a

Original Article ISSN 0101-2061 (Print)

Food Science and Technology ISSN 1678-457X (Online)

DOI: https://doi.org/10.1590/fst.64720

Protein quality assessment of commercial whey protein supplements commonly consumed

in Turkey by in vitro protein digestibility-corrected amino acid score (PDCAAS)
Halime PEHLIVANOĞLU1* , Hamide Feyza BARDAKÇI2, Mustafa YAMAN2

Abstract
Whey protein is preferred in sports nutrition because of its rich essential amino acids and protein digestibility rate compared
to other protein sources. The purpose of this study was to determine the amino acid content and in vitro protein digestibility of
whey protein supplements and evaluate their protein quality using the in vitro protein digestibility-corrected amino acid score
(PDCAAS) method. The amount of amino acids were determined using high performance liquid chromatography (HPLC)
and the protein digestibility was determined by in vitro simulated gastrointestinal tract. A high levels of glycine and very low
levels of cysteine were detected in some samples. The ratio of glycine in 4 out of 14 samples ranged between 7.4 and 40.3%. The
ratio of essential amino acids and branched chain amino acids in all samples were less than the reference protein. The in vitro
protein digestibility of whey proteins ranged from 50.4 to 79.6%, which was very low compared with the values indicated in
the literature. The PDCAAS values of whey proteins were very low and ranged from 0.08 to 0.71 in the samples. Based on these
results, manufacturers should revise their processing techniques in order to provide high quality whey protein.
Keywords: whey protein; amino acid; digestibility; quality; PDCAAS.
Practical Application: Protein quality assessment of commercial whey protein supplements by in vitro protein digestibility-
corrected amino acid score (PDCAAS).

1 Introduction
Athletes, bodybuilders, and physically active individuals (Rankin & Darragh, 2006). In addition, the amount of branched
extensively consume protein-rich products such as whey, soy, casein, chain amino acids (BCAAs) (leucine, isoleucine, and valine) in
and egg (Williams, 2005). Among these, whey and casein are the the structure of whey protein is higher than other protein sources
preferred proteins in terms of quality (Tipton et al., 2004). Whey (Pennings et al., 2011; Khanam et al., 2013). Unlike other amino
and casein are mostly derived from cow milk. Cow milk protein acids, BCAAs are not metabolized in the liver and are used directly
consists of 80% casein and 20% whey protein. Whey protein is by skeletal muscle. These BCAAs serve as an effective source of
consisted of β-lactoglobulin (β-LG) and α-lactoglobulin (α-LG) energy for the muscles during exercise as well as a ready source
and a small amount of immunoglobulin (Bobe et al., 1998). β-LG for the synthesis of muscle proteins (Shimomura et al., 2004;
has 162 amino acids and the most distinctive feature of whey Nicastro et al., 2012). Among the BCAAs, leucine also functions
protein compared to casein is that it has 5 cysteines in its structure as a critical regulator of translation initiation in muscle protein
(Sava et al., 2005). This significant difference is used to determine synthesis. Whey protein contains a higher amount of leucine
the quality difference between casein and whey protein (European than casein (Norton & Layman, 2006). Another important
Union, 1990; Ballin, 2006). feature of whey protein compared to other proteins is that it
is easier to digest and absorb in the body. Therefore, it is a
Whey protein is commercially produced from cheese whey.
good source to address immediate protein requirements after
Cheese whey is the liquid resulting from the separation of caseins
exercise or physical activity (Tang et al., 2009). Whey protein is
and milk fat during the cheesemaking process (Alves et al., 2019).
available as hydrolysate, isolate, and concentrate (Lucena et al.,
The quality and nutritional composition of whey depends on the
2007). Whey protein hydrolysate is a type of protein separated
cheese making processes, the type of milk used, environmental
into its own amino acids, although the price is higher than
conditions and animals. The use of cheese whey is of industrial
other proteins because the production method is more costly
interest, due to the large amount produced, and availability of
(Pouliot et al., 1999).
nutrients such as lactose, soluble proteins, lactic acid and B
vitamins (Fangmeier et al., 2019). Therefore, cheese whey can The quality of the protein in a food is evaluated by the rate
be used in the formulation of whey dairy beverages, ricotta, of digestion in the small intestine (Dangin et al., 2001). Protein
whey concentrate and milk blends (Guimarães et al., 2018, digestibility is an important factor when determining the amount
2019;Trindade et al., 2019).Whey protein is especially preferred of protein absorbed in the small intestine after digestion and reflects
in sports nutrition because of its rich essential amino acid (EAA) the effectiveness of the protein’s use in the diet (World Health
content and protein quality compared to other protein sources Organization, 2007). The data regarding the digestibility of proteins

