Carrageenan type effect on soybean oil/soy protein isolate emulsion employed as fat replacer in panela-type cheese

Abstract

In order to modify the fatty acid profile of panela-type cheese (a Mexican fresh cheese), emulsified soybean oil with soy protein isolate and different carrageenan (iota, kappa or lambda) was employed as fat replacer. The replacement of milk fat in panela-type cheese resulted in higher cheese yield values and moisture content, besides a concomitant lower fat phase and higher protein content, due to a soy protein isolate in emulsified soybean oil. Fat replacement resulted in a harder but less cohesive, spring and resilient texture, where differences in texture could be attributed to the specific carrageenan-casein interactions within the rennet coagulated cheese matrix. The FTIR analysis showed that the milk fat replacement changed the fatty acid profile, also in function of the type of carrageenan employed. Lambda carrageenan containing emulsions improved moisture retention and maintained the textural properties of panela-type cheese.

Full text

GRASAS Y ACEITES 66 (4)
October–December 2015, e097
ISSN-L: 0017-3495
doi: http://dx.doi.org/10.3989/gya.0240151
Carrageenan type effect on soybean oil/soy protein isolate
emulsion employed as fat replacer in panela-type cheese
E. Rojas-Nerya, N. Güemes-Verab, G.O. Meza-Marqueza and A. Totosausa,*
aFood Science Lab & Pilot Plant, Tecnológico Estudios Superiores Ecatepec. Av. Tecnológico esq.
Av. Central s/n, Ecatepec 55210, Estado de México, MEXICO
bInstituto de Ciencias Agropecuarias, Universidad Autónoma Estado Hidalgo.
Av. Universidad km. 1, Tulancingo 43600, Hidalgo, MEXICO
*Corresponding author: alfonso.totosaus@gmail.com
Submitted: 10 February 2015; Accepted: 09 April 2015
SUMMARY: In order to modify the fatty acid profile of panela-type cheese (a Mexican fresh cheese), emulsi-
fied soybean oil with soy protein isolate and different carrageenan (iota, kappa or lambda) was employed as fat
replacer. The replacement of milk fat in panela-type cheese resulted in higher cheese yield values and moisture
content, besides a concomitant lower fat phase and higher protein content, due to a soy protein isolate in emulsi-
fied soybean oil. Fat replacement resulted in a harder but less cohesive, spring and resilient texture, where differ-
ences in texture could be attributed to the specific carrageenan-casein interactions within the rennet coagulated
cheese matrix. The FTIR analysis showed that the milk fat replacement changed the fatty acid profile, also in
function of the type of carrageenan employed. Lambda carrageenan containing emulsions improved moisture
retention and maintained the textural properties of panela-type cheese.
KEYWORDS: Carrageenan; Emulsified soybean oil; Fat replacement; Fourier Transform Infra-Red Spectroscopy;
Panela-type cheese; Textural profile analysis
RESUMEN: Efecto del tipo de carragenina en emulsiones de aceite de soja/aislado de proteína de soja utilizadas
como sustituto de grasa en quesos tipo panela. Para modificar el perfil de ácidos grasos de los quesos tipo panela
(queso fresco popular en México), se utilizó aceite de soja emulsionado con aislado de proteína de soja y diferen-
tes carrageninas (iota, kappa o lambda) como sustituto de la grasa. Reemplazar la grasa de la leche en el queso
tipo panela resultó en mayor rendimiento quesero y mayor contenido de humedad, además de una concomitante
menor fase grasa y mayor contenido de proteína, debido al aislado de proteína de soja en el aceite de soja emul-
sionado. La sustitución de la grasa dio como resultado una textura más dura, pero menos cohesiva, elástica y
resiliente, donde estas diferencias podrían ser atribuidas a la interacción especifica entre carrageninas-caseinas
en la matriz coagulada del queso. El análisis de FTIR muestra que reemplazar la grasa de la leche cambia el
perfil de ácidos grasos, también en función del tipo de carragenina empleado. Las emulsiones con lambda carra-
geninas mejoraron la retención de humedad y mantuvieron las propiedades de textura del queso tipo panela.
PALABRAS CLAVE: Aceite de soja emulsionado; Análisis del perfil de textura; Carrageninas; Infrarrojo con
Transformada de Fourier; Queso tipo panela; Sustituto de grasa
Citation/Cómo citar este artículo: Rojas-Nery E, Güemes-Vera N, Meza-Marquez GO, Totosaus A. Carrageenan type effect
on soybean oil/soy protein isolate emulsion employed as fat replacer in panela-type cheese. Grasas Aceites 66 (4): e097.
http://dx.doi.org/10.3989/gya.0240151.
Copyright: © 2015 CSIC. This is an open-access article distributed under the terms of the Creative Commons
Attribution-Non Commercial (by-nc) Spain 3.0 Licence.

2 • E. Rojas-Nery, N. Güemes-Vera, G.O. Meza-Marquez and A. Totosaus
Grasas Aceites 66 (4), October–December 2015, e097. ISSN-L: 0017–3495 doi: http://dx.doi.org/10.3989/gya.0240151
1. INTRODUCTION
Dairy products like cheese are an important source
of essential nutrients (mainly proteins, besides cal-
cium, magnesium and potassium, riboflavin and
vitamin B12). Nonetheless, dairy products like cheese
are one of the major contributors of saturated fatty
acid (SFA) intake, compounds associated with the
development of chronic diseases such as cardiovas-
cular disease, congestive heart failure, and obesity
(Livingstone et al., 2012). Consumer awareness of
dietary fat has increased and the demand for low-fat
foods, including cheese, has grown substantially, with
a strong influence on the market. The removal of the
fat from the casein network in low-fat cheese results
in the formation of a more compact casein network
that released more water and became tougher. To
improve the textural characteristics of low-fat cheese,
the moisture levels in curd must be increased (Banks,
2004). To compensate for these changes, fat substi-
tutes or fat mimics can be employed. Fat substitutes
are generally lipid-based macromolecules that physi-
cally and chemically resemble fats and oils such as
sucrose fatty acid esters and polyesters, carbohydrate
fatty acid esters, various emulsifiers (such as mono-
and di-glycerides, lecithin), and structured lipids. Fat
mimics are generally carbohydrate-based (modified
starches and hydrocolloids) or protein-based macro-
molecules that are designed to mimic the organolep-
tic and physical properties of fats generally via the
binding of water (Johnson etal., 2009).
