Academic literature on the topic 'FLAVONOID’S PATTERN'

Many samples of Monoclea gottschei from Central and South America and the Caribbean Islands have been compared for their flavonoids. The species exhibits the most diverse flavonoid pattern ever observed for a liverwort. From different chemotypes 28 flavones were isolated: 8 aglycones, 3 monoglycosides, 9 diglycosides, 6 triglycosides and 2 oligo- or polyglycosides. The chromatographic and spectroscopic data of previously unidentified compounds are presented. Samples of M . forsteri from New Zealand have been analyzed for flavonoids too, but their flavone patterns are much less diverse: 1 aglycone, 1 polyglycoside and 2 further mono-or-diglycosides were isolated. The differences in flavonoid production of both species are an additional feature for the distinction of the two Monoclea species

During the chemotaxonomic study of 16 species of Stemonoporus, three flavonoid aglycones (flavonol quercetin, flavonol kaempferol and flavone apigenin) and glycosides (quercetin 3- glucoside, quercetin 3-rutinoside and apigenin 5-glucoside) were isolated while flavonol myricetin, flavone luteolin and proanthcyanidin were not detected in any of the species surveyed. The isolated flavonoids can be used as chemotaxonomic markers. The species of Stemonoporus can be regarded as advanced in flavonoid pattern because of the absence of myricetin and loss of proanthocyanidins. The data of the flavonoid patterns and the outcome of cluster analysis are taxonomically useful to resolve the controversies over the systematic arrangement of the species and suggest the need for a revision of classification of the genus Stemonoporus. Key words: Dipterocarpaceae; Flavonols; Chemotaxonomy; Cluster analysis. Journal of Natural History MuseumVol. 24, 2009Page: 146-155

SUMMARY We report on the first case in which sequestered secondary plant compounds determine an insect’s external appearance in the ultraviolet spectrum and thereby influence visually mediated mate choice. Larvae of the common blue butterfly Polyommatus icarus specifically sequester flavonoids in different amounts and types, depending on the part or species of food plant. During late pupal development the majority of ultraviolet-absorbing flavonoids are deposited in the wing scales. The flavonoid content of the larval diet thereby determines ultraviolet wing patterns. In laboratory and field experiments, male butterflies clearly preferred flavonoid-rich, ultraviolet-absorbing female dummies. This preference is mediated visually by the ultraviolet pattern of the wings. Food-plant parts and species vary in value as a food source, so ultraviolet wing patterns may signal mate quality and are not a species-specific characteristic. We discuss the use of principal component analysis in analysing spectral data in the context of visual communication. We propose the alternative application of confidence intervals of averaged spectra as a novel straightforward statistical method for comparing groups of spectra in a manner that is independent of assumptions about the visual system of the receiver. In addition, they can be used to give confidence intervals to derived measures of colour such as quantum catch by photoreceptors.

AbstractFlavonoids are a group of phenolic secondary metabolites in plants that have important physiological, ecological and economic value. In this study, using the desert plant Artemisia sphaerocephala Krasch. as the sample material, the content and components of the total flavonoids in its seeds at seven different developmental stages were determined. In addition, the genes involved in flavonoid metabolism were identified by full-length transcriptome sequencing (third-generation sequencing technology based on PacBio RS II). Their expression levels were analysed by RNA-seq short reading sequencing, to reveal the patterns and regulation mechanisms of flavonoid accumulation during seed development. The key results were as follows: the content of total flavonoids in mature seeds was 15.05 mg g−1, including five subclasses: flavonols, chalcones, flavones, flavanones and proanthocyanidins, among which flavonols accounted for 45.78%. The period of rapid accumulation of flavonoids was 40–70 d following anthesis. The high expression of phenylalanine ammonia-lyase (PAL), 4-coumarate-CoA ligase (4CL) and UDP-glucose:flavonoids 3-o-glucosyltransferase (UF3GT) promoted the accumulation of total flavonoids, while the high expression of flavonoids 3′-hydroxylase (F3′H) and flavonols synthase (FLS) made flavanols the main component. Transcription factors such as the MYB-bHLH-WDR (MBW) complex and Selenium-binding protein (SBP) directly regulated the structural genes of flavonoid metabolism, while C2H2-type zinc finger (C2H2), Zinc-finger transcription factor (GATA), Dehydration-responsive element binding (DREB), Global Transcription factor Group E protein (GTE), Trihelix DNA-binding factors (Trihelix) and Phytochrome-interacting factor (PIF) indirectly promoted the synthesis of flavonoids through hormones such as brassinoidsteroids (BRs) and abscisic acid (ABA). These results provided valuable resources for the application of related genes in genetics and breeding.

