Review of techniques to manufacture micro-hydrogel particles for the food industry and their applications

Journal of Food Engineering 119 (2013) 781–792

Contents lists available at ScienceDirect

Journal of Food Engineering journal homepage: www.elsevier.com/locate/jfoodeng

Review

Review of techniques to manufacture micro-hydrogel particles for the food industry and their applications Heather M. Shewan, Jason R. Stokes ⇑ School of Chemical Engineering, University of Queensland, Brisbane, QLD 4072, Australia

a r t i c l e

i n f o

Article history: Received 22 March 2013 Received in revised form 27 June 2013 Accepted 28 June 2013 Available online 9 July 2013 Keywords: Microgel Nanogel Core–shell Microparticle Nanoparticle Biopolymer Protein Polysaccharide Emulsion Atomisation Microfluidics Phase separation Crosslink Rheology Encapsulation Satiety Targeted delivery Controlled delivery Spheroids Fibre

a b s t r a c t Microgels are ‘soft’ microscopic cross-linked polymeric particles that are being increasingly exploited in a variety of industries for rheology control, encapsulation and targeted delivery. They are valued because of the ability to tune their functionality to address specific applications in oil recovery, coatings, drug delivery, cosmetics, personal care and foods. Food microgels are typically biopolymer hydrogels in the form of microspheres, nanospheres (also called nanogels), spheroids and fibres. The utilisation of engineered microgels in foods has so far been limited, despite their great potential to address several needs in the food industry, including: satiety control, encapsulation of phytonutrients and prebiotics, texture control for healthier food formulations (e.g. reduced fat products), and targeting delivery to specific areas in the digestive tract. We review the scientific and patent literature on the utilisation and manufacturing methods for producing microgels with an emphasis on micro-hydrogels for food applications. Ó 2013 Elsevier Ltd. All rights reserved.

Contents 1. 2.

3.

Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Microgel properties and applications . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.1. Flow behaviour, rheology and texture control . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.2. Encapsulation and targeted delivery . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Manufacturing methodologies and approaches . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.1. Moving from precursor to microgel . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.1.1. Physical gelation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.1.2. Chemical gelation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.2. Physical formation of precursor droplets – emulsion route . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.2.1. Droplet deformation and breakup theory . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.2.2. Microgel formation from homogenisation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.2.3. Microfluidics . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

⇑ Corresponding author. Tel.: +61 7 3365 4361; fax: +61 7 3365 4199. E-mail addresses: [email protected] (H.M. Shewan), [email protected] (J.R. Stokes). 0260-8774/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.jfoodeng.2013.06.046

782 782 782 784 784 785 785 785 785 786 786 786

782

H.M. Shewan, J.R. Stokes / Journal of Food Engineering 119 (2013) 781–792

3.2.4. Membrane . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Physical formation of precursor droplets – atomisation route . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.3.1. Theory of drop break-up in air . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.3.2. Spinning disk atomisation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.3.3. Spray nozzle . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.3.4. Extrusion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.3.5. Spray drying and spray cooling . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.3.6. Rehydration of spray-dried biopolymer solutions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.4. Physical formation of precursor droplets – microparticulation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.4.1. Shear gels . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.4.2. Microparticulated proteins. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.5. Physicochemical formation of precursor droplets . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.5.1. Coacervation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.5.2. Phase separated polymers . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Conclusion and outlook . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3.3.

4.

1. Introduction Micro hydrogels are ‘soft’ microscopic particles consisting of cross-linked polymeric molecules. They are valued for their functionality and ability to tune physical properties in industrial applications including oil recovery, paint and surface coating, controlled drug delivery, cosmetics, personal care, pharmaceuticals and foods (Tan et al., 2010; Stokes, 2011). The particles are swollen in solvent and possess a rich set of functionalities and suspension properties that can be exploited for rheological and texture control as well as encapsulation and/or targeted delivery. Exploitation of the unique properties of microgels in food and beverage applications are yet to be fully realised, which is partially due to a lack of awareness of their potential and the perceived limitations for their large scale manufacture. To address this issue and encourage further research and development in the field, we review routes to manufacture food-grade micro- and nano-hydrogel particles (which we simply refer to here as microgels) and highlight their potential benefits in an exciting variety of food applications. The incorporation of solvent into the polymer network structure of microgels makes them unique and highly exploitable across many industries. The network structure is usually viscoelastic and responsive to variations in its environment (e.g. solvent quality) that allows the microgel to swell or de-swell accordingly. For example, adjusting solvent quality to cause de-swelling can be used to drive solid-to-liquid transitions in microgel suspensions while it can also be used to release encapsulated ingredients. Their rheological and encapsulation/release properties can be tuned for specific applications through variations in the composition, size, shape, cross-link density, and surface properties of the microgels. While microgels are tuneable to obtain a range of different functionalities for non-food and food applications (for examples, see Fernandez-Neives et al., 2011), the degree of modifications to chemical structure and surface properties are more limited in food applications due to the need for the microgels to be safe to eat. The major consequence is that microgels for food applications are (typically) based on semi-rigid polymers that are present in nature, i.e. polysaccharides, while synthetic microgels are (typically) based on flexible polyelectrolytes. As a consequence, processes for producing food-grade microgels are very different from their synthetic counter-parts and provide unique challenges, and there are many properties of food-grade microgels that are distinctly different to those exploited in other industries. Interestingly, the most widely used microgels are in fact food grade and naturally occurring: starches! While starch granules are hard in their natural state, upon heating they swell in what is referred to as gelatinization. In this state the soft granules can be considered to be microgels

787 787 787 787 788 788 788 789 789 789 789 790 790 790 790 791

since they are comprised of cross-linked carbohydrate polymers. Starches are universally used in food for texture and rheology control, and are a rich topic for review in their own right. Thus they are considered outside the scope of this review. This review includes a brief presentation on microgel properties and applications followed by a more thorough evaluation of the scientific and patent literature on the main methods applicable to non-starch biopolymer microgel manufacture. The utilisation of engineered microgels in foods has been limited due to the difficulty in manufacturing food-grade biopolymer microgels using a consistent, cost effective and scalable process (Gouin, 2004). We review commercially viable processes for manufacturing food grade microgels at an industrial scale. 2. Microgel properties and applications 2.1. Flow behaviour, rheology and texture control The rheology of colloidal microgel suspensions uniquely shares characteristics of both polymer solutions and hard-sphere suspensions (Wolfe and Scopazzi, 1989; Stokes, 2011). Microgel suspensions show increased viscosity and shear thinning at relatively low solids content like polymers, but they are inherently particles and are thus governed by many of the same factors as hard colloidal spheres. However, their specific volume can alter in response to changes in solvent quality in much the same way as polymers alter their conformation with solvent quality. We summarise here the key rheological-based properties of microgels noting that a more detailed review is provided in Stokes (2011). The viscosity of hard sphere suspensions increases with increasing phase volume until it diverges towards infinity at a so-called maximum packing fraction governed by the particle size distribution, and corresponding to 0.64 for monodisperse hard spherical particles, as described by commonly used models of Krieger and Dougherty (1959) and Quemada (1977). The viscosity of nonattractive microgel suspensions follows a similar functional relationship to that of hard spheres, but the maximum packing fraction can exceed 0.64 and even go beyond unity. This is possible because microgels contain solvent in their network structure, and so their specific volume is defined under dilute concentrations where they are fully swollen. Large effective phase volumes can be obtained for the softer microgels because they deform and compress at high phase volumes. Fig. 1 depicts the viscosity and linear viscoelastic properties of microgel dispersions and their structural arrangement as either a gel or soft glass at high phase volume. Colloidal scale microgel particles can be designed to possess long range attractive forces that cause aggregation and flocculation

H.M. Shewan, J.R. Stokes / Journal of Food Engineering 119 (2013) 781–792

So glass

Gel

G’

Viscosity

Decreasing Parcle Modulus

G’’

Liquid-like dispersion

Solid-like dispersion 1

φ/φm

Fig. 1. Schematic of rheological behaviour of microgel suspensions across a range of phase volumes (U) and their underlying microstructure in the form of either a softglass or gel. Viscous behaviour of microgels occurs up to maximum packing fraction and elastic solid-like behaviour above maximum packing (Um).

at low volume fractions. Like any colloid, the viscosity and rheology is largely controlled by the strength of interaction potential between particles and the resulting network structure. In this case, like other attractive colloids, the viscosity approaches infinity at lower maximum packing fraction than that for spheres with nonattractive surface forces. When microgels are densely packed, such that the effective phase volume is above the maximum packing fraction, they display characteristics of both a viscoelastic polymer gel and an attractive colloidal suspension; that is, they display solid-like linear viscoelastic properties but they also flow as a fluid when a sufficient shear stress is applied (i.e. the yield stress). The storage modulus of dense suspensions increases with increasing particle modulus, which is considered to be controlled by Hertzian contact mechanics (Evans and Lips, 1990; Adams et al., 2004). Microgels are commonly used for rheological control as suspensions can undergo a transition from liquid-like to solid-like behaviour, with an increase in phase volume. This unique ability can be exploited by increasing the volume of the microgels in suspension, either by adding a greater percentage of microgels or by inducing swelling via a change in pH or solvent conditions. Microgels be can be tuned in a variety of ways that influence their functionality and flow properties. As depicted in Fig. 2, particle properties that can be manipulated include: the degree of crosslinking which affects particle modulus; particle size and size distribution; polymer type which controls their responsiveness to the environment for example pH, temperature or enzymatic response; particle surface charge; and particle shape from spherical to elongated rods and

