Chemical Processes - ASAGA · 2017-06-11 · -the first step is the dispersion of the core material in an aqueous cationic polymer (gelatin) solution at 40-60°C-A polyanion (gum

Microencapsulation Methods

CSIRO FOOD AND NUTRITION

Mary Ann Augustin & Luz Sanguansri

Short Course on Micro- and Nano-encapsulation of Functional Ingredients in Food ProductsWorld Congress on Oils & Fats and 31st Lectureship Series31st Oct – 4th November 2015, Rosario, Argentina

Chemical Processes

Outline

• Biopolymer Gels

• Emulsions• Single emulsions• Multilayer emulsion• Double emulsions

• Liposomes

• Molecular Inclusion

• Testing and Characterisation

Micro and Nanoencapsulation Technologies | Augustin & Sanguansri2 |

Biopolymer gels

Single Biopolymer

Mixed Biopolymer Complexes

Chemical Processes_ Microencapsulation | Augustin & Sanguansri3 |

Biopolymeric particles

Biopolymer gels – protects entrapped core and reduce diffusions rate of active until triggered by an external stimulus

• Hydrogels (Discussed in Lecture 3)

• Assembled polymer molecules – polymer gel network

• e.g. alginate, protein

• Biopolymer complexes

• Coacervates

• Complexes between oppositely charged polymers

• e.g. gelatin – gum acacia, whey protein - xanthan

Microspheres


Coacervation

Chemical Processes_ Microencapsulation | Augustin & Sanguansri

The phase separation of a single polyelectrolyte or a mixtureof polyelectrolytes from a solution and deposition of theagglomerated colloidal particles (i.e. the matrix material) onan immiscible active core results in the formation of a simplecoacervate or a complex coacervate

Madene et al (2006) Int. J. Food Sci. Technol. ; Augustin & Hemar ( 2009) Chem Soc Rev

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Factors affecting physical propertiesParticle size determined by core size (emulsion size) and wall thicknessDurability - determined by size and wall thicknessDensity - primarily determined by load

Factors affecting stabilityWall thickness - loadWall matrix - oxygen and water vapor permeabilityWater activity - and its implications on Tg

Complex Coacervation for Encapsulation of Oils

Madene et al (2006) Int. J. Food Sci. Technol. 41, 1-21

Complex coacervation is coacervation caused by the interaction of two oppositely charged colloids

Example: Use of coacervates for encapsulation of fish oil

In the encapsulation process:

-the first step is the dispersion of the core material in an aqueous cationic polymer (gelatin) solution at 40-60°C

-A polyanion (gum arabic) is then added

- pH of the system are adjusted so that a liquid coacervate forms (pH 4.0-4.5)

- system is cooled and the gelatin in the coacervate gels forms a rubbery gel

- an aldehyde is added (eg glutaraldehdye) which forms cross-links with the amino groups of the proteins

-aldehyde treated capsules are dried



Coacervate (gelatin-gum arabaic) with stearidonic acid soybean oil core

Control Transglutaminase Maillard Reaction

Ifeduba and Akoh (2015) Food Hydrocolloids, 51, 136-145

Oxidative StabilityMaillard Reaction > Control > Transglutaminase

Thermal StabilityMaillard Reaction > Transgluatminase > Control

Encapsulation of bovine serum albumin in a complex coacervate


Vandenberg et al (2004) J Controlled Release

Protein retention during manufacture and 24-h acid incubation of chitosan-coated alginate microcapsules produced by varying the alginate concentration.

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Complex coacervate


Cartoon of two self-assembled complexes of gum arabic (white ribbon ) and β-lactoglobulin (dark spheres). The complex as a whole is considered as a new colloidal entity. Protein is 4 nm in diameter and the gum arabic has a diameter of approximately 50 nm.

De Kruif et al. (2004) Curr Opin Colloid Interface Sci

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Electrostatic complexation between gelatin and pectin at low pH


(a) gelatin and pectin molecules exist as individual molecules in solution due to electrostatic repulsion;

(b) gelatin–pectin soluble complexes are formed due to electrostatic attraction between positive patches on gelatin and negative patches on pectin;

(c) soluble complexes merge and form gelatin–pectin sub-units;

(d) hydrogel particles form due to coalescence of sub-units;

(e) setting of internal structure as temperature cools down

B Wu & McClements (2015) Food Res Int, 72, p.231-240

Biopolymeric Systems - 1Bio-polymer systems Method of complex or particle formation ReferenceBeta-lactoglobulin complex

Complex formed by binding of lipophilic molecules to hydrophobic pockets on their surface

Liang et al 2008; Zimet & Livney 2009

Beta lactoglobulin-pectin

Molecular clusters formed at pH values where there is slight electrostatic attraction for encapsulation of omega-3 fatty acids, Vitamin D2

Zimet & Livney 2009; Ron et al 2010

Protein-polyphenol co-assemblies

Complex formed by heating above thermal denaturation where there is weak attraction between protein molecules.

