Received: 16 October 2020 Revised: 7 May 2021 Accepted: 9 May 2021

DOI: 10.1111/1750-3841.15799

CONCISE REVIEWS & H YPOTH ESES IN FOOD SCIENCE

How does milk fat replacement influence cheese analogue

microstructure, rheology, and texture profile?

Valeria Giha María José Ordoñez Ruby Alejandra Villamil

Science Faculty, Pontificia Universidad

Javeriana, Bogotá, Colombia
Correspondence
Ruby Alejandra Villamil, Science Faculty,
Pontificia Universidad Javeriana, Bogotá
110111, Colombia.
Email: villamil.r@javeriana.edu.co
Abstract: Over the past few years, the market for cheese substitutes has been
growing on account of the simple and cost-effective production of these cheese-
like products. It is well established that the functional properties of cheeses are
directly related to their composition. Therefore, the variation of fat in cheese sub-
stitutes certainly affects the characteristics of the cheeses. The purpose of this
review was to summarize the latest research on the effects of milk fat replace-
ment with vegetable oils on the rheological, textural, and microstructural prop-
erties of cheese analogues. The findings suggest that the primary effects of modi-
fying fat in cheese analogues are associated with an alteration in the interactions
among the components of the protein matrix, which varies because of milk fat
extraction. Overall, changes in the functional properties of analogous cheeses
will depend on the type of oil, the percentage of fat modification, and the type of
cheese produced.

KEYWORDS
cheese matrix, cheese microstructure, fat globule size, rheological behavior, texture parameters

1 INTRODUCTION One of the challenges of these analogues is to exhibit

Cheese analogues are products intended to replace tradi-
tional cheeses, in which the fat and/or protein of the milk
is replaced by nondairy alternatives (Fox et al., 2017). The
market for these cheese-like products has been growing
because they are profitable and easy to produce compared
to traditional cheeses (Transparency Market Research,
2020). The formulation of cheese analogues has been pro-
moted to meet consumer demands (Ferrão et al., 2016). The
milk fat substitution with vegetable oils is a trend in the
manufacture of cheese-like products (Bermúdez-Aguirre
& Barbosa-Cánovas, 2011; Calligaris et al., 2013; Hughes
et al., 2012; Villamil et al., 2021) since it can improve
the fatty acid profile of the final cheese, contributing to
a healthier ratio between saturated and unsaturated fats
ratio (Achachlouei et al., 2013; Villamil et al., 2020). How-
ever, the increase of the nutritional quality implies the
incorporation of food sources that represent a high cost of
the final product (Fox et al., 2017).
similar functionality of the original cheese because an
alteration in the fat profile of the cheese may affect its phys-
ical properties and stability (Awad et al., 2014; Ennis &
Mulvihill, 2001; Ehsannia & Sanjabi, 2016; Kaminarides &
Stachtiaris, 2000). Among the technological properties of
cheeses, rheological, textural, and microstructural charac-
teristics are fundamental to increase the versatility of the
end use applications of the cheese analogues (Hennelly
et al., 2005; Liu et al., 2008; Mounsey & O’Riordan, 2001;
Montesinos-Herrero et al., 2006). Hence, the formulations
of cheese analogues always tend drastically to avoid modi-
fications of the original technological characteristics of the
cheese after replacing the milk fat with vegetable oil; there-
fore, this will depend on the type of cheese and its charac-
teristics, as well as the intended purpose of the end product
(Kapoor & Metzger, 2008).
The concept of ‘‘rheological properties’’ for cheeses
refers to the capacity of cheese to maintain its inte-
grated structure (Fox et al., 2017). It is of considerable