Lonicera maackii (Caprifoliaceae) is a large, upright shrub with fruits that contain many bioactive compounds. Flavonoids are common active substances in L. maackii. However, there is a dearth of information about the accumulation of these flavonoids and their possible medicinal value. We used targeted metabolomics analysis based on ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) to analyze five developmental stages of L. maackii fruit. A total of 438 metabolites were identified in the five developmental stages, including 81 flavonoids and derivatives. The 81 flavonoids included 25 flavones and derivatives, 35 flavonols and derivatives, two isoflavones, three cyanidins and derivatives, eight procyanidins, and eight flavanones. In addition, we outlined the putative flavonoid biosynthesis pathway and screened their upstream metabolites. More importantly, we analyzed the accumulation patterns of several typical flavones and flavonols. The results reported here improved our understanding of the dynamic changes in flavonoids during fruit development and contributed to making full use of the medicinal value of L. maackii fruit.

Flavonoids are the biggest compound from the phenolic group that has the function of antioxidants. One of the plants with flavonoids is patat (Phrynium capitatum), which is usually used as food wrapping material. This study aims to quantify total flavonoid content, antioxidant activity and identify flavonoids from patat leaves ethyl acetate extract. Patat leaves sample cleaned, dried, and grinded. Grinded patat leaves were macerated gradually for 2 × 24 hours with n-hexane, ethyl acetate, and methanol solvent. The extract undergoing total flavonoids test guided with AlCl3 and CH3COONa. Extract with the biggest flavonoids content proceed to antioxidant activity test by SOD (superoxide dismutase) method and analyzed for the flavonoid structure by UHPLC MS/MS (ORBITRAP HRMS). The biggest total flavonoids are ethyl acetate extract with 8.678 mg QE/g; then methanol extract with 5.296 mg QE/g; and n-hexane extract was not tested because of negative results in the qualitative test. The antioxidant activity of the ethyl acetate extract of patat leaves is classified as inactive, with an Ic50 value of 488.299 ppm. The low antioxidant activity is due to the sample, which is still a matrix containing many compounds. The flavonoid compounds identified by its fragmentation pattern were kaempferide, formononetin, and pinostrobin.

A wide variety of plant-derived compounds, including the polyphenolic flavonoids, is present in the human diet or is consumed for medicinal reasons. Epidemiological and animal studies tend to suggest a protective effect of flavonoids against cardiovascular diseases and some types of cancer. Although flavonoids have been studied for about 50 years, the cellular mechanisms involved in their biological activity are still largely unknown. Antioxidant properties of the flavonoids have been postulated as a mechanism for putative protective effect against cardiovascular disease. Nevertheless, these properties alone are not sufficient to explain the anti-carcinogenic potential of these polyphenols. The mechanisms by which the molecules interact with cells or are absorbed by them are very important for determining the intracellular concentration and distribution of the metabolites to internal organs. With the exception of the cells lining the gastrointestinal tract, all other cells in the body are only exposed to flavonoid metabolites and degradation products. No previous studies have addressed this aspect of cellular exposure, except for some methylated metabolites. Within the last decade, reports on flavonoid activities have been largely associated with enzyme inhibition and anti-proliferative activity. From our recent work on the human colon cancer cell line HT29 and comparison with published studies, structure–function relationships demonstrate that antioxidant, enzyme inhibitor or anti-proliferative activities are dependent on particular structure motifs. The present review also presents a summary of mechanistic data on a few selected compounds.

