Journal Pre-proof Tenderness and histochemistry of muscle tissues from Eriocheir sinensis Long Zhang, Wenli Wang, Fen Zhou, Yao Zheng, Xichang Wang PII:
To appear in:
Received Date: 6 December 2018 Revised Date:
24 October 2019
Accepted Date: 26 October 2019
Please cite this article as: Zhang L., Wang W., Zhou F., Zheng Y. & Wang X., Tenderness and histochemistry of muscle tissues from Eriocheir sinensis, Food Bioscience (2019), doi: https:// doi.org/10.1016/j.fbio.2019.100479. This is a PDF file of an article that has undergone enhancements after acceptance, such as the addition of a cover page and metadata, and formatting for readability, but it is not yet the definitive version of record. This version will undergo additional copyediting, typesetting and review before it is published in its final form, but we are providing this version to give early visibility of the article. Please note that, during the production process, errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. © 2019 Published by Elsevier Ltd.
Tenderness and Histochemistry of Muscle Tissues from
Long Zhang#, Wenli Wang#, Fen Zhou, Yao Zheng, Xichang Wang*
College of Food Science & Technology, Shanghai Ocean University, Shanghai 201306, PR
Running Title: Tenderness of Eriocheir sinensis
# These authors contributed equally to this study.
*Corresponding author: Xichang Wang
Address: No.999, Huchenghuan Rd, Nanhui New City, Pudong New District, Shanghai, P.R.
Tel: +86 21 61900051; Fax: +86 21 61900054;
E-mail address: [email protected]
Texture is the most important factor influencing consumer satisfaction with crab
meat palatability. A valuable micro method using particle size analysis was developed
to qualitatively determine cooked meat tenderness of different muscle tissues. The test
was successfully used with Chinese mitten crab (Eriocheir sinensis), horse crab
(Portunus trituberculatus), manila clam (Ruditapes philippinarum), red swamp
crawfish (Procambarus clarkii), and crucian carp (Carassius auratus). It was
observed that the method produces tenderness values that correlate (R2 = 0.910) well
with the sensory evaluation (p < 0.01). These results suggested that the breaking rate
of muscle fiber using the entropy weighted coefficient method can be quantified to
reflect meat tenderness, which is very suitable for cooked meat with low-fat, low
connective tissue, and meat which is hard to cut into specific shapes. The
histochemical and textural properties were different for the different edible muscle
tissues. Tenderness had a high correlation with moisture, myofibrillar proteins and the
water holding capacity. Furthermore, the myofibrillar protein (about 70 g/100 g crude
protein) was the main determinant of tenderness. This study suggested that it might be
possible to develop a more widely applicable method for evaluating the cooked
texture of crustaceans.
Keywords: Chinese mitten crab, Eriocheir sinensis, horse crab, Portunus
trituberculatus, manila clam, Ruditapes philippinarum, red swamp crawfish,
Procambarus clarkii, crucian carp, Carassius auratus, tenderness
Texture is the most important factor influencing consumer satisfaction for crab
meat palatability. The texture of crab meat is also an important part of the
comprehensive evaluation of crab quality. The outstanding texture characteristic of
aquatic products is their tenderness, but the water environment, circulation, and other
aspects will significantly affect the tenderness of crab meat, and there are also
differences in different muscle tissues. Tenderness is an important quality attribute
determined by the ease of chewing (Boleman et al., 1997). From the perspective of
consumer perception, it has been widely used in the evaluation of meat quality. The
tenderness of meat can be expressed as resistance to tooth pressure or how much force
it takes to bite through a piece of meat with molars. It can also be described by a large
number of descriptors, including mushy, crumbly, mealy, or elastic (KaiĆ and ŽGur,
2017; Marino et al., 2011).
