Effect of the Type of Emulsifying Salt

on Microstructure and Rheological Properties of

“Requeijão Cremoso” Processed Cheese Spreads
Clarissa R. da Cunha, Maria Regina Alcântara, and Walkiria H. Viotto

Abstract: The role of different types of emulsifying salts—sodium citrate (TSC), sodium hexametaphosphate (SHMP),
sodium tripolyphosphate (STPP) and tetrasodium pyrophosphate (TSPP)—on microstructure and rheology of “requeijão
cremoso” processed cheese was determined. The cheeses manufactured with TSC, TSPP, and STPP behaved like con-
centrated solutions, while the cheese manufactured with SHMP exhibited weak gel behavior and the lowest values for
E: Food Engineering &

the phase angle (G”/G’). This means that SHMP cheese had the protein network with the largest amount of molecular
Physical Properties

interactions, which can be explained by its highest degree of fat emulsification. Rotational viscometry indicated that
all the spreadable cheeses behaved like pseudoplastic fluids. The cheeses made with SHMP and TSPP presented low
values for the flow behavior index, meaning that viscosity was more dependent on shear rate. Regarding the consistency
index, TSPP cheese showed the highest value, which could be attributed to the combined effect of its high pH and
homogeneous fat particle size distribution.
Keywords: fat emulsification, microstructure, peptization, rheology, spreadable processed cheese

Introduction casein to act as an emulsifier at the water–oil interface (Zehren

“Requeijão cremoso” is a traditional Brazilian spreadable processed and Nusbaum 1992). The transformation of the casein network
cheese which presents a fluid consistency and a high fat content into a stable colloidal dispersion is accompanied by an increase
(minimum 55% on a dry weight basis), and can be essentially in viscosity of the molten mass, and this phenomenon is known
classified as an oil-in-water emulsion. Traditionally the product is as cremification (Zehren and Nusbaum 1992; Guinee and others
made by the fusion of the fresh curd obtained by direct acidifica- 2004).
tion of heated milk (after removal of the whey and washing), but Currently, citrates, monophosphates, and polyphosphates are
some manufacturers also use curds obtained by lactic fermentation widely used in the manufacture of processed cheese, and depend-
and/or rennet coagulation. The fusion is achieved through the ad- ing on the desired action they can be used alone or combined
dition of emulsifying salts (ESs), usually followed by the addition in different proportions (Berger and others 1989). The degree
of cream and/or butter. The resulting mass is then cooked at 85 of hydration of the para-casein and the size distribution of the
to 95 ◦ C/2 to 7 min, and hot filled into plastic or glass jars. emulsified fat globules has considerable influence on the struc-
“Requeijão cremoso” cheese is usually spread over toasts or cream ture and rheology of the processed cheese (Guinee and others
crackers using a knife, and also used as a food ingredient for pizza 2004). Understanding the effect of each salt on cheese rheolog-
and as a filling for stuffed foods (pasta, pies, nuggets, sandwiches, ical characteristics, such as the elastic and the viscous moduli, is
etc.). The product has increasing importance in the national mar- important to explain the structure of the final product. Moreover,
ket: from 2004 to 2007, its production increased 52%, achieving the knowledge of the flow behavior of the materials allows the
47000000 kg and placing it as the second most consumed cheese correct selection of equipments (pumps and filling machines) and
in Brazil (ABIQ 2008). optimization of their operational conditions.
In processed cheeses, the type of ES influences the casein pep- There are few studies on the role of ESs on the rheological
tization and the degree of fat emulsification, parameters that affect characteristics of processed cheeses. Most of them include differ-
the quality of the final product. Peptization, defined as the sepa- ent raw materials, processing conditions and cheese composition,
ration of the large hydrophobic aggregates of casein into smaller therefore leading to conflicting results (Dimitreli and others 2005;
units during the heating and stirring of the mass, increases casein Dimitreli and Thomareis 2009; Sádlı́kova and others 2010). The
hydration, and exposes its polar and nonpolar groups, allowing use of anhydrous ingredients makes it possible to use always the
same source of protein and fat, avoiding variation in cheese com-
position and allowing cheese to be treated as a model system where
the variables are limited and well known. This approach might be
MS 20120049 Submitted 1/10/2012, Accepted 5/9/2012. Author da Cunha and useful to provide a more general understanding about the effect of
Viotto are with the Dept. of Food Technology, Faculty of Food Engineering, State Univ. individual ESs on the rheology of processed cheeses.
of Campinas – UNICAMP, P.O. Box 6121, Campinas-SP 13083–862, Brazil. The objective of the present study was to evaluate the role of dif-
Author Alcântara is with the Inst. of Chemistry, Univ. of São Paulo, P.O. Box 26077,
São Paulo, SP 05599–970, Brazil. Direct inquiries to author da Cunha (E-mail:
ferent types of ES (sodium citrate [TSC], sodium hexametaphos-
[email protected]). phate [SHMP], sodium tripolyphosphate [STPP], and tetrasodium
pyrophosphate [TSPP]) on the microstructure and rheological