Received 24 Nov., 2020

Accepted 19 Jan., 2021
1
Faculty of Veterinary Medicine, Namik Kemal University, Tekirdağ, Turkey
2
Department of Nutrition and Dietetics, Faculty of Health Sciences, İstanbul Sabahattin Zaim University, İstanbul, Turkey
*Corresponding author: [email protected]

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Original Article
Protein quality of whey protein supplements

are limited due to ethical restrictions. Therefore, a simulated in vitro phosphate dehydrate (NaH2PO4.2H2O) and 0.88 g disodium hydrogen
protein digestibility model is a widely used method for determining phosphate dihydrate (Na2HPO4.2H2O) were weighed into a 1 L
protein digestibility. The method simulates the digestive process in flask and dissolved with deionized water. Then, the pH of the buffer
the human gastrointestinal tract via pepsin and pancreatic enzymes solution was adjusted to 6.8-6.9 and filtered through a 0.22 µm CA
(Tavano et al., 2016). filter under vacuum. The mobile phase gradient program used in the
separation of amino acids is given in Table 1. The wavelength was
In 1991, the FAO/WHO (Food and Agricultural organization of
254 nm. The separation was achieved with a Gemini-NX 5u C18
the United Nations, 1991) suggested using the protein digestibility-
110 Å, 4.6 x 250 mm column (Phenomenex, CA, USA) with a
corrected amino acid score (PDCAAS) for evaluating food protein
flow rate of 1 mL/min. The column oven temperature was 40 °C.
quality. According to the PDCAAS method, the nutritional quality
of a food protein is determined by the content of the EAAs and
total tract digestibility of crude protein. The amino acid score is Cysteine analysis
calculated by dividing each EAA content by the reference values Cysteine analysis was performed according to the method specified
for 2- to 5-year-old children and adults, respectively. In calculating by ISO 13903 (International Organization for Standardization, 2005).
PDCAAS, the first limiting amino acid score is multiplied by protein First, an oxidation mixture was prepared with 0.5 mL hydrogen
digestibility. PDCAAS score of 1 represents that the protein after peroxide mixed with 4.5 mL performic acid-phenol solution (4.73 g
digestion provides 100 percent of the EAAs required by the organism of phenol in 89% formic acid and 11% water). This solution was
(World Health Organization, 2007). incubated at room temperature for 1 h in order to form performic
Quality protein intake in athletes and physically active individuals acid. Then, the mixture was cooled in an ice-water bath before adding
is important for their performance. There is limited data about the to the sample. Homogenized sample (0.5 g) was weighed into a 10 mL
quality of whey protein supplements available on the market. The glass test tube with 5 mL oxidation mixture and refrigerated at 0 °C
aim of this study was to determine the amino acid content and in for 16 h. After this step, the sample was hydrolyzed, derivatized,
vitro protein digestibility of whey protein supplements and evaluate and analyzed by HPLC (Section 2.2.1 and 2.2.2.).
their protein quality using in vitro PDCAAS methods.
Tryptophan analysis
2 Materials and methods Tryptophan has an indole structure, which is completely
decomposed under acidic conditions; therefore, basic hydrolysis
2.1 Sampling is applied during analysis. The protein hydrolysis and HPLC
In this research, all whey protein supplements (14) were obtained determination method for tryptophan described by Eroğlu et al.
from food supplement market and fitness centers in Istanbul, Turkey. (2016) was used with some modifications. First, 0.5 g sample was
added to a 50 mL schott glass bottles. Next, 20 mL 5 N NaOH
solution was added and hydrolyzed in oven at 120 °C for 12 h. The
2.2 Amino acid analysis samples were cooled to room temperature and filtered through an
ashless filter paper. Next, 1 mL filtered sample was put into a 250 mL
Protein hydrolysis and derivation beaker and adjusted to pH 6.3 using 1 M HCI solution. Then, the
volume was adjusted to 100 mL with deionized water and filtered
Protein hydrolysis and the HPLC determination method for
through a 0.45 µm CA filter before injecting into the HPLC.
amino acids described by Bidlingmeyer et al. (1984) was used with
some modifications. First, 0.5 g sample was added to a 50 mL
schott glass bottle. Then, 20 mL 6 N HCl solution was added and HPLC tryptophan determination
hydrolyzed in an oven at 120 °C for 24 h. The hydrolyzed sample The HPLC system included a Shimadzu Nexera-i device with
was cooled to room temperature and filtered through an ashless a RF-20A fluorescence detector (Shimadzu Corporation, Kyoto,
filter paper. After that, 0.2 mL filtrate was moved to a 10 mL glass Japan). The mobile phase composed of 90% ammonium acetate
test tube and evaporated to dryness at 50 °C under nitrogen. The (0.033 M) and 10% ACN. Then, the pH was adjusted to 6.3 ± 0.1
sample was washed twice with 0.2 mL distilled water and 0.2 with ortho-phosphoric acid and filtered through a 0.22 µm CA
mL acetonitrile (ACN) to remove the acid under nitrogen. The filter under vacuum. The fluorescence detector excitation and
derivatization phase of amino acids utilized 0.5 mL coupling solution emission wavelengths were 280 and 340 nm, respectively. The
(ACN:MeOH:TEA, 100:50:20, v/v) and 0.1 mL Edman’s Reagent separation was achieved with a Gemini-NX 5u C18 110 Å, 4.6
(1.2% phenylisothiocyanate in ACN) in a test tube incubated in an x 250 mm column (Phenomenex, CA, USA) with a flow rate of
oven at 40 °C for 30 min. The derivatized sample was evaporated 1 mL/min. The column oven temperature was 30 °C.
to dryness at 40 °C under nitrogen and then washed two times
with 0.2 mL ACN. After that, 5 mL 0.02 M ammonium acetate
solution was added and filtered with a 0.45 µm cellulose acetate Table 1. HPLC Gradient program for amino acid analysis.
(CA) filter before injecting into the HPLC. Time (min) Mobile Phase A (%) Mobile Phase B (%)
0.01 100 0
HPLC determination of amino acids 13.00 84 16
22.00 64 36
A Shimadzu Nexera-i HPLC with a Shimadzu UV-20A UV- 26.00 60 40
Vis detector (Shimadzu Corporation, Kyoto, Japan) was used for 26.01 40 60
separation of amino acids. The mobile phase consisted of buffer 33.00 100 0
solution (A) and ACN (B). Mobile Phase A: 0.78 g sodium dihydrogen 40.01 100 0