783

fibres. Microgels can be designed to shrink or swell by creating an imbalance in the osmotic pressure between the inside and outside of the microgel (Lyon and Fernandez-Nieves, 2012). This imbalance can be brought about by altering temperature, pH or ions present in the suspension. Through careful manipulation of their effective phase volume, internal structure (particle modulus), surface properties and size, microgel suspensions can be designed to exhibit a range of responses under shear, including constant viscosity (Newtonian), shear thinning, yield stress, and shear-thickening (Wolfe and Scopazzi, 1989; Frith and Lips, 1995; Frith et al., 1999; Adams et al., 2004; Frith, 2010). At high shear rates the viscosity of microgel suspensions is generally less than that for polymer solutions with a similar low shear viscosity. Also, microgel suspensions do not possess the non-linear viscoelasticity and extensional properties observed with polymer solutions (Stokes, 2011). Such non-linear viscoelastic properties can be undesirable in some food systems, such as beverages and dressings, from both a processing and consumer experience perspective. Microgel suspensions may offer an alternative way in which to obtain desired shear thinning and/or yield stresses without undesirable elastic properties (Stokes, 2011). An example of where the tuneable properties of microgels have been widely utilised for advanced rheological control is in personal care products (e.g. carbopol microgels are used to form clear topological ‘gels’) and paints, as reviewed in (Stokes, 2011). For coating applications microgels made it possible to remove harmful volatile organic compounds from paint formulations. Analogous to this non-food application, microgels may also enable fat reduction in processed-foods. Fat replacement is a priority goal for the food industry due to rising obesity levels in consumers and rising prices of food grade oils due to their association with fuel. One such product that is commercially available is a microgel called Simplesse™, which is formed from microparticulated cross-linked whey protein (see Section 3.4.2). This was originally patented in 1988 (Singer et al., 1988) and purported to be a fat replacer in products including spreads, ice-cream and yoghurt (Singer et al., 1989, 1992). Sandoval-Castilla et al. (2004) found that the addition of microparticulated whey protein to reduced fat yoghurt results in textural characteristics matching those of full fat yoghurt. Karaca et al. (2009) showed that upon addition of Simplesse™ to low and no fat ice-cream, sensory scores were similar or better than that of the full fat product when viscosity of the two products is matched. The rheology and sensory characteristics of a low fat mayonnaise with microgels in the form of shear (fluid) gels have been patented and studied by Norton et al. (2009). They show that by using soft spherical fluid gels in place of the oil droplets found in commercial full fat mayonnaise, they can closely match both the rheological profile at shear rates up to 300 s1 and the sensory properties assessed by a consumer panel. Microgels also have the potential to: lower energy density of foods (Sagis et al., 2008); improve mouth feel; add shear stability; reduce sedimentation or creaming during transport and storage; stabilise emulsion droplets (Hedges, 2009); and coat sensitive products with thin transparent protective

Fig. 2. Schematic showing characteristics of microgels.

784

H.M. Shewan, J.R. Stokes / Journal of Food Engineering 119 (2013) 781–792

Fig. 3. Schematic showing types of encapsulation.

coating. The traditional use of starch for numerous functions in foods also demonstrates the potential for non-starch microgels. 2.2. Encapsulation and targeted delivery Encapsulation can be used to protect sensitive ingredients (actives) inside the microgel (carrier), and target delivery of this ingredient to the desired site in the body. For example, in pharmaceuticals the adhesion of the microgel to the inside of the mouth can enable the slow release of a drug (Kutyla et al., 2013). Encapsulation in microgels can be achieved by embedding the active directly within the polymer network (dispersed) or within an immiscible droplet phase within the microgel, as depicted in Fig. 3 (Augustin and Hemar, 2009; Dubey et al., 2009). Microgels and the suspension format can be tuned to the potential application, the desired release rate, and breakdown profile. There are several key parameters commonly used as a measure of encapsulation efficiency and performance, each of which must be matched to the desired behaviour of the microgel, for example ‘in mouth release’ of an active will need to be very slow for protection of a nutraceutical but very fast for sensory perception of a flavouring ingredient (Gouin, 2004). These parameters include: the amount of active encapsulated within the microgel compared to that remaining on the particle surface; the quality of the active; and the release rate during production, storage and usage (Augustin and Hemar, 2009; Jun-xia et al., 2011; Najafi et al., 2011). Type of carrier material, amount of carrier and encapsulation method all affect these measures and there is often a balance required between release rates in storage compared to those in use, and ratio of active to carrier material. Encapsulation and targeted delivery is most commonly associated with the pharmaceutical industry where there is extensive research into biocompatible polymer microgels of synthetic origin. Although many of these synthetic compounds cannot be added to food products we still have much to learn from these applications. Synthetic microgels can be functionalised to be responsive to light, pH and temperature. The responsiveness can be used to release a drug or other compound at the desired point in the human body. An example of this is magnetic nanoparticles coated with block copolymer that have an open structure at room temperature that allows a drug to be loaded, but a more condensed structure at body temperature releasing the drug slowly as it passes through the intestinal tract (Neoh and Kang, 2010). There are many modes for tuning microgels during manufacture to give a desired drug release rate at a specific delivery site (Schmidt et al., 2006). Highly tuneable microgels can be exploited in nutraceutical food applications by delivering functional ingredients while: masking unpleas-

ant flavours and aromas; controlling release location and rate; increasing satiety; and protecting sensitive and volatile ingredients (Chen et al., 2006; Dubey et al., 2009; Li et al., 2012). Air (or other gases) can be added to microgels to make them either neutrally buoyant (low volume of air) or to reduce their energy density (high volume of air). While the density of microgels is closely matched to that of the solvent (water), a slight density mismatch leads to sedimentation over time. In addition, encapsulation of dense components leads to enhanced sedimentation of the microgels. Microgel suspensions that are stable against creaming and sedimentation can thus be made by tuning the volume of air to match the microgel density with the fluid density (Shewan and Stokes, 2012). A similar result can also be achieved by incorporating a small volume of oil in the microgel. Microencapsulation is commonly used for protecting sensitive and volatile food components from attack (Gouin, 2004; Augustin and Hemar, 2009; Dubey et al., 2009). For example, microencapsulation has recently been developed to protect viable probiotics in dairy desserts to ensure they are available at a beneficial level of 106–109 cfu/g at the time of consumption (Homayouni et al., 2008; Mohammadi et al., 2011). Probiotics L. Casei and B. Lactis have been encapsulated in granular scale alginate microgels via an emulsion gelation route (Homayouni et al., 2008). In ice-cream the encapsulated probiotics had significantly higher survival rates than as free cells during both the freezing and storage processes whilst sensory properties were maintained. Estrada et al. (2011) effectively encapsulated salmon oil in microgels to increase the level of polyunsaturated fatty acids in yoghurt from 1.76% to 7.50% without affecting yogurt quality (pH, syneresis and colour). To achieve this they spray dried an emulsion containing salmon oil, gum arabic and maltodextrin. Satiety control is being targeted by manufacturers to improve the health status of consumers. Li et al. (2012) suggest that encapsulation of lipophilic compounds in large microgels could modulate satiety response by two mechanisms: (1) large microgels would be transported more slowly through the stomach and intestinal tract and; (2) the rate of release of lipophilic compounds from the biopolymer shell would be limited by the large size. The potential application of microgels in food is broad and can take many avenues. The main limitation at this time is designing or adapting manufacture techniques to be economically viable and only using ingredients which are safe for consumption. The increasing demand for healthy and functional foods can only improve the cost/benefit ratio.

3. Manufacturing methodologies and approaches Microgel manufacture techniques can be classified by looking at the type of precursor or the macro-scale process required to initiate particle formation. Defining microgel manufacture techniques by precursor gives three classes (Fernandez-Neives et al., 2011): (1) Monomer; (2) Polymer; or (3) macro-gel. For microgels made from synthetic polymers, the starting point is usually a monomer. The starting point for biopolymer microgels is usually the polymer or macro-gel; either the polymer is gelled into a microparticle or they are formed by mechanical comminution of macro-gels. Comminution after and during gelation has been particularly successful for producing anisotropic and irregularly shaped biopolymer microgels. Classification by particle formation mechanism is a more useful grouping for biopolymer microgel manufacture. It also gives three classes, those formed by: (1) Homogenous nucleation; (2) Mechanical/physical methods (emulsification, atomisation or microparticulation); or (3) Chemical/thermodynamic methods (coacervation, phase separation). Homogenous nucleation is not relevant to

