food and bioproducts processing 9 1 ( 2 0 1 3 ) 429439

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Food and Bioproducts Processing

journal homepage: www.elsevier.com/locate/fbp

Evaluation of the rheological, textural, microstructural and

sensory properties of soy cheese spreads

Qinghui Li, Yuelan Xia, Li Zhou, Jingli Xie

State Key Laboratory of Bioengineering Reactor, Department of Food Science and Technology, School of Biotechnology, East China
University of Science and Technology, Shanghai 200237, PR China

a b s t r a c t

In this work, different processes including lactic acid bacteria fermentation, glucono--lactone (GDL) coagulation
and enzymatic hydrolysis were applied to produce soy cheese spread without any milk solid and/or alkaline metal
caseinates added. All the soy cheese spread samples were studied with respect to the differences in the textural
properties utilizing TPA and rheological behavior, besides their microstructural features and sensory assessment.
In addition, ureaSDS-PAGE assay was used to investigate the proteolysis in enzymatic hydrolysis process. As a result,
the soy cheese spread sample produced by the combined use of GDL and lactic acid bacteria fermentation together
with enzymatic hydrolysis process, was observed with better spread-ability, more acceptable sensory features, and
stable homogeneous structure than other samples.
2013 The Institution of Chemical Engineers. Published by Elsevier B.V. All rights reserved.

Keywords: Soy cheese spread; Glucono--lactone; Lactic acid bacteria; Enzymatic hydrolysis; UreaSDS-PAGE

1. Introduction described in unfavorable terms with a negative stereotype

(Schyver and Smith, 2005). A typical result is, that many west-
Soybeans are excellent source of high-quality protein and has ern people do not adapt to the diet of soy foods, such as tofu
many uses in human nutrition (Liu, 1997). Consumption of soy and preserved beancurd, which are traditional foods of most
foods and the incorporation of soymilk and its by-products in Asian populations.
human diets are increasing due to their reported benecial It might make a difference, if soybeans were used to make
effects on nutrition and health (Rinaldoni et al., 2012). These food products which are readily accepted by most people. For
effects include lowering of plasma cholesterol, prevention of this purpose, cheese-like product may be a relatively ideal and
cancer, diabetes, osteoporosis, and obesity, and protection suitable form of soy food.
against bowel and kidney disease, and relief of menopausal Various soy cheeses are made in many countries, but as
problems (Anderson et al., 1999; Friedman and Brandon, 2001; many rely on the use of heat and salt/lactic acid (magne-
Shah et al., 2007). Soybean proteins are used in human foods sium sulfate, calcium sulfate or glucono--lactone) to produce
in a variety of forms, including infant formulas, ours, protein the gel, the texture of the nal products is often unsatis-
isolates and concentrates, and texturizing bers. Soy foods factory (Kohyama and Nishinari, 1993). Related researches
include cheese, drinks, miso, tempeh, tofu, salami, and vege- of soy-based cheese analogs have attracted much attention
tarian meat substitutes. New soy foods are continually being recently (Otieno et al., 2005). Some researches focused on mak-
developed (Singh et al., 2000). ing cheese from mixtures of cow milk and soymilk, and trying
Although the health benets of soy foods have generated to increase the proportion of soymilk in the blend. The qual-
an increase in consumer awareness, especially among the ity of the cheese is proportionally reduced signicantly with
people who were unfamiliar with soy foods, the taste, texture, increased level of soymilk in the blend (Rani and Verma, 1995).
and visual appearance which are all considered components Some researchers have carried out studies on spreadable pro-
of soy foods physical sensory characteristics, in general, are cessed cheese analogs made with vegetable fat (Bachmann,
2001; Cunha et al., 2010), and some others have tried to blend
various amounts of tofu, oil, salt, carrageenan, pectin and