Meat tenderness can be measured using instruments or humans (as sensory or
consumer groups). The instrumental methods for measuring shear force (SF),
including slice shear force (SSF) and Warner-Bratzler shear force (WBSF), have been
widely used in meat tenderness evaluation. WBSF is used to measure the strength
needed for whole muscle fiber shearing (AMSA, 2015). This method requires that the
sample be cut to a specified size so that the core perpendicular to the muscle fiber can
be cut completely. However, crab meat is hard to cut into specific shapes so it cannot
meet the sample requirements for instrumental measurements. Moreover, it has been
shown that shear force (both WBSF and SSF) does not properly reflect tenderness 3
differences among muscles (King et al., 2009; Rhee et al., 2004). Therefore, it is
inappropriate to use SF to compare tenderness differences among muscles (AMSA,
2015). The measurement of myofibril fragment lengths (MFL) after homogenization
is a method to microscopically predict meat tenderness. The myofibril sediment is
measured using phase contrast microscopy by counting the number of sarcomere
segments of the average length of each fibril using 100 fibrils (Moholisa et al., 2017;
MØller et al., 1973; Pool and Klose, 1969). MØller et al. (1973) observed that
mechanical measurements of fragments reflected the physical properties of the muscle
tissue and had a significant correlation with meat tenderness. Based on the micro
method, it might be possible to reflect tenderness or fragility differences among
muscles using the rate of change of particle size over multiple time gradients using a
Malvern Mastersizer. Since the method is not a one-time break to detect the breaking
force, the breaking rate is monitored using different breaking times with the same
breaking force, which is more consistent with multiple breaks during chewing, and
might more objectively reflect the degree of fragility of different muscle fibers.
For marine crabs species, snow crab (Chionoecetes opilio) (Hayashi et al., 1981),
blue crab (Callinectes sapidus) (Çelik et al., 2004; Gökoðlu and Yerlikaya, 2003;
Hernández-Robledo et al., 2016; Martínez et al., 2014), green crab (Carcinus maenus
(Skonberg and Perkins, 2002), Carcinus mediterraneus (Cherif et al., 2008)), brown
crab (Cancer pagurus) (Barrento et al., 2010), warty crab (Eriphia verrucosa) (Kaya
et al., 2009), mangrove crab (Scylla serrata) (Castilho et al., 2007), Atlantic spider
crab (Maja brachydactyla) (Marques et al., 2010), swimming crab (Ovalipes
trimaculatus) (Dima, et al., 2012), stone crab (Platyxanthus patagonicus) (Dima et al.,
2012), and mud crab (Scylla serrata) (Benjakul and Sutthipan, 2009), other than
nutrition and flavor characteristics, previous studies have mainly focused on the
muscle histomorphology, as well as changes in the physicochemical and sensory
properties during thermal processing and frozen storage. However, few studies have
focused on the texture properties of cooked crab meat, which could generally be
measured using the water holding capacity and sensory evaluation as the standard.
There is currently no quantitative method for determining tenderness of crab meat.
This study used E. sinensis, which is quite popular among Chinese consumers
due to its unique flavors and fine texture. The annual production of E. sinensis has
increased sharply from 649,000 tonnes in 2011 to 812,000 tonnes in 2016 (China
Fishery Bureau, 2012, 2017). The purpose of this study was to develop a method for
evaluating the tenderness of cooked crab meat and investigate the correlation between
tenderness and histochemistry of edible muscle tissues from E. sinensis, as well as
identify the important histochemical compounds affecting the tenderness of different
muscle tissues from E. sinensis.
2. Materials and methods
aminomethane (Tris) and E-51 resin were obtained from Sinopharm Chemical
N,N,N´,N´-tetramethylethylenediamine (TEMED), sodium dodecyl sulphate (SDS),
and high and low molecular weight markers were bought from Bio-Rad Laboratories
(Hercules, CA, USA). Carnoy's solution was bought from Beijing Solarbio Science &
Technology Co., Ltd. (Beijing, China). HPLC-grade glutaraldehyde, 100% ethanol,
and acetone were purchased from Sigma Aldrich Chemical Co. (St. Louis, MO, USA).
All other chemicals were analytical grade from Ampel Lab Technologies Inc.