Fresh cheese was elaborated from natural cows’
milk, pasteurized, non-acidified, and with an elevated
water content (up to 58%). In 2014, cheeses pro-
duction in Mexico was around 312,082 tons, where
fresh and panela cheese represented 17.8 and 15.7%,
respectively (SIAP, 2014). The higher consumption
of fresh cheese calls for an improvement in quality
characteristics by offering low fat fresh-type prod-
ucts. Processed cheese products manufactured by
blending various edible oils, proteins and other ingre-
dients can offer an attractive food-based delivery
vehicle for lipids such as polyunsaturated fatty acids
or omega-3 fatty acids (Ye etal., 2009). The substitu-
tion of milk fat by vegetable oils can contribute to a
healthier saturated/unsaturated fat balance in cheese
(Yu and Hammond, 2000, Fathi Achachlouei etal.,
2013, Lobato-Calleros etal., 2002, 2003, 2007), by
lowering cholesterol levels and impacting human
nutrition (Kesenkas et al., 2009). Soybean is the
dominant oilseed produced in the world, due to its
favorable agronomic characteristics, high-quality
protein, and valuable edible oil, contributing to the
half of all oilseeds produced worldwide. Soybean oil’s
particular fatty acid composition is higher in linoleic
acid and lower in linolenic acid, in comparison with
the other major vegetable oils, both essential fatty
acids for humans and therefore of dietary importance
(Wang, 2002). In the same manner, soy protein ingre-
dients possess appropriate functional properties for
food applications due their high nutritional value
and their ability to form and stabilize emulsions
(Kinsella, 1979). In this view, since vegetable oils had
been employed to replace fat in cheese elaborations,
a viable alternative is to use an emulsion elaborated
with soybean oil and soy protein isolate. The incor-
poration of polysaccharides to a protein solution
improves the stability of oil droplets against cream-
ing (Uruakpa and Arnfield, 2005). Polysaccharides
such as carrageenans are widely employed in the
dairy industry due to their specific reactivity with
casein micelles (Drohan et al. 1997, Langendorff
etal., 1999, 2000). The incorporation of carrageen-
ans in soybean oil/soy protein isolate emulsion,
could, on one hand, results in a more stable emul-
sion during and after cheese processing, given that
a synergistic effect between soy protein isolate and
carrageenans had been reported (Molina Ortiz etal.,
2004). On the other hand, the presence of different
types of carrageenans in emulsified soybean oil will
affect the physicochemical and textural properties of
panela-type cheese in a different way, due to reactiv-
ity between caseins and carrageenans. The aim of this
work was to determine the effect of milk fat replace-
ment with emulsified soybean oil, employing soy pro-
tein isolate and different carrageenans (iota, kappa or
lambda) on the physicochemical, textural and FTIR
spectroscopy of panela-type cheese.
2. MATERIALS AND METHODS
2.1. Emulsi ed soy oil and panela-type cheese
elaboration
Soy protein isolate Appensol ISL (DVA Mexicana,
Naucalpan, Mexico) was dissolved in distilled water
(5% w/v) before being mixed with different types of
carrageenans (1%, w/v): kappa-carrageenan Gelcarin
GP8612, iota-carrageenan Viscarin SD389 or lambda-
carrageenan Viscarin GP209 (FMC BioPolymers,
Philadelphia, USA). To each protein-carrageenan
suspension, Nutrioli® pure soy oil (Ragasa Industrial
S.A de C.V., Monterrey, México) was incorporated
in a proportion of 80:20 (v/v) and mixed employing
an Oster homogenizer (Sumbean Mexicana, Mexico,
12,000 rpm) until obtaining a homogenous ‘mayon-
naise’. The emulsified oil was stored in plastic bags at
−20 °C until use as a milk fat replacer.
Raw milk was obtained from dairy facilities
of the Universidad Autonoma Estado Hidalgo at
Tulancingo, Mexico. Milk (4.37% fat content) was
filtered and pasteurized (63 °C for 30 min) then
cooled at 42 °C. Part of the milk was skimmed in
an Elecrem 3 cream separator (Fresnes, France) to
a final milk fat content of 0.2%. Whole (full fat)
and skimmed milk were mixed in order to obtain

Carrageenan type effect on soybean oil/soy protein isolate emulsion employed as fat replacer in panela-type cheese • 3
Grasas Aceites 66 (4), October–December 2015, e097. ISSN-L: 0017–3495 doi: http://dx.doi.org/10.3989/gya.0240151
different milk fat proportions, incorporating the
different emulsified soy oil formulations to replace
and compensate for milk fat, according to Table 1.
The different milk batches with different percent-
ages of milk fat replaced with emulsified soybean
oil/soy protein isolate and iota, kappa or lambda
carrageenan were kept at a constant temperature
of 40 °C and mixed before the addition of 0.01%
v/v of Cuamex® rennet (Chr. Hansen de México S.