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Pehlivanoğlu; Bardakçı; Yaman

In vitro Protein Digestibility (IVPD) and PDCAAS Table 2. Declared and measured amounts of protein in whey supplements.
The in vitro protein digestibility was performed based on Sample Declared (g/100 g) Measured (g/100 g) % of declared
method described by Pasini et al. (2001) with some modifications. 1 71 64.8 ± 3.6 91.2
Samples containing 250 mg protein were weighed in a 50 mL 2 62.5 58.8 ± 3.2 94.1
plastic falcon tube. First, 20 mL 0.1 N HCI solution was added 3 75.8 68.8 ± 3.8 90.7
to the samples and then pepsin enzyme (2800 units/mg, enzyme/ 4 78.5 70.8 ± 3.9 90.1
protein ratio of 1:30 (w/w)) was added. The digestion was carried 5 82 71.8 ± 4.0 87.5
out at 37 °C for 30 min in a shaking water bath. After that, 5 mL 6 79.1 69.8 ± 3.9 88.2
phosphate buffer solution (1 M) was added and adjusted to pH 7 7 79.1 71.8 ± 4.0 90.7
with 0.05 NaOH solution. Then, pancreatin (4x U.S.P., enzyme/ 8 73 70.8 ± 3.9 96.9
8 62 72.8 ± 4.0 117.3
protein ratio of 1:25, (w/w)) solution was added and incubated at
10 79 73.8 ± 4.1 93.4
37 °C for 2 h in a shaking water bath. After this step, the digestion
11 86 74.8 ± 4.1 86.9
was stopped by adding 1 mL TCA (20%, w/v), the solution was
12 73 54.8 ± 3.0 75.1
centrifuged at 8000 g for 5 min, and the pellet was analyzed for 13 80 70.8 ± 3.9 88.5
nitrogen content using the Kjeldahl method (Association of Official 14 76 66.8 ± 3.7 87.9
Analytical Chemists, 2012). İn vitro protein digestibility-corrected Values are means ± standard deviation, n = 3. The different letters in the same column
amino acid scoring (PDCAAS) calculated as follows: indicate statistical differences between samples (ANOVA p < 0.05, Tukey’s test).

