Proteins – Functionality & Application

Anneke Martin

Content

Functionality

Techno-functionality of proteins

Interfacial properties

Gelation properties

Type of protein networks

Water holding

Texturizing of proteins

Available methods

Extrusion

Industrial examples

Three areas of functionality

Physiological functionalityPhysiological functionality Nutritional functionalityNutritional functionality

Physical functionalityPhysical functionality

…protein

structure & conformation

proteinstructure &

conformation

Types of proteins – based on structure/shape

Globular proteins

(e.g. b-lactoglobulin, soy protein)

Fibrillar proteins

(e.g. gelatin, meat)

Random coil proteins

(e.g. caseins)

Other proteins (macropolymers)

(e.g. gluten)

Different classes of ingredient functionality

Functionality Property Example

Techno-functional SolubilitySolubility

Precipitation

Bulk rheologyThickening

GellingTexturizing

Surface activityFoaming

Emulsifying

Sensory Binding of lipids/flavors

Bio-functional NutritionalDigestibilityAllergenicity

Anti-microbial

PhysiologicalACE inhibition

Opoid activity, etc.

Techno-functional properties of proteins

Function Mechanism Food

solubility hydrophilicity beverages

viscosity water bindinghydrodynamic size

soups, gravies, dressings

water binding hydrogen bonding meat/sausages, cakes, breads

gelation network formation meats, sausages, pasta, baked goods

elasticity hydrophobic interactionsdisulfide crosslinks

meat products, bakery products

emulsification interfacial adsorptionfilm formation

sausages, soups, dressings, desserts

foaming interfacial adsorptionfilm formation

whipped toppings, cakes, mousse, nougat

fat and flavor binding hydrophobic bonding bakery products

Solubility

0 M NaCl

0.5 M

0.2 M

Renkema et al.

β-lactoglobulin

Franco et al. (2011) Fluid Phase Equil 306:242

[NaCl]

0.02

0.01

0.005

0.001

soy protein

Changes in solubility

Insolubility arises from aggregation which is caused by:

Heat (unfolding � exposure hydrophobic groups � more attraction)

Change in pH (at iso-electric point no net charge � no repulsion)

Enzymatic hydrolysis (exposure of hydrophobic groups � more attraction)

Association with non-protein compounds (lipids, flavors, polysaccharides)

NB Presence of salts is needed to solubilise meat proteins!

Interfacial properties of proteins

Most proteins have hydrophilic and hydrophobic parts

Protein unfolds at interface and decreases interfacial tension

Due to charge proteins act as stabiliser at the air/water and oil/water

interface

air

fat

----

--

---

--

-

--

� Negative charge causes electrostatic repulsion and stabilizing effect

� Addition of salts (e.g. Na+) decreases repulsion

Interfacial properties of proteins

Interface, role of protein

Steric repulsion/charge

Effect of e.g. salt, pH for stability

Destabilisation process

Practical example

Examples of foods with interfaces

solid liquid gas

solidsolid suspension: fruit ice, chocolate

solid emulsion, gel:jellies, cheese, processed meat

products

solid foam: foam candy, bread,

baked products

liquid sol, suspension:

orange juice, acidic beverages

emulsion: milk, mayonnaise, margarine, french

dressing

foam: meringues, whipped cream, beer foam

gas aerosol aerosol

DISPERSED PHASE

CO

NT

INU

OU

SP

HA

SE

Sausage emulsion – comminuted meat

Contains ~30% fat present in small droplets

Stable homogeneously distributed fat droplets are positive for

juiciness & tenderness

Gelation properties (globular proteins)

Heat treatment � unfolding � aggregation � gelation

Gelation kinetics, type of gel and gel strength are a.o. influenced by:- temperature-time- pH- presence of salts- protein concentration

NOT every protein denatures and forms gels!

Gelation properties

gelatin

whey protein

soy protein

MOLECULARDENATURATION/AGGREGATION

GELATION NETWORK

Mechanisms of network formation

Heat-induced gels

High Temperature (whey, soy, egg white)

Low temperature (gelatin)

Cold -set gels, pre-heat treatment followed by:

Acidification (yoghurt)

Enzyme induced, e.g. rennet (cheese)

Addition of salts, e.g. Ca2+ (tofu)

High pressure induced

Combination of pressure and temperature

Type of network:• fine/coarse

stranded• particledetermines rheological and eating properties

Methods: from molecule to food product

Chromatography

Thermal analysis

Circular dichroism

SDS Page

< 10 nm 20-500 nm 1-500 µm mm-cm >mm-cm

Information on: -structure-unfolding vs. native-denaturation temp.

