- Email: [email protected]
mastic gum microsized particles composites
ARTICLE IN PRESS
FOOD
HYDROCOLLOIDS Food Hydrocolloids 22 (2008) 854–861 www.elsevier.com/locate/foodhyd
The structural characteristics and mechanical properties of biopolymer/mastic gum microsized particles composites Christos Mavrakis, Vassilis Kiosseoglou Laboratory of Food Chemistry and Technology, School of Chemistry, Aristotle University of Thessaloniki, 54124 Thessaloniki, Greece Received 20 December 2006; accepted 31 March 2007
Abstract Biopolymer/mastic gum (MG) particles composites were prepared by incorporating microsized MG particles in gel matrices based on gelatin, egg white or polysaccharides and their mechanical properties as influenced by MG content and particle size were investigated. Observation studies under the microscope were conducted to determine the structural features of the MG composites. The findings were combined with data on protein adsorption to MG particle surface to discuss the mechanical properties of the gels, which differed substantially depending on the biopolymer involved in gel network structure development. A mechanism is put forward to explain the role of MG particles as filler materials of biopolymer gel matrices. r 2007 Elsevier Ltd. All rights reserved. Keywords: Composite gel; Filler effects; Protein; Polysaccharide; Mechanical properties
1. Introduction The structure and rheological behaviour of a great number of traditional semi-solid foods depend to an appreciable extent on the presence and interactions within the food system between biopolymeric molecules, namely proteins or polysaccharides. Additionally, due to their vast diversity, both in structure and functionality, biopolymers are extremely useful as food additives to scientists involved in product development in designing new foods that meet the consumer demand for cheaper and healthier products which at the same time exhibit novel structural and textural characteristics (Aguilera & Stanley, 1999; Kiosseoglou, 2005). When the biopolymeric molecules interact, usually through non-covalent bonds, a network structure develops that entraps and immobilizes the solvent as well as all the other low-molecular weight constituents of the system. The gel structure differs depending on the type of polymer involved. Protein molecules normally tend to interact, Corresponding author. Tel.: +302310997834; fax: +302310997779.
E-mail address: [email protected] (V. Kiosseoglou). 0268-005X/$ - see front matter r 2007 Elsevier Ltd. All rights reserved. doi:10.1016/j.foodhyd.2007.03.011
following heat treatment and unfolding, at temperatures above their denaturation point. Polysaccharides, on the other hand, as a rule produce gel network structures when a solution is quenched either in the absence or presence of gelation aids (e.g. Ca2+, sugar, etc.) (Morris, 1998). Gelatin, a degradation product of collagen, in spite of being a protein behaves like a polysaccharide and produces gel networks on cooling gelatin solutions of a relatively low concentration. Molecular interactions in gelatin networks rely on the formation of junction zones where molecular segments from different gelatin molecules interact and form an extensive triple helical structure (Guo, Colby, Lusignan, & Howe, 2003). Many semi-solid food systems are composites having a gel network structure where a particulate material, the filler, is embedded. The rheological properties of the filled network structure as well as the textural characteristics of the food could be significantly affected by the presence of particles such as meat fibers, starch granules or emulsified oil droplets. Parameters such as the filler size and shape and the strength of filler surface–gel matrix interaction should be taken into account when considering the reinforcing effect of a filler particle on a biopolymer
ARTICLE IN PRESS C. Mavrakis, V. Kiosseoglou / Food Hydrocolloids 22 (2008) 854–861
network (Aguilera, 1992; Anton, Le Denmat, Beaumal, & Pilet, 2001; Thoufeek Ahamed, Singhal, Kulkami, Kale, & Palb, 1996). Reinforcing fillers of inorganic nature, including hydroxyapatite clay, glass and carbon particles or nanotubes are widely used to improve the mechanical properties of products such as fiberglass, rubber or natural and synthetic polymer matrices (Frogley, Ravich, & Wagner, 2003; Gauthier, Chazeau, Prasse, & Cavaille, 2005; Langley, Martin, & Ogin, 1994).When suitably incorporated, these fillers may bring about a substantial improvement of the mechanical properties of the gel structure due to effective load transfer from the matrix to the much harder filler particle through the particle surface which should interact with the matrix. The choice, however, for fillers, suitable for use in food systems, is very limited. One such filler of organic nature is microcrystalline cellulose which as reported by Kasapis (1999) may reinforce the structure of gelatin gels. Another material that could also be used in this respect is mastic gum (MG), which is the dried exudate obtained from the tree Pistacia lentiscus L., (Anacardiaceae), a native of the East Mediterranean and also of the Middle East countries. Chios MG is obtained in the form of drops from bark incisions on the Chios variety of the tree cultivated in the south of the island of Chios (Greece). The drops are then allowed to dry out in the air. The gum, a rather hard and brittle material at room temperature, has been finding use since ancient times as an antiseptic in medicine, as food antioxidant and as chewing gum base. The annual production yield of MG ranges between 100 and 150 tons and is used mainly as chewing gum base although a plasticizer is usually added to reduce its undesirable hardness and adhesiveness (Kehayoglou, Doxastakis, & Kiosseoglou, 1996). Additionally, the gum is exploited as flavouring additive in an increasingly expanding number of traditional foods and drinks (ice-cream, confectionery, baked products, liqueurs, etc.). As experience shows, MG in certain semi-solid products (ice-cream or baked products) behaves not only as flavouring agent but also as texture modifier, acting probably as filler of the protein-based gel matrixes, and could, therefore, be exploited as a natural food additive in order to develop novel structures that appeal to the consumer. The aim of the present work was to investigate the role of Chios MG as filler material of food biopolymer gel network structures, which determine to a large extent the mechanical and textural characteristics of most of semisolid food products. As the gum is a non-polar material its mode of action as filler will depend on the adsorption behaviour of the biopolymeric molecules to the filler particle surface and, therefore, on the molecular structure of the biopolymer involved. The ultimate objective of the study was to establish whether MG, apart from its outstanding flavouring effect in certain food products, may also affect their mechanical properties when present at relative low concentrations.
