Amino acid composition and nutritional value evaluation of Chinese chestnut ( Castanea mollissima Blume) and its protein subunit

Abstract

The amino acid composition, nutritional value and proteins subunit of chestnuts (Castanea mollissima Blume) from three regions of China (Henan, Hunan, and Guangdong) were investigated. Experimental results showed that the albumin fraction dominated the chestnut protein composition, but globulin and prolamin were nondetectable. All the Chinese chestnut proteins had a nutritionally balanced amino acid composition. Their amino acid score (AAS) could reach the FAO/WHO (2013) requirement for essential amino acids for older children, adolescents and adults. Leucine was the first limiting amino acid for all the Chinese chestnut protein isolates and digestible indispensable amino acid scores (DIAAS) were more than 90 (Leu). The sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) spectra showed that different Chinese chestnut protein isolates had almost the same band components, which were mainly comprised of seven polypeptide segments with the molecular weights of (91–93) kDa, (70–72) kDa, (53–55) kDa, 37 kDa, (27–33) kDa, 20 kDa and (5–15) kDa. The amino acid compositions and the abundance of low molecular weight proteins indicated that Chinese chestnut could be utilized as a good source of plant protein for human nutrition.

Full text

Amino acid composition and nutritional value
evaluation of Chinese chestnut (Castanea
mollissima Blume) and its protein subunit
Fang Yang,†
a
Xingjian Huang,†
b
Conglan Zhang,
c
Mei Zhang,
a
Chao Huang
a
and Hao Yang *
a
The amino acid composition, nutritional value and proteins subunit of chestnuts (Castanea mollissima
Blume) from three regions of China (Henan, Hunan, and Guangdong) were investigated. Experimental
results showed that the albumin fraction dominated the chestnut protein composition, but globulin and
prolamin were nondetectable. All the Chinese chestnut proteins had a nutritionally balanced amino acid
composition. Their amino acid score (AAS) could reach the FAO/WHO (2013) requirement for essential
amino acids for older children, adolescents and adults. Leucine was the first limiting amino acid for all
the Chinese chestnut protein isolates and digestible indispensable amino acid scores (DIAAS) were more
than 90 (Leu). The sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) spectra
showed that different Chinese chestnut protein isolates had almost the same band components, which
were mainly comprised of seven polypeptide segments with the molecular weights of (91–93) kDa, (70–
72) kDa, (53–55) kDa, 37 kDa, (27–33) kDa, 20 kDa and (5–15) kDa. The amino acid compositions and
the abundance of low molecular weight proteins indicated that Chinese chestnut could be utilized as
a good source of plant protein for human nutrition.
Introduction
Chestnut is an important edible fruit, which has a long growing
history over 2000 years in China. It contains considerable
essential fatty acids, bers, saccharides, vitamins, and
minerals.
1,2
In addition, chestnut is also an excellent source of
plant protein. The average crude protein of 100 g fresh chestnut
fruit was estimated to be 3.5 g, which represented approxi-
mately 9.2% of the recommended daily intake (RDI) for females
and 7.6% of the RDI for males.
3
It is well known that plant
protein is an alternative to animal protein for human nutrition,
which contributes about 65% of the per capita supply of protein
within the scope of the global. Furthermore, plant protein can
be used as a complete and well-balanced source of amino acids
to meet the physiological needs of the human body.
4,5
A recent
study showed that the predominant free amino acids in
chestnut were aspartic acid, asparagine and glutamic acid.
6,7
Therefore, chestnut has a great potential to be used as a nutri-
tional food.
In addition to amino acids, protein subunits have caused
extensive attention because amino acids from single peptides or
peptide containing protein hydrolysates are easier to be absor-
bed than those from equivalent mixtures of free amino acids.
8,9
Besides, protein subunits have relationship to their specic
bioactivity.
10–12
For example, Wang and Ng isolated a novel
antifungal protein from Chinese chestnut seeds and the active
protein exhibited a molecular mass of 37 kDa.
13
Collada et al.
puried a 20 kDa protein from cotyledons of recalcitrant
chestnut seeds. The isolated protein could increase the rena-
turation yields of chemically denatured citrate synthase and
prevent the irreversible thermal inactivation.
