information on bsc and msc thesis projects at the laboratory ......bsc and msc thesis projects –...

L A B O R A T O R Y o f F O O D C H E M I S T R Y

Information on BSc and MSc thesis projects

at the Laboratory of Food Chemistry

Last update 2/3/2017 Go to www.fch.wur.nl ( education)

BSc and MSc thesis projects – Laboratory of Food Chemistry

1

Table of Contents

CHOOSING AND PERFORMING A THESIS RESEARCH PROJECT AT THE LABORATORY OF FOOD CHEMISTRY ..... 2

PART 1: QUALITY AND/OR PROCESSING OF FOOD (RAW) MATERIALS ............................................................. 4

EFFECT OF THE MAILLARD REACTION ON PROTEIN DIGESTIBILITY (DAIRY SUBJECT) ................................................................. 4 THE ROLE OF MAILLARD REACTIONS IN PROTEIN FOAM STABILITY (DAIRY SUBJECT) ............................................................... 4 INGREDIENTS, INTERFACES AND ICE CREAM (DAIRY SUBJECT) ............................................................................................ 5 FREE RADICAL INTERMEDIATES PRODUCED DURING PRIMARY LIPID OXIDATION IN O/W FOOD EMULSIONS ................................. 6 NEW ANALYTICAL TECHNIQUES FOR RESEARCH ON FOOD .................................................................................................. 6

PART 2: ENZYMATIC OR FUNGAL UPGRADING OF FOOD (RAW) MATERIALS .................................................... 7

UPGRADING LIGNIN – FUTURE AS NATURAL INGREDIENT FOR FOOD & FEED OR NATURAL CHEMICALS? ...................................... 7 CAN LACCASE-MEDIATOR SYSTEMS UPGRADE LIGNIN: NEW NATURAL INGREDIENTS? ............................................................. 8

PART 3: HEALTH ASPECTS OF FOOD ................................................................................................................. 9

ENHANCING THE PRODUCTION OF PLANT METABOLITES WITH ANTIMICROBIAL ACTIVITY ......................................................... 9 STRUCTURE ACTIVITY RELATIONSHIPS OF ISOFLAVONOIDS AS ANTIMICROBIAL AGENTS .......................................................... 10 ANTIMICROBIAL PHYTOCHEMICALS FROM BRASSICA ..................................................................................................... 11 EFFECTS OF PROCESSING OF PROTEINS ON THEIR IMMUNOGENICITY: ALLERGY AS A MODEL (DAIRY SUBJECT) ............................ 12 THE FATE OF HUMAN MILK OLIGOSACCHARIDES (HMO) IN INFANT GUT (DAIRY SUBJECT) .................................................. 13 N-GLYCANS IN BOVINE AND HUMAN MILK (DAIRY SUBJECT) ........................................................................................... 13 TOWARD CONTROLLED STEERING OF MICROBIOTA AND IMMUNITY IN INFANTS BY NON-DIGESTIBLE CARBOHYDRATES ................. 14 BEHAVIOUR AND DIGESTIBILITY OF STARCH IN PIGS ....................................................................................................... 15 CARBS CAN MAKE THE DIFFERENCE: HOW PECTINS FUEL IMMUNITY .................................................................................. 16 THE EFFECT OF PROTEIN STRUCTURE ON DIGESTION DYNAMICS (DAIRY SUBJECT) ................................................................. 17 THE EFFECT OF DAIRY PROTEIN COMPOSITION ON PROTEIN DIGESTION KINETICS .................................................................. 18

PART 4: NEW AND/OR IMPROVED INGREDIENTS FOR FOOD .......................................................................... 19

USE OF ENZYMATIC CROSSLINKING TO PRODUCE PROTEIN NANOPARTICLES (DAIRY SUBJECT) ................................................. 19 UNDERSTANDING POLYSACCHARIDE INTERACTIONS IN FRUITS AND VEGETABLES .................................................................. 20 ISOMALTO/MALTO-POLYSACCHARIDES (IMMPS): NOVEL POLYSACCHARIDES FROM STARCH. .............................................. 21

PART 5: EDUCATIONAL DEVELOPMENT .......................................................................................................... 22

DESIGN AND/OR EVALUATION OF DIGITAL LEARNING MATERIAL FOR FOOD CHEMISTRY ........................................................ 22 DEVELOPING NEW LEARNING ACTIVITIES OR IMPROVING EXISTING LEARNING ACTIVITIES ....................................................... 22


2

Choosing and performing a thesis research project at the

Laboratory of Food Chemistry

Purpose of a thesis research project

Your thesis research project is an important part of your

study. In the past years you have gained knowledge in

food science with the aid of teachers and supervisors

during lectures, laboratory classes, computer classes,

etc. You learned obtaining information by literature

searches, internet searches, performing experiments,

evaluating your results and writing reports and/or giving

presentations. Also, you have started to a certain extent

to develop a scientific way of reasoning on how to solve

problems. The purpose of a thesis project is to further

develop this skill to analyse and solve a problem in a

scientific way. In addition, a thesis project also perfectly

serves the objective of obtaining knowledge/insight in a

specific area of research.