Corresponding author. Tel.: +86 021 64251803; fax: +86 021 64251803.
E-mail address: jlxie@ecust.edu.cn (J. Xie).
Received 31 March 2012; Received in revised form 17 January 2013; Accepted 4 March 2013
0960-3085/$ see front matter 2013 The Institution of Chemical Engineers. Published by Elsevier B.V. All rights reserved.
http://dx.doi.org/10.1016/j.fbp.2013.03.001
430 food and bioproducts processing 9 1 ( 2 0 1 3 ) 429439
maltodextrin to develop soy based cream cheese products
which were trans fat free, and which had textural properties
similar to those of commercial dairy cream cheese (Zulkurnain
et al., 2008). However, the main problem remains that prod-
ucts made from soybeans tend to have an undesirable beany
taste. Furthermore, the texture of spreads made from soybean
sources usually have a grainy texture in comparison to the
smooth, uniform texture normally associated with spreads
made from dairy cheeses (Hashimoto and Sunada, 1976).
Thus, it was the interest of the authors to develop differ-
ent technical processes for making soy cheese spreads with
good functional properties, avor characteristics, and stable
homogeneous structure, but without any milk solids added.
The role of lactic acid bacteria in soymilk fermentation is
to produce acid and avor, and to remove undesirable beany
taste (Chou et al., 2006; Wang et al., 2003), which mainly comes
from the presence of n-hexanal and pentanal (Scalabrini
et al., 1998), and furthermore to improve the nutritional pro-
le of soymilk. Considerable amounts of atulence-causing
oligosaccharides in soy foods such as rafnose and stachyose
limit soy foods biological value and acceptability (de Ftima
Viana et al., 2005). The hydrolysis of these oligosaccharides
requires -galactosidase, an enzyme that hydrolyzes the -
galactoside bond in such indigestible sugars. Fermentation
of soy products with microorganisms that possess high -
galactosidase activity has been found to minimize the content
of atulence-causing oligosaccharides in soy products. Bi-
dobacteria and lactobacilli such as Lactobacillus fermentum and
Lactobacillus acidophilus have been reported to produce varying
levels of -galactosidase. Fermentation, especially with Bi-
dobacteria, also renders the soy proteins more digestible and
reduces the content of soy oligosaccharides (Bozanic et al.,
2008). In addition, probiotics have been reported to exhibit pro-
teolytic activity to produce shorter-chain peptides and amino
acids to meet their requirement for growth (Liong et al., 2009).
Coagulation is the basic process of cheese production. To
cause the coagulation of milk, rennet is added, and casein in
milk is changed into paracasein by the enzymatic hydrolysis
of rennet, which results in gluco macro peptide (GMP) being
cleaved from the casein micelles. Due to interaction between
remaining paracasein and calcium salts, milk curd is formed.
Unlike cow milk, soymilk does not contain caseins, and
thus the rennet does not work in soymilk in relation to curd
formation. Thus the common cheese production process is
not available in soy cheese production. Although there is
some research on the coagulation of soy protein induced by
various proteolytic enzymes, the coagulum induced by such
enzymes is weak and with an incompact structure (Aoyama
et al., 2000; Fuke et al., 1985; Inouye et al., 2002; Murata et al.,
1987; Yasuda et al., 1999). For this reason, soybean protein
is usually coagulated by bacterial cultures or by acids and
salts such as calcium sulfate, magnesium sulfate or glucono-
-lactone (GDL). GDL is a compound used to produce tofu
with good coagulation ability and also contributes toward
the formation of silky and good-avored curd (Kohyama and
Nishinari, 1993). Nevertheless, the soy cheese analog made
from pressing the curd which is formed by soybean protein
gathered when adding bacterial cultures or salts always has
a grainy and loose texture (Hashimoto and Sunada, 1976).
The objective of this work is to develop optimal processes
for the soy cheese spreads production, by using lactic acid
bacteria and GDL together with enzymatic hydrolysis process.
Moreover, the evaluation of the processes was made by textu-
ral properties, microstructural features and sensory features
of soy cheese samples made through different production pro-
cesses.
2. Materials and methods
2.1. Preparation of starter culture
Two probiotic strains of Lactobacillus acidophilus NCFMTM and
Bidobacterium lactis HOWARUTM Bido were purchased from
the Danisco Deutschland GmbH. A starter culture of NCFMTM
and HOWARUTM Bido in the ratio 1:1 was prepared by inocu-
lating the two strains (0.06%, w/v) in sterilized skim milk (12%
TS) under neutralizing conditions and incubated at 37 C for
1618 h. The milk had been stored under 4 C before use.
2.2. Soy cheese spread manufacture
Commercial soybeans (approximately 36.3% protein, 18.4%
fat and 26% carbohydrate), from Northeast China, were pur-
chased from a local supermarket. Soymilk was prepared
according to the method described as follow:
Soybeans were soaked in 0.1% (w/v) sodium bicarbonate
solution (solid:liquid = 1:5, w/v) over night, before they were
heated to 85 C for 10 min. Then the soybeans were washed
with fresh water and peel was removed, and ground in water
at 95 C (soybeans:water = 1:10, w/v) with a soymilkgrinder
(JYDZ-33B, Joyoung Co. Ltd., China), 11,000 rpm for 4 min. The
resultant slurry was then ltrated manually with a muslin
cloth to obtain soymilk while the soy residue was discarded.
The cooked soymilk was added with 0.1% (w/v) carrageenan
(GRINDSTED Carrageenan CH515, Danisco A/S, Denmark) and
2.5% (w/v) coconut oil with a melting point at 23 C, and then
homogenized at 25 MPa.
Four different soy cheese spread samples (hereinafter to be
referred as SCS-A, SCS-B, SCS-C, and SCS-D) were prepared by
distinct production processes as follows:
SCS-A: In the rst step, the soymilk was inoculated with 3%
(v/v) starter culture and incubated in thermostatic 37 C water
bath. After coagulation, the curd was cut vertically and hori-
zontally with cheese knives and the temperature was raised
to 45 C over 15 min. Then whey was drained and curd was
transferred to a mold (15 cm 15 cm 5 cm) lined with cheese-
cloth. The mold had perforations on the sides and bottom. The
curd was pressed in a controlled and incremental manner to
consolidate it to produce rm soy cheese A (hereinafter to be
referred as FSC-A). In the second step, the FSC-A was weighed
and heated in water bath to 70 C for 5 min, then cooled to
40 C and 0.1% (w/w) proteolytic enzyme papain (8 105 U/g,
Pangbo Biological Engineering Co. Ltd., China) was added with
slow stirring for 5 min. After heated again in water bath to