PDCAAS = ((mg of first limiting amino acid in 1 g test protein)/

(mg of the same amino acid in 1 g reference protein))*protein
while the ratio was very low in one sample (6.0%) compared to
digestibility (World Health Organization, 2007).
the reference value (16%). When we compared the proportion of
serine, proline and arginine with the reference values, there were
2.3 Statistical analysis no big differences were found in samples. The ratio of glycine
All analyses were performed in triplicate, and the average ranged between 1.5 and 4.2% in 10 out of 14 samples, whereas
value was used with standard deviation. Significant differences other samples were very high compared to the reference (2%) and
were assessed using ANOVA Tukey’s test (p < 0.05). ranged between 7.4 and 40.3%. As seen from the results, glycine
had the most varied results (Figure 1). As seen from the table, the
proportion of histidine in 13 out of 14 samples was less than the
3 Results and discussion reference value (2%). The proportion of threonine ranged between
1.7 and 9.2% in 13 out of 14 samples, whereas it was very low in
3.1 Total protein content in whey proteins one sample (0.3%) compared to the reference value (4.5%). When
The measured and declared amounts of total protein in whey phenylalanine ratios were compared, no big differences were found
protein supplements are shown in Table 2. The amount of measured between samples. However, the ratio of phenylalanine, except in
total protein ranged from 54.8 ± 3.0 to 74.8 ± 4.1 g/100 g. The one sample (4.2%), was below the reference (3.5%). The ratio of
declared amount of protein on the labels ranged between 62 and methionine ranged between 0.4 and 2.6%, and lysine between
86 g/100 g. The measured total protein amount ranged from 75.1 1.3 and 15.3% in samples. The ratio of methionine was below the
to 117.3% of the amount listed on the whey protein packaging. The reference value (2%) in 11 out of 14 samples and lysine was below
protein content in 6 of 14 samples was less than 10% of the declared the reference value (10.5%) in 10 out of 14 samples.
value, whereas in one sample it was more than 10% of the declared When we evaluated the amino acid composition in whey
amount. Almeida et al. (2015) compared the amount of protein in protein supplements, the most significant difference was found in
whey protein (WP) supplements consumed in the WP-USA and the proportions of glycine compared to other amino acids. The ratio
Brazil. The average amount of protein was 83% in the WP-USA, of glycine in 4 out of 14 samples ranged between 7.4 and 40.3%.
whereas in WP-Brazil it was 63.36%. In the same study, the quality When the amino acid ratios in foods are examined, the largest
of whey protein supplements consumed in the WP-USA was higher ratio is in collagen protein. The main content of collagen consists
than in WP-Brazil. When we evaluated our results, total protein of hydroxyproline, proline, and glycine (Gauza-Włodarczyk et al.,
content in whey protein supplements consumed in Turkey were 2017). The ratio of glycine in 100 g protein is 33% (Paz-Lugo et al.,
higher than 69%. The protein content in whey protein supplements 2018). In another study reported by Van Klinken & Mook (1990),
collected in Turkey was between that in the WP-USA and WP-Brazil. the ratio of glycine, proline, hydroxyproline, and alanine is 31.7,
11.6, 10.1, and 11.3%, respectively, in 100 g protein. In our study, the
3.2 Amino acid composition amount of glycine was very high in four samples. This difference may
be due to the presence of collagen in the whey protein supplements.
The proportions of each amino acid in 100 g protein from
However, further studies are needed to confirm this. In addition,
whey supplements are shown in Table 3. In our study, each amino
the ratios of aspartic acid, alanine, tyrosine, valine, isoleucine,
acid ratio was compared with the results of the study conducted
leucine, tryptophan, and cysteine were below the reference values.
by Pennings et al. (2011). When we compared the proportion of
aspartic acid, alanine, tyrosine, BCAAs (valine, isoleucine and The most other significant change was observed in the ratio
leucine), tryptophan and cysteine with the reference, the ratio of of cysteine in 100 g protein. According to the European legislation
these amino acids in all samples were below the reference values. published in 1990, “Commission Regulation (EEC) No. 2921/90,”
The proportions of glutamic acid ranged between 6.0 and 22.0%. the value of cysteine in whey protein and casein are 3.0% and 0.3%,
The ratio of glutamic acid was above 10.7% in 12 out of 14 samples respectively, and these reference values were used for whey protein