A thesis research project at the Laboratory of Food Chemistry

After you have chosen a research project, you will start-up this project together with

your supervisor. The supervisor is usually a researcher from our Laboratory. In short,

you will make a research plan, perform experiments by using our equipment and give

some presentations to your fellow-students. The project is finalised with a report and a

final presentation (colloquium).

A thesis research project in Food Chemistry is related to a specific (bio)chemical food

product or biomass conversion issue, which on its turn is related to an industrial

application. In this way we do not solve an industrial problem, as this is the task of our

graduates once they work in industry. Instead, we aim at understanding the mechanisms

behind the problems and we aim to approach this in an academic way. With this

knowledge/insight students are enabled to address (yet unknown) future problems, once

they are graduated.

Within the Laboratory of Food Chemistry we focus our research on a limited number of

topics. By doing so we are able to maintain our internationally well recognized position.

This is also of benefit for students, as on the one hand their research project is well

embedded and on the other hand it is a good reference regarding future employment. As

a consequence the number of areas of research possible for thesis projects is somewhat

restricted. In addition, to guarantee an optimal supervision, it is not always possible to

choose the subject of your interest. This can be due to the fact that other students have

already chosen the subject and for optimal supervision we want to restrict the number of

students per supervisor.

The main research topics of our Laboratory are (see also Table of Contents):

Part 1: Quality and/or processing of food (raw) materials

Part 2: Enzymatic or fungal upgrading of food (raw) materials

Part 3: Health aspects of food

Part 4: New and/or improved ingredients for food

Part 5: Educational development

Within these topics we deal with several foods and food raw materials such as dairy

products, cereals, fruits, vegetables, legumes, beverages (coffee, tea, beer), etc.


3

Choosing a thesis research project

First, formulate for yourself what specific knowledge/skills you want to learn during the

thesis project. If you are e.g. interested in enzymology use this aspect for making your

choice.

Second, is the background of the subject motivating you? If you do not like the

background of the subject it will decrease the chance of success. Be aware that although

it might be interesting to work on an industrial problem or product, it is the underlying

way of performing research that is most important. By doing this you are, as stated

above, enabled to address any industrial future problem. In order to acquire a solid

background in food chemistry this can usually be trained better if one works with specific

components of food rather than with the complete food product itself.

On the following pages you can find the description of the research topics followed by

information on specific thesis projects.

I am sure you will be able to find a subject that fits you. Many students have preceded

you and enjoyed their thesis projects.

prof.dr.ir. Harry Gruppen.

Chair of Food Chemistry


4

Part 1: Quality and/or processing of food (raw) materials

Effect of the Maillard reaction on protein digestibility (dairy subject)

The Maillard reaction occurs during thermal processing of food such as baking, roasting,

brewing and etc. It can also happen by design to improve the techno-functional

properties of proteins, e.g. emulsion and foam stability. In the early stage of the

reaction, the Maillard reaction is between the amino group of a protein and the reducing

end of a carbohydrate (glycation). In the later stage, the Maillard reaction can lead to

protein cross-linking (aggregation). Previous research showed that both phases influence

the protein enzymatic hydrolysis. Proteases are characterized by specificity and

selectivity. It is expected that the trypsin hydrolysis (specific for lysine/arginine, the

same as glycation) will be negatively affected by glycation. Other proteases (non-

lysine/arginine specific) might also be affected by glycation and/or aggregation. We

would like to study how much the Maillard reaction influences protein enzymatic

hydrolysis and to understand the mechanism of it at a molecular level.

In vitro enzymatic hydrolysis is used to

mimic gastrointestinal digestion.

Analytical tools such as pH-stat and LC-

MS are used to identify and quantify the

glycation/aggregation and peptide

formation.

Thesis project:

• The effect of substrate concentration on protein enzymatic hydrolysis

The role of Maillard reactions in Protein Foam stability (dairy subject)

The foam stability of proteins is of relevance to many food products. While many studies

have been performed to find the link between the properties of the proteins and the

resulting foam such a link has not yet been established. In recent research evidence was

found for the effect of Maillard induced aggregation of proteins in the formation and

stabilization of foam. In this way the functional properties of the proteins could be

boosted significantly in a relatively simple process.

The Maillard reaction starts by linking one saccharide to a free amino-group of a

protein. We observed that as a result of this reaction the proteins started to form

oligomers and small aggregates. Exactly these oligomers and aggregates will be the

subject of study. By selective treatment, or modification of proteins well-defined ‘model’

systems will be produced. These systems should be characterized to the type of

aggregation that was obtained (size, size-distribution, charge or hydrophobicity).

Thesis projects:

Firstly the effect of conditions (protein type, sugar type, temperature / time) will

be studied. Also other reactions than the Maillard reaction could be used to induce

oligomer formation. After formation and characterization of the chemical

properties, some assays can be used to test the changes in foam properties of the

modified variants.


5

Ingredients, Interfaces and Ice cream (dairy subject)

Ice cream consists for 50 % of air. The air bubbles are stabilised by mixtures of dairy

proteins and low molecular weight (LMW) surfactants. In the production, first the LMW

surfactants adsorb to the interface. Later in the process, the LMW surfactants are

displaced from the interface by the adsorption of proteins. If novel ingredients are used,

this subtle balance between adsorption and desorption is easily disturbed, which can lead

to undesired product properties.