Amaranth plants contain abundant betalains and flavonoids. Anthocyanins are important flavonoids; however, they cannot coexist in the same plant with betalains. Blue light influences metabolite synthesis and hypocotyl elongation; accordingly, analyses of its effects on betalain and flavonoid biosynthesis in Amaranthus tricolor may provide insight into the distribution of these plant pigments. We analyzed the betalain and flavonoid content and transcriptome profiles in amaranth hypocotyls under blue light and dark conditions. Furthermore, we analyzed the expression patterns of key genes related to betalains and flavonoids. Amaranth hypocotyls were shorter and redder and showed higher betalain and flavonoid content under blue light than in dark conditions. Key genes involved in the synthesis of betalains and flavonoids were upregulated under blue light. The gene encoding DELLA was also upregulated. These results suggest that blue light favors the synthesis of both betalains and flavonoids via the suppression of bioactive gibberellin and the promotion of DELLA protein accumulation, which also suppresses hypocotyl elongation. The metabolite profiles differed between plants under blue light and dark conditions. These findings improve our understanding of the environmental cues and molecular mechanisms underlying pigment variation in Amaranthus.

Stamen tea from Nelumbo nucifera Gaertn. (or the so-called sacred lotus) is widely consumed, and its flavonoids provide various human health benefits. The method used for tea preparation for consumption, namely the infusion time, may affect the levels of extractable flavonoids, ultimately affecting their biological effects. To date, there is no report on this critical information. Thus, this study aims to determine the kinetics of solid liquid extraction of flavonoid from sacred lotus stamen using the traditional method of preparing sacred lotus stamen tea. Phytochemical composition was also analyzed using high-performance liquid chromatography (HPLC). The antioxidant potential of stamen tea was also determined. The results indicated that the infusion time critically affects the concentrations of flavonoids and the antioxidant capacity of sacred lotus stamen tea, with a minimum infusion time of 5–12 min being required to release the different flavonoids from the tea. The extraction was modeled using second order kinetics. The rate of release was investigated by the glycosylation pattern, with flavonoid diglycosides, e.g., rutin and Kae-3-Rob, being released faster than flavonoid monoglycosides. The antioxidant activity was also highly correlated with flavonoid levels during infusion. Taken together, data obtained here underline that, among others, the infusion time should be considered for the experimental design of future epidemiological studies and/or clinical trials to reach the highest health benefits.

Flavonoids are plant secondary metabolites that are well known for their health-promoting properties as nutraceuticals in diets. Bioavailability and biological activities of flavonoids vary among the individual subclasses with different patterns of substitution, inclusive of glycosylation, to their basic structures. Many flavonoids exist as glycosides in plants. This study investigated the possibility of glycosylation of flavonoids through biotransformation using filamentous fungi as whole-cell biocatalysts. Microbial transformations of ten flavonoids (four flavones, four flavonols, a flavanone, and an aurone) were performed in cultures of Mucor hiemalis KCTC 26779. As a result, a flavonoid glycoside was obtained which has not been described previously. The chemical structure of this product was elucidated as 6,2′-dimethoxyflavonol-3- O-β-d-glucopyranoside by analyzing 1-dimensional and 2-dimensional-nuclear magnetic resonance spectral and high-resolution electrospray ionization mass spectral data. This compound could be useful for further biological and bioavailability studies, as well as expanding the library of flavonoid derivatives.