C 2012 Institute of Food Technologists

 R

E176 Journal of Food Science r Vol. 77, Nr. 8, 2012 doi: 10.1111/j.1750-3841.2012.02797.x
Further reproduction without permission is prohibited
Effect of emulsifying salt on rheology . . .

characteristics of “requeijão cremoso” cheese manufactured us- Microstructure

ing anhydrous ingredients. Microstructure of processed cheeses was assessed using scan-
ning electron microscopy. The samples were prepared using the
method described by Marchesseau and others (1997), adapted to
Material and Methods the conditions of “requeijão cremoso” cheese, as described by
Gallina (2005). Initially the samples were placed in disposable sy-
Processing of “requeijão cremoso” made from anhydrous ringes and the plunger carefully pushed down to form “threads”
ingredients of processed cheese. The “threads” were fixed with 2.5% glu-
The processed cheeses used in this study were the same as those taraldehyde in a 0.1 M phosphate buffer (pH 7.2) for 4 h at
described in an earlier study (Cunha and Viotto 2010), where room temperature, then cut with a knife and fixed for more 9 h,
the ingredients and manufacture conditions have been detailed, as washed in 0.1 M phosphate buffer and postfixed with 1% OsO4 in
well as the methods used to evaluate cheese composition and the 0.1 M phosphate buffer for 8 h. After further washing with 0.1 M
degree of casein dissociation. The processed cheese manufacture phosphate buffer, the samples were dehydrated in a graded series
flow diagram is shown in Figure 1. of ethanol (50%, 70%, 90%, 95%, and 100%), followed by 100%

E: Food Engineering &

Physical Properties

Figure 1–Schematic illustration showing the manufacturing stages of “requeijão cremoso” processed cheeses made with different emulsifying salts:
TSC (trisodium citrate); SHMP (sodium hexametaphosphate); TSPP (tetrasodium pyrophosphate); and STPP (sodium tripolyphosphate). The amount of
each ingredient was calculated and adjusted to result in a final product with 63% moisture, 10.5% protein, 1% NaCl, and 0.83% emulsifying salt. The
amount of lactic acid added was that required to reduce pH of the mixture caseinate + water to 5.3. Font: Cunha and Viotto (2010). 209 x 279 mm
(300 × 300 DPI).

Vol. 77, Nr. 8, 2012 r Journal of Food Science E177

 Effect of emulsifying salt on rheology . . .

