Title Promoter Affiliations Abstract "Influence of product and process factors on the bioaccessibility and bioavailability of carotenoids in plant-based food systems" "Marc Hendrickx" "Food and Microbial Technology (CLMT)" "Fruits and vegetables are part of a healthy diet as they are important in maintaining good health as well as to reduce coronary heart diseases, strokes and certain types of cancer. Carotenoids are lipophilic micronutrients present in all fruits and vegetables. They are located in plastids of the plant cell, where they can be associated with other components. Due to their high level of bio-encapsulation and low solubility in an aqueous phase, absorption by the human body is rather low. In previous research, carotenoid bioaccessibility appeared to be the highest in food systems where natural barriers were destroyed through processing, to which lipids were added and whereby the carotenoids were transferred to the lipid fraction added prior to digestion. In this context, emulsification of a carotenoid-enriched oil phase could be explored as an adequate delivery system for carotenoids. Due to their lipophilic nature, the absorption pathway of carotenoids is strongly linked to the one of lipid digestion. However, it can be questioned to what extent emulsion characteristics play a role in lipid digestibility and how this can influence carotenoid bioaccessibility.Therefore, this doctoral project focused on exploring the effect of specific emulsion characteristics on lipid digestibility, micelle formation and carotenoid bioaccessibility. For this, an in vitro digestion approach was proposed that enables the study of digestion time (kinetic approach). From an analytical perspective, multiple lipid digestion species were evaluated for the first time, to obtain detailed insight in the lipid digestion process. Furthermore, this study aimed to link the evolution of lipid digestion and carotenoid bioaccessibility to (micro)structural changes occurring along the digestive tract.In this context, the effect of (i) the unsaturation degree of the emulsified oil phase, (ii) the gastrointestinal stability of emulsions stabilized by small molecule surfactants and (iii) the presence, location and structure of fiber on lipid digestion, mixed micelle formation and carotenoid bioaccessibility was investigated. Hereto, oil-in-water emulsions were formulated with 5% (w/v) carotenoid-enriched oil and stabilized by one or two emulsifiers (0-1.5% w/v), such as Tween 80, sucrose esters, citrus pectin or phosphatidylcholine. All emulsions studied, presented small initial oil droplet sizes (smaller than 2 µm). The emulsions were submitted to in vitro digestion during which the small intestinal phase was simulated kinetically. In this way, the time dependency of lipolysis, micelle formation and carotenoid bioaccessibility could be evaluated, allowing to gain more detailed insight in the mechanism behind these processes.Different emulsion characteristics led to different reaction rate constants as well as different final extents of triacylglycerol hydrolysis and carotenoid micellarization. For example, oils rich in polyunsaturated fatty acids (soybean or linseed oil) were digested slower and sometimes to a lesser extent in comparison to olive oil rich in monounsaturated fatty acids. This can be partially attributed to the more bended structure of polyunsaturated fatty acids, which might retard and/or hinder lipase hydrolysis. This doctoral project showed that not only the oil droplet size of the initial emulsion is of importance, but also the gastrointestinal stability of these oil droplets. More specifically, emulsions unstable in the gastric phase presented a larger particle size at the beginning of the small intestine, resulting in a slower and eventually incomplete lipolysis. The work regarding pectin was divided into two parts. First, the emulsifying properties of citrus pectin with different degree of methylesterification were evaluated, after which the effect of the presence of pectin in emulsions was studied on the gastrointestinal stability of oil droplets as well as on the kinetics of lipolysis and carotenoid bioaccessibility. It was quantitatively shown that citrus pectin is capable of reducing the interfacial tension of an oil droplet regardless its structure, evidencing its adsorption at the oil-water interfaces and surface-active properties. The results of the digestion study showed that the pectin structure influenced the lipolysis rate, while the pectin location in the emulsion had a large impact on lipolysis extent. In this sense, the presence of pectin at the oil-water interface led to a fast lipolysis, but low extent. This suggests