Avui dia millorar la productivitat, la salut i el benestar animal en els sistemes intensius de producció és bàsic per assegurar la seva sostenibilitat. A més, ens enfrontem a la reducció de l'ús d'antibiòtics pel creixement dels bacteris multirresistents. En conseqüència, és essencial investigar i desenvolupar noves alternatives juntament amb un nou enfocament del metabolisme, la salut i el benestar animal necessari per abordar aquests importants desafiaments. Els flavonoides cítrics són polifenols que posseeixen propietats antiinflamatòries, antioxidants i antimicrobianes, i han mostrat efectes prometedors en estudis previs realitzats en vedells d'engreix. D'altra banda, el creixent coneixement sobre una xarxa de comunicació entre el tracte gastrointestinal, la microbiota i el sistema nerviós central, l'eix intestí-cervell, proposa que la inflamació, la microbiota i la dieta estarien involucrades en la modulació de la conducta animal i alimentària. Així, l'objectiu d'aquesta tesi va ser estudiar els beneficis de suplementar flavonoides cítrics en vedells d'engreix alimentats amb dietes altes en concentrat sobre la productivitat, la conducta alimentària i animal, aprofundint en els possibles mecanismes d'acció relacionats amb l'eix intestí-cervell. El primer estudi va avaluar els efectes dels flavonoides cítrics sobre la conducta alimentària (menjadora uniboca), paràmetres productius, salut de la paret ruminal i comportament animal en vedells Holstein alimentats amb concentrat granulat. Els flavonoides van reduir la ingesta de concentrat i van modificar la conducta alimentària, reduint el percentatge de menjades grans. A més, es va millorar la salut de la paret ruminal i el comportament animal. En base a aquests resultats, els següents estudis van avaluar les possibles interaccions de suplementar flavonoides cítrics amb l'espai de menjadora, presentació (granulat vs. farina) i composició (nivell de greix) del pinso. També es va estudiar l'expressió de gens relacionats amb l'eix intestí-cervell, com receptors de nutrients, de neurotransmissors i diferents reguladors de la inflamació en l'epiteli ruminal i, en l'últim estudi, duodenal. En tots els estudis, els vedells suplementats amb flavonoides van dedicar més temps als comportaments alimentaris. Si bé els paràmetres productius i la ingesta de concentrat no es van veure afectats quan el concentrat es va subministrar granulat utilitzant menjadores multiespai, els flavonoides van reduir la ingesta de concentrat quan es va subministrar en farina, sense afectar el rendiment i millorant l'eficiència. En els diferents estudis, el comportament animal també va millorar, reduint comportaments orals no nutritius i les interaccions agressives i sexuals. A més, l'estudi macroscòpic de la paret ruminal realitzat en escorxador va mostrar un color més clar en els vedells suplementats amb flavonoides en els diferents estudis. Per contra, l'expressió gènica en l'epiteli ruminal va ser diferent entre estudis, sent l'expressió de gens relacionats amb la detecció de nutrients i el comportament diferent depenent de la presentació del concentrat (pellet o farina) i la seva composició (nivell de greix) en vedells suplementats amb flavonoides cítrics. L'expressió d'aquests gens en l'epiteli duodenal també va ser modificada en vedells alimentats amb dietes altes en greix. En conclusió, la suplementació amb flavonoides cítrics va modificar la conducta alimentària en els vedells, disminuint el percentatge de menjars grans o augmentant el temps dedicat comportaments alimentaris. Aquesta modulació de la conducta alimentària, juntament amb la salut de la paret ruminal podria explicar la millora del comportament en aquests vedells. A més, la suplementació amb flavonoides va modificar l'expressió de gens en l'epiteli ruminal i duodenal que podrien estar relacionats amb la conducta alimentària i animal. La presentació del concentrat (granulat vs. farina) i el nivell de greix van afectar a aquesta expressió gènica de receptors de nutrients, pèptids i hormones, i de molècules pro-inflamatòries, probablement com a resultat del metabolisme dels flavonoides en el rumen.
Hoy en día mejorar la productividad, la salud y el bienestar animal en los sistemas intensivos de producción es básico para asegurar su sostenibilidad. Además, nos enfrentamos a la reducción del uso de antibióticos por el crecimiento de bacterias multirresistentes. Consecuentemente, es esencial investigar y desarrollar nuevas alternativas y, posiblemente, un nuevo enfoque del metabolismo, salud y bienestar animal es necesario para abordar estos importantes desafíos. Los flavonoides cítricos son polifenoles que poseen propiedades antiinflamatorias, antioxidantes y antimicrobianas, y han mostrado efectos prometedores en estudios previos con terneros de cebo. Por otro lado, el creciente conocimiento sobre una red de comunicación entre tracto gastrointestinal, microbiota y sistema nervioso central, el eje intestino-cerebro, propone que inflamación, microbiota y dieta podrían estar involucradas en la modulación de la conducta animal y alimentaria. Así, el objetivo de esta tesis fue estudiar los beneficios de suplementar flavonoides cítricos en terneros de cebo alimentados con dietas altas en concentrado sobre el rendimiento productivo, la conducta alimentaria y animal, profundizando en los posibles mecanismos de acción relacionados con el eje intestino-cerebro. El primer estudio evaluó los efectos de suplementar flavonoides cítricos sobre la conducta alimentaria (comedero uniboca), parámetros productivos, salud de la pared ruminal y comportamiento animal en terneros Holstein alimentados con concentrado granulado. Los flavonoides redujeron la ingesta de concentrado y modificaron la conducta alimentaria, reduciendo el porcentaje de comidas grandes. Además, se mejoró la salud de la pared ruminal y el comportamiento animal. En base a estos resultados, los siguientes estudios evaluaron las posibles interacciones de suplementar flavonoides cítricos con el espacio de comedero, presentación (granulado vs. harina) y composición (nivel de grasa) del concentrado. Además, se estudió la expresión de genes relacionados con el eje intestino-cerebro, como receptores de nutrientes, de neurotransmisores y diferentes reguladores de la inflamación en el epitelio ruminal y, en el último