Narcissus is an economically important crop with a large number of species, hybrids and/or varieties in cultivation. It is one of the most popular ornamental plants. The Netherlands is among the... Show moreNarcissus is an economically important crop with a large number of species, hybrids and/or varieties in cultivation. It is one of the most popular ornamental plants. The Netherlands is among the top producers of Narcissus in the world. The ornamental value of Narcissus is quite well known but they can also be a valuable resource of natural compounds. Among these compounds, galanthamine has been successfully commercialized as medicine for the treatment of early to mild Alzheimer’s Disease (AD). The characteristic symptoms of this disease are loss of memory and impairment of multiple cognitive and emotional functions. The objective was to find out how galanthamine levels could be enhanced to convert this floricultural crop into a medicinal crop. There are different factors that influence the production of galanthamine and other metabolites in the bulbs. Effect of various factors such as preanalytical methods, bulb location and age along with effect of biotic and abiotic factors on the growth and yield were studied. From the results, it is evident that factors have detrimental effect on the total bulb yield and galanthamine levels. Show less
Plant exudates appeared long time ago in the history line of life in the early Devonian. They are typical saps produced by specialized cells. They can be naturally excreted outside or right after... Show morePlant exudates appeared long time ago in the history line of life in the early Devonian. They are typical saps produced by specialized cells. They can be naturally excreted outside or right after plants are damaged or infected. Through time, diverse cultures recognized them as valuable sources of natural medicines. Of various types of plant exudates, the most outstanding are gums, mucilage, essential oils, oleoresins, resins, phloem like saps, and latexes. Regardless of their commercial importance, in the aspect of biology, plant exudates are believed to be involved in plant defense mechanisms, but their roles still remain unclear. In this thesis, based on diverse experiments, many features of plant exudates were unveiled. Interestingly, even if plant exudates possess a broad chemical diversity, their specialized biological functions resulted in less variation of their metabolomes as compared with their bearing tissues. This lower variation indicates that plant exudates might partake of a general-broad defense against herbivores and pathogens in their bearing tissues. This defense involves both mechanical and chemical traits which might complement and modulate each other, eventually resulting in a chemo-mechanical defense layer. Of the diverse chemicals in plant exudates, terpenes are one of the most common metabolites in these saps. Show less
The major and ultimate aim of metabolomics is to obtain an answer to a specific biological or clinical question. For that, many strategies have been applied in this field, including NMR and MS... Show moreThe major and ultimate aim of metabolomics is to obtain an answer to a specific biological or clinical question. For that, many strategies have been applied in this field, including NMR and MS-based approaches. CE-MS is one of the techniques in this field but remains underappreciated because of issues such as compromised sensitivity and poor reproducibility. In this thesis, we proposed standardized protocols for CE-MS studies using a sheathless interface and applied them in the metabolic analysis of ultra-small biological samples, such as low numbers of HepG2 cells. Another CE-MS technique used in this thesis is the traditional sheath liquid CE-MS, and it was successfully used in the metabolic profiling of plasma samples. We also introduced CE-MS with both interfaces in the field of nanosciences with our efforts in revealing the adsorption characteristics of polar metabolites to the surface of nanomaterials. This work clearly demonstrates that CE-MS can be reliably performed for metabolomics studies with acceptable repeatability. The use of sheathless CE-MS interface enables us to have in-depth profiling for ultra-small samples, and may become a powerful tool in the emerging field of single-cell analysis. Show less
We showed that constitutive and inducible chemical and morphological defenses against Western flower thrips differ between tomato and chrysanthemum plants. Furthermore, we demonstrated that both... Show moreWe showed that constitutive and inducible chemical and morphological defenses against Western flower thrips differ between tomato and chrysanthemum plants. Furthermore, we demonstrated that both plant species respond differently to bacteria-derived defense elicitors, such as the phytotoxin coronatine. This highlights the plant species-specificity of these interactions and the possible limitation for the use of pathogen-associated molecular patterns to enhance the plant immune system. This study thus provides knowledge and novel strategies for Western flower thrips control. Yet, further comprehensive work is needed to evaluate the influence of these induction strategies on plant fitness. Show less