14
Chestnut is geographically distributed in three major areas:
Europe with Castanea sativa Miller, Asia with Castanea crenata
Sieb. et Zucc. (Japan) and Castanea mollissima Blume (China
and Korea), and North America with Castanea dentata (Marsh.)
Borkh. As far as we know, the nutritional value evaluation and
the protein subunit of fresh Chinese chestnut (Castanea mol-
lissima Blume) has not been reported yet. In this study, the
nutritional properties of chestnut (Castanea mollissima Blume)
from three different regions of China were evaluated, which
included amino acid composition, amino acid score (AAS), the
predicted protein efficiency ratio (PER), essential amino acid
index (EAAI), and digestible indispensable amino acid score
(DIAAS).
15
Their difference in subunit components was also
a
Key Laboratory for Green Chemical Process of Ministry of Education, Wuhan Institute
of Technology, Wuhan, 430205, China. E-mail: hyang@wit.edu.cn; Tel:
+86-13377876683
b
Key Laboratory of Environment Correlative Dietology, Ministry of Educatio n,
Huazhong Agricultural University, Wuhan, 430070, China
c
Department of Biological Engineering, Hubei University Zhixing College, Wuhan,
430011, China
†Co-rst author contributed equally to this work.
Cite this: RSC Adv.,2018,8,2653
Received 3rd December 2017
Accepted 5th January 2018
DOI: 10.1039/c7ra13007d
rsc.li/rsc-advances
This journal is © The Royal Society of Chemistry 2018 RSC Adv.,2018,8,2653–2659 | 2653
RSC Advances
PAPER

investigated by sodium dodecyl sulphate polyacrylamide gel
electrophoresis (SDS-PAGE). The purpose of this study is to
provide a fundamental information of amino acid composition
and nutritional value of Chinese chestnut. In addition, it also
provides a reference for future study on the relationship
between protein subunit structure and functionality of chestnut
protein.
Materials and methods
Materials
Chestnuts (Castanea mollissima Blume) from three different
regions of China from North to South (Henan, Hubei, and
Guangdong) with a 23to 34latitude span, harvested in the fall
of 2014, were used in this study. They were purchased fresh
either from chestnut processors or from the distributors, and
they were kept in a cooler for no more than 2 weeks if long
distance shipment was required. The acquisition was done
through local contacts (collaborators or colleagues) to ensure
that only regionally grown chestnuts were collected. Hence,
chestnut samples were named as their geographic growing
locations from north to south in China: HEN (Henan province),
HUN (Hunan province), and GD (Guangdong province). The
three different regions (Henan, Hunan and Guangdong) were at
the same longitude (113 degree), and the average temperatures
in the fall season (August–October) were 20.6 C, 23.8 C, and
26.2 C, respectively. Their average precipitations were 641 mm,
1400 mm, and 1777 mm, respectively.
Fresh chestnuts (1000 g) per region were previously cross-cut
on the top according to a previous work,
16
then they were peeled
with a knife and the nuts were rapidly cut into small. Three
independent tests (replications) were performed. In each
replication, 20–30 shelled nuts were diced into small pieces,
and approximately 200 g of the mixture was milled in a Model
FW80 high-speed grinder (Teste Instrument Co., Ltd., Tianjin,
China) to obtain ne particles. They were placed in sealed
plastic bags, kept at 2 C and analyzed within seven days.
Chemicals and reagents were of analytical grade and ob-
tained from Sinopharm (Shanghai, China) and Sigma Chemical
(St. Louis, MO, USA). Water used in the experiments was
ultrapure deionized water.
Proximate chemical analysis
The moisture content of chestnuts was analyzed by gravimetric
method using a drying oven (DHG-9075A, Yiheng, Shanghai,
China) at 101 2C until a constant weight was obtained. The
ash content and total protein nitrogen were analyzed using
AOAC methods (AOAC, 2000). Sample digestion was carried out
in a digestion system sealed with a cork, and copper was used as
a catalyst. The percentage of nitrogen was converted to crude
protein by multiplying with 5.30, which was specic for
chestnut fruit.
17
Protein fractionation
Different chestnut protein fractions were isolated by sequentially
extracting defatted chestnut powder with different solvents
according to the method described by Chavan et al.