estudio, duodenal. En todos los estudios, los terneros suplementados con flavonoides dedicaron más tiempo a diferentes comportamientos alimentarios. Si bien los parámetros productivos y la ingesta de concentrado no se vieron afectados cuando el concentrado se suministró en gránulos utilizando comederos multiespacio, los flavonoides redujeron la ingesta de concentrado cuando se suministró en harina, sin afectar el rendimiento y mejorando la eficiencia. En los diferentes estudios, el comportamiento animal también mejoró, reduciendo comportamientos orales no nutritivos y las interacciones agresivas y sexuales. Además, el estudio macroscópico de la pared ruminal realizado en matadero mostró un color más claro en los terneros suplementados con flavonoides cítricos en los diferentes estudios. Por el contrario, la expresión génica en el epitelio ruminal fue diferente entre estudios, siendo la expresión de genes relacionados con la detección de nutrientes y el comportamiento diferente dependiendo de la presentación del concentrado (pellet o harina) y su composición (nivel de grasa) en terneros suplementados con flavonoides cítricos. La expresión de estos genes en el epitelio duodenal también fue modificada en terneros alimentados con dietas altas en grasa. En conclusión, la suplementación con flavonoides cítricos modificó la conducta alimentaria en los terneros, disminuyendo el porcentaje de comidas grandes o aumentando el tiempo dedicado comportamientos alimentarios. Esta modulación de la conducta alimentaria, junto con la salud de la pared ruminal podría explicar la mejora del comportamiento en estos terneros. Además, la suplementación con flavonoides modificó la expresión de genes en el epitelio ruminal y duodenal que podrían estar relacionados con la conducta alimentaria y animal. La presentación del concentrado (granulado vs. harina) y el nivel de grasa afectaron a esta expresión génica de receptores de nutrientes, péptidos y hormonas, y de moléculas pro-inflamatorias, probablemente como resultado del metabolismo de los flavonoides en el rumen.
Nowadays, improving productivity, health and animal welfare in livestock reared under intensive conditions is mandatory for the sustainability of our production systems. Furthermore, reducing the use of antibiotics due to the constant growth of resistant bacteria is a critical issue to face. Consequently, the research and development of new proposals beyond traditional solutions applied are essential, and probably also a new approach of animal metabolism, health and well-being is needed to tackle all these important challenges. Citrus flavonoids are polyphenols that possess interesting anti-inflammatory, antioxidant, and antimicrobial properties and have showed promising effects in previous research on beef cattle. Otherwise, recently the knowledge of a communication network between gastrointestinal tract, microbiota, and the central nervous system, the gut-brain axis, has increased. Thus, inflammation, microbiota, and diet have been proposed to be involved in animal behavior and eating pattern modulation. Therefore, the main objective of this thesis was to study the possible benefits of supplementing citrus flavonoids in bulls fattened under intensive conditions and fed highconcentrate diets on performance and productivity, but also their possible effects on eating and animal behavior and to study the possible mode of action related with the gutbrain axis. Thus, a first study evaluated the effects of citrus flavonoids supplementation on eating pattern (using a single-space feeder), performance parameters, rumen wall health, and animal behavior in Holstein bulls fed high-concentrate diets in pellet form. Citrus flavonoids reduced feed intake and modified eating pattern, reducing the percentage of large meal sizes. Additionally, rumen wall health and animal behavior were improved. Based on these results, the following studies evaluated the possible interactions of citrus flavonoids supplementation with feeder space, concentrate presentation (pellet vs. meal) and composition (fat level). Furthermore, the expression of genes involved in the gutbrain axis crosstalk, such as nutrient sensing receptors, some neurotransmitters receptors and different inflammation regulators were studied in the epithelium of rumen and, in the last study, of duodenum epithelium as well. When bulls were supplemented with citrus flavonoids devoted more time to perform feeding behaviors throughout the different studies. Whilst performance parameters and concentrate intake were not affected when concentrate was fed in pellet form using multi-space feeders, concentrate intake was reduced when citrus flavonoids were supplemented in a concentrate fed in meal form without impairing performance, so improving efficiency. In all studies, animal behavior was also improved, by reducing oral non-nutritive behaviors, and aggressive and sexual interactions. Moreover, in all studies, the macroscopical rumen wall study performed at the slaughterhouse showed lighter color for bulls supplemented with citrus flavonoids. Conversely, ruminal gene expression differed among the studies, so the expression of genes involved in nutrient sensing and behavior in bulls supplemented with citrus flavonoids were different depending on the concentrate presentation (pellet or meal), and composition (fat level inclusion). The expression of these genes in the duodenum epithelium when bulls were fed high-fat diets was also modified. In conclusion, citrus flavonoids supplementation modified eating pattern in Holstein bulls, decreasing the percentage of large meal sizes or increasing time devote to feeding events. This modulation of the eating pattern, together with an improvement in rumen wall might be involved in the improvement of animal behavior of bulls supplemented with citrus flavonoids. Moreover, flavonoid supplementation differently modified the expression of genes in the rumen and duodenum epithelium that could be related with eating pattern and animal behavior regulation, although concentrate presentation (pellet vs. meal) and fat level might affect these effects on gene expression of different nutrient sensing, peptides and hormones receptors, along with some proinflammatory molecules, probably as result of the effect of rumen fermentation on flavonoid metabolism.