855
2. Materials and methods 2.1. Materials Chios MG granules were bought from the local market. The granules were ground to powder for 5 min in a mill fitted with knives and the resulting powder was fractionated into samples consisting of particles of various sizes with the aid of a set of sieves of mesh sizes 125, 90, 75 and 45 mm on a mechanical vibrator operated for 15 min. Agar, carrageenan (mainly k-carrageenan), gelatin (type A, from porcine skin, 93% w/w protein content), gum arabic and spray-dried egg white (85% w/w protein content) were obtained from Sigma Chemical Co. (St. Louis, MO). Tween 40 was a product of Fluka AG (Buchs, Switzerland). All the chemicals used in the experiments were of analytical grade. 2.2. Particle size determination The MG powder fractions were dispersed in a 1% (w/v) Tween solution in distilled water under continuous stirring for at least 1 h to obtain 3% (w/v) MG dispersions. The particle size distribution was determined by applying a laser scattering technique with the aid of the Malvern Mastersizer 2000 (Malvern Instruments Ltd., Malvern, Worcestershire, UK). The mean particle size, d3,2 and d4,3, is referred to as apparent mean size since most of the particles were not spherical. 2.3. Gel preparation Gel samples based on gelatin were prepared by dissolving 3 g of protein in 100 ml phosphate buffer (1/10 M Na2HPO4 and KH2PO4) of pH 7 with the aid of a magnetic stirrer and heating at 80 1C for 2 h. The MG powder was then incorporated under continuous stirring for 2 h and the resulting dispersion was transferred to cylindrical cells of 1 cm diameter made from aluminium foil and reinforced with plastic tape. The cells were placed at 3 1C for 24 h, then cut open, and cylindrical samples of 1 cm height were obtained. To prepare the agar and carrageenan gels, 2 g of gum arabic were initially dissolved in 100 ml buffer followed by the dispersion of the MG powder. The agar or carrageenan (3 g) was then dissolved at 80 1C under continuous stirring for 1 h, the dispersions were placed in cylindrical cells and cooled at 3 1C for 24 h as describe above. The gel based on egg white was prepared by first dissolving the white in the buffer to obtain a 10% solution in protein. Following dispersion of the MG powder under continuous stirring for 2 h, cylindrical cells were filled with the MG dispersion and heated at 90 1C for 3 min. The cells were cut open, following storage for 24 h at room temperature, and cylindrical samples 1 cm in height were obtained. 2.4. Compression test The cylindrical gel samples were subjected to a 80% compression at a speed of 0.5 mm/s using the Stable Micro
ARTICLE IN PRESS 856
C. Mavrakis, V. Kiosseoglou / Food Hydrocolloids 22 (2008) 854–861
System TA-XT2i Texture Analyzer (Surrey, England) equipped with a 2.5 cm diameter cylinder. Bulging of sample middle part was prevented by applying vegetable oil to the plates in contact with the sample surfaces. The force–time curves obtained from the analyzer were converted into compression stress-Hency strain ones (van Vliet, 1999). Liquid loss under pressure was determined (Funami, Yada, & Nakao, 1998) from the liquid expressed from a 5 mm thick gel sample compressed (20%) for 2 min between layers of filter paper with the aid of Texture Analyser. 2.5. Optical microscopy Optical microscopy images of MG dispersions in biopolymer solutions before and after heating were obtained by an Axiolab A reflected light microscope (Zeiss, Berlin, Germany) equipped with a Power Shot G2 photographic camera (Canon, Tokyo, Japan). 2.6. Amount of protein adsorbed to MG surface To determine the amount of protein adsorbed per unit particle surface, solutions of gelatin and egg white were prepared and MG powder of a mean particle size, d3,2 45–75 mm was dispersed with the aid of a magnetic stirrer for 30 min. Following centrifugation at 10,000g for 30 min, the protein content of the supernatant was determined according to Lowry, Rosebrough, Farr, and Randall (1951). The amount of protein adsorbed per unit surface was calculated using Gs ðmg=m2 Þ ¼ GT =ST , where GT is the amount of protein adsorbed in the system and ST is the total particle surface derived by applying the equation: S T ¼ 6V =d 3;2 , where V is the MG volume obtained assuming that the density of MG is 1.06 g/cm3.
solution, are presented in Fig. 1. The particle diameters are reported as apparent diameters and were calculated from particle size dispersion data provided by the Malvern sizer. The mean diameter values corresponded very satisfactorily to those expected according to the mesh size of the sieves used for particle fractionation. The dispersions of MG particles in gelatin and agar gel systems, along with their respective dispersions in cold biopolymer solutions, are illustrated in Fig. 2, while Fig. 3 exhibits MG dispersions in egg white solutions and gels. The marked differences observed between the composites based on the three biopolymeric materials apparently reflected differences in the mechanism behind gel network formation and probably in the extent and strength of interaction between the biopolymer matrix and the MG particle surface. MG particles appeared to become satisfactorily dispersed in both gelatin and gum arabic solutions (Fig. 2A and C, respectively). When, however, agar was introduced in the latter, aided by the application of heat, the particles melted and turned into spheres while at the same time they flocculated and formed large particle aggregates (Fig. 2D). Particle flocculation was not observed in the case of gelatin MG dispersions except that the particles upon heating also acquired a spherical shape remaining, however, fully dispersed in the gelatin matrix (Fig. 2B). This apparently is an indication that the MG particles were better protected against flocculation during heating by the gelatin molecules than by gum arabic/agar. As shown in Fig. 4, gelatin adsorbed to the MG particle surface and the surface protein load reached a plateau at higher protein concentrations corresponding to surface coverage of around 3 mg/m2. Gelatin in solution possesses a randomly coiled structure. Upon adsorption to solid surfaces a number of protein chain segments, described as ‘‘trains’’, attach to the surface leaving free loops and molecular ‘‘tails’’ extending into the bulk solution (Braithwaite, Luckham, & Howe, 1999). Gelatin attachment to hydrophilic surfaces is mainly governed by electrostatic interactions while adsorption on non-polar surfaces is mainly the result of hydrophobic binding that
2.7. Statistical analysis
3. Results and discussion 3.1. Structure of biopolymer/MG composites
160 140 d3.2 and d4.3 (μm)
All experiments were repeated at least three times and the data were analyzed using the one-way ANOVA program. The level of confidence was 95%. Significant differences between means were identified by the LSD procedure using the statistical software package SPSS v. 8.0 for Windows (SPSS Inc., Chicago, IL).
120 100 80 60 40 20 0
The mean MG particle diameters, d3,2 and d4,3, of the powder fractions, determined by applying the laser light scattering technique to MG dispersions in 1% Tween
90-125μm
75-90μm
45-75μm
<45μm
Fig. 1. Apparent mean particle size [d3,2 ( ) and d4,3 (&)] of the MG particle fractions determined by laser light scattering.
ARTICLE IN PRESS C. Mavrakis, V. Kiosseoglou / Food Hydrocolloids 22 (2008) 854–861
857
Fig. 2. Optical microphotographs of MG particle (mesh size o45 mm) dispersions in 3% gelatin solution (A) and gel (B) or in 2% gum arabic solution (C) and in 3% agar gel (D).
Fig. 3. Optical microphotographs of 10% egg white solution (A), of 6% (w/v) MG particles dispersion in the egg white solution (B and C for mesh size o45 and 90–125 mm, respectively) and of a 10% egg white gel containing 6% MG particles of mesh size 90–125 mm (D).
overcomes the electrostatic repulsion when both the particle surface and the protein molecule carry charges of the same sign (Kamyshny, Toledano, & Magdassi, 1999;
Thomas, Kellaway, & Jones, 1991). The presence of this adsorbed protein layer at the particle surfaces may then protect the particles against flocculation probably
ARTICLE IN PRESS C. Mavrakis, V. Kiosseoglou / Food Hydrocolloids 22 (2008) 854–861
858
6
Γs (mg/m2)
5 4 3 2 1 0 0
0.1
0.2 0.3 0.4 concentration (%w/v)
0.5
0.6
and the gel matrix. As preliminary experiments have shown, gum arabic also adsorbed to the particle surface producing a well-dispersed and stable system. Once, however, agar was introduced by heating, particle flocculation took place suggesting that depletion phenomena (Dickinson, 2003) might have operated due to the exclusion of agar molecules from a area near the particle surface as they do not adsorb to the surfaces, either directly or indirectly, through interaction with the adsorbed gum arabic molecules. Egg white is a mixture of soluble proteins of globular nature with the exception of ovomucin, which tends to
Fig. 4. Gelatin (m) and egg white protein (’) surface load, Gs, versus protein concentration for MG particle dispersions (mesh size 45–75 mm) in protein solutions.
relative elasticity modulus [-]
16 14 12 stress [kN.m-2]
3.5
10 8
3 2.5 2 1.5 1 0.5 0
6
control
+ 3% MG (45-75μm)
+ 6% MG (45-75μm)
+ 12% MG (45-75μm)
control
+ 3% MG (45-75μm)
+ 6% MG (45-75μm)
+ 12% MG (45-75μm)
4 2
1.8 1.6 0
0.2
0.4
0.6
0.8 1 strain [-]
1.2
1.4
1.6
1.8
140
stress [kN.m-2]
120
relative stress [-]
0
1.4 1.2 1 0.8 0.6 0.4
100
0.2
80
0
60 40
1.2
20
0
0.2
0.4
0.6
0.8 1 strain [-]
1.2
1.4
1.6
1.8
Fig. 5. Stress versus strain curves, (A) for egg white and gelatin ((m) and (’), respectively) and (B) for agar and carrageenan ((m) and (’), respectively) gels (filled symbols) and for the same gels containing 6% MG particles of mesh size 45–75 mm (open symbols).
relative strain [-]
1
0
0.8 0.6 0.4 0.2 0 control
through steric repulsive interactions, originating from overlapping ‘‘tail’’ and ‘‘loop’’ segments of gelatin molecules adsorbed on neighbouring particles that approach each other to very short distances, resulting in a system with MG particles evenly dispersed in the protein solution
+ 3% MG (45-75μm)
+ 6% MG (45-75μm)
+12% MG (45-75μm)
Fig. 6. Effect of MG concentration on the relative elasticity modulus (A) and relative stress (B) and strain (C) at fracture values for egg white (&), gelatine (B), agar (J) and carrageenan (n) gels containing MG particles of mesh size 45–75 mm.