that pectin does not hinder lipase adsorption, yet it may interact with other components within the intestinal juices, such as bile salts, inhibiting a complete lipid digestion. In all digestion studies performed, a strong interrelation was observed between triacylglycerol hydrolysis, micelle formation and carotenoid bioaccessibility.The data obtained in the digestion studies discussed above were first modelled using a single response empirical model (fractional conversion model). In this single response approach, all lipid digestion species evaluated were modelled independently. However, they are part of a common reaction scheme and are interrelated. Therefore, in the last part of this doctoral project, it was aimed to build a mechanism-based multiresponse model. Using the iterative process of multiresponse kinetic modelling and the data obtained previously in this work, it was possible to propose a generally applicable reaction scheme of lipid digestion, which could be translated to a set of corresponding differential equations describing the evolution of all evaluated lipolysis species together. This multiresponse model is expected to have a higher reliability and applicability compared to the selected single response empirical model applied before. The mechanistic multiresponse kinetic model proposed, was successful to describe all datasets obtained in this work.From this work, it can be concluded that emulsion design can be of great interest to tailor lipid digestion and carotenoid bioaccessibility. Moreover, if one wants to manipulate carotenoid bioaccessibility, one can focus on influencing lipolysis in case carotenoids are incorporated in an emulsified lipid fraction prior to digestion." "Use of Microorganisms for Carotenoids Delivery: Next Generation of Probiotics for Cardiovascular Disease" "Lynn Vanhaecke" "Department of Veterinary Public Health and Food Safety, Institut National de la Recherche Agronomique, University of Naples Federico II, Organic Waste Systems (Belgium), BioActor (Netherlands), French National Institute for Agricultural Research, Biopolis (Spain), ProDigest (Belgium)" "CARODEL aims to valorise the result from the previous FP7 COLORSPORE, project, in which initial isolation and characterization work was performed on Bacillus strains producing gastric-stable carotenolds. As the stability in the gastrointestinal Iract (GIT), antioxidant activity and biosavailability of particular Bacillus carotenoids was shown to be higher than those of common dietary carotenoids, the conclusions from COLORSPORE provided strong and compelling reasons to suppor further development and commercialisation of these bacteria-derived carotenoids" "Honest signaling in birds: Can carotenoids promote steroidogenesis?" "Marcel Eens" "Behavioural Ecology & Ecophysiology" "The general aim of the proposal is to test if the supplementation of carotenoids and/or cholesterol can promote the synthesis of T that regulate the uptake of carotenoids in the cells of the trait. This is in line with the finding that carotenoids can regulate the steroidogenic activity in humans (Wickenheisser et al. 2005; Angwafor and Anderson 2008) and other mammals (Folman et al. 1983; Chew 1993; Chew et al. 2001; Campbell et al. 2006). Indirect evidence shows that providing zebra finches Taeniopygiaguttata with carotenoids enhances their escape flight ability (Blount and Matheson 2006), suggesting that carotenoids may stimulate the production of androgen with anabolic effects on the flight muscles. Similar evidence comes from a recent discovery that European starlings Sturnus vulgaris supplemented with carotenoids increase their song rate and that starlings supplemented with both carotenoids and T show an enhanced nest-oriented song compared to birds treated just with T (VanHout, Eens and Pinxten 2011). Moreover providing cholesterol to zebra finches enhances bill carotenoids (McGraw and Parker 2006) suggesting a promotion of T by cholesterol is the first step of the complex steroidogenic process." "Maternal investment in yolk hormones and carotenoids: sources of variation and effects in wild rockhopper penguins." "Maud Poisbleau" "Behavioural Ecology & Ecophysiology" "Mothers can influence the phenotype and performance of their offspring by adjusting their deposition of egg components to environmental conditions and according to their own physiological state. Birds, and rockhopper penguins (Eudyptes chrysocome) in particular, have excellent characteristics which can be used to study the sources of variation in maternal investment in eggs and their potential effects on embryonic survival and development. Rockhopper penguins exhibit a reversed egg size