H.M. Shewan, J.R. Stokes / Journal of Food Engineering 119 (2013) 781–792

biopolymer microgels and is not considered here; we instead focus on physical methods and physicochemical routes (Vilstrup, 2001; Dubey et al., 2009). The main physical approaches involve gelling the droplet phase of an emulsion or spray. These are highlighted in the next section and include the theory for creating emulsions; this is not only a manufacturing method in its own right, when combined with a gelation step as covered in Section 3.1, but also the precursor to many of the chemical methods. The physicochemical route, reviewed in the final section, has two distinct paths: coacervation, involving attraction between mixed polymers; and phase separation, involving a degree of repulsion between mixed polymers. Each of these methods has particular advantages or limitations and the method chosen largely depends on both the chosen polymer and the application. 3.1. Moving from precursor to microgel Cross-linking is a discrete step required for formation of microgels and it is initiated via chemical addition, pH change, enzymes or temperature. The type and quantity of cross-links can dictate particle stability, particle morphology, suspension microstructure and rheology. Initiation of cross-linking is required to form a gel particle from a droplet created by either physical or physicochemical methods. Cross-linking mechanisms for polymers can be divided into two distinct groups known as physical and chemical gelation. The route used for gelation of biopolymer droplets to form microgels depends on the biopolymer used and on the application. In contrast to commonly used synthetic polymers, most gelling-biopolymers gel through physical associations. However, chemical bond formation by addition of chemicals or enzymes can be used subsequent to physical gelation to form a more stable microgel. 3.1.1. Physical gelation Formation of biopolymer gels can occur when a network structure forms via non-covalent bonds such as hydrogen bonding, electrostatic interactions and hydrophobic interactions. These physical associations are strong enough to form stable gels under specific conditions (for example, temperature or pH) however they are less stable than covalent bonds and are often reversible. The particular route of physical gelation is complex and still an area of active research as it is dependent on both biopolymer conformation and solution conditions (Clark and Amici, 2003; Rinaudo, 2006). In addition, more than one type of interaction can occur during polymer gelation. For example: the gelation of gellan and carrageenan rely on a balance between attractive hydrogen bonds and electrostatic repulsions of ionic groups on the chain; and protein selfassociation requires both hydrophobic attraction and formation of covalent disulfide bonds (Rinaudo, 2006; Matalanis et al., 2011). 3.1.1.1. Hydrogen bonds. Inter- and intra-chain hydrogen bonds involve loose dipolar interactions between hydrogen atoms and electronegative atoms such as oxygen (Rinaudo, 2006). These reversible bonds are strengthened with decreasing temperature (Jones and McClements, 2010). Hydrogen bonding is the main mechanism of gel formation in random coil biopolymers such as agarose, carrageenan and bacterial gellan which, when cooled, begin to form fibrous helical structures then aggregate to form a gel (Clark and Amici, 2003). Gelation of gelatine also involves stabilising hydrogen bonds to form the triple helical structure although gelatine does not undergo the aggregation step (Clark and Amici, 2003). 3.1.1.2. Hydrophobic interactions. Hydrophobic interactions are especially important in gelation of proteins and modified polysaccharides where polymer chains have both hydrophilic and

785

hydrophobic parts (Rinaudo, 2006; Matalanis et al., 2011). When these amphiphilic molecules are dispersed in water the hydrophilic parts interact with the water and the hydrophobic parts interact with one another to form a hydrophobic domain, for example folding of a globular protein to bring all the non-polar parts to the centre (Rinaudo, 2006; Jones and McClements, 2010). 3.1.1.3. Ionic interactions. Ionic or electrostatic interactions occur where there is an attraction between charged polysaccharides and other ionic species in the solution such as counter-ions, ionic surfactants or polymers. These interactions can be repulsive or attractive and can result in phase separation or coacervation reactions respectively (Rinaudo, 2006; Jones and McClements, 2010). Ionic interactions are affected by the solution pH in relation to the isoelectric point of the biopolymer, the types of ions present, temperature and the salt concentration (Jones and McClements, 2010). For gelation to occur by this mechanism a specific salt is often required, for example: addition of calcium ion for alginate gelation. 3.1.2. Chemical gelation Chemical cross-linking involves the formation of covalent bonds. Chemical cross-linking of proteins such as gelatine is achieved using glutaraldehyde, transglutaminase (TGA) and genipin. We review transglutaminase and genipin because they can potentially be used in foods. However we do not cover glutaraldehyde as it is a toxic chemical and so it has limited application for food and pharmaceutical products. TGA is an enzyme which acts as a catalyst to the reaction between the c-carbonyl group of a glutamine residue and the e-amino group of a lysine residue (Bertoni et al., 2006). There are two sources of TGA, microbial (mTGA) and mammalian tissue (tTGA). tTGA has a higher reactivity than mTGA but it has the disadvantage that it requires calcium to be present for its activity. The process and amounts of TGA used affect cross-link stability over time, and it is generally observed that if an insufficient amount of TGA is added prior to thermal physical gelation then a weaker gel forms (Babin and Dickinson, 2001). Gelatine gels cross-linked with TGA have been shown to be stable under: thermal stress, up to 37 °C; mechanical stress, up to 345 mmHg (Liu et al., 2009); and in the form of microgel capsules (Prata et al., 2008). TGA cross-linking has the potential to allow microgels to be produced with a variety of moduli and functionalities. Genipin is another available chemical cross-linking agent which reacts with free amine groups via two different mechanisms: (1) nucleophilic attack by an amine group resulting in the formation of a heterocyclic amine; (2) nucleophilic substitution reaction, which results in the replacement of the aldehyde group formed during mechanism (1) with a secondary amide linkage (Liang et al., 2004; Chiou et al., 2006). Liang et al. (2003, 2004) found that although glutaraldehyde and genipin have the same cross-linking mechanism, genipin is significantly less toxic. The reaction time of genipin is highly variable and affected by pH and both protein and genipin concentrations. The time taken to form a stable gel using genipin cross-linking occurred after: 165 min with gelatine (Liang et al., 2003); a two week period for Pollock gelatine samples; and 5 days for porcine gelatine (Chiou et al., 2006). A major disadvantage of genipin is that the cross-linking reaction results in a dark blue/black coloured gel which limits its use in foods (Liang et al., 2003). 3.2. Physical formation of precursor droplets – emulsion route Emulsification can be used for many combinations of biopolymers and oils. The shear or energy required for small droplet formation can be provided by a large range of homogenisers and

786

H.M. Shewan, J.R. Stokes / Journal of Food Engineering 119 (2013) 781–792

high pressure mixing devices, as well as recently developed smallscale devices based on microfluidics and flow through membranes. Equipment choice should be made taking into account the fact that, in general, the greater the shear applied to a particular system the smaller the droplet that is formed. Droplet size also depends on such factors as the exact flow geometry, viscosity ratio of the phases and the interfacial tension between them. Discussion of the emulsification processes most applicable to manufacture of microgels for food applications follows a brief introduction to emulsion theory. 3.2.1. Droplet deformation and breakup theory Droplet breakup mechanisms have been well established in atomisation and emulsification. It has been shown that control over the particle size and modulus will depend on hydrocolloid concentration, ratio of both volume and viscosity of the continuous and dispersed phases, emulsifier type, shear conditions and temperature (Esposito et al., 1996; Ugwoke and Kinget, 1998). To create an emulsion, energy is required to form droplets of one phase suspended in another immiscible phase with a large surface area between the two phases (Dickinson, 2009). To bring about drop break-up the shear applied must be greater than the interfacial forces holding the drop together. This force is described by the Laplace pressure (PL) which is the difference in pressure at the concave side of a curved phase boundary and that at the convex side and depends on drop radius (R) and interfacial tension (cint). The drop breaks if the capillary number Ca, defined in Eq. (1) as the ratio of the shear stress (s) over the half Laplace pressure (PL = 2cint/ R) exceeds some critical value (Cac) that depends on the viscosity ratio (p) between the dispersed and continuous phases, as shown in Eq. (2) (Leal-Calderon et al., 2007). The relationship between the capillary number and viscosity ratio and drop breakup under different flow conditions for single droplets is shown in Fig. 4. Under shear flow, drop breakup of single droplets will not occur for p > 4. This observation is valid for droplet volume fractions of up to 30% (Leal-Calderon et al., 2007).

Ca ¼

p¼

Rs

cint

gd gc

ð1Þ

ð2Þ

Interfacial surface tension can be reduced by addition of an emulsifier, in the form of a surfactant, hydrocolloid or protein, to control droplet size and prevent coalescence of drops after formation (Surh et al., 2006; Leal-Calderon et al., 2007; Dickinson, 2009). Dickinson

Fig. 4. Capillary number against viscosity ratio showing condition of rupturing in quasi static-conditions for a simple shear flow. Graph produced from the following C2 equation from (de Bruijn, 1989): log Cacrit ¼ C 1 þ a log p þ log plog where C1, C2 pmax and a are constants. A schematic representation of drop morphology during break up dependent on viscosity ratio A (p < 0.2); B1 and B2 (0.03

3.8) is also shown, with permission (Rumscheidt and Mason, 1961).