ARTICLE IN PRESS C. Mavrakis, V. Kiosseoglou / Food Hydrocolloids 22 (2008) 854–861
form a weak gel-like structure responsible for the mucous appearance of the white. When the dehydrated egg white was dispersed in water a number of threadlike aggregates were spotted against a homogeneous background under the microscope (Fig. 3A). These aggregates may constitute dispersed ovomucin network structures as well as ovalbumin–conalbumin aggregates formed during egg white dehydration. In fact, as Weijers, van de Velde, Stijnman, van de Pijpekamp, and Visschers (2006) point out, the latter are responsible for the opaque appearance of gels prepared by heating the white, which is an indication of a non-homogeneous gel network matrix made up of randomly aggregating, denatured protein molecules. Incorporation of MG particles in egg white solution resulted in phase separation with the particles aggregating and forming a thread-like network spanning the total volume of the system. The thickness of the treads appears to depend on the particle size as the large-sized MG particles produced a less dense and more coarse flocculated network (Fig. 3B and C). Following heating and gel matrix formation, the initial structure did not change except that the MG particles became more spherical (Fig. 3D). 3.2. Mechanical properties and particle surface–matrix interaction When the gel samples were subjected to compression, their structure yielded at a strain value that depended on the type of biopolymer involved in gel formation. The polysaccharide gels fractured at a relatively lower strain value than the protein gels (Fig. 5). When MG was incorporated the gel resistance to compression was modified, depending on the biopolymer involved in net-
859
work structure development. Thus, MG particles at a concentration level of 6%, appeared to have a reinforcing effect on the structure of egg white gels since the samples yielded at a higher stress, than those of the control, while the slope of the stress–strain curve became steeper indicating an increase in gel elasticity. The reinforcing effect of MG on gelatin-based gels was less obvious as the value of the stress at fracture was not affected while the increase in the slope of the stress–strain curve was only marginal (Fig. 5A). In the case of polysaccharide gels, on the other hand, incorporation of MG resulted in a marked reduction of the gel resistance to compression (Fig. 5B). The relative change with MG content of stress/strain at fracture and elasticity modulus, determined from the initial slope of the stress–strain curve at small deformation, is exhibited in Fig. 6. In general, the strain at fracture appears to decrease with MG content for all the gel samples, the decrease being more spectacular in the case of agar. The relative stress at fracture increased with the MG content only in the case of the egg white gels. An even more spectacular increase was observed for Young’s elasticity modulus indicating that the particles functioned as active fillers and reinforced the gel network structure. Finally, the mechanical strength of the polysaccharide gels was inversely correlated with MG content and a negative filler effect was the outcome of MG incorporation, indicating that the network structure was interrupted by the presence of MG particles. As was discussed in the previous section, heating of egg white above the denaturation temperature of its constituent proteins, led to partial protein unfolding followed by interaction and gel network formation. The opaque appearance of the resulting gel indicates that non-ordered interaction between the unfolded protein molecules took
Table 1 Strain and stress at fracture and elasticity modulus values of egg white, gelatin, carrageenan and agar gels containing 6% MG particles of varying mean size Strain []
Stress (kN m2)
Elasticity modulus (kN m2)
Egg white 10%w/v +6% MG (90–125 mm) +6% MG (75–90 mm) +6% MG (45–75 mm)
0.8570.007a 0.7070.011b 0.7170.026b 0.7670.010c
7.770.15a 8.270.07b 8.570.06c 11.870.10d
Gelatin 3%w/v +6% MG (90–125 mm) +6% MG (75–90 mm) +6% MG (45–75 mm)
1.2570.012a 0.9670.005b 0.9870.007c 1.0970.005d
12.570.50a 9.070.34b 9.370.27b 12.770.13a
Carrageenan 3%w/v +6% MG (90–125 mm) +6% MG (75–90 mm) +6% MG (45–75 mm)
0.5170.001a 0.4570.002b 0.4270.019c 0.4170.001c
121.870.32a 93.870.72b 82.170.74c 81.270.98c
253.273.54a 222.972.50b 206.472.90c 206.072.64c
Agar 3%w/v +6% MG (90–125 mm) +6% MG (75–90 mm) +6% MG (45–75 mm)
0.2170.001a 0.2070.005b 0.1870.003c 0.1270.004d
40.670.34a 30.971.50b 25.970.41c 5.670.05d
187.072.72a 158.971.47b 145.472.47c 49.170.25d
a–d
Different letters indicate significant difference *po0.05.
8.070.51a 10.770.11b 12.370.25c 14.370.31d 4.670.18a 5.270.10a,b 5.570.20b 5.670.52b
ARTICLE IN PRESS 860
C. Mavrakis, V. Kiosseoglou / Food Hydrocolloids 22 (2008) 854–861
place, producing a non-homogeneous gel structure with regions of high polymer concentration and ones nearly devoid of polymer (Doi & Kitabatake, 1997). MG particles tended to accumulate at the polymer-depleted regions where they assembled and produced a thread-like structure embedded in the primary egg white gel matrix (Fig. 3). The formation of this secondary network, in turn, reinforces the gel structure as it can bear the stress transferred from the primary network, through extensive interaction between the gel network and the MG particle surfaces, which is a prerequisite for embedded particles to act as active fillers of gel structure (Aguilera, 1992). When the mean particle size was decreased, the MG particle thread-like structure became denser with thinner particle chains, leading to a higher mechanical strength (Table 1). Gelatin produces homogeneous, translucent and elastic gels. Due to adsorption of gelatin molecules to the particle surface, the MG particles became fully dispersed both in the gelatin solution and the gel network structure. Their incorporation, however, did not appear to have any effect on the mechanical properties of gelatin gel. A similar effect was also reported by Langley and Green (1989) for gelatin gels incorporating glass beads at relatively low concentration levels and only above 20% the glass spheres acted as active filler of the matrix. In contrast to egg white proteins which when adsorb to the particle surfaces unfold only to a limited extent, due to the presence in their molecules of disulfide bridges, the highly flexible gelatin molecules are expected to spread at the surface. As a result of this, interactions with the network matrix should only be confined to those molecular segments that constitute the ‘‘tails’’ of the adsorbed molecules and may become involved in triple helix formation with the matrix proteins (Fig. 7A). The lack of extensive interaction between the particle surfaces and the network should also explain the relatively marginal influence of MG particle size on the mechanical properties of the gelatin gels (Table 1). In the case of egg white, on the other hand, extensive hydrophobic and disulfide bond formation may take place between the adsorbed protein molecules and those involved in gel structure development, leading to filler reinforcement effects by the incorporated MG particles at relatively lower concentrations compared to gelatin (Fig. 7B). Another striking difference between the gelatin that possesses a random coil structure and the globular proteins such as ovalbumin is their behaviour upon adsorption to surfaces from solutions when the system is heat treated at temperatures above the denaturation point of ovalbumin. As Hagolle, Launay, and Relkin (1998) reported globular protein denaturation and aggregate formation results in a dramatic increase of the amount of protein adsorbed per unit surface in straight contact to gelatin (Braithwaite et al., 1999) where a slight decrease in protein adsorption takes place. Polysaccharides, like agar and carrageenan, produce hard, elastic and brittle gel network structures. Incorporation of MG appears to interrupt the gel structure with the
Fig. 7. Schematic representation of MG particle surfaces- biopolymer network structure interaction in egg white (A), gelatin (B) and agar (C) gels.