18
Briey,
defatted and dried chestnut powder was fully dissolved in
distilled water and extracted over 45 min at room temperature,
then the suspension was centrifuged at 4200 rpm for 20 min. The
residues were re-extracted twice with the same solvent. The
recovered ltrates were combined and designated the “water-
soluble fraction (albumin)”. The residue from aqueous solu-
tion extraction was further fully dissolved with 0.5 M NaCl
solution (pH 7.0) in order to obtain globulin. The residue from
above salt solution extraction was further fully dissolved with
70% (v/v) ethanol solution at 65 C in a shaking water bath in
order to obtain prolamin. The residue from above ethanol
solution extraction was further fully dissolved with 0.1 M sodium
hydroxide in order to obtain glutelin. Filtrates containing the
desired protein fractions were dialyzed against distilled water for
48 h at 4 C and separately lyophilized. Protein content in each
fraction was determined according to AOAC 2000. All lyophilized
chestnut protein fractions were then stored in the refrigerator (4
C) in airtight plastic bottles until further use.
Amino acid composition and evaluation of nutritional
parameters
Two milligrams of defatted chestnut power samples were
hydrolyzed with 0.5 mL 6 M HCl in a sealed ampoule containing
8 mL phenol (for protection of tyrosine) and 0.25 mLnorleucine
(catalogue no. N8513, Sigma, as an internal standard) for 24 h at
110 C under vacuum. The acid hydrolysate was dried completely
using a Speedvac concentrator (Savant Instrument, Farmingdale,
NY) and the dry residue was re-dissolved in 0.5 mL of citrate
buffer (Beckman A303084, CA). The sample was ltered through
a0.45mm nylon lter before being analyzed with an automated
Amino Acid Analyzer (Hitachi 835-50, Japan). Sulphur-
containing amino acids, cystine and methionine were deter-
mined aer a pre-hydrolysis oxidation with performic acids.
19
Cystine was analyzed according to Okuno's report.
20
The
contents of different amino acids recovered were presented as g/
100 g protein and were compared with the FAO/WHO (2013)
reference pattern. The ratio of essential to total amino acids was
reported as E/T(%). The essential amino acid (EAA) score was
calculated by the method of FAO/WHO as shown below:
21
EAA ¼mg of EAA in 1 g of test protein
mg of EAA in 1 g of egg protein 100
Essential amino acid index (EAAI) was calculated by the
method of FAO/WHO as shown below:
EAAI ¼ffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffiffi
Yn
i¼1EAAi
n
q
The predicted protein efficiency ratio (PER) values were
calculated from their amino acid composition based on three
equations developed by Chavan et al.,
22
as given below:
PER-1 ¼0.684 + 0.456(Leu) 0.047(Pro)
PER-2 ¼0.468 + 0.454(Leu) 0.105(Tyr)
2654 |RSC Adv.,2018,8,2653–2659 This journal is © The Royal Society of Chemistry 2018
RSC Advances Paper

PER-3 ¼1.816 + 0.435(Met) + 0.780(Leu)
+ 0.211(His) 0.944(Tyr)
The amino acid score (AAS) was calculated by the method of
FAO/WHO as shown below:
AAS ¼
mg of AA in 1 g of test protein
mg of AA in 1 g of the FAO=WHO reference pattern
100
The digestible indispensable amino acid (IAA) reference
ratio was calculated to determine DIAAS of the samples, and
each IAA reference ratio was calculated according to the
following equation:
Digestible indispensable amino acid score (DIAAS) was
calculated as shown below:
23
DIAAS (%) ¼100 lowest value of digestible IAA reference ratio
Electrophoresis
Total protein isolate and protein fractions of three different
Chinesechestnuts(HEN,HUN,andGD)wereallusedfor
sodium dodecyl sulphate polyacrylamide gel electrophoresis
(SDS-PAGE) studies. SDS-PAGE was performed with 12.5% (v/v)
bisacrylamide gel and 4% (v/v) stacking gel in a DYCZ-24DN