70 C for 3 min, it was transferred to a blender (JYL-B031, Joy-
oung Co. Ltd., China) and blended with 2% water (in relation
to the initial weighed rm soy cheese weight, w/w), 10% (w/w)
coconut oil, 2% (w/w) emulsifying salt (sodium tripolyphos-
phate (STPP):trisodium phosphate (TSP):sodium citrate = 2:2:1)
and 0.3% (w/w) salt, stirring at 1500 rpm for 25 min. At last, the
sample SCS-A was obtained and lled into glass jars, kept at
4 C.
SCS-B: The soymilk was coagulated with glucono--lactone
(GDL, 0.25%, w/v) at 50 C for 3 h. After the resulting curd was
cut vertically and horizontally with cheese knives and drained
of whey, it was transferred to a mold (15 cm 15 cm 5 cm)
lined with cheesecloth. The curd was pressed in a controlled
and incremental manner to consolidate it to produce rm soy
 food and bioproducts processing 9 1 ( 2 0 1 3 ) 429439
cheese B (hereinafter to be referred as FSC-B). Then the sample
SCS-B was made according to the same process mentioned in
the second step of the manufacture of sample SCS-A.
SCS-C: After the soymilk was inoculated with 3% (v/v)
starter culture and incubated at 37 C in thermostatic water
bath until the acidity thereof reached 0.20% for approximately
1.5 h, 0.25% (w/v) GDL was added. Then the soymilk was
heated to 50 C until the curd was formed. The curd was cut
vertically and horizontally with cheese knives and whey was
drained. Then it was pressed to produce rm soy cheese C
(hereinafter to be referred as FSC-C). Then the sample SCS-C
was made according to the same process mentioned in the
second step of the manufacture of sample SCS-A.
SCS-D: The coagulation of soymilk and mold pressing pro-
cess were the same as that of the sample SCS-C. After the
rm soy cheese D (hereinafter to be referred as FSC-D) was
weighed and heated in water bath to 70 C for 5 min, it was
directly transferred to the blender without proteolysis process
and blended with 2% water (in relation to the initial weighed
rm soy cheese weight, w/w), 10% (w/w) coconut oil, 2% (w/w)
emulsifying salt (sodium tripolyphosphate (STPP):trisodium
phosphate (TSP):sodium citrate = 2:2:1) and 0.3% (w/w) salt,
stirring at 1500 rpm for 25 min. Then the sample SCS-D was
obtained and lled into glass jars, kept at 4 C.
2.3. Chemical analyses
All SCS samples, with their respective previous FSC matrixes,
and a commercial dairy cream cheese spread sample (DCCS)
purchased from local supermarket (Processed Cream Cheese
Spread, produced by Arla Foods Amba, Denmark) were
analyzed in triplicate, and average results were used for eval-
uation. Fat was determined by a modied Babcock method
(Marshall, 1992). To determine protein content, total nitrogen
content of the cheese was measured by the Kjeldahl method
(AOAC, 2000) and a factor of 5.71 was used to convert nitrogen
content to protein content. Ash content was determined by
using a mufe furnace at 550 C and moisture, pH and titrat-
able acidity were determined according to AOAC (2000).
2.4. UreaSDS-PAGE (ureasodium dodecyl
sulfate-polyacrylamide gel electrophoresis)
All SCS samples and their previous FSC matrixes were tested
using ureaSDS-PAGE assay to measure the level of proteol-
ysis. 10 mg of each sample were respectively added to 0.5 ml
of loading buffer solution containing 20% (v/v) glycerol, 0.2%
(w/v) SDS, 0.063 M TrisHCl, 6 M urea, 4% (v/v) 2-ME, and 0.2%
(w/v) bromophenol blue. The mixtures were shaken for 5 min
in order to dissolve by a vortex mixer (Vortex-5, Kylin-Bell
Lab Instruments Co. Ltd., China), and incubated for 30 min at
25 C. After heating for 5 min in boiling water bath and allowed
to cool, the suspensions were centrifuged at 10,000 rpm for
10 min. The amount loaded into the gel was 20 l. The protein
molecular weight marker (TaKaRa BIO Inc., Japan) was also
heated for 5 min in a boiling water bath and allowed to cool
before use, and 5 l of this was loaded into the gel.
The ureaSDS-PAGE of samples was performed using a
Mini-PROTEAN Tetra system (Bio-Rad Laboratories Inc., USA)
with stacking gel of 5% and separating gel of 12.5% acrylamide.
The separation was carried out at 120 V constant voltages for
60 min and then stained with 0.1% Coomassie Brilliant Blue
R-250 for 45 min. Destaining was done with a solution of 5%
ethanol and 10% acetic acid.
431
2.5. Texture prole analyses
Specimens stored at 4 C were cut into cuboids
(5 cm 5 cm 1.5 cm) before that they were assayed via
measurement of the forcetime curve using a TA.XT-plus
apparatus (Stable Micro Systems Ltd., UK). A 5 kg-load
cell was selected and duly calibrated in advance with a
2 kg-weight. The probe used was P/25 (25 mm Dia cylinder
aluminum), and TPA tests were performed in triplicate.
A typical mastication testing prole was thus imple-
mented, which involves two consecutive compressions at
(controlled) room temperature (25 C); the compression dis-
tance was 0.75 cm (50%), and the test speed was 1.0 mm s1 .
The tests generated a plot of force vs. time from which tex-
tural parameters were automatically calculated. Hence, it
was possible to measure hardness, adhesiveness, springiness,
cohesiveness, gumminess, chewiness, and resilience of the
DCCS and SCS specimens.
2.6. Rheological analyses
The dynamic oscillatory test was used to measure the dynamic
viscoelastic properties of the samples. The measurements
were obtained by using a controlled-strain rheometer (AR-
G2, TA Instruments, New Castle, DE, USA) equipped with a
temperature controller. A scoop of each sample was dug out
with an iron spoon, and loaded between two 25-mm, par-
allel plate geometry with a gap of 1800 m. Then a special
scraper scrapes off the rest part of the sample. A thin layer
of parafn oil was gently applied to the edge of the exposed
sample to prevent moisture loss. Dynamic strain sweep test
was performed to determine the linear viscoelastic region
(LVR) such that all oscillation tests were performed within this
region (Zulkurnain et al., 2008). The storage modulus (G ) and
loss modulus (G ) were determined during a frequency sweep
which increased from 0.1 to 100 Hz at a constant of 0.5% strain
over a period of 10 min. All tests were conducted in triplicate
at 70 C.
2.7. Scanning electron microscopy
Samples for SEM were cut from the interior of each cheese with
a razor blade into a rectangular strip of 4 mm 4 mm 2 mm
size. The cheese strips were xed by immediately immers-
ing in a freshly prepared 2.5% (v/v) glutaraldehyde solution
and stored for 6 h at refrigerated temperature (46 C). Sam-
ples were cleaned with the physiological saline (pH 7.2) three
times for 10 min each. Then samples were dehydrated by
subsequent transfer, for 10 min each, in a series of ethanol
concentrations (10%, 20%, 40%, 60%, 80%, 90%, and 100%),