The Wnt signaling system is an intricate network of cellular communication pathway that is crucial for tissue homeostasis, development, and disease. It is named after a family of secreted proteins called Wnts, which binds to the receptor on the cellular surface to activate intracellular signaling cascades. This system has vital role in growth related conditions and tumours, particularly as a main factor in proliferation and spread of colorectal cancer (CRC), which is correlated with the build-up of β-catenin in cells due to glycogen synthase kinase-3β (GSK-3β) inactivation. Recently, several polyphenolic substances from the naturally occurring flavonoid family were examined for their potential to inhibit Wnt signaling and found that they exhibit potent anti-oxidant and anti-carcinogenic activities. This thesis identifies the mechanism by which the flavonoids like silibinin, eriodictyol and quercetin attaches to β-catenin and Wnt protein, determining more effective natural inhibitor of the disease-causing protein. For clarify the binding efficiency of flavonoids, a comparative molecular docking analysis was carried out and their interaction with the respective protein molecule is done in this study

Recently, there has been an increasing interest in health-promoting products which are also natural and safe for consumption because the consumer market has been searching for a healthy lifestyle. This global market trend has driven the food industry to invest in developing innovative products containing bioactive components. Flavonoids are a group of phenolic compounds of low molecular weight, consisting of 15 carbon atoms. Their alterations in the heterocyclic ring’s substitution pattern generate six subclasses: flavonols, flavanols, flavones, flavanones, isoflavones and anthocyanins. Also, different studies have reported that diets rich in flavonoids provide numerous benefits associated with health-promoting effects by reducing the risk of development of chronic diseases such as cardiovascular diseases, diabetes type II and some types of cancers. These effects have been related to their biological properties which also include other activities such as colorant effects (e.g., anthocyanins), transforming them into potential food additives with desirable capacities. Therefore, this review aims to revise the classes of flavonoids and their main biological properties as well as the most used extraction techniques applied for obtaining these compounds, their bioavailability and the application to formulate new natural food additives.