(2009) defines an effective emulsifier for two different situations: (1) turbulent flow conditions such as those in a homogeniser; and (2) quiescent conditions. In quiescent conditions, mass transfer of the surfactant to the drop surface occurs by diffusion, meaning that the most rapidly absorbing species are small molecular weight surfactants and individual protein molecules. Under turbulent flow conditions, transport to the interface is dominated by convection favouring large macromolecules and colloidal particles. 3.2.2. Microgel formation from homogenisation Microgels can simply be formed by gelling the aqueous phase of a water-in-oil emulsion. Gelation may be brought about during emulsification or after emulsification. As the droplet phase gels, its viscosity increases dramatically and this impacts the size distribution and shape of the microgel (Ellis and Jacquier, 2009a); for example excessive shearing during gelation can result in anisotropic microgels (‘shear gels’, Section 3.4.1). Emulsion–gelation is a particularly easy technique to use at the laboratory scale. Water-in-oil emulsification followed by a cooling gelation step, has been used by Adams et al. (2004) and Loret et al. (2007) to produce agar microgels and by Ellis and Jacquier (2009a) to form jcarrageenan microspheres. Creating food emulsions at industrial scale is common practice and hence it is possible to use the same technology to scale up the above processes to produce microgels of similar size to drop sizes found in regular food emulsions. Mofidi et al. (2000) used an emulsion based process for large scale production of alginate microgels (1–500 lm in size) where emulsion droplets containing alginate are dispersed in oil followed by cross-linking of the droplet phase by addition of calcium chloride. The limitation of this method is the extended time required to remove oil and surfactant from the microgel using a series of centrifugation and washing steps. This disadvantage can be overcome by use of two novel methods covered later in this paper in shear gelation (Section 3.4.1) and water in water emulsions (Section 3.5.2). 3.2.3. Microfluidics Microfluidic devices are a very small scale system most appropriate for producing monodisperse, uniform microgels, as well as single and double (w/o/w or o/w/o) emulsions for laboratory scale testing. The continuous phase passes through a small channel and past a nozzle through which the dispersed phase is pumped, an example of which is shown in Fig. 5a. The channels are generally formed by soft lithography of polymeric materials (Oh et al., 2008) and are either hydrophobic or hydrophilic depending whether an o/w or a w/o emulsion is being produced. The spontaneous detachment and relaxation into the spherical drop are driven by interfacial tension (Leal-Calderon et al., 2007) which can present a challenge in adapting microfluidics for use with highly viscous biopolymer solutions (Zhang et al., 2007). It is unlikely that microfluidics can be up scaled appropriately and economically for general food use, but it can be used to obtain valuable insights into controlling parameters necessary for the design of large scale processing equipment (Tran et al., 2011). At the laboratory scale, microfluidics can be used to produce microgels with specific shapes and internal structures. They also have the advantage of being monodisperse which enables the rate of release of actives such as flavours or nutraceuticals to be predicted and tuned for desired product characteristics (Zhang et al., 2007; Amici et al., 2008). The second main advantage is that this is one of the few reliable and repeatable methods for producing double emulsions such as water-in-oil-in-water (w/o/w) with monodisperse included droplets and monodisperse particles (Matalanis et al., 2011), as shown on the right of Fig. 3. Microfluidics is particularly suited to high value and specialist applications rather that large volume low value products.

H.M. Shewan, J.R. Stokes / Journal of Food Engineering 119 (2013) 781–792

787

Fig. 5. Microgel manufacture methods at: (a) lab scale, for example microfluidics to form alginate microgels reprinted with permission from (Amici et al., 2008); (b) Pilot scale, for example membrane emulsification followed by gelation reprinted with permission from (Vladisavljevic´ and Williams, 2005); and (c) industrial scale-spinning disk apparatus showing jets and drop break up reprinted with permission from (Southwest Research Institute, 2011).

One example of a larger scale device using insights from microfluidics is described by Maan et al. (2011). They use a novel method known as EGDE (Edge based Droplet GEneration), to produce monodisperse droplets of 10 lm in size. This design allows a significantly higher throughput than previous method due in part to its low susceptibility to fouling and has been used to produce whey protein microgels in sunflower oil. This technique shows promise for the food industry although it still requires further development as the physical EDGE equipment capable of producing 1 m3/h would require factory floor space of 0.3 m3 (van Dijke et al., 2010). 3.2.4. Membrane Membrane emulsification to form microgels involves pushing an aqueous solution through the membrane pores into an oil phase or air (typically), as demonstrated in Fig. 5b. Membrane pore size is reported as being the most significant parameter in determining the final droplet size (Tran et al., 2011) with trans-membrane pressure (a function of membrane design and the viscosity of the disperse phase) also having a significant influence on droplet size and size distribution (Liu et al., 2003). Membrane techniques have similar advantages to that of microfluidic techniques with additional benefit of energy efficiency (Yuan et al., 2009). Microgels ranging in size from 10 to 100 lm, corresponding to 2–10 times the membrane pore diameter, have been produced via membrane emulsification (Tran et al., 2011). A wide variety of membranes have been used to produce emulsions, including Shirasu porous glass, ceramic, silicone based nickel and stainless steel, with the choice of membrane being dependent on whether the disperse phase is hydrophilic or hydrophobic (Liu et al., 2003; Vladisavljevic´ and Williams, 2005; Bao et al., 2007; Yuan et al., 2009; Tran et al., 2011). The main advantages of this method are that it is useful for producing monodisperse spherical microgels with or without encapsulated ingredients as shown on the left of both Figs. 2 and 3. However, despite recent advances in membrane technology the process flow rate is still limited (Tran et al., 2011) and further developments are required to scale-up appropriately. 3.3. Physical formation of precursor droplets – atomisation route The production of microgels by atomisation of a biopolymer solution involves forming droplets in air, typically by breaking up a liquid stream using natural (Rayleigh) flow instabilities, ultrasonics or electrostatics. Gelation can be instigated by air temperature or through the penetration of a liquid stream or droplets directly into a gelation-promoting liquid. However, impact into another liquid can result in flattened and/or anisotropic microgels. There are a range of technologies available for atomisation including: airless atomisation, involving a stream of fluid under pressure that exits

an orifice as a jet which then breaks up into a stream of droplets; or spray to air-atomisation where the fluid exits with co-flowing jets of compressed air (Stokes, 2012). 3.3.1. Theory of drop break-up in air Break up of liquid streams in laminar flow conditions occurs through a Rayleigh instability. Small waves form at the interface of the stream of fluid and propagate along its length. These waves eventually cause the stream to break into spherical droplets, minimising the surface energy of the fluid (Malkin and Isayev, 2006). The maximum length of the stream before drop breakup (L) is dependent on the stream speed (u0), stream radius (Rs), density (q), and surface tension (cint), and can be determined as follows;

L ¼ 8:46u0

qR3s cint

!1=2 ð3Þ

This only holds for Newtonian liquid jets under laminar flow conditions; break up is significantly more complex when elastic fluids are used or when flow is turbulent. According to van Hoeve et al. (2010), a uniform stream is formed that leads into the Rayleigh instability when the Weber number (We, defined by Eq. (4)) of the fluid is less than 4 and the Weber number of the gas (Weg) is greater than 0.2. However these values should be used with caution as exact limiting values have not been agreed upon (Delteil et al., 2011). An alternative is to use the Weber, Ohnesorge (Oh) and Reynolds (Re) numbers described by Eqs. (4)–(7) in combination as shown in Fig. 6 (Sander and Weigand, 2008) to define the operating conditions for uniform droplet formation.

We ¼

qRS u20 cint

Weg ¼

qg We q

ð4Þ

ð5Þ

qRS u0 g

ð6Þ

pffiffiffiffiffiffiffi We Oh ¼ Re

ð7Þ

Re ¼

3.3.2. Spinning disk atomisation Spinning disk atomisation utilises a flow of liquid across spinning disk with drop break-up, due to Rayleigh instabilities, occurring at the edge of the disk, shown in Fig. 5c. The nozzle size, rate of spinning and flow can be controlled to give a defined particle size distribution and an active ingredient can be encapsulated by concurrent flow of the active and encapsulating fluid across the disk.

788

H.M. Shewan, J.R. Stokes / Journal of Food Engineering 119 (2013) 781–792

Fig. 6. Plot of Ohnesorge against the Reynolds numbers indicating mechanisms of drop breakup in each region reproduced with permission from (Sander and Weigand, 2008) with examples of flow in each regime reprinted with permission from (Lee et al., 2008).

This method has the advantage that it has very high production rates (Gouin, 2004) however particles are generally highly polydisperse. Senuma and Hilborn (1999) present a technique for modifying a standard spinning disk by adding: a sloped surface to minimise unstable flow patterns; teeth at the edge; and a method of collection based on flight distance of different sized particles to give a monodisperse particle size distribution. They found that the viscosity of the solution fed over the disk affects particle size distribution with higher viscosities resulting in a wider distribution. Spinning disk atomisation is commonly used in industrial spray drying/cooling processes and is a simple, efficient and cost effective method for producing spherical or elongated microgels with or without encapsulated ingredients (Dubey et al., 2009). 3.3.3. Spray nozzle This method produces droplets by passing a flow of liquid and air concurrently through a nozzle at a high flow rate. Droplet break up occurs due to fluid jet instabilities and can be controlled by adjusting air and polymer solution flow rates, polymer solution concentration and surface tension. Perrechil et al. (2011) verified this by producing j-carrageenan–sodium caseinate microgels in a range of shapes including spherical, ellipsoidal, long strings and irregular particles. Although single nozzles are commonly used in industrial spray drying the disadvantage of this method is the likelihood of blockages occurring in the spray nozzle particularly for viscous biopolymers (Gouin, 2004; Burey et al., 2008). 3.3.4. Extrusion Extrusion involves projecting an emulsion of core and coating material though a die; at microscale this requires extrusion of the jet through a nozzle with a diameter on the order of 100 lm down to a few microns (Burey et al., 2008). Jet break-up extrusion forms a laminar jet by forcing a polymer solution from the nozzle tip. One of several techniques may be applied to break the fluid jet into monodisperse droplets including: electrostatic generation, jet cutting and acoustic jet excitation. There are two modes for electrostatic drop break up: (1) dripping mode in which the fluid is gently pushed through the nozzle and a low current applied and; (2) jet mode at higher velocity to form a smooth stable jet which requires a higher electric current to break-up droplets. Mode (1) has very limited production rates whereas in mode (2) high productivity can be achieved to produce microgels of 1–15 lm (Tran et al., 2011). Jet cutting is carried out by use of cutting wires on a rotating tool. This is a simple method that is capable of production rates greater than 10 times that of the

other break-up methods and allows particle size to be controlled, however particle size is large (>150 lm) (Prusse et al., 2000). Acoustic jet excitation utilises vibration to create instabilities in the fluid jet to cause breakup. Particles from nano to millimetre size are possible by varying polymer concentration, flow rate vibration wavelength and cooling rate or distance to a ‘hardening’ bath (Tran et al., 2011). Microgels have been successfully produced from alginate, pectin, chitosan and gelatine using the jet cutting method (Prusse et al., 2000) and large scale jet extrusion has been used successfully to encapsulate fish oils (Southwest Research Institute, 2011). This is another simple method for formation of monodisperse microgels or microcapsules with potential for scale up for the food industry. Walther et al. (2004) have shown that highly anisotropic particles can be made by extruding hot carrageenan through a needle into cold oil then forcing both phases through a narrow channel. In the channel the drop hardens into a specific shape dependent on temperatures and flow rates of both the carrageenan and oil phases. They produced microgels that ranged from being slightly elliptical to ‘star’ shaped, as shown on the right of Fig. 3. The variation of shape was shown to influence microgel suspension rheology.