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R wileyonlinelibrary.com/journal/jfds J. Food Sci. 2021;86:2802–2815.
PHYSICAL PROPERTIES OF CHEESE ANALOGUES
importance since it can affect the degree (or length)
of chewing, which may affect the product acceptability
(O’Callaghan & Guinee, 2004). Some studies have demon-
strated that modification of fat can result in a signifi-
cantly higher melting area (Totosaus et al., 2017). Addi-
tionally, the incorporation of emulsified vegetable oils into
the matrix may affect the amount and distribution of fat
droplets in the protein network, leading to changes in
the microstructure and textural behavior of the cheese
(Lobato-Calleros et al., 2002, 2003). One study reported
that fat modification resulted in a harder cheese with
decreased fracturability (Bermúdez-Aguirre & Barbosa-
Cánovas, 2011), which may contribute to diminished con-
sumer acceptability. Overall, fat is important for taste,
appearance, and texture of foods, contributing to adjust-
ment of solids, impact of particle size, consistency, color,
and proper balance of flavor characteristics (Lucca & Tep-
per, 1994). As shown, different studies have revealed how
the total or partial replacement of milk fat by vegetable oils
can affect the physical properties of cheeses. For this rea-
son, this work aims to assess how the rheological, textural,
and microstructure properties of cheese analogues may be
affected by the substitution of milk fat with vegetable oils.
2 GENERAL VIEW OF CHEESE
ANALOGUES
Cheese analogues can be arbitrarily classified into three
categories: cheese analogues, filled cheeses, and tofu-
based cheeses. They can also be classified based on the
ingredients used in their formulation and manufactur-
ing processes, as partially dairy or non-dairy, depend-
ing on whether the fat and/or protein components are
derived from dairy or vegetable sources (Fox et al., 2017).
Cheese analogues are formulated and produced with spe-
cific nutritional and/or functional properties, according to
the market and consumer needs (Bachmann, 2001). The
main advantage of these cheese-like products is their versa-
tility, as these substitute products can be designed to meet
special dietary needs through changes in their formulation
(e.g., lactose free, low in calories, low in saturated fat and
cholesterol and even vitamin and mineral enriched).
Cheese analogues are intended to replace traditional
cheeses; hence, they should not be nutritionally inferior
to cheese. Promoters of cheese-like products claim nutri-
tional advantages over genuine cheeses. Some researchers
have addressed the use of unsaturated fat in dairy cheese
in order to improve the lipid profile of the cheeses (Living-
stone et al., 2012). However, there is a growing scientific
consensus in favor of milk fat due to its specific physio-
logical functions (Gómez-Cortés et al., 2018). The presence
of medium- and short-chain fatty acids provides energy
2803
to different vital organs such as muscles, heart and liver,
among others (Miciński et al., 2012). Also, it has been con-
ferred preventive properties against certain diseases such
as ulcerative colitis and cancer in addition to having anti-
inflammatory and antibacterial effects (Tan et al., 2014). In
spite of these properties, the use of unsaturated fats has
shown to offer certain health advantages such as the pre-
vention of oxidative modification of low-density lipopro-
tein (LDL) in the vascular endothelium (Aguilera et al.,
2001). It has been demonstrated that a higher intake of sat-
urated fatty acids can be reflected in a significant increase
in LDL and total cholesterol compared to a diet rich in
food sources of unsaturated fatty acids (Briggs et al., 2017).
In addition, by substituting milk fat for vegetable oils, the
analogues are nearly cholesterol free (Bachmann, 2001).
The substitution and/or fortification of cheeses with the
incorporation of vegetable oils seems to be useful in the
prevention of different non-transmissible diseases while
representing a competitive advantage in the food indus-
try. The substitution of milk fat or milk protein in an ana-
logue cheese is not the same as committing fraud or adul-
teration of the product since any change in the composi-
tion of the milk is reported on the packaging of the cheese
analogue or substitute. Milk is often subject of fraud in
order to obtain economic benefits such as adding water to
increase the volume of the milk and thus sell it at a higher
price with a lower nutritional value (Handford et al., 2016).
In the case of some cheese analogues, milk components
are often replaced by other products with the objective of
providing a nutritional benefit to consumers. However, as
mentioned above, changing the matrix can lead to varia-
tions in the technological and instrumental characteristics
of the cheese. In this paper, we discuss the different effects
of modifying milk fat by vegetable oil on analogue cheeses.
3 IMPORTANCE OF FAT
REPLACEMENT IN CHEESE ANALOGUES
Today, a wide variety of cheese (e.g., fresh cheese, matured
cheese, spreadable cheese, and processed cheese, among
others) is used as ingredients for cooking and recipes. The
technological properties of these cheeses will depend on
their end use application. For example, a cheese with excel-
lent melting ability and springiness is desirable for pizza
(Guinee, 2016); but on the contrary, for other food prepara-
tions such as Palak paneer, a high degree of meltability may
be undesirable. Each cheese variety has properties that
depend on the manufacturing procedures, the treatment of
the milk, and the manufacturing technique (Lamichhane
et al., 2018). In addition, the cheese composition, such as
its dry matter, fat, protein, and calcium content, is a deter-
mining factor in its behavior (Mehta, 2018). Therefore, at
2804
the moment of developing an analogue cheese by replac-
ing milk fat with vegetable oil, one must be considering