2.2 Muscle samples
Twenty males (used for all analyses, 110 ± 4 g) and 6 females (only used for
biometric data analysis, 110 ± 4 g) E. sinensis were harvested in April, 2018, by a
crab company (Chongming Island, Shanghai, China), and promptly tied individually
using cotton rope and transported alive to the laboratory within 2 h in a styrofoam box
with refrigeration (~4oC). The muscle tissues (including claw, abdomen and leg
muscles), gonads, and hepatopancreas as the edible parts were individually removed
and weighed (BT224S, Sartorius Lab Instruments GmbH & Co. KG, Goettingen,
Germany) in a 4-7oC chill storage. For each tissue, yield was calculated as follows:
Yield = each edible part wet weight/body wet weight×100
Edible Contribution = each edible part weight/sun of edible parts weight×100
Each muscle was subsequently collected, mixed, vacuum packed and stored at -70oC for further analysis (a maximum of 4 wk).
P. trituberculatus (male, n = 8, 165 ± 5 g), R. philippinarum (n = 60, 20 ± 1 g), P.
clarkia (n = 50, 27 ± 2 g), C. auratus (n = 10, 155 ± 5 g) were purchased at the
Shanghai seafood market in April, 2018. Whole live samples were steamed at 100oC
for 20 min, then various cooked meat of E. sinensis (from claws, legs, and abdomen),
P. trituberculatus (from abdomen), R. philippinarum (from adductor), P. clarkii (from
abdomen), and C. auratus (from dorsal body) were manually separated using tweezers
with the help of laboratory members and used for tenderness measurements.
2.3 Proximate composition analysis
The moisture was determined using drying in the oven at 105oC until a constant
weight was achieved according to analytical method No. 950.46 (AOAC, 2000). The
crude protein was determined using the Kjeldahl method No. 920.153 (AOAC, 2000).
The conversion factor of total nitrogen to crude protein was 6.25. The fat was
determined using the Soxhlet extraction method No. 960.39 (AOAC, 2000). The ash
was determined using ashing for 8 to 12 h at 550oC according to analytical method No.
928.08 (AOAC, 2000).
2.4 Fractionation of nitrogenous constituents
Proteins in raw crab muscles (including claws, abdominal and legs muscles)
were classified separately according to solubility using the method of Hashimoto et al.
(1979). All operations were done at 4oC. All muscle samples (2.5 g) were put into 10
volumes of buffer A (0.05 M NaCl, 20 mM Tris-HCl, pH = 7.0), and samples were
homogenized at 14,000 rpm for 1.5 min (HG-200, Hsiangtai Machinery Industry Co.,
Ltd., Taipei, Taiwan). Samples were stirred for 0.5 h using a mechanical stirrer (RT10,
IKA®-Werke GmbH & Co., KG, Staufen, Germany) at a speed of 300 rpm. The
extract was centrifuged using a refrigerated centrifuge for 10 min at 12,000×g (9,960
rpm in a JA-12 rotor, Avanti® J-26XP, Beckman Coulter, Inc., Palo Alto, CA, USA).
Centrifugation was done 3 times and all supernatants were combined. PCA was added
to the solution and the final concentration was 5% (w/v). The mixture was centrifuged
at 12,000×g for 5 min at 4oC. The precipitate was mainly the sarcoplasmic proteins
and the remaining supernatant was considered to be non-protein nitrogen, which
included other nitrogen compounds, including amino acids and small peptides.
The residue after extraction with buffer A was stirred with 10 volumes of buffer
B (0.5 M NaCl, 20 mM Tris-HCl, pH = 7.0) for 0.5 h, and centrifuged at 12,000×g for
10 min. The extraction was done 3 times. The supernatants were combined into the
myofibrillar protein fraction. Ten volumes of 0.1 M NaOH were added to the
precipitate, and the mixture was stirred continuously for 2 h. The mixture was then
centrifuged at 12,000×g for 10 min. The extraction was done 4 times. The 4
supernatants were combined into the alkali-soluble protein fraction. The final
precipitate was the stroma protein fraction (alkali-insoluble protein fraction).
Total Kjeldahl nitrogen contents (AOAC, 2000) of the 5 fractions were obtained
along with their protein patterns.