The power of personalized nutrition lies in being able to conduct clinical research on healthy people while capturing metabolic markers sensitive to the impact of environmental and metabolic... Show moreThe power of personalized nutrition lies in being able to conduct clinical research on healthy people while capturing metabolic markers sensitive to the impact of environmental and metabolic stressors (e.g. diet, changing sex hormones and the menstrual cycle). Using clinical biomarkers, metabolomics, and diet interventions with intake analyses, we demonstrated the metabolic impact of vegan and animal diet interventions using fasting plasma analysis after 48 hours and using postprandial plasma analysis after meals and snacks. Sexually dimorphic responses were differentiated using proteomics and pathway analyses in two larger, sex-balanced cohorts. Finally, clinical biomarker and metabolomics analyses identified metabolic subtypes across menstrual cycle phases. Although challenges with integrating –omics technology and nutrition remain, the fundamental information generated from these research studies may provide a foundation for future novel personalized nutrition strategies. Show less
Climate change is a challenge for both current and future generations. New biological resources have to be developed in order to meet the demand for energy as well as the demand for food. One way... Show moreClimate change is a challenge for both current and future generations. New biological resources have to be developed in order to meet the demand for energy as well as the demand for food. One way of doing this is to make use of so-called smart crops, as they have improved yields and a small environmental footprint. Model plant organisms, such as Arabidopsis thaliana, can contribute to the development of smart crops as they have been studied extensively by the research community. The purpose of this research is to unravel the pathways leading to the enhanced growth characteristics phenotype of two phenotypically engineered Arabidopsis mutants from a systems biology perspective, by using metabolomics. In metabolomics, metabolites are studied both qualitatively and quantitively under specific growth conditions. Since metabolites are the end products of cellular processes, the metabolome is most closely related to the phenotype of plants. To obtain the metabolic profile directly from the leaves of Arabidopsis, high-resolution magic angle spinning (HR-MAS) NMR is used. Combined with multivariate analysis, biomarkers, related to the mutant phenotype, are determined. The biological interpretation of the biomarkers will be performed in a systems biology approach to obtain a model to explain the mutant phenotype. Show less
This thesis provides insights into the mechanisms of renal I/R injury based on human kidney transplantation (i.e. the status of delayed graft function: DGF). A severe energetic crisis... Show moreThis thesis provides insights into the mechanisms of renal I/R injury based on human kidney transplantation (i.e. the status of delayed graft function: DGF). A severe energetic crisis differentiates DGF kidneys from adequately functioning controls. Although intact beta-oxidation, aerobic glycolysis and glutaminolysis provide Krebs Cycle intermediates, these intermediates are not able to enter the mitochondrial Krebs cycle. Hence, dysfunctional mitochondria disable efficient ATP production leading to the metabolic incompetence that causes DGF and underlies renal I/R injury. This finding sheds a whole new light on I/R injury and explains why ATP-dependent therapeutics are ineffective as treatment for I/R injury. A major difference in the vulnerability of mitochondria to ischemia and reperfusion between rodents and humans was found. This could explain the current differences in effectiveness of therapies in the experimental versus the clinical setting. Big cohort studies give insights in donor, recipient and transplant-procedure variables and challenge the reluctance towards the use of DCD donor kidneys. New preventive strategies could limit I/R injury by preserving mitochondria (hypothetically with peptide SS-31 or activation of mitochondrial aldehyde dehydrogenase). This will overcome the detrimental effects of I/R injury on graft function and survival - thereby increasing the success rate of kidney transplantation. Show less
In recent decades, the use of a systems-based view of life has provided key insight into fundamental processes with respect to biology. In life sciences, important paradigm shifts are the way in... Show moreIn recent decades, the use of a systems-based view of life has provided key insight into fundamental processes with respect to biology. In life sciences, important paradigm shifts are the way in which we approach health and disease. Although modern medicine has traditionally emphasized pathology and acute conditions, our current understanding is that different interventions are needed for treating and preventing chronic disease. To design better interventions, new diagnostic tools are urgently needed in order to create new opportunities for achieving personalized health and medicine. The focus in diagnosis is, therefore, shifting from measuring single biomarkers such as glucose and cholesterol to creating complex maps of the dynamic patterns underlying regulatory processes. Moreover, the notion of "health" is viewed in a holistic context using biochemistry as a basis and then expanding this basis to include the psycho-social environment, including the individual's worldview. In the field of diagnostics, a highly promising new tool has recently emerged based on ultra-weak photon emission from biological systems, including all living cells. The aim of this thesis was to explore the applications of UPE and correlate it with biochemistry in order to obtain a deeper understanding of the processes that occur in living systems. Show less
