorting-associated protein were also impacted.

Sexual selection theory tries to explain evolution of apparently useless traits which mainly developed in males of numerous species. One such trait is also rich and vibrant coloration, typical for many of the bird species. These traits are difficult to be falsified, and therefore they honestly signal quality of the individual. Carotenoid coloration reflects the health condition and melanin coloration the social status, even though this traditional division might not be as strict according to the latest studies. Apart from these ways of maintaining honesty, recent studies are focusing more also on the effect of hormones, mainly two steroids: male sexual hormone - testosterone -, and the "stress" hormone - corticosterone. Both hormones could positively influence male's sexual traits such as ornamental coloration. On the other hand, elevated levels of these hormones possess risk to the organism (higher energetic expenditure, chronic stress), therefore also might potentially become costly. Using photographs of birds in standardized conditions as well as spectrophotometry I analyzed the plumage coloration of males of the Yellowhammer (Emberiza citrinella). Concentrations of testosterone and corticosterone deposited in feathers were analyzed using the LC-MS/MS. Also, for males in breeding season 2015,...

Corynebacterium poinsettiae mutant strains blocked in carotenoid biosynthesis were obtained by treatment with the mutagen N-methyl-N1-nitro-N-nitrosoguanidine. Additional carotenoid (Crt) mutant strains were obtained from a previous study conducted in our laboratory. Fifty-nine Crt mutants affected in carotenoid biosynthesis were examined by a normal phase high performance liquid chromatography (HPLC) system. Pigment extracts of Crt mutants and C. poinsettiae wild type strains were resolved by an isocratic system with hexane:acetone:dicholoromethane, 11.35:1.73:1.00 (by vol.) as the eluting solvent. In addition to the five major peaks, twelve minor peaks were observed in the wild type C. poinsettiae strain used in this study. Crt mutant and wild type strain peak heights were measured from the individual chromatograms and the peak height data set created was analyzed using the Statistical Analysis System program to perform a cluster analysis. The cluster analysis revealed five carotenoid mutant groups. Carotenoid pigments which accumulated or were absent in each of the cluster groups are reported. Cluster group 1 mutants (CrtA) are blocked in the dehydrogenase(s) which is(are) responsible for the dehydrogenations between phytoene and lycopene. Cluster group 2 mutants (CrtB) appear to be blocked at a second dehydrogenase specific for the dehydrogenation from C.p. 470 to C.p. 496. Cluster group 3 mutants (CrtC) are blocked at a cyclization step in the pathway which involves cyclization of C.p. 496 to C.p. 470 and which may cyclize C.p. 473 to C.p. 450. The genes CrtA and CrtB map only 0.5 map units from each other while CrtA and CrtC map 2.1 map units from one another. Mutants which accumulate end products but which lack certain precursors indicate a branched pathway for pigment biosynthesis exists in this organism. Media for the formation, fusion and regeneration of C. poinsettiae protoplasts are reported and a protocol for the use of these media in genetic crosses of strains blocked in carotenoid biosynthesis is described. While isolating antibiotic resistant mutants useful in genetic analyses, novobiocin resistant mutants were observed to have a distinctly different colony pigment phenotype as compared to the wild type strain. HPLC chromatograms of a novobiocin resistant strain showed a distinctly different carotenoid pigment profile. The results provide evidence for differential gene expression in the carotenoid biosynthetic pathway when these mutants are grown in the presence of novobiocin.

No description available.

It has been well documented that fruit and vegetable (F/V) intake is linked to lower risk of mortality and chronic disease. Raman resonance spectroscopy is a valid indicator of F/V intake and quantifies that intake by measuring skin carotenoid levels. In this study, 46 children, participated in a 6-week feeding study wherein they were randomly assigned to one of two groups: 1) consuming a high-carotenoid (HC) breakfast/snack food, or 2) consuming a placebo bar, every day. The HC food contained 4.3mg carotenoids per serving and the placebo contained none. Skin carotenoids were measured every two weeks using a BioPhotonic scanner. The treatment group had a mean increase in scanner score of 5,802 Raman intensity units which was significantly higher than the mean increase of the placebo group, 1,771. In this study we found that consumption of 120 gm of a high-carotenoid food significantly increased skin carotenoid levels in children ages 5-18 over a 6 week period. Packaging type as well as storage conditions play a role in preserving carotenoids which are sensitive to light, temperature, and oxygen. Care must be taken when choosing packaging and storage conditions for foods containing carotenoids. Two shelf-life studies, one at room-temperature and another in frozen storage, were conducted on the HC breakfast/snack food to determine the best method of packaging to maintain quality and preserve carotenoids. The food was randomly packed into one of three packaging types for both studies. The packages used in the frozen study were then randomly assigned to one of three freezer storage methods. Room-temperature samples were analyzed on days 3, 7, 10, and 14 and samples in frozen storage were pulled every month for 5 months. Measures of water activity, moisture content, color values (L*a*b*, chroma, and hue), and carotenoid content were analyzed in each sample. At the conclusion of each study, L*, b*, chroma, and hue were significantly affected by packaging type. No significant associations were found in any other measures. From this study we draw the conclusion that cellophane packaging or packaging with a N2 backflush would be the best options for use with this food to best retain its quality.

京都大学 / 0048 / 新制・課程博士 / 博士(農学) / 甲第19039号 / 農博第2117号 / 新制||農||1032(附属図書館) / 学位論文||H27||N4921(農学部図書室) / 31990 / 京都大学大学院農学研究科応用生物科学専攻 / (主査)教授 菅原 達也, 教授 澤山 茂樹, 教授 佐藤 健司 / 学位規則第4条第1項該当 / Doctor of Agricultural Science / Kyoto University / DFAM

Allopolyploids are formed by interspecific hybridization and whole genome duplication, with the resulting organism contains multiple distinct subgenomes in one nucleus. Subgenomic interactions result in massive genetic and epigenetic reconstruction, contributing to variable phenotypic traits noted in newly formed allopolyploids. To better understand these mechanisms in the context of molecular biology, evolution, and plant breeding, plant biologists study the model organism Brassica napus (farmed as canola or oilseed rape). With white-flowering and yellow-flowering progenitors, flower color phenotype of B. napus presents an opportunity to examine subgenomic interactions. CAROTENOID CLEAVAGE DIOXYGENASE 4 (CCD4) is known to play a major role in determining flower color phenotype of carotenoid-synthesizing angiosperms. Here, we investigate the genetic and epigenetic role of CCD4 orthologs and their role in flower color phenotype of B. napus.