nanotechnology in food production - a potential risk or a risky potential

8/13/2019 Nanotechnology in Food Production - A Potential Risk or a Risky Potential

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Aidan NoormanUtrecht UniversityDecember 2008

Supervisors:

Raymond Pieters, PhDInstitute for Risk Assessment SciencesUtrecht University, Utrecht, the Netherlands

Nico van Be!en, PhDInternational Life Science Institute, Brussels, Belgium

Nanotechnology in food production: apotential risk or a risky potential?

"n the potentia o# emusi#ication techni$ues to%enerate nanostructures #or the use in #ood and the

ris& and sa#ety considerations


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Nanotechnoo%y in #ood production: a potentia ris& or a ris&y potentia' ( i

Nanotechnology in food production:A potential risk or a risky potential?

"n the potentia o# emusi#ication techni$ues to %enerate nanostructures#or the use in #ood and the ris& and sa#ety considerations

)aster thesis by:

Aidan Noorman)Sc pro%ram *o+icoo%y and nvironmenta -eath.acuty o# Biomedica Sciences, Utrecht University, Utrecht, the Netherandse-mail: /ANoorman1studentsuun
mailto:[email protected]:[email protected]


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Nanotechnoo%y in #ood production: a potentia ris& or a ris&y potentia' ( ii

Abstract

The purpose of this report was to investigate the safety assessment of emulsification techniques and the variety

of emulsion-based nanostructures (EBNS) that can be obtained and subsequently used in the food industry

Environmental issues will not be addressed

The number of nanotechnology applications is increasing and it is e!pected that the food industry will be the

newest field in which nanotechnology will be applied " conservative si#e definition of $%% nm was chosen for

ris& assessment purposes 'ells are capable of ta&ing up nanostructures of up to $%% nm in si#e and

nanostructures can be engineered with certain properties that could mimic effects of smaller si#ed

nanostructures

evices to produce emulsions and EBNS are already used in the food industry (homogenisers) while other

methods are still being developed which are more efficient (low-energy methods) " large variety of

nanostructures can be obtained with emulsification techniques such as simple emulsions lipid nanostructures

solid nanostructures etc

*ery little is &nown about the effects of nanostructures on the gastrointestinal tract Nanostructures in the body

mostly accumulate in the liver and &idneys where the effects are the most pronounced Surface properties are

very important as they can determine the fate function and possible ris&s of nanostructures +ore research is

needed in which other non-metallic and non-carbon-based nanostructures are (orally) tested

,ecommendations for a food nanostructure-specific ris& assessment include the goal of the nanostructure

(intended or unintended e!posure) consideration of physicochemical properties relevant to food nanostructures

(eg solubility) the history of safe use and a minimum amount of testing to ensure safety " ris& assessment

paradigm is proposed which incorporates these recommendations


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Nanotechnoo%y in #ood production: a potentia ris& or a ris&y potentia' ( iii

Table of contents

1. INTRODUCTION-----------------------------------------------------------------------1

2. EMULSIONS AND APPLICATIONS-----------------------------------------------4

3. SAET! ASSESSMENT O NANOSTRUCTURES IN OOD--------------12

4. CONCLUSIONS------------------------------------------------------------------------22

". REERENCES--------------------------------------------------------------------------23


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ntroduction ( 3

1. Introduction

Nanotechnology and nanoscience deal with the development application and production of materials with si#es

less than %% nm +aterials with such dimensions posses unique properties that are not e!hibited by larger

structures of the same material ./berd0rster et al 1%%$b2 ,esearch and development into nanotechnology is anincreasing business 3n the 4S the budget of the National Nanotechnology 3nitiative (NN3) has increased from

5676 million in 1%% to 56 billion in 1%%8 and is estimated to increase to 59 billion in 1%%: .NN3 1%%;2 3n

Europe funding is mostly provided by national governments and totals around or now many scientists and companies are more interested in the technological and

economical benefits that nanotechnology may hold for the future .,enn and ,oco 1%%72

evelopment of nanotechnology occurs in many different fields and disciplines such as electronics cosmetics

pharmaceutics agriculture etc /ne of the newer fields where nanotechnology is ma&ing an entrance is food 3t is

&nown that a large part of the general population is already e!posed to nanotechnology via cosmetics .Thomas

et al 1%%72 and now an even larger part may be e!posed through the food chain 3n >ebruary 1%%; an online

survey by the ?oodrow ?ilson 3nternational 'entre for Scholars (??3'S) identified over 7%% products that

contain some form of nanomaterial .??3'S 1%%;2 /f the 7%% products 7: where found to be related with

food this includes mostly supplements storage devices and &itchen equipment but this is e!pected to be more

since this is only an online survey and no supermar&et products have been tested

There are four ma@or areas in food production to which nanotechnology can be appliedA food processing

pac&aging and storage food safety and functional foods (food with additional heath-promoting functions) >ood

pac&aging and storage is so far the most promising area for nanotechnology and some of the innovations include

o!ygen barriers .Erlat et al 1%% Sorrentino et al 1%%82 antimicrobial films .,him et al 1%%72 and thermalbuffering techniques for perishable foods .Cohnston et al 1%%;2 >ood safety mostly includes detection and

elimination of foodborne pathogens and to!ins .Branen et al 1%%8 Dang et al 1%%;2 Nanotechnology in food

processing and functional foods is raising both interests and concerns because this involves direct application

into the food itself The use of nanostructures in food can affect the appearance smell and taste of a product

.raveland-Bi&&er and de Fruif 1%%7 ,odrGgue# =atino et al 1%%;2 or reduce costs andHor environmental

burden .*inc#e and *atai 1%%62

Some of the techniques and principles to generate nanostructures are actually not new "n e!ample is the

generation of nanofibers /ne way to ma&e nanofibers is through a process called electrospinning The

technology for this was already invented in the early :%%s for polymer filaments and in the ::%s the technique

was refined to produce nanofibers .Ii and Jia 1%%62 ?ith these fibres it was possible to produce nanofilters

.,ama&rishna et al 1%%72 that eventually spawned the technology for membrane emulsification .'harcosset et

al 1%%6 Fampers 1%%82

This report will focus on nanostructures that can be obtained through emulsification Emulsification is the

mi!ing of two immiscible fluids and is a process that has been used for a very long time in food processing

Emulsification is mainly used in dairy products and beverages but nowadays it is also possible to create

nanostructures with it The capabilities of older techniques and principles to generate nano-emulsions and

emulsion-based nanostructures (EBNS) will be investigated for their use in food "s EBNS can have various

compositions and characteristics .aufin et al 1%%2 different ris&s have been associated with nanostructures