3.3.5. Spray drying and spray cooling The most common example of atomisation proven at large scale and commonly used in the food industry is spray drying. It has the advantage of forming a convenient dry powder product with extended shelf life and low storage and transport costs (Burey et al., 2008; Augustin and Hemar, 2009; Jones and McClements, 2010). Spray drying involves atomisation of liquid feed into a stream of hot air to evaporate the solvent – which in the case of food applications is water – followed by separation of the dried particles (Gouin, 2004; Burey et al., 2008; Augustin and Hemar, 2009; Dubey et al., 2009). Atomisation can be achieved by passing the fluid through a nozzle of a certain size or by passing the solution over a spinning disk. The particle size can be controlled by: selection of the nozzle or spinning disk; concentration and viscosity of the feed; and feed flow rate. Spray drying is suitable for volatile and heat sensitive ingredients due to the very short exposure of the particle to hot air (Augustin and Hemar, 2009; Jones and McClements, 2010). Disadvantages are the limited ability to use with non-water soluble or high viscosity biopolymers and the possible breakdown of the porous particle during rehydration (Gouin, 2004).

H.M. Shewan, J.R. Stokes / Journal of Food Engineering 119 (2013) 781–792

Spray cooling, which is the reverse of spray drying, involves atomisation of liquid via a heated nozzle and the use of cool air to solidify droplets into microgels; in this case the microgel is usually a lipid (Gouin, 2004; Augustin and Hemar, 2009). This is the least expensive encapsulation technology and is routinely used for the encapsulation of hydrophilic ingredients such as mineral salts, enzymes, and flavours (Gouin, 2004; Augustin and Hemar, 2009). Bhandari (2009) developed and patented a method for producing microgels utilising atomisation. In this technique a cross-linkable biopolymer is atomised from the nozzle at the top of an enclosed vessel similar to spray drying. In this case, instead of the gelation occurring through contact with hot air, it occurs within the vessel on contact of the droplets with a cross-linking reagent that is atomised from a nozzle at the base of the vessel. The example given is alginate droplets that gel on contact with calcium chloride droplets; this is considered a simple process for encapsulating actives such as fish oil. The method has been shown to produce uniform particles with a controlled particle size with little risk of contamination. 3.3.6. Rehydration of spray-dried biopolymer solutions Burey et al. (2009) describe a novel method for producing microspheres utilising spray drying. This method involves spray drying a cross-linkable solution in disordered (non-aggregated) form to produce solid particles that can be rehydrated at a temperature below the gelation temperature to form a discrete gel particle 2–30 times the size of the dry particle (Gidley and Hedges, 1998). This process can be used for biopolymers including gellan, carrageenan and agar. If gelation is more rapid than dissolution, then gelled particles are predicted to form that are dependent on the kinetics of these two processes. Fast gelation leads to the formation of small (10 lm) particles, while slow gelation results in a biopolymer solution that forms a weak continuous gel after an extended time (Burey et al., 2009). Varying these hydration conditions allows control over hydrated particle size and particle modulus and potential for tailoring their use in specific applications from encapsulation to texture modification (Stokes, 2012). This is another simple and easily scalable technique for large volume production of food grade microgels. 3.4. Physical formation of precursor droplets – microparticulation 3.4.1. Shear gels Shear gels, also referred to as fluid gels, are formed by shearing during gelation of biopolymers that would otherwise form a network structure if allowed to gel under quiescent conditions (Norton et al., 1999; Altmann et al., 2004). The particle size of these sheared gels can be varied by increasing shear rate to give smaller particle sizes. If the shear rate is greater than the relaxation time of the polymer droplet (also dependent on the continuous phase viscosity) then non-spherical droplets (see right hand side of Fig. 2) will be formed. Both agarose and j-carrageenan shear gels have been produced using a pin stirrer in a jacketed vessel (Gabriele et al., 2009, 2010). Application of shear during cooling resulted in spherical microgels although the shear rate, cooling rate and final temperature affected the exact size and shape of the gels and whether they were prone to aggregation. Another method involves application of shear to a biopolymer gel to comminute it into small particulates, which are typically polydisperse in size and highly irregular shapes; this is referred to as a ‘broken gel’ (Ellis and Jacquier, 2009b; Garcia et al., 2011). Jimenez-Colmenero et al. (2012) suggest that a broken gel formed by grinding a gelled konjac, carrageen and corn starch mixture could be used as a thermally stable substitute for pork back fat in meat products.

789

Particle morphology is of interest as it has a strong influence on the rheology of the shear gel suspensions (Ellis and Jacquier, 2009b; Gabriele et al., 2010; Stokes, 2012). Anisotropic drops can be formed by application of high shear rates during gelation. These sheared-gel suspensions with anisotropic particles tend to possess a low, yet finite, yield stress that allows ingredients such as herbs and oil to be suspended while still being pourable. Use of sheargels also eliminates the negative mouthfeel and stringiness associated with polymer-based thickeners and starches (Stokes and Frith, 2008). In terms of processing equipment and scale up, this method has the same advantages and disadvantages as the basic emulsion gelation method without having to separate microgels from oil, consequently it has potential to be used on a large scale.

3.4.2. Microparticulated proteins Protein bulk gels are formed by heating a thermally gelling protein solution so that the protein denatures by unfolding and then aggregates upon cooling to form a network gel structure (Stokes, 2012). Globular proteins can be denatured by heating to form microgels with hydrophobic attraction and disulphide formation causing them to self-associate (Matalanis et al., 2011). Protein can be denatured by acidification or heating followed by gelation by cooling, addition of ions such as iron or calcium or complexation with a polysaccharide (Chen et al., 2006; Jones et al., 2010a; Schmitt et al., 2011). The morphology of the microgels formed will be dictated by the native protein and also process conditions with possible shapes including fibrillar spherical or anisotropic ranging from nano size (40 nm) to beads (2 mm) (Chen et al., 2006; Jones and McClements, 2010). In general, fibrillar microgels are formed when the gelation process occurs at a pH far from the isoelectric point of the protein while particle gels are formed by gelation close to the isoelectric point (Augustin and Hemar, 2009). Two examples of protein microgels produced at the laboratory scale are from whey protein and from a lysozyme/ovalbumin mix (Yu et al., 2006; Schmitt et al., 2011). The whey protein isolates were mixed with water, adjusted to pH 5.7–5.8, heated to 85 °C without stirring before being rapidly cooled to 4 °C (Schmitt et al., 2011). It was concluded from this work that microgel formation was a function of the ratio between a and b lactoglobulin, and to a lesser extent on the mineral composition. Yu et al. (2006) followed the complexation route to produce microgels of size 100 nm with a predominantly lysozyme core and ovalbumin shell. The process used to generate these microgels involved mixing under acidic conditions, increasing to pH 10.3 while stirring, then heating to 80 °C (Yu et al., 2006). This process is very simple and easily scalable requiring only heating and mixing vessels currently used in the food industry. Microparticulates can also be formed by applying shear to interrupt the aggregation process of random coil proteins (Singer and Dunn, 1990; Singer, 1996; Sirikulchayanont et al., 2007) with both the process and product being the subject of patents (Singer et al., 1988; Kruesemann et al., 2010). As previously mentioned this process is currently operating at commercial scale for the production of SimplesseTM. Simultaneous homogenization and pasteurisation produces protein particles or protein gels ranging in size from 0.01 to 100 lm, although the exact particle morphology will depend on the solution pH, ionic strength and protein concentration (Dissanayake et al., 2010; Schmitt et al., 2011; Stokes, 2012). Microparticulates have been formed from a range of proteins including egg albumin, whey protein, mung bean protein, soy protein and gelatine (Singer et al., 1988; Singer and Dunn, 1990; Ziegler, 1992; Sirikulchayanont et al., 2007; Jones and McClements, 2010). Saglam et al. (2011) utilise a novel two-step emulsification process to produce microparticulate with >20% protein concentration.

790

H.M. Shewan, J.R. Stokes / Journal of Food Engineering 119 (2013) 781–792

3.5. Physicochemical formation of precursor droplets The physicochemical methods for producing microgels utilise forces such as electrostatic interactions or steric exclusion effects to separate mixed polymer solutions into a droplet phase and a continuous phase (Jones et al., 2010b; Matalanis et al., 2011). This route for microgel formation is commonly used in conjunction with one of the physical methods previously described. Physicochemical reactions, in particular complex formation and attractive interactions, between actives and carriers such as polysaccharides or proteins can also be used as a precursor to microgel formation to encapsulate an active ingredient (Matalanis et al., 2011). Biopolymer microgels can be formed from a single biopolymer solution by changing solution conditions such as pH or ionic strength to promote self-association, for example simple coacervation described in Section 3.5.1 (Matalanis et al., 2011). Mixed biopolymer solutions with opposite charge can also be used as a starting point for microgel formation as in complex coacervation and phase separation.