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Table 3. Proportions of each amino acids (in 100 g protein) in whey protein supplements.
Sample Asp Glu Ser Gly His Arg Thr Ala Pro Tyr Val Met Ile Leu Phe Lys Trp Cys
1 10.8 ± 0.7 20.2 ± 1.3 5.6 ± 0.3 2.8 ± 0.2 2.1 ± 0.2 2.87 ± 0.18 8.3 ± 0.8 4.2 ± 0.4 4.0 ± 0.3 3.0 ± 0.3 3.7 ± 0.2 2.1 ± 0.2 3.9 ± 0.4 9.3 ± 0.9 3.3 ± 0.3 11.4 ± 1.1 2.0 ± 0.2 2.7 ± 0.3
2 6.0 ± 0.4 17.8 ± 1.1 4.2 ± 0.3 2.2 ± 0.1 1.8 ± 0.2 2.15 ± 0.13 5.2 ± 0.5 2.7 ± 0.3 4.9 ± 0.3 3.2 ± 0.3 2.8 ± 0.2 1.8 ± 0.2 2.3 ± 0.2 6.6 ± 0.7 2.6 ± 0.3 6.0 ± 0.6 1.7 ± 0.2 2.7 ± 0.3
Original Article

3 7.1 ± 0.4 13.6 ± 0.8 3.6 ± 0.2 2.0 ± 0.1 1.2 ± 0.1 1.87 ± 0.12 5.1 ± 0.5 2.8 ± 0.3 3.4 ± 0.2 2.0 ± 0.2 2.5 ± 0.2 1.4 ± 0.1 2.4 ± 0.2 6.0 ± 0.6 2.2 ± 0.2 7.2 ± 0.7 1.7 ± 0.2 1.5 ± 0.1
4 10.8 ± 0.7 22.0 ± 1.4 5.6 ± 0.3 2.6 ± 0.2 2.0 ± 0.2 2.49 ± 0.15 9.2 ± 0.9 4.3 ± 0.4 4.4 ± 0.3 2.9 ± 0.3 3.7 ± 0.2 2.2 ± 0.2 4.4 ± 0.4 9.2 ± 0.9 3.1 ± 0.3 12.4 ± 1.2 1.8 ± 0.2 2.3 ± 0.2
5 10.1 ± 0.6 18.6 ± 1.1 5.1 ± 0.3 2.4 ± 0.1 1.8 ± 0.1 2.41 ± 0.15 7.0 ± 0.7 3.7 ± 0.4 3.9 ± 0.3 2.4 ± 0.2 3.3 ± 0.2 1.9 ± 0.2 3.6 ± 0.4 8.0 ± 0.8 2.7 ± 0.3 10.8 ± 1.1 1.4 ± 0.1 2.2 ± 0.2
6 7.0 ± 0.4 17.9 ± 1.1 3.9 ± 0.2 7.4 ± 0.5 1.6 ± 0.1 2.07 ± 0.13 4.9 ± 0.5 2.9 ± 0.3 2.9 ± 0.2 2.1 ± 0.2 3.2 ± 0.2 1.5 ± 0.1 3.5 ± 0.3 10.1 ± 1.0 2.2 ± 0.2 15.3 ± 1.5 1.2 ± 0.1 0.1 ± 0.0
7 5.9 ± 0.4 16.6 ± 1.0 3.2 ± 0.2 7.7 ± 0.5 1.1 ± 0.1 1.75 ± 0.11 3.4 ± 0.3 0.7 ± 0.1 3.0 ± 0.2 2.2 ± 0.2 3.0 ± 0.2 1.3 ± 0.1 3.1 ± 0.3 7.6 ± 0.7 2.0 ± 0.2 5.9 ± 0.6 1.3 ± 0.1 0.1 ± 0.0
8 6.3 ± 0.4 13.4 ± 0.8 3.0 ± 0.2 40.3 ± 2.5 1.7 ± 0.1 1.75 ± 0.11 4.3 ± 0.4 2.8 ± 0.3 4.4 ± 0.3 2.7 ± 0.3 2.9 ± 0.2 1.4 ± 0.1 1.9 ± 0.2 5.3 ± 0.5 2.7 ± 0.3 5.5 ± 0.5 1.2 ± 0.1 0.6 ± 0.1
9 2.3 ± 0.1 6.0 ± 0.4 1.1 ± 0.1 22.9 ± 1.4 0.3 ± 0.0 3.86 ± 0.24 0.3 ± 0.0 0.6 ± 0.0 6.5 ± 0.4 0.6 ± 0.1 1.9 ± 0.1 0.4 ± 0.0 0.3 ± 0.0 1.4 ± 0.1 1.5 ± 0.2 1.3 ± 0.1 0.2 ± 0.0 1.0 ± 0.1
10 8.6 ± 0.5 15.4 ± 1.0 3.8 ± 0.2 2.0 ± 0.1 1.4 ± 0.1 1.73 ± 0.11 5.6 ± 0.6 3.1 ± 0.3 3.4 ± 0.2 2.1 ± 0.2 2.8 ± 0.2 1.6 ± 0.2 3.0 ± 0.3 8.2 ± 0.8 2.2 ± 0.2 8.8 ± 0.9 1.6 ± 0.2 1.4 ± 0.1
11 6.9 ± 0.4 10.7 ± 0.7 4.9 ± 0.3 4.2 ± 0.3 1.8 ± 0.1 4.06 ± 0.25 3.2 ± 0.3 3.3 ± 0.3 2.0 ± 0.1 3.0 ± 0.3 3.6 ± 0.2 2.6 ± 0.3 2.3 ± 0.2 4.8 ± 0.5 4.2 ± 0.4 5.5 ± 0.5 1.4 ± 0.1 2.2 ± 0.2
12 5.0 ± 0.3 12.2 ± 0.8 2.8 ± 0.2 1.5 ± 0.1 1.5 ± 0.1 1.83 ± 0.11 2.8 ± 0.3 2.0 ± 0.2 2.6 ± 0.2 2.3 ± 0.2 2.3 ± 0.1 1.2 ± 0.1 2.0 ± 0.2 4.2 ± 0.4 1.9 ± 0.2 4.7 ± 0.5 1.9 ± 0.2 2.9 ± 0.3
13 8.0 ± 0.5 14.0 ± 0.9 3.8 ± 0.2 2.0 ± 0.1 1.4 ± 0.1 1.78 ± 0.11 5.5 ± 0.5 2.8 ± 0.3 3.0 ± 0.2 1.8 ± 0.2 2.5 ± 0.2 1.3 ± 0.1 2.7 ± 0.3 5.8 ± 0.6 2.0 ± 0.2 7.6 ± 0.8 1.7 ± 0.2 0.8 ± 0.1
14 6.8 ± 0.4 19.0 ± 1.2 3.3 ± 0.2 1.8 ± 0.1 1.4 ± 0.1 1.71 ± 0.11 4.7 ± 0.5 2.9 ± 0.3 2.7 ± 0.2 2.0 ± 0.2 2.5 ± 0.2 1.4 ± 0.1 2.7 ± 0.3 5.7 ± 0.6 1.9 ± 0.2 5.7 ± 0.6 1.8 ± 0.2 2.4 ± 0.2
Values are means ± standard deviation, n = 3.
Protein quality of whey protein supplements