Therefore the emphasis of this report will be on the safety assessment of EBNS The first half of this report will


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ntroduction ( 2

provide a short elaboration of the methods to generate nanostructures and their possible impact on food The

second half of this report is on the ris& assessment aspect for nanotechnology in food " ris& assessment was

recently developed by the Scientific 'ommittee on Emerging and Newly-3dentified Kealth ,is&s (S'EN3K,)

that applies to nanotechnology in general 'ritical views and recommendations will be provided based on

possibilities of EBNS in order to ma&e the tiered ris& assessment more specific to the food industry

1.1 S#$e %ef#n#t#ons

3n the scientific community there is a general tendency to define structures with at least one dimension that

measures less than %% nm as a nanostructure >or to!icological and safety assessment purposes this definition

may not be sufficient 'ells are capable of ta&ing up particles of 9%% nm this include modes not involving

receptors Iately a process has been described that can even ta&e up $%% nm particles .arnett and Fallinteri

1%%72 "lso the LengineeredM aspect of %% nm engineered nanostructures can give similar capabilities as O%%

nm nanostructures So in order to set a strict definition of the term LnanoM that can be used throughout the whole

scientific community a consensus will have to be reached between to!icologists and scientists dealing with

structural engineering of nanostructures

+a&ing agreements on a general definition of the term nano is not easy even in a specific field There are

several reasons for this /ne is the fact that the si#e definition of the word nano differs between scientists

Emulsions are categorised based on their droplet si#es >rom the commonly used definition of nano we would

e!pect that Lnano-emulsionsM are O%% nm in their droplet si#es Several scientists do indeed use this definition

.+eleson et al 1%%62 but there are also other scientists that employ a different si#e (range) such as 1%-1%% nm

.Solans et al 1%%$ Tadros et al 1%%62 or even %%-%%% nm .?indhab et al 1%%$2

3n the field of emulsion research the label Lnano-emulsionM where we assume approaches the general definition

of LnanoM (O%% nm) there are multiple synonyms such as mini-emulsion submicron emulsions ultrafine

emulsion and microemulsions .Solans et al 1%%$2 Iately the term nano-emulsion has been gaining popularity

due to the LnanoM-hype in society and the scientific community Thus not only is the very definition of the term

nano being debated also the terminology has not been standardised

3n light of all these aspects it will be hard to define a LtrueM nano-emulsion " search in the literature for the term

Lnano-emulsionM and several of its synonyms yielded nanostructures with average si#es that were above the %%

nm 'onsidering this result the upta&e capacity of cells and the fact that nanostructures can be engineered to

have certain properties it would seem appropriate to also include the structures that are %% nm and larger in

si#e

The ne!t step would be to determine a proper upper limit >rom a to!icological and ris& assessment point of

view upta&e si#es of 9%% nm or even $%% nm are an important aspect to consider >urthermore a distinction is

often made between smaller (O$%% nm) and larger ($%% nm) micron particles ./berd0rster et al 1%%$b *ega-

*illa et al 1%%;2 while others do consider $%% nm as the upper limit ."nton et al 1%%;2 >or now a

conservative upper limit of $%% nm will be used for food ris& assessment purposes

The idea of implementing nanotechnology in food is new and so we can e!pect that the number of literature

sources with respect to engineered nanostructures in food to be limited 3n this report we will describe EBNS less

than $%% nm in si#e that may be suitable for food applications >or the assessments of potential dangers both in

vitroand in vivostudies of EBNS (O$%% nm) are used Kowever other non-EBNS studies are used in order todescribe effects of various properties "lso pharmaceutical studies with nano-emulsion formulations will be


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musions and appications ( 5

2. Emulsions and applications

Emulsion is the mi!ture of two immiscible liquids where one liquid is dispersed (dispersed phase eg in

droplets) in the other (continuous phase) " nano-emulsion is an emulsion with droplet si#es that range up to

several hundred nm >or reasons discussed in the introduction the upper limit of $%% nm will be used Nano-emulsions can present characteristics and properties which depend on the composition and preparation method

3n this chapter a general principle of nanostructure generation will be e!plained Ne!t several methods to

generate nanostructures will be described "n elaboration will be given on the &ind of nano-emulsions and

EBNS that can be obtained by these methods The chapter will be concluded with a critical view on what aspects

of nanostructure generation and developments can be considered new

2.1 Nano fabcat#on

3n general the production of nanostructures can be accomplished in two waysA via a bottom-up approach or a

top-down approach The top-down approach is the down-si#ing of macrostructures This is usually accomplished

by applying a force in the form of shear and impact evices capable of this are milling devices and

homogenisers "dvantage of this method is that the amount of force and thus the si#e can be easily controlled

Top-down approaches are currently the most widely used methods in large scale industries .Sanguansri and

"ugustin 1%%72

The bottom-up approach deals with the manipulation andHor controlled (self-)assembly of individual molecules

to create nanostructures This method of production is not new in natureA in the body proteins and en#ymes are

formed by amino acids which themselves are controlled by ribosomes E!amples of man-made assembled

nanostructures are the various carbon nanostructures such as nanotubes and fullerenes This process is more

tedious and more susceptible to failure than the top-down approach due to the heavy dependence on thephysicochemical properties and the environment in which the reactions ta&e place The advantage however is

that there is a far greater chemical variety .Seeman and Belcher 1%%12 3t is e!pected that the use of this

approach will further increase as nanotechnology matures .Sanguansri and "ugustin 1%%72

2.2 E'(ls#ons as te')late

The creation of nano-si#ed structures can be accomplished in many

different ways /ne of them is by basing it on an emulsion-

template Emulsions are a versatile system that can be used for

many different products including standard food emulsions

formation of nanospheres or powders nanobeads for processing or

flavour enhancement etc .*ladisavl@eviP and ?illiams 1%%$2

'haracteristics of an emulsion depend for a great deal on the si#e of

the droplets >or e!ample emulsions with large droplets ($%% nm)

have a mil&y appearance whereas emulsions with small droplets

(O1%% nm) are clear (seefig )

There are many different &inds of emulsions and many different

ways to produce them Kere an e!planation is given of some of the

methods that are capable of producing nano-emulsions Several ofthese methods are already in use by the food industry but e!planations are also given of some new methods that

Figure 1+ampe o# the di##erence bet6een a 47nm nanoemusion 9A and a norma 3000 nmemusion 9B )odi#ied #rom ;Soans et al, 2007