Ingredients Interfaces Ice cream

T0

T1

To control the stability of foam in mixed protein – surfactant systems, the interactions of

proteins and surfactants in the interface needs to be understood. In this project, you will

investigate how the chemical properties of proteins and LMW surfactants alter the

adsorption and desorption of these compounds in mixed systems and relate it to the

stability of foam made from mixed solutions.

Thesis projects:

Investigation of interactions of mixed systems of proteins and LMW surfactants in

bulk solution

o Structure and physico-chemical properties of proteins (e.g. CD, DSC and

protein charge)

o Foam and interfacial properties of mixed systems

You will determine these properties in a well-defined system of dairy proteins and a non-

ionic surfactant in order to investigate the formation of hydrophobic complexes of protein

and surfactants.

The project will involve techniques to determine foam properties (FoamScan), interfacial

properties (automated drop tensiometer) and bulk characteristics (CD, ESI-MS,

Nanosizer).


6

Free radical intermediates produced during primary lipid oxidation in O/W food

emulsions

Food emulsions (O/W) such as mayonnaise and salad dressing are very prone to

deterioration caused by lipid oxidation. The dispersed oil droplets contain unsaturated

fatty acid chains that can be readily oxidized via diverse mechanisms.

The first pathway of this oxidation is a free radical chain reaction, leading to

hydroperoxide formation. Degradation of these hydroperoxides leads to the formation of

so-called secondary oxidation products (e.g. aldehydes, epoxides), which are responsible

for the rancidity and off-flavor of O/W food emulsion products. The mechanistical

pathways of lipid oxidation have been partly unraveled in bulk oils. However, in food

emulsions, both chemical and colloidal effects come into play. Unravelling of these

mechanisms is hampered by lack of knowledge on unstable radical intermediates.

This project will focus on the initial stage of primary oxidation and the transition stage

between primary and secondary oxidation. Here we will aim to identify and quantify the

radical intermediates by Electron Spin Resonance (ESR). This technique allows for the

detection of stable radicals. In lipid oxidation, formed radicals are very reactive and

short-lived, making it impossible to analyze them as such. To tackle this problem, so-

called spin-traps can be used. The emulsion matrix adds significant complexity to the

applicability of a spin-trap, which needs to be considered when selecting the right spin-

trap. Ultimately, the ESR-method will be applied in shelf-life tests, where we want be

able to resolve the formation of specific radicals in time.

ESR-facilities are located in the Helix building at the WUR-campus. Samples will be

generated in collaboration with Unilever R&D Vlaardingen.

Thesis projects

Identification and quantification of free radical intermediates produced during

primary lipid oxidation in O/W food emulsions by ESR

Isolation and identification of hydroperoxides (LC-MS + NMR)

New analytical techniques for research on food

During the last years a lot of new techniques and technologies were introduced which

allows the food chemist to monitor reactions which take place in agricultural products

during storage and processing. Nowadays it is possible to monitor chemical reactions in

food products which was impossible only a few years ago. The discipline of proteomics

would not have evolved so quickly without the development of advanced mass

spectrometry techniques. Therefore, it is now possible to determine the structure of

complex mixtures of bio molecules.

The techniques which are available and used at the Laboratory of Food Chemistry are

described well. But for the introduction of these new techniques for the monitoring of

different components it is sometimes necessary to carry out methodology research. For

example: the quantification of components when new HPLC detectors are used, the

better understanding of the fragmentation reactions which take place during the

ionization of oligosaccharides in the LC-MS or the improvement of HPLC separation of

complex mixtures of polysaccharides. The student should have a good perception of

analytical techniques.

Thesis projects:

Application of LC-MS, MALDI-TOF MS, HPLC.


7

Part 2: Enzymatic or fungal upgrading of food (raw) materials

Upgrading lignin – future as natural ingredient for food & feed or natural

chemicals?

Upgrading lignin, one of the most abundant polymers in nature, has already been a challenge for

decades. Excitingly, with recent and new analytical possibilities, revealing the black box of lignin

comes into reach! Hence, now, possibilities to upgrade lignin may open up new ways towards natural

ingredients for food & feed products or as natural chemicals, e.g. as anti-oxidants or anti-microbials.

Biological conversion of lignin, or lignin-rich raw

materials, plays a role during the cultivation of

mushrooms. Vice versa, fungi are an interesting

tool to modify, degrade or convert lignin. The

exact effect of (various) fungi on the lignin

structure is unknown. In addition, it is known that

certain fungal species produce enzymes, which

may alter or degrade lignin structures – however,

underlying mechanisms are unknown.

To accommodate development of new ingredients

from lignin, the relation between the lignin

structure and possible enzyme or fungal effects

on this structure needs to be fully understood.

Hereto, the chemical structure and quantification

of lignin is required for which isolation of

(various) lignin fractions, a highly challenging

task, is aimed at.