mini-Protein electrophoresis (Sixty-one Instrument, Beijing,
China). The electrophoresis buffer consisted of 125 mM Tris,
5 M urea at pH 6.8, 0.2% (v/v) SDS, 20% (v/v) glycerol, and
0.01% (w/v) bromophenol blue. Defatted chestnut sample
(6 mg) was mixed with 420 mLextractionbuffer and incubated
for 1 h. The extraction buffer was composed of 50 mM Tris, 5 M
urea at pH 8.0, 0.2% (v/v) SDS, 2% (w/v) reducing agent (2-
mercaptoethanol). The samples were then mixed with 420 mL
electrophoresis buffer. This solution was heated at 95 C
for 5 min with constant shaking. Aliquots of 6 mLofprepared
sample were loaded into each well. The electrophoretic sepa-
ration was conducted at 200 V for 50 min. The gels were
stained with Coomassie blue solution for 30 min with
constant shaking, and then destained twice with 45% (v/v)
ultrapure water, 45% (v/v) methanol and 10% (v/v) acetic
acid solution respectively for 1 h, and once with 5% (v/v) acetic
acid and 22.5% (v/v) methanol solution for 12 h. The gels were
scanned by a Sharp JX-330 scanner (Amersham Biosciences,
Quebec), and the integrated intensities of electrophoresis
protein bands were analyzed with Image Master ID Elite so-
ware (Version 2.0, Amersham Biosciences). The protein
composition was calculated as the total areas under all the
peaks.
24
Statistical analysis
All experiments were replicated three times (n¼3) and each
employed a new batch of ground chestnuts as an independent
test. Analyses of data from all replications were performed by
a one-way ANOVA test, followed by Duncan's new multiple
range test with a 0.05 signicance level using Statistical
program 10.0 (Stat SoInc., Tulsa, OK, USA).
Results and discussion
Proximate chemical analysis
The proximate chemical compositions of the chestnut fruits
from three different regions of China (HEN, HUN, and GD) were
shown in Table 1. Chestnut from Hunan province had signi-
cantly higher moisture content than the samples from Henan
and Guangdong province (P< 0.05). All chestnut fruits had
a moisture content greater than 46%, while the mean moisture
content was 47.58%, which was lower than that of Spanish
chestnuts (54.00%).
25
The American chestnut variety had
a similar moisture content.
26
The average ash contents of these
Chinese chestnut fruits (1.59%) were similar to or less than the
Turkish chestnuts (1.00–3.20%).
27
The crude protein contents were 8.12% (HEN), 9.74% (HUN)
and 7.54% (GD) respectively, which were similar with that re-
ported by Pereira-Lorenzo (4.50–9.60%).
25
Soluble protein
content calculated by the protein fractions ranged from 4.86%
(GD) to 6.68% (HUN), which were also consistent with the report
of Pereira-Lorenzo.
25
Soluble protein content was lower than the
total protein contents in Table 1 because of the presence of
insoluble protein. The average soluble protein content of
Chinese chestnut fruits was 5.70% and there was not a deni-
tive geographic pattern. That is to say, the regional variation was
found to be relatively small.
Protein fractionation
Osborne solubility-based protein fractionation data (Table 2)
indicated that albumin (71.62% of the total soluble protein)
Table 1 Proximate chemical analysis (g/100 g)
b
of chestnut fruits from
three different regions of China (HEN, HUN, and GD)
Region Moisture Ash Crude protein Soluble protein
HEN 46.43 0.31
a
1.81 0.09
a
8.12 0.13
a
5.57 1.83
a
HUN 49.75 0.26
a
1.44 0.08
a
9.74 0.24
a
6.68 0.04
a
GD 46.55 1.01
a
1.51 0.04
a
7.54 0.22
a
4.86 0.05
a
Ave 47.58 1.88 1.59 0.20 8.47 1.14 5.70 0.92
a
Means (n¼3) standard deviations within a column with different
superscripts differ signicantly (P< 0.05).
b
For moisture, the unit is
on a wet weight basis; for all other components, the unit is on a dry
weight basis.