A. de C. V., México). The curd was left to settle for
30 min and cut into 1 cm3 cubes. The cut curd was
then subjected to mild stirring during 30 min. The
whey was drained and the upper layer of the whey
and salt (0.8% w/v) were mixed with the curd. The
curd was molded in 2 kg containers. The individual
cheeses were pressed and the remaining whey was
expelled. After 90 min, the cheeses were weighed to
determine yield, and vacuum packed until further
analysis. Each cheese formulation was elaborated
in duplicate.
2.2. Cheese yield, moisture, fat phase and total
protein content
Cheese yield (%) was calculated as the percent
weight of the finished cheese divided by the weight
of the milk employed for each batch (Drake etal.,
1996). Cheese moisture was determined according
to the AOAC Official Method No. 926.08 (AOAC,
1998) employing aluminum pans (ca. 3 g of sample)
dehydrated in an oven at 100 °C for 4 h. Fat con-
tent was determined by the acid butyric method of
Van Gulik according to the ISO 3433 (ISO, 2011).
Total protein content was determined in agreement
with the AOAC Official Method 926 (AOAC, 1998)
employing a conversion factor of 6.38. All analyses
were performed in triplicate.
2.3. Textural pro le analysis
The panela-type cheese texture analysis was per-
formed in a texture analyzer LFRA 4500 (Brookfield
Engineering, Middleboro, MA, USA). Samples were
obtained from the middle of the whole cheese blocks
(4x4 cm and 2 cm height) which were kept in plastic
bags to avoid moisture loss at room temperature. The
samples were consecutively compressed two times
(30%) with a 7 cm diameter acrylic disk with a
5- second waiting period at a crosshead speed of
1 mm/s. From the force-deformation curves tex-
ture profile parameters were calculated as follows:
hardness (force necessary to attain a given deforma-
tion, maximum force), adhesiveness (work neces-
sary to overcome the attractive forces between the
surface of the food and the surface of other materi-
alswith which the food comes in contact), cohesive-
ness (strength of theinternal bonds making up the
body of the product), springiness (degree to which a
product returns to its original shape once it has been
compressed), and resilience (capacity to recover its
original shape after compression) (Szczesniak, 1963;
Bourne, 1978). The results are the mean of at least
three reproducible runs per cheese batch.
2.4. Fourier transformation infra-red spectroscopy
analysis
A Fourier Transform Infrared spectrophotom-
eter PerkinElmer model Spectrum GX (Perkin
Elemer Inc, Walthman, MA, USA) equipped with
a horizontal attenuated total reflection accessory
and diamond point was employed to determinate
changes in cheeses composition. Sixty-four scans
were coded at a nominal resolution of 4 cm−1 in the
spectral region 4000–550 cm−1. Single beam spectra
of the samples were collected against a background
of air and presented in absorbance units. The cheese
samples were placed in the HATR accessory to give
total crystal coverage, cleaning the crystal between
samples.
2.5. Experimental design and data analysis
In order to determinate the effect of milk fat re plac-
ement in panela-type cheese employing emulsified
soy oil, a complete factorial design was employed.
The proposed model for the data was:
yij=μ+αi+βj+∈ij Eq. (1)
where yij represents the variable response for the i-th
level of fat replacement (0, 25, 50 and 75%), at the
j-th type of carrageenan (kappa, iota or lambda);
μ is the overall mean; αi and βj are the main effects
TABLE 1. Milk fat replacement with emulsified soybean oil/soy protein isolate and carrageenans
Replacement (%)
Whole milk liters
(4.37% fat)
Total fat in milk
after skimming (g)
Skim milk liters
(0.2% fat)
Emulsified soybean/soy protein isolate and
carrageenans added to replace milk fat (g)
Total
fat (%)
0 4 174.8 0 0 4.37
25 3 131.1 1 44.6 3.28
50 2 87.4 2 89.2 2.18
75 1 43.7 3 133.8 1.09

4 • E. Rojas-Nery, N. Güemes-Vera, G.O. Meza-Marquez and A. Totosaus
Grasas Aceites 66 (4), October–December 2015, e097. ISSN-L: 0017–3495 doi: http://dx.doi.org/10.3989/gya.0240151
of the level of substitution and carrageenan type;
and …ij is the residual or error terms assumed to be
normally distributed with zero mean and variance
σ2 (Der and Everitt, 2002). The results were ana-
lyzed according to the PROC ANOVA procedure in
SAS Software v 8.0 (SAS System, Cary). Significant
differences means were determined by the Duncan
means test.
3. RESULTS
3.1. Yield, moisture, fat phase and protein
The use of emulsified oil to replacemilkfatre -
sulted in a significantly (p<0.05) higher panela-
type cheese yield, where a major proportion of
replacement (upto 75%) resulted in higher yield
values. In same manner, emulsified oil with lambda
carrageenan increased the cheese yield significantly
(p<0.05) (Table2). Higher yield values were related
to an increase in cheese moisture content andreten-
tion of this water into the coagulated system.Panela-
type cheese moisture was significantly (p<0.05)
higher when milk fat was replaced with emulsi -
fied oil above 50%. Lambda carrageenan containing
emulsions resulted in significantly (p<0.05) higher
cheese moisture contents (Table 2). As fat was
replaced with emulsified oil, the fat phase decreased
in the panela-type cheese. Replacing 75% of milk fat
with emulsified oil resulted in significantly (p<0.05)
lower fat content, with higher fat content in control
(full-fat) samples. For the carrageenan type, emul-
sified oil with lambda carrageenans resulted in sig-
nificantly (p<0.05) lower percentages of fat in the
cheese (Table2). The replacement of milk fat with
emulsions made with soy protein isolate increased
the protein content in the cheese. Total protein was
significantly (p<0.05) higher when 75% of milk fat
was replaced with emulsified oil. Lower protein con-
tent was obtained in control samples. For the carra-
geenan type, kappa and iota carrageenan treatments
resulted in significantly (p<0.05) higher protein
contents (Table 2).