Jones et al 2010a, 2010b; Shpigelman et al 2010

Soy-Zein complex Complex formed by heating above thermal denaturation where there is strong repulsion between protein molecules to promote unfolding and self association but not extensive aggregation.

Chen & Subirade 2009; Nicolai & Durand 2007

Caseinate Complexes formed by binding lipophilic molecules that remain dispersed in aqueous solutions

Semo et al 2007

Casein micelles-hydrophobic nutraceuticals

Molecular clusters formed from casein micelles capable of encapsulating non-polar molecules

Portnaya et al 2006; Semo et al 2007.

Gliadin Particles formed by solvent desorption or nanoprecipitation

Duclairoir et al 1999;


Augustin & Sanguansri 2015, Book Chapter, in press

Biopolymeric Systems - 2Bio-polymer systems

Method of complex or particle formation

Reference

Alginate, pectin (in calcium solution); chitosan (in tripolyphosphate solution); Whey protein-alginate gel beads (in CaCl2 solution)

Particles formed by extrusion or injection of biopolymer solution into another solution as gelling agent

Amici et al 2008; Liu et al 2006; Shin et al 2007; Sheu 1993. Matalanis et al 2011; Wichchukit et al 2013

Whey protein Emulsification-internal gelation process and particles formed by cold-set gelation

Egan et al 2013

Alginate; alginate-whey protein; casein micelles-rennet

Particles formed by utilizing W/O emulsion as template to produce biopolymer with specific dimensions (i.e. homogenization in an oil phase under controlled conditions and particles separated, washed and dried).

Matalanis et al 2011; Reis et al 2006; Chen & Subirade 2006

Gelatin-sodium alginate polyelectrolyte complex

Particles formed by polyelectrolyte complex between Gelatin A and sodium alginate

Devi & Kakati 2013



Biopolymeric Systems - 3Bio-polymer systems Method of complex or particle formation ReferenceCasein, casein micelles Particles formed by protein gelation under controlled

aggregation, adjusting pH close to isoelectric point and adding multivalent counter ions, rennet or enzymes

Cooper et al 2010, DeJong & Koppelman 2002; Huppertz & de Kruif 2008; Song et al 2010

Starch components and derivatives (amylose, maltodextrin, cycoldextrin)

Helices with hydrophobic interior binding non-polar molecules with appropriate molecular dimensions through hydrophobic interactions (e.g. fatty acids and ionic surfactants)

Wangsakan et al 2001, 2004a, 2004b; Zabar et al 2010;

Proteins & polysaccharides

By electrostatic attraction between molecules with opposite electrical charges causing them to associate with each other, by heating under conditions where they form molecular complexes.

Jones & McClements 2010

Proteins & polysaccharides

By strong repulsion between two different biopolymers which occurs when one or both biopolymers are uncharged or have similar charges. Different microstructures are formed by shear and gelation.

Matalanis et al 2011

Beta lactoglobulin –polysaccharide (sodium alginate, low methoxy pectin or high methoxy pectin)

Pressure induced gelation, phase separation triggered by unfolding and/or aggregation

Dumay et al (1999)

Chitosan Emulsion chemical crosslinking method (vanillin as crosslinker)

Peng et al (2010)



Emulsions



Structured emulsions

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McClements & Li (2010) Adv Colloid & Interface Sci, 159, 213-228

Emulsion-based systems - 1• Conventional emulsions

• Simple oil-in-water emulsion

• Filled Hydrogels

• Droplets within a gelled matrix

• Multi-layered emulsions

• Layer-by-layer deposition of oppositely charged

polyelectrolyte onto a primary emulsion droplet

• Microemulsions

• Spontaneously formed transparent dispersions (5-100 nm)

• Nanoemulsions

• Metastable dispersions containing droplets <100 nm


Emulsion systems

CoreDispersed phase

EncapsulantContinuous phase

180 nm 90 nm 60 nm 30 nmNanoemulsions

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Emulsion-based systems - 2

• Liposomes

• Spherical bilayer vesicles

• Self-assembled polar lipid structures

• Stable dispersions - cubosomes, hexosomes

• Solid lipid nanoparticles

• Particle formed above melting temperature of fat and cooled

• Double emulsions

• W/O/W; O/W/O


Double emulsions

Self-assembled structures

Liposome

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Simple Emulsions


Encapsulant material & encapsulation technique influencesthe degree of stabilisation of spray-dried emulsions