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can serve as a potential alternative to the current ones Emulsions used in the e!planations here are the standard

emulsion consisting of water and oil

2.2.1 High-energy methods

The methods of producing nano-emulsions can be roughly divided into two categoriesA high-energy and low-energy methods Kigh-energy methods are now being used in the industry The name is derived from the fact

that the energy that is applied to brea& up the droplets is very high but this method may not always be efficient

.Tadros et al 1%%62 Two high-energy methods will be e!plained that are currently used in the food industry

11 Kigh pressure homogeni#ation

Kigh pressure homogenisation is a technique that employs shear stresses to brea& up droplets The fluid with

(preformed) emulsions is forced through microchannels where e!treme shear stresses are generated due to the

sudden restriction of the flow under high pressure combined with acceleration compression turbulence impact

and eventually rapid pressure drop >igure 1shows several different types of high pressure homogenisation

systems which all wor& with the same principles .Schultz et al 1%%62 +icrofluidi#er systems are the most

effective in producing nano-emulsions .+eleson et al 1%%6 =innamaneni et al 1%%92

Figure 2 Di##erent types o# hi%h pressure homo%enisationsystems )odi#ied #rom ;Schut et al, 2005


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musions and appications ( =

4ltrasonication is a technology that has been used on a laboratory scale for many years But recently (O$ years)

ultrasound devices have been developed for industrial uses

2.2.2 Low-energy methods

Beside the more traditional high-energy methods for nano-emulsions there are also methods that do not requirelarge amounts of applied energy the so-called low energy methods Three of such methods have been described

in the literature and will be briefly e!plained here as potential alternative to the high energy methods etailed

descriptions of the processes governing the reactions are beyond the scope of this report and only basic

e!planations will be given

111 The solvent displacement method

Solvent displacement utilises the physicochemical properties of components to form nano-emulsions Solvent

displacement consists of three components >irst a solute (eg oil) is dissolved in a solvent (eg ethanol) and

afterwards a third component (eg water) is added The third component is miscible with the solvent but not with

the solute uring the addition of the third component small droplets are formed containing the soluteHsolvent

mi! Spontaneous emulsification can occur when diffusion of the third component into the soluteHsolvent droplet

causes the solvent to be displaced from the droplet leaving behind small nuclei of the solute Since the third

component is immiscible with the solute nano-si#ed emulsions are rapidly formed

This method of producing emulsions was termed the Lou#o effectM because the original authors compared the

processes with the addition of water to ou#o a common alcoholic beverage in reece .*itale and Fat# 1%%92

The advantage of solvent displacement is that it does not require surfactantsHemulsifiers "lso the whole process

occurs rapidly and over the entire volume which ma&es it interesting for industrial applications

1111 =hase inversion temperature=hase inversion temperature (=3T) utilises the physicochemical properties of the surfactant to obtain nano-

emulsions or EBNS 3t is based on the interruption of the transition of an oil-in-water (/H?) phase into a water-

in-oil (?H/) phase >or instance when the temperature rises the surfactant is modified in such a way that it

e!hibits equal affinity for both fluids (eg water and oil) This is when the /H? phase starts to go over into a

?H/ phase uring the transition between phases nano-emulsions are formed due to low interfacial tension

Kowever coalescence is e!tremely fast which will eventually result in a coarse emulsion .Solans et al 1%%$2

To prevent coalescence the temperature is suddenly and rapidly lowered in order to preserve the current state

(ie the transition state where nano-emulsions have formed) ue to the temperature change the surfactant

e!hibits normal affinity for one of the fluids ."nton et al 1%%;2

1119 +embrane emulsion

+embrane emulsification does not involve physicochemical properties of the components but instead uses a

membrane with nanopores " liquid-to-be-dispersed is slowly forced through the membrane +echanical

agitation usually in the form of a flow is used to detach the newly formed droplets at the membrane pores

+embrane emulsification requires less energy than high pressure homogenisation and ultrasound ma&ing it an

interesting option for large scale use " disadvantage is the low flu! of the liquid-to-be-dispersed through the

membrane which means a longer production time .'harcosset et al 1%%6 *ladisavl@eviP and ?illiams 1%%$2


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musions and appications ( >

2.3 A))l#cat#ons of e'(ls#on base% nanost&(ct(&es

The methods described above are capable of producing a wide variety of nanostructures "side from the usual

emulsions various other nanostructures can be obtained >igure 9shows several nanostructures that can be

obtained based on an emulsion template 3n this section an e!planation is given of the general features of the

various EBNS and how they can be obtained

Figure Di##erent types o# emusions and BNS obtainabe throu%h variousmethods o# emusi#ication )odi#ied #rom ;?adisav@evi and iiams, 2007


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encapsulation purposes .+ellema et al 1%%7 Su et al 1%%;2 The preparation of such emulsions is first done

with high energy methods to obtain as small as possible inner-emulsions and afterwards methods that are gentler

for the final emulsion are used The use of gentle methods is done to prevent the LinnerM emulsion from

coalescing /ther fragilities in this system are ris& of oil droplet coalescence rupture of oil droplets and lea&age

to and from the inner emulsion .*an er raaf et al 1%%$2 +embrane emulsion seems to be the best suitedemulsion technique for the preparation of the final emulsion

2.3.2 Nano-emulsions as carriers nanoca!sules

Nano-emulsions can be engineered in such a way that they can function as carriers "side from being used for

improved product features nano-emulsions can now be used during preparation processes or as nutritional

additives There are many different forms of nano-carriers including the normal emulsions double emulsions

and encapsulated emulsions

Normal emulsions (/H? and ?H/) are the simplest e!ample of a carrier The desired content can be easily

included within the emulsions provided that the content is miscible with the to-be-dispersed phase Kowever

most emulsions will not last long in the intestine due to the harsh conditions present there Therefore most

emulsions are encapsulated with proteins or polymers .'hen et al 1%%7a2

There are several different techniques to encapsulate emulsions /ne of them is the layer-by-layer technique

(IbI) IbI is based on application of oppositely charged molecules .echer ::82 " primary emulsion is

made that e!hibits a certain charge at the surface of the droplet /ppositely charged molecules are then added

which will adsorb to the droplet surface due to electrostatic attraction " second layer can be applied with an

opposite charge The cycle can be repeated if necessary

/ther encapsulation techniques include the incorporation of the encapsulation-monomers or -proteins in the

droplet before emulsion formation "fter an initiation signal usually temperature the monomers precipitate and

migrate to the waterHoil interface where polymerisation occurs as a result of a reaction with the water or a

component in the water .Tiar&s et al 1%%2

"n advantage of these encapsulation techniques is that they can be applied after the emulsion has formed /lder

methods used solid particles on which the different layers were applied To use the capsule the core had to be

dissolved which required harsh conditions ,emoval of the dissolved core is also not always fully achieved The

resulting shell should also be able to open up if it is to be used as a delivery system .Su&horu&ov et al 1%%62