acetone. The samples were dried in a CPD 030 critical point dryer Results and Discussion
(Bal-tec AG, Balzers, Liechtenstein), fractured at room tempera-
ture, mounted on aluminium stubs with silver glue, coated with Characterization of the “requeijão cremoso” cheese
gold (240 s/40 mA) using the SCD 050 sputterer (Bal-tec AG, The chemical compositions of “requeijão cremoso” cheeses
Balzers, Liechtenstein) and observed under the JEOL JSM- have been previously reported by Cunha and Viotto (2010).
5800LV scanning electron microscope (JEOL-USA, Inc., Peabody, At the end of manufacture, “requeijão cremoso” cheeses had
Mass., U.S.A.) at 10 kV. the following mean composition: moisture 63.23 ± 0.36%, fat
23.82 ± 0.46%, fat in a dry wet basis 64.82 ± 0.82%, salt
Rheological properties 1.05 ± 0.04%, protein 10.51 ± 0.25%, nonprotein nitrogen
The mechanical spectra of the samples were obtained using 0.02 ± 0.00%, and total calcium 160.3 ± 1.5%. The degree of
a Paar Physica MCR 300 rheometer (Anton Paar GmbH, Graz, casein dissociation for the cheeses made with SHMP, TSPP, TSC,
Áustria), with a 5 cm diameter, 2◦ stainless steel cone–plate system. and STPP were, respectively, 69.2 ± 2.8%, 89.0 ± 1.6%, 86.6 ±
The truncation of the cone (gap) was 100 μm. The temperature 1.6%, and 85.5 ± 2.1%. Figure 2 shows pH and percentage soluble
of the samples was maintained at 10 ◦ C, on the bottom plate, calcium/total calcium of the processed cheeses as a function of the
by a Peltier system. Sufficient sample was held by a spatula, a type of ES used (adapted from Cunha and Viotto 2010).
E: Food Engineering &

second spatula was then used to gently push the sample on to the The pH and the degree of casein dissociation of the cheese man-
Physical Properties

bottom plate of the rheometer. The plate was raised slowly to the ufactured with SHMP were significantly (P < 0.05) lower than
measuring position. Excess samples were trimmed off the edges of those of the other cheeses. These findings are in agreement with
the cone using a plastic spatula. the results reported by Dimitreli and others (2005), Dimitreli and
In the oscillatory tests, G’ (elastic modulus), G” (viscous mod- Thomareis (2009), and Sádlı́kova and others (2010). The value for
ulus), and η∗ (complex viscosity) were measured at frequencies the percentage soluble calcium/total calcium was also significantly
ranging from 0.01 to 10 Hz at a strain of 0.3 Pa, which was (P < 0.05) lower for the SHMP cheese as compared to the oth-
within the linear viscoelastic region (determined by preliminary ers. In general, cheeses made with TSPP, TSC, and STPP showed
experiments). a more similar composition, although slight differences could be
In the rotational tests, the flow curves of the samples were observed, particularly with respect to pH, which was higher for
obtained using the controlled shear rate mode of the rheometer TSPP cheese than to TSC cheese, and to the percentage soluble
at a shear rate range of 0 to 100 per s. The total run time was 8 calcium/total calcium, which was higher for STPP cheese than
min. The data for shear stress (τ ) as a function of the deformation to the other 2. The reasons for the differences found with respect
rate (dγ /dt) were fitted to the power-law model τ = k(dγ /dt)n , to these parameters have been extensively discussed in an earlier
where k is the consistency index and n the flow behavior index. study (Cunha and Viotto 2010).

Experimental design Microstructure of the “requeijão cremoso” cheese made

A random block experimental design was adopted. The factor with different ESs
studied was the type of ES used (TSC, SHMP, STPP, and TSPP). TSC, TSPP, STPP, and SHMP used as ESs gave an efficient
Four trials were carried out in triplicate, giving a total of 12 exper- emulsification for all processed cheese samples.
iments. The results were evaluated using the analysis of variance Figure 3 shows the microstructure (5000 × magnification) of
(ANOVA), using the Tukey test to verify differences between the processed cheeses manufactured with TSC (a), SHMP (b), TSPP
means (P < 0.05). The analyses were performed using a statis- (c), and STPP (d). The following typical processed cheese struc-
R
tures can be seen in all samples: a continuous protein matrix per-
tical program (Minitab 12.1 for Windows, Minitab Inc., State
College, Pa., U.S.A.). meated by fat globules and, eventually, by water (small irregular
shaped spaces, not covered by protein). The fat globules presented
predominantly spherical shapes, uniformly distributed through-
out the protein matrix, in agreement with results previously ob-
tained for processed cheeses (Tamime and others 1990; Mistry and
Anderson 1993).
Cheese made with SHMP presented smaller fat globules than the
other cheeses (Figure 3). Photomicrographs show also that cheeses
made with SHMP and TSPP presented a more homogenous dis-
tribution, whilst cheese made with STPP and TSC presented a
wider range of particle size. Previously reported data on the fat
particle size distribution of the same cheeses, measured using a
particle size analyser, showed similar results (Cunha and Viotto
2010).
The presence of a large number of small fat globules indicates
that SHMP was more efficient in promoting fat emulsification,
which seems contradictory since cheese made with SHMP also
showed the lowest degree of casein dissociation. As discussed by
Figure 2–pH and percentage soluble calcium/total calcium of the pro- Cunha and Viotto (2010), this apparently conflicting result could
cessed cheeses as a function of the type of emulsifying salt: trisodium be explained by the hypothesis that, during manufacture, SHMP
citrate (TSC); sodium hexametaphosphate (SHMP); tetrasodium pyrophos-
phate (TSPP); pentasodium tripolyphosphate (STPP). Adapted from previ-
promoted an effective peptization of the casein, which sounds
ously reported results (Cunha and Viotto 2010). 209 × 297 mm (300 × reasonable since the calcium binding capacity increases with the
300 DPI). number of phosphate anions in the chain. The solubilization of