In this project a newly developed method for lignin analysis (pyrolysis coupled to gas chromatography

with mass spectrometric detection (py-GC/MS)) is employed as tool for both quantification and

structural characterisation of lignin. While py-GC/MS-characterisation of lignin is in place,

quantification requires further development of the method.

Different strategies might be employed to upgrade/alter lignin, such as enzyme incubation, fungal

growth and chemical degradation.

Thesis projects:

Mild isolation and characterisation of lignin from various raw materials, including mapping lignin/ phenolic compounds-like structures in the various fractions obtained.

Method development for quantitative lignin analysis with py-GC/MS.

Effect of fungal growth or enzymes for upgrading/altering lignin (examples of relevant analytics are py-GC/MS, HPSEC or UPLC-MS).


8

Can laccase-mediator systems upgrade lignin: new natural ingredients?

Lignin is a polymeric aromatic structure in plant cell walls, and is one of the most

abundant polymeric materials in nature. Its insolubility and recalcitrance to enzymatic

breakdown make lignin breakdown one of the largest challenges in biorefinery.

Although challenging, enzymatic breakdown of lignin may open up new ways towards

natural ingredients for food & feed products or as natural chemicals, e.g. as anti-oxidants

or anti-microbials.

Laccases are among the few enzymes that show activity towards lignin. These enzymes

couple the reduction of O2 to the one-electron oxidation of aromatic substrates. For the

oxidation of the majority of the lignin subunits, it is necessary to combine laccase with a

mediator (a small molecule that acts as electron carrier between enzyme and substrate).

The initial oxidation of lignin by laccase-mediator systems has been suggested to result

in a competition between lignin depolymerization, ketone formation at the Cα atoms, or

lignin polymerization. Currently, it is poorly understood how this competition is balanced.

Mediator

Mediator·O2

H2O

Polymerization?Depolymerization?Ketone formation?

Before laccase-mediator systems (LMS) can be applied to result in new natural

ingredients, we need to be able to predict the effect of these systems on the substrate

structure.

This project aims at understanding how laccase-mediators act on lignin or lignin-like

structures. The focus is on the competition between polymerization, depolymerization

and Cα oxidation of lignin, as initiated by laccase-mediator systems.

Attention is paid to the influence of mediator types, lignin structure and reaction

conditions.

Thesis projects:

Understand the effect of LMS on model dimers, that represent structures of

industrial lignins (UHPLC-MS).

Understand the influence of mediator type on LMS reactions, including product

profiling (UHPLC-MS, O2 consumption).


9

Part 3: Health aspects of food

Enhancing the production of plant metabolites with antimicrobial activity Prenylation as a tool to diversify and enhance antimicrobial properties of phenolic compounds

Plants of various species are known to produce defence compounds when they are

exposed to stress. These defence compounds can possess antimicrobial activity. The

process of prenylation, substitution with a

prenyl-group (C5), can be performed in vivo or in vitro. We are working with plants of

the family Leguminosae, currently focussed on peanuts which produce prenylated

stilbenoids during micro-malting combined with fungal stress. Prenylation plays a key

role in the functionality of phenolic compounds and has been shown to modulate

antimicrobial activity of flavonoids and stilbenoids. In addition, we are applying similar

methodologies on benzoxazinoids, a class of defence compounds found in several species

of grains (e.g. wheat, rye, barley).

To summarise, we will employ different approaches to produce antimicrobial (prenylated)

compounds:

In vivo:

- Micro-malting: germination of plant seeds combined with exposure of seedlings to

fungal growth.

In vitro:

- Prenyltransferases (PTs): Prenyltransferase enzymes can be produced by

genetically modified E. coli. PTs can then be used to attach prenyl-groups to

various phenolic compounds.

- Chemical prenylation: Prenylation of flavonoids and stilbenoids via organic

synthesis.

Thesis projects:

Thesis projects will include a combination of practical work in the fields of chemistry and

microbiology.

Micro-malting seeds with the germinator combined with fungal challenging. This

will be followed by extraction, fractionation, purification, and characterisation of

phenolic compounds by (preparative) liquid chromatography (LC) and mass

spectrometry (MS).

Targeted production of prenylated phenolic compounds with microbial

prenyltransferases (PTs). This will involve practical work in genetic modification of

Germinator used in micro-malting,

normally used in the beer brewing

industry for pilot-scale malting

Example of a prenylation reaction, the stilbenoid resveratrol is substituted with a prenyl-group.


10

bacteria and purification of the produced PTs. Prenylated phenolics will be

analysed by LC-MS.

Organic synthesis of prenylated phenolic compounds. Optimisation of the

production of (specific) prenylated flavonoids or stilbenoids. Products will be

analysed by LC-MS and NMR. The product molecules will be purified using

preparative LC.

Antimicrobial testing of the aforementioned compounds, and investigation of the

structure-activity relationship and mechanism of antimicrobial activity.

Structure activity relationships of isoflavonoids as antimicrobial agents

At present there is a great concern about

the emergence of multidrug resistant

bacterial strains, and continuous research

efforts are being devoted for the

development of new and effective

antimicrobial agents. Many plants from the

Leguminosae family have been used for

medicinal purposes since ancient times in

many parts of the world. Therefore,

substantial attention has been focused on

the exploration and utilization of legume

phytochemicals as alternatives of traditional

antimicrobial agents.