Digestible IAA reference ratio ¼mg of digestible IAA in 1 g pretein of food
mg of the same dietary IAA in 1 g of the reference protein
This journal is © The Royal Society of Chemistry 2018 RSC Adv.,2018,8,2653–2659 | 2655
Paper RSC Advances

were the most dominant, followed by glutens (28.38%). The
globulin and prolamin were nondetectable in all the Chinese
chestnut fruits. The albumin content of HUN was highest
(5.24%), followed by HEN (3.76%) and GD (3.35%). The ratios of
albumin to gluten of the three chestnut proteins (HEN, HUN,
and GD) were 2.1, 3.7, and 2.3, respectively. This obvious
difference makes it possible to distinguish different sources of
chestnut proteins. The absence of globulins in the Chinese
chestnut protein is quite different from those from beans
(63.93%)
27
and cowpea seeds (66.6%),
28
and it maybe the
distinctive features of Chinese chestnut protein.
Amino acid composition and nutritional value evaluation
The amino acid composition of chestnut fruits from three
different regions of China (HEN, HUN, and GD) were shown in
Table 3, while their AAS and DIAAS were shown in Table 4. All
the Chinese chestnut fruit proteins were found to be rich in
aspartic acid (16.6–20.1 g/100 g protein) and glutamic acid
(15.8–17.0 g/100 g protein), which agreed well with previous
reports.
29
According to Phat et al.,
30
the high levels of aspartic
and glutamic acids were responsible for the special favour and
taste. The sulphur-containing amino acid content (methionine
and cystine) was 3.9, 3.8 and 3.8 g/100 g protein for HEN, HUN
and GD, respectively, which indicated that the Chinese chestnut
fruits had higher content of the sulphur-containing amino acids
than that of FAO/WHO (2013) standard mode for older children,
Table 2 Protein fractionation yields
b
of chestnut fruits from three
different regions of China (HEN, HUN, and GD)
Osborne protein
fraction Albumin Globulin Prolamin Gluten
HEN 67.50 0.02
a
0 0 32.50 0.02
a
HUN 78.44 0.02
a
0 0 21.56 0.01
a
GD 68.93 0.02
a
0 0 31.07 0.01
a
Ave 71.62 5.95 0 0 28.38 5.95
a
Means (n¼3) standard deviations within a column with different
superscripts differ signicantly (P< 0.05).
b
For protein fractionation
yields, the data are percentage of total soluble protein.
Table 3 Amino acid composition, E/T(100%), EAAI, and predicted
protein efficiency ratio (PER) of chestnut fruits from three different
regions of China (HEN, HUN, and GD) (g/100 g of protein isolate)
Amino acid HEN HUN GD
FAO/WHO (2013) standard
mode for older children,
adolescents and adults
Ile 3.9 3.8 4.2 3.0
Leu 6.8 6.7 7.4 6.1
Lys 6.3 6.3 6.4 4.8
Met 1.8 1.6 1.9
Cys 2.1 2.2 1.9
Total sulphur
amino acids
3.9 3.8 3.8 2.3
Tyr 2.6 2.8 3.4
Phe 3.9 4.0 5.0
Total aromatic
amino acids
6.5 6.8 8.4 4.1
Thr 4.1 4.0 4.4 2.5
Val 5.8 4.7 4.9 4.0
His 2.9 2.8 2.7 1.6
Trp 1.2 1.3 1.0 0.66
Total essential
amino acids
38.5 37.4 40.5 27.5
Asp 18.3 19.1 16.6
Glu 16.0 15.8 16.5
Ser 4.3 4.1 4.6
Gly 4.7 4.6 4.9
Arg 7.5 9.3 6.8
Ala 5.6 6.0 6.1
Pro 5.2 3.7 3.9
Total non-essential
amino acids
62.3 62.6 59.4
E/T(%) 38.5 37.4 40.5
EAAI 77.7 76.0 79.0
PER-1 2.2 2.2 2.5
PER-2 2.4 2.3 2.5
PER-3 2.7 2.3 2.2
Table 4 Score of chestnut fruits from three different regions of China
(HEN, HUN, and GD). (a) AAS of chestnut fruits from three different
regions of China (HEN, HUN, and GD). (b) DIAAS of chestnut fruits from
three different regions of China (HEN, HUN, and GD)
c
(a)
Amino acids score HEN HUN GD
Ile 130.00
b
126.67 140.00
Leu 111.48
a
109.84
a
121.31
a
Lys 131.25 131.25 133.33
Met + Cys 169.57 165.22 165.22
Phe + Tyr 158.54 165.85 204.88
Thr 164.00 160.00 176.00
Val 145.00 117.50
b
122.50
b
Trp 181.82 196.97 151.52
His 181.25 175.00 168.75
Total essential amino acids 140.00 136.00 147.27
(b)
Digestible IAA reference
ratio
d
HEN HUN GD
Ile 1.09 1.06 1.17
Leu 0.93 0.91 1.01
Lys 1.05 1.05 1.07
Met + Cys 1.45 1.41 1.71
Phe + Tyr 1.32 1.38 1.71
Thr 1.26 1.23 1.35
Val 1.16 0.94 0.98
Trp 1.64 1.77 1.36
His 1.27 1.23 1.18
DIAAS (%)
e
93 (Leu) 91 (Leu) 98 (Val)
a
Means the rst limited amino acid.