ENCAPSULANT MATERIAL + OIL

⇓

Emulsification

⇓

STABILISED LIQUID EMULSION

⇓

Spray Drying

⇓

SPRAY-DRIED EMULSION


Preparation of simple emulsions

Spray dried canola oil emulsions (40% oil) stabilised by Caseinate – Sugar (1:2) Blends

Type of carbohydrate used in combination with protein has influence on the efficiency of encapsulation

Powders made from 40% TS emulsion homogenised at 18MPa

Augustin & Sanguansri, (2010) Book Chapter IN Oxidation in foods and beverages and antioxidant applications and Nutraceuticals, (Eds Decker, Elias and McClements)


Preparation of filled hydrogels



Multilayer Emulsions


Multi-layered emulsions



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Multilayered fish oil emulsion (casein-fucoidan encapsulant)

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Note: Fucoidan (sulfated fucan, is an anionic polysaccharide mainly composed of L-fucose and sulphate groups, which is found in brown algae and sea cucumber. Chang & McClements (2015) Food Hydrocolloids, 51, 252 -260

Multilayer emulsions for oil encapsulation –Physical stability


Linseed oil-in-water emulsions stabilized by a whey protein isolate (WPI) – sodium alginate (SA)

• At pH 6–7 SA did not adsorb on WPI-coated droplets promoting depletion flocculation.

• At pH 5 emulsions remain stable as a WPI–SA bilayer was formed around the droplets.

• At pH 4 bridging flocculation and droplet aggregates promoted emulsion instability

Fiormonti et al. (2015) Food Hydrocolloids, 43, 8-17

Limonene emulsions stabilised by ovalbumin fibrils and high methoxyl pectin (layer by layer deposition)


Release of active depends on number of layers

Humblet-Hua et al. (2011) Food Hydrocolloids, 25, 307-314

CLSM (5 layered capsule) –ThT stain of OVA


Processes occurring digestion of emulsion droplets

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Multilayer emulsions stabilised by lactoferrin –dietary fiber: Lipid Digestibility in vitro

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The presence of a dietary fiber coating around the initial lipid droplets stabilised by lactoferrin had little influence on the total extent of lipid digestion in simulated intestinal fluid

Tokle et al. (2012) Food & Function, 3, 58-66

Double Emulsions


Double emulsions

Benichou et al. (2004) Adv Coll Interfacial Sci,

Double emulsions are complex liquid dispersion systems known also as `emulsions of emulsions´, in which the droplets of one dispersed liquid are further dispersed in another liquid. The inner dispersed globule/droplet in the double emulsion are separated (compartmentalized) from the outer liquid phase by a layer of another phase


Release of Vitamin B1 from double emulsions

Effect of the external aqueous phase pH on the release profile of vitamin B1 from multiple emulsions stabilized with WPI/xanthan gum (4/0.5) as the external emulsifier (pH 7, ●; pH 4, ▴; pH 2, ▪).

Benichou et al. (2004) Adv Coll Interfacial Sci.



Gelled inner dispersed phase (w(1)/o/w(2)) multiple emulsions as fat replacers

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Opperman et al. (2015) Food Hydroclloids, 48, 17-26

Gelled inner phase (gelatin or w multiple emulsions are able to withstand shear and heat → Opportunities as potential fat replacers in foods


Filled hydrogel microspheres based on electrostatic complexation (O/W1/W2) emulsions

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Lipid droplets are trapped within hydrogel microspheres (W1), which themselves are suspended within a continuous aqueous phase (W2).

Delivery systems for lipophilic agents that need to be released in the mouth

Zhang et al. (2015) Food Hydrocolloids, 44, 345-352

Liposomes


Liposomes

• Liposomes • are spherical bilayer vesicles formed

from dispersion of polar lipids in aqueous solvents

• have been widely studied for their ability to act as drug delivery vehicles by shielding reactive or sensitive compounds prior to release at desired sites in the body

• Liposomes may also be used • to encapsulate sensitive food

components

scienceinyoureyes.memphys.sdu.dk


Liposomes in the Food Industry

Liposome entrapment has been shown to stabilize encapsulated, materials against a range of environmental and chemical charges, including enzymatic and chemical modification, as well as buffering against extreme pH, temperature, and ionic strength changes.

Liposomes have been used

• to deliver food flavors and nutrients

• to deliver food additives

• to incorporate food antimicrobials that could aid in the protection of food products against growth of spoilage and pathogenic microorganisms

Taylor et al. (2005) Critical reviews in food science and nutrition 45, 587-605


Liposomes


• Liposomes are generally prepared with phospholipids or glycolipids

• Liposomes are obtained by dispersing the polar lipids in an aqueous phase under high shear forces.