3.5.1. Coacervation Coacervation is a chemical method for producing polymer droplets in suspension and is defined by Kissel et al. (2006) as ‘‘the macromolecular aggregation process brought about by partial desolvation of fully solvated macromolecules’’. It is the separation of two liquid phases into one concentrated colloidal phase, being the coacervate, and another highly dilute colloidal phase (de Kruif et al., 2004). There are two types of coacervation, ‘simple’ and ‘complex’. Simple coacervation involves only one polymer and phase separation is brought about by addition of a salt, pH or temperature change, which occurs when alginate is mixed with calcium for example. Complex coacervation involves two polymers and phase separation is brought about by anion– cation interactions, the most commonly cited example of this being gelatine and gum arabic. Coacervation uses the principle of difference in ionic forces to cause the polymer(s) to form droplets and drop out of solution. A key to this is knowledge of the iso-electric point (PI) of the polymers and adjusting the formulation accordingly. The actual mechanisms are complex and vary for the different protein–polysaccharide mixtures, as reviewed by Turgeon et al. (2007). Electrostatic effects and other weak energy interactions, especially hydrogen bonding, play a very important role during complex formation/coacervation between proteins and polysaccharides. Hydrophobic interactions can also make a significant contribution to formation of complexes and coacervates between oppositely charged biopolymers (Turgeon et al., 2007). Although the mechanisms are complex, the method has the advantage that specialist equipment is not required to complete the process and hence it is easily scalable. Vilstrup (2001) discusses the wide ranging uses of the coacervation technique in food applications. There is a range of literature available covering biopolymer coacervates, many of which are directly applicable for food applications (de Kruif et al., 2004). The complexities are real, as discussed above, however the technique is well established for protein–polysaccharide and cationic–anionic polysaccharide mixtures, examples of which include whey protein or gelatine with gum arabic and chitosan-alginate (Gouin, 2004; Augustin and Hemar, 2009). The main disadvantage in this method is associated particularly with the use of gelatine and other biopolymers that require covalent cross-linking (as discussed in Section 3.1.2) to form a gel that will remain stable under extreme conditions such as when mixed with acidic food ingredients or when subjected to extremes of temperature during sterilisation or freezing.

3.5.2. Phase separated polymers If an aqueous solution contains two different kinds of biopolymer molecules that have sufficient repulsive force between them, it will separate into two aqueous phases; one that is rich in biopolymer A and depleted in biopolymer B, and vice versa (Norton and Frith, 2001; McClements, 2010). The conditions of electrostatic or steric repulsion that drive this can be adjusted to form a droplet of one phase dispersed in a continuous medium of the other (Stokes et al., 2001; McClements, 2010). A biopolymer mixture will undergo phase separation to minimise the free energy of the system, with the morphology of the final mixture dependent on the temperature, molecular ordering and the phase volume of each of the components (Norton and Frith, 2001; Butler and Heppenstall-Butler, 2003). The included phase forms spherical droplets of 2–100 lm when phase separation occurs under quiescent conditions (Wolf et al., 2000; Norton and Frith, 2001). In some cases shear is required to bring about phase separation or to form anisotropic particles from phase separated mixtures. Microgel particles can then be produced by cross-linking of the biopolymer in the droplet phase. Applying shear to a phase separated mixture and cooling, drops of various shapes and sizes are formed including spherical, ellipsoidal, short and long fibrils, as well as irregular shaped particles (Wolf et al., 2000, 2001; Leng and Turgeon, 2007). Phase separated biopolymer mixtures are referred to as water in water (w/w) emulsions, and they are governed by the same physical principles, including the rules for droplet break-up and coalescence, as for conventional water in oil emulsions (Wolf et al., 2000; Leng and Turgeon, 2007), but their interfacial tension is several orders of magnitude lower (Stokes et al., 2001). The advantages of microgel formation from w/w emulsions are threefold; the ability to form microgels without the addition of any surfactant; the formation of the microgels without an oil phase and; the ability to manipulate particle shape from spherical to highly anisotropic. These three advantages result in a biocompatible microgel that can be easily manufactured to impart the required mouthfeel and flow behaviour on a food product (Norton and Frith, 2001). Gelatine/maltodextran mixtures (Stokes et al., 2001; Butler and Heppenstall-Butler, 2003), gellan/ carrageenan (Wolf et al., 2000, 2001) and gelatine/guar mixtures (Wolf et al., 2000) are those most commonly reported in the literature.

4. Conclusion and outlook The methods used for microgel manufacture are many and varied and the suitability depends on the hydrocolloid or polymer carrier as well as the functionality, particle size and particle size distribution required. Many of the reviewed methods for producing microgels are accessible at industrial scale, although those methods that produce microgels with a monodisperse size distribution are currently best suited to laboratory scale manufacture only. Microgels in the food industry have been primarily targeted towards fat replacement, rheology control, texture modification and encapsulation. However, there are still many opportunities not yet realised in the food industry, for example, synthetic microgels are designed to be highly responsive yet this has not been addressed for food-grade microgels due to the restrictions in the ability to artificially functionalise biopolymers. The major limitations preventing widespread use of microgels in the food industry can be summarised as being due to: compatibility with foods; control when mixed with a range of ingredients; return on investment (added value compared to capital costs); and practicality. These are typical challenges faced with any ‘new’ technology in the food industry. The development of microgels for the food industry has only really been pursued in recent times and

H.M. Shewan, J.R. Stokes / Journal of Food Engineering 119 (2013) 781–792

there is considerable scope for major development in their use and manufacture. With the major need by manufacturers to enhance the consumer benefits in processed foods, microgel technologies may well create exciting opportunities in the future in the same way synthetic microgels revolutionised the coatings industry. Microgels can address several needs of the food industry; satiety control, encapsulation of phytonutrients and prebiotics, texture control for healthier food formulations (e.g. reduced fat products), and targeting delivery to specific areas in the digestive tract. Plenty of other exciting applications and benefits are likely to emerge with further research and development in their fundamental properties and manufacturing routes. References Adams, S., Frith, W.J., Stokes, J.R., 2004. Influence of particle modulus on the rheological properties of agar microgel suspensions. Journal of Rheology 48 (6), 1195–1213. Altmann, N., Cooper-White, J.J., Dunstan, D.E., Stokes, J.R., 2004. Strong through to weak ‘sheared’ gels. Journal of non-Newtonian Fluid Mechanics 124 (1–3), 129– 136. Amici, E., Tetradis-Meris, G., de Torres, P., Jousse, F., 2008. Alginate gelation in microfluidic channels. Food Hydrocolloids 22 (1), 97–104. Augustin, M.A., Hemar, Y., 2009. Nano- and micro-structured assemblies for encapsulation of food ingredients. Chemical Society Reviews 38 (4), 902–912. Babin, H., Dickinson, E., 2001. Influence of transglutaminase treatment on the thermoreversible gelation of gelatin. Food Hydrocolloids 15 (3), 271–276. Bao, D.C., Zhang, H.A., Liu, X.D., Zhao, Y.J., Ma, X.J., Yuan, Q., 2007. Preparation of monodispersed polymer microspheres by SPG membrane emulsificationsolvent evaporation technology. Journal of Dispersion Science and Technology 28 (3), 485–490. Bertoni, F., Barbani, N., Giusti, P., Ciardelli, G., 2006. Transglutaminase reactivity with gelatine: perspective applications in tissue engineering. Biotechnology Letters 28 (10), 697–702. Bhandari, B., 2009. Device and Method for Preparing Microparticles. PCTWO2009062254-A1 Australia. Burey, P., Bhandari, B.R., Howes, T., Gidley, M.J., 2008. Hydrocolloid gel particles: formation, characterization, and application. Critical Reviews in Food Science and Nutrition 48 (5), 361–377. Burey, P., Bhandari, B.R., Howes, T., Gidley, M.J., 2009. Gel particles from spray-dried disordered polysaccharides. Carbohydrate Polymers 76 (2), 206–213. Butler, M.F., Heppenstall-Butler, M., 2003. Phase separation in gelatin/dextran and gelatin/maltodextrin mixtures. Food Hydrocolloids 17 (6), 815–830. Chen, L.Y., Remondetto, G.E., Subirade, M., 2006. Food protein-based materials as nutraceutical delivery systems. Trends in Food Science and Technology 17 (5), 272–283. Chiou, B.-S., Avena-Bustillos, R.J., Shey, J., Yee, E., Bechtel, P.J., Imam, S.H., Glenn, G.M., Orts, W.J., 2006. Rheological and mechanical properties of cross-linked fish gelatins. Polymer 47 (18), 6379–6386. Clark, A.H., Amici, E., 2003. The formation and properties of biopolymer gels, In: Dickinson, E.V.T. (Ed.), Food Colloids, Biopolymers and Materials, pp. 35–48. de Bruijn, R.A., 1989. Thesis – Deformation and Breakup of Drops in Simple Shear Flows. University of Eindhoven, Eindhoven. de Kruif, C.G., Weinbreck, F., de Vries, R., 2004. Complex coacervation of proteins and anionic polysaccharides. Current Opinion in Colloid and Interface Science 9 (5), 340–349. Delteil, J., Vincent, S., Erriguible, A., Subra-Paternault, P., 2011. Numerical investigations in Rayleigh breakup of round liquid jets with VOF methods. Computers and Fluids 50 (1), 10–23. Dickinson, E., 2009. Hydrocolloids as emulsifiers and emulsion stabilizers. Food Hydrocolloids 23 (6), 1473–1482. Dissanayake, M., Kelly, A.L., Vasiljevic, T., 2010. Gelling properties of microparticulated whey proteins. Journal of Agricultural and Food Chemistry 58 (11), 6825–6832. Dubey, R., Shami, T.C., Rao, K.U.B., 2009. Microencapsulation technology and applications. Defence Science Journal 59 (1), 82–95. Ellis, A., Jacquier, J.C., 2009a. Manufacture of food grade kappa-carrageenan microspheres. Journal of Food Engineering 94 (3–4), 316–320. Ellis, A., Jacquier, J.C., 2009b. Manufacture and characterisation of agarose microparticles. Journal of Food Engineering 90 (2), 141–145. Esposito, E., Cortesi, R., Nastruzzi, C., 1996. Gelatin microspheres: influence of preparation parameters and thermal treatment on chemico-physical and biopharmaceutical properties. Biomaterials 17 (20), 2009–2020. Estrada, J.D., Boeneke, C., Bechtel, P., Sathivel, S., 2011. Developing a strawberry yogurt fortified with marine fish oil. Journal of Dairy Science 94 (12), 5760– 5769. Evans, I.D., Lips, A., 1990. Concentration dependence of the linear elastic behaviour of model microgel dispersions. Journal of the Chemical Society, Faraday Transactions 86 (20), 3413–3417. Fernandez-Neives, A., Wyss, H., Mattson, J., Weitz, D.A., 2011. Microgel Suspensions – Fundamentals and Applications. Wiley-VCH Verlag, Weinheim.