Het doel van dit proefschrift was daarom de onderliggende biochemische processen gerelateerd aan migraine te onderzoeken. Het eerste gedeelte (Hoofdstukken 2 en 3) beschrijft twee onderzoeken... Show moreHet doel van dit proefschrift was daarom de onderliggende biochemische processen gerelateerd aan migraine te onderzoeken. Het eerste gedeelte (Hoofdstukken 2 en 3) beschrijft twee onderzoeken waarin gekeken is naar de biochemische constitutie van de hersenen van migraine patiënten door middel van proton magnetische resonantie spectroscopie (1H-MRS) en diffusie gewogen spectroscopie (DWS). In het tweede gedeelte (Hoofdstukken 4 en 5) wordt een systematische review met meta-analyse beschreven om een overzicht te geven van eerdere inspanningen om in verschillende lichaamsvloeistoffen biochemische verschillen te detecteren die relevant zijn voor migraine. Daarnaast wordt er een studie beschreven waarin is gekeken naar biochemische verschillen in liquor van migraine patiënten en gezonde controles. In het derde gedeelte worden twee methodologische studies beschreven waarin (Hoofdstukken 6 en 7) de biochemische stabiliteit van liquor en het effect van vervuiling met ethanol werden onderzocht. Show less
The work described in this thesis presents part of a framework that can be used to extract detailed disease biological information from peripheral tissue. This framework is based on the... Show moreThe work described in this thesis presents part of a framework that can be used to extract detailed disease biological information from peripheral tissue. This framework is based on the central dogma of biology “DNA to RNA to protein” and on a systems biology approach that aims to produce synergetic data whose disease pathological, prognostic and predictive value is greater than the sum of the individual experiment results. HD patients are often characterized by a multifaceted clinical profile, consisting of several symptoms and variable disease progression rates. Therefore, a systems approach such as the one described above is expected to be the most effective in identifying potential treatments and predictive biomarkers that will be most informative for the different patient subpopulations. Show less
Treating chronic diseases such as rheumatoid arthritis and type 2 diabetes mellitus is a hot topic that has been discussed widely and investigated extensively, but never solved, due in part to... Show moreTreating chronic diseases such as rheumatoid arthritis and type 2 diabetes mellitus is a hot topic that has been discussed widely and investigated extensively, but never solved, due in part to their high complexity. Integrating disease-related information using a systems approach may help improve our knowledge of stages of the disease, thus improving the accuracy of diagnosing chronic disease. With respect to integrative thinking, traditional Chinese medicine (TCM)‒based concepts may provide a suitable holistic model, as TCM describes disease syndromes/phenotypes as an experience-based reference from the systems level. Systems-based metabolomics provides a comprehensive picture of small molecular metabolites as a readout and provides biological interpretations of the pathophysiology of disease. The rapid, highly sensitive, non-invasive measurement of ultra-weak photon emission (UPE) -- which measures spontaneously emitted photons at the surface of the skin--has been proposed for supporting TCM-based diagnostics and for reflecting the whole body’s physiological and pathological status. Combining metabolomics with TCM-based diagnostics will provide a robust model for investigating the biological processes that underlie UPE. This thesis aimed to investigate system-wide perturbations by using/combining metabolomics, UPE and TCM-based diagnostics, to provide i) a systems view of chronic disease, and ii) personalized phenotyping guided by TCM-based principles. Show less
On the basis of systems thinking, in this thesis metabolomics, Chinese medicine (CM), as well as Western medicine (WM) were combined to achieve a more comprehensive systems diagnosis of patients... Show moreOn the basis of systems thinking, in this thesis metabolomics, Chinese medicine (CM), as well as Western medicine (WM) were combined to achieve a more comprehensive systems diagnosis of patients with chronic diseases. Specifically, metabolomics has been applied to characterize patients by small molecule profiles, which can be applied in phenotyping patients and matching these with optimal therapies. Chinese and Western medicine have different perspectives on diagnosis and could be highly complementary to each other, so combining both could have advantages in personalized diagnosis. Therefore, in this thesis a combination of systems approaches is used, including metabolomics, CM-based diagnosis principles as well as WM to provide systems diagnosis of patients with chronic diseases, in particular rheumatoid arthritis (RA). With systems diagnosis, we could better identify potential biomarkers to predict the WM therapeutic response of patients with RA and monitoring disease progression. Show less