Encapsulation of emulsions avoids these problems by loading before the shell is formed

2.3.3 Nanoli!osomes

Nanocapsules can also be formed using lipid bilayers ie nanoliposomes 3n general nanoliposomes can be

obtained through high- and low-energy techniques >or low energy techniques liposome generation is based on

the =3T method where the addition of water has a temperature that is below the melting point of the surfactant

The addition of water allows for the crystallisation of the shell thus creating a capsule around the emulsion

.Keurtault et al 1%%12

"dvantages of nanoliposomes are that they can be readily made from natural or food-grade lipids They can also

be used to target specific areas in the food matri! .+o#afari et al 1%%72 Nanoliposomes are already used as a

food processing aid .=iard et al :;72 and newer applications will mainly be as carriers in the food additives

business +ore advanced forms of lipid nanocapsules are multi-vesicular lipid capsules These are similar to the

double emulsions but now in the form of liposomes .+o#afari et al 1%%72


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musions and appications ( C

2.3." #olid nano!articles

Kigh- and low-energy methods are also produce solid nano-si#ed particles This class of structures consists of a

homogeneous structure throughout the whole particle whereas the nanocapsules have a (hollow) core-shell

structure Nanoparticles can function as carriers by imbedding the desired substance into the particle itself

.3shihara et al 1%%;2 or by attaching it to the surface of the particle ."tyabi et al 1%%; 'hauvierre et al

1%%92

=roduction of solid nanoparticles is done by dissolving monomers which are used as the hydrophobic component

of the mi!ture @ust li&e oil is in water "pplication of a high-energy method will cause the monomers to

polymerise which will then form droplets =olymerisation is accomplished by molecules that are added to the

mi!ture These molecules start reacting with the monomers causing them to form polymers .Thic&ett and

ilbert 1%%82 roplets are entirely composed of polymerised monomers and are stabilised by surfactants

thereby becoming (solid) nanoparticles

"nother type of solid nanoparticles is the solid lipid nanoparticle (SIN) These particles already have a large

potential in the pharmaceutical industry as carrier =roduction is simple and is usually done with high-energy

methods .*ladisavl@eviP and ?illiams 1%%$2 Basically the lipid is mi!ed with water or another immiscible

fluid at a temperature that is above the melting point of the lipid The mi!ture is put through a high-energy

device and subsequently cooled down to crystalli#e the lipids "dvantages of SIN are similar to the solid

nanoparticles with respect to carrier capabilities SIN can be made from lipids which ma&es them potentially

less bioto!ic than solid nanoparticles

2.3.$ %olloidosomes

'olloidosomes are emulsions with nanoparticles loc&ed together on the interface forming a shell (fig 6)

.insmore et al 1%%12 Both components (inner-emulsion and shell nanoparticles) can be made from emulsions

Shell formation can be prepared through electrostatic interactions This preparation can be considered surfactant-

free to a certain e!tent .Sa&ai 1%%;2 'olloidosomes as a whole structure cannot be considered a nanostructure

because their si#e is far greater than $%% nm and can even range into the %-%% m But the nanoparticles that

form the shell can be well below $%% nm

'olloidosomes can be theoretically multifunctional with the nanoparticles on the shell as well as the inner-

emulsion itself acting as a delivery system Kowever reports of its use have not yet appeared which is most

li&ely due to this being a recent development


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Figure ! ooidosome, an advanced product o# emusions "n the e#t is a schematic vie6 o# a cooidosome and on theri%ht scannin% eectron microscope ima%e o# a 30 Em vacuum dried cooidosome )odi#ied #rom ;Dinsmore et al, 2002


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Nanostructures are becoming more comple! in order to perform certain functions Nanostructured coatings

dispersion (eg emulsions) and nano-bul& materials such as nanometals -ceramics and -polymers are fairly

simple compared to the structures that are currently being designed So-called second generation nanostructures

are more comple! forms of the older nanostructures .,enn and ,oco 1%%7 ?eiss 1%%82 These include the

double emulsions multi vesicular lipid nanocapsules SIN and colloidosomes +ost of these systems can be

distinguished from the first generation by their multi-functional capability and because of this they are most

li&ely capable of more comple! interactions during food production andHor in the body .,enn and ,oco 1%%7

Sanvicens and +arco 1%%;2

3n summary nanostructures are naturally present in our food 3n the past attempts to improve food

characteristics unintentionally lead to increased nanostructure e!posure whereas nowadays nanostructures are

actively introduced in order to improve food characteristics New methods are being developed to generate

increasingly more comple! nanostructures ue to the possibility of more comple! interactions of nanostructures

e!tra attention may be needed for their safety assessment especially in the food industry where EBNS can be

used as food additives processing aids or as a nutritional supplement


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Sa#ety assessment o# nanostructures in #ood ( 32

. "afety assessment of nanostructures in food

3.1 Potent#al s+s

+any people have doubts about nanotechnology concerning its safety because so little is &nown 3f

nanotechnology is going to be used on a large scale in the food industry then the ris&s that can be associated with

nanostructures will need to be characterised This section will focus on the identification of possible ris&s of

nanostructures that can be used in the food industry =ossible ris&s and &nown effects will be identified from the

preparation methods up to the immune system 3n recent years numerous papers have described the ris&s of

engineered nanomaterials E!amples of these materials are carbon nanotubes fullerenes and metallic

nanoparticles "lso in the pharmaceutical field are there many reviews on ris& assessment of nanomaterials and

delivery systems "ssessments of nanoformulations used in the food industry however have hardly been

performed ue to the lac& of food ris& assessments of nanostructures ha#ard assessments of nanoformulations

in the (oral) pharmaceutical research will also be used

3.1.1 &m!ortance of !re!aration methods

The composition and components of the nanostructures are important determinants of to!icity " comparison of

EBNS made of non-to!ic components and potentially to!ic components such as cationic molecules indicate

to!icity related to the materials and not to the fact that it is an EBNS .?eyenberg et al 1%%82