791

Frith, W.J., 2010. Mixed biopolymer aqueous solutions – phase behaviour and rheology. Advances in Colloid and Interface Science 161 (1–2), 48–60. Frith, W.J., Lips, A., 1995. The rheology of concentrated suspensions of deformable particles. Advances in Colloid and Interface Science 61, 161–189. Frith, W.J., Lips, A., Norton, I.T., 1999. Steady shear flow properties of spherical microgel suspensions. Structure and Dynamics of Materials in the Mesoscopic Domain, 207–218. Gabriele, A., Spyropoulos, F., Norton, I.T., 2009. Kinetic study of fluid gel formation and viscoelastic response with kappa-carrageenan. Food Hydrocolloids 23 (8), 2054–2061. Gabriele, A., Spyropoulos, F., Norton, I.T., 2010. A conceptual model for fluid gel lubrication. Soft Matter 6 (17), 4205–4213. Garcia, M.C., Alfaro, M.C., Calero, N., Munoz, J., 2011. Influence of gellan gum concentration on the dynamic viscoelasticity and transient flow of fluid gels. Biochemical Engineering Journal 55 (2), 73–81. Gidley, M.J.R., Hedges, N.D., 1998. Suspensions of Gelled Biopolymers. 5738897A. United States. Gouin, S., 2004. Microencapsulation: industrial appraisal of existing technologies and trends. Trends in Food Science and Technology 15 (7–8), 330–347. Hedges, N.D.S., 2009. Particle stabilised emulsion composition. 20090087464 United States. Homayouni, A., Azizi, A., Ehsani, M.R., Yarmand, M.S., Razavi, S.H., 2008. Effect of microencapsulation and resistant starch on the probiotic survival and sensory properties of synbiotic ice cream. Food Chemistry 111 (1), 50–55. Jimenez-Colmenero, F., Cofrades, S., Herrero, A.M., Fernandez-Martin, F., RodriguezSalas, L., Ruiz-Capillas, C., 2012. Konjac gel fat analogue for use in meat products: comparison with pork fats. Food Hydrocolloids 26 (1), 63–72. Jones, O.G., Lesmes, U., Dubin, P., McClements, D.J., 2010a. Effect of polysaccharide charge on formation and properties of biopolymer nanoparticles created by heat treatment of [beta]-lactoglobulin–pectin complexes. Food Hydrocolloids 24 (4), 374–383. Jones, O.G., Decker, E.A., McClements, D.J., 2010b. Comparison of protein– polysaccharide nanoparticle fabrication methods: impact of biopolymer complexation before or after particle formation. Journal of Colloid and Interface Science 344 (1), 21–29. Jones, O.G., McClements, D.J., 2010. Functional biopolymer particles: design, fabrication, and applications. Comprehensive Reviews in Food Science and Food Safety 9 (4), 374–397. Jun-xia, X., Hai-yan, Y., Jian, Y., 2011. Microencapsulation of sweet orange oil by complex coacervation with soybean protein isolate/gum Arabic. Food Chemistry 125 (4), 1267–1272. Karaca, O.B., Guven, M., Yasar, K., Kaya, S., Kahyaoglu, T., 2009. The functional, rheological and sensory characteristics of ice creams with various fat replacers. International Journal of Dairy Technology 62 (1), 93–99. Kissel, T., Maretschek, S., Packhauser, C., Schnieders, J., Seidel, N., 2006. Microencapsulation Techniques for Parenteral Depot Systems and Their Application in the Pharmaceutical Industry. In: Benita, S. (Ed.), Microencapsulation, Methods and Industrial Applications, second ed. CRC Press, Boca-Raton. Krieger, I.M., Dougherty, T.J., 1959. A mechanism for non-Newtonian flow in suspensions of rigid spheres. Transactions of the Society of Rheology 3 (1), 137– 152. Kruesemann, D., Nordanger, J., Krusemann, D., 2010. Treating Whey Protein Concentrate by Microparticulation Useful in Food Industry, Comprises Heat Treating Protein Concentrate in Tube Heat Exchanger Elements At Elevated Pressure and Mechanical Processing the Protein Concentrate. WO2008063115A1. Kutyla, M.J., Boehm, M.W., Stokes, J.R., Shaw, P.N., Davies, N.M., McGeary, R.P., Tuke, J., Ross, B.P., 2013. Cyclodextrin-crosslinked poly(acrylic acid): adhesion and controlled release of diflunisal and fluconazole from solid dosage forms. AAPS Pharm Science Technology 14 (1), 301–311. Leal-Calderon, F., Schmitt, V., Bibette, J., 2007. Emulsion Science, Basic Principles, second ed. Springer Science+Business Media, LLC., New York. Lee, L.Y., Lim, L.K., Hua, J., Wang, C.-H., 2008. Jet breakup and droplet formation in near-critical regime of carbon dioxide–dichloromethane system. Chemical Engineering Science 63 (13), 3366–3378. Leng, X.J., Turgeon, S.L., 2007. Study of the shear effects on the mixture of whey protein/polysaccharides – 2: application of flow models in the study of the shear effects on WPI/polysaccharide system. Food Hydrocolloids 21 (7), 1014– 1021. Li, Y., Kim, J., Park, Y., McClements, D.J., 2012. Modulation of lipid digestibility using structured emulsion-based delivery systems: comparison of in vivo and in vitro measurements. Food and Function 3 (5), 528–536. Liang, H.C., Chang, W.H., Liang, H.F., Lee, M.H., Sung, H.W., 2004. Crosslinking structures of gelatin hydrogels crosslinked with genipin or a water-soluble carbodiimide. Journal of Applied Polymer Science 91 (6), 4017–4026. Liang, H.C., Chang, W.H., Lin, K.J., Sung, H.W., 2003. Genipin-crosslinked gelatin microspheres as a drug carrier for intramuscular administration: in vitro and in vivo studies. Journal of Biomedical Materials Research Part A 65A (2), 271– 282. Liu, X.D., Bao, D.C., Xue, W.M., Xiong, Y., Yu, W.T., Yu, X.J., Ma, X.J., Yuan, Q., 2003. Preparation of uniform calcium alginate gel beads by membrane emulsification coupled with internal gelation. Journal of Applied Polymer Science 87 (5), 848– 852. Liu, Y., Kopelman, D., Wu, L.Q., Hijji, K., Attar, I., Preiss-Bloom, O., Payne, G.F., 2009. Biomimetic sealant based on gelatin and microbial transglutaminase: an initial