E178 Journal of Food Science r Vol. 77, Nr. 8, 2012

Effect of emulsifying salt on rheology . . .

Table 1 shows the mean values obtained for G’, G”, η∗ , and
casein allowed it to act as a true emulsifier, but after that prob-
ably occurred the formation of a new network, composed of phase angle (G”/G’) for the processed cheeses manufactured with
re-associated casein and insoluble calcium phosphate. In the case
different types of ES. The type of ES significantly influenced
(P < 0.05) all the parameters evaluated.
of TSC, TSPP, and STPP, the re-association of the casein was lim-
ited, possibly because of the higher pH and the chemical nature ofThe cheeses manufactured with TSC, TSPP, and STPP
the salts, which explains the higher degrees of casein dissociation.
(Figure 4a, 4c, and 4d) presented typical behavior of concen-
The greater homogeneity in particle size distribution found for
trated solutions: G” was greater than G’ at low frequencies, G’
TSPP cheese as compared to cheeses made with STPP and TSC was greater than G” at higher frequencies, and η∗ was strongly
have been also discussed by Cunha and Viotto (2010), and could dependent on the frequency. At higher frequencies, there was
be explained by the slightly higher pH showed by TSPP cheese insufficient time for the intermolecular bonds to break during
(Figure 2). During manufacture, the higher pH improved the cal-the oscillation period, and thus the network behaved like a gel,
cium sequestering ability of the salt (Guinee and others 2004; presenting characteristics more typical of solids than of liquids
(G’ > G”). At lower frequencies, the molecular chains had suffi-
Dimitreli and others 2005), allowing casein to dissociate more ef-
fectively and to act as a more efficient emulsifier than in TSCcient time to undergo re-arrangements and flow.
or STPP cheeses. During cooling, a certain level of casein re- It can be seen that the frequency in which transition occurred

E: Food Engineering &

association probably occurred for the 3 samples, but the extension
was smallest for the cheese manufactured with TSPP, followed by

Physical Properties
and the mechanisms of re-association were possibly different de-
the cheeses manufactured with TSC and STPP, respectively. This
pending on the type of salt, which would explain the results found
means that the molecular interactions were probably stronger in the
for the degree of casein dissociation and the percentage soluble
cheese manufactured with TSPP than in those manufactured with
calcium/total calcium. TSC or STPP, much lower frequencies being required to allow for
molecular re-arrangements to occur. This result can be explained
Influence of the type of ES on rheological properties of by the size and distribution of the fat globules. In the cheeses
“requeijão cremoso” cheese manufactured with STPP and TSC, the fat globules were larger
Oscillatory tests. Figure 4 shows the mechanical spectra of and showed less uniformity in size than in the cheese manufactured
the processed cheeses manufactured with different ESs, whereas with TSPP (see Figure 3). The emulsified fat globules covered

Figure 3–Scanning electron microscopy of “requeijão cremoso” cheeses manufactured with different emulsifying salts: (a) trisodium citrate (TSC), (b)
sodium hexametaphosphate (SHMP), (c) tetrasodium pyrophosphate (TSPP), (d) pentasodium tripolyphosphate (STPP). 5000 ×, at 10 kV. Black arrows
indicate protein matrix and white arrows indicate fat globules. Scale bar = 1 μm. 150 × 113 mm (300 × 300 DPI).