b
Means the second limited amino
acid.
c
Indispensable amino acid (IAA) digestibility coefficients are
based on predicted human values obtained from pig data.
32
d
Digestible IAA reference ratio (digestible IAA in 1 g protein of
chestnut fruit/mg of the same dietary indispensable amino acid in 1 g
of the reference protein).
e
DIAAS for chestnut fruit (lowest value of
the ‘digestible IAA reference ratio’expressed as % for each reference
pattern).
2656 |RSC Adv.,2018,8,2653–2659 This journal is © The Royal Society of Chemistry 2018
RSC Advances Paper

adolescents and adults (2.3 g/100 g). Total aromatic amino acid
content of the three chestnut proteins was higher than the
requirement of FAO/WHO (2013) for older children, adolescents
and adults, while total essential amino acid content was higher
than the requirement of FAO/WHO (2013) for older children,
adolescents and adults and all E/Tvalues were about 37%,
which was considered adequate for an ideal protein (FAO/WHO,
2013). The greater the EAAI, the more balanced amino acid
composition and the higher quality and efficiency of the
protein.
31
The EAAI of the three chestnut proteins were all
higher than 100, which suggested that the amino acid compo-
sition of Chinese chestnut is superior to FAO/WHO (2013)
standard. The predicted PER-1 and PER-2 values (Table 3) of GD
were higher than those of HEN and HUN, suggesting that GD
might have higher digestibility.
Based on the analyse above, leucine was the rst limiting
amino acid for all the Chinese chestnut proteins (Table 4). The
second limiting amino acid was isoleucine for HEN, whereas for
HUN and GD, the second limiting amino acid was valine,
respectively. However, total essential amino acid scores for the
three Chinese chestnut proteins could reach the FAO/WHO
requirement (2013) for older children, adolescents and adults.
As a result, the essential amino acid composition of Chinese
chestnut protein was reasonable, which was similar with that of
Chinese kabuli and desi chickpea (Cicer arietinum L.) cultivars.
9
DIAAS was calculated to evaluate the protein quality of
Chinese chestnut fruits. It is a new and advanced recommended
method of measuring protein quality (FAO, 2013) and is on the
basis of the lowest amount of the digestible dietary indispens-
able amino acid per unit of the dietary protein. Table 4(b)
Fig. 1 (a) SDS-PAGE spectra of the chestnut fruits protein from three different regions of China*, and (b) SDS-PAGE spectra of chestnut fruits
albumin and glutelin fractions**.*(1) HUN; (2) GD; (3) HEN. **(1) albumin of HUN; (2) albumin of HEN; (3) albumin of GD; (4) glutelin of HUN; (5)
glutelin of HEN; (6) glutelin of GD.
This journal is © The Royal Society of Chemistry 2018 RSC Adv.,2018,8,2653–2659 | 2657
Paper RSC Advances

presents the DIAAS values for the proteins in the analyzed three
chestnut fruits. True ileal digestibility coefficients for indis-
pensable amino acids in these proteins were obtained from
growing pigs,
32,33
which have been recommended by the FAO if
true ileal amino acid digestibility data derived from humans
were absent.
