International Dairy Journal 137 (2023) 105519

Contents lists available at ScienceDirect

International Dairy Journal

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

The effects of the use of whey protein as a fat replacer on the

composition, proteolysis, textural, meltability, microstructural, and
sensory properties of reduced-fat Boru-type Künefe cheese during
storage
Mustafa Kadir Esen a, *, Nuray Güzeler b
a
Food Technology Pr., Department of Food Processing, Vocational School of Mut, Mersin University, Mersin, Turkey
b
Department of Food Engineering, Faculty of Engineering, Cukurova University, Adana, Turkey

a r t i c l e i n f o a b s t r a c t

Article history: Boru-type Künefe cheeses were produced from standardised cow milk (fat ratios: 2, 1.5, 1, 0.5%) using
Received 9 April 2022 whey protein concentrates (whey protein ratios: 0, 0.5, 1, 1.5%, respectively) and stored at 4
C for 90
Received in revised form days. The composition, proteolysis, texture, meltability, microstructure, and sensory properties of
29 September 2022
cheeses were examined during storage. The effects of using different fat and whey protein ratios were
Accepted 4 October 2022
Available online 8 October 2022
determined to be significant (p <0.05) on the composition (except pH), texture (except springiness and
cohesiveness), meltability and sensory properties of cheeses. Storage period affected significant (p <0.05)
pH, titration acidity, fat content, meltability values, odour score, hardness, texture (except springiness)
and proteolysis properties. It was determined that there were differences in the microstructure of the
cheeses. Overall, the cheeses produced with 0.5% whey protein and 1.5% fat milk were similar to the
control cheese and determined the most suitable cheese in terms of some properties.
© 2022 Elsevier Ltd. All rights reserved.

1. Introduction
Various local cheeses are produced in Turkey with different
traditional production techniques depending on many factors
(Tekinşen & Elmalı, 2006). One of the important cheese types in the
Eastern Mediterranean Region of Turkey is Künefe cheese (Karaca &
Ocak, 2016). Künefe cheese is generally produced from goats' milk,
sometimes cows' milk or a mixture of both, and is consumed fresh
and without salt (Karaca & Kırdar, 2010, 2016). Its name is Künefe
cheese because this cheese is used in the Künefe dessert in
southern provinces (Kamber & Terzi, 2008). In addition, Künefe is
the name of a special dessert in this region, made by Tel Kadayıf (a
sweet pastry with a texture like that of shredded phyllo dough)
(Karaca & Ocak, 2016). Another type of Künefe cheese is Boru-type
Künefe cheese, which is dry cooked by adding emulsifying salts to
the curd. Applying heat treatment to the curd prevents the cheese
from containing pathogenic microorganisms and increases its
storage. Therefore, Boru-type Künefe cheese is preferred in Künefe
* Corresponding author.
E-mail address: mkesen@mersin.edu.tr (M.K. Esen).
desserts (M. S. Çayır & D. Taş, personal communication, June 15,
2019).
It is stated that low-calorie and low-fat food markets show
dynamic growth due to the increasing demand for low-fat food
products (Chavan, Khedkar, & Bhatt, 2016). Because of the growing
consumer trend for low-fat products, the production of reduced- or
low-fat cheeses has substantially increased since 1980 (Koca &


Metin, 2004; Molina, Alvarez, Ramos, Olano, & Lo
pez-Fandin
~ o,
2000). In cheese, removal or reduction of fat causes textural,
functional and flavour defects such as bland, firm, rubbery body,
poor meltability, and undesirable colour (Akın & Kırmacı, 2015;
Koca & Metin, 2004; McMahon, Alleyne, Fife, & Oberg, 1996; Mistry,
2001; Romeih, Michaelidou, Biliaderis, & Zerfiridis, 2002;
Sipahioglu, Alvarez, & Solano-Lopez, 1999). The use of fat replacers
in cheese production could solve these defects by softening the
body of reduced-fat cheeses and improving their texture, functional
properties, and yield (Nateghi et al., 2012). Therefore, fat replacers
are used to replace fat fully or partially in reduced-fat foods (Drake,
Boylston, & Swanson, 1996; Kavas, Oysun, Ozer, & Uysal, 2004).
Whey proteins have been considered a fat replacer due to their
high nutritional value, biological and functional properties, and
various performance properties similar to fat (Hanafy, Ghanimah,