MG particles of a relatively small size bringing about an even greater structure breakdown (Table 1). This should be attributed to the complete lack of particle surface–network structure interaction and to accumulation of MG particle aggregates in isolated pockets within the network leading to a decrease in gel strength (Fig. 7C). The smaller the size of the MG particles the more extensive the gel structure interruption since the MG particles are expected to become more effectively distributed within the gel matrix. Although the application of large deformation tests employed in this investigation, combined with observations under the optical microscope, have provided some useful results regarding the topology and the mechanical properties of the biopolymer/MG particles composites, additional systematic studies in these systems using more sophisticated techniques such as dynamic oscillation and stress
ARTICLE IN PRESS C. Mavrakis, V. Kiosseoglou / Food Hydrocolloids 22 (2008) 854–861
relaxation are required in order to elucidate the effect of the filler on the composite properties. Additionally, there is a need to carry out further studies based on theoretical modeling from the area of synthetic polymer research on the effect of MG particles with a relatively large size on the biopolymer gel network structure. A successful protocol has now been established (Kasapis, 1995; Kontogiorgos, Ritzoulis, Biliaderis, & Kasapis, 2006) that allows the prediction of the effect of the continuous matrix reinforcement by spherical filler particles of biomaterial composites. The changes in morphology of the filler brought about by heat treatment may also affect the gel network properties and could, therefore, constitute an additional area of future research. 4. Conclusions Incorporation of MG particles at relatively low concentrations in biopolymer gel matrices resulted in composite gels exhibiting structural features and mechanical properties that differed depending on the type of biopolymer involved in gel structure development. Active filler effects were observed only in the case of egg white protein/ MG particles composites while the effect of incorporation of MG in gels based on gelatin was only a marginal one. This difference in behaviour was attributed to extensive particle surface–egg white protein interaction involving disulfidic bridges as well as hydrophobic bonds. Nonhomogenous dispersions of MG particles which flocculated due to depletion effects, operating between the gum arabicstabilized MG particles and the agar or carrageenan solutions, were possibly responsible for the negative filler effects observed in the case of polysaccharide gel/MG particles composites. References Aguilera, J. M. (1992). Generation of engineered structures in gels. In H. G. Schartzberg, & R. W. Hartel (Eds.), Physical chemistry of foods (pp. 387–421). New York, Basel and Hong Kong: Marcel Dekker, Inc. Aguilera, J. M., & Stanley, W. D. (1999). Food structuring. In J. M. Aguilera, & W. D. Stanley (Eds.), Microstructural principles of food processing and engineering (pp. 228–239). Maryland: Aspen Publishers, Inc. Anton, M., Le Denmat, M., Beaumal, V., & Pilet, P. (2001). Filler effects of oil droplets on the rheology of heat-set emulsion gels prepared with egg yolk and egg yolk fractions. Colloids and Surfaces B: Biointerfaces, 21, 137–147. Braithwaite, J. C. G., Luckham, F. P., & Howe, M. A. (1999). Study of a solvated adsorbed gelatin layer using a modified force microscope. Journal of Colloid and Interface Science, 213, 525–545. Dickinson, E. (2003). Hydrocolloids at interfaces and the influence on the properties of dispersed systems. Food Hydrocolloids, 17, 25–39. Doi, E., & Kitabatake, N. (1997). Structure and functionality of egg proteins. In S. Damodaran, & A. Paraf (Eds.), Food proteins and their
861
applications (pp. 325–340). New York, Basel and Hong Kong: Marcel Dekker, Inc. Frogley, D. M., Ravich, D., & Wagner, H. D. (2003). Mechanical properties of carbon nanoparticle-reinforced elastomers. Composites Science and Technology, 63, 1647–1654. Funami, T., Yada, H., & Nakao, Y. (1998). Thermal and rheological properties of curdlan gel in minced pork gel. Food Hydrocolloids, 12, 55–64. Gauthier, C., Chazeau, L., Prasse, T., & Cavaille, Y. J. (2005). Reinforcement effects of vapour grown carbon nanofibres as fillers in rubbery matrices. Composites Science and Technology, 65, 335–343. Guo, L., Colby, H. R., Lusignan, P. C., & Howe, M. A. (2003). Physical gelation of gelatin studied with rheo-optics. Macromolecules, 36, 10009–10020. Hagolle, N., Launay, B., & Relkin, P. (1998). Impact of structural changes and aggregation on adsorption kinetics of ovalbumin at the water/air interface. Colloids and Surfaces B: Biointerfaces, 10, 191–198. Kamyshny, A., Toledano, O., & Magdassi, S. (1999). Adsorption of hydrophobized IgG and gelatin onto phosphatidyl choline-coated silica. Colloids and Surfaces B: Biointerfaces, 13, 187–194. Kasapis, S. (1995). Phase separation in hydrocolloid gels. In S. E. Hill, & J. R. Mitchell (Eds.), Biopolymer mixtures (pp. 193–224). Nottingham, UK: Nottingham University Press. Kasapis, S. (1999). The elastic moduli of the microcrystalline cellulose— Gelatin blends. Food Hydrocolloids, 13, 543–546. Kehayoglou, H. A., Doxastakis, I. G., & Kiosseoglou, D. V. (1996). Modification of compressive behaviour of Chios mastiche gum. Journal of Texture Studies, 27, 81–92. Kiosseoglou, V. (2005). Protein interactions and structure development in processed food systems containing emulsified oil/fat. In P. A. Riley (Ed.), Food research, safety and policies (pp. 185–197). New York: Nova Science Publishers, Inc. Kontogiorgos, V., Ritzoulis, C., Biliaderis, C. G., & Kasapis, S. (2006). Effect of barley beta-glucan concentration on the microstructural and mechanical behaviour of sodium caseinate gels. Food Hydrocolloids, 20, 749–756. Langley, R. K., & Green, L. M. (1989). Compression strength and fracture properties of model particulate food composites in relation to their microstructure and particle–matrix interaction. Journal of Texture Studies, 20, 191–207. Langley, R. K., Martin, A., & Ogin, L. S. (1994). The effect of filler volume fraction on the fracture toughness of a model food composite. Composites Science and Technology, 50, 259–264. Lowry, O. H., Rosebrough, N. J., Farr, A. L., & Randall, R. J. (1951). Protein measurement with the folin phenol reagent. Journal of Biological Chemistry, 193, 265–275. Morris, V. J. (1998b). Gelation of polysaccharides. In S. E. Hill, D. A. Ledward, & J. R. Mitchell (Eds.), Functional properties of food macromolecules (pp. 151–226). Gaithersburg, MD: Aspen Publishers, Inc. Thomas, M., Kellaway, W. I., & Jones, E. B. (1991). Gelatin film formation at the solid/aqueous interface. International Journal of Pharmaceutics, 73, 185–190. Thoufeek Ahamed, N., Singhal, R. S., Kulkami, P. R., Kale, D. D., & Palb, M. (1996). Studies on Chenopodium quinoa and Amaranthus paniculatas starch as biodegradable fillers in LDPE films. Carbohydrate Polymers, 31, 157–160. van Vliet, T. (1999). Reological classification of foods and instrumental techniques for their study. In A. Rosenthal (Ed.), Food texture, measurement and perception (pp. 65–96). Maryland: Aspen Publishers. Weijers, M., van de Velde, F., Stijnman, A., van de Pijpekamp, A., & Visschers, W. R. (2006). Structure and rheological properties of acidinduced egg white protein gels. Food Hydrocolloids, 20, 146–159.