how the change in the composition of the traditional prod-
uct can affect its technological properties and thus its final
use.
Fat is a fundamental component of cheese. Cheese ana-
logues made with altered fat contents may be unsuitable
in terms of overall quality (Rogers et al., 2010). Fat replace-
ment can impact the ripening biochemistry, microstruc-
ture and sensorial properties, due to the fatty acid com-
position, mainly unsaturated fatty acids (Moatsou et al.,
2019). Additionally, the texture defects are the most com-
monly described in low-fat cheeses (Lobato-Calleros et al.,
1997). Suitable vegetable oils are the main source of bioac-
tive lipids; hence, these oils have been incorporated at dif-
ferent levels as substitutes for milk fat in a variety of cheese
analogues (e.g., Table 1). Further, the microstructure, fol-
lowed by rheological properties and texture profile analysis
(TPA), is the primary variable to study.
3.1 Microstructure
Cheese microstructure is a complex matter composed of
a protein matrix that entraps fat, water, minerals, and
bacterial colonies and their metabolites (Burdikova et al.,
2015). Microstructure is a fundamental aspect in the devel-
opment of technological and textural properties of the
cheeses (Everett & Auty, 2017) for which it is necessary to
fully understand how the components of the cheese inter-
act with each other (Mehta, 2018).
Cheese microstructure varies according to composi-
tional factors, coagulants (e.g., enzymes, lactic acid bacte-
ria or direct acidification), degree of ripening, and storage
conditions (Mehta, 2018; Watkinson et al., 2001). For exam-
ple, in acid coagulation cheese, fat globules are usually less
coalesced compared to rennet curd cheese. For acid curd,
the food matrix is composed of casein (instead of calcium
phosphate paracasein), showing a lower melting degree
and a lower volume fraction of the casein particles due
to the high moisturize and decreased casein concentration
(Fox et al., 2017). On the other hand, rennet curd cheese has
a matrix composed of a hydrated calcium phosphate para-
casein network that occludes the fat phase which occurs as
coalesced globules (Guinee, 2016).
The evaluation of microstructure includes the assess-
ment of the fat globule size and distribution and the
type of crystals present in the casein network (Fox et al.,
2017; Kapoor & Metzger, 2008). Different microscopic tech-
niques have been used for this purpose, being confocal
laser scanning microscopy (CLSM) and scanning electron
microscopy (SEM) the most frequently used (El-Bakry &
Sheehan, 2014).
PHYSICAL PROPERTIES OF CHEESE ANALOGUES
As can be observed in Table 2, the findings showed that
cheese analogues with milk fat substitution with vegetable
oils have an increase in fat globule size (Abd El-Wahed &
Hassanien, 2019; Cunha et al., 2010) and a decrease in the
number of globules present (Cunha et al., 2013; Lee et al.,
2015; Leong et al., 2020) in a less uniform distribution (Abd
El-Wahed & Hassanien, 2019; Cunha et al., 2010). In con-
trast, some studies have shown a decrease in the size of
the fat globules (Cunha et al., 2013; Leong et al., 2020).
Although there is limited information regarding pores in
cheese analogues, very few of the revised studies show data
related to this aspect. These studies showed that cheese
analogues with partial or total milk fat substitution pre-
sented a higher number of pores in the cheese (Ramel &
Marangoni, 2018) and an increase in the size of the pores
compared to traditional cheeses (Leong et al., 2020).
3.1.1 Fat globules
Some studies have evaluated the effects of replacing milk
fat with vegetable oils regarding fat globule size and dis-
tribution (Abd El-Wahed & Hassanien, 2019; Cunha et al.,
2010, 2013; Lee et al., 2015; Leong et al., 2020). Overall, fat
globules are mostly spherical and evenly distributed in the
protein matrix. The matrix is affected when fat is modified,
either by replacement or fortification. For example, some
cheese analogues have shown an increase in the diameter
of their fat globules, which could affect the integrity of the
curd (Everett & Olson, 2003). Among the selected studies,
as shown in Table 2, it was possible to observe changes
related to this same aspect. In processed cheese spread
analogues with 50% or 100% substitution of milk fat with
peanut cream, through direct oil inclusion, the number of
fat globules increased, as well as their diameter, and they
were less evenly distributed (Abd El-Wahed & Hassanien,
2019). Cunha et al. (2013) showed similar results with a
model of requeijão cremoso spreadable cheese analogue
with a 23% fat replacement through direct oil inclusion of
partially hydrogenated soybean fat or soybean oil, which
showed an increase in diameter as well. These character-
istics differ from traditional cheeses which generally show
numerous small particles of fat dispersed in a uniform pro-
tein network (Cunha et al., 2010). However, some studies
presented different results showing, instead, a decrease in
the size of the fat globules. Leong et al. (2020) showed that
in two cheddar-like analogues with partial substitution of
milk fat with canola oil, very small fat droplets distributed
in the matrix were observed. This result can be related
to the oil inclusion method used since, in the aforemen-
tioned studies (Abd El-Wahed & Hassanien, 2019; Cunha
et al., 2010, 2013), the researchers used direct oil inclusion,
whereas in this study, an emulsion made by ultrasound
TA B L E 1 Summary of studies assessing texture, rheology and/or microstructure of cheese analogues with substitution of milk fat by vegetable oils
Type of cheese Milk fat replacer Fat replacement Assessed parameters Reference
Cheddar cheese analogue Canola oil 100% Rheological properties and TPA (Leong et al., 2020)
Kashar cheese analogue Palm oil 0%, 50%, and 100% TPA (Kavak & Karabiyik,
2020)
PHYSICAL PROPERTIES OF CHEESE ANALOGUES