https://doi.org/10.1016/j.idairyj.2022.105519
0958-6946/© 2022 Elsevier Ltd. All rights reserved.
M.K. Esen and N. Güzeler
Hassanein, & Hashim, 2016; Vidigal et al., 2012). Whey protein has
potential functional benefits such as high solubility, water binding
capacity, emulsification, stabilisation, good appearance, taste and
texture (Hanafy et al., 2016; Korhonen, Pihlanto-Lepp€ €,
ala
Rantama €ki, & Tupasela, 1998). As to nutritional benefits, whey
protein can decrease the energy content of foods when used as a fat
substitute, raise the protein level, and balance the amino acid
profile (Korhonen et al., 1998). There has been no study on using fat
replacers in Boru-type Künefe cheese.
This study determined the effects of using whey protein as a fat
replacer on the composition, proteolysis, textural, meltability,
microstructural, and sensory properties of reduced-fat Boru-type
Künefe cheese during storage. Further, whey was evaluated to
reduce or eliminate defects in reduced fat Boru-type Künefe
cheeses and obtain a product with properties close to fresh Künefe
cheeses and ensure that consumers would consume a healthy,
nutritious, and dietary cheese.
2. Material and methods
2.1. Materials
Raw cows' milk, pH 6.76 ± 0.01, with the following gross
composition 12.21 ± 0.60% (w/v) dry matter, 4.03 ± 0.67% (w/v) fat,
2.92 ± 0.03% (w/v) protein and 0.18 ± 0.01% (w/v) lactic acid was
supplied by the Animal Husbandry section of the Agriculture Fac-
ulty, Cukurova University (Adana, Turkey). Skimmed whey protein
concentrate powder (WPC35) containing 35% protein was obtained
from Enka Milk and Food Products Inc. (Konya, Turkey). Fermen-
tation was achieved using cheese rennet [1/16,000 MCU mL1
(200 IMCU mL1), liquid form, chymosin/pepsin ratio:  85/15;
_
Mysecoren, Maysa Gida, Istanbul, Turkey]. Cooking was achieved
using emulsifying salts (Kasomel 3112 and Kasomel 2185), whose
trademark is Kasomel salts contain various phosphate and citrate
_
salt combinations (Maysa Gida, Istanbul, Turkey).
2.2. Cheesemaking
Three independent Boru-type Künefe Cheese production trials
were carried out at Research and Application Farm, Food Branch,
Milk and Dairy Products Business, Faculty of Agriculture, Çukurova
University.
Complete replacement of fat with whey protein powder in
cheese can cause certain defects in cheese production. For this
reason, in the study, four different cheese types were produced by
applying the fat and whey protein ratios at certain rates, and with
the aim that the sum of total dry matters of the milk was the same
in all milk types.
Raw milk with 4.03 ± 0.67% (w/v) fat content was divided into
four groups for standardisation after it arrived at the factory. The fat
content of milk was standardised to 2% (A) for full-fat and 1.5% (B),
1% (C), 0.5% (D) by adding skim milk for fat-reduced cheese pro-
duction. The first group (A) were the control samples, and whey
protein concentrate was not added. Whey protein concentrate was
added at 0.5%, 1% and 1.5%, respectively, to groups B, C and D.
Dispersion of whey proteins in milk was according to McMahon
et al. (1996). Whey protein concentrate were dispersed in a
portion (approximately 30%) of the milk at 4
C. Milk was heated to
80
C for 10 min and then cooled to 30
C; then the remainder of the
milk was added. After the mixing process, cheese rennet was added
to each batch and left to curdle for 40 min at 33 ± 1
C. Then, the
curd was cut with a curd knife in the shape of 2 cm and let stand
until draining of whey. The cheese curd was pressed and fermented
for 1e2 h until it reached pH 5.4e5.5. The resulting cheese was
named fresh Künefe cheese. Then, it was taken to the dry cooking
2
International Dairy Journal 137 (2023) 105519
machine, 0.5% Kasomel 3112 and 0.5% Kasomel 2185 emulsifying
salts were added, and the cooking process was applied at 70 ± 1
C
for 15 min. After the cooking process, the cheeses were put into
cylindrical plastic moulds. After the cheese rested, they were vac-
uum and shrink packaged in Cryovac bags and stored at þ4
C for
90 days (a flow diagram is given in Supplementary material Fig. S1).
Cheese analyses were carried out at Cukurova University Faculty
of Agriculture, Department of Food Engineering, Dairy Technology
Research Laboratory and Çukurova University Central Research
Laboratory.
2.3. Raw cow milk and whey analyses
The pH values of raw cow milk and whey samples were deter-
mined using a pH meter (Model: WTW pH3110 Set 2 type; Weil-
heim, Germany). Acidity determination was made according to the
alkali titration method, and results were determined in terms of %
lactic acid. The total solid content of the samples (%) was deter-
mined by the gravimetric method (TS, 2002), fat content by the
Gerber method (IDF, 2008) and protein content by the Kjeldahl
method (IDF, 2014).
2.4. Cheese analyses
2.4.1. Composition and proteolysis
Cheese yield performance was calculated as the amount of
cheese (kg) produced from 100 kg of milk (Koçak, 2007). Compo-
sition of cheeses was measured on days 1, 15, 45 and 90. The pH of
grated cheese (10 g) homogenised with 10 mL of distilled water was
measured using a pH meter (Inolab pH 720, WTW GmbH, Weil-
heim, Germany) (Hannon et al., 2003). Titration acidity was
determined by alkaline titration (TS, 1995). The total solid content
of the cheeses (%) was determined by the gravimetric method (IDF,
2004). Fat content was determined by the Gerber method (IDF,
2008). Fat content in dry matter was calculated as follows:
Fat content in dry matter (%) ¼ [fat (%)/dry matter (%)]  100.
Total nitrogen and protein content (total N  6.38) was deter-
mined by the Kjeldahl method (IDF, 2014). Protein content in dry
matter was calculated as follows:
Protein content in dry matter (%) ¼ [protein (%)/dry matter
(%)]  100.
The water-soluble nitrogen (WSN) ratio was also determined
according to the Kjeldahl method described in Kuchroo and Fox
(1982) and Hayaloglu (2003). The ripening degree was calculated
as (WSN/TN)  100. Total free amino acid ratios of cheeses were
determined spectrophotometrically by the Cd-ninhydrin method
according to the method specified by Doi, Daisuke, and Matoba
(1981) and the method applied by Folkertsma and Fox (1992).
2.4.2. Textural properties
Textural properties of the cheeses were evaluated using a
TA.XTPlus Texture Analyser (Stable Micro Systems Ltd., Godalming,
Surrey, UK) on the 1st, 15th, 45th and 90th days of storage. For
texture profile analysis (TPA), cheese samples were cut into cylin-
ders of 10 mm and 25 mm in diameter, wrapped in plastic and
allowed to come to room temperature (±20
C). Test conditions; P/
36 cylinder stainless steel probe; test speed 1 mm s1; pre-test
speed 5 mm s1, post-test speed 1 mm s1; compression (strain)
25%; time pause was applied as 5 s. From the resulting graphs, the
values for hardness, cohesiveness, springiness and chewiness of
cheese blocks were analysed using the Exponent Stable Micro
M.K. Esen and N. Güzeler
Systems Version 6.1.16.0 equipment software (Stable Micro Systems
lu, 2002).
Ltd) (Kahyaog
2.4.3. Meltability
Meltability of the cheeses was determined on the 1st, 15th, 45th
and 90th days of storage according to the method of Poduval and