FOOD
HYDROCOLLOIDS Food Hydrocolloids 22 (2008) 854–861 www.elsevier.com/locate/foodhyd
The structural characteristics and mechanical properties of biopolymer/mastic gum microsized particles composites Christos Mavrakis, Vassilis Kiosseoglou Laboratory of Food Chemistry and Technology, School of Chemistry, Aristotle University of Thessaloniki, 54124 Thessaloniki, Greece Received 20 December 2006; accepted 31 March 2007
Abstract Biopolymer/mastic gum (MG) particles composites were prepared by incorporating microsized MG particles in gel matrices based on gelatin, egg white or polysaccharides and their mechanical properties as influenced by MG content and particle size were investigated. Observation studies under the microscope were conducted to determine the structural features of the MG composites. The findings were combined with data on protein adsorption to MG particle surface to discuss the mechanical properties of the gels, which differed substantially depending on the biopolymer involved in gel network structure development. A mechanism is put forward to explain the role of MG particles as filler materials of biopolymer gel matrices. r 2007 Elsevier Ltd. All rights reserved. Keywords: Composite gel; Filler effects; Protein; Polysaccharide; Mechanical properties
1. Introduction The structure and rheological behaviour of a great number of traditional semi-solid foods depend to an appreciable extent on the presence and interactions within the food system between biopolymeric molecules, namely proteins or polysaccharides. Additionally, due to their vast diversity, both in structure and functionality, biopolymers are extremely useful as food additives to scientists involved in product development in designing new foods that meet the consumer demand for cheaper and healthier products which at the same time exhibit novel structural and textural characteristics (Aguilera & Stanley, 1999; Kiosseoglou, 2005). When the biopolymeric molecules interact, usually through non-covalent bonds, a network structure develops that entraps and immobilizes the solvent as well as all the other low-molecular weight constituents of the system. The gel structure differs depending on the type of polymer involved. Protein molecules normally tend to interact, Corresponding author. Tel.: +302310997834; fax: +302310997779.
E-mail address: [email protected] (V. Kiosseoglou). 0268-005X/$ - see front matter r 2007 Elsevier Ltd. All rights reserved. doi:10.1016/j.foodhyd.2007.03.011
following heat treatment and unfolding, at temperatures above their denaturation point. Polysaccharides, on the other hand, as a rule produce gel network structures when a solution is quenched either in the absence or presence of gelation aids (e.g. Ca2+, sugar, etc.) (Morris, 1998). Gelatin, a degradation product of collagen, in spite of being a protein behaves like a polysaccharide and produces gel networks on cooling gelatin solutions of a relatively low concentration. Molecular interactions in gelatin networks rely on the formation of junction zones where molecular segments from different gelatin molecules interact and form an extensive triple helical structure (Guo, Colby, Lusignan, & Howe, 2003). Many semi-solid food systems are composites having a gel network structure where a particulate material, the filler, is embedded. The rheological properties of the filled network structure as well as the textural characteristics of the food could be significantly affected by the presence of particles such as meat fibers, starch granules or emulsified oil droplets. Parameters such as the filler size and shape and the strength of filler surface–gel matrix interaction should be taken into account when considering the reinforcing effect of a filler particle on a biopolymer
ARTICLE IN PRESS C. Mavrakis, V. Kiosseoglou / Food Hydrocolloids 22 (2008) 854–861
network (Aguilera, 1992; Anton, Le Denmat, Beaumal, & Pilet, 2001; Thoufeek Ahamed, Singhal, Kulkami, Kale, & Palb, 1996). Reinforcing fillers of inorganic nature, including hydroxyapatite clay, glass and carbon particles or nanotubes are widely used to improve the mechanical properties of products such as fiberglass, rubber or natural and synthetic polymer matrices (Frogley, Ravich, & Wagner, 2003; Gauthier, Chazeau, Prasse, & Cavaille, 2005; Langley, Martin, & Ogin, 1994).When suitably incorporated, these fillers may bring about a substantial improvement of the mechanical properties of the gel structure due to effective load transfer from the matrix to the much harder filler particle through the particle surface which should interact with the matrix. The choice, however, for fillers, suitable for use in food systems, is very limited. One such filler of organic nature is microcrystalline cellulose which as reported by Kasapis (1999) may reinforce the structure of gelatin gels. Another material that could also be used in this respect is mastic gum (MG), which is the dried exudate obtained from the tree Pistacia lentiscus L., (Anacardiaceae), a native of the East Mediterranean and also of the Middle East countries. Chios MG is obtained in the form of drops from bark incisions on the Chios variety of the tree cultivated in the south of the island of Chios (Greece). The drops are then allowed to dry out in the air. The gum, a rather hard and brittle material at room temperature, has been finding use since ancient times as an antiseptic in medicine, as food antioxidant and as chewing gum base. The annual production yield of MG ranges between 100 and 150 tons and is used mainly as chewing gum base although a plasticizer is usually added to reduce its undesirable hardness and adhesiveness (Kehayoglou, Doxastakis, & Kiosseoglou, 1996). Additionally, the gum is exploited as flavouring additive in an increasingly expanding number of traditional foods and drinks (ice-cream, confectionery, baked products, liqueurs, etc.). As experience shows, MG in certain semi-solid products (ice-cream or baked products) behaves not only as flavouring agent but also as texture modifier, acting probably as filler of the protein-based gel matrixes, and could, therefore, be exploited as a natural food additive in order to develop novel structures that appeal to the consumer. The aim of the present work was to investigate the role of Chios MG as filler material of food biopolymer gel network structures, which determine to a large extent the mechanical and textural characteristics of most of semisolid food products. As the gum is a non-polar material its mode of action as filler will depend on the adsorption behaviour of the biopolymeric molecules to the filler particle surface and, therefore, on the molecular structure of the biopolymer involved. The ultimate objective of the study was to establish whether MG, apart from its outstanding flavouring effect in certain food products, may also affect their mechanical properties when present at relative low concentrations.