Processed cheese spread Peanut cream 50% and 100% Microstructure and TPA (Abd El-Wahed &
analogues Hassanien, 2019)
Model processed cheese Canola oil 26% Microstructure, rheological (Ramel & Marangoni,
imitation product properties and TPA 2018)
Model processed cheese Sunflower oil 0%, 10%, 20%, 30%, and Microstructure and rheological (Lee et al., 2015)
analogue 40% properties
Model processed cheese Coconut fat and palm oil 99% Rheological properties and TPA (Hanáková et al., 2013)
analogues
Requeijão cremoso spreadable Partially hydrogenated soybean fat 23.2% Microstructure, rheological (Cunha et al., 2013)
cheese analogue and soybean oil properties, and TPA
Requeijão cremoso spreadable Hydrogenated vegetable fat 25% and 50% Microstructure and TPA (Cunha et al., 2010)
cheese analogue
Oaxaca type cheese analogue Soya bean oil 0%, 25%, 50%, and 75% Rheological properties and TPA (Totosaus et al., 2017)
White Turkish cheese Corn oil 0%, 50%, 60%, 100%, TPA (Arslan et al., 2014)
analogue and
Kashar cheese analogue Palm oil 0% and 100% Microstructure, rheological (Dinkçi et al., 2011)
properties, and TPA
White Turkish cheese Interesterified fat (palm, palm 0%, 50%, 60%, 100%, TPA (Arslan et al., 2010)
analogue kernel, and soybean oil) and 100%
Abbreviation: TPA, texture profile analysis.
2805
2806
TA B L E 2 Results of microstructure of cheese analogues manufactured with different vegetable oils
Pores Number of Sphericity of
diameter pores oil droplets Coalescence globules
Increased – Increased Increased
– – Decreased – Decreased
– Increased – –
– – – No change
– – – –
– – – –
– – – –
was applied that could modify the size of the fat drops in
the matrix thus making them smaller. The findings corre-
late with what was mentioned by Lobato-Caballeros et al.
(1997) where it was explained that the diameter of the fat
globules is related to the shear forces applied during the
cheese analogue preparation and the fatty acid composi-
tion of the used oil. This explained how the elaboration of
a previous emulsion by ultrasound decreased the diame-
ter of the fat globules in comparison with the direct incor-
poration. However, fat globules in a pasta filata Kashar
cheese analogue with total replacement of milk fat with
palm oil with a direct oil inclusion method also presented
smaller and more uniform fat globules (Dinkçi et al., 2011).
Nevertheless, it must be highlighted that the homogeniza-
tion process performed after the direct inclusion (wherein
the milk was heated and homogenized at 55–60◦ C and 120
bars) can again be related to the shear forces present at the
time of production.
Furthermore, findings in a study presented by Lee et al.
(2015) showed that the samples of a processed cheese
spread analogue with a lower percentage of fat (10%–20%)
had small and regular shaped fat globules scattered in the
protein matrix. In contrast, samples with a higher fat per-
centage (30%–40%) had very large fat globules (some of
them larger than 20 mm) interspersed with smaller ones.
This may be related to the process of coalescence: a higher
percentage of fat led to a greater number of fat globules.
Thus, the globules are more likely to coalesce and form
larger due to their proximity to each other.
3.1.2 Pore microstructure
A pore is a structural unit defined as an open space
between many protein links on the microstructure surface
of cheeses (El-Bakry & Sheehan, 2014). These areas corre-
spond to regions where air holes are present (Ong et al.,
2012). Some of the parameters related to pore microstruc-
PHYSICAL PROPERTIES OF CHEESE ANALOGUES
Fat globule
Number of fat Fat globule uniform
size distribution References
– Decreased – (Leong et al., 2020)
Increased Decreased (Abd El-Wahed &
Hassanien, 2019)
– – – (Ramel &
Marangoni, 2018)
– Decreased No change (Lee et al., 2015)
– Increased Increased (Cunha et al., 2013)
Decreased Increased Decreased (Cunha et al., 2010)
– Decreased Increased (Dinkçi et al., 2011)
ture are pore number, pore area, and pore diameter
(Impoco et al., 2011). As observed in Table 2, the reviewed
articles evaluated two parameters related to pores in the
microstructure of cheese analogues. A study conducted by
Ramel and Marangoni (2018) showed an increase in the
number of pores after replacing 26% of the milk fat with
canola oil for a model processed cheese. The reduction
or complete removal of the fat from the milk increases
the number of air spaces in the cheese matrix, which
represents the fat globules that were removed during the
processing of the cheese analogues (Cunha et al., 2010).
Regarding the size of pores, Leong et al. (2020) reported
that pore diameters increased in a cheddar cheese ana-
logue with full-fat (100%) replacement with canola oil,
compared to a traditional cheddar cheese. This result can
again be related to the space left by the milk fat glob-
ules when removed from the matrix. Lobato-Calleros et al.
(2007) show that the formation of numerous spaces in the
protein network was originally occupied by oil drops from
the extracted milk fat and this occurrence allowed the for-
mation of a very porous and open protein structure of
the cheese analogues (Ramel & Marangoni, 2018; Leong
et al., 2020). Despite the importance of the microstruc-
ture, findings are very limited and show variability because
each article presented a different type of cheese which can
modify the characteristics of the microstructure as can be
seen in Table 2. As expected, the different types of cheese
show changes that correspond to variations in the protein
matrix. According to the results, for spreadable type ana-
logues the size of the fat globules tends to increase, while
in more solid analogues such as Cheddar, Kashar and pro-
cessed cheeses, the size of the globules decreases.
3.2 Rheology
In food science, rheology is applied to evaluate the
deformation and flow of food materials. Rheological
PHYSICAL PROPERTIES OF CHEESE ANALOGUES
TA B L E 3 Results of rheology of cheese analogues manufactured with different vegetable oils
Melting
Spreadability Meltability point Elasticity
Decreased Decreased Increased Decreased
– Decreased – –
– – – Decreased LF,
increased HF
– – – Increased CF,
decreased PO
Decreased Decreased – –
– – – Decreased A, no
change B
– Increased – –
– Decreased – –
Notes: A. 25% milk fat replacement; B. 50% milk fat replacement.
Abbreviations: CF, coconut fat; HF, high frequencies; LF, low frequencies;; PO, palm oil treatment.
measurements are used to define the consistency of certain
products in terms of viscosity and springiness, attributes
that can determine the flow behavior of the material
(Muthukumarappan & Swamy, 2017). In the case of the
meltability characterization, the Schreiber test is the test
most applied (Kapoor & Metzger, 2008). For viscoelastic
properties, the most commonly utilized are the Arnott test,
Schreiber melt test, and Dynamic stress rheometry (DSR).
The rheological properties of a product allow one to con-
trol its quality during processing as they are closely related
to the textural, sensorial, and microstructural attributes of
foods (Muliawan & Hatzikiriakos, 2007). The direct rela-
tionship of these technological properties lies in the fact
that the rheology and texture of cheese depends on its
structure and internal environment, such as its pH and
temperature (O’Callaghan & Guinee, 2004). Therefore, the
study of the microstructure provides an approach to the
