Dissertation committee:
Assistant Professor Maria Halabalaki1
Professor Leandros A. Skaltsounis1
Associate Professor Theodora Psaltopoulou2
Professor Luc Pieters3
Professor Judith M. Rollinger4
Professor Emmanuel Mikros5
Professor Evangelos Gikas6
1Division of Pharmacognosy and Naturals Products Chemistry, Department of Pharmacy School of Pharmacy, National and Kapodistrian University of Athens, Greece.
2Department of Hygiene, Epidemiology and Medical Statistics, School of Medicine, National and Kapodistrian University of Athens, Greece
3Natural Products & Food Research and Analysis (NatuRA), Department of Pharmaceutical Sciences, University of Antwerp, Belgium.
4Division of Pharmacognosy, Department of Pharmaceutical Sciences, Faculty of Life Sciences, University of Vienna
5Division of Pharmaceutical Chemistry, School of Pharmacy, National and Kapodistrian University of Athens, Greece.
6Division of Analytical Chemistry, Department of Chemistry, National and Kapodistrian University of Athens, Greece.
Summary:
Based on world health organization (WHO) most recent data, chronic diseases constitute the leading cause of deaths in modern societies. Chronic disease rates are accelerating globally, advancing across every region, and pervading all socioeconomic classes. Cardiovascular diseases (CVD), cancer, obesity, and chronic obstructive pulmonary disease are the most frequently disorders in western populations and linked by common and preventable biological risk factors. It has been found that ageing and disease processes induce inflammation that decreases physical function and increases the risk for ageing-associated diseases. In this regard, high blood pressure, high blood cholesterol and overweight are the main factors which are related to major behavioral risk factors and specifically unhealthy diet, physical inactivity, and tobacco use.
After many years of research, scientists have postulated that food and human nutrition are regarded as the main axis for human well-being and are strictly associated with improved life quality and increased life expectancy. It first came to scientific attention with the seven countries study, in which results supported those Mediterranean countries of southern Europe have lower mortality rate from CVDs than in North America and northern Europe. Analyzing thoroughly these observations and starting from the famous quote “Let food be the medicine, and let medicine be the food” until to recent advances and clinical observations, is evident that diet frames the balance of human system and the key-feature for healthy-ageing is prevention. It has been proved that human dietary habits could serve as nutritional shield against age-related diseases and promote healthy ageing. The findings that diet interventions can increase lifespan, set dietary plants a promising source of chemical entities that promote healthy span and/or delay ageing. Towards this purpose several foods or food-based products have been studied for exploration of their biological impact as well as their content in phytonutrients. The recent advances in medicine and the increased research around natural products (NPs) have awaken the health-consciousness of people and the adaption of well-being lifestyle approaches. To-date Mediterranean diet (MD) is in the limelight of epidemiologists and nutritionists interest and year-by-year new data arise regarding the positive impact of MD lifestyle and several associations between specific foods and diseases prevention5. The philosophy and the basics of such diets has been adopted and adjusted in synchronous patterns to cover the modern way of life and needs.
Nutraceuticals is one of the recent elements introduced in our daily routine, while are defined as edible plants, dietary supplements and medicinal food, with a distinctive health impact in either prevention and/or treatment of disease. In general, there is a plethora of definitions regarding nutraceuticals; yet the concept in all terminologies remain same and underlines their contribution to health maintenance and diseases prevention. Herbal/medicinal plants, also known as functional foods, usually serve as source for nutraceuticals production. These plants are rich in phytochemicals which apart from forming the shield of plants against several exogenous factors act also as protectors from human diseases. Flavonoids, tannins, phenolic acids, alkaloids, terpenoids, essential oils, saponins are few of NPs chemical classes with proven advantageous effects to human health.
In the search for new biologically active substances, targeted bioassays play important role; in vitro and in vivo assays are mainly developed for extracts and/or pure compounds biological evaluation. However, despite the technological breakthrough that NPs drug discovery experiences and in spite of enormous efforts and investments, many active molecules in vitro fail when tested in vivo. This is mainly caused due to limited bioavailability and metabolisation processes (mostly under conditions of administration via the enteral route), which engender bio-transformations to molecules before reaching their final target. Intestinal absorption, hepatic metabolism, distribution in body tissues, bioconversion by the colonic microbiota with formation of colonic metabolites, and elimination are the main efficacy influencing factors. This is a very important drawback which, despite the impressive progress of the last years, does not allow the rationalization and the effectiveness of NPs discovery process. In addition, it is frequently observed that traditional medicinal plant extracts with a high level of ethnopharmacological and even clinical evidence, fail to show cell-based activity. The misleading “inactivity” of many NPs isolated from plants is likely caused because those are often pro-drugs, e.g., glycosides, which must undergo in vivo metabolic conversion (activation). Some well-known examples include the metabolic activation by gut bacteria of the isoflavone daidzein to the active metabolite equol, or of the phenolic compound salicin from willow bark to salicylic acid. Towards this purpose, new metabolisation assays have been developed, which mimic the metabolisation of compounds in the human body. One such example is the gastro-intestinal dialysis model (GIDM-colon) including microbial fermentation in the colon. It is important to highlight that although these methods are used in pharmacological research, nonetheless they have not been used so far in NPs research.
Therefore, the overall aim of the current thesis is to introduce an approach starting from the characterization of edible plants and/or foods, evaluation of their biological activity and finally metabolization study of certain bioactives. More specifically in the current thesis the phytochemical profile of several foodstuffs has been investigated incorporating new technologies and methods in NPs discovery and isolation in order to investigate the metabolism of natural entities coming from dietary plants, which promote healthy ageing. This is achieved by an integrated approach as illustrated in Figure 1. In parallel, NPs biotransformation reactions and metabolism in human system and explore the possible active forms of metabolites focusing on olive oil bioactives. Towards this direction an in vitro metabolism model was employed to study the behaviour and the bioavailability.The thesis is divided into two chapters. The first chapter focuses on biological screening via in vitro and in vivo assays of dietary plants with interesting phytochemical profile and promising biological results. Biological tests were centered between two key biological aspects, namely accumulating ROS elicited damages and the development of age-related metabolic diseases. A new approach is introduced by the parallel in vivo and in vitro testing and dereplication study of extracts and constituents thereof, with sophisticated analytical techniques in a successive and integrated manner.
Chapter 2 aims towards a detailed investigation into the absorption, metabolism and microflora-dependent transformation of olive polar constituents, the Gastro-Intestinal Dialysis Model with Colon (GIDM-Colon) was applied. Specifically, in this chapter the anatomy and physiology of human alimentary canal, are described. The human gut microbiota and their significance in human health is extensively discussed, while the modulation of gut microbiota through dietetic maneuvers including olive polyphenols is also commented on. Successively, the so far ADMET assays and metabolisation protocols are discussed and the increasing need of in vitro assays that simulates human metabolism is presented. Moreover, the in vitro GIDM-Colon Model that was incorporated in this study is broadly described and statistical evaluation of GIDM-Colon results of each tested compound and TPF are discussed.