Mistry (1999). For this purpose, 10 g grated Boru-type Künefe
cheese was placed in a test tube (32  250 mm) and packed to form
a plug at the bottom. After marking the height of the sample, the
tube was covered using aluminium foil, and several holes were
opened to allow hot gas to escape during heating. The tubes were
placed vertically in a refrigerator at 4
C for 30 min. Then, they were
placed horizontally in an oven heated at 104
C for 60 min. After the
tubes were removed from the oven, they were left to cool for
30 min at room temperature on a smooth surface. Then, the cheese
flow distances were measured as meltability in mm.
2.4.4. Microstructure
Microstructures of the cheeses were determined using a scan-
ning electron microscopy (Quanta 650 Field Emission SEM, FEI,
Hillsboro, Oregon, USA) on the 1st and 90th days of storage. Cheese
samples were cut into small blocks (1 cm3) and left for 1 night in a
3.0% glutaraldehyde solution, prepared 0.1 M with sodium phos-
phate buffer (pH: 7). After washing four times with buffer solution,
it was post-fixed in 1% osmium tetroxide for 1 h, then washed with
buffer solution four times, and then dehydrated with a graded
aqueous ethanol series (40, 50, 60, 70, 80, 90, and 100%). The dried
samples were adhered to staples with carbon-coated double-sided
tape. Finally, samples were coated with gold-palladium (the “Bal-
Tec SDC 050” instrument (Balzers, Liechtenstein)) and examined
with a scanning electron microscope (Quanta 650 Field Emission
SEM) (Hayat, 1981; Şanlı, Sezgin, Şenel, & Benli, 2011).
2.4.5. Sensory evaluation
The cheeses were evaluated by 11 trained panellists (6 females,
5 males between 25 and 50 years of age) on days 1, 15, 45 and 90 for
appearance (scale 0e5), texture (scale 0e5), odour (scale 0e5),
taste (scale 0e5) and overall acceptability (scale 0e20). The sensory
evaluation form used was adapted from TS (1989) and Mutluer
(2007).
2.5. Statistical analysis
An analysis of variance (ANOVA) of SPSS statistics software
(SPSS package program, version 22.0, SPSS Inc., Chicago, IL, USA)
was used for the statistical analysis of data. Differences among the
treatments were statistically measured by Duncan's multiple range
tests. Analysis was applied for 1, 15, 45 and 90 days of storage. The
results were evaluated as significant at p <0.05 (Guo, Xie, Wang,
Zou, & Tang, 2018).
3. Results and discussion
3.1. Chemical characteristics of raw cow milk and whey
The chemical properties of the raw cow milk and whey samples
are shown in Table 1. There was no significant difference among
treatments for pH, titratable acidity values, and total solid content
(p >0.05). Significant differences were observed in the fat and
protein content of the raw cow milk and whey samples (p <0.05).
Since the fat contents were adjusted by standardisation, the fat
contents were generally determined at the desired level. In addi-
tion, total dry matter in the milk reached its target and therefore
there were no significant differences among samples. It was
observed that the protein contents of raw cows' milk increased in
3
International Dairy Journal 137 (2023) 105519
direct proportion to the added whey protein concentrate ratio. As
the ratio of whey protein concentrate added to raw milk increased,
the protein content in whey also increased, although significant
differences were only found between the whey for cheeses A and D.
Similar observations to the present study have been obtained by
Aykut (2003) and Kırmacı (2006) for a Feta type cheese, and White
cheese, respectively.
3.2. Compositional properties and proteolysis measurements of the
cheeses
During cheese production, approximately 24% of milk protein is
lost in the whey (whey protein) (Lucey & Kelly, 1994). Improvement
in cheese yield may be attributed to adding whey protein disper-
sions to milk (Hanafy et al., 2016). The cheese yield was between 8
and 9% for all cheeses (Table 2). There were significant differences
in cheese yields among treatments. Cheese yield generally de-
creases in reduced-fat cheese production (Anvari & Joyner, 2018). If
the fat content was kept constant and only whey protein was added
to the cheese, an increase in cheese yield would be expected.
However, as the fat content was reduced and whey protein was
added, there were irregular changes in the cheese yield. Extra
lactose from WPC was not expected to be incorporated into the final
cheese, therefore, the decrease in final cheese yields may be due to
this. In addition, the data presented suggest that the yields are
diminished when WPC replaces fat, resulting in a negative eco-
nomic impact, but only at high levels of replacement, because only
the yield for D cheese was significantly lower than A cheese.
Compositional properties and proteolysis measurements of
Boru-type Künefe cheeses are shown in Table 2. Results for pH were
similar for all cheeses at each storage time. The pH values of all
cheeses decreased during storage; however, this was significant
only for cheeses A and B (p <0.05). The decrease in pH values of the
cheeses during storage was similar to other studies (El-Nimer,
Omar, Ahmed, & Mostafa, 2017; Nazari, Mortazavi, Hesari, &
Tabatabaei Yazdi, 2019).
Titration acidity gives the total amount of acid, but pH indicates
the strength of acidity. It is known that these two methods are
inversely related to each other (Hayalog lu, Topçu, & Koca, 2011).
The use of different fat and whey protein ratios was significant in
the titration values of the cheeses at the 90th day of storage (p
<0.05); the TA was higher in the D cheese than in the rest of
cheeses. It has been reported that a decrease in fat content, as well
as an increase in protein-based replacer, increases titration acidity
€
in cheeses (El-Sonbaty & El-Sisey, 2010; Oztekin, 2003). Changes in
titration acidity values between the 1st and 90th of the storage in
all samples were statistically significant (p <0.05). The increase in
titratable acidity was likely caused by organic acid formation pri-
marily by lactic acid producing bacteria which increased during
storage (Di Pierro, Sorrentino, Mariniello, Giosafatto, & Porta, 2011).
El-Aidie, Ghita, El-Dieb, and El-Garhi (2019) obtained similar re-
sults to the present study for low-fat Edam cheese.
There were differences (p <0.05) in the total solid content, be-
tween cheeses A and D due to the use of different fat and whey
protein ratios. It was determined that cheese without whey protein
added had the highest total solid contents on all days of storage. In
traditional cheese-making, whey proteins are not typically bound
up in the cheese matrix and so whey proteins are lost in the whey.
However, whey proteins may be incorporated in a native form or
denatured state into cheese (Hinrichs, 2001). Fat replacers are
known for their water-binding capacity, which may in turn explain
the lower total solids content found in the cheeses containing whey
protein than in cheeses without added whey protein (Kavas et al.,
2004). One of the most important strategies for improving the
functional properties of low-fat cheese is to increase its moisture
M.K. Esen and N. Güzeler International Dairy Journal 137 (2023) 105519