2.5 SDS–polyacrylamide gel electrophoresis (SDS–PAGE)
The protein profiles of raw crab muscles were determined using SDS-PAGE as
described by Laemmli (1970) with slight modifications. The protein samples were
dissolved in electrophoresis buffer solution (containing 2% SDS, 8 M carbamide, 2%
β-mercaptoethanol, 50 mM Tris-HCl, pH = 8.0) and heated at 100oC for 3 min. The
samples (10 µL, contained ~10 µg of protein) were loaded on homemade 7.5%
separating gel and 5% stacking gel, and electrophoresis was carried out with a
constant current of 15 mA/gel. After gel electrophoresis, the gel was stained with
0.1% Coomassie brilliant blue R-250 in a mixed solution of 45% methanol and 10%
acetic acid for 30 min and destained with 45% methanol and 9% acetic acid for 2 h.
Nine molecular weight markers, including myosin heavy chain from rabbit skeletal
muscle (200 kDa), β-galactosidase (116 kDa), phosphorylase B (97.2 kDa), serum
albumin (66.4 kDa), ovalbumin (44.2 kDa), carbonic anhydrase (29 kDa), trypsin
inhibitor (20.1 kDa), lysozyme (14.3 kDa) and aprotinin (6.5 kDa), were used for
estimating protein sizes.
2.6 Histological analysis
Specimen preparation for light microscopy (LM) was done using the method of
Lee et al. (2012). Raw muscles were placed for 40 min in Carnoy's solution,
embedded in paraffin wax, sliced to a thickness of 4-6 µm using a microtome
(RM2235, Leica Microsystems Vertrieb GmbH, Wetzlar, Germany), and stained with
hematoxylin and eosin. Histological observations were done using an optical
microscope (DMI3000, Leica Microsystems) using 100 and 400 × magnification.
Specimens were prepared for transmission electron microscope (TEM) using the
method of Zhen et al. (2018). Muscle tissues (~1 mm3 cube) were separated using a
sharp blade from raw muscles, then fixed in 2.5% glutaraldehyde for 3 h at 4oC,
rinsed in 0.1 M sodium phosphate buffer, and then post-fixed in 1% osmium tetroxide
(OsO4) solution for 2 h at 4oC. After fixation, 0.1 M phosphate buffer was used to
wash the specimens 3 times for 20 min, the specimens were dehydrated for 15 min
each with a series of ethanol and acetone solutions (50, 70 and 90% ethanol, 90%
ethanol:90% acetone at a 1:1 ratio, 90% acetone, and 100% acetone each 3 times). The
samples were embedded in E-51 resin and sliced into ultra-thin sections (60 nm in
thickness). The samples were placed on copper mesh (200 mesh/inch) and stained
with 3% uranyl acetate and 3% lead citrate. The specimens were observed using a
TEM (CM-120, Philips Electronics Ltd., Eindhoven, The Netherlands) at 60 kV.
2.7 Low field nuclear magnetic resonance (LF-NMR) measurements
LF-NMR was used to measure the distribution of water in muscle tissue, which
was done using a Niumag Benchtop Pulsed NMR Analyzer MesoMR23-060H-I
(Niumag Electric Corp., Shanghai, China) operating at a resonance frequency for
protons of 21 MHz. The measurements were done using the method reported by Shao
et al. (2016). The relaxation times using low-field 1H NMR were measured in 3.0 g
portions of fillet placed in sampling tubes (70 mm in diameter). Spin-spin relaxation
times (T2) were collected using a Carr-Purcell-Meiboom-Gill pulse sequence with the
echo times of 500 µs and 2,000 collected echoes at 25oC. T2b, T21 and T22 relax in the
range of 0-10, 30-100, and >200 ms, respectively. The scan was repeated 3 times.
2.8 Water holding capacity (WHC)
Centrifugal and cooking loss reflected the WHC (Li et al., 2017). The WHC was
measured using centrifuge drip and cooking loss. Approximately 3.0 g of raw crab
muscle was weighed in a dry, clean centrifuge tube. The crab muscle was then
centrifuged at 12,000×g at 4oC for 15 min, the liquid at the top of tubes was removed,
and the meat samples were weighed to determine the centrifuge drip. The following
equation was used:
Centrifuge Drip = 100×(1−A/B)
where A is the final weight of the sample, and B is the initial weight of the sample.