Previous studies have found these plants to be rich in isoflavonoids. Isoflavonoids are

phenolic plant secondary metabolites, and are produced as part of the plant defence

system. Isoflavonoids have been shown to exhibit interesting biological and

pharmacological properties, such as antioxidant, antibacterial, antiviral and anti-

inflammatory activity. These properties seem to be highly structure-dependent.

Prenylation is a type of substitution commonly found in bioactive isoflavonoids. Some

prenylated isoflavonoids have shown strong inhibitory activity towards antimicrobial

resistant strains, as well as synergy with traditional antimicrobials. However, the exact

relation between chemical structure (e.g. number, type and position of substituents in the isoflavonoid skeleton) and bioactivity is not yet well understood.

In order to develop novel and effective antimicrobials it is necessary to fully understand

these structure-activity relationships (SAR), and elucidate the main chemical features

necessary to have good antimicrobial activity. Different strategies might be employed to

enhance the antimicrobial potential of these compounds (e.g. fermentation, enzymatic prenylation).

Thesis projects

In general, thesis projects will include practical work from both chemistry and

microbiology.

Characterization and purification of antimicrobial isoflavonoids present in seed

extracts by chromatographic techniques, such as UPLC-MS and preparative HPLC,

in order to determine the main structure-activity relationships.

Elucidation of the mode of action of the main antimicrobial compounds using

different biochemical assays.


11

Antimicrobial Phytochemicals from Brassica

There is a growing interest in finding novel antimicrobial agents because of the increasing

incidence of antibiotic resistance. Nonetheless, finding new potential antimicrobial agents

against pathogenic Gram-negative bacteria is a challenge due to their specific feature,

called efflux pump system. Previous research in Laboratory of Food Chemistry has shown

that prenylated (iso)flavonoids from elicited legume sprouts do not show growth

inhibition in Gram-negative bacteria, e.g. Escherichia coli when there is no efflux pump

inhibitor being used.

Therefore, we are now investigating antimicrobial property of other types of plant

secondary metabolites from different plant family, namely Brassicaceae. We are also

extending the investigation on antimicrobial activity against yeast and fungi which cause

food spoilage.

Brassica is characterized by glucosinolates (GSL) and sulfur-containing alkaloid. Both are

involved in the plant defense giving an indication that they are potentially antimicrobial.

Furthermore, GSL is hydrolysed in a presence of myrosinase, a thioglucosidase. In

neutral pH, it forms isothiocyanates (ITC) which have been reported to have

antimicrobial property. However, there is lack of systematic study on their mode of action

as well as structure-activity relationship.

Besides, we are also interested in studying the hydrolysis of GSL and the stability of ITC

being formed. For this, we need to design an analytical procedure to monitor the

presence of ITC both qualitatively and quantitatively using UHPLC-MS analysis.

Thesis projects

In general, thesis projects will include practical work from both chemistry and

microbiology.

Develop an analytical method to simultaneously monitor GSL, ITC, and sulfur-

containing alkaloids both qualitatively and quantitatively using UHPLC-MS

Study the GSL hydrolysis by myrosinase and the stability of ITC in aqueous

solution upon different temperatures and pHs

Study the mode of action of the main antibacterial compounds

Study the antifungal activity of ITC, GSL, and alkaloids against yeast and fungi

and their mode of action


12

Effects of processing of proteins on their immunogenicity: allergy as a model (dairy subject)

Food allergy develops in two phases: a sensitisation phase and an elicitation phase, the

latter resulting in clinical symptoms of allergic disease. Sensitisation is the phase during

which immunological priming to the inducing allergen occurs, leading to the evolution of

aTh2-biased immune response and the production of IgE-antibody.

Skewing of adaptive immune response to a Th2-type phenotype and IgE-antibody

production necessarily are preceded by an innate immune response to the allergen.

Activation of receptors on cells of the innate arm of the immune system is crucial for the

initiation of adaptive immune responses.

Processing changes the structural and chemical properties of proteins, and hence of

allergens. Proteins denature, aggregate, bind to lipid structures, and undergo

glycosylation and or glycation (Maillard reaction). These processing-related structural and

chemical changes will influence how the immune system will respond, possibly leading to

eventual allergenicity.

The aim of this project is to study the effect of processing-induced alterations in proteins

(unfolding, fibril formation, aggregation, Maillardation) on the response that they provoke

in (innate) immune cells, such as macrophages and dendritic cells.

Thesis projects:

Characterisation of processing-induced structural alterations in (model) food

proteins (fluorescence, circular dichroïsm, size, electrophoretic behaviour, etc.)

Analysis of immune response to such alterations by exposing model cell cultures

to the proteins by immunologic methods such as phagocytosis and cytokine

production


13

The Fate of Human Milk Oligosaccharides (HMO) in Infant Gut (dairy subject)

Milk is the first food to fulfil the need for

the neonates. It provides some potentially

significant physiological and biological

functions such as prebiotic, antipathogenic

and immunomodulatory activities that are

partly resulted from oligosaccharides.