Table 1
Composition of the raw cow milk and whey from the cheeses.a

Component pH TA (LA%) DM (%) Fat (%) Protein (%)

Raw milk
PA 6.78 ± 0.15a 0.17 ± 0.02a 10.19 ± 0.17a 2.12 ± 0.13a 2.93 ± 0.03b
PB 6.71 ± 0.10a 0.17 ± 0.01a 10.14 ± 0.11a 1.60 ± 0.20b 3.20 ± 0.45ab
PC 6.80 ± 0.01a 0.17 ± 0.02a 10.18 ± 0.01a 1.08 ± 0.08c 3.57 ± 0.11a
PD 6.67 ± 0.09a 0.17 ± 0.02a 9.95 ± 0.31a 0.58 ± 0.28d 3.62 ± 0.33a
Whey
WA 6.17 ± 0.05a 0.12 ± 0.02a 7.13 ± 0.39a 0.78 ± 0.08a 0.88 ± 0.08b
WB 6.20 ± 0.27a 0.14 ± 0.17a 7.18 ± 0.23a 0.68 ± 0.02a 1.17 ± 0.13ab
WC 6.23 ± 0.20a 0.14 ± 0.01a 7.28 ± 0.15a 0.62 ± 0.03a 1.20 ± 0.30ab
WD 6.20 ± 0.08a 0.12 ± 0.02a 7.15 ± 0.32a 0.38 ± 0.16b 1.42 ± 0.34a
a
Abbreviations are: PA, raw cow milk standardised to 2% fat and added 0% whey protein concentrate, PB, raw cow milk standardised to 1.5% fat and added 0.5% whey protein
concentrate, PC, raw cow milk standardised to 1% fat and added 1% whey protein concentrate, PD, raw cow milk standardised to 0.5% fat and added 1.5% whey protein
concentrate; WA, whey from cheese A; WB, whey from cheese B; WC, whey from cheese C; WD, whey from cheese D; DM, dry matter; TA, titratable acidity in terms of % lactic
acid. Means in the same column followed by different superscript lowercase letters were significantly different p <0.05.

Table 2
Compositional properties and proteolysis measurements of Boru-type Künefe cheeses during storage.a