3.2. Texture pro le analysis
For cheese hardness, the replacement of 25 or
50% milk fat with emulsified soybean oil resulted in
a significantly (p<0.05) harder texture. In the same
way, the panela-type cheese texture was significantly
(p<0.05) harder when iota or kappa carrageenan was
employed in the emulsified oil (Table 3). Panela-type
cheese adhesiveness was not significantly (p>0.05)
different for the percent of milk fat replacement with
emulsified oil. For the carrageenan type, the emul-
sified oil containing iota or kappa carrageenan was
significantly (p<0.05) more adhesive (Table 3).
In contrast, elastic related textural parameters
(cohesiveness, springiness and resilience) were re -
duced when emulsified oil was employed to replace
milk fat. The cohesiveness of cheeses was significantly
(p<0.05) lower when milk fat was replaced. In same
manner, incorporating iota or kappa carrageenan to
cheese elaboration significantly (p<0.05) decreased
cheese cohesiveness (Table 3). The springiness of the
panela-type cheese was significantly (p<0.05) lower
when emulsified oil replaced milk fat. Iota or kappa
carrageenan in emulsified soybean oil employed to
replace milk fat decreased cheese springiness sig-
nificantly (p<0.05) as well (Table 3). Significantly
(p<0.05) lower resilience values were also observed
when emulsified soybean oil replaced milk fat. Iota
or kappa carrageenan in emulsified oil decreased
sample resilience significantly (p<0.05) (Table 3).
TABLE 2. Physicochemical properties of fat-reduced panela-type cheese employing emulsified oil with carrageenans
Carrageenan type in emulsified
soybean oil/soy protein isolate
Milk fat
substitution (%) Yield (%) Moisture (%) Fat (%)
Total protein
(%)
Control 0 16.41±0.00d,C 56.84±2.03c,C 30.40±0.21a,A 12.41±0.30c,C
Iota 25 17.18±0.00c,B 57.88±2.51b,B 27.00±1.20b,B 12.39±0.20c,A
50 17.47±0.17b,B 58.71±2.25a,B 26.50±0.60c,B 11.83±0.14b,A
75 17.60±0.00a,B 58.92±1.91a,B 25.50±0.25d,B 14.11±0.03a,A
Kappa 25 15.50±0.00c,B 58.74±1.95b,B 27.20±0.10b,B 12.48±0.36c,A
50 15.40±0.00b,B 59.14±2.03a,B 26.80±0.30c,B 13.54±0.31b,A
75 16.22±0.00a,B 59.81±2.38a,B 25.64±0.30d,B 13.41±0.05a,A
Lambda 25 16.41±0.00c,A 59.62±2.52b,A 26.90±1.20b,C 12.60±0.18c,B
50 17.70±0.00b,A 60.40±2.12a,A 25.85±1.73c,C 13.65±0.17b,B
75 19.08±0.00a,A 60.72±1.71a,A 25.18±0.30d,C 12.21±0.17a,B
a,b,c,dMeans with the same letter in the same column are not significantly (p>0.05) different for the percent of milk fat substitution.
A,B,CMeans with the same letter in same column are not significantly (p>0.05) different for the carrageenan type.

Carrageenan type effect on soybean oil/soy protein isolate emulsion employed as fat replacer in panela-type cheese • 5
Grasas Aceites 66 (4), October–December 2015, e097. ISSN-L: 0017–3495 doi: http://dx.doi.org/10.3989/gya.0240151
3.3. Fourier transformation infra-red spectroscopy
analysis
Figure 1 shows the MIR spectra for the different
panela-type cheeses formulated with the different milk
fat substitutions (0, 25, 50 and 75%) for each emul-
sified oil containing the different carrageenan types
(iota, kappa and lambda). Arrows indicate intensity
of absorption bands as a result of the increase in milk
fat substitution by emulsified oil. MIR spectra present
differences in the absorbance magnitude for each one
of the panela-type cheese samples, since each band
or spectral peak is directly related to the substitution
and carrageenan type (iota, kappa or lambda). This
reflects the changes in cheese chemical composition
for the different milk fat replacement and the differ-
ent carrageenans employed in emulsified oil formula-
tion, contrasting the differences at the 2100–950 cm−1
region for iota (Fig. 1-a), kappa (Fig. 1-b) or lambda
(Fig. 1-c) carrageenans.
4. DISCUSSION
4.1. Yield, moisture, fat phase and protein
The replacement of milk fat with emulsified oil
affected the physicochemical properties of panela-
type cheese. The main effect was regarding water
retention improvement in the coagulated cheese
curd. Since soybean oil emulsions were emulsified
and stabilized with soy protein isolate and car-
rageenans, the hydration properties and interac-
tions of both theese macromolecules with caseins
(before, during and after being rennet coagulated)
seems to be the explanation for this behavior,
resulting in higher yield due to higher moisture
retention.
On one hand, the use of emulsified soybean oil
implies the presence of additional water in the sys-
tem, along with proteins and polysaccharides that
can retain water in the system and/or interact with
milk proteins during cheese processing. Giroux etal.
(2013) reported that higher moisture content was
found in model cheese made from double emulsions,
resulting in the incorporation of larger oil droplets,
increasing the openness of the cheese matrix and cre-
ating more interstitial space, providing a reservoir for
retaining additional moisture in cheese. Fat globules
are occluded in the paracasein network pores ofthe
cheese, physically limiting the aggregation ofthesur-
rounding paracasein network, reducing proteinmatrix
contraction and moisture expulsion (Fox etal., 2000;
Lobato-Calleros etal., 2007). Milk fat replacement is
related to the generation of larger interstitial spaces,
due to larger oil droplets, resulting in higher mois-
ture retention. The moisture content of cheese was
inversely proportional to milk fat content, and milk
fat content reduction in the cheese increased moisture
content (Romeih etal., 2002). In addition, the more
retained water resulted in higher cheese yield in sam-
ples with emulsified oil replacing milk fat. In the same
manner, the control full-fat samples had higher fat
contents than cheese made with emulsion since emul-
sions contained a lower true fat fraction, resulting in
lower fat content and higher protein content (Giroux
etal., 2013). When more emulsified oil was incorpo-
rated to replace milk fat, a higher protein content was
observed due to the soy protein isolate employed to
formulate the emulsions. Higher protein content in
low fat cheese increased the water binding capacity
of the cheese matrix (Fathi Achachlouei etal., 2013).