792

H.M. Shewan, J.R. Stokes / Journal of Food Engineering 119 (2013) 781–792

in vivo investigation. Journal of Biomedical Materials Research Part B – Applied Biomaterials 91B (1), 5–16. Loret, C., Frith, W.J., Fryer, P.J., 2007. Mechanical and structural properties of maltodextrin/agarose microgels composites. Applied Rheology 17 (3). Lyon, L.A., Fernandez-Nieves, A., 2012. The Polymer/Colloid Duality of Microgel Suspensions, In: Johnson, M.A., Martinez, T.J. (Eds.), Annual Review of Physical Chemistry, vol. 63, pp. 25–43. Maan, A.A., Schroen, K., Boom, R., 2011. Spontaneous droplet formation techniques for monodisperse emulsions preparation – perspectives for food applications. Journal of Food Engineering 107 (3–4), 334–346. Malkin, A.Y., Isayev, A.I., 2006. Rheology – Concepts, Methods, and Applications. ChemTec Publishing, Toronto. Matalanis, A., Jones, O.G., McClements, D.J., 2011. Structured biopolymer-based delivery systems for encapsulation, protection, and release of lipophilic compounds. Food Hydrocolloids 25 (8), 1865–1880. McClements, D.J., 2010. Emulsion design to improve the delivery of functional lipophilic components. Annual Review of Food Science and Technology 1, 241– 269. Mofidi, N., Aghai-Moghadam, M., Sarbolouki, M.N., 2000. Mass preparation and characterization of alginate microspheres. Process Biochemistry 35 (9), 885– 888. Mohammadi, R., Mortazavian, A.M., Khosrokhavar, R., da Cruz, A.G., 2011. Probiotic ice cream: viability of probiotic bacteria and sensory properties. Annals of Microbiology 61 (3), 411–424. Najafi, M.N., Kadkhodaee, R., Mortazavi, S.A., 2011. Effect of drying process and wall material on the properties of encapsulated cardamom oil. Food Biophysics 6 (1), 68–76. Neoh, K.G., Kang, E.T., 2010. Responsive surfaces for biomedical applications. Mrs Bulletin 35 (9), 673–681. Norton, I.T., Frith, W.J., 2001. Microstructure design in mixed biopolymer composites. Food Hydrocolloids 15 (4–6), 543–553. Norton, I.T., Jarvis, D.A., Foster, T.J., 1999. A molecular model for the formation and properties of fluid gels. International Journal of Biological Macromolecules 26 (4), 255–261. Norton, I.T., Spyropoulos, F., Heuer, E.A.K., Le Reverend, B.J.D., Cox, P.W., 2009. Microstructure Engineering of Healthy Everyday Foods. In: 5th International Symposium on Food Rheology and, Structure, pp. 10–14. Oh, J.K., Drumright, R., Siegwart, D.J., Matyjaszewski, K., 2008. The development of microgels/nanogels for drug delivery applications. Progress in Polymer Science 33 (4), 448–477. Perrechil, F.A., Sato, A.C.K., Cunha, R.L., 2011. [kappa]-Carrageenan–sodium caseinate microgel production by atomization: critical analysis of the experimental procedure. Journal of Food Engineering 104 (1), 123–133. Prata, A.S., Zanin, M.H.A., Ré, M.I., Grosso, C.R.F., 2008. Release properties of chemical and enzymatic crosslinked gelatin-gum Arabic microparticles containing a fluorescent probe plus vetiver essential oil. Colloids and Surfaces B: Biointerfaces 67 (2), 171–178. Prusse, U., Dalluhn, J., Breford, J., Vorlop, K.D., 2000. Production of spherical beads by JetCutting. Chemical Engineering and Technology 23 (12), 1105–1110. Quemada, D., 1977. Rheology of concentrated disperse systems and minimum energy-dissipation principle. 1. Viscosity–concentration relationship. Rheologica Acta 16 (1), 82–94. Rinaudo, M., 2006. Non-covalent interactions in polysaccharide systems. Macromolecular Bioscience 6 (8), 590–610. Rumscheidt, F.D., Mason, S.G., 1961. Particle motions in sheared suspensions XII. Deformation and burst of fluid drops in shear and hyperbolic flow. Journal of Colloid Science 16 (3), 238–261. Sagis, L.M.C., de Ruiter, R., Miranda, F.J.R., de Ruiter, J., Schroen, K., van Aelst, A.C., Kieft, H., Boom, R., van der Linden, E., 2008. Polymer microcapsules with a fiberreinforced nanocomposite shell. Langmuir 24 (5), 1608–1612. Saglam, D., Venema, P., de Vries, R., Sagis, L.M.C., van der Linden, E., 2011. Preparation of high protein micro-particles using two-step emulsification. Food Hydrocolloids 25 (5), 1139–1148. Sander, W., Weigand, B., 2008. Direct numerical simulation and analysis of instability enhancing parameters in liquid sheets at moderate Reynolds numbers. Physics of Fluids 20 (5). Sandoval-Castilla, O., Lobato-Calleros, C., Aguirre-Mandujano, E., Vernon-Carter, E.J., 2004. Microstructure and texture of yogurt as influenced by fat replacers. International Dairy Journal 14 (2), 151–159. Schmidt, H.T., Kroczynski, M., Maddox, J., Chen, Y., Josephs, R., Ostafin, A.E., 2006. Antibody-conjugated soybean oil-filled calcium phosphate nanoshells for targetted delivery of hydrophobic molecules. Journal of Microencapsulation 23 (7), 769–781. Schmitt, C., Bovay, C., Vuilliomenet, A.-M., Rouvet, M., Bovetto, L., 2011. Influence of protein and mineral composition on the formation of whey protein heatinduced microgels. Food Hydrocolloids 25 (4), 558–567.

Senuma, Y., Hilborn, J.G., 1999. Key parameters for monodispersed polymer microspheres with spinning disk atomization. Materials Research Innovations 3 (1), 42–49. Shewan, H.M., Stokes, J.R., Provisional 2012. Food Additive. 2012902202 Australia. Singer, N.S., 1996. Microparticulated Proteins as Fat Mimetics. In: Jones, S.A., Roller, S. (Eds.), Handbook of Fat Replacers. CRC press LLC, Boca Raton, Florida. Singer, N.S., Dunn, J.M., 1990. Protein microparticulation – the principle and the process. Journal of the American College of Nutrition 9 (4), 388–397. Singer, N.S., Latella, J., Yamamoto, S., 1992. Reduced Fat Yoghurt Having Fat Wholly or Partly Replaced by Micro-particulated Protein. US5096731-A. Singer, N.S., Wilcox, R., Podolski, J.S., Chang, H.H., Pookote, S., Dunn, J.M., Hatchewell, L., Hatchwell, L., 1989. Mayonnaise and Salad Dressings for Use in Desserts, Sauces, Icing, Pipe Fillings, Spreads, Dips, etc. WO8905587-A2. Singer, N.S., Yamamoto, S., Latella, J., 1988. Protein Product Base. US 47342875. USA. Sirikulchayanont, P., Jayanta, S., Pradipasena, P., Miyawaki, O., 2007. Characteristics of microparticulated particles from mung bean protein. International Journal of Food Properties 10 (3), 621–630. SouthwestResearchInstitute, 2011. Micro/Nano Encapsulation, San Antonio, Texas. Stokes, J.R., 2011. Rheology of Industrially Relevant Microgels. In: FernandezNeives, A., Wyss, H., Mattson, J., Weitz, D.A. (Eds.), Microgel Suspensions – Fundamentals and Applications. Wiley-VCH Verlag, Weinheim. Stokes, J.R., 2012. Food Biopolymer Gels, Microgel and Nanogel Structures, Formation and Rheology. In: Bhandari, B.Y.R. (Ed.), Food Materials Science and Engineering. Wiley-Blackwell, UK. Stokes, J.R., Frith, W.J., 2008. Rheology of gelling and yielding soft matter systems. Soft Matter 4 (6), 1133–1140. Stokes, J.R., Wolf, B., Frith, W.J., 2001. Phase-separated biopolymer mixture rheology: prediction using a viscoelastic emulsion model. Journal of Rheology 45 (5), 1173–1191. Surh, J., Decker, E.A., McClements, D.J., 2006. Properties and stability of oil-in-water emulsions stabilized by fish gelatin. Food Hydrocolloids 20 (5), 596–606. Tan, B.H., Pelton, R.H., Tam, K.C., 2010. Microstructure and rheological properties of thermo-responsive poly(N-isopropylacrylamide) microgels. Polymer 51 (14), 3238–3243. Tran, V.-T., Benoit, J.-P., Venier-Julienne, M.-C., 2011. Why and how to prepare biodegradable, monodispersed, polymeric microparticles in the field of pharmacy? International Journal of Pharmaceutics 407 (1–2), 1–11. Turgeon, S.L., Schmitt, C., Sanchez, C., 2007. Protein–polysaccharide complexes and coacervates. Current Opinion in Colloid and Interface Science 12 (4–5), 166– 178. Ugwoke, M.I., Kinget, R., 1998. Influence of processing variables on the properties of gelatin microspheres prepared by the emulsification solvent extraction technique. Journal of Microencapsulation 15 (3), 273–281. van Dijke, K.C., Schroen, K., van der Padt, A., Boom, R., 2010. EDGE emulsification for food-grade dispersions. Journal of Food Engineering 97 (3), 348–354. van Hoeve, W., Gekle, S., Snoeijer, J.H., Versluis, M., Brenner, M.P., Lohse, D., 2010. Breakup of diminutive Rayleigh jets. Physics of Fluids 22 (12). Vilstrup, P., 2001. Microencapsulation of Food Ingredients, first ed. Leatherhead Publishing, London, England. Vladisavljevic´, G.T., Williams, R.A., 2005. Recent developments in manufacturing emulsions and particulate products using membranes. Advances in Colloid and Interface Science 113 (1), 1–20. Walther, B., Hamberg, L., Walkenstrom, P., Hermansson, A.M., 2004. Formation of shaped drops in a fast continuous flow process. Journal of Colloid and Interface Science 270 (1), 195–204. Wolf, B., Frith, W.J., Norton, I.T., 2001. Influence of gelation on particle shape in sheared biopolymer blends. Journal of Rheology 45 (5), 1141–1157. Wolf, B., Scirocco, R., Frith, W.J., Norton, I.T., 2000. Shear-induced anisotropic microstructure in phase-separated biopolymer mixtures. Food Hydrocolloids 14 (3), 217–225. Wolfe, M.S., Scopazzi, C., 1989. Rheology of swellable microgel dispersions: influence of crosslink density. Journal of Colloid and Interface Science 133 (1), 265–277. Yu, S.Y., Yao, P., Jiang, M., Zhang, G.Z., 2006. Nanogels prepared by self-assembly of oppositely charged globular proteins. Biopolymers 83 (2), 148–158. Yuan, Q.C., Aryanti, N., Gutierrez, G., Williams, R.A., 2009. Enhancing the throughput of membrane emulsification techniques to manufacture functional particles. Industrial and Engineering Chemistry Research 48 (19), 8872–8880. Zhang, H., Tumarkin, E., Sullan, R.M.A., Walker, G.C., Kumacheva, E., 2007. Exploring microfluidic routes to microgels of biological polymers. Macromolecular Rapid Communications 28 (5), 527–538. Ziegler, G.R., 1992. Process for producing microparticulated protein and the product thereof. US 5 (147), 677.