The cooking loss of crab meat was measured after steaming. The samples (3.00 g)
were placed in a 5 × 7 cm2 polyethylene bag (Jiangyin Honghao Packaging Materials
Co., Ltd., Jiangyin, Jiangsu, China), then steamed at 100oC for 3 min and immediately
cooled on ice. The meat samples were removed from the bag and placed in a 50 mL
centrifuge tube with a 0.25 mm diameter filter screen (purchased at the Xinxing
experimental equipment market in Haimen, Jiangsu, China) to allow additional drip
for 10 min at 4oC. The cooking loss equation was:
Cooking Loss = 100×(1−A/B)
where A is the weight after cooking, and B is the weight before cooking.
2.9 Instrumental tenderness measurement
Meat samples (3 g) from cooked E. sinensis (including claws, abdomen, and
legs), P. trituberculatus, R. philippinarum, P. clarkii, and C. auratus were finely
blade-minced and added to 5 volumes (v/w) of buffer A (0.05 M NaCl, 20 mM
Tris-HCl, pH = 7.0) in a 50 mL centrifuge tube, then homogenized for 30, 45, 60, 75,
and 90 s using the homogenizer at a speed of 8,000 rpm in the centrifuge tube. Each
homogenate was used for particle size analyses using a Malvern Mastersizer (MS2000,
Malvern Instruments Ltd., Malvern, UK).
With constant stirring, the samples were diluted to 1:500 with distilled water in
the sample cell at 20oC. The particle size was measured in the range of 0.02-2,000 µm.
The specific parameters (Gao et al., 2012) were set as follows: the refractive index of
default particle (glass: 1.520), the particle absorbance value (0.1), and the refractive
index of water (1.330). Each measurement was repeated 3 times. The data are
represented by characteristic parameters, including D[3,2], D[4,3], d(0.1), d(0.5), and
d(0.9). D[3,2] is the Sauter mean particle diameter, which refers to the average
diameter of the equivalent sphere with the same specific surface area as the actual
particle group. D[4,3] is the volumetric mean particle diameter, which represents the
equivalent sphere of the same volume as the actual particle group. d(0.1), d(0.5), and
d(0.9) are the volumetric diameters at which 10, 50, and 90% of the particles are
smaller (Liu et al., 2008).
After treatment with different homogenization times (30, 45, 60, 75, and 90 s),
the rate of the particle size changes of the homogenate were measured. The
corresponding weighted coefficient of each characteristic parameter was obtained
using the entropy weighted coefficient method (Jia and Zhao, 2010; Zhong, 2017).
The characteristic of the entropy weighted coefficient method is that the weighted
coefficient of each characteristic parameter is directly calculated using the rate of the
particle size changes with each characteristic parameter, and the subjective judgment
of the decision maker is not introduced. The instrument tenderness (total breaking rate)
equation was as follows:
Instrument Tenderness = aKD[3,2]+ bKD[4,3]+ cKd(0.1)+ dKd(0.5)+ eKd(0.9)
where a, b, c, d, and e are the corresponding weighted coefficient of Ki, Ki is the rate
of the particle size changes with each characteristic parameter.
Finally, the correlation of instrument tenderness value and sensory tenderness
value was measured to verify the accuracy and reliability of the instrument for
evaluating meat tenderness.
2.10 Sensory analysis
Sensory analysis was done on cooked meat from the 5 species. Sensory analysis
for tenderness was done using a 10-member trained descriptive attribute panel that
was screened, selected, and trained in accordance with the guidelines for the sensory
evaluation of meat of the American Meat Science Association (AMSA, 2015). The
panel consisted of 10 members aged 20-30, 3 males and 7 females from Shanghai
Ocean University. Sensory experiments were carried out in a normal sensory
laboratory, which complied with international standards for test rooms (ISO, 2014).