rheological behaviors of cheeses since it represents the spa-
tial distribution of the components (casein, minerals, fat,
moisture and dissolved solutes such as sucrose, lactic acid,
soluble salts, and peptides) and the level of intra- and inter-
molecular attractions (Fox et al., 2017).
Rheology represents the properties associated with mas-
tication that can influence the sensory perception of
cheeses and its suitability for different consumers. There-
fore, changes in the rheological characteristics of a prod-
uct may affect consumers’ acceptance. These aspects of the
analogue cheeses alert the manufacturer to the needs of
the consumer since the desired rheological properties must
be imitated in the analogues in order to be accepted in/by
the food industry (Guinee, 2011). The rheological charac-
terization of cheeses is performed by subjecting the prod-
ucts to stress so as to observe their deformation behavior.
Depending on their deformation properties, they can be
2807
Viscosity Flow behavior References
– – (Leong et al., 2020)
– – (Ramel & Marangoni,
2018)
Increased LF, No change (Lee et al., 2015)
decreased HF
– – (Hanáková et al., 2013)
Increased No change (Cunha et al., 2013)
– – (Cunha et al., 2010)
– – (Totosaus et al., 2017)
– – (Dinkçi et al., 2011)
classified as ideal elastic solids, ideal viscous liquids, or vis-
coelastic (Fox et al., 2017). Cheese is generally classified as a
viscoelastic material because its stress–strain ratio exhibits
both solid and liquid behavior (Guinee, 2011).
The rheological properties of cheese analogue can be
observed in the studies presented in Table 3, where the
results show that major samples solidify at higher tempera-
tures than traditional cheeses (Cunha et al., 2013; Lee et al.,
2013). A cross-over of G′′ (viscous modulus) and G′ (elas-
tic modulus) can also be observed where the frequency
decreased as fat concentration increased. This behavior
had been previously observed (Guinee & O’Kennedy, 2009;
Gliguem et al., 2011; Lee et al., 2013; Wang et al., 2011).
Regarding meltability, most studies show that when milk
fat is replaced by vegetable fat, there is a tendency to a
decreased meltability (Cunha et al., 2013; Leong et al.,
2020; Ramel & Marangoni, 2018). On the contrary, one of
the studies reported an increase in meltability after milk
fat replacement (Totosaus et al., 2017). Considering the
results presented above, one must take into account how
the total or partial substitution of milk fat by vegetable
oils can affect the rheological characteristics of cheeses to
develop analogues that conserve traditional technological
properties.
3.2.1 Viscoelastic properties
Processed cheeses have viscoelastic properties that can be
modified according to their composition, and especially by
their fat content (Fox et al., 2017). In a study conducted
by Lee et al. (2015), various models of processed cheese
spreads with different concentrations of milk fat replace-
ment with sunflower oil (0%–40%), added through direct
2808
incorporation, were produced. In this study, the viscoelas-
tic properties were evaluated using a frequency sweep
model, which is a widely used test as it allows one to deter-
mine the viscoelastic properties of a sample as a function of
time. In this test, high frequencies are used to imitate fast
motion on short time scales, while low frequencies simu-
late slow motion on long time scales or at rest. This allows
the determination of G′′ and G as a function of frequency
(Janmey et al., 2007). Findings show that increasing fat
concentrations develop a higher G′′ and lower G′ at low
frequencies and lower G′′ with higher G′ at high frequen-
cies for all samples (Lee et al., 2015). This is related to the
fact that protein interactions in the cheese matrix can be
modified after a heating process, leading to changes in vis-
cosity (Guinee & O’Kennedy, 2009; Lee & Anema, 2009;
Lee et al., 2013). The same behavior was present in another
investigation (Cunha et al., 2013) where a requeijão cre-
moso cheese spread analogue was developed by replacing
23.3% of its milk fat with hydrogenated soybean fat or soy-
bean oil. The results showed that the type of oil signifi-
cantly influenced all the parameters studied. At the higher
frequencies, a typical solid behavior could be observed
because of inefficient breaking of intermolecular links of
protein over time, while at lower frequencies, the molec-
ular chains had more chance to undergo rearrangement
and flow (Cunha et al., 2013; Lee et al., 2015). The rheolog-
ical behavior of the samples studied is potentially caused
by a high degree of emulsification that is characterized by
a decrease in diameter and an increase in the number of fat
globules, thereby producing a larger surface area and more
protein–protein bonds in the matrix (Guinee, 2007). This
is due to the nature of the emulsion of the oil in water in
which the fat globules are covered with casein micelles and
behave like pseudo proteins, incorporating themselves into
the structure of the protein matrix (Guinee, 2003; Michal-
ski et al., 2002, 2004). These data are consistent with other
studies that show analogue samples solidify at higher tem-
peratures (Guinee & O’Kennedy, 2009; Lee et al., 2013).
Reports have shown that removing the fat from the casein
matrix generates a larger paracasein network, thus causing
an increase in the viscosity of the cheese emulsion system
(Urgu et al., 2019). It has also been observed that the size
of the fat globules within the cheese matrix can determine
the elasticity of the cheese, being more elastic with smaller
fat globules (Michalski et al., 2004). In the study conducted
by Cunha et al. (2010), a significant decrease in the elastic-
ity of the analogue cheese with 25% replacement of milk
fat by vegetable oil was observed, which can be related to
the increase in the size of fat droplets, as shown in Table 3.
However, when the replacement of milk fat by vegetable
oil was increased to 50%, the elasticity did not show signif-
icant differences from the traditional cheese, which may
indicate that this characteristic not only depends on the
PHYSICAL PROPERTIES OF CHEESE ANALOGUES
size but also on the uniform distribution of the fat droplets.
Since this uniform distribution decreased after the replace-
ment of milk fat, it allowed the presence of areas with
protein interaction that favored the elasticity of the ana-
logue cheese (Pereira et al., 2001). Moreover, in a study
conducted by Hanáková et al. (2013), in analogous mod-
els of processed cheese with a 99% substitution of milk fat
by coconut fat or palm oil, an increase in the G′′ modulus
was observed for all samples, with an increasing tempera-
ture during cooling; the lowest G′ values were found in the
products with palm oil. This could be related to the pres-
ence of fatty acids with lower melting points, mainly oleic
acid (Cunha et al., 2010).
3.2.2 Meltability
Cheese meltability is a very important functional prop-
erty for those cheeses that are used as an ingredient in
heat treated meals, such as pizza, cheeseburgers, lasagna,
cheese sticks, and breads (Altan et al., 2005). Their melting
capacity is determined by the composition of the cheese
itself, its dry matter, its fat, protein, and calcium content.
The melting properties can vary among different types of
cheeses, as they have different production methods and
end-use applications (Fox et al., 2017). The reviewed stud-