Thesis project:

Oligosaccharides present in human milk

and infant faeces will be identified and

quantified by CE-LIF, MALDI-TOF-MS,

HILIC-MS, etc. Mechanisms of

metabolisation, conversion and utilisation

of HMO in infant gut will be proposed.

N-Glycans in bovine and human milk (dairy subject)

Milk is the first source of nutrition for newborns. Newborns have an immature immune

system and milk contains many biofunctional components, such as human milk

oligosaccharides (HMOs) and proteins, that are known to be involved in the development

of the immune system. HMOs not only provide nutrition to the infants but also have

shown to influence the composition of the gut microbiome, modulate the immune

system, and help protect against pathogens. Proteins play a pivotal role in protecting the

gut mucosa against pathogens and are also known to play an important role in the

protection of the mammary gland. Since the proteins in milk are highly glycosylated,

glycans are considered to be a major contributor to this immune system development.

Thesis project:

This project will focus on the release of glyco-tails from the proteins. Recently, there is a

method developed in our lab for the glycation of cow milk proteins which needs

validation. After this the method will be used to analyse the glycosylation patterns of

proteins present in milk from different mothers during lactation.

N-glycans can be detected/identified in bovine milk by LC-MS and MALDI-TOF-MS


14

Toward controlled steering of microbiota and immunity in infants by non-

digestible carbohydrates

Bacteria colonizing the infant mucosa guide the development of a balanced immune

system and also support maturation of the gut-barrier. Mother-milk has been considered

the golden standard for guiding this colonization. Mother-milk contains energy sources

for microbiota and also supports immune function directly. For those infants where

mother-milk is not a feasible option, cow-milk formulas

supplemented with non-digestible carbohydrates (NDC)

are used. An important function of these NDCs is a

preferred support of Th1-responses responsible for

fighting infections. However, recently it has been found

that not all NDCs currently applied support Th1-responses

and that induced responses are dependent on the

composition of NDCs.

The goal of this project is the design of tailored NDC-

supplements for healthy and disease-prone infants who

do not receive mother-milk. In this way the immune

response development with respect to T-cell skewing and

epithelial barrier function will be supported. As a result

the risk of developing allergies or gastrointestinal

disorders in a later stadium of life will be lowered.

Thesis projects:

Characterization and quantification of NDCs by MALDI-TOF-MS, HILIC-MS etc.

Additional fractionation using AEC, SEC or preparative HILIC-MS will be performed

to unravel the different structures present in the highly complex mixtures.

Following the fate of selected (fractions of) NDCs during in vitro batch

fermentation using infant faecal inoculum by HILIC-MS and SCFAs analysis using

HPLC and GC. Unknown glycosidic NDC products will be identified by HILIC-MS.

Analysis of NDCs, their degradation products and SCFAs produced after incubation

in a co-culture system mimicking the infant-gastrointestinal tract (UMC

Groningen)


15

Behaviour and digestibility of starch in pigs

In a world with a rapidly growing population with

a higher demand for animal protein, the global

meat consumption is on the rise. To meet these

increasing demands, efficient and sustainable use

of raw ingredients becomes more and more

important.

Starch is the main storage carbohydrate in plants

and forms an important source of energy in the

diet of humans and animals. Many different

sources of starch are used in food and feed. This

project focusses on studying the chemical and

physical properties of starch from different

sources, and their physical behaviour and

digestibility in a pig model. By gaining a better

insight in the types of starch and their relations

to pig digestibility, more precise models can be

developed to predict the feeding value of the

starch, which will lead to a more efficient use of

the energy in starch.

The main questions to be answered in this project, is how and where different starches

that are present in (processed) feed are digested within a pigs digestive tract, and how

this effects the pig's performance. This project consist of a combination of in vitro (lab)

digestion trials, which will be done to adapt human digestible and resistant starch assays

to the animal situation, and in vivo experiments (in pigs), where digesta of differently fed

animals will be analysed.

Thesis topics:

Within this project various thesis topics are available for BSc as well MSc students.

Analysis of the effect of the feed matrix and feed processing on starch

digestibility, using newly developed in vitro digestion models.

Study chemical characteristics of partly digested starch samples from an in vivo

digestibility trial with pigs (February 2016).

Comparing in vivo intestinal samples from a starch digestibility trial with pigs with

samples from an in vitro digestibility assays (February 2016).


16

Carbs can make the difference: how pectins fuel immunity

A disbalance in microbiota communities in the intestine is implicated in a large number of

Western diseases. This correlates with low intake of dietary fibre in Western diets. Pectin

is a dietary fibre that might be essential for prevention of Western diseases. Recently,

evidence has been found that beneficial effects of pectin are highly dependent on its

chemical structure and the effects go beyond prebiotic effects. Earlier research has been

demonstrated that low methyl esterified pectins have direct, microbiota-independent

effects due to their direct interaction with TLRs in the small intestine. High methyl

esterified pectins mainly impact the microbiota in the colon to enhance formation of

pectic oligosaccharides and SCFA.