855
2. Materials and methods 2.1. Materials Chios MG granules were bought from the local market. The granules were ground to powder for 5 min in a mill fitted with knives and the resulting powder was fractionated into samples consisting of particles of various sizes with the aid of a set of sieves of mesh sizes 125, 90, 75 and 45 mm on a mechanical vibrator operated for 15 min. Agar, carrageenan (mainly k-carrageenan), gelatin (type A, from porcine skin, 93% w/w protein content), gum arabic and spray-dried egg white (85% w/w protein content) were obtained from Sigma Chemical Co. (St. Louis, MO). Tween 40 was a product of Fluka AG (Buchs, Switzerland). All the chemicals used in the experiments were of analytical grade. 2.2. Particle size determination The MG powder fractions were dispersed in a 1% (w/v) Tween solution in distilled water under continuous stirring for at least 1 h to obtain 3% (w/v) MG dispersions. The particle size distribution was determined by applying a laser scattering technique with the aid of the Malvern Mastersizer 2000 (Malvern Instruments Ltd., Malvern, Worcestershire, UK). The mean particle size, d3,2 and d4,3, is referred to as apparent mean size since most of the particles were not spherical. 2.3. Gel preparation Gel samples based on gelatin were prepared by dissolving 3 g of protein in 100 ml phosphate buffer (1/10 M Na2HPO4 and KH2PO4) of pH 7 with the aid of a magnetic stirrer and heating at 80 1C for 2 h. The MG powder was then incorporated under continuous stirring for 2 h and the resulting dispersion was transferred to cylindrical cells of 1 cm diameter made from aluminium foil and reinforced with plastic tape. The cells were placed at 3 1C for 24 h, then cut open, and cylindrical samples of 1 cm height were obtained. To prepare the agar and carrageenan gels, 2 g of gum arabic were initially dissolved in 100 ml buffer followed by the dispersion of the MG powder. The agar or carrageenan (3 g) was then dissolved at 80 1C under continuous stirring for 1 h, the dispersions were placed in cylindrical cells and cooled at 3 1C for 24 h as describe above. The gel based on egg white was prepared by first dissolving the white in the buffer to obtain a 10% solution in protein. Following dispersion of the MG powder under continuous stirring for 2 h, cylindrical cells were filled with the MG dispersion and heated at 90 1C for 3 min. The cells were cut open, following storage for 24 h at room temperature, and cylindrical samples 1 cm in height were obtained. 2.4. Compression test The cylindrical gel samples were subjected to a 80% compression at a speed of 0.5 mm/s using the Stable Micro
ARTICLE IN PRESS 856
C. Mavrakis, V. Kiosseoglou / Food Hydrocolloids 22 (2008) 854–861
System TA-XT2i Texture Analyzer (Surrey, England) equipped with a 2.5 cm diameter cylinder. Bulging of sample middle part was prevented by applying vegetable oil to the plates in contact with the sample surfaces. The force–time curves obtained from the analyzer were converted into compression stress-Hency strain ones (van Vliet, 1999). Liquid loss under pressure was determined (Funami, Yada, & Nakao, 1998) from the liquid expressed from a 5 mm thick gel sample compressed (20%) for 2 min between layers of filter paper with the aid of Texture Analyser. 2.5. Optical microscopy Optical microscopy images of MG dispersions in biopolymer solutions before and after heating were obtained by an Axiolab A reflected light microscope (Zeiss, Berlin, Germany) equipped with a Power Shot G2 photographic camera (Canon, Tokyo, Japan). 2.6. Amount of protein adsorbed to MG surface To determine the amount of protein adsorbed per unit particle surface, solutions of gelatin and egg white were prepared and MG powder of a mean particle size, d3,2 45–75 mm was dispersed with the aid of a magnetic stirrer for 30 min. Following centrifugation at 10,000g for 30 min, the protein content of the supernatant was determined according to Lowry, Rosebrough, Farr, and Randall (1951). The amount of protein adsorbed per unit surface was calculated using Gs ðmg=m2 Þ ¼ GT =ST , where GT is the amount of protein adsorbed in the system and ST is the total particle surface derived by applying the equation: S T ¼ 6V =d 3;2 , where V is the MG volume obtained assuming that the density of MG is 1.06 g/cm3.
solution, are presented in Fig. 1. The particle diameters are reported as apparent diameters and were calculated from particle size dispersion data provided by the Malvern sizer. The mean diameter values corresponded very satisfactorily to those expected according to the mesh size of the sieves used for particle fractionation. The dispersions of MG particles in gelatin and agar gel systems, along with their respective dispersions in cold biopolymer solutions, are illustrated in Fig. 2, while Fig. 3 exhibits MG dispersions in egg white solutions and gels. The marked differences observed between the composites based on the three biopolymeric materials apparently reflected differences in the mechanism behind gel network formation and probably in the extent and strength of interaction between the biopolymer matrix and the MG particle surface. MG particles appeared to become satisfactorily dispersed in both gelatin and gum arabic solutions (Fig. 2A and C, respectively). When, however, agar was introduced in the latter, aided by the application of heat, the particles melted and turned into spheres while at the same time they flocculated and formed large particle aggregates (Fig. 2D). Particle flocculation was not observed in the case of gelatin MG dispersions except that the particles upon heating also acquired a spherical shape remaining, however, fully dispersed in the gelatin matrix (Fig. 2B). This apparently is an indication that the MG particles were better protected against flocculation during heating by the gelatin molecules than by gum arabic/agar. As shown in Fig. 4, gelatin adsorbed to the MG particle surface and the surface protein load reached a plateau at higher protein concentrations corresponding to surface coverage of around 3 mg/m2. Gelatin in solution possesses a randomly coiled structure. Upon adsorption to solid surfaces a number of protein chain segments, described as ‘‘trains’’, attach to the surface leaving free loops and molecular ‘‘tails’’ extending into the bulk solution (Braithwaite, Luckham, & Howe, 1999). Gelatin attachment to hydrophilic surfaces is mainly governed by electrostatic interactions while adsorption on non-polar surfaces is mainly the result of hydrophobic binding that
2.7. Statistical analysis
3. Results and discussion 3.1. Structure of biopolymer/MG composites
160 140 d3.2 and d4.3 (μm)
All experiments were repeated at least three times and the data were analyzed using the one-way ANOVA program. The level of confidence was 95%. Significant differences between means were identified by the LSD procedure using the statistical software package SPSS v. 8.0 for Windows (SPSS Inc., Chicago, IL).
120 100 80 60 40 20 0
The mean MG particle diameters, d3,2 and d4,3, of the powder fractions, determined by applying the laser light scattering technique to MG dispersions in 1% Tween
90-125μm
75-90μm
45-75μm
<45μm
Fig. 1. Apparent mean particle size [d3,2 ( ) and d4,3 (&)] of the MG particle fractions determined by laser light scattering.
ARTICLE IN PRESS C. Mavrakis, V. Kiosseoglou / Food Hydrocolloids 22 (2008) 854–861
857
Fig. 2. Optical microphotographs of MG particle (mesh size o45 mm) dispersions in 3% gelatin solution (A) and gel (B) or in 2% gum arabic solution (C) and in 3% agar gel (D).
Fig. 3. Optical microphotographs of 10% egg white solution (A), of 6% (w/v) MG particles dispersion in the egg white solution (B and C for mesh size o45 and 90–125 mm, respectively) and of a 10% egg white gel containing 6% MG particles of mesh size 90–125 mm (D).
overcomes the electrostatic repulsion when both the particle surface and the protein molecule carry charges of the same sign (Kamyshny, Toledano, & Magdassi, 1999;
Thomas, Kellaway, & Jones, 1991). The presence of this adsorbed protein layer at the particle surfaces may then protect the particles against flocculation probably
ARTICLE IN PRESS C. Mavrakis, V. Kiosseoglou / Food Hydrocolloids 22 (2008) 854–861
858
6
Γs (mg/m2)
5 4 3 2 1 0 0
0.1
0.2 0.3 0.4 concentration (%w/v)
0.5
0.6
and the gel matrix. As preliminary experiments have shown, gum arabic also adsorbed to the particle surface producing a well-dispersed and stable system. Once, however, agar was introduced by heating, particle flocculation took place suggesting that depletion phenomena (Dickinson, 2003) might have operated due to the exclusion of agar molecules from a area near the particle surface as they do not adsorb to the surfaces, either directly or indirectly, through interaction with the adsorbed gum arabic molecules. Egg white is a mixture of soluble proteins of globular nature with the exception of ovomucin, which tends to
Fig. 4. Gelatin (m) and egg white protein (’) surface load, Gs, versus protein concentration for MG particle dispersions (mesh size 45–75 mm) in protein solutions.
relative elasticity modulus [-]
16 14 12 stress [kN.m-2]
3.5
10 8
3 2.5 2 1.5 1 0.5 0
6
control
+ 3% MG (45-75μm)
+ 6% MG (45-75μm)
+ 12% MG (45-75μm)
control
+ 3% MG (45-75μm)
+ 6% MG (45-75μm)
+ 12% MG (45-75μm)
4 2
1.8 1.6 0
0.2
0.4
0.6
0.8 1 strain [-]
1.2
1.4
1.6
1.8
140
stress [kN.m-2]
120
relative stress [-]
0
1.4 1.2 1 0.8 0.6 0.4
100
0.2
80
0
60 40
1.2
20
0
0.2
0.4
0.6
0.8 1 strain [-]
1.2
1.4
1.6
1.8
Fig. 5. Stress versus strain curves, (A) for egg white and gelatin ((m) and (’), respectively) and (B) for agar and carrageenan ((m) and (’), respectively) gels (filled symbols) and for the same gels containing 6% MG particles of mesh size 45–75 mm (open symbols).
relative strain [-]
1
0
0.8 0.6 0.4 0.2 0 control
through steric repulsive interactions, originating from overlapping ‘‘tail’’ and ‘‘loop’’ segments of gelatin molecules adsorbed on neighbouring particles that approach each other to very short distances, resulting in a system with MG particles evenly dispersed in the protein solution
+ 3% MG (45-75μm)
+ 6% MG (45-75μm)
+12% MG (45-75μm)
Fig. 6. Effect of MG concentration on the relative elasticity modulus (A) and relative stress (B) and strain (C) at fracture values for egg white (&), gelatine (B), agar (J) and carrageenan (n) gels containing MG particles of mesh size 45–75 mm.