15
It is considered as ‘Excellent’, if DIAAS $100%;
‘Good’, if DIAAS between 75% and 99%; and ‘Low’if DIAAS <
75%. When the FAO (2013) reference pattern for older children,
adolescents and adults was employed, the scores ranged from
91% (Leu) in HUN to 98% (Val) in HUN. DIAAS values for the
three chestnut fruits were close and were all higher than 90%,
which could be identied as ‘good’quality. It should be noticed
that most of the plant proteins are of ‘low’quality (DIAAS <
75%).
34,35
The DIAAS value of the Chinese chestnut fruits is
higher than many other plant proteins, for example, rice (DIAAS
¼73%), wheat our (DIAAS ¼51%), and lentils (DIAAS ¼54%).
This indicates that the protein of chestnut fruit is a good kind of
plant protein and it contains a surplus amount of indispensable
amino acids (total essential amino acids >37%), which can
greatly improve the developing countries' typical low-quality
diets.
Electrophoresis (SDS-PAGE)
SDS-PAGE spectra of Chinese chestnut proteins were shown in
Fig. 1, which indicated that the molecular weight distributions
of all the Chinese chestnut proteins were similar. Fig. 1(a)
showed that the three Chinese chestnut proteins were mainly
comprised of seven polypeptide segments having the molecular
weights of (91–93) kDa, (70–72) kDa, (53–55) kDa, 37 kDa, (27–
33) kDa, 20 kDa, and (5–15) kDa. Fig. 1(b) showed that the
albumins from all the Chinese chestnut proteins were mainly
comprised of four polypeptide segments with the molecular
weights of Alb-1 (32 kDa), Alb-2 (27 kDa), Alb-3 (20 kDa) and Alb-
4(5–15 kDa). The glutelin from the three chestnut proteins were
mainly comprised of six polypeptide segments, which had the
molecular weights of Glu-1 (92 kDa), Glu-2 (71 kDa), Glu-3 (54
kDa), Glu-4 (37 kDa), Glu-5 (33 kDa) and Glu-6 (30 kDa).
The relative protein quantity in Table 5 showed that the
major proteins present in the Chinese chestnut were in the
range of 5–20 kDa (the bands 6 and 7) and 27–53 kDa (the bands
3, 4, and 5). The result was similar as that of S´
anchez-Vioque
et al.,
36
who reported that the major components of chickpea
protein were in the range of 40–47 kDa and 24–25 kDa. Total
relative protein quantity of the bands 5, 6, and 7, which con-
tained the albumins fraction of Chinese chestnut proteins was
78.3% for HEN, 72.2% for HUN and 74.7% for GD, whereas total
relative protein quantity of the bands 1, 2, 3 and 4 was 21.7% for
HEN, 27.8% for HUN and 25.3% for GD. Interestingly, the
albumins of the three Chinese chestnut proteins were very high
(more than 70%) and they were made up of subunits with low
molecular weight (5–32 kDa). It was thought that the lower
molecular weight may be helpful to improve water solubility
and digestibility of the chestnut protein.
Conclusions
To sum up, amino acid components of chestnut fruits from
three different regions of China (HEN, HUN and GD) were
analyzed in this paper to evaluate the nutritional value of
Chinese chestnuts. Albumin and gluten were the two major
protein components of Chinese chestnut, while globulin and
prolamin were absent. Leucine was found to be the limiting
amino acids for all the Chinese chestnut proteins, so chestnut
should be combined with leucine-rich food in future food
research and development to get better digestibility. According
to DIAAS based on the FAO/WHO (2013) requirement of the
essential amino acids for older children, adolescents and
adults, Chinese chestnut (DIAAS > 90%) was a good source of
plant protein with a better digestibility than other plant
proteins. SDS-PAGE indicated that the three Chinese chestnut
proteins were rich in albumins with low molecular weight
subunits (5–32 kDa) and they showed almost the same band
components. Therefore, Chinese chestnut protein can be
utilized as a good source of plant protein for human nutrition.
Conflicts of interest
There are no conicts to declare.
Acknowledgements
This study was supported by the National Natural Science
Foundation of China (Grant No. 31701604) and Scientic
Research Foundation of Wuhan Institute of Technology (Grant
No. k201633). Fang Yang expresses her gratitude for the post-
doctoral opportunity at the University of Kentucky where
a portion of the study was conducted.
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