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Figure 1. HPLC chromatogram of sample 8 and sample 1.

Table 4. Amount and proportions of essential amino acids (EAA) and branched chain amino acids (BCAA) in whey proteins.
Sample EAA (mg/100 g) BCAA (mg/100 g) EAA (%) % BCAA
1 28447 ± 1572bc 10901 ± 602abc 44.0 ± 4.1a 16.9 ± 1.6ab
2 16972 ± 938e 6882 ± 380d 29.0 ± 2.7cd 11.7 ± 1.1cde
3 19537 ± 1080de 7456 ± 412d 28.5 ± 2.7cd 10.9 ± 1.0de
4 32467 ± 1794a 12225 ± 675a 46.0 ± 4.3a 17.3 ± 1.6a
5 27734 ± 1533bc 10712 ± 591bc 38.8 ± 3.6ab 15.0 ± 1.4abc
6 29189 ± 1613ab 11664 ± 644ab 42.0 ± 3.9ab 16.8 ± 1.6ab
7 19750 ± 1091de 9812 ± 542c 27.6 ± 2.6cd 13.7 ± 1.3bcd
8 17762 ± 982de 7129 ± 394d 25.2 ± 2.3cd 10.1 ± 0.9e
9 5368 ± 297g 2666 ± 147f 7.4 ± 0.7e 3.7 ± 0.3f
10 24889 ± 1376c 10328 ± 571bc 33.9 ± 3.2bc 14.0 ± 1.3abcd
11 20638 ± 1140d 7986 ± 441d 27.7 ± 2.6cd 10.7 ± 1.0de
12 11513 ± 636f 4634 ± 256e 21.1 ± 2.0d 8.5 ± 0.8e
13 20588 ± 1138de 7773 ± 430d 29.2 ± 2.7cd 11.0 ± 1.0de
14 17557 ± 970de 7246 ± 400d 26.4 ± 2.5cd 10.9 ± 1.0de
Values are means ± standard deviation, n = 3. The different letters in the same column indicate statistical differences between samples (ANOVA p < 0.05, Tukey’s test).