ies presented highly variable results. Most findings showed
a decrease in the melting capacity of the cheese analogues
with milk fat replacement. Furthermore, only one showed
an increase in the melting capacity of the cheese analogues
(Totosaus et al., 2017). It should be noted that all the stud-
ies used a modified Schreiber fusion test to evaluate melta-
bility as a functional property, thus the variability of the
results was not due to the type of study performed.
Overall, the authors described a decrease in the melta-
bility that is attributed to different causes. One of them is
the incorporation of vegetable oil by means of an emul-
sion whose composition includes protein as an emulsi-
fier. To carry out the substitution of milk fat with vegetal
oils, emulsions are used to avoid the separation phase. For
example, in the study conducted by Leong et al. (2020),
a double emulsion and a single emulsion were prepared
using an ultrasound method to incorporate canola oil into
the milk matrix in order to replace milk fat. This method
allows preparing nanoemulsions that are characterized by
having fat globules of nanometric sizes which lead to a
greater stability. The emulsions prepared in the study were
composed of casein which could increase the interactions
among/within the protein network, thus decreasing the
melting capacity of the cheese analogues. This possibility
may be related to the function that fat globules have in
the cheese matrix as described by different authors, where
it is established that fat reduction allows a higher degree
PHYSICAL PROPERTIES OF CHEESE ANALOGUES
of fusion between the casein chains during gel formation,
which affects the fusion capacity of the product by creating
rigid protein bonds (Fox et al., 2017; Gunasekaran & Ak,
2002). The fat globules present in the protein network of
the cheese matrix act as fillers, which inhibit the binding of
the casein chains, which then increases their fusion capac-
ity (Gunasekaran & Ak, 2002). Accordingly, one of the fac-
tors that may be associated with the decrease in meltabil-
ity is the structure of the matrix. Likewise, the homoge-
nization of the vegetable oil after its inclusion can decrease
the size of the fat droplets in the milk matrix of analogue
cheeses resulting in a greater amount of protein interac-
tions and therefore a smaller meltability as was observed in
Cunha et al. (2013) whereby the vegetable oil was homoge-
nized with a homogenizer-grinder after its direct inclusion.
A further important aspect when replacing the milk fat
in cheese is the melting point of the oil involved, as this
can change the characteristics of the final product when
subjected to heat. Dinkçi et al. (2011) replaced 100% of the
milk fat with palm oil which contains palm stearin; this
component has a high melting point and therefore, the
fusion capacity of this cheese analogues was lower than
a traditional pasta filata Kashar cheese. All these results
differ from those of Totosaus et al. (2017), in which, as
per observation, they reported an increase of the melta-
bility of a pasta filata Oaxaca type cheese with 25%, 50%,
and 75% replacement of milk fat by soybean oil. This
result was related to the change in the moisture content
of cheese analogues by increasing or decreasing the inclu-
sion of vegetable oil which was not reported in the stud-
ies already described. The evaluation of the behavior of
the oil included for the milk fat replacement is mandatory,
as it can modify the technological properties of the end
product.
3.3 Texture
Texture is defined as a group of physical properties that
arise from the structural elements of a food type, perceived
mainly by the sensation of touch. It is directly related to
the deformation, disintegration, and flow of a food item
subjected to a force (Lu, 2013). The textural component is
determined by the response of the cheese to the applied
stresses during chewing, such as compression between the
molars or shearing in the mouth (Fox et al., 2017). There-
fore, a change in this parameter may affect sensory accep-
tance of a product. To measure this parameter, a wide
range of destructive and non-destructive methodologies
has been performed in fresh and processed foods (Chen
& Opara, 2013). For cheeses, texture analysis is usually
evaluated through compression tests such as texture pro-
file analysis (TPA). The TPA is an instrumental test origi-
2809
nally developed to provide objective measurements of tex-
ture parameters. It was designed as a two-cycle compres-
sion performed to simulate chewing (Nishinari et al., 2013).
All textural parameters (e.g., hardness, springiness, adhe-
siveness, gumminess, chewiness, and cohesiveness) were
evaluated among the different articles reviewed (Table 4)
in which the most evaluated texture parameter was hard-
ness.
Low-fat cheeses generally have a harder texture than
full-fat cheeses because the absence of fat concentrates
the other components, generating rigid protein bonds that
improve the physical strength of the cheese (Leong et al.,
2020). Most studies reported an increase in hardness after
milk fat substitution for vegetable oils (Abd El-Wahed &
Hassanien, 2019; Arslan et al., 2010; Cunha et al., 2010;
Leong et al., 2020; Ramel & Marangoni, 2018; Totosaus
et al., 2017), while some reported a decrease in this param-
eter (Hanáková et al., 2013; Kavak & Karabiyik, 2020).
A study using cheddar cheese analogues with full-fat
canola oil substitution showed a decrease in hardness
compared to the control cheese; furthermore, the ched-
dar cheese analogue also showed an increase in hardness
when a double emulsion was used (Leong et al., 2020).
Similar trends were also observed in different research
on cheese analogues (Fenelon & Guinee, 2000; Kesenkaş
et al., 2009). The softness of the single emulsion samples
can be attributed to the fact that canola oil is liquid at
room temperature (Leong et al., 2020). Similar trends were
also observed in different research on cheese analogues
(Fenelon & Guinee, 2000; Kesenkaş et al., 2009). Accord-
ing to Lobato-Calleros et al. (1998), raising the propor-
tion of saturated fatty acids of the fat content can increase
hardness in the product, and this is consistent within the
literature. A study conducted by Hanáková et al. (2013)
showed a decrease in the hardness in a model of processed
cheese after replacing 99% of the milk fat with coconut fat
and palm oil. This can also relate to results presented by
Dinkçi et al. (2011) wherein a pasta filata Kashar cheese
containing a 100% replacement of milk fat with palm oil
was significantly softer than the cheese with whole fat.
Other authors observed the same pattern with a fresh soft
cheese analogue containing 100% sunflower oil in which it
was explained that the inclusion of fat allows the protein
bonds to break down, which reduces rigidity and provides
smoothness and a softer texture (Abd El-Salam, 2015).
In contrast, the spreadable cheese analogue with 25%
and 50% milk fat replacement with hydrogenated vegetable
fat showed an increase in its hardness (Cunha et al., 2010).
This result was associated with a larger diameter of its
fat globules, which means that there was a higher pro-
tein mass per unit area of fat. A high protein density has
been related to higher levels of hardness since the pro-
tein matrix is the structural component of the product
2810 PHYSICAL PROPERTIES OF CHEESE ANALOGUES