defatted 3 times in chloroform for 30 min each and stored
in a glass tube containing isoamyl acetate. When all the
samples were ready, they were freeze fractured in liquid
nitrogen, thawed in isoamyl acetate, and nally dried in
a critical point carbon dioxide drying apparatus (Denton
Vacuum, Inc., Cherry Hill, NJ). The dried specimens were
mounted on aluminum stubs using 2-sided sticky tape and
coated with a thin layer of gold in an ion sputter (Hitachi
E-1045 ion sputter, Hitachi Ltd., Tokyo, Japan). Microstruc-
ture examination of these specimens was done using a
JSM-6380LV scanning electron microscope (JEOL, Japan). Spec-
imens made from replicates of each type of sample were
compared to assure that artifacts were identied in pictures
selected.
432 food and bioproducts processing 9 1 ( 2 0 1 3 ) 429439
2.8. Sensory analyses
An acceptance sensory panel assessed the coded experimen-
tal DCCS and all SCS specimens at random, according to
the methodology described by Meilgaard et al. (1999). The
panel consisted of 10 members, specically trained for dairy
product sensory analyses, with ages ranging from 20 to 40
years old. Specimen pieces were placed into airtight plas-
tic containers, and conditioned at room temperature for
15 min before testing (so as to guarantee that samples were
tested while still fresh). Duplicated samples were assessed
at room temperature (20 C) and under white light after pre-
vious conrmation of microbiological safety, using a 9-point
structured hedonic scale described by Meilgaard et al. (1999),
with some modications. According to the 9-point struc-
tured hedonic scale, the acceptance test was carried out for
the attributes of appearance (1 = very rough; 9 = very smooth
and homogeneous), color (1 = colorless and gray; 9 = dark and
bright), avor (1 = undetectable; 2 = very intense), creaminess
(1 = undetectable; 9 = very intense), rmness (1 = very soft;
9 = very hard), spread-ability (1 = not spreadable; 9 = very good)
and overall impression (1 = very bad; 9 = liked very much). For
all the attributes, 9-point scale was dened as that the high-
est value indicates the highest degree of preference. A plastic
knife and some dry bread were offered together with the sam-
ple, for the evaluation of spread-ability. Moreover, crackers
and water were accessible to all panelists, to clean the palate
whenever appropriate.
2.9. Data presentation and statistical analyses
All data presented are mean values of triplicates, obtained
from three separate experiments, unless stated otherwise.
Analyses of variance were carried out where indicated using
SAS version 8.02 (SAS Institute Inc., Cary, NC, USA). Compar-
isons were considered signicantly different if P < 0.05.
3. Results and discussion
3.1. Chemical prole
Fig. 1 shows the change of the pH and titrable acidity (TA%)
of the soymilk, fermented by inoculating 3% starter culture
Fig. 1 Change of the pH and titrable acidity (TA %) of the
soymilk fermented by inoculating 3% starter culture of
NCFMTM and HOWARUTM Bido in the ratio 1:1 during the
coagulating process.
during the coagulating process. It indicated that the soymilk
was a good medium to support extensive growth of lactic acid
bacteria, thus the lactic acid bacteria fermentation method
was potential to coagulate the soymilk. The result is consistent
with Farnworth et al. (2007).
The chemical composition of the SCS samples, their pre-
vious FSC matrixes, and the DCCS sample were illustrated by
Table 1. As a result of the addition of fat, the fat content of SCS
samples was higher than that of their previous FSC matrixes,
respectively. However, comparing to the dairy cream cheese
spread sample, all the soy cheese spread samples had lower
fat content. Although dairy products were well acknowledged
as a source of protein, our results showed that the protein
content of the soy cheese spread samples was almost triple of
protein content of the commercial dairy cream cheese spread
sample. This observation is consistent with Zulkurnain et al.
(2008), which indicated that soy is a nutritionally complete
protein with an unusually well-rounded amino acid prole as a
plant protein, and contains adequate quantities of all essential
amino acid.
For the FSC matrixes, the FSC-A sample, which was made
by lactic acid bacteria fermentation method, had the highest
fat and protein content, but the lowest pH. While the FSC-B
sample, made by GDL use only process, was with the low-
est fat and protein content, but the highest pH. The FSC-C
and FSC-D samples, made by the combined methods of GDL
and lactic acid bacteria fermentation, showed an intermediate
value between FSC-A and FSC-B. The differences of chemical
features among the cheese samples were mainly attributed
to the different coagulation processes. The metabolism of
starter cultures and their enzymes led to some complicated
biochemical reactions including the degradation of proteins,
carbohydrates and fats in the curd (Ahmad et al., 2008),
which resulted in protein and fat particles were degraded into
smaller size, and could distribute evenly in or bound by the
network structure of cheese samples. In contrast, the coagu-
lation of the soymilk caused by GDL directly, led to more loss of
proteins and fats since the network structure crushed during
pressing process.
SCS-A, SCS-B and SCS-C were respectively made from FSC-
A, FSC-B and FSC-C by the same process. However, the SCS-B
displayed the lowest fat and protein content, and the highest
pH, and the SCS-C turned to with the highest fat and protein
content and the lowest pH, meanwhile the SCS-A showed a
intermediate value between the SCS-B and SCS-C. It indicated
GDL method created a poor fat and protein retention capacity,
and high pH. On the other hand, lactic acid bacteria fermenta-
tion method brought about a better fat and protein retention
capacity, and lower pH. Yet there was an interesting result
about the combination of GDL and lactic acid bacteria fermen-
tation methods, which realized best fat and protein retention
capacity, and the closest pH to the dairy cream cheese spread.
In addition, the signicant differences (P < 0.05) in fat, pro-
tein and moisture contents between sample SCS-C and SCS-D
suggested that additional enzymatic hydrolysis process was a
key factor for the chemical composition of the cheese samples.
3.2. Proteolysis (ureaSDS-PAGE)
UreaSDS-PAGE was used to illustrate the proteolysis of all SCS
samples and FSC matrixes. According to the gel image showed
in Fig. 2, same distinct bands from 15 to 100 kDa were
observed in all FSC matrixes (FSC-A, FSC-B, FSC-C, and FSC-
D) and the SCS-D sample, whereas in other three SCS samples
 food and bioproducts processing 9 1 ( 2 0 1 3 ) 429439 433