Surfactants are an important component of EBNS which influence particle si#e and distribution Kigh pressure

homogenisation and ultrasound generally have a broad si#e distribution which is often overcome by adding more

surfactant 3f the surfactant is &nown to have adverse effects at relevant doses then the preparation method may

be a cause for concern 'ertain preparation and purification methods can help reduce to!icity mainly by

removing e!cessive amounts .Keydenreich et al 1%%92 /ne disadvantage is that these preparation methodsthemselves can affect the si#e properties of the EBNS so careful consideration is needed for a balance between

ris&s and benefits

" good preparation can also avoid adverse effects due to impurities Non-food grade materials are often used to

solubilise or dissolve ingredients which may become entrapped together with the active ingredient .+o#afari et

al 1%%72 3t has been shown in vitrothat metal impurities in carbon nanotubes are responsible for the generation

of reactive o!idative species and inflammatory reactions in lung cells and macrophages =urified nanotubes did

not display such effects .=uls&am! et al 1%%82 Thus beside the intrinsic to!icological properties of the

components preparation methods of EBNS are equally important to consider

3.1.2 Nanostructures in the gastrointestinal tract

The gastrointestinal tract (3T) is the first ma@or contact point with nanostructures The first large obstacle is the

stomach where the high acidity cleavage en#ymes and the mucus layer can prevent nanostructures from entering

the body But this does not provide a full protection 4pta&e of various nanostructures was observed throughout

the 3T .esai et al ::7 Cani et al ::6 ?ang et al 1%%7 ?ang et al 1%%82 4pta&e occurs mostly

through specialised membranous epithelial (+) cells in the =eyerMs =atches .esai et al ::7 Cani et al ::62

/ther cells and mechanisms such as enterocytes and paracellular transport respectively also ma&e it possible

for nanostructures to pass the intestinal epithelium .Iomer et al 1%%12

The 3T is capable of ta&ing up particles of up to $% m in si#e .Kussain et al 1%%2 which is well above thesi#e of nanostructures Kowever upta&e of nanostructures is dependent on the nanostructure and its surface


17/27


properties (see subsection 96) /ral administration of carbon-based nanotubes fullerenes and quantum dots

were found not to be ta&en up by the intestines and were eventually recovered in the faeces .eng et al 1%%8

Nel et al 1%%72 indicating that si#e is not the main factor driving upta&e

Effects of nanostructures on the 3T are not yet fully &nown /nly slight inflammation of the stomach and

intestines in mice was observed after oral administration of nano #inc and titanium dio!ide (Ti/ 1) .?ang et al

1%%7 ?ang et al 1%%82 3n humans it is thought that e!posure to nano-si#ed structures can contribute to 3T

diseases such as inflammatory bowel disease and 'rohnMs disease .Iomer et al 1%%12 /ther than some

inflammation no other adverse effects of nanostructures in the 3T are &nown etailed ha#ard characterisations

of the 3T and the &nowledge of the effects on susceptible people will be essential before engineered

nanostructures are allowed to be used in food

3.1.3 'iodistribution

"fter being ta&en up in the 3T nanostructures will distribute throughout the body Studies of nano Ti/ 1and

#inc indicate that the liver is the primary site of accumulation .Cani et al ::6 Sugibayashi et al 1%%; ?ang

et al 1%%72 =rominent effects are hydropic degeneration and slight necrosis of hepatocytes >urthermore

increased levels of "IT "ST "I= IK and cholesterol esters indicated liver in@ury and dysfunction in mice

.?ang et al 1%%7 ?ang et al 1%%82 Ti/1 however is easily eliminated from the body .Sugibayashi et al

1%%;2 3n a safety evaluation of Ti/1nanoparticles in food most of the Ti/1accumulated in the liver and was

gradually eliminated from the body (mice 9%U decrease month after administration) No symptoms or

noticeable behaviour changes were observed

Fidneys are the second largest site of accumulation for Ti/ 1and #inc nanoparticle Kistopathological effects are

swelling of the renal glomerulus and proteinic liquid in the renal tubules .?ang et al 1%%7 ?ang et al 1%%82

/ther minor accumulation sites include the spleen heart and lung tissues The heart tissue showed no

morphological alterations after oral e!posure of Ti/1and #inc but blood serum levels of IK and V-KBK

were higher compared to controls .?ang et al 1%%7 ?ang et al 1%%82 IK and V-KBK are often used as

mar&ers of cardiovascular damage Ti/1 and #inc also accumulated in the spleen and lung but no

histopathological effects were observed .Cani et al ::6 ?ang et al 1%%82

3n summary the main sites of accumulation are the liver and &idneys where the effects are also the most

prominent /ther organs may also be affected to a lesser degree Ti/1was shown to be gradually eliminated

from the body and thus effects may be reversible These studies were done with nano Ti/ 1and #inc but no

chronic oral e!posure studies of other food nanostructures and EBNS are &nown

3.1." (ffects of surface !ro!erties

Surface properties of nanostructures are a special issue >irst of all the smaller the particle the higher its

surface-to-volume ratio becomes ie smaller nanostructures have relatively more surface area than larger

structures This ma&es them potentially more reactive Secondly the surface can be engineered to e!hibit certain

properties or functions The combination of these two aspects is a reason why nanostructures in food raise so

many concerns " few e!amples of adverse effects due to surface properties are discussed below

"n important factor for upta&e or internalisation of the particles is surface charge To enhance the upta&epotential of nanostructures their surfaces are often modified with molecules with a certain charge =articles


18/27


.>lorence ::8 +ao et al 1%%$2 and emulsions .Dang and Benita 1%%%2 with a positive charge are more easily

absorbed by the 3T The positive charge however can have a to!ic effect on cells =ositive charged or cationic

molecules can enter cells through various cell mechanisms .Jia et al 1%%;2 or by disrupting the lipid membrane

.Ieroueil et al 1%%;2 " wide variety of commonly used nanostructures was investigated for the potential to

create (nano)holes in the lipid bilayer of cells 3t seems that regardless of the shape si#e composition chargedensity and deformability all tested cationic nanostructures were capable of disrupting the bilayer by either

membrane thinningHerosion hole formation or e!pansion .Ieroueil et al 1%%;2 Kole formationHe!pansion can

lead to lea&age of the cell content and even cell death ?hether cell death is attributed to the lea&age or to the

nanostructure itself is still unclear

4pta&e of cationic nanostructures by cells disrupts certain cell processes and leads to cell death ,ecently 7% nm

cationic polystyrene nanospheres have been shown in vitroto induce mitochondrial damage in macrophages and

bronchial epithelial cells followed by apoptosis and necrosis respectively .Jia et al 1%%;2 4pta&e and cell

death mechanisms differed between the two cell lines Jia et al also tested polystyrene nanospheres on cells of

the liver adrenal gland and microvascular endothelium Nanospheres were ta&en up by these cells but in

contrary to macrophages and bronchial epithelial cells did not induce mitochondrial damage increased 'a 1Wflu!