Fruit constitutes a major component of our diet, providing fiber, vitamins, minerals, and many other phytonutrients that promote good health. Fleshy fruits, such as tomatoes, already contain high levels of several of these ingredients. Nevertheless, efforts have been invested in increasing and diversifying the content of phytonutrients, such as carotenoids and flavonoids, in tomato fruits. Increasing levels of phytonutrients, such as lycopene, is highly justified from the perspective of the lycopene extraction industry due to cost effectiveness reasons. Diversifying phytonutrients, in particular those that contribute to fruit color, could potentially provide an array of attractive colors to our diet. Our major goal was to devise a novel strategy for developing tomato fruits with enhanced levels of phytochemicals known to promote good health with special emphasis on lycopene content. A further important goal was to analyze global gene expression of selected genetic lines produced throughout this study in order is to dissect the molecular mechanisms regulating phytonutrients accumulation in the tomato fruit. To achieve these goals we proposed to: 1. combine, by classical breeding, engineered polyamine metabolism with photomorphogenic high pigment mutants in order generate tomato plant with exceptionally high levels of phytonutrients; 2. use gene transfer technology for genetic introduction of key genes that promote phytonutrient accumulation in the tomato fruit, 3. Analyze accumulation patterns of the phytonutrients in the tomato fruit during ripening; 4. Analyze global gene expression during fruit ripening in selected genotypes identified in objectives 1 and 2, and 5. Identify and analyze regulatory mechanisms of chloroplast disassembly and chromoplast formation. During the 3 years research period we have carried out most of the research activities laid out in the original proposal and our key conclusions are as follows: 1. the engineered polyamine metabolism strategy proposed by the US collaborators can not increase lycopene content either on its own or in combination with an hp mutant (hp-2ᵈᵍ); 2. The hp-2ᵈᵍ affects strongly the transcriptional profile of the tomato fruit showing a strong tendency for up- rather than down-regulation of genes, 3. Ontology assignment of these miss-regulated genes revealed a consistent up-regulation of genes related to chloroplast biogenesis and photosynthesis in hp-2ᵈᵍ mutants throughout fruit development; 4. A tendency for up-regulation was also usually observed in structural genes involved in phytonutrientbiosynthesis; however this up-regulation was not as consistent. 5. Microscopic observations revealed a significantly higher number of chloroplasts in pericarp cells of mature-green hp-2ᵈᵍ/hp-2ᵈᵍ fruits in comparison to their normal fully isogenic counterparts. 6. The relative abundance of chloroplasts could be observed from early stages of fruit development. Cumulatively these results suggest that: 1. the overproduction of secondary metabolites, characterizing hp-2ᵈᵍ/hp-2ᵈᵍ fruits, is more due to chloroplast number rather then to transcriptional activation of structural genes of the relevant metabolic pathways, and 2. The molecular trigger increasing metabolite levels in hp-2ᵈᵍ mutant fruits should be traced at early stage of fruit development.