Table 1 Chemical composition of the SCS samples (SCS-A, SCS-B, SCS-C, and SCS-D), their respective previous FSC
matrixes (FSC-A, FSC-B, FSC-C, and FSC-D), and commercial dairy cream cheese spread sample (DCCS).
Sample Fat (g/100 g) Total protein Moisture content Salt content Ash content pH
content (g/100 g) (g/100 g) (g/100 g) (g/100 g)

FSC-A 20.37 0.02a 22.39 0.02a 56.29 0.56a 0.53 0.01a 4.68 0.02a
FSC-B 18.51 0.02b 19.72 0.01c 57.58 0.30a 0.26 0.01b 5.81 0.03b
FSC-C 20.24 0.02a 20.11 0.02b 55.26 0.16a 0.96 0.01c 4.80 0.02a
FSC-D 20.24 0.02a 20.11 0.02b 55.26 0.16a 0.96 0.01c 4.80 0.02a
SCS-A 25.32 0.02a 17.60 0.02c 49.62 0.27b 0.3 0a 1.72 0.01d 6.55 0.01b
SCS-B 23.94 0.02a 17.10 0.02d 51.09 0.36b 0.3 0a 1.30 0.01d 7.20 0.02c
SCS-C 26.74 0.02a 19.48 0.03b 48.29 0.43b 0.3 0a 1.56 0.01d 6.54 0.02b
SCS-D 24.80 0.02a 17.20 0.02d 59.15 0.20a 0.3 0a 1.53 0.01d 6.63 0.02b
DCCS 28.74 0.02a 6.40 0.02c 54.48 0.18a 0.7 0a 1.93 0.01d 6.30 0.02b