The addition of a fat replacer increased the cheese
yield probably due to its higher moisture retention
ability (Sahan etal., 2008). The presence of both soy
TABLE 3. Textural profile analysis of fat-reduced panela-type cheese employing emulsified oil with carrageenans
Carrageenan type in emulsified
soybean oil/soy protein isolate
Milk fat
substitution (%) Hardness (N) Adhesiveness (N)
Cohesiveness
(dimensionless)
Springiness
(dimensionless) Resilience
Control 0 31.40±0.82b,C 0.75±0.40a,B 0.39±0.03a,A 0.80±0.01a,A 0.76±0.01a,A
Iota 25 45.19±3.83a,A 0.70±0.10a,A 0.38±0.01b,C 0.77±0.01b,C 0.74±0.00b,C
50 45.81±8.43a,A 0.75±0.13a,A 0.34±0.04c,C 0.75±0.00c,C 0.72±0.02c,C
75 27.87±4.50b,A 0.74±0.77a,A 0.37±0.02c,C 0.75±0.00c,C 0.67±0.02d,C
Kappa 25 41.13±2.17a,A 0.80±0.17a,A 0.34±0.01b,C 0.76±0.01b,C 0.71±0.02b,C
50 29.30±1.52a,A 0.79±0.16a,A 0.31±0.01c,C 0.75±0.00c,C 0.64±0.03c,C
75 39.25±2.93b,A 0.76±0.17a,A 0.26±0.02c,C 0.75±0.03c,C 0.59±0.04d,C
Lambda 25 27.80±1.21a,B 0.66±0.08a,C 0.37±0.03b,B 0.80±0.02b,B 0.75±0.00b,B
50 39.63±0.89a,B 0.67±0.05a,C 0.39±0.01c,B 0.73±0.02c,B 0.75±0.01c,B
75 32.16±2.27b,B 0.70±0.05a,C 0.39±0.03c,B 0.74±0.04c,B 0.74±0.02d,B
a,b,c,d means with same letter in same column are not significantly (p>0.05) different for the percent of milk fat substitution.
A,B,C means with same letter in same column are not significantly (p>0.05) different for the carrageenan type.

6 • E. Rojas-Nery, N. Güemes-Vera, G.O. Meza-Marquez and A. Totosaus
Grasas Aceites 66 (4), October–December 2015, e097. ISSN-L: 0017–3495 doi: http://dx.doi.org/10.3989/gya.0240151
protein isolate and carrageenans helped to increase
water retention and yield as well as protein content.
On the other hand, the associative interaction
between carrageenans and casein micelles depends
on the carrageenan’s conformation. At neutral pH,
kappa and iota carrageenans in helical conforma-
tion can stabilize milk proteins at a low concentra-
tion since the addition of carrageenan affected the
formation of rennet-induced gels, mainly attributed
to electrostatic interactions between kappa casein
the positive patch and negative sulfate groups of
carrageenans (Snoeren et al., 1976;Thaiudomand
Goff,2003; Gu etal., 20 05; Cor redi getal., 2 011; Wa ng
et al., 2014). Iota, kappa and lambda carrageenans
adsorb onto casein micelles forming a cross-linking
network below the coil-helix transition temperature
(60 °C), probably due to the bridging by the helical
parts of carrageenan chains (Dalgleishand Morris,
1998; Langendorff et al., 2000; Gu et al., 2005).
Carrageenan conformation results in differences
FIGURE 1. FTIR spectra for the different panela-type cheeses formulated with emulsified soybean oil/soy
protein isolate and (a) iota-carrageenan, (b) lambda-carrageenan, and (c) kappa-carrageenan.

Carrageenan type effect on soybean oil/soy protein isolate emulsion employed as fat replacer in panela-type cheese • 7
Grasas Aceites 66 (4), October–December 2015, e097. ISSN-L: 0017–3495 doi: http://dx.doi.org/10.3989/gya.0240151
between adsorption behaviors due to different charge
densities. Both charge density (lambda>iota>kappa)
and polysaccharide conformation contribute tothe
structure forming event (Gu etal., 2005; Wangetal.,
2014). Kappa and iota are in the helix form at ambi-
ent temperature, whereas lambda is in the random
coil conformation (Nilsson and Piculell, 1991; Gu
et al., 2005). Kappa carrageenan adsorption onto
casein micelle is thermally reversible, whereas iota-
carrageenan adsorption onto casein micelle is irre-
versible (Ceˇrníková et a l., 2008). The most charged
lambda carrageenan does not show transition from
coil to helix, being that it is a non-gelling carrageenan
(Nilson and Piculell, 1991; Corredig etal., 2011).
4.2. Texture pro le analysis
In general, fat replacement resulted in a harder
and less ductile texture, where higher moisture
retained in the cheese structure (as yield related to
weight gain) and lower fat content affected texture.