In this thesis, metabolomics is used to study the role of the host-virus interaction on a metabolic level. A special emphasis is directed on the role of inflammation and oxidative stress on... Show more In this thesis, metabolomics is used to study the role of the host-virus interaction on a metabolic level. A special emphasis is directed on the role of inflammation and oxidative stress on the metabolic level, as part of the innate immune response against viral infection. We chose respiratory syncytial virus (RSV) and hepatitis B virus (HBV) as candidate viruses to metabolically study their role in acute respiratory infection and chronic hepatitis B infection. Secondly we also investigated infant metabolic and immunological consequences of in utero exposure to antiretroviral intervention and human immunodeficiency virus (HIV). Collectively, established targeted metabolomics approaches in conjunction with newly developed metabolomics methodologies and complemented with other “omics” techniques, were used to address pertinent questions related to host metabolic functioning and alterations during viral infection. In vitro RSV studies together with in vivo patient based studies relating to chronic HBV infection and in utero exposure too antiretroviral and HIV were used to address these questions. The work is divided into three research parts containing: i. the analytical methodology development work, ii. in vitro based metabolomics and iii. patient based metabolomics. Show less
The explosive increase in infections by pathogens is a major problem in the clinic today. The theme of this thesis was to find novel antibiotics from actinomycetes. Next-generation... Show more The explosive increase in infections by pathogens is a major problem in the clinic today. The theme of this thesis was to find novel antibiotics from actinomycetes. Next-generation sequencing revealed that the biosynthetic potential of actinomycetes had been grossly underestimated. In this thesis, different antibiotics-eliciting strategies, including microbial cocultivation, streptomycin-resistant mutation, overexpression of pathway-specific activator, variation of culture conditions, were utilized to enforce fluctuations in the production of bioactive compounds in actinomycetes, after which, NMR-based metabolic profiling was used to facilitate uncovering those elicited molecules. This pipeline allowed the discovery of new antibiotics involving various chemical skeletons, such as 7-prenylisatin, methoxylated isocoumarins, endophenazines, and C-glycosylpyranonaphthoquinones. On the other hand, genome-mining methodology enabled the discovery of a group of endophenasides and leucanicidin in Kitasatospora sp. MBT66, whereby the rhamnosylation of both scaffold are executed by a same promiscuous glycosyltransferase. Last but not least, a novel antibiotic termed lugdunomycin with unprecedented chemical scaffold, as well as a number of new angucycline-type antibiotics, were characterized from Streptomyces sp. QL37. The biosynthetic pathway of lugdunomycin was deciphered by genetic knockout and OSMAC (One Strain MAny Compound) strategy. In summary, this thesis explores an interface of genomics and metabolomics to accelerate new antibiotics discovery. Show less
Advances in technology have turned modern biology into a data-intensive enterprise. The advent of high-output technologies like Microarrays and Next-generation sequencing technologies has resulted... Show moreAdvances in technology have turned modern biology into a data-intensive enterprise. The advent of high-output technologies like Microarrays and Next-generation sequencing technologies has resulted in researchers grappling not just with huge volumes but also multiple types of data. While generation and storage of high-quality data are an important research focus, it is increasingly recognized that translating data into actionable information and insight is a critical research challenge. To infer reliable conclusions from the data, it is often necessary to integrate large amounts of heterogeneous data with different formats and semantics. Given the breadth and volume of data involved, this goal is best achieved through automated methods and tools for data integration and workflow management. This thesis presents automated strategies that combine bioinformatics and statistical methods to identify novel biomarkers in high-throughput OMICs datasets pertaining to the metabolic syndrome and to gain mechanistic insight into the underlying biological processes. An underlying theme in this thesis is data-driven approaches that generate plausible hypothesis which is followed by experimental verification. Show less
The research described in this thesis was aimed at analyzing the regulation of the channeling of carbon into different terpenoid pathways including the terpenoid indole alkaloids (TIA) and their... Show moreThe research described in this thesis was aimed at analyzing the regulation of the channeling of carbon into different terpenoid pathways including the terpenoid indole alkaloids (TIA) and their iridoid precursors in Catharanthus roseus cell suspension cultures. Of the different terpenoid classes; monoterpenoid (C10; TIA), triterpenoid (C30; sterol), and tetraterpenoid (C40; carotenoid), the latter was the largest pool. The production of carotenoids and TIA through the methyl-erythritol phosphate (MEP) pathway was increased after jasmonic acid elicitation, while the sterols from the mevalonate (MVA) pathway were not increased. Mevalonate feeding only increased the sterols. The effect of geraniol feeding alone and in combination with jasmonic acid elicitation resulted in increased levels of loganic acid and strictosidine, respectively. Catharanthus roseus cell suspension cultures overexpressing geraniol synthase in the plastid or cytosol were successfully developed and metabolic analysis by NMR-based metabolomics showed different metabolic effects related to the subcellular compartmentation of geraniol synthase. In conclusion, reallocation of C5 units between MEP and MVA pathway seems difficult, but both have different regulation which could be further studied to increase alkaloid production. Show less