Values are means standard errors; n = 3. Means pertaining to the same parameter (i.e. in the same column), with the same superscript (ad),
do not differ signicantly (P > 0.05) from each other.

Fig. 2 UreaSDS-PAGE analyses of SCS samples (SCS-A, SCS-B, SCS-C, and SCS-D) and their previous FSC matrixes (FSC-A,
FSC-B, FSC-C, and FSC-D).

(SCS-A, SCS-B, and SCS-C) with enzymatic hydrolysis, no clear

bands were observed, which indicated that the soy proteins in
such samples were extensively hydrolyzed by the papain, and
degraded further into peptide fragments with lower molecular
mass which were not detected by ureaSDS-PAGE. FSC-B was
noted by two exceptional distinct bands above 100 kDa but not
existed in the gel of FSC-A, FSC-C and FSC-D, which should
due to the proteolysis of FSC-A, FSC-C and FSC-D caused by
the starter cultures.
Soy protein was generally regarded as the storage pro-
tein held in discrete particles called protein bodies, and with
big molecular size (Shewry et al., 1995). Therefore, the prod-
ucts made from soybean sources usually had a grainy texture
(Hashimoto and Sunada, 1976). Similarly, the problem also
arose in present study and disturbed the development of
present soy cheese spread products. The results of elec-
trophoresis showed that the enzymatic hydrolysis process was
able to degrade the large soy protein molecules into peptide
fragments with smaller size, which could settle the problem
of grainy texture.

3.3. Evaluation of the rheological and textural proles Fig. 3 The viscoelastic linearity behavior of all soy cheese
spread samples (SCS-A, SCS-B, SCS-C, and SCS-D) and the
According to the results of dynamic strain sweep test dairy cream cheese spread sample (DCCS) determined by
(Fig. 3), the critical strain limits of the SCS samples were the dynamic strain sweep test.
434 food and bioproducts processing 9 1 ( 2 0 1 3 ) 429439
approximately between 1.2% and 4.1%, whereas the critical
strain limit of the DCCS sample was approximately 0.8%. The
value of the DCCS sample is consistent for the majority of
dairy cheese spread, of which the linear viscoelastic strain
limit is typically 1% or less (Gunasekaran and Ak, 2002). The
critical strain limits of the SCS samples were higher than that
of the DCCS sample, which could be owing to their lower fat
content and higher protein content. In dairy cheese spread,
solidication of fat during cooling contributes to the rigidity
of the casein matrix and reduces the exibility of structure.
Moreover, carrageenan plays an important role in the gelling
properties of the soy protein to form carrageenan mixed gel,
which apparently enhances the structural integrity of the soy
cheese spread (Zulkurnain et al., 2008).
The critical strain limits of the SCS samples followed the
order, SCS-C > SCS-D > SCS-B > SCS-A, which indicated that the
sample SCS-C, made by combined use of GDL and lactic acid
bacteria fermentation methods, exhibited a more stable and
less easily fractured structural system than other samples.
However, when the lactic acid bacteria fermentation method
or GDL coagulation method was processed separately, more
easily fractured structural system formed and even the lactic
acid bacteria fermentation method carried out the most break-
able structure. In addition, the enzymatic hydrolysis process
was able to improve the stability of samples structural sys-
tem by comparison of SCS-C and SCS-D, which might be a
result of that the large soy protein molecules were degraded
into smaller size by enzymatic hydrolysis, which allowed
new formation of bands in the hydrocolloid polymer net-
work and had more internal cohesive force (Medureira et al.,
2010).
The results of the dynamic frequency sweep test for the
DCCS and all the SCS samples are shown respectively in
Figs. 48. It was observed that the elastic modulus (G ) were
higher than the viscous modulus (G ) for both DCCS and SCS
samples, suggesting that both DCCS and SCS samples exhib-
ited a solid-like behavior and own dense structure. Similar
behaviors have been reported in various commercial cream
cheeses by Kealy (2006) and also reported in soy cream cheese
by Zulkurnain et al. (2008). In addition, the G values of
both DCCS and SCS samples increased with the increasing
Fig. 4 Evaluation of the dynamic rheological properties of sample DCCS according to the dynamic frequency sweep test.
angular frequency over the range of 0.1100 rad/s, whereas
the G values decreased at low angular frequency over the
range of 0.11.0 rad/s then increased as the angular frequency
increased over the range of 1.0100 rad/s.
The viscoelastic behavior of the DCCS and SCS samples
indicated that the sample SCS-B, which was made by GDL
method, exhibited less elastic behavior and had less internal
cohesive force compared with the SCS-A sample made by lac-
tic acid bacteria fermentation method, while sample SCS-C
made by combined use of GDL and lactic acid bacteria fermen-
tation methods exhibited a intermediate value between them.
According to Medureira et al. (2010), the higher rate of acidi-
cation probably account for more elastic and viscous of the
matrices. In present work, lactic acid bacteria fermentation
led to a high rate of acidication, whereas GDL coagulation
presented a lower rate of acidication. This might cause the
viscoelastic behavior difference between SCS samples. More-
over, DCCS exhibited a G value close to the SCS-A, which
might be the result of lactic acid bacteria fermentation.
Table 2 shows the textural properties obtained by TPA test.
According to the results, the textural properties of the DCCS
and SCS samples were signicantly different (P < 0.05) from
each other. The value of cohesiveness displayed the follow-
ing order: SCS-A > DCCS > SCS-C > SCS-B, which was coinciding
with the results of the G values. Comparing to the DCCS
sample, the SCS samples exhibited evidently lower values of
adhesiveness and chewiness, and higher resilience. It sug-
gested that the spread-ability and chewiness of SCS samples
might be lower than that of DCCS. However, in contrast, the
values of adhesiveness, gumminess, and chewiness for the
SCS-C sample were signicantly higher (P < 0.05) than those of
the sample SCS-A and SCS-B, suggesting that SCS-C presented
optimal textural properties in all SCS samples. Comprehen-
sive analysis of the textural proles suggested that the lowest
value of textural properties were observed in SCS-B, while
SCS-A exhibited the highest hardness, but other parame-
ters of textural properties were not prominent. Furthermore,
SCS-D showed lower values of all textural properties except
chewiness than SCS-C, suggesting that enzymatic hydrolysis
process was able to improve textural properties of the SCS
samples.
 food and bioproducts processing 9 1 ( 2 0 1 3 ) 429439 435