Properties Cheese Day

1 15 45 90

Cheese yield (%) A 9.40 ± 0.10ab null null null

B 9.12 ± 0.10bc null null null
C 9.69 ± 0.17a null null null
D 8.90 ± 0.26c null null null
pH A 5.76 ± 0.05aK 5.73 ± 0.04aK 5.70 ± 0.04aKL 5.62 ± 0.04aL
B 5.73 ± 0.07aK 5.70 ± 0.06aKL 5.68 ± 0.05aKL 5.61 ± 0.05aL
C 5.69 ± 0.08aK 5.67 ± 0.08aK 5.65 ± 0.08aK 5.58 ± 0.06aK
D 5.68 ± 0.08aK 5.66 ± 0.08aK 5.63 ± 0.08aK 5.57 ± 0.06aK
TA (LA%) A 1.05 ± 0.23aL 1.33 ± 0.12aK 1.36 ± 0.11aK 1.49 ± 0.02bK
B 1.08 ± 0.23aL 1.42 ± 0.18aK 1.45 ± 0.18aK 1.51 ± 0.01bK
C 1.18 ± 0.26aL 1.50 ± 0.13aKL 1.48 ± 0.16aKL 1.52 ± 0.02bK
D 1.20 ± 0.32aL 1.53 ± 0.15aKL 1.54 ± 0.12aKL 1.60 ± 0.07aK
TSC (%) A 44.93 ± 1.45aK 45.09 ± 1.10aK 43.39 ± 1.84aK 43.06 ± 1.29aK
B 41.51 ± 2.90abK 43.40 ± 1.93aK 41.89 ± 1.53aK 39.51 ± 2.65abK
C 41.58 ± 0.98abK 41.92 ± 0.81abK 41.25 ± 0.88aK 40.83 ± 1.08abK
D 37.77 ± 3.96bK 39.12 ± 2.95bK 41.48 ± 1.38aK 36.82 ± 3.67bK
Fat (%) A 14.42 ± 0.63aK 13.67 ± 0.38aKL 13.17 ± 1.01aL 13.00 ± 0.00aL
B 10.58 ± 1.13bK 9.92 ± 0.72bK 10.33 ± 1.18bK 9.42 ± 0.14bK
C 7.67 ± 0.63cK 7.17 ± 1.01cK 7.08 ± 1.01cK 7.75 ± 0.43cK
D 4.00 ± 2.25dK 3.83 ± 2.52dK 3.67 ± 2.27dK 4.92 ± 1.23dK
Fat in DM (%) A 32.14 ± 2.43aK 30.33 ± 1.40aK 30.33 ± 1.65aK 30.21 ± 0.91aK
B 25.66 ± 4.19abK 22.88 ± 2.02bK 24.63 ± 2.22aK 23.91 ± 1.72bK
C 18.45 ± 1.54bK 17.07 ± 2.13bK 17.16 ± 2.27bK 18.99 ± 1.29cK
D 10.44 ± 5.73cK 9.78 ± 6.59cK 8.97 ± 5.73cK 13.60 ± 4.27dK
TN A 4.06 ± 0.05cM 4.44 ± 0.08abL 4.39 ± 0.08bL 4.61 ± 0.10aK
B 4.29 ± 0.06bK 4.32 ± 0.08bK 4.25 ± 0.13bK 4.40 ± 0.20aK
C 4.50 ± 0.10aM 4.54 ± 0.08aLM 4.76 ± 0.06aK 4.65 ± 0.05aKL
D 4.30 ± 0.10bL 4.45 ± 0.05abKL 4.66 ± 0.15aK 4.64 ± 0.13aK
Protein (%) A 25.88 ± 0.29cM 28.35 ± 0.52abL 28.01 ± 0.51bL 29.41 ± 0.62aK
B 27.39 ± 0.38bK 27.54 ± 0.49bK 27.14 ± 0.83bK 28.07 ± 1.28aK
C 28.71 ± 0.64aM 28.97 ± 0.50aLM 30.35 ± 0.38aK 29.67 ± 0.32aKL
D 27.43 ± 0.64bL 28.37 ± 0.29abKL 29.73 ± 0.93aK 29.62 ± 0.84aK
Protein in DM (%) A 57.64 ± 1.55bM 62.90 ± 2.57bL 64.64 ± 3.63bKL 68.32 ± 0.86bK
B 66.16 ± 3.72aKL 63.55 ± 3.40bL 64.80 ± 1.92bL 71.11 ± 1.62bK
C 69.06 ± 1.07aL 69.10 ± 0.38abL 73.59 ± 1.11aK 72.69 ± 1.90bK
D 73.05 ± 5.99aK 72.76 ± 4.64aK 71.70 ± 2.32aK 80.85 ± 5.98aK
WSN (%) A 0.19 ± 0.02cK 0.21 ± 0.01cK 0.20 ± 0.01cK null
B 0.22 ± 0.01abM 0.28 ± 0.01bL 0.48 ± 0.02aK null
C 0.20 ± 0.01bcM 0.28 ± 0.01bL 0.49 ± 0.01aK null
D 0.24 ± 0.01aM 0.32 ± 0.01aL 0.43 ± 0.02bK null
RD (%) A 4.77 ± 0.39bcK 4.68 ± 0.30cK 4.59 ± 0.26dK null
B 5.16 ± 0.26abM 6.53 ± 0.06bL 11.17 ± 0.25aK null
C 4.51 ± 0.17cM 6.24 ± 0.15bL 10.35 ± 0.16bK null
D 5.53 ± 0.08aM 7.23 ± 0.15aL 9.27 ± 0.33cK null
TFAA (Abs 507 nm) A 0.086 ± 0.008dN 0.134 ± 0.005cM 0.175 ± 0.004bL 0.197 ± 0.004cK
B 0.104 ± 0.006cN 0.138 ± 0.003cM 0.176 ± 0.004bL 0.211 ± 0.006bK
C 0.134 ± 0.006bN 0.158 ± 0.008bM 0.185 ± 0.007bL 0.232 ± 0.007aK
D 0.170 ± 0.004aN 0.193 ± 0.005aM 0.213 ± 0.009aL 0.234 ± 0.002aK
a
Abbreviations are: TSC, total solid content; TA, titratable acidity in terms of % lactic acid; TN, total nitrogen; WSN, water-soluble nitrogen; RD, ripening degree; TFAA, total
free amino acids. Boru-type Künefe cheeses were made with: A, 2% fat milk (control); B, 0.5% whey protein and 1.5% fat milk; C, 1% whey protein and 1% fat milk; D, 1.5% whey
protein and 0.5% fat milk. Means in the same column or row followed by different superscript lowercase or uppercase letters, respectively, were significantly different p <0.05.