Substitution of part of the milk fat with non-milk fat
modified textural properties of the processed cheese
since fat globule size increased with the decrease in
the distribution uniformity within the protein matrix
(Cunha etal., 2010). Cheese structure is altered with
a decrease in fat content, and lower-fat cheese has a
more compact protein matrix with less open spaces
than full-fat cheese since interstitial spaces are occu-
pied by fat globules. When fat content is reduced,
longer areas of uninterrupted protein matrix with
less uniformly dispersed fat globules are formed
(Guinee and McSweeney, 2006). This is associated
with hard texture even when the moisture content
is high (Gunasekaran and Ak, 2000). Fat reduction
in fresh white cheese resulted in more complete pro-
tein zones comprising the protein structure, increas-
ing the degree of protein molecules cross-linking
in the three dimensional network, and increasing
resistance to deformation (Lobato-Calleros et al.,
2007). In same manner, cheese elaborated with milk
fat was more cohesive than the vegetable fat blend
cheese (Dinkçi et al., 2011), and cheese cohesive-
ness decreases with fat content (Gunasekaran and
Ak, 2000). It has been reported that the addition of
palm oil decreased cheese hardness, probably due to
the presence of fatty acids with lower melting point
(Cunha et al., 2010). Some fat mimics have been
found to enhance the uniformity of fat distribution
in reduced-fat cheese (Drake etal., 1996). Differences
in texture could be attributed to the interactions of
hydrocolloids with the rest of the matrix besides the
ability of the given hydrocolloids to form gels (asin
the case of kappa and iota-carrageenan, forming
harder cheeses, followed by lambda) (Hanáková
etal., 2013). An increasing concentration of kappa
and iota carrageenans (more emulsified oil replac-
ing milk fat) enhances interactions between car-
rageenan chains and leads to the formation of a
denser network structure which increases the rigid-
ity of processed cheese (Ceˇrníková etal., 2008). In
panela-type cheese at the experimental conditions
employed, milk fat substitution resulted in a harder
and less cohesive texture.
4.3. Fourier transformation infra-red spectroscopy
analysis
In the MIR spectra of panela-type cheese formu-
lated with different percentages of milk fat substitu-
tion with emulsified soybean oil/soy protein isolate
with iota, kappa or lambda carrageenans, the same
typical absorption bands can be observed but with
different magnitude, reflecting changes in the com-
position of panela-type cheese. Changes in cheese IR
spectra had been employed to determine changes in its
composition, presenting several typical peaks and the
assignments different wavelength ranges for the con-
tributions of the hydroxyl groups, acids, esters, amide I
and amide II, aliphatic chains of fatty acids and acidic
amino acids, at specific regions: 3873–3000cm−1 for
O–H stretching modes of water absorbing, −C−H
stretching in fatty acids (3000–2800 cm−1), −C=O of
acids and esters (1750–1650 cm−1), amide I andamide
II of proteins (1650–1450cm−1), esters and aliphatic
chains of fatty acids (1460–1150cm−1) and C=O and
C−C stretching of acids (1200–800 cm−1), respec-
tively (Cuibus et al., 2014). As can be observed, in
the 1200–950 cm−1 region, bands seem to be closer
with tension vibration of C–O–C bonds, a char-
acteristic vibration attributed to carbohydrate content
(Al-Jowder et al., 1999). The higher the percent of
milk fat substitution (25–75%), the higher the bands’
magnitude. This is a higher amount of C–O–C
functional groups (from iota, kappa or lambda car-
rageenans) incorporatedinto cheese (bythe emul-
sified oilreplacing milk fat). In the same manner,
the absorbance of C–O (~1175 cm−1) and C=O
(~1750cm−1) of the ester bonds of triacylglycerols
and the acyl chain C–H (3000–2800 cm−1) are com-
monly used to determine fat. The infra-red bands
appearing in the 3000–2800cm−1 region are particu-
larly useful because they are sensitive to the content,
the conformation and the packing of the triglycerides
(Casal and Mantsch, 1984; Dufour and Riaublanc,
1997). Since the milk fat replacement with 25% of
emulsified oil spectra is different to that with 75%
of milk fat replacement, the differences in the spec-
tra can be employed to identify carrageenan addition
to cheese.
5. CONCLUSIONS
Fat content influences the volume and continu-
ity of the casein matrix, which is interrupted by fat
globules. When milk fat was replaced by emulsi-
fied soybean oil, structural changes resulted in an
increase in moisture enhanced by the soy protein

8 • E. Rojas-Nery, N. Güemes-Vera, G.O. Meza-Marquez and A. Totosaus
Grasas Aceites 66 (4), October–December 2015, e097. ISSN-L: 0017–3495 doi: http://dx.doi.org/10.3989/gya.0240151
isolate and carrageenan-casein interactions retain-
ing more water (decreasing curd volume due to water
expelling). The panela-type cheese texture resulted
as expected in a harder but less plastic structure, i.e.,
although a harder cheese was obtained, the lower
cohesiveness, springiness and resilient values com-
pensate the texture profile. Milk fat replacement
changed the fatty acid profile, where the type of car-
rageenan also affected the MIR spectra. Lambda
carrageenan containing emulsions improved mois-
ture retention and maintained the textural properties
of panela-type cheese.
ACKNOWLEDGEMENTS
Rojas-Nery thank Conacyt, México for the grant
for his graduate studies.
REFERENCES
Al-Jowder O, Kemsley EK, Wilson RH. 2002. Detection of
adulteration in cooked meat products by MIR-Infrared
Spectroscopy. J. Agric. Food Chem. 50, 1325–1329. http://
dx.doi.org/10.1021/jf0108967.
AOAC, 1998. Official Method of Analysis of AOAC International
(16th Ed.), Washington, DC.
Banks JM. 2004. The technology of low-fat cheese manufacture.
Int. J. Dairy Technol. 57, 199–207. http://dx.doi.org/10.1111/
j.1471-0307.2004.00136.x.
Bourne MC. 1978. Texture profile analysis. Food Technol. 32,
62–66, 72.
Casal HL, Mantsch HH. 1984. Polymorphic phase behavior of
phospholipid membranes studied by infrared spectroscopy.