Fig. 5 Evaluation of the dynamic rheological properties of sample SCS-A according to the dynamic frequency sweep test.

Fig. 6 Evaluation of the dynamic rheological properties of sample SCS-B according to the dynamic frequency sweep test.

Fig. 7 Evaluation of the dynamic rheological properties of sample SCS-C according to the dynamic frequency sweep test.
436 food and bioproducts processing 9 1 ( 2 0 1 3 ) 429439

Fig. 8 Evaluation of the dynamic rheological properties of sample SCS-D according to the dynamic frequency sweep test.

Table 2 The data of textural properties obtained by TPA test.

Sample Hardness Adhesiveness Springiness Cohesiveness Gumminess Chewiness Resilience

SCS-A 397.456a
846.244 a
0.910a
0.595a
140.685a
128.081 a
0.041a
SCS-B 217.407b 663.147b 0.896b 0.487b 129.367b 115.895a 0.046a
SCS-C 378.365c 1384.114c 0.915a 0.567c 245.656c 186.704b 0.039b
SCS-D 370.213c 1289.201c 0.906a 0.539d 204.003d 222.509c 0.046a
DCCS 301.682d 2187.313d 0.911a 0.573c 205.428d 232.061d 0.018c

Means pertaining to the same parameter (i.e. in the same column), with the same superscript (ad), do not differ signicantly (P > 0.05) from
each other.

3.4. Evaluation of the microstructural prole
Figs. 1013 show the microscopy photographs of the
microstructure of the SCS samples, respectively, with the
same magnitude (5000). In typical scanning electron
microscopy of cheese (Fig. 9), a protein matrix is visible with
various forms and sizes of open spaces, representing the fat
globules that were extracted during sample preparation for
analysis (Cunha et al., 2010).
An open, irregular and porous network was observed in the
microstructure of each sample. Unlike traditional processed
cheese, the fat globules of soy cheese spread did not dis-
tribute uniformly throughout the protein matrix. Fig. 10 shows
that the sample SCS-A exhibited a compact protein network
with a spot of big open spaces. It indicated that fat glob-
ules were large and unevenly dispersed in the sample SCS-A,
which resulted in the sample SCS-A lacking a homogeneous
and smooth texture. The sample SCS-B was observed to own
a loose and inhomogeneous network, and some small open
spaces unevenly distributed on the surface (Fig. 11). It indi-
cated that the crosslinked interactions between proteins in

Fig. 10 Typical scanning electron microscopy (SEM) of soy

Fig. 9 Typical scanning electron microscopy (SEM) of dairy cheese spread sample SCS-A, which was made by GDL
cheese spread sample DCCS: magnitude 5000; at 15 kV. method and modied by papain; magnitude 5000; at
Bar = 5 m. 15 kV. Bar = 5 m.
 food and bioproducts processing 9 1 ( 2 0 1 3 ) 429439 437

Fig. 11 Scanning electron microscopy (SEM) of the soy
cheese spread sample SCS-B, which was made by lactic
acid bacteria fermentation method and modied by papain;
magnitude 5000 at 15 kV. Bar = 5 m.
Fig. 13 Scanning electron micrograph (SEM) of the soy
cheese spread sample SCS-D, which was made by
combined use of GDL and lactic acid bacteria fermentation
methods but not modied by papain; magnitude 5000 at
15 kV. Bar = 5 m.

in the microstructure of the sample SCS-D, which resulted in

an irregular and porous network. Furthermore, many aggre-
gated protein particles appeared in the microstructure of the
sample SCS-D, implying that sample SCS-C tended to own a
compact, spreadable, and homogeneous texture whereas the
sample SCS-D tended to have an inhomogeneous texture with
some minute particulate matters in it.