4
M.K. Esen and N. Güzeler
content sufficiently to provide a ratio of moisture to protein in the
lower fat cheese that is equal or higher than its full-fat counterpart
(Broadbent, McMahon, Oberg, & Welker, 2001; Koca & Metin,
2004). Similar results to the present study have been obtained by
Akın and Kırmacı (2015), El-Nimer et al. (2017) and El-Salam (2015).
The fat content of cheese milk affected (p <0.05) the fat content
and fat content in dry matter of cheeses. It was found that the
decrease in the fat content and fat content in the dry matter of the
cheeses compared with the fat content in the raw milk was at the
desired level.
The use of different fat and whey protein ratios significantly
influenced total nitrogen and protein contents of the cheeses on the
1st, 15th and 45th days of storage (p <0.05). The total nitrogen and
protein contents of cheeses C and D were statistically different from
each other only on the 1st day of storage (p <0.05), while the values
were similar between these cheeses for the other days of storage.
The protein content in dry matter in the cheese D was significantly
different to that of control cheese (A) at all days of storage (p <0.05).
In addition, protein in dry matter of cheeses C and D were close to
each other on the 1st, 15th and 45th days of storage (p >0.05).
Higher protein level in the cheese would be expected due to the
incorporation of the protein-based fat mimetic into the cheese
matrix (Drake et al., 1996). It was observed that whey protein
concentrates positively contributed to the total nitrogen, protein,
and protein content in the dry matter of the cheeses.
The effects of storage time on the total nitrogen and protein
content of cheeses were statistically significant in the cheese
samples except for cheese B (p <0.05). It was observed that the
protein content in dry matter of cheeses generally increased (p
<0.05). Although there were minor changes (p >0.05) in the total
solid content during storage, it significantly affected the protein
content in dry matter of cheeses (p <0.05).
It was observed that as the fat content of the cheeses decreased,
the water-soluble nitrogen (WSN) were higher than the control
sample (p <0.05) and the WSN of the cheeses increased (p <0.05)
during storage (except cheese A). In a study on white-brined
cheese, an increasing trend was observed in WSN production in
cheeses made with reduced-fat milk. The activity of rennet and/or
starter or non-starter proteolytic enzymes may be altering the
compositional effects of low-fat cheeses (Romeih et al., 2002). In a
study, it was reported that the reason for the increase in water-
soluble nitrogen up to the 15th day during ripening and then its
decrease might be that the water-soluble nitrogen in cheese has
passed into brine (Aykut, 2003). Depending on proteolysis, water-
soluble protein continuously increases during the ripening period
€
(Oner , 2018). The increase in water-soluble nitrogen ra-
& Sarıdag
tios during storage was also observed in other studies (Akın &
Kırmacı, 2015; Basiony, El-Nimer, & El-Gandour, 2017). Since the
degree of ripening of the cheeses was calculated by proportioning
the total amount of water-soluble nitrogen to the total amount of
nitrogen, the results for RD were similar to that obtained for WSN. It
can be said that the concomitant reduction of fat and addition of
whey protein concentrate in cheeses increases the degree of
ripening (p <0.05). Considering that the raw materials and com-
positions used in the production of cheeses are different, it can be
said that the difference in ripening degree is expected (Ayar, 1996;
Bayram, 2018). The effects of storage time on the ripening degrees
of the cheeses were found to be statistically significant in all sam-
ples (p <0.05). The increase in ripening degrees is generally a result
of the activity of proteases and proteinases synthesised by micro-
organisms or enzymes naturally present or intentionality added
into the milk (Ayar, 1996).
It was observed that total free amino acid values increased (p
<0.05) with the increase of whey protein rates in cheeses, and the
total free amino acid values of the cheeses increased (p <0.05)
5
International Dairy Journal 137 (2023) 105519
during storage. Free amino acids (FAAs) are the end products of
proteolysis released from peptides by the action of peptidases. They
accumulate during ripening and subsequently are then metab-
olised, to varying extent by the enzymes of the cheese microflora
into volatile or non-volatile compounds, which contribute to
flavour development (Michaelidou, Katsiari, Voutsinas,
Polychroniadou, & Alichanidis, 2007). Sahan, Yasar, Hayaloglu,
Karaca, and Kaya (2008), in a study on the use of fat replacers,
stated that the total amino acid values of cheeses increased with
the use of fat replacers. Guinee, Auty, and Fenelon (2000) reported
in a study on Cheddar cheeses that the relationship between fat and
ripening time significantly affected free amino acid concentrations,
with the rate of increase of FAAs with time being least in the case of
full-fat Cheddar cheeses and greatest in the case of the low-fat
Cheddar cheeses.
3.3. Textural and meltability properties of the cheeses
The results of the textural properties of the cheeses are shown in
Table 3. There were differences (p <0.05) in the hardness values,
between cheeses A and D due to the use of different amounts of fat
and whey protein. Since fat breaks up the protein matrix and plays
the role of lubricant to provide smoothness and a softer texture
(Romeih et al., 2002), cheese A was generally significantly softer
than the other cheeses during storage (p <0.05). Moisture, protein,
fat content and the pH value of the environment have an important
role in the hardness values of cheeses (Bulut-Solak, 2013). More
non-interrupted protein zones form in the cheese as the fat content
is reduced. As a result, a higher degree of crosslinking of protein
molecules occurs resulting in three-dimensional networks, exhib-
iting high resistance to deformation (Lobato-Calleros et al., 2007).
In the light of this information, the low-fat cheese (D) was the
hardest cheese in line with its high-protein content in dry matter.
In addition, the hardness values of cheese B were found close to the
control cheese. In the literature, it was reported that the use of
different fat replacers decreased the hardness of low-fat cheese
(Akın & Kırmacı, 2015; Koca & Metin, 2004; Romeih et al., 2002).
During storage, there was a regular decrease (p <0.05) in the
hardness values of cheeses C and D, while fluctuations occurred (p
<0.05) in the other cheeses. It is thought that the reason for this
decrease is probably the proteolytic and other enzymatic reactions
that take place in the cheese system (Bulut-Solak, 2013). A study
conducted by Yaşar (2007) stated that the hardness values of
Kashar cheese decreased depending on ripening period. The stor-
age period (15th and 45th day) significantly influenced the
springiness values of cheese A (p <0.05). Topcu, Bulat, and Ozer €
(2020) emphasised that the main factors affecting the springiness
of cheeses are protein hydrolysis, reduction of free water and so-
lidification of milk fat. It was thought that the differences in the
springiness values for the cheese A during storage might be due to
its higher fat content in comparison with the other cheeses.
The addition of different whey protein concentrate amounts to
reduced-fat cheese milk did not statistically affect the cohesiveness
values of the cheeses (p >0.05). Also, the storage time was not
significant on the cohesiveness values of all cheese samples (p
>0.05).
The addition of different whey protein concentrate amounts to
reduced-fat cheese milk caused significant differences (p <0.05) in
chewiness values of the cheeses on the 1st and 45th days of storage.
Kırmacı (2006) stated that chewiness values of cheeses are signif-
icantly different according to the fat content of milk and the use of
fat replacers. While the chewiness value were similar in cheese A (p
>0.05) during storage, it was lower in cheeses C and D on the 90th
day of storage compared with the first day. In different studies, it
was reported that chewiness values of cheeses generally decrease
M.K. Esen and N. Güzeler International Dairy Journal 137 (2023) 105519

Table 3
Textural, meltability and sensory properties of Boru-type Künefe cheeses during storage.a