The test rooms are designed to do sensory evaluations using known and controlled
conditions with a minimum of distractions and to reduce the effects that psychological
factors and physical conditions can have on the panelists judgement. The testing area
is separate to reduce interference, such as from odour and noise. The colour of the
walls and furnishings of the testing area is light neutral grey, and the lighting is
uniform, free from strong shadows. Prior to sensory evaluation, the panelists had
thoroughly discussed 18 aquatic meats including fish, shrimp, shellfish and crab with
words to identify the extremes and middle of the rating scale. As training progressed,
the panelists were able to identify other points along the rating scale. The tenderness
was measured using a quantitative descriptive analysis (QDA) scale with a 9-point
descriptive scale (1 = Extremely Tough, 2 = Very Tough, 3 = Moderately Tough, 4 =
Slightly Tough, 5 = Neither Tender nor Tough, 6 = Lightly Tender, 7 = Moderately
Tender, 8 = Very Tender, 9 = Extremely Tender).
2.11 Statistical analysis
Results were shown as mean values ± standard deviation (SD). The one-way
analysis of variance (ANOVA) with Duncan's multiple range test was used to analyze
the data. The Statistical Package for the Social Sciences (SPSS for Windows: SPSS
Inc., Chicago, IL, USA) was used for statistical analysis and Pearson's method was
used for the correlation analysis. A p-value < 0.05 was considered statistically
significant. However, 0.01 was also used for some of the data to indicate the greater
significance of the differences. All analyses were repeated 3 times.
3. Results and Discussion
3.1 Proximate compositions
The average meat yield from E. sinensis was ~37%, in which muscle made the
greatest contribution, followed by the hepatopancreas, and gonads (Table 1). The
muscle contributions from E. sinensis were higher than those compared with other
marine crab species. For example, Barrento et al. (2010) reported that the muscle and
hepatopancreas of brown crabs were responsible for the majority of meat yield, with
muscle being a larger percentage in male crabs compared to females. The Atlantic
spider crab had an average meat yield of ~17%, in which the hepatopancreas had the
highest contribution, followed by muscle tissue and the gonads (Marques et al., 2010).
This study confirmed that males contributed a higher proportion of muscle from claws
than females of the same size. This species shows sexual dimorphism, displayed in
the larger claws of males compared with females. 14
Based on wet weight, the proximate compositions among all the tissues from E.
sinensis show that claw muscle had the lowest moisture and the highest crude protein
and ash (Table 2). On the other hand, the muscle from the abdomen had the highest
moisture and fat. The total fat in the three tissues was far below 5%, a value generally
considered to characterize a low-fat food. All muscle tissues had crude protein as the
major component. Based on dry weight, the abdomen muscle had the highest crude
protein, followed by claw muscle and leg muscle tissues. The abdomen muscle also
had the highest fat compared to the claw and leg muscles. The differences of ash were
not significant among the different muscles.
These results are consistent with muscle being the structural tissue with the
highest crude protein and poor fat. Previous studies, such as brown crab (Barrento, et
al., 2010) and warty crab (Kaya et al., 2009), are also consistent with these results.
For example, the claw muscle of brown crab from both sexes had 74.6-77.8%
moisture, 16.4-20.5% crude protein, 0.2-0.4% fat, and 1.9-2.2% ash (Barrento, et al.,
2010). For warty crab meat, the crude protein and crude lipid were 19.7 and 0.66
g/100 g wet weight (Kaya et al., 2009). The texture of muscle tissue is mainly
attributed to macromolecular substances. Because crude protein is the major
component of muscle, it has a leading role in the texture of crab meat.
3.2 Distribution of nitrogenous constituents
The various tissues of E. sinensis had different nitrogen distributions (Table 3)
and protein patterns (Fig. 1). The non-protein nitrogen-containing compounds were
mainly related to taste compounds, including amino acids, nucleotides, and peptides.
The sarcoplasmic fraction extracted from muscle tissue is made up of enzymes,
principally glycolytic enzymes, but other enzymes are present as well, including
enzymes related to the pentose shunt and auxiliary enzymes such as creatine kinase
and AMP deaminase (Strasburg et al., 2017). The non-protein nitrogen had a wet
weight of 0.17-0.25 g/100 g, and the sarcoplasmic fraction had a wet weight of
3.2-3.9 g/100 g (Table 3). Fig. 1 shows that proteins with a low molecular weight
(below 97 kDa) were present in the sarcoplasmic fractions of all tissues tested.