TA B L E 4 Results of texture of cheese analogues manufactured with different vegetable oils

Hardness Adhesiveness Cohesiveness Springiness Chewiness Gumminess References
Decreased SE, – – – – – (Leong et al., 2020)
increased DE
Decreased – – Decreased Decreased Decreased (Kavak & Karabiyik,
2020)
Increased Increased Increased No change Increased Increased (Abd El-Wahed &
Hassanien, 2019)
Increased – – – – – (Ramel & Marangoni,
2018)
Decreased – – – – – (Hanáková et al., 2013)
Increased Increased – No change – Increased (Cunha et al., 2013)
Increased Increased – – – – (Cunha et al., 2010)
Increased Increased Decreased Decreased – – (Totosaus et al., 2017)
C and D – – No change E and F E and F (Arslan et al., 2014)
Decreased,E and increased increased
F Increased
Decreased No change Decreased No change Decreased Decreased (Dinkçi et al., 2011)
Increased – – No change Increased Increased (Arslan et al., 2010)
Notes: C. 75% milk fat replacement with soya bean oil; D. 2% fat content with 50% milk fat replacement with vegetable oil; E. 2.5% milk fat content with 60% fat
replacement vegetable oil; F. 1.5% fat content with 100% milk fat replacement vegetable oil.
Abbreviations: DE, double emulsion treatment; SE, single emulsion treatment.