Vol. 77, Nr. 8, 2012 r Journal of Food Science E179

 Effect of emulsifying salt on rheology . . .

with casein function as large protein (or pseudo protein) particles, the other cheeses, indicating that, in this sample, the molecular
which are incorporated into the protein network (Marchesseau interactions were even weaker.
and others 1997; Michalski and others 2002; Guinee and others The cheese manufactured with SHMP (Figure 4b) showed be-
2004), increasing gel strength. The larger the diameter and the havior typical of a weak gel: G’ greater than G” throughout the
lesser the uniformity of the fat globules, the smaller the surface area frequency range, both moduli being little dependent on the fre-
available for interaction and the less the amount of protein–protein quency, and η∗ fell sharply with the increase in frequency. This in-
interactions, justifying the higher frequencies of transition found dicated that the cheese manufactured with SHMP had a more elas-
in the cheeses manufactured with TSC and STPP as compared to tic structure with stronger molecular interactions than the other
those manufactured with TSPP. In the cheese manufactured with samples. The small size of the fat particles found in this sample
STPP (Figure 4d), the frequency in which the transition from must have contributed to this, since the smaller the diameter, the
“liquid” to “solid” behavior occurred was much higher than in greater the number of protein–protein bonds.
The type of ES significantly influenced the phase angle (G”/G’)
(Table 1). In decreasing order of G”/G’, the cheeses manufactured
were as follows: STPP > TSC > TSPP > SHMP. A high value
for the phase angle indicates a less structured and more peptized
E: Food Engineering &

protein network. The result found could be explained by the size

Physical Properties

of the fat globules of the samples. The cheese manufactured with

SHMP showed the smallest fat globule diameter, which made the
network firmer and more structured, with a low value for G”/G’.
The cheeses manufactured with STPP and TSC showed lower
values for G’ (elastic modulus) and η∗ (complex viscosity) as com-
pared to the others (Table 1). This result could also be explained
by the larger size and reduced uniformity of the fat globules found
in these samples (see Figure 3).
The size of the fat globules was not the only factor that in-
fluenced the elastic modulus, since, despite the difference in size
of the fat globules of the cheeses manufactured with TSPP and
SHMP (Figure 3), there was little difference between them with

Table 1– Mean values (n = 3) for G’ (elastic modulus), G” (vis-

cous modulus), η∗ (complex viscosity), and G”/G’ (phase angle)
of the processed cheeses manufactured with different emulsify-
ing salts: sodium hexametaphosphate (SHMP), tetrasodium py-
rophosphate (TSPP), trisodium citrate (TSC), and pentasodium
tripolyphosphate (STPP). The reported values are related to the
frequence 5.11 Hz. Values in parenthesis indicate standard de-
viation related to 3 replicates.
Sample SHMP TSPP TSC STPP
a a b
G’ (Pa) 1493 1616 779 427b
(97) (129) (48) (22)
G” (Pa) 730b 1196a 680b 464b
(64) (122) (77) (15)
η∗ (Pa.s) 51.8a 62.7a 32.2b 19.6b
(4.6) (5.9) (3.5) (0.8)
G”/G’ (o ) 0.49d 0.74c 0.88b 1.09a
(0.03) (0.04) (0.05) (0.03)
a,b,c,d
Means with the same letter, in the same line, do not differ significantly (P > 0.05).