Biochim. Biophys. Acta 779, 382–401. http://dx.doi.org/
10.1016/0304-4157(84)90017-0.
Ceˇrníková M, Bunˇ kaa F, Pavlínek V, Breˇzina P, Hrabeˇ J, Valáseˇka
P. 2008. Effect of carrageenan type on viscoelastic proper-
ties of processed cheese. Food Hydrocolloid. 22, 1054–1061.
http://dx.doi.org/10.1016/j.foodhyd.2007.05.020.
Corredig M, Sharafbafi N, Kristo E. 2011. Polysaccharide-
protein interactions in dairy matrices, control and design of
structures. Food Hydrocolloid. 25, 1833–1841. http://dx.doi.
org/10.1016/j.foodhyd.2011.05.014.
Cuibus L, Maggio R, Mures¸an V, Diaconeasa Z, Fetea F, Socaciu
C. 2014. Preliminary Discrimination of Cheese Adulteration
by FT-IR Spectroscopy. Bull. UASVM Food Sci. Technol 71,
142–146. http://dx.doi.org/10.15835/buasvmcn-fst:10795.
Cunha CR, Dias AI, Viotto WH. 2010. Microstructure, texture,
colour and sensory evaluation of a spreadable processed
cheese analogue made with vegetable fat. Food Res. Int. 43,
723–729. http://dx.doi.org/10.1016/j.foodres.2009.11.009.
Dalgleish DG, Morris ER. 1988. Interactions between carrageen-
ans and casein micelles: electrophoretic and hydrodynamic
properties of the particles. Food Hydrocolloid. 2, 311–320.
http://dx.doi.org/10.1016/S0268-005X(88)80028-6.
Der G, Everitt BS. 2002. A Handbook of Statistical Analyses Using
SAS. Chapman & Hall, London.
Dinkçi N, Kesenkas¸ H, Seçkin AK, Kinik O, Gönç S. 2011.
Influence of a vegetable fat blend on the texture, micro-
structure and sensory properties of kashar cheese. Grasas y
Aceites 62, 275–283. http://dx.doi.org/10.3989/gya.091810.
Drake MA, Herrett W, Boylston TD, Swanson BG. 1996. Lecithin
improves texture of reduced-fat cheeses. J. Food Sci.
6, 639–642. http://dx.doi.org/10.1111/j.1365-2621.1996.
tb13176.x.
Drohan DD, Tziboula A, McNulty D, Horne D.S. 1997. Milk
protein-carrageenan interactions. Food Hydrocolloid. 11,
101–107. http://dx.doi.org/10.1016/S0268-005X(97)80016-1.
Dufour É, Riaublanc A. 1997. Potentiality of spectroscopic meth-
ods for the characterization of dairy products. II. Mid infra-
red study of the melting temperature of cream triacylglycerols
and of the solid fat content in cream. Lait 77, 671–681. http://
dx.doi.org/10.1051/lait:1997648.
Fathi Achachlouei B, Hesari J, Azadmard Damirchi S,
Peigham bardoust SH, Esmaiili M, Alijani S. 2013.
Production and characterization of a functional Iranian
white brined cheese by replacement of dairy fat with vege-
table oils. Food Sci. Technol. Int. 19, 389–398. http://dx.doi.
org/10.1177/1082013212455341.
Fox PF, Guinee TP, Cogan TM, McSweeney PLH. 2000.
Fundamentals of Cheese Science. Aspen Publishers Inc.,
Gaithersburg.
Giroux HJ, Constantineau S, Fustier P, Champagne CP,
St-Gelais D, Lacroix M, Britten M. 2013. Cheese fortifica-
tion using water-in-oil-in-water double emulsions as car-
rier for water soluble nutrients. Int. Dairy J. 29, 107–114.
http://dx.doi.org/10.1016/j.idairyj.2012.10.009.
Gu YS, Decker EA, McClements DJ. 2005. Influence of pH and
carrageenan type on properties of b-lactoglobulin stabi-
lized oil-in-water emulsions. Food Hydrocolloid. 19: 83–91.
http://dx.doi.org/10.1016/j.foodhyd.2004.04.016.
Guinee TP, McSweeney PLH. 2006. Significance of milk fat in
cheese, in Fox PF, McSweeney PLH (Ed.) Advanced Dairy
Chemistry, Volume 2: Lipids, 3rd ed, Springer, New York,
pp. 377–440.
Gunasekaran S, Ak MM. 2000. Cheese Rheology and Texture.
CRC Press, Boca Raton.
Hanáková Z, Bunˇka F, Pavlínek V, Hudecˇková L, Janiš R. 2013.
The effect of selected hydrocolloids on the rheological
properties of processed cheese analogues made with veg-
etable fats during the cooling phase. Int. J. Dairy Technol.
66, 484–489. http://dx.doi.org/10.1111/1471-0307.12066.
ISO, 2011. ISO 3433:2008: Cheese- Determination of fat content-
Van Gulik method. International Organisation for
Standardisation, Geneva.
Johnson ME, Kapoor R, McMahon DJ, McCoy DR,
Narasimmon RG. 2009. Reduction of sodium and fat levels
in natural and processed cheeses: scientific and technological
aspects. Compr. Rev. Food. Sci. F. 8, 252–268. http://dx.doi.
org/10.1111/j.1541-4337.2009.00080.x.
Kesenkas H, Dinkçia N, Kemal Seçkin A, Kinika Ö, Gönça S.
2009. The effect of using a vegetable fat blend on some
attributes of kashar cheese. Grasas Aceites 60, 41–47.
http://dx.doi.org/10.3989/gya.032408.
Kinsella JE. 1979. Functional properties of soy proteins. J. Am.