Properties Cheese Day

1 15 45 90
bKL bK bLM
Hardness (g) A 4146 ± 471 4238 ± 511 3222 ± 579 2949 ± 450aM
B 3834 ± 229bL 4897 ± 559abK 4484 ± 61aK 3404 ± 197aL
C 5310 ± 303aK 5172 ± 477aK 4206 ± 507aL 3383 ± 313aM
D 5657 ± 380aK 5245 ± 170aK 4789 ± 538aK 3635 ± 623aL
Springiness A 0.87 ± 0.04aKL 0.83 ± 0.00aL 0.99 ± 0.14aK 0.88 ± 0.04aKL
B 0.87 ± 0.04aK 0.94 ± 0.19aK 0.93 ± 0.07aK 0.89 ± 0.01aK
C 0.86 ± 0.02aK 0.93 ± 0.15aK 0.88 ± 0.07aK 0.86 ± 0.02aK
D 1.02 ± 0.25aK 0.84 ± 0.02aK 0.90 ± 0.07aK 0.89 ± 0.01aK
Cohesiveness A 0.80 ± 0.04aK 0.78 ± 0.03aK 0.83 ± 0.04aK 0.81 ± 0.02aK
B 0.80 ± 0.04aK 0.80 ± 0.01aK 0.84 ± 0.05aK 0.80 ± 0.00aK
C 0.81 ± 0.02aK 0.78 ± 0.07aK 0.84 ± 0.05aK 0.80 ± 0.01aK
D 0.80 ± 0.03aK 0.75 ± 0.09aK 0.85 ± 0.03aK 0.82 ± 0.01aK
Chewiness (g) A 2881 ± 481bcK 2741 ± 443aK 2614 ± 146bK 2116 ± 486aK
B 2689 ± 116cLM 3671 ± 863aK 3493 ± 363abKL 2417 ± 168aM
C 3699 ± 324abK 3726 ± 511aK 3086 ± 394abK 2344 ± 247aL
D 4553 ± 759aK 3313 ± 366aKL 3690 ± 818aKL 2664 ± 490aL
Meltability (mm) A 96.17 ± 9.31aM 138.83 ± 10.89aK 143.50 ± 3.12aK 120.83 ± 3.82aL
B 93.33 ± 5.11aM 140.67 ± 8.81aK 135.17 ± 7.29aK 107.50 ± 1.32bL
C 70.50 ± 3.28bM 114.83 ± 6.71bK 112.83 ± 6.53bK 87.17 ± 0.29cL
D 68.00 ± 6.08bM 79.87 ± 5.00cL 90.67 ± 5.53cK 71.33 ± 0.58dLM
Appearance A 4.12 ± 0.39aK 4.46 ± 0.13aK 4.53 ± 0.25aK 4.57 ± 0.20aK
B 4.39 ± 0.26aK 4.34 ± 0.08aK 4.55 ± 0.38aK 4.22 ± 0.13bK
C 4.37 ± 0.07aK 3.99 ± 0.28bK 4.39 ± 0.32aK 4.04 ± 0.04bK
D 4.37 ± 0.44aK 4.00 ± 0.07bK 4.13 ± 0.57aK 4.15 ± 0.03bK
Texture A 4.18 ± 0.39aK 4.60 ± 0.23aK 4.28 ± 0.42aK 3.87 ± 0.47aK
B 4.37 ± 0.37aK 4.48 ± 0.04aK 4.45 ± 0.12aK 4.25 ± 0.08aK
C 4.26 ± 0.13aK 3.84 ± 0.35bK 4.33 ± 0.32aK 4.19 ± 0.15aK
D 4.09 ± 0.47aK 4.15 ± 0.24abK 3.90 ± 0.70aK 4.06 ± 0.06aK
Odour A 4.39 ± 0.09aKL 4.76 ± 0.25aK 4.72 ± 0.24aK 4.03 ± 0.38aL
B 4.24 ± 0.49abK 4.63 ± 0.28aK 4.58 ± 0.30aK 4.16 ± 0.09aK
C 4.05 ± 0.28abL 4.55 ± 0.15aK 4.61 ± 0.20aK 3.97 ± 0.13aL
D 3.67 ± 0.17bL 4.62 ± 0.34aK 4.41 ± 0.34aK 4.20 ± 0.13aK
Taste A 4.26 ± 0.36aK 4.63 ± 0.07aK 4.42 ± 0.16aK 3.93 ± 0.57aK
B 4.28 ± 0.47aK 4.55 ± 0.16aK 4.26 ± 0.43aK 3.84 ± 0.77aK
C 3.98 ± 0.30aK 4.44 ± 0.31abK 3.85 ± 0.31aK 4.02 ± 0.36aK
D 3.92 ± 0.10aK 4.02 ± 0.37bK 3.74 ± 0.81aK 3.64 ± 0.09aK
Overall acceptance A 16.95 ± 1.14aK 18.45 ± 0.50aK 17.96 ± 0.92aK 16.41 ± 1.53aK
B 17.28 ± 1.52aK 18.00 ± 0.47aK 17.84 ± 0.95aK 16.47 ± 0.90aK
C 16.67 ± 0.46aK 16.81 ± 0.48bK 17.18 ± 0.95aK 16.22 ± 0.37aK
D 16.06 ± 1.00aK 16.79 ± 0.86bK 16.19 ± 2.08aK 16.06 ± 0.19aK
a
Boru-type Künefe cheeses were made with: A, 2% fat milk (control); B, 0.5% whey protein and 1.5% fat milk; C, 1% whey protein and 1% fat milk; D, 1.5% whey protein and
0.5% fat milk. Means in the same column or row followed by different superscript lowercase or uppercase letters, respectively, were significantly different p <0.05.