or lysosomal disruption indicating cell-specific to!icities and mechanisms

The severity of adverse effects can have a charge (as in dose)-response relation irect cell-polymer contact

assays demonstrated increasing cytoto!icity with increasing charge density of polyesters in mouse fibroblasts

.4nger et al 1%%82 Thus a certain dose-response relationship of nanostructures may be set up based on the

charge

The use of cationic molecules as building bloc&s for nanostructures can still be to!ic despite it going through

several processes =rocessing cationic lipids into %%-$%% nm SIN does not reduce their capability to decrease

cell viability .Keydenreich et al 1%%92 Though caution is needed when inferring in vitroresults 'omparative

studies between in vitro and in vivopulmonary to!icity results of nanoparticles demonstrated little correlation

.Sayes et al 1%%82 ?hether this holds true for other non-pulmonary to!icity studies is not &nown

?ith such potential to!ic effects in mind certain aspects of nanostructures will need e!tra caution /ne technique

that deals with surface charge is the IbI technique which ma&es it possible to coat emulsions and EBNS

.echer ::8 il et al 1%%;2 The application of this technique may unintentionally increase the to!icity of

nanostructures and possibly nullifying the intended effect of carrier systems eg reduced to!icity 3t is

interesting to note that the application of positively charged molecules on nanostructures may decrease oral

bioavailability +ucus in the stomach is negatively charged and so positively charged molecules may become

entrapped in it .Koet et al 1%%62 "lternatively some scientists believe that this can also increase intestinal

lifetime and facilitate upta&e by presenting nanostructures to the 3T epithelial cells ."rbXs et al 1%%1 'hen et

al 1%%7a Kussain et al 1%%2 This issue is still under debate 3mportant characteristics of nanostructures

especially surface properties are to a certain e!tent accompanied with negative effects

The intestinal environment with its high acidity and cleavage en#ymes may change the nanostructure surfaces in

a certain way 3nflammatory effects and cell death were induced by pathogen-associated molecular patterns

(="+=) con@ugated to food additive-grade nano-Ti/1 ="+= may inadvertently adsorb to the surface of Ti/1

."shwood et al 1%%82 The gut flora is a potentially large source of ="+=s Thus even though a nanostructure

can be harmless at first in or e) vivoevents may change its current status Table summarises a few of thepotential ris&s that can be induced as a result of in ore) vivo(metabolic) processes and interactions 4ltimately


19/27


the proteins that are present or adsorbed to the surface determine what is actually presented to cells .'edervall et

al 1%%82

Nanostructures in the 3T do not necessarily have to be ta&en up to e!ert an effect on other organs /ne e!ample

is the effect on the pK buffering system .'hen et al 1%%7b2 'hen et al.investigated the effects of nano- and

micro-copper Severe effects (I$%was 69 mgH&g body weight for nano-copper as compared to $%%% mgH&g

for micro-si#ed copper) and damage to liver &idney and spleen where observed 'hen et al.hypothesi#ed that

depletion of KWin the gastric @uice due to higher reactivity of nano-copper lead to the formation of bicarbonate

ions (K'/9-) K'/9

-is hardly e!creted by the &idney due to renal disorders and so accumulation occurs that

could lead to other metabolic and physiologic statesHdisorders such as al&alosis in this case

#able 1 +ampes o# materias used in #ood and in nanostructures that can induce undesired e##ects due to interactionsRe#erences are in vivostudies uness indicated other6ise

$aterial Food use %ossible effect &eference

P/ .ood additive mmune evasion, mayenhance un6anted

biodistribution

;"berdFrster et al,2007bG Sanvicens and

)arco, 2008or e!ample Salman et al. .1%%82 tested

orally administered thiamine-coated and non-coated nanoparticles in mice and showed that the thiamine-coating

elicited stronger 3g and 3g1a titres Non-coated nanoparticles induced a typical Th1 response indicating that

coated nanoparticles are capable of shifting towards a more balanced response This also further indicates the

importance of surface properties ie surface can determine functionality of the nanostructure

Nanostructures may also affect diseases based on deregulation of the immune system Nano metals and ceramics

can bias the defenceHinflammatory capabilities of macrophages .Iucarelli et al 1%%62 Si/1nanoparticles biased

naive macrophages towards pro-inflammatory reactions by selectively inducing production of 3I-Y and TN>-V


20/27

Sa#ety assessment o# nanostructures in #ood ( 3=

Ti/1was shown to increase e!pression of viral TI, receptors TI,9 and TI,8 Nanoparticles made from cobalt

induced a cyto&ine profile that resembles e!perimental and clinical autoimmunity in vitro.=etrarca et al 1%%72

No reports are &nown that focus on nanostructures in food and the effect on susceptible sub@ects eg 3T

disease models

/n the other hand there are reports describing immune suppression by nanostructures 3n a mouse model ofpeanut allergy nanoparticles of plasmid N" and chitosan reduced 3gE levels plasma histamine and vascular

lea&age Secretory 3g" and serum 3g1a were found after treatment .,oy et al :::2 This may imply a

positive effect of (functionalised) nanostructures that can be used in food Kowever this feature may also raise

concerns about undesired and e!cessive immune suppression To date there are no e!tensive reports on this

matter further research is needed

3.1.3 *ther routes of e)!osure

/ther routes such as inhalation may become increasingly more relevant as more food applications are going to

be developed for nanostructures Before food ends up on our plates it will undergo several processes E!posure

through inhalation can occur with coo&ing processes or when the nanostructure is in a powder form Effects of

nanostructures on the respiratory tract have been e!tensively described .ill et al 1%%8 +Zhlfeld et al 1%%;

/berd0rster et al 1%%$b2 3nterestingly it was already &nown si!ty years ago that nanostructures (O$%% nm) can

be ta&en up and transported up the olfactory bulb and nerve ./berd0rster et al 1%%$b2 /berd0rster et al..1%%$2

noted that these studies were performed on rodents and questioned the relevance to humans 3ndeed the human

olfactory mucosa comprises only $U of the total nasal mucosal surface as opposed to $%U in rats Thus relative

e!posure would seem to be far less for humans Nevertheless this still may be a concern for people with pets