quality. The ratio of cysteine in all samples was less than the reference protein ranged from 3.7 to 17.3%. The reference ratio of BCAA is
value. In particular, the ratio of cysteine was very low (0.1-1.0%) reported as 28% (Pennings et al., 2011) and 22.5% (Hulmi et al.,
in the samples with a high ratio of glycine (7.4-40.3%). In a recent 2010). In our results, the ratio of BCAA in all samples was less
study conducted by Gauza-Włodarczyk et al. (2017) cysteine was than the reference values. EAA and leucine (as BCAA) levels are
not found in fish skin (FS), collagen, and bovine Achilles tendon associated with higher muscle protein synthesis (Luiking et al.,
(BAT) collagen and was very low in bone collagen (0.39%). In our 2016). BCAAs are metabolized directly by skeletal muscle. Because
study, we found very high amounts of glycine in 4 out of 14 samples BCAAs are digested faster than other amino acids, they can serve
and a very low amount of cysteine was found compared to reference as a highly effective source of muscle energy during exercise
whey protein. In the literature, collagen contains a high amount of and as a ready source for muscle protein synthesis after exercise
glycine and very low amounts of cysteine. It is thought that these (Nicastro et al., 2012; Shimomura et al., 2004). BCAA and EAA
4 samples may contain some collagen and/or casein. However, values were very low in our study. According to the results, the
further studies are needed to confirm this. use of these products both as a source of fast energy and muscle
protein synthesis may be insufficient.
In Table 4, the amount of EAA ranged from 5368 to 32467 mg/100
g in samples, whereas the proportion ranged from 7.4 to 46.0%.
3.3 Protein Digestibility-Corrected Amino Acid Score
The ratio of EAA is reported as 46.5% (Pennings et al., 2011) and
46.3% (Hulmi et al., 2010). When our results were compared to (PDCAAS)
the reference, the ratio was between 38.8 and 46.0% in 4 out of The protein digestibility results of whey proteins are summarized
14 samples and these results were close to the reference values. in Table 5 and ranged from 50.4 to 79.6%. Almeida et al. (2015)
In the other 10 samples, the ratio was below 33.9% compared to compared the protein digestibility of whey protein supplements
the reference value. The amount of BCAA ranged between 2666 consumed in the WP-USA and WP-Brazil. The in vitro digestibility
and 12225 mg/100 g in samples, while the proportion in 100 g of whey protein supplements was 91.7% in a sample collected

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Table 5. Amino acid score, protein digestibility, and protein digestibility-corrected amino acid score (PDCAAS) of whey protein supplements.
His Ile Leu Lys Met+Cys Phe + Tyr Thr Try Valin Digestibility
Ref * PDCAAS
15.0 30.0 59.0 45.0 22.0 38.0 23.0 6.0 39.0 (%)
bc
Original Article