Oil Chem. Soc. 56, 242–258. http://dx.doi.org/10.1007/
BF02671468.
Langendorff V, Cuvelier G, Launay B, Michon C, Parker A, de
Kruif CG. 1999. Casein micelle/iota-carrageenan interactions
in milk: Influence of temperature. Food Hydrocolloid. 13,
211–218. http://dx.doi.org/10.1016/S0268-005X(98)00087-3.
Langendorff V, Cuvelier G, Michon C, Launay B, Parker A, de
Kruif CG. 2000. Effects of carrageenan type on the behav-
iour of carrageenan/milk mixtures. Food Hydrocolloid. 14,
273–280. http://dx.doi.org/10.1016/S0268-005X(99)00064-8.
Livingstone KM, Lovegrove JA, Givens DI. 2012. The impact of
substituting SFA in dairy products with MUFA or PUFA
on CVD risk: evidence from human intervention studies.
Nutrit. Res. Rev. 25, 193–206. http://dx.doi.org/10.1017/
S095442241200011X.
Lobato-Calleros C, Ramírez-Santiago C, Osorio-Santiago VJ,
Vernon-Carter EJ. 2002. Microstructure and texture of
manchego cheese-like products made with canola oil,
lipophilic, and hydrophilic emulsifiers. J. Texture Stud.
33, 165–182. http://dx.doi.org/10.1111/j.1745-4603.2002.
tb01343.x.
Lobato-Calleros C, Reyes-Hernández J, Beristain CI, Hornelas-
Uribe Y, Sánchez-García JE, Vernon-Carter EJ. 2007.
Microstructure and texture of white fresh cheese made
with canola oil and whey protein concentrate in partial or
total replacement of milk fat. Food Res. Int. 40, 529–537.
http://dx.doi.org/10.1016/j.foodres.2006.10.011.
Lobato-Calleros C, Velázquez-Varela J, Sánchez-García J, Vernon-
Carter EJ. 2003. Dynamic rheology of Mexican manchego
cheeselike products containing canola oil and emulsifier
blends. Food Res. Int. 36, 81–90. http://dx.doi.org/10.1016/
S0963-9969(02)00111-4.

Carrageenan type effect on soybean oil/soy protein isolate emulsion employed as fat replacer in panela-type cheese • 9
Grasas Aceites 66 (4), October–December 2015, e097. ISSN-L: 0017–3495 doi: http://dx.doi.org/10.3989/gya.0240151
Molina Ortiz SE, MC Puppo, JR Wagner. 2004. Relationship
between structural changes and functional properties of soy
protein isolates–carrageenan systems. Food Hydrocolloid. 18,
1045–1053. http://dx.doi.org/10.1016/j.foodhyd.2004.04.011.
Nilsson S, Piculell L. 1991. Helix-coil transitions of ionic polysac-
charides analysed within the Poisson-Boltzmann cell model.
4. Effects of site specific counterion binding. Macromol. 24,
3804–3811. http://dx.doi.org/10.1021/ma00013a010.
Romeih EA, Michaelidou A, Biliaderis CG, Zerfiridis GK.
2002. Low-fat white-brined cheese made from bovine milk
and two commercial fat mimetics: chemical, physical and
sensory attributes. Int. Dairy J. 12, 525–540. http://dx.doi.
org/10.1016/S0958-6946(02)00043-2.
Sahan N, Yasar K, Hayaloglu AA, Karaca OB, Kaya A. 2008.
Influence of fat replacers on chemical composition, pro-
teolysis, texture profiles, meltability and sensory proper-
ties of low-fat Kashar cheese. J. Dairy Res. 75, 1–7. http://
dx.doi.org/10.1017/S0022029907002786.
SIAP. 2014. Sistema de Información Agroalimentaria y Pesquera,
Boletín de Leche octubre-diciembre 2014. SAGARPA,
México. Available at URL: http://www.siap.gob.mx/wp-
content/uploads/boletinleche/boletLecheOct-dic_2014.pdf.
Snoeren THM, Paynes TAJ, Jeunink J, Both P. 1975. Elec-
trostatic interaction between κ-carrageenan and κ-casein.
Milchwissenschaft. 30, 391–396.
Szczceniak AS. 1963. Classification of textural characteristics.
J. Food Sci. 28, 385–389. http://dx.doi.org/10.1111/j.1365-
2621.1963.tb00215.x.
Thaiudom S, Goff HD. 2003. Effect of kappa-carrageenan on milk
protein polysaccharide mixtures. Int Dairy J. 13, 763–771.
http://dx.doi.org/10.1016/S0958-6946(03)00097-9.
Uruakpa FO, S.D. Arntfield. 2005. Emulsifying characteris-
tics of commercial canola protein–hydrocolloid systems.
Food Res. Int. 38, 659–672. http://dx.doi.org/10.1016/j.
foodres.2005.01.004.
Wang F, Liu X, Hu Y, Luo J, Lv X, Guo H, Ren F. 2014. Effect
of carrageenan on the formation of rennet-induced casein
micelle gels. Food Hydrocolloid. 36, 212–219. http://dx.doi.
org/10.1016/j.foodhyd.2013.10.004.
Wang T. 2002. Soy bean oil, in Gunstone FD (Ed.) VegetableOils
in Food Technology. Blackwell Publishing Co., London,
18–58.
Ye A, Cui J, Taneja A, Zhu X, Singh H. 2009. Evaluation of
processed cheese fortified with fish oil emulsion. Food Res.
Int. 42, 1093–1098. http://dx.doi.org/10.1016/j.foodres.2009.
05.006.
Yu L, Hammond EG. 2000. Production and characterization of
a Swiss cheese-like product from modified vegetable oils.
J.Am. Oil Chem. Soc. 77, 917–924. http://dx.doi.org/10.1007/
s11746-000-0145-y.