dimorphism, brood reduction, a unique reversed hatching asynchrony, and large differences in breeding success among years and populations. Therefore, I can study variation in maternal investment on all possible levels: within the population, within clutches and among seasons and populations. I will first explore the sources of variation in investment in eggs (in terms of layingdate, mass, and levels of androgens, carotenoids and antibodies) according to female quality (e.g. arrival and laying dates and weights, blood parameters). Various approaches will be used to examine the contribution of male quality. Effects on embryo survival and development will be studied mainly via experimental manipulation of androgen levels. Finally, my long-term study and collaboration with groups working on other rockhopper populations will enable assessment of inter-annual and populational variation in maternal investment." "Carotenoid bioaccessibility in fruit and vegetable based systems" "Marc Hendrickx" "Centre for Food and Microbial Technology" "Carotenoids are important micronutrients that have been linked with several health benefits. To exert the health benefits, carotenoids must be absorbed and reach their site of action. The absorption process of carotenoids consists of various steps including release from the food matrix, solubilization into a lipid phase, incorporation into micelles (i.e. bioaccessibility) and uptake through the intestine epithelia. However, previous investigations have proven that the transfer of carotenoids into the oil phase during digestion that is a prerequisite for their bioaccessibility is limited by matrix related factors that hamper carotenoid release and their transfer into oil. Consequently, the fraction of carotenoids that is actually absorbed, and serves health related functions (i.e. bioavailability), was also proven to be generally low. In this context, high pressure homogenization (HPH) and thermal processing in the presence of oil were investigated as alternative strategies that might improve carotenoid transfer from fruit and vegetable matrices to oil phase and potentially enhance their bioaccessibility.Hereto, the effect of HPH (10, 30, 50, 70, 100 MPa) and thermal processing (80-120 °C, 20 min; 95-110 °C, 0-40 min) on the transfer of carotenoids (lycopene, α-carotene and β-carotene) from tomato and carrot based matrices to the oil phase was investigated. The effect of carotenoid bioencapsulation in determining carotenoid release from the matrix and subsequent transfer into the oil phase was investigated by decomposing tomato and carrot matrices into a chromoplast and a cell cluster fraction. Subsequently, the effect of storage conditions on the stability of carotenoids and lipids in shelf-stable tomato and carrot-based systems was investigated. Finally, the kinetics of the micellar incorporation of lycopene and its cis isomers, β-carotene and its cis isomers as well as α-carotene and lipid digestion products, free fatty acids (FFAs) and monoglycerides (MAGs) during in vitro digestion of low-fat oil in water emulsions was investigated. Moreover, the relation between carotenoid bioaccessibility and lipid digestion was elucidated.During HPH, carotenoid transfer to oil was clearly increased by increasing homogenization intensity. This is the result of the increased shearing and contact between the matrix and the oil phase, which could facilitate the transfer of carotenoids from the matrix into oil. During thermal processing treatment conditions of >100 °C for 10 min were necessary to significantly favour carotenoid transfer to oil (≥75%). HPH at ≥ 50 MPa could achieve a similar carotenoid transfer efficiency in the chromoplast fractions. The cell wall represented a limiting factor for carotenoid transfer during processing. This suggests that in order to achieve maximum carotenoid transfer to oil, the food systems should be disrupted to a level where the cell wall is broken. The selective transfer of a particular carotenoid during processing depended on its chemical structure (e.g. transfer of lycopene" "A new methodological approach to understand how structural barriers govern carotenoid bioaccessibility in plant based foods." "Marc Hendrickx" "Centre for Food and Microbial Technology" "The general objective of the present research project is to understand how structural barriers govern carotenoid release and bioaccessibility in fruit and vegetable based food systems. Because research intended to unravel the underlying mechanisms that determine carotenoid bioaccessibility is hampered by using real food products, various plant dispersion model systems with well-characterised properties will be generated based on