1 1.37 ± 0.08de 1.30 ± 0.07e 1.57 ± 0.09de 2.53 ± 0.14b 2.18 ± 0.10c 1.65 ± 0.09d 3.61 ± 0.20a 3.37 ± 0.19a 0.94 ± 0.05f 75.4 ± 4.2 0.71 ± 0.04a
2 1.20 ± 0.07d 0.78 ± 0.04e 1.12 ± 0.06d 1.32 ± 0.07cd 2.05 ± 0.11b 1.52 ± 0.08c 2.24 ± 0.12b 2.85 ± 0.16a 0.71 ± 0.04e 50.4 ± 2.8f 0.32 ± 0.02e
3 0.77 ± 0.04g 0.80 ± 0.04fg 1.01 ± 0.06ef 1.59 ± 0.09c 1.34 ± 0.02d 1.09 ± 0.06e 2.23 ± 0.12b 2.85 ± 0.16a 0.64 ± 0.04g 79.6 ± 4.4b 0.50 ± 0.03b
4 1.27 ± 0.07de 1.47 ± 0.08d 1.55 ± 0.09d 2.75 ± 0.15b 2.07 ± 0.11c 1.56 ± 0.09d 4.00 ± 0.22a 2.92 ± 0.16b 0.96 ± 0.05e 67.9 ± 3.8bcde 0.65 ± 0.04a
5 1.15 ± 0.06d 1.21 ± 0.07d 1.36 ± 0.08d 2.39 ± 0.13b 1.87 ± 0.10c 1.33 ± 0.07d 3.03 ± 0.17 a 2.38 ± 0.13b 0.85 ± 0.05e 60.8 ± 3.4ef 0.51 ± 0.03b
6 1.08 ± 0.06de 1.17 ± 0.06d 1.70 ± 0.09c 3.38 ± 0.19a 0.75 ± 0.04f 1.14 ± 0.06d 2.14 ± 0.12b 2.07 ± 0.11b 0.82 ± 0.05ef 64.7 ± 3.6cde 0.48 ± 0.03b
7 0.71 ± 0.04e 1.04 ± 0.06d 1.28 ± 0.07bc 1.32 ± 0.07b 0.63 ± 0.04e 1.10 ± 0.06cd 1.45 ± 0.08b 2.14 ± 0.12a 0.76 ± 0.04e 59.4 ± 3.3ef 0.38 ± 0.02de
8 1.11 ± 0.06c 0.64 ± 0.04e 0.89 ± 0.05d 1.21 ± 0.07c 0.88 ± 0.04d 1.43 ± 0.08b 1.86 ± 0.10a 2.03 ± 0.11a 0.74 ± 0.04de 79.3 ± 4.4ab 0.51 ± 0.03b
9 0.18 ± 0.01f 0.11 ± 0.01g 0.24 ± 0.01e 0.29 ± 0.02de 0.67 ± 0.03a 0.55 ± 0.03b 0.11 ± 0.01g 0.31 ± 0.02d 0.49 ± 0.03c 74.5 ± 4.2bcd 0.08 ± 0.00f
10 0.91 ± 0.05ef 1.01 ± 0.06e 1.38 ± 0.08d 1.94 ± 0.11c 1.34 ± 0.06d 1.14 ± 0.06de 2.43 ± 0.13b 2.74 ± 0.15a 0.73 ± 0.04f 66.7 ± 3.7cde 0.48 ± 0.03b
11 1.15 ± 0.06de 0.75 ± 0.04f 0.81 ± 0.04f 1.22 ± 0.07cd 2.16 ± 0.10a 1.90 ± 0.10b 1.40 ± 0.08c 2.37 ± 0.13a 0.93 ± 0.05ef 61.4 ± 3.4ef 0.46 ± 0.03bc
12 0.95 ± 0.05d 0.67 ± 0.04e 0.71 ± 0.04e 1.05 ± 0.06cd 1.87 ± 0.08b 1.12 ± 0.06cd 1.21 ± 0.07c 3.15 ± 0.17a 0.58 ± 0.03e 64.4 ± 3.6cde 0.37 ± 0.02de
13 0.92 ± 0.05d 0.89 ± 0.05d 0.99 ± 0.05d 1.69 ± 0.09c 0.92 ± 0.05d 1.01 ± 0.06d 2.40 ± 0.13b 2.82 ± 0.16a 0.64 ± 0.04e 63.4 ± 3.5de 0.40 ± 0.02cd
14 0.90 ± 0.05e 0.90 ± 0.05e 0.97 ± 0.05e 1.26 ± 0.07d 1.71 ± 0.08c 1.02 ± 0.06de 2.05 ± 0.11b 2.97 ± 0.16a 0.63 ± 0.03f 59.2 ± 3.3ef 0.37 ± 0.02de
The different letters in the same rows indicate statistical differences between samples (ANOVA p < 0.05, Tukey’s test). *Reference values (mg/kg) for adult according to 2007 WHO/FAO/UNU report (World Health Organization, 2007).
Protein quality of whey protein supplements

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Original Article
Pehlivanoğlu; Bardakçı; Yaman

in the WP-USA and 88.4% in a sample from WP-Brazil. When processing techniques to provide high quality whey protein to
we evaluated our results, in vitro protein digestibility in whey consumers.
protein supplements from Turkey were lower than in the USA
and Brazil. The low protein digestibility may be attributed to the References
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