Table 2–Mean values (n = 3) for n (flow behavior index), k (consis-

tency index) and R2 of the processed cheeses manufactured with
different emulsifying salts: sodium hexametaphosphate (SHMP),
tetrasodium pyrophosphate (TSPP), trisodium citrate (TSC), and
pentasodium tripolyphosphate (STPP). Values in parenthesis in-
dicate standard deviation related to 3 replicates. Experimental
data obtained in rotational tests (shear stress × shear rate) were
fitted to the power-law model: τ = k(dy/dt)n , where τ = shear
stress (Pa), k = consistency index (Pa.sn ), (dy/dt) = shear rate
(per s) and n = flow behavior index (dimensionless).
Sample SHMP TSPP TSC STPP
b b a,b
n 0.24 0.24 0.30 0.36a
(0.03) (0.03) (0.01) (0.02)
Figure 4–Mechanical spectra of the processed cheeses manufactured with k (Pa.sn ) 242.7c 701.7a 409.0b 232.9c
different emulsifying salts: (a) trisodium citrate (TSC); (b) sodium hex- (23.0) (66.8) (33.9) (18.8)
ametaphosphate (SHMP); (c) tetrasodium pyrophosphate (TSPP); (d) pen- R2 0.979 0.928 0.955 0.970
tasodium tripolyphosphate (STPP). The results showed refer to the 2nd
replicate. 209 × 297 mm (300 × 300 DPI).
a,b,c
Means with the same letter, in the same line, do not differ significantly (P > 0.05).

E180 Journal of Food Science r Vol. 77, Nr. 8, 2012

Effect of emulsifying salt on rheology . . .

respect to G’. In SHMP cheese, the high values for G’ were due Conclusions
to the combined effects of small fat droplets, low degree of casein The type of ES influenced the microstructure and the rheologi-
dissociation and low percentage soluble calcium/ total calcium, cal properties of processed cheese. SHMP was the most efficient fat
which resulted in a more elastic protein network. In cheese made emulsifier, resulting in the network with the smallest fat globules
with TSPP, on the other hand, the high values of G’ could be ex- size. The cheeses manufactured with TSC, TSPP, and STPP be-
plained both by a better emulsification in comparison with TSC haved like concentrated solutions, while the cheese manufactured
and STPP cheeses, and by its higher pH in comparison with with SHMP exhibited weak gel behavior and the lowest values for
SHMP cheese. Increasing pH increases the negative charge of the the phase angle G”/G’. This means that the protein network with
protein molecules, causing their repulsion and expansion. In pro- the largest amount of molecular interactions was obtained for the
cess cheese, ionic repulsion in the pH range 5.7 to 6.0, instead cheese made with SHMP, which is in agreement with the highest
of totally dispersing proteins, enhances different type of interac- degree of fat emulsification found for this sample. Rotational vis-
tions, such as noncovalent bonds (hydrogen bonds, hydrophobic, cometry indicated that all the samples behaved like pseudoplastic
and electrostatic interactions) (Marchesseau and others, 1997), thus fluids. Because of the strongest network structure of cheese made
increasing elasticity. with SHMP, it also presented a low value for the flow behavior
Rotational tests. Table 2 shows the mean values for n and k index, meaning that viscosity was more dependent on shear rate.

E: Food Engineering &

for the cheeses manufactured with different ESs. The type of ES With respect to the consistency index, the cheese manufactured

Physical Properties
significantly (P < 0.05) influenced the 2 parameters evaluated. with TSPP showed the highest value as compared to the other
All the cheeses presented n values below 1, indicating that the samples, indicating a higher viscosity that could be attributed to
apparent viscosity decreased with increase in the deformation rate the combined effect of its high pH and homogeneous fat particle
applied, that is, they presented the behavior of pseudoplastic fluids. size distribution. For the cheese industry, an improved knowledge
The cheese manufactured with STPP showed the highest value for about how each type of ES affects the structure and the rheologi-
n, indicating that the proteins were freer to orientate themselves in cal properties of processed cheese will permit a most appropriate
the direction of flow than in the cheeses manufactured with TSPP selection of the blend of ESs and a better control of the texture
and SHMP. This result is in agreement with the less structured pro- characteristics of the final product.
tein network showed by this sample in oscillatory measurements
(Table 1). Cheese made with TSC presented an intermediate value Acknowledgements
for n, meaning that its protein network was slightly firmer than This research was carried out with the financial support of the
that of STPP cheese but less structured than those of SHMP or State of São Paulo Research Foundation, FAPESP, Brazil and the
TSPP cheeses, which is consistent with the intermediate value of Brazilian National Research Council, CNPQ, Brazil.
G”/G’ showed by this sample (Table 1).
The cheese manufactured with SHMP showed a low value for References
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