The myofibrillar proteins from invertebrate muscles have extraordinarily large,
thick filaments mainly composed of paramyosin (PM) forming a core and myosin as a
surface protein. Myosin and actin (AC) are the main participants in muscle
contraction (Shelud’Ko et al., 1999). The myofibrillar fraction was dominant in three
muscle tissues with significant differences of content. The myofibrillar fraction from
three muscle tissues showed similar SDS-PAGE patterns, showing myosin heavy
chain (MHC) and AC as the major proteins. Myosin is the most abundant protein in
the myofibril and provides an important contribution to the structure and tensile
strength of meat (Marino et al., 2013). PM, which has a molecular weight of 100 kDa,
was also observed, and the PM in the claw muscle was higher than the content in
either the abdomen or leg muscle. Based on the functions of these muscles, PM could
contribute to the strength of the corresponding muscle tissue. Moreover, thermally
induced gelation of PM from scallop smooth adductor muscle was very firm and
brittle. The PM thermal gel was made at temperatures above 30°C using a two-step
increase in elastic (G′) and viscous (G′′) moduli, these values were higher than in gels
produced from actomyosin at a high temperature (Fukuda et al., 2006).
Alkali soluble components arise from polymerized myofibrillar proteins
(Karnjanapratum et al., 2013). Proteins with a molecular weight of ~35, 45, 100, and
200 kDa were observed in the alkali-soluble fraction, corresponding to tropomyosin
(TM), AC, PM, and MHC, respectively. Insoluble components were regarded as
stroma proteins. The connective tissues encasing individual muscle fibers
(endomysium), muscle fiber bundles (perimysium), and muscle (epimysium)
contributed to the “background toughness” of meat (Xiong, 2018). It should be noted
that the stroma proteins in the muscle tissues of E. sinensis was low. Collagen is a
major component of stroma proteins of connective tissue (Xiong, 2018). Mizuta et al.
(1994) reported that the muscles of prawn and shrimp, which had relatively high
collagen, tended to be firm, whereas the muscles in crabs were generally less firm and
the collagen was typically lower. High molecular weight (above 200 kDa) proteins
were still observed in the stroma fraction (Fig. 1).
3.3 Observations of muscle fiber structure using LM and TEM
To better understand the contribution of muscle fiber and connective tissue to
meat texture, it is important to clarify the distribution of these tissues. The transverse
and longitudinal sections of the raw muscle tissues were observed (Fig. 2A) using an
optical microscope. Muscle fiber and connective tissue were stained bright red and
light red, respectively (Fig. 2A). Connective tissue was relatively poor in areas
primarily consisting of muscle tissue. A similar pattern was observed for the muscles
from prawn and shrimp, which showed a dense distribution of collagen fibers, while
the muscles from crabs had a sparse distribution as reported by Mizuta et al. (1994).
Those findings were consistent with the very low muscle stroma protein observed in
the nitrogen distribution analyses. The muscle bundle diameter of muscle from the
abdomen was the largest, followed by the claw and leg (Fig. 2A).
Further observations of the ultrastructure of the raw muscles using longitudinal
sections allowed for the determination of striated muscles formed by overlap of thin
and thick filaments (Fig. 2B). Franzini (1970) observed that the length of thin and
thick filaments was highly variable within a single crab’s fiber, therefore, a
comparison made on the basis of a few random measurements would not valid.
Sarcomere length is a measure of the state of postmortem raw muscle shortening
(Locker and Hagyard, 1963), however, in this study, the sarcomere changes of raw
muscle in the post-mortem period were not investigated.
In transverse sections, the lattice distribution formed by thick microfilaments was
visible, with thin filaments surrounding the hexagonally arranged thick filaments (Fig.