different fractions isolated from tomato and carrot tissue. The plant dispersion model systems will be build up by a (tomato or carrot) serum phase and a (tomato or carrot) particle phase containing the carotenoids. Structural barriers in both the serum and particle phase will be focus points. The general objective can be translated into two specific hypotheses: - The structural characteristics of pectin present in the serum phase play an important role in determining carotenoid bioaccessibility in fruit and vegetable based food products. - The localisation of carotenoids in plant based food products (i.e. the level of bio-encapsulation) determines to a large extent their bioaccessibility in fruit and vegetable based food products." "The effect of structural build-up of plant-based food emulsions on carotenoid bio-accessibility and in vitro digestion of fat." "Marc Hendrickx" "Centre for Food and Microbial Technology" "Vegetable-based foods such as soups and sauces, are good sources of water, fibers (e.g. pectin), micronutrients and lipids. In foods, lipids are often considered as negative since an excess uptake of lipids is linked with a risk of obesity. It should however be noted that lipids are important macronutrients providing energy and essential fatty acids and facilitating the uptake of lipid soluble micronutrients such as carotenoids.The general objective of this PhD was to gain more insight in the relation between in vitro β-carotene bioaccessibility and lipid digestion and to understand the specific role of the structural build-up of plant-based emulsions in β-carotene bioaccessibility and lipid digestion. To reach these objectives, all experimental chapters studied the influence of different emulsifiers in an oil-in-water emulsion containing 5% β-carotene enriched olive oil (no natural barriers such as chromoplast membrane and/or cell wall). The emulsifiers used were citrus pectin (CP) (added in a concentration of 1% or 2%, with a degree of methyl-esterification (DM) of 14%, 30%, 57%, 66% or 99%), sugar beet pectin (added in a concentration of 1% with a DM of 32% or 58%), L-α-phosphatidylcholine (added in different concentrations: 1%, 2%, 3% or 4%), or a combination of citrus pectin and phosphatidylcholine.Based on the results, no general relation was found between the in vitro β-carotene bioaccessibility and lipid digestion. It seems that the type, the amount and the surface activity of the present emulsifiers are determining which compounds can be incorporated into the micelles. The DM and concentration of citrus pectin were influencing the micellar incorporation of both β-carotene and lipids, whereas the DM of sugar beet pectin was not. Probably, in this type of pectin, the other pectin properties are more important than the DM. By increasing the phosphatidylcholine concentration, the lipid micellar incorporation remained whereas the β-carotene bioaccessibility increased. This lead to the idea that phosphatidylcholine may dissolve β-carotene so that an increase of phosphatidylcholine concentration in the micelles leads to an increase of β-carotene bioaccessibility. When phosphatidylcholine was present together with citrus pectin as emulsifier, the β-carotene bioaccessibility was dependent on the pectin DM whereas the lipid digestion remained again. Since carotenoids and lipids have different structures and polarities, their incorporation may be different, which can be used to engineer targeted (digestive) functionalities in food products. Sugar beet pectin can be chosen as emulsifier if high lipid digestion and reasonable β-carotene bioaccessibility are desired, whereas 4% phosphatidylcholine can be chosen if high β-carotene bioaccessibility and lower lipid digestion are desired. The lowest lipid digestion was found in the emulsions containing pectin with a high DM (2%CP99) or with a low DM (2%CP14) as emulsifier." "SLEEP IN THE CITY: How does artificial light at night affect EEG-based measures of sleep (CitySleep)?" "Marcel Eens" "Behavioural Ecology & Ecophysiology" "Sleep is an adaptive state of inactivity, which plays critical functions including replenishing energy and neurological recovery. Artificial light at night (ALAN) is ubiquitous in the urbanizing world, and has the potential to substantially alter sleep patterns. Our understanding of how ALAN affects sleep in wild animals is seriously limited, because past studies have relied on behavioural metrics of sleep, which cannot distinguish different types of sleep (rapid eye movement (REM) versus slow wave sleep (SWS)), or accurately quantify sleep intensity (amount of slow waves within SWS). In this study (CitySleep), the Experienced Researcher (ER) will use