/ther important effects on the lungs are in general inflammation and fibrosis generation of reactive o!idative

species various effects on macrophages and cardiovascular effects "gain surface properties can have a large

influence ./berd0rster et al 1%%$b2 +ost of the assessment studies have used metallic or carbon-based

nanostructures and not much emphasis is given to EBNS that are used in the pharmaceutical or food industry

+ore research is needed using EBNS that can also be used in the food industry

" third ma@or route of potential upta&e is through dermal e!posure S&in contact with nanostructures (in food)

can occur during handling or preparation of food ,is& assessments of cosmetics mainly sunscreens and lotions

have shown however that nanostructures do not penetrate the living s&in .Nohyne+ et al 1%%82 +oreover it

was concluded that nano-si#ed structures do not significantly penetrate deeper than micro-si#ed structures

although the definition of micro-si#ed structures was not clearly e!plained by Nohyne& et al

3.2 A foo%-s)ec#f#c s+ assess'ent

3n 1%%8 the E4 Scientific 'ommittee on Emerging and Newly-3dentified Kealth ,is&s (S'EN3K,) developed a

tiered ris& assessment methodology for nanomaterials based on an e!posure assessment algorithm (fig $)

.S'EN3K, 1%%82 The S'EN3K, ac&nowledges that both assessments deal with nanostructures in general and

that there may be a need for more industry-specific ris& assessments Kere several aspects will be discussed that

can ma&e the e!posure assessment algorithm by the S'EN3K, more specific to the food industry


21/27

Sa#ety assessment o# nanostructures in #ood ( 3>

Figure ' +posure assessment a%orithm #rom SN-R ;SN-R,200>ood nanostructures larger than %% nm should also be considered in ris& assessments Nanoparticles in the air

can penetrate deeper into the lungs depending on their si#e Nanostructures in food do not have this limitation

ma&ing it easier to come in contact with cells Though a proper upper limit still needs to be determined

"nother physicochemical property agglomeration may be a potential concern "gglomeration (or coalescence)

can cause nano-emulsions to lose their beneficial effects >ood manufacturers can avoid this by applying

molecules capable of repulsing other particles .Studart et al 1%%82 Kowever such procedures can alter other

properties of the nanostructures .Firchner et al 1%%$2 and in vivo behaviours (eg increased circulation

capabilities) .arnett and Fallinteri 1%%72 consequently introducing new ris&s 3n food ris& assessments it may

therefore be important to assess the effect of anti-agglomeration measures

/ne important aspect of nanotechnology in the food industry is that there can be an intended e!posure

'urrently manufacturers try to minimise the amount of processing and enhance food nutrition >or instance by

reducing the transport medium the upta&e of nutrients in the gastrointestinal tract will be improved .esai et al

::8 Cani et al ::%2


22/27


There are two types of nanostructures with an intended e!posureA ) nanostructures which are involved in

nutritional enhancement This is mostly in the form of carrier systems The goal of such nanostructures is to have

a high as possible bioavailability These structures will concern both occupational and consumer safety

1) Nanostructures which are present in the end-product but have a food-improvement or cosmetic related

function such as enhancement of taste or smell This also includes food spoilage and antimicrobial protection."n et al 1%%;2 These nanostructures should not be ta&en up into the body This may raise concerns for

manufacturers Nano fragrance molecules for e!ample can be ta&en up by cells in the lung and nasal cavity

./berd0rster et al 1%%$b2 Bioavailability of these nanostructures should be as low as possible

Nanostructures that do not have any intended e!posures will most li&ely be used as processing aids or as a

functionalised coating on equipment This can be for e!ample nanocapsules protecting an en#yme during the

production process andHor allowing gradual release of the en#yme afterwards .=iard et al :;72 ?hen these

nanostructures remain in the end-product measures may be necessary to insure that consumer e!posure (and

bioavailability) is &ept at a minimum especially when adverse effects are &nown

&n vitrotesting is well suited as an initial screening for adverse effects but a recent comparison of in vitroand in

vivopulmonary tests indicated little correlation .Sayes et al 1%%82 'haracteristics of nanostructures (eg si#e

si#e distribution charge etc) can be altered by the food matri! processing of the food matri! and the 3T To

describe the effects as accurately as possible in vivotests need to be carried out with the matrices that (will)

house the nanostructures ./berd0rster et al 1%%$a2 "lthough with this approach many complications arise that

may obscure the effects of the nanostructure >eeding high levels of whole foods to test animals can lead to

nutritional imbalances and difficulties with establishing causal relations .'onstable et al 1%%82 To reduce

unnecessary animal testing in vitrostudies should get priority over in vivostudies 3f in vitrostudies indicate

adverse effects at relevant doses then in vivoe!periments should be done to confirm results &n vitroand in vivo

studies need to be developed that correlate with each other

Effects of chronic e!posure are harder to determine than acute effects To provide a certain level of safety on the

long term measures have been devised that ta&e into account a lifetime e!posure /ne measure discussed by the

S'EN3K, is the threshold of to!icological concern (TT') 3t provides guidance in the form of a level of

e!posure or inta&e per person per day below which no significant ris& to humans is e!pected to e!ist The

determination of a TT' value is done by comparing the molecular structure with other structural analogues and

the &nowledge gained from the general to!icity database from the past si!ty years .Froes et al 1%%$ Froes and

Fo#ianows&i 1%%1 +unro et al 1%%;2 This method is beneficial with respect to limited resources time and

e!pertise and is very useful when there is little or no data available of the substance >or nanostructures

however the TT' may not be an appropriate guidance value Nanomaterials are a relatively new phenomenon

and so there are no other substances to compare with 3f nanostructures are made from e!isting components such

as chitosan then a TT' value can be derived from chitosan to!icity (bul&) data But the effect of nano-si#ing

chitosan is not &nown and therefore some minimal amount of ( in vivo) testing may still be required "lso due to

their comple!ity different forms of nanostructures can present different ris&s depending on their manufacturing

process .Iin 1%%82 The lac& of &nowledge of nanostructures will ma&e it difficult to apply a TT' concept in

ris& assessments Fnowledge on nanostructure characteristics and preferably also dose-response relations will

therefore need to be generated first

Because comparison with structural analogues may not always be possible an alternative to the TT' may be

needed The acceptable daily inta&e ("3) may fulfil this role for now The "3 is also a guidance value for the

daily inta&e of a substance over the entire lifetime .Iu :;;2 The difference between TT' and "3 is that the