2B). The average diameters of thick microfilaments of the claw, abdomen and leg
were 21 ± 2, 23 ± 3, and 19 ± 2 nm (p < 0.05), respectively. The thick microfilaments
were rarely observed in a regular pattern in the three E. sinensis. On the other hand,
Squire et al. (1981) reported the thick microfilaments of vertebrate skeletal muscle
could form a regular lattice arrangement with 6 thick microfilaments arranged
equidistantly around each thick microfilament. Additionally, in other studies, the
reseachers observed that a more or less hexagonally shaped thick filament lattice was
invariably present in ultrathin transverse sections of marine crabs (Cohen and Hess,
1967; Franzini, 1970; Fourtner and Sherman, 1972; Hoyle, 1978). It was speculated
that this was closely related to the maturity of the crab and the tenderness of the
muscle. As the crab matures, the lattice regularity of the thick microfilaments
increased, and the tenderness of the muscle decreased. In future research, the
arrangement of thick microfilaments in cooked muscle should be studied.
3.4 Water distributions and WHC
The muscle tissue water was split into three segments for more detailed analyses.
Fig. 3 shows the water distributions with NMR, and WHC of the raw samples. It was
observed that the entrapped water was the highest, followed by bound water and free
water. Amongst all the raw muscles, muscle from the abdomen had the highest free
water, which could contribute to the higher moisture. The WHC of meat can be
defined as its ability to retain water and this has a direct influence on its tenderness
and yield, thereby influencing its economic value (Guichard, 2006). Centrifuge drip
and cooking loss are the primary indicators of WHC of raw and cooked meat,
respectively (Silva et al., 2015). The highest centrifuge drip and cooking loss were
observed in the abdomen muscle. The majority of the water that was affected by the
process of converting muscle to meat was the entrapped water. Bertram et al. (2002)
reported that more mobile myowater within the meat (extra-myofibrillar water) was to
be considered as the source of the potential drip from the meat. From the indices of
centrifuge drip and cooking loss, the claw muscle showed the highest WHC compared
to the abdomen and leg muscle.
3.5 Instrumental measurement and sensory evaluation of meat tenderness
Fig. 4 shows the distribution of particle size with different homogenization times.
The curve shifted to the left with the increasing of homogenization times, and the
specificity of the curve shape of different aquatic animals is shown. To quantify
tenderness, the variation of particle size with different characterization parameters is
shown in Fig. 5. The absolute value of the linear slope equation represents the
breaking rate of meat fiber with each characteristic parameter. The entropy weighted
coefficient method was used to calculate the corresponding weighted coefficient of
the breaking rate with each characteristic parameter and to obtain the instrument
tenderness value (Table 4). The weighted coefficients of D[3,2], D[4,3], d(0.1), d(0.5),
and d(0.9) were 27.9, 9.3, 40.3, 18.0, and 4.4%, respectively. The corresponding
instrumental tenderness values were ranked as: ES-claw > ES-leg > ES-abdomen >
CA-dorsal > PC-abdomen > PT-abdomen > RP-adductor. These results showed that
the breaking rate of the 3 edible muscles of E. sinensis was higher than that of other
species with the same homogenization speed, and the relationship between the
breaking rate of the muscles of E. sinensis was: claw meat > leg meat > abdomen
meat. In other words, crab meat was more tender than other species, and the claw
meat was the most tender, followed by leg meat. The tenderness correlation (R2 =
0.910) between the instrumental measurement and sensory evaluation was highly
significant (p < 0.01) (Fig. 6). Therefore, the breaking rate using the entropy weighted
coefficient method analysis could be quantified to reflect the tenderness of different
muscle tissues. When the same mechanical homogenization was used to shear the
meat, the particle size of the muscle fiber became smaller as the breaking times
increased. Moreover, for meat of different tenderness, the rate of reduction rate of
muscle fiber particle size was different, and the meat was more tender, and the
breaking rate was larger. The raw material properties were the basis to establish the
method of instrumental tenderness measurement. Therefore, this micro method is very
suitable for cooked meat with low-fat, low connective tissue, and meat which is hard
to cut into specific shapes compared with the shear force method.
3.6 Correlation analysis
The correlation analysis between various histochemical and textural indices of E.
sinensis are shown