ARTICLE IN PRESS C. Mavrakis, V. Kiosseoglou / Food Hydrocolloids 22 (2008) 854–861
form a weak gel-like structure responsible for the mucous appearance of the white. When the dehydrated egg white was dispersed in water a number of threadlike aggregates were spotted against a homogeneous background under the microscope (Fig. 3A). These aggregates may constitute dispersed ovomucin network structures as well as ovalbumin–conalbumin aggregates formed during egg white dehydration. In fact, as Weijers, van de Velde, Stijnman, van de Pijpekamp, and Visschers (2006) point out, the latter are responsible for the opaque appearance of gels prepared by heating the white, which is an indication of a non-homogeneous gel network matrix made up of randomly aggregating, denatured protein molecules. Incorporation of MG particles in egg white solution resulted in phase separation with the particles aggregating and forming a thread-like network spanning the total volume of the system. The thickness of the treads appears to depend on the particle size as the large-sized MG particles produced a less dense and more coarse flocculated network (Fig. 3B and C). Following heating and gel matrix formation, the initial structure did not change except that the MG particles became more spherical (Fig. 3D). 3.2. Mechanical properties and particle surface–matrix interaction When the gel samples were subjected to compression, their structure yielded at a strain value that depended on the type of biopolymer involved in gel formation. The polysaccharide gels fractured at a relatively lower strain value than the protein gels (Fig. 5). When MG was incorporated the gel resistance to compression was modified, depending on the biopolymer involved in net-
859
work structure development. Thus, MG particles at a concentration level of 6%, appeared to have a reinforcing effect on the structure of egg white gels since the samples yielded at a higher stress, than those of the control, while the slope of the stress–strain curve became steeper indicating an increase in gel elasticity. The reinforcing effect of MG on gelatin-based gels was less obvious as the value of the stress at fracture was not affected while the increase in the slope of the stress–strain curve was only marginal (Fig. 5A). In the case of polysaccharide gels, on the other hand, incorporation of MG resulted in a marked reduction of the gel resistance to compression (Fig. 5B). The relative change with MG content of stress/strain at fracture and elasticity modulus, determined from the initial slope of the stress–strain curve at small deformation, is exhibited in Fig. 6. In general, the strain at fracture appears to decrease with MG content for all the gel samples, the decrease being more spectacular in the case of agar. The relative stress at fracture increased with the MG content only in the case of the egg white gels. An even more spectacular increase was observed for Young’s elasticity modulus indicating that the particles functioned as active fillers and reinforced the gel network structure. Finally, the mechanical strength of the polysaccharide gels was inversely correlated with MG content and a negative filler effect was the outcome of MG incorporation, indicating that the network structure was interrupted by the presence of MG particles. As was discussed in the previous section, heating of egg white above the denaturation temperature of its constituent proteins, led to partial protein unfolding followed by interaction and gel network formation. The opaque appearance of the resulting gel indicates that non-ordered interaction between the unfolded protein molecules took
Table 1 Strain and stress at fracture and elasticity modulus values of egg white, gelatin, carrageenan and agar gels containing 6% MG particles of varying mean size Strain []
Stress (kN m2)
Elasticity modulus (kN m2)
Egg white 10%w/v +6% MG (90–125 mm) +6% MG (75–90 mm) +6% MG (45–75 mm)
0.8570.007a 0.7070.011b 0.7170.026b 0.7670.010c
7.770.15a 8.270.07b 8.570.06c 11.870.10d
Gelatin 3%w/v +6% MG (90–125 mm) +6% MG (75–90 mm) +6% MG (45–75 mm)
1.2570.012a 0.9670.005b 0.9870.007c 1.0970.005d
12.570.50a 9.070.34b 9.370.27b 12.770.13a
Carrageenan 3%w/v +6% MG (90–125 mm) +6% MG (75–90 mm) +6% MG (45–75 mm)
0.5170.001a 0.4570.002b 0.4270.019c 0.4170.001c
121.870.32a 93.870.72b 82.170.74c 81.270.98c
253.273.54a 222.972.50b 206.472.90c 206.072.64c
Agar 3%w/v +6% MG (90–125 mm) +6% MG (75–90 mm) +6% MG (45–75 mm)
0.2170.001a 0.2070.005b 0.1870.003c 0.1270.004d
40.670.34a 30.971.50b 25.970.41c 5.670.05d
187.072.72a 158.971.47b 145.472.47c 49.170.25d
a–d
Different letters indicate significant difference *po0.05.
8.070.51a 10.770.11b 12.370.25c 14.370.31d 4.670.18a 5.270.10a,b 5.570.20b 5.670.52b
ARTICLE IN PRESS 860
C. Mavrakis, V. Kiosseoglou / Food Hydrocolloids 22 (2008) 854–861
place, producing a non-homogeneous gel structure with regions of high polymer concentration and ones nearly devoid of polymer (Doi & Kitabatake, 1997). MG particles tended to accumulate at the polymer-depleted regions where they assembled and produced a thread-like structure embedded in the primary egg white gel matrix (Fig. 3). The formation of this secondary network, in turn, reinforces the gel structure as it can bear the stress transferred from the primary network, through extensive interaction between the gel network and the MG particle surfaces, which is a prerequisite for embedded particles to act as active fillers of gel structure (Aguilera, 1992). When the mean particle size was decreased, the MG particle thread-like structure became denser with thinner particle chains, leading to a higher mechanical strength (Table 1). Gelatin produces homogeneous, translucent and elastic gels. Due to adsorption of gelatin molecules to the particle surface, the MG particles became fully dispersed both in the gelatin solution and the gel network structure. Their incorporation, however, did not appear to have any effect on the mechanical properties of gelatin gel. A similar effect was also reported by Langley and Green (1989) for gelatin gels incorporating glass beads at relatively low concentration levels and only above 20% the glass spheres acted as active filler of the matrix. In contrast to egg white proteins which when adsorb to the particle surfaces unfold only to a limited extent, due to the presence in their molecules of disulfide bridges, the highly flexible gelatin molecules are expected to spread at the surface. As a result of this, interactions with the network matrix should only be confined to those molecular segments that constitute the ‘‘tails’’ of the adsorbed molecules and may become involved in triple helix formation with the matrix proteins (Fig. 7A). The lack of extensive interaction between the particle surfaces and the network should also explain the relatively marginal influence of MG particle size on the mechanical properties of the gelatin gels (Table 1). In the case of egg white, on the other hand, extensive hydrophobic and disulfide bond formation may take place between the adsorbed protein molecules and those involved in gel structure development, leading to filler reinforcement effects by the incorporated MG particles at relatively lower concentrations compared to gelatin (Fig. 7B). Another striking difference between the gelatin that possesses a random coil structure and the globular proteins such as ovalbumin is their behaviour upon adsorption to surfaces from solutions when the system is heat treated at temperatures above the denaturation point of ovalbumin. As Hagolle, Launay, and Relkin (1998) reported globular protein denaturation and aggregate formation results in a dramatic increase of the amount of protein adsorbed per unit surface in straight contact to gelatin (Braithwaite et al., 1999) where a slight decrease in protein adsorption takes place. Polysaccharides, like agar and carrageenan, produce hard, elastic and brittle gel network structures. Incorporation of MG appears to interrupt the gel structure with the
Fig. 7. Schematic representation of MG particle surfaces- biopolymer network structure interaction in egg white (A), gelatin (B) and agar (C) gels.