With the persistent efforts to improve healthcare while the costs are ever-increasing, there is a growing demand for healthcare innovations including faster and cheaper analytical methods to... Show moreWith the persistent efforts to improve healthcare while the costs are ever-increasing, there is a growing demand for healthcare innovations including faster and cheaper analytical methods to support clinical decisions. The in-depth phenotyping of a biological system, such as comprehensive small-molecule profiling, will be key for breakthroughs in biomedical and pharmacological research towards the development of personalized medicine. In this thesis the development of new methods towards high-throughput, mass spectrometry-based profiling of small molecules in complex biological samples is presented. One option for high-throughput mass spectrometric profiling is direct infusion in which no separation techniques are employed prior to MS detection. With the pre-analytical part being a major bottleneck in a typical direct-infusion MS-based analytical workflow, the focus was set on the development of new sample pretreatment and sampling procedures, including miniaturized and automated concepts with the potential for high-throughput application. Show less
The world continues to be plagued by numerous diseases and health complications. Unfortunately, current drug discovery efforts are lagging behind compared to the pace of emergence of new diseases... Show moreThe world continues to be plagued by numerous diseases and health complications. Unfortunately, current drug discovery efforts are lagging behind compared to the pace of emergence of new diseases or the development of resistance to current therapies. Medicinal plants especially for people in developing countries are sometimes the only remedies available for the prevention and treatment of diseases. Modern science has also come to recognize the importance of medicinal plants in drug discovery and development given the contributions that this sector has made to drug discovery. As such, several pharmacognostic/natural products chemistry techniques have been developed to aid research on the medicinal properties of plants. The application of these techniques is very useful in two main areas in that they facilitate the validation of the medicinal properties of folk remedies and in the discovery of new remedies from natural products. This thesis focused on the evaluation of the medicinal properties of Vernonia guineensis Benth. (Asteraceae), which is commonly called Ginseng in Cameroon. The species was found to possess antimicrobial, antiplasmodial, antiproliferative/anticancer and anthelmintic effects. Results obtained and reported in this thesis will form a useful foundation for further research into the medicinal potential of this plant Show less
In this thesis new algorithms and methods that enable the de novo identification of metabolites have been developed. The aim was to find methods to propose candidate structures for unknown... Show moreIn this thesis new algorithms and methods that enable the de novo identification of metabolites have been developed. The aim was to find methods to propose candidate structures for unknown metabolites using MSn data as starting point. These methods have been integrated into a semi-automated pipeline to identify new human metabolites. The discovery of new metabolites will improve our capability to understand disease via its metabolic fingerprint, to develop personalized treatments and to discover new drugs. In addition, the cheminformatics methods presented in this thesis increase our understanding on the properties of human metabolites. The research described in this thesis has shown that the success of de novo metabolite identification relies on the synergy between analytical chemistry methods (i.e. LC-MSn) and cheminformatics tools. Show less
The detailed description of the chemical compounds present in organisms, organs/tissues, biofluids and cells is the key to understand the complexity of biological systems. The small molecules ... Show moreThe detailed description of the chemical compounds present in organisms, organs/tissues, biofluids and cells is the key to understand the complexity of biological systems. The small molecules (metabolites) are known to be very diverse in structure and function. However, the identification of the chemical structure of metabolites is one of the major bottlenecks in metabolomics research. Hence, the annotation and the structure elucidation of the metabolites are essential to understand the biological system under study. Actually, no single analytical platform exists that can measure and identify all existing metabolites. Multistage mass spectrometry (MSn) is a powerful analytical technique that helps identifying all these metabolites. This technique provides detailed structural information of the unknown metabolite by fragmenting the metabolite and its fragments recursively. However, only computational tools can provide a fast and straightforward analysis of the large amount of complex data that is generated by using MSn spectrometry. The aim of this thesis was to develop a novel semi-automatic approach for the identification of metabolites using MS n data. Furthermore, these tools were to be integrated into a pipeline to assign identities to unknown metabolites present in databases but especially to unknown metabolites not present in a database Show less