(Lobato-Calleros et al., 1998). The main structure of cheese
is casein and its micelles are bound together and form
a three-dimensional matrix which gives structure to the
product. The compound that holds these proteins together
is calcium which helps to maintain a stable structure.
Therefore, adding more protein, calcium, or decreasing the
percentage of fat, which increases the concentration of pro-
teins in the matrix, can generate greater rigidity in the
cheese. This could also be related to the increased hardness
values that were obtained by Arslan et al. (2010, 2014) with
a white fresh type of cheese. As the fat content is reduced,
more areas of uninterrupted protein comprise the struc-
ture of the cheese, so the hardness can be increased. Over-
all, fat replacement can increase hardness because of the
change in cheese composition and structural variation of
the matrix (Abd El-Wahed & Hassanien, 2019; Cunha et al.,
2013; Ramel & Marangoni, 2018; Totosaus et al., 2017). Fur-
ther, few of the reviewed articles reported the springiness
parameter (Kavak & Karabiyik, 2020; Totosaus et al., 2017).
In cheeses, springiness is essentially conferred by the pro-
tein matrix (Fox et al., 2017); hence, this may be the reason
why no significant differences were found among the sam-
ples with respect to fat content. Few changes were observed
for the values of this texture parameter. The springiness of
the Oaxaca pasta filata cheese samples decreased signifi-
cantly when milk fat was replaced by emulsified soybean
oil (Totosaus et al., 2017). This same result was seen by
Kavak and Karabiyik (2020) when replacing 50% and 100%
of the milk fat in a pasta filata Kashar cheese with palm oil.
The reduction in springiness may have taken place during
cheese ripening as a result of the proteolytic breakdown of
the protein matrix (Jaster et al., 2019).
Regarding adhesiveness, it could be observed that all
the reviewed articles showed an increase in this parame-
ter for the cheese analogues developed with partial milk
fat replacement as shown in Table 4. These results can be
related to the size of the fat globules since the smaller their
size, the greater the adhesiveness (Fox et al., 2017). The
increase in the protein density and the protein interactions
influence the degree of adherence between the product and
the contact surface (Lobato-Calleros et al., 1997).
Additionally, an increase in the gumminess of the
cheeses was observed in all the studies that evaluated this
textural parameter except for two (Dinkçi et al., 2011; Kavak
& Karabiyik, 2020). These results can be explained by con-
sidering that gumminess is a secondary parameter of TPA,
which is conditioned by hardness and cohesiveness; so, as
hardness increases, gumminess also increases in each of
the analogous cheeses (Yusof et al., 2019).
Regarding chewiness, higher values of this parameter in
the cheese analogues were reported (Arslan et al., 2010,
2014; Abd El-Wahed & Hassanien, 2019). The analogues
that showed these results had a lower percentage of total
fat compared to the control cheeses. This may relate to the
findings described by Mistry (2001), who states that in low
fat variants, there is an inadequate breakdown of casein
and therefore, the cheese appears to have a relatively firm
texture.
PHYSICAL PROPERTIES OF CHEESE ANALOGUES
Regarding cohesiveness, Abd El-Wahed and Hassanien
(2019) reported an increase in this parameter in a spread-
able cheese analogue. This is consistent with the findings
from Lobato-Calleros et al. (1997) which state that a struc-
ture of the protein network and the interactions between
fat and casein influence the degree of consistency between
the product and the surface in contact with it. However,
other studies reported a significantly lower cohesiveness
(Dinkçi et al., 2011; Totosaus et al., 2017). In a study con-
ducted by Totosaus et al. (2017), when milk fat was replaced
by 75% with soya bean oil emulsion in an Oaxaca cheese
analogue, cohesiveness decreased. Further, Dinkçi et al.
(2011) also reported a decrease in cohesiveness after milk
fat replacement by vegetable oil. These differences may be
due to vegetable oil and milk fat linkages, which have dif-
ferent intermolecular associations of casein in the para-
casein network of cheese (Abd El-Salam, 2015). There are
differences between the hydrophobic characteristics of the
types of fat used, the type of cheese and the cheese man-
ufacturing process (Lamichhane et al., 2018). In general
terms, the texture parameter that reports the greatest vari-
ability is hardness and this may be related to the variety of
cheeses evaluated, as shown in Table 1. Likewise, all the
parameters were altered from the fat modification, so one
must identify the adequate level of fat replacement to pro-
vide ideal characteristics.
The variations among the results can be attributed to the
different test conditions in which the compression speed,
the time between compressions, and the diameter of the
cylinders varied. The type of oil used, the type of cheese
produced, and the type of inclusion for which the substitu-
tion was made must also be considered. This would affect
the result of the texture characteristics.
4 STRENGTHS AND LIMITATIONS
This was the first review that sought/aimed to show the
effects of fat modification in analogue cheeses focusing on
the effects on rheology, texture, and microscopy. The high
variability of cheese types in the market makes it difficult to
unify results in terms of ideal attributes. In addition, there
is a lot of variability in the manufacturing techniques of the
cheeses, the type of oil used, the percentage of fat inclusion
and the ingredients of the cheese, as well as the evaluation
techniques employed. Therefore, drawing detailed conclu-
sions and establishing the level of association among stud-
ies is complex. The need of further research on the proper-
ties of fat modification in analogous cheeses must be high-
lighted. For future studies, construing the results from the
intrinsic properties of the original cheese to decrease the
variance between types of cheese is suggested.
2811
5 CONCLUDING REMARKS AND
FUTURE PERSPECTIVES
Few studies in the literature specifically address the effects
of milk fat substitution by vegetable oils on the rheolog-
ical, microstructural, and textural profile of cheese ana-
logues. The compiled data showed that partial or total
fat substitution can significantly affect the technological
properties of cheeses. Changes may vary depending on
the type of oil used and the percentage of fat replace-
ment applied. Regarding microstructure, fat globule size
may vary. Some studies show globules tend to increase
their diameters; thus, they might also decrease, which
was/could be attributed to the breakage of structural bonds
during processing, hence creating different size particles.
With respect to the number and size of the pores, the
increased porosity observed represented the fat globules
that were removed. Rheological measurements showed
most cheese analogues with fat substitution presented G′–
G′′ cross-over behavior at which the frequency decreased
as fat concentration replacement increased. Moreover, the
melting properties of the cheese analogues decreased due
to the increase in the protein interactions caused by the
decrease in the size of the fat cells and their homogeniza-
tion in the milk. Overall, most cheese analogues showed
significant changes in all textual parameters except in
springiness.
This review evidence that there is a great diversity of
variables present in studies on cheese analogues, which
could be reflected in the different results found for each
of the parameters evaluated. It is necessary to carry out
more research with defined methods that can contribute
to the unification of the outcomes. However, the previous
studies are a great tool with which to start improving the
attributes of analogue cheeses in attempting to preserve a
greater percentage of the technological and instrumental
characteristics of the imitated cheese. It is valid to empha-
size the importance of continuing with the development of
different products with fat modification that increases the
offer of healthy cheeses in the market to promote a healthy
lifestyle.
AC K N OW L E D G M E N T S
The authors would like to express their gratitude to Min-
isterio de Ciencia Tecnología e Innovación (MinCiencias)
because two of the authors have been supported by the
Young Researchers Program. This work was also supported
by the Pontificia Universidad Javeriana, Bogotá, Colombia.
AU T H O R CO N T R I B U T I O N S
Valeria Giha: Methodology; Writing-original draft. María
José Ordoñez: Writing-original draft; Writing-review and
2812
editing. Ruby Alejandra Villamil: Conceptualization;
Funding acquisition; Resources; Supervision; Writing-
review and editing.
CONFLICTS OF INTEREST
The authors declare no conflict of interest.
ORCID
Ruby Alejandra Villamil https://orcid.org/0000-0002-
2513-2929
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How to cite this article: Giha, V., Ordoñez, M. J.,
& Villamil, R. A. How does milk fat replacement
influence cheese analogue microstructure,
rheology, and texture profile? J Food Sci,
2021;86:2802–2815.
https://doi.org/10.1111/1750-3841.15799