MG particles of a relatively small size bringing about an even greater structure breakdown (Table 1). This should be attributed to the complete lack of particle surface–network structure interaction and to accumulation of MG particle aggregates in isolated pockets within the network leading to a decrease in gel strength (Fig. 7C). The smaller the size of the MG particles the more extensive the gel structure interruption since the MG particles are expected to become more effectively distributed within the gel matrix. Although the application of large deformation tests employed in this investigation, combined with observations under the optical microscope, have provided some useful results regarding the topology and the mechanical properties of the biopolymer/MG particles composites, additional systematic studies in these systems using more sophisticated techniques such as dynamic oscillation and stress
ARTICLE IN PRESS C. Mavrakis, V. Kiosseoglou / Food Hydrocolloids 22 (2008) 854–861
relaxation are required in order to elucidate the effect of the filler on the composite properties. Additionally, there is a need to carry out further studies based on theoretical modeling from the area of synthetic polymer research on the effect of MG particles with a relatively large size on the biopolymer gel network structure. A successful protocol has now been established (Kasapis, 1995; Kontogiorgos, Ritzoulis, Biliaderis, & Kasapis, 2006) that allows the prediction of the effect of the continuous matrix reinforcement by spherical filler particles of biomaterial composites. The changes in morphology of the filler brought about by heat treatment may also affect the gel network properties and could, therefore, constitute an additional area of future research. 4. Conclusions Incorporation of MG particles at relatively low concentrations in biopolymer gel matrices resulted in composite gels exhibiting structural features and mechanical properties that differed depending on the type of biopolymer involved in gel structure development. Active filler effects were observed only in the case of egg white protein/ MG particles composites while the effect of incorporation of MG in gels based on gelatin was only a marginal one. This difference in behaviour was attributed to extensive particle surface–egg white protein interaction involving disulfidic bridges as well as hydrophobic bonds. Nonhomogenous dispersions of MG particles which flocculated due to depletion effects, operating between the gum arabicstabilized MG particles and the agar or carrageenan solutions, were possibly responsible for the negative filler effects observed in the case of polysaccharide gel/MG particles composites. References Aguilera, J. M. (1992). Generation of engineered structures in gels. In H. G. Schartzberg, & R. W. Hartel (Eds.), Physical chemistry of foods (pp. 387–421). New York, Basel and Hong Kong: Marcel Dekker, Inc. Aguilera, J. M., & Stanley, W. D. (1999). Food structuring. In J. M. Aguilera, & W. D. Stanley (Eds.), Microstructural principles of food processing and engineering (pp. 228–239). Maryland: Aspen Publishers, Inc. Anton, M., Le Denmat, M., Beaumal, V., & Pilet, P. (2001). Filler effects of oil droplets on the rheology of heat-set emulsion gels prepared with egg yolk and egg yolk fractions. Colloids and Surfaces B: Biointerfaces, 21, 137–147. Braithwaite, J. C. G., Luckham, F. P., & Howe, M. A. (1999). Study of a solvated adsorbed gelatin layer using a modified force microscope. Journal of Colloid and Interface Science, 213, 525–545. Dickinson, E. (2003). Hydrocolloids at interfaces and the influence on the properties of dispersed systems. Food Hydrocolloids, 17, 25–39. Doi, E., & Kitabatake, N. (1997). Structure and functionality of egg proteins. In S. Damodaran, & A. Paraf (Eds.), Food proteins and their
861
applications (pp. 325–340). New York, Basel and Hong Kong: Marcel Dekker, Inc. Frogley, D. M., Ravich, D., & Wagner, H. D. (2003). Mechanical properties of carbon nanoparticle-reinforced elastomers. Composites Science and Technology, 63, 1647–1654. Funami, T., Yada, H., & Nakao, Y. (1998). Thermal and rheological properties of curdlan gel in minced pork gel. Food Hydrocolloids, 12, 55–64. Gauthier, C., Chazeau, L., Prasse, T., & Cavaille, Y. J. (2005). Reinforcement effects of vapour grown carbon nanofibres as fillers in rubbery matrices. Composites Science and Technology, 65, 335–343. Guo, L., Colby, H. R., Lusignan, P. C., & Howe, M. A. (2003). Physical gelation of gelatin studied with rheo-optics. Macromolecules, 36, 10009–10020. Hagolle, N., Launay, B., & Relkin, P. (1998). Impact of structural changes and aggregation on adsorption kinetics of ovalbumin at the water/air interface. Colloids and Surfaces B: Biointerfaces, 10, 191–198. Kamyshny, A., Toledano, O., & Magdassi, S. (1999). Adsorption of hydrophobized IgG and gelatin onto phosphatidyl choline-coated silica. Colloids and Surfaces B: Biointerfaces, 13, 187–194. Kasapis, S. (1995). Phase separation in hydrocolloid gels. In S. E. Hill, & J. R. Mitchell (Eds.), Biopolymer mixtures (pp. 193–224). Nottingham, UK: Nottingham University Press. Kasapis, S. (1999). The elastic moduli of the microcrystalline cellulose— Gelatin blends. Food Hydrocolloids, 13, 543–546. Kehayoglou, H. A., Doxastakis, I. G., & Kiosseoglou, D. V. (1996). Modification of compressive behaviour of Chios mastiche gum. Journal of Texture Studies, 27, 81–92. Kiosseoglou, V. (2005). Protein interactions and structure development in processed food systems containing emulsified oil/fat. In P. A. Riley (Ed.), Food research, safety and policies (pp. 185–197). New York: Nova Science Publishers, Inc. Kontogiorgos, V., Ritzoulis, C., Biliaderis, C. G., & Kasapis, S. (2006). Effect of barley beta-glucan concentration on the microstructural and mechanical behaviour of sodium caseinate gels. Food Hydrocolloids, 20, 749–756. Langley, R. K., & Green, L. M. (1989). Compression strength and fracture properties of model particulate food composites in relation to their microstructure and particle–matrix interaction. Journal of Texture Studies, 20, 191–207. Langley, R. K., Martin, A., & Ogin, L. S. (1994). The effect of filler volume fraction on the fracture toughness of a model food composite. Composites Science and Technology, 50, 259–264. Lowry, O. H., Rosebrough, N. J., Farr, A. L., & Randall, R. J. (1951). Protein measurement with the folin phenol reagent. Journal of Biological Chemistry, 193, 265–275. Morris, V. J. (1998b). Gelation of polysaccharides. In S. E. Hill, D. A. Ledward, & J. R. Mitchell (Eds.), Functional properties of food macromolecules (pp. 151–226). Gaithersburg, MD: Aspen Publishers, Inc. Thomas, M., Kellaway, W. I., & Jones, E. B. (1991). Gelatin film formation at the solid/aqueous interface. International Journal of Pharmaceutics, 73, 185–190. Thoufeek Ahamed, N., Singhal, R. S., Kulkami, P. R., Kale, D. D., & Palb, M. (1996). Studies on Chenopodium quinoa and Amaranthus paniculatas starch as biodegradable fillers in LDPE films. Carbohydrate Polymers, 31, 157–160. van Vliet, T. (1999). Reological classification of foods and instrumental techniques for their study. In A. Rosenthal (Ed.), Food texture, measurement and perception (pp. 65–96). Maryland: Aspen Publishers. Weijers, M., van de Velde, F., Stijnman, A., van de Pijpekamp, A., & Visschers, W. R. (2006). Structure and rheological properties of acidinduced egg white protein gels. Food Hydrocolloids, 20, 146–159.