state-of-the-art neurologgers to obtain electroencephalogram (EEG) data on sleep in wild great tits (Parus major) exposed to ALAN. She will obtain data from free-living nestlings and from adults in semi-natural aviaries. Great tits sleep in nest boxes that can be experimentally exposed to ALAN, and have served as a model species in behavioural sleep studies. The ER will work with the University of Antwerp's Behavioural Ecology and Ecophysiology (BECO) Group, which has developed methods to manipulate ALAN inside boxes, and has high-quality publications on how ALAN affects sleep behaviour. She will receive expert training on implanting neurologgers from Prof. A. Vyssotski (University of Zurich), and training on interpreting EEG data from Dr. N. Rattenborg's Avian Sleep Research Group (Max Planck Institute). She will contribute expert knowledge of urban ecology, stress physiology and bird handling, and introduce neurologgers to the BECO Group, facilitating a major advance in research methodology. Results will be disseminated through top-tier publications, international conferences and public engagement, and used to advance scientific knowledge and motivate environmental policy changes. The ER will gain skills that will propel her research to a higher level and allow her to secure a permanent research position." "Integrative and experimental study of the effects of artificial light exposure at night during development in birds in the real world: merging mechanistic approaches with short- and long-term health and fitness consequences." "Marcel Eens" "Behavioural Ecology & Ecophysiology" "Artificial light at night (ALAN) has been underexplored as an anthropogenic stressor compared to chemical and noise pollution and it can be considered a threat to biodiversity. Because organisms have evolved with the periodicity of light dark cycles, ALAN may affect multiple aspects of behaviour and physiology. Changes in behaviour and physiology associated with ALAN may have particularly strong effects early in life, when developmental trajectories are sensitive to stressful conditions. Yet, a major gap in knowledge involves the effects of ALAN during development, particularly in natural populations. Using birds as a model, we will perform the first study to comprehensively examine how exposure to ALAN affects physiological stress in nestlings in the short- and long-term. We will examine whether early life light exposure affects aging rate, as indicated by telomere dynamics. Finally, we will assess whether exposure to ALAN early in life has enduring effects on the phenotype and fitness (reproduction, survival) of adults. We will use experimental approaches, involving manipulations of light inside nestboxes and cross-fostering experiments. We will also collect correlational data, using a population exposed to heterogeneous light regimes. By elucidating effects of ALAN on developing organisms and assessing mitigating strategies (such as part-night lighting), our study will motivate action to cope with the consequences of living in lighted environments." "Metal Pollution and Oxidative Stress: Exploring effects on aging rate, behavior and fitness." "Marcel Eens" "Behavioural Ecology & Ecophysiology" "Toxic pollutants increasingly threaten the integrity of natural populations. Metal pollution can have particularly detrimental effects on organisms and entire ecosystems. One potent means by which metal pollution may affect organisms is by elevating oxidative stress, resulting in biomolecular damage and fitness declines. However, the long-term behavioral and fitness implications of metal-induced oxidative stress are poorly understood. Using great tits (Parus major) as a model species, I propose to comprehensively explore how metal pollution affects oxidative stress, aging rate, behavior and fitness. I will perform the first study to examine whether metal-induced oxidative stress affects aging rate, on the molecular level as indicated by telomere degradation, and in terms of sexual signals and fitness traits. I will also explore effects of metal pollution on risk-taking behavior, which may be altered via effects of oxidative stress on neural function and life history decisions. I will study nest box populations of great-tits located across a metal pollution gradient. Experimental approaches will include exposing nestlings to metals and antioxidants, a parental risk-taking experiment, and measuring exploratory behavior in the laboratory. This study will make a pioneering contribution to evolutionary biology by testing the oxidative stress theory of aging in the context of metal pollution, and have critical importance to ecotoxicology and conservation biology."