protein structures as delivery vehicles in foods

7/29/2019 Protein Structures as Delivery Vehicles in Foods

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Chapter 5ProteinStructuresasDeliveryVehiclesinFoods

Paul Smith1,2 andMarkPlunkett11YK I Institutefor SurfaceChemistry, Box 5607,SE 114 86

Stockholm,Sweden2Current address: Cargill R& D Centre Europe, Havenstraat 84,

B1800Vilvoorde, Belgium

Protein structures are important for providingmany propertiesof foods. Hydrophobins have recently been extracted fromfilamentous fungi. These are amphiphilic molecules withunusual surface-active properties. These can be used in orderto make unique nano-structures in foods and also to givedifferent behaviour in other applications. There are agreatmany opportunities and challenges for food scientists.Structures could be used for incorporation of ingredients or tocreatenovel textures and products.

2009American Chemical Society

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90Introduction

Proteins have many different roles in foods. They are very importantnutritional components of the diet. Certain proteins are surface active. They arewidely used in food systems where a surface-active component is required.Obvious examples are inmilk and dairy products. These are generally low-techfoods and a great deal of specific functionality is hot needed. It is the otheringredients that people have looked for in developing newer foods. Howeverspecific active proteins exist that can be developed and applied to give newfunctionalities and opportunities. One such proteins are the hydrophobins.

Hydrophobins are proteins that exist on the outside of filamentous fungi.They were initially discovered because they were stable to boiling duringextraction (7). This is very unusual for such a complicated material. Thestructure of the proteins has been well characterized (2) and reviews of theliterature have been published. The hydrophobins tend to be found on theoutside of the fungi. The proteins are all about lOkDa in size and contain alarge proportion of hydrophobic amno acids. The main unifying feature is thepresenceof8Cys residues.

Study has shown that different hydrophobins are formed and expressed atdifferent stages of the fungi's life. They are expressed all through the life offungi. The properties and biological roles are fully described in differentreviews (5-5). However it can be seen that they have two main roles. They helpfungi survive and adapt to the environment and they also have various structuralroles.

The 3D structureof a hydrophobin has been discovered relatively recently.The relationship between the structureand the properties is very interesting. Thestructure is illustrated in Figure 1.

It can be seen that the molecule is amphiphilic with one hydrophobic andone hydrophilicpart. This means that when the molecules are placed togetherthey can form different self-assembled structures. This work will describe theproperties of the different structures and suggest ways in which they can beincorporated and used in foods.

ExperimentalThe experimental work was performed using a variety of different pieces of

equipment.Hydrophobin II was extracted and manufactured at VTT Biotechnology,

Espoo, Finland. It was supplied as a gift. Sodiumcaseinatewas purchased fromAr ia Foods, Stockholm, Sweden. Lipids were obtained from Sigma Chemicals.


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In Micro/Nanoencapsulation of Active Food Ingredients; Huang, Q., el al.;

ACS Symposium Series; American Chemical Society: Washington, DC, 2009.


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92Solutions of different quantities of hydrophobin in milli Q water were

prepared and degassed. They were used as fresh. Varying concentrations wereused up to 5mg/ml.

Foam stability was measured using a Turbiscan Classic, Formulaction,Toulouse, France. A ir was bubbled into the solutions and the density of thefoamwas measured for up to 17 days.

Contact angle measurements on the solutions were performed using thependant drop technique.

Surface rheology measurements were performed using a surface rheometerfrom K S V Instruments, Helsinki, Finland. A Langmuir-Blodgett trough wasobtained fromthe same source.

ResultsOn investigation of the samples by Mastersizer we saw very clear evidence

of structuring within the system (Figure 2). This was also apparent afterextensive degassing of the system. Thiswas seen repeatedly. If weconsider thesize of the structures then we see that they appear to be of a scale for smallvesicles, which can agglomerate in the system. The larger structures arepresumablyagglomerations.

Contact angle measurements showed that the hydrophobins were surfaceactive, with surface activity increasingwith concentration (Figure 3). Despitethis, attempts at using the materials for emulsification were unsuccessful. Anextremely large amount of work was given over to the attempt to manufactureemulsions, either water in oi l or oi l in water, but this was ultimatelyunsuccessful. In conjunctionwith other surface-active proteins such as sodiumcaseinate, emulsification was possible. These emulsions did not differsignificantly in character from sodium caseinate stabilized systems and sopresumably it is the sodium caseinate that is responsible for the effect.

However it was found that the hydrophobins had very strong foamingeffects. Foams made at 5 mg/ml of additive were stable for over a month.Decay was measured with a turbiscan and over 17 days only relatively smallchanges were seen with the 5 mg/ml solution. Even at lower concentrations,very good and stable foams were seen that lasted for several days at 0.01 mg/ml.This indicates that the hydrophobins have very strong interactions and aprofanity for the air / water interface.

Because of this strong foam building ability and the poor emulsificationproperties it was decided to use surface rheology in order to study the behavior.On performing these measurements it was found that extremely lowconcentrations of hydrophobin in water were necessary in order to be able toachieve any measurements at all. In fact concentrations lower than lxl0"6mg/mlwere needed.


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97A t these low concentrations, a two dimensional gel was formed between the

particles. These were relatively strong interactions between the individualmolecules. These have resulted in the interactions that are seen. As can bededuced from Figure 4thesefilms are very strong and elastic gel.

ConclusionsWe can see that the behavior of the hydrophobins is very interesting.

Although the properties that have been studied are bulk ones it is in fact thebehavior at the nanoscale that is having a significant effect. The evidence isthatsmall structures of hydrophobins are forming. These can then partition to the airwater interface where the amphiphilic nature of the molecule causes them tointeract to form strong elastic films. There is clearly little partition to any oil/water interface. This indicates that the specific nano-scale interactions of theparticles are important. In order to further establish the use and applicability ofthesesystems further work is needed.

The results also reveal the complexity and functionality of the hydrophobinmolecules and their applicability in fungi. It seems that the film buildingfunctionality is extremely critical.

AcknowledgementsW e would like to thank Rauni Seppanen, Per Claesson and Eva Blomberg

for interesting discussions. Hans Ringblom, A nnika Dahlman and Anne-MarieHrdin provided experimental help. We are grateful to V I N N O V A and T E K E Sfor financial support.

References1. Wessels J .G., de Vries O.M., Asgeirsdottir S.A., Springer J . J . Gen.

Microbiol. 1991, 137, 2349-2345.2. Linder M.B., Szilvay G.R., Nakari-SetlT , Pentil M.E., FEMS

Microbiol. Rev. 2005,29, 879-896.3. Wsten . ., van Wetter W . A . , Lugones L . G . , van deer M ei H .C . , Burscher

H. J . , Wessels J .G . , Curr. Biol., 1999, 9,85-88.4. TalbotN . J . , Nature, 1999, 398, 295-296.5. Scholtmeijer K ., Wessels J .G.,Wsten H.A. , Appl. Microbiol. Biotechnol,

2001, 56, 1-8


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protein structures as delivery vehicles in foods

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