Light scattering

Electron microscopy

Information on: -aggregation-size of structures

Confocal microscopy

Light microscopy

Rheology

Information on: -size of structures-properties of network at small deformation-ingredient interaction

Texture analyzer

Microscopy

Texture analyzer

Sensory panel

Information on: -properties of network at large deformation related to eating properties-microstructure

Information on: -properties of network at large deformation-sensory properties, liking

Typical methods

Texture Analyzer – large deformation – eating properties

Rheometer – small deformation – gelation kinetics

Recoverable energy is high for elastic materials

Rheological properties protein networks

Link to hardness, elasticity

Structure (particle versus stranded network) versus water binding

Short chains versus long chains (winegum, young versus old cheese)

Typical methods

TA

Rheometer

Microscopy

Light scattering

Egg white protein – ovalbumin

Parameter

[NaCl] 0.2 M -

Type gel particle stranded

Fracture stress (kPa) 70 37

RE (%) 45 75

TIFN 2013

Vegetarian burger (Javaanse schijf)

Role of egg white protein: binding water and holding mass together

Texture analyzer and sensory panel are used to determine differences

TNO 2013

Soy protein – effect of salt type

Aggregates with Ca > Mg

Coarseness gels MgCl2 >MgSO4

Both anion and cation determine

structure of protein network

Effect on water holding, hardness & eating

properties

Urbonaite (2013) TIFN

Water binding/holdingRelevant for juiciness, tenderness

Water holding capacity in meatWater holding capacity in meat proteins

[NaCl] ↑

plasma

[gelatin]

TIFN 2013 TNO 2010

Texture analyzer: hardness, elasticity, serum release

Serum release during deformation is a measure for

juiciness and flavour perception

The perceived juiciness is the result of the amount of

serum that is pushed out of the meat matrix while

chewing and the ability of the tissue to bind water , which

is affected by the salt content.

Eating properties

Sausages with high serum release were perceived significantly saltier than

those with little serum release

Sausages with a high serum release were perceived more juicy than those

with a low serum release.

interconnected pores

separate pores

protein continuous system

Microstructure controls serum release � Serum release relates positively to salt/flavour perception

vdBerg (2007) Food Hydrocoll 21:961

Gluten

80% of gluten consists of gliadin and glutenin

Gluten are not soluble in water

Gives structure to e.g. bread and pasta

Forms strong reversible elastic network and entraps air bubbles

during proving and baking

Glutenfree

Gliadin fraction is said to be responsible for coeliakie

Replacement of gluten by hydrocolloids in bread does not result in

satisfying products

Gliadin fractions can be replaced by protein particles made from whey

protein or gelatin

protein particles flour+water+protein

Van Riemsdijk et al. (2011) J Cereal Sci

Replacement of meat proteins by plant protein

Meat protein – fibrillar structure

Plant proteins – globular or other protein

unfolding of globular proteins and alignment of protein aggregates into fibrillar structures

TUNE FUNCTIONALITY OF PROTEIN !

Structuring of proteins

Formation of fibrillar-like structures of protein to mimic meat

Electrospinning

Shear cell

Fibril formation at low pH

Extrusion

Examples: Valess, Ojah (Beeter), texturized protein (Solae)

Electrospinning

Spinning of proteins in food-grade way is difficult

For use in food fibers need to be collected and/or

aligned (in progress)

Upscaling of process needs attention

Nieuwland et al. 2013 TNO

gelatin + 15% WPI + 12.5% WPI + 10% WPI

Shear cell

Flow induced structuring of 30% caseinate in

combination with enzymatic crosslinking

Fibrous structures of µm-mm

Manski et al. (2008) Food Hydrocolloids 22: 587

Not performed yet with plant proteins?

Scaling up?

Fibril formation

Extensive heating and shearing at low pH renders protein fibrils

Unique property: very low critical gelling concentration

Drawbacks: yield (conversion) + upscaling

1000 nm

1000 nm

WPI fibrils Ovalbumin fibrils

Munialo (2013)TIFN

Extrusion

Thermal and mechanical energy input at low to medium

moisture conditions

Protein concentrates or flours are mixed with water, pushed

through a cylinder (turning screw). Moisture is evaporated

forming dry fibrous, porous granules or chunks

Texturized vegetable protein (TVP)

Microfibrillar protein network due to unfolding, orienting and thermal

crosslinking (temperature well above denaturation temp.)

Dry texturized proteins with high water binding & swelling capacity

Extrusion of soy protein

During extrusion denaturation and chemical

reactions decrease solubility

Moisture content affects solubility

28% moisture 60% moisture

buffer

denaturing agents to increase solubility (e.g. SDS, urea, mercapto-ethanol)

Chen et al. (2011) Food Sci Techn 44:957

Extrusion of other plant proteins

Literature shows that pea protein can be extruded and has similar

properties as extruded soy proteins

Nothing available yet on extruded lupine protein

Role of gluten and starch in extrusion of plant proteins under

investigation in Cluster project TNO

Valess

milk proteins + polysaccharides (alginate from sea weed)

extensive mixing � phase separation of biopolymers

formation of structure through addition of Ca2+

Patent by Kweldam US2011244090

Ojah Beeter

Sold by ao Vegetarische slager

Based on plant proteins (and probably polysaccharides from

seaweed)

glutenfree

Structure formation by sheeting/lamination of pressed heated protein

Questions?