The aim of our research is to fully characterize bioactive pectins and their intermediate

degradation products upon fermentation by enzymatic fingerprinting techniques.

Pectin molecules from various sources will be applied and enzymatically tailored with a

combined beneficial effect. The already available data on pectin health effects will serve

to produce a mixture of pectins with a desired molecular weight and methyl esterification

pattern, which will be tested (in collaboration with the University Medical Center

Groningen) in mice with a temporary disrupted small-intestinal barrier-function and mice

with a disrupted colonic-integrity. Results obtained will be used to further tailor pectin

structure to yield optimal bioactivity.

This study will contribute to the understanding of the structure-function relationship of

pectins.

Thesis topics:

Enzymatic tailoring bioactive pectic polymers.

Chemical characterization of tailored pectins using enzymatic fingerprinting.

Identifying and quantifying intermediate fermentation products from mice digesta

& faeces of pectin fed mice.


17

The effect of protein structure on digestion dynamics (dairy subject)

Enzymatic protein hydrolysis finds its interest in protein digestion and availability in the

digestive tract as well as in industrial application of protein hydrolysates as food

ingredient. To be able to predict the exact outcome (hydrolysate composition) of an

enzymatic hydrolysis and thereby predict the bio- and techno-functional properties, the

activity of the proteases should be well understood. Nowadays, the activity of proteases

is mainly explained by its specificity, which indicates after which amino acids the

protease can cleave the peptide bond (e.g. trypsin can cleave after lysine and arginine).

However, in practice it was observed that not all the possible cleavage sites are

hydrolysed to the same extent or some even not at all (DHmax,theoretical≠DHmax,experimental).

This phenomena can be explained by the selectivity of the protease, which indicates the

preference for certain possible cleavage sites within a given specificity (Butré, 2014).

As the selectivity can have a significant influence on the final outcome of the protein

hydrolysis, it is important to understand the mechanism behind selectivity in order to

predict the outcome. It is hypothesized

that in general selectivity can be

explained by the ‘accessibility of’ and

‘affinity to’ a certain cleavage site. It is

thought that the accessibility can be

related to the tertiary structure of the

substrate protein as well as possible

aggregation, and affinity can be

explained by the primary structure

together with possible exposed charges.

The aim of this study is to investigate

the mechanistic factors determining the

selectivity of proteases by performing in vitro protein hydrolysis, using commercially

available proteases and mainly milk proteins as substrate. After hydrolysis, the

hydrolysate composition (peptides and/or intact protein) will be analysed using RP-UPLC-

UV-MS. From the composition over time, the selectivity can be calculated which can then

be related to the starting conditions/substrate properties.

Thesis projects:

Describing the relation between tertiary structure of the substrate protein and the preference for intact protein (zipper-) or released peptides (one-by-one mechanism).

Describing the effect of tertiary structure as well as primary structure of different

substrate proteins on the selectivity of trypsin.

One-by-one

Zipper


18

The effect of dairy protein composition on protein digestion kinetics

Proteins are digested in the gastrointestinal tract by proteases, ultimately leading to tri-

and di-peptides and free amino acids (AA) that can be absorbed by the intestinal

epithelium. The epithelium releases free AA into the blood stream. The kinetics of protein

digestion and absorption appear to influence the fate of the AA systemically, i.e.

incorporation in protein or oxidation and secretion as urea.

Protein digestion and absorption kinetics appear to depend on the protein composition. For

example it has been shown in healthy volunteers that the postprandial AA plasma peak

after whey protein ingestion is higher than after casein ingestion. This is usually attributed

to the differences in gastric emptying between the proteins due to their distinct

physicochemical behavior in the stomach. However, we have shown previously that in an

in vitro digestion model enzymatic protein hydrolysis kinetics are also different between

whey and casein proteins. This may contribute to the overall protein digestion and

absorption kinetics as reflected in postprandial AA peak as well.

Enzymatic protein hydrolysis depends on the primary structure of the proteins. Whey and

casein milk fractions are mixtures of proteins with unique primary structures. It is of

interest to further the understanding of the effect of protein composition of protein

mixtures on the overall enzymatic protein hydrolysis kinetics.

Thesis project:

In vitro digestion experiments will be perfomed to simulate gastrointestinal

conditions. Protein hydrolysis kinetics of dairy protein (mixtures) will be monitored

by SDS-PAGE, OPA, HP-SEC and UPLC.

Experiments will be performed at the life-science lab at Nutricia Research on Utrecht

Science Park.

The life-science lab is a ML-II laboratory, therefore vaccination (Hep A+B, DTP) is

mandatory. Not having valid vaccinations means they need to be arranged, via Nutricia

Research, at the latest 6 wks before start.


19

Part 4: New and/or improved ingredients for food

Use of enzymatic crosslinking to produce protein nanoparticles (dairy subject)

To obtain good product properties, one can choose different types of proteins, coming

from different sources. An alternative is to ‘pre-process’ the proteins to produce protein

isolates with altered functionality. In addition to traditional steps (heating, Maillard),

enzymatic crosslinking can be used in this way.