This project was based on the hypothesis that nematodes interacting with plants as either parasites or beneficial saprophytes are attracted to their host by natural products. This concept was supported by numerous observations that parasitic nematodes are attracted to root exudates. Our overall goal was to identify nematode sensory compounds from root exudates and to use that information for reducing nematicide applications. We applied skills of the investigators to achieve three specific objectives: 1) Identify nematode behavioral cues (e.g., attractants or repellents) in root exudates; 2) Identify new natural nematicidal compounds; and 3) Combine a natural attractant and a nematicide into a nematode trap. Because saprophytic nematodes benefit plants by mineralizing organic matter, we sought compounds attractive primarily to parasitic nematodes. The project was constructed on several complementary foundations. First, data from Dr. Spiegel’s lab showed that under aseptic conditions Ditylenchus dipsaci, a parasite on onion, is attracted to certain fractions of onion root exudates. Second, PI Phillips had a sizeable collection of natural plant products he had identified from previous work on Rhizobium-legume interactions, which could be tested “off the shelf”. Third, Dr. Ferris had access to aseptic and natural populations of various saprophytic and parasitic nematodes. The project focused on five nematode species: D.dipsaci, Heterodera avenae, and Tylenchulussemipenetransat ARO, and Meloidogyne javanicand Caenorhabditis elegans at UCD. Ten pure plant compounds, mostly flavonoids, were tested on the various nematode species using six different assay systems. Results obtained with assorted test systems and by various scientists in the same test systems were essentially irreproducible. Many convincing, Many convincing, i.e. statistically significant, results in one system or with one investigator could not be repeated with other assays or different people. A recent report from others found that these compounds, plus another 30, were inactive as attractants in three additional parasitic nematode species (Wuyts et al. Nematology 8:89- 101, 2006). Assays designed to test the hypothesis that several compounds together are required to attract nematodes have thus far failed to find a reproducibly active combination. In contrast to results using pure plant compounds, complex unfractionated exudates from aseptic onion root reproducibly attracted D. dipsaci in both the ARO and UCD labs. Onion root exudate collection, separation into HPLC fractions, assays using D. dipsaci and MS-MS experiments proceeded collaboratively between ARO and UCD without any definitive identification of an active compound. The final active fraction contained two major molecules and traces of several other compounds. In the end, analytical studies were limited by the amount of onion root exudate and the complexity of the purification process. These tests showed that aseptic plant roots release attractant molecules, but whether nematodes influence that release, as insects trigger release of attractants from plants, is unknown. Related experiments showed that the saprophyte C. elegans stimulates its prey, Pseudomonas bacteria, to increase production of 2, 4-diacetylphloroglucinol (DAPG) a compound that promotes amino acid exudation by plant roots. It is thus possible that saprophytic nematodes are attracted primarily to their bacterial or fungal prey and secondarily to effects of those microorganisms on root exudation. These observations offer promising avenues for understanding root-zone interactions, but no direct routes to controlling nematodes in agriculture were evident. Extracts from two plant sources, Chrysanthemum coronarium and Sequoia sempervirens, showed nematicidal activity at ARO and UCD, respectively. Attempts to purify an active compound from S. sempervirens failed, but preliminary results from C. coronarium are judged to form a potential basis for further work at ARO. These results highlight the problems of studying complex movement patterns in sentient organisms like nematodes and the issues associated with natural product isolation from complex mixtures. Those two difficulties combined with complications now associated with obtaining US visas, slowed and ultimately limited progress on this project. As a result, US investigators expended only 65% of the $207,400 originally planned for this project. The Israeli side of the project advanced more directly toward its scientific goals and lists its expenditures in the customary financial report.