3.5. Sensory prole

The mean score attributed to each of the parameters

Fig. 12 Scanning electron microscopy (SEM) of the soy
cheese spread sample SCS-C, which was made by
combined use of GDL and lactic acid bacteria fermentation
methods and modied by papain; magnitude 5000; at
15 kV. Bar = 5 m.
the sample SCS-B was weak, leading to its poor textural prop-
erties, low fat and protein retention capacity. According to
Figs. 12 and 13, both the samples SCS-C and SCS-D had good
crosslinked protein network structures, however the sample
SCS-C had a more homogeneous and compact protein net-
work. It was also observed that a lot of open spaces occurred
evaluated: appearance, color, avor, creaminess, rmness,
spread-ability and overall impression is shown in Table 3.
Both the DCCS and SCS samples received mean scores
above 6.0 for all the attributes evaluated, evidencing that they
were all well accepted by the panelists. Comparing to the DCCS
sample, all the SCS samples presented lower score of overall
impression by the panelists. However, the SCS-C samples was
signicantly (P < 0.05) preferred than the other SCS products
by the panelists, indicating the soy cheese spread produced by
the combination of GDL and lactic acid bacteria fermentation
process was more accepted. The SCS-B sample received lower
mean scores of all features than those of the SCS-A sample,
indicating that the soy cheese spread made by GDL method
got less sensory acceptance.
There were no signicant differences (P > 0.05) between the
SCS-D and DCCS samples with respect to color and rmness.

Table 3 Mean scores for the attributes of appearance, color, avor, creaminess, rmness, spread-ability and overall
impression for the DCCS and SCS samples.
Parameters Samples

SCS-A SCS-B SCS-C SCS-D DCCS

Appearance 7.1 0.6a 7.0 0.6a 7.3 0.7b 6.8 0.7c 7.5 0.5d
Color 6.9 0.4b 6.8 0.6a 7.2 0.6c 7.4 0.5d 7.5 0.4d
Flavor 6.9 0.6a 6.7 0.7a 7.2 0.8b 6.5 0.9c 7.8 0.6d
Creaminess 7.3 0.5a 7.1 0.7c 7.5 0.5b 7.2 0.8d 7.5 0.4b
Firmness 6.5 0.6b 6.0 0.7c 6.3 0.5a 6.1 0.6d 6.2 0.5d
Spread-ability 7.1 0.6c 6.9 0.6c 7.5 0.5b 6.3 0.5a 8.0 0.6d
Overall impression 6.8 0.6b 6.6 0.7b 7.4 0.5c 6.3 0.5a 7.7 0.5d

Values are means standard errors; n = 10. Means pertaining to the same parameter (i.e. in the same row), with the same superscript (ad), do
not differ signicantly (P > 0.05) from each other.
438 food and bioproducts processing 9 1 ( 2 0 1 3 ) 429439
However, for the attributes of appearance, avor and spread-
ability, the SCS-D sample achieved signicantly (P < 0.05) lower
scores than the DCCS sample and the other three SCS samples.
Considering the SCS-D was observed an inhomogeneous tex-
ture with some minute particulate matters inside and lack of
the enzymatic hydrolysis process, it could be concluded that
the application of enzymatic hydrolysis was useful to improve
the sensory properties of the SCS samples.
4. Conclusions
Different coagulation processes in the manufacture of the
SCS samples resulted in signicant differences in textural
and sensory properties. According to the present work, the
SCS samples did not get the same overall impressions as the
DCCS sample by the panelists. However, the SCS-C sample,
which was made by combined GDL and lactic acid bacteria
fermentation methods, exhibited a more stable and less eas-
ily fractured structural system and achieved higher scores of
sensory acceptance than other SCS samples. Moreover, the
ureaSDS-PAGE and sensory analysis indicated that enzymatic
hydrolysis process was able to degrade the large soy protein
molecules into smaller fragments to resolve the problem of
grainy texture occurred in the soy cheese spread, and improve
the stability and sensory acceptance. Rheological studies
demonstrated that the soy cheese spread samples developed
in present study exhibited a solidlike behavior. Moreover,
chemical analysis showed that the soy cheese spread con-
tained more protein than the DCCS, which prospect the soy
cheese spread as a nutritional food for the people used to
vegetarian food or foods made from soybean.
Acknowledgments
Part of this work was supported nancially by Unigreen Food
Pty Ltd., Australia. Authors also wished to thank Danisco
(China) Co. Ltd. (Shanghai, China) for supplying starter culture
and other food ingredients.
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