• Liposomes can entrap both sensitive oil and water-soluble components

• Liposomes protects the sensitive components prior to their release.

http://www.endovasc.com/images/graphics/

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Liposome entrapment of Fe for fortification of milk

• Lecithin is used microencapsulation of Fe

• This allows effective Fe fortification of milk – preventing undesirable interactions

of Fe with fat – Improves bioavailability – Masks taste of Fe

0

2

4

6

8

10

12

14

Bio-availability (% Fe absorption)

FeSO4 inwater

FeSO4 inmilk

lipsomalprep inmilk

Boccio et al. Nutr. Rev (1997)

Chemical Processes_ Microencapsulation | Augustin & Sanguansri38

Molecular Inclusion

39 | Materials_ Microencapsulation | Augustin & Sanguansri

OHO

O

HO OH

HOOO

O

O

O

O

OO

OO O

OOH

OH

OH

OH

OH

OH

OH

OH

OHOH

HO

HOHOHO

HO

HO β-Cyclodextrin

Hydrophobicinterior

HO

Materials_ Microencapsulation | Augustin & Sanguansri40 |

Inclusion Complexes

Factors affecting inclusion• Molar ratio of “includable” substances - competitive

basis of inclusion (in a food system vs. an encapsulation system)

• “Desire” to be in CD as opposed to solvent system -driven in by different molecular forces including: polarity (guest vs. solvent), ionic nature, and temperature

• Molecular size and configuration - eugenol fits -isoeugenol does not fit

G. Reineccius, 2001

Materials_ Microencapsulation | Augustin & Sanguansri

Flavour encapsulation

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Complexation behaviour of 13 volatile flavour compounds (α-pinene, β-pinene, camphene, eucalyptol, limonene, linalool, p-cymene, myrcene, menthone,

menthol, trans-anethole, pulegone and camphor) with cyclodextrin was compared↓

A 1:1 inclusion complex for all↓

α-CD and γ-CD gave generally lower stability constants than β-CDs

Ciobanuet al. (2013) Food Res Int, 53, 110-114

Testing and Characterisation


Tests on Microcapsules - Characteristaion

• Physical Characterisation• Particle sizing (light scattering)• Structure on various length scales (light microscopy, scanning electron

microscopy)• Charge (zeta potential)• Dynamic vapour sorption & uptake/release of water• Differential scanning calorimetry (glass transition temperature)

• Chemical composition • Gross composition• Content of core (eg omega-3 fatty acids, flavour compounds)

• Other measures (depending on microcapsules)• Free-fat / Surface Fat ( in oil powders)• Viability of probiotics (in encapsulated probiotics)


Tests on Microcapsules –Stability

• Physical stability & Particle integrity• Changes in physical characteristics ( eg particle size and morphology)• Physical stability - Robustness to compression (eg Instron)• Characteristics of powders ( eg increase in free fat of powders, increase in

stickiness of powders, caking, colour)

• Chemical stability • Core stability

– Kinetics of degradation of core as a function of the environment (eg temperature, moisture)

• Changes to encapsulant /matrix material– Change in glass transition temperature due to moisture uptake


Further Reading• Augustin, M.A. and Hemar, Y., 2009. ‘Nano-structured assemblies for encapsulation of food ingredients’. Chemical

Society Reviews, 38, pp. 902-912.

• Garti, N. ed., 2008. ‘Delivery and controlled release of bioactives in foods and nutraceuticals’, Woodhead Publishing Limited, Cambridge, England.

• Gouin, S., 2004. Microencapsulation: industrial appraisal of existing technologies and trends. Trends in Food Science and Technology, 15 (7-8), pp. 330-347.

• deKruif, G.G., Weinbreck, C.G. and de Vries, R. 2004. ‘Complex coacervation of proteins and anionic polysaccharides’. Current Opinion in Colloid & Interface Science, 9, pp. 340-349.

• Madene, A., Jacquot, M., Scher, J. and Desobry, S., 2006. ‘Flavour encapsulation and controlled release – a review’. International Journal of Food Science and Technology, 41 (s1), pp 1-21.

• McClements, D.J., Decker, E.A., Park, Y. and Weiss, J., 2009. Structural design principles for delivery of bioactive components in nutraceuticals and functional foods. Critical Reviews in Food Science and Nutrition, 49, pp. 577-606.


Thank youCSIRO Food & NutritionMary Ann AugustinResearch Group Leadert +61 3 9731 3486e [email protected]

Documents

Chemical Processes - ASAGA · 2017-06-11 · -the first step is the dispersion of the core material in an aqueous cationic polymer (gelatin) solution at 40-60°C-A polyanion (gum