during storage and ripening (Cankurt, 2015; Kırmacı, 2006; Yüksel,
2019).
The melting properties of Künefe cheese are very important
because this cheese is used as an ingredient in Künefe desserts in
Turkey. The values for meltability of cheeses during storage are
presented in Table 3. Meltability values of cheeses were significantly
(p <0.05) influenced using different fat and whey protein ratios
during storage. The decrease in meltability was associated with
whey proteins acting as active fillers in the casein network, causing
a stronger mixed gel structure and the fibrous structure these pro-
€
teins form around fat globules (Ozkan, 2012; Şalvarcı, 2015). Pro-
teins do not melt, but their interactions with each other can change
and form an effect that we call melt (Lucey, Johnson, & Horne, 2003).
Similar results were observed for Kashar (Koca & Metin, 2004;
Sahan et al., 2008), and Mozzarella (El-Tahra, Ismail, & El-Metwally,
2008) cheeses. The fact that the meltability values of cheese B
generally were close to control cheese during storage was thought to
support the reduction of the fat ratio in cheeses and the addition of
whey protein concentrate at certain rates. Although the meltability
values showed irregular changes (p <0.05) during storage, an in-
crease was observed in all cheeses at the end of storage compared
with the first day. The increase in cheese meltability is attributed to
proteolysis occurring in cheese during storage. The protein matrix
absorbs moisture held by the fat serum channels during storage.
6
This situation allows the rearrangement of proteins, which increases
hydration around the matrix. Then, as the proteins are hydrated,
they flow more easily when heated, thus increasing the meltability
of the cheeses (Dave, Sharma, & McMahon, 2003). Studies have
reported that the meltability values of different cheese varieties
increase during storage (El-Tahra et al., 2008; Rudan, Barbano, Yun,
& Kindstedt, 1999; Yaşar, 2007).
3.4. Microstructure of the cheeses
The microstructure of cheeses is presented in Fig. 1 (see also
Supplementary material Fig. S2). It was observed that the use of
different fat and whey protein proportions affected the microstruc-
ture of cheeses. It was determined that the size and number of fat
globules spread in the protein matrix decreased compared with the
control cheese. This situation continued as the fat rate in cheese
decreased, and the amount of whey protein concentrate added
increased. It was observed that the protein network from cheese A to
cheese D becomes denser and creates a more compact and softer
image. When cheese D was examined, it was determined that the
protein matrix was more transparent than the other samples and the
fat globules were less. The protein-dominated microstructure of low-
fat cheese forms harder and rubbery cheese. In low-fat cheeses, there
are fewer fat globules within the protein matrix, and the globules are
M.K. Esen and N. Güzeler
Fig. 1. Microstructure on 1st and 90th days of Boru-type Künefe cheeses (magnification, 1000) made with: A, 2% fat milk (control); B, 0.5% whey protein and 1.5% fat milk; C, 1%
whey protein and 1% fat milk; D, 1.5% whey protein and 0.5% fat milk.
usually smaller than in full-fat cheese (Mistry & Anderson, 1993;
Sipahioglu, Alvarez, & Solano-Lopez, 1999). It was observed that the
voids in the microstructures of cheeses A and B decreased on the
90th day of storage and had a flatter structure. Therefore, the size
and number of fat globules in cheeses A and B decreased.
When the microstructures of cheeses C and D were examined,
there was a slight increase in the void size and number on the 90th
7
International Dairy Journal 137 (2023) 105519
day of storage. It was observed that the microstructure of the
cheeses expanded during storage. It was thought that this was due
to increased proteolysis (Oluk, Güven, & Hayaloglu, 2013) and the
resulting weakening of the protein network (Nazari et al., 2019). El-
Aidie et al. (2019) reported in a study on low-fat Edam Cheeses that
some of the void space appeared elongated due to ripening because
of the complete curd fusion owing to the breakdown of the casein
M.K. Esen and N. Güzeler
matrix. El-Nimer et al. (2017) stated in a study of Gouda-like
cheeses that after 90 days of ripening, the microstructure of all
treatments was changed greatly owing to the action of the ripening
agents and altered chemical composition.
3.5. Sensory properties of the cheeses
The sensory panel's assessment of cheese quality during storage
is given in Table 3. The data show that the appearance, texture,
odour, taste and overall acceptance of Boru-type Künefe cheeses
were rarely affected by the whey protein concentrate used in
cheesemaking. The fact that the sensory properties are close to the
control cheese supports fat reduction and the addition of whey
protein concentrate. Cheese with 0.5% whey protein and 1.5% fat
milk scored close to the control cheese in sensory properties as well
as in texture, meltability and microstructure properties. All cheeses
were judged as acceptable products by the panellists during stor-
age. Similar studies reported that different fat replacers improve
the sensory characteristics of various reduced and low-fat cheeses
(Akın & Kırmacı, 2015; Romeih et al., 2002; Sipahioglu et al., 1999;
Zalazar et al., 2002).
4. Conclusions
The results of this study indicated that the reduction of fat and
the use of the whey protein affected the composition, proteolysis,
textural, meltability, microstructural, and sensory properties of
reduced-fat Boru-type Künefe cheese during storage. In this work,
it was verified that the diminution of fat and increase of whey
protein concentrate (especially cheese B) allowed for obtaining a
low-fat cheese similar to full-fat cheese. Except for the control
cheese, the most suitable cheese in terms of some properties was
determined as Boru-type Künefe cheese with 0.5% whey protein
and 1.5% fat milk. Because of consumers' demand for low-calorie
foods, these cheeses may be an alternative to fat cheeses. These
findings may contribute to the dairy industry providing advantages
of Boru-type Künefe cheeses. These advantages are the suitability of
cheeses for industrial production, the more extended storage
period of cheese compared with fresh Künefe cheese, the use of fat
in production for cream production, and the utilisation of whey to
obtain protein. Also, these results will contribute to the reduced-fat
cheese literature. Further research is needed to investigate other fat
replacers in reduced-fat Künefe cheeses. In addition, the charac-
terisation of Boru-type Künefe cheeses, in which only the fat is
decreased (without addition of WPC), would be interesting to verify
the influence of the addition of WPC on the characteristics of fat-
reduced cheeses.
Credit author statement
Mustafa Kadir Esen: Conceptualization, Methodology, Formal
analysis, Investigation, Writing - Original Draft, Visualization.
Nuray Güzeler: Conceptualization, Methodology, Validation,
Resources, Writing - Review & Editing, Supervision.
All authors approved the final manuscript and accepted to be
held responsible for the content.
Declaration of competing interest
None.
8
International Dairy Journal 137 (2023) 105519
Acknowledgements
This study from a doctoral dissertation (Çukurova University)
was supported by Çukurova University, Unit of Scientific Research
Projects, Adana, Turkey (Project number FDK-2019-12145).
Appendix A. Supplementary data
Supplementary data to this article can be found online at
https://doi.org/10.1016/j.idairyj.2022.105519.
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