23/27

Sa#ety assessment o# nanostructures in #ood ( 3C

"3 relies on identification of the highest dose level where no effect occurs and subsequent application of a

safety margin whereas the TT' assures safety based on sufficiently low e!posure (in the absence of specific

to!icity data) .+unro et al 1%%;2 Thus the advantage of an "3 over the TT' is that it is more specific to a

substance The "3 is li&ely to be a precursor to the TT' because it is derived after the determination of adverse

effects The TT' is more of a predictive form of ha#ard assessment and can be more efficient than the "3 if thenanostructure is composed of structures with already e!isting to!icity data "3 values are currently being used

for food additives and @udging of the potentials of nanostructures the first applications will most li&ely be as

food additives

/ne issue that may need addressing in the future but is not e!plicitly mentioned in the S'EN3K, report is the

possibility of multi-functionality The currently &nown nanostructures (nanotubes metallic nanoparticles) are

relatively simple but future nanostructures are e!pected to be more comple! (delivery systems eg

colloidosomes fig 7) .,enn and ,oco 1%%7 Sanvicens and +arco 1%%; ?eiss 1%%82 ,enn and ,oco .1%%72

predict that the ne!t generation of nanostructures will involve assembly and networ&ing systems adding up to

comple!ity and multi-functionality 3n nano-food processing this can possibly include miniature LfactoriesM

capable of several (en#ymatic) processing steps at the same time But should we therefore be afraid of Lmulti-

to!icityM[ This will most li&ely depend on the nanostructure capabilities 3t may be that the safety of multi-

functional nanostructures will have to be @udged on each separate function and its capability as a whole

Figure (+ampe o# a #ictitious muti#unctiona nanostructure .i%ure #rom;Sanvicens and )arco, 2008igure 8proposes a food nanotechnology ris& assessment paradigm that incorporates the aspects discussed

above " sequence of questions is as&ed to determine the need for further e!tensive testing This paradigm builds

on the e!isting ris& assessment by the S'EN3K, (fig $) and incorporates several of its elements >urthermore


24/27


this paradigm is only intended for nanostructures that are in contact with food and does not address

environmental issues

The ris& assessments set up by the S'EN3K, focuses more on physicochemical properties as a rationale for

safety testing (fig $) The proposed ris& assessment on the other hand starts by identifying the purpose of the

nanostructure (ie intended versus unintended) 3f a nanostructure is only used during food production andremoved afterwards from the end-product (ie unintended) then no detailed assessments are needed ( >ig 8 left

column) This is to prevent unnecessary and costly testing The only other concern of these types of

nanostructures is limited to occupational safety

>or nanostructures that remain in the end-product further testing may depend on assessment of the &nown data

on the physicochemical properties (fig8 second column) This is a similar procedure as in figure $ Enhanced

solubility or reduced agglomeration potential can increase bioavailability These and other properties can give a

good indication of the bioavailability potential without doing any e!tensive testing

"lterations to the nanostructure throughout its life should also be ta&en into consideration This includes physical

changes but also adsorption of upta&e enhancing molecules " changing environment li&e the 3T or (domestic)

heating can change such properties and can even create different routes of e!posure Therefore also other routes

such as inhalation should be considered Besides adverse effects due to upta&e effects not related to upta&e such

as irritation or inflammation may also need to be considered

Nanostructures with high bioavailability should be sub@ect to at least some (initial) tests to ensure a minimum

level of consumer safety (fig 8 third column) The initial testing should also include further characterisation of

other un&nown physicochemical properties Effects can be compared to the corresponding bul& material Bul&

material guidelines can be followed only when effects are of the same &ind and less severe at the same mass

concentration This is to avoid manufacturers bypassing further testing as a result of e!isting bul& form data

aps in the legislation should not be a reason to avoid testing even though the bul& form of the nanostructure has

been in use for a long time (eg ,"S history of safe use) " full ris& assessment should be performed when

adverse effects of the nanostructure are more severe than the bul& form or when tests indicate different (new)

&inds of adverse effects


25/27


26/27

oncusions ( 22

!. *onclusions

/ne of the purposes of this report was to investigate the possibilities to generate various nanostructures through

emulsification methods that can be used in the food industry +ethods to produce nanostructures are based on

old techniques and principles +anufacturers are developing more efficient methods to produce EBNS therebyincreasing consumer e!posure

" wide variety of EBNS can be obtained through emulsification These nanostructures can be functionalised to

perform certain functions such as en#yme delivery This aspect differentiates engineered nanostructures from

natural nanostructures which are already present in food in the form of proteins bacteria single molecules (eg

vitamins) etc Nanostructures that are currently being developed are becoming more comple! which enables

them to perform multiple functions These multi-functional EBNS may require special attention in ris&

assessments

The other purpose and main focus of this report was to identify possible ris&s that may arise due to

nanostructures in food and to determine important aspects for a ris& assessment that is specific to food

nanostructures Surface properties are probably the most important aspect of nanostructures to consider Surface

molecules can determine the fate function and possible ris&s of nanostructures There is still very little &nown

on the ris&s of nanostructures especially when it comes to effects on the 3T There have been a lot of in vitro

studies conducted and relatively few (oral) in vivostudies =ulmonary to!icity studies of nanoparticles indicated

little correlation Similar investigations need to be conducted for other non-pulmonary studies "lso most

studies have used metallic or carbon-based nanoparticles and no e!tensive studies have been conducted using

other &inds of nanostructures Effect of nanostructures on hypersensitivity and disease models (as susceptible

sub@ects) will also need more research

,ecommendations for a food nanostructures-specific ris& assessment areA ) to base assessments on the purpose

of the nanostructure ie intendedversus unintendede!posure Some nanostructures are intended to be ta&en up

whereas others are only used during production which does not need e!tensive testing

1) The need for testing based on physicochemical properties differs for nanostructures in food compared to those

used in other industries Solubility can increase upta&e and depending on the purpose may be undesired

"ssessments should also include nanostructures larger than %% nm because cells are capable of ta&ing up larger

si#ed nanostructures Nanostructures can be LengineeredM with certain properties which may have similar adverse

effects as smaller si#ed nanostructures

9) " minimum amount of testing for nanostructures with a potential of being ta&en up (based on

physicochemical properties) whether it is intended or not This is to provide a minimum level of safety >urther

testing if needed can provide guidance values such as "3 and may even set up a to!icity database that can be

used for other nanostructures Such a database will be helpful for a TT' concept which may help in further

reducing (animal) testing

3mplementation of these recommendations in a ris& assessment will ma&e it more specific to for nanostructures

in food The ris& assessment paradigm presented in figure 8incorporates these recommendations in the form of a

series of questions of which the answers determine the need for further testing


27/27

Re#erences ( 24

'. &eferences

nanotechnology in food production - a potential risk or a risky potential

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