Currently, peroxidase enzymes have been used to crosslink different

types of proteins. Initial experiments have shown that protein nano-

particles of different sizes (10-80 nm) can be obtained. For these

nanoparticles a significant increase in foam stability was observed. One

of the possible reasons for this is that the particles get stuck between

air-bubbles in a foam, similar to fat globules as in for instance ice-cream.

The aim of this project is to obtain insight in how the size and structure

of crosslinked protein nanoparticles (1) can be controlled by the reaction conditions, and

(2) affect the foam stabilizing properties of the protein.

Thesis projects:

Comparison of heat-induced aggregates to enzymatically cross-linked proteins.

Understanding the relation between reaction conditions and the structure / size of

the formed products (protein nanoparticles)

Study the use of alternative enzymes / substrates in the formation of protein

nanoparticles with different properties.


20

Understanding polysaccharide interactions in fruits and vegetables

The primary plant cell wall in fruits and vegetables is composed of a complex mixture of

pectin, hemicellulose and cellulose. It is proposed that in the current cell wall model two

separate networks co-exist: a

pectic network and a

hemicellulose/cellulose

network. For long time it was

believed that these two

networks are independent but

more and more evidence is

found that interactions

between these 2 networks

exist. However, the nature of

the interactions and the cell

wall components involved are

unknown, species-dependent,

and both hints for covalent and

non-covalent interactions are

found.

Therefore the aim of this project is to understand the architecture of the plant cell wall:

are these polysaccharides linked to each other and if they are, by what kind of linkage?

Thesis projects:

In this project we will address this question in several approaches:

Enzymatic degradation: by using purified enzymes we degrade the hypothesized

crosslink between the pectin and hemicellulose/cellulose network and characterise

solubilised polysaccharides.

Chemical extraction: degrading specific linkages by cross-link degradation and

characterisation of solubilised polysaccharides

Both topics are addressed by characterisation of the extracted material with techniques

such as HPSEC, HPAEC, UPLC-MS and MALDI-TOF-MS.


21

IsoMalto/Malto-Polysaccharides (IMMPs): novel polysaccharides from starch.

Starch is a natural polysaccharide that makes up a large part of the human diet. The

compound itself is biodegradable and used in a wide variety of applications. Due to its

relatively low price and its high availability starch is an attractive substrate for

(enzymatic) modification.

The discovery of the GtfB enzyme (4,6-α-glucanotransferase) opens up a new way to

modify starch. This enzyme is able to alter the intrinsic properties of starch by cleaving

α-1,4 glycosidic linkages and introducing α-1,6 glycosidic linkages into the

polysaccharide.

The products of this enzymatic modification are named IsoMalto/Malto-Polysaccharides

(IMMPs), due to the nature of their linkage composition. IMMPs can be considered as a

new generation of polysaccharides with added functionality for products in and outside

the food industry, such as: food fibres, nutraceuticals, texturizers, replacement of

synthetic polymers, biomedical materials and many other possible applications.

The aim of this project is to make the carbohydrates, to map the properties and to

design optimal products for further applications.

Thesis projects:

In depth chemical analysis of novel polysaccharides, including; molecular weight,

ratio of α-1,4: α-1,6 linkages and directed (enzymatic) modification.

Physical analysis of novel polysaccharides, including; viscosity, elasticity, gel

strength, glass transition point and directed (enzymatic) modification.


22

Part 5: Educational development

Design and/or evaluation of digital learning material for Food Chemistry

Since 2000, the Laboratory of Food Chemistry has been designing and developing digital

learning materials. These materials are used in different courses in the field of food

chemistry. With computer applications, learning material can be designed that can reach

certain learning goals in a more efficient way compared to traditional teaching methods.

The possibilities of digital techniques are amongst others easy visualization, incorporation

of movies or animations, the possibility to personalize the computer program, giving

immediate feedback or hints, and interaction, motivation and activation of students.

This project deals with three different fields: Food Chemistry, Didactics, and Information

and Communication Technology (ICT). It is the combination of these three fields that

makes this project interesting. The goal is to design and/or evaluate useful digital

learning materials. You will learn about educational theories, about using computer

technologies and about a certain subject within food chemistry.

Thesis project:

Designing digital learning material for a course in the field of food chemistry.

Depending on the interest of the student and the needs of the teachers we can

describe the topic for the thesis project together. Contact [email protected] if

you want to know more about this project.

Developing new learning activities or improving existing learning activities

The Laboratory of Food Chemistry is continuously working to improve our education. One

important development is to develop activating learning activities. Learning activities are

for example laboratory activities, group work, class room activities during lectures,

training questions within readers, etc.

This thesis subject deals with two different fields: Food Chemistry and Didactics. You will

learn how to design and evaluate useful learning activities, by incorporating education

theories, for a certain subject within the field of food chemistry. This means that you will

also gain knowledge about that specific food chemistry subject

Thesis project:

Designing learning material for activating learning activities for a course in the

field of food chemistry. Depending on the interest of the student and the needs of

the teachers we can describe the topic for the thesis project together. Contact

[email protected] if you want to know more about this project.

mailto:[email protected]

mailto:[email protected]

information on bsc and msc thesis projects at the laboratory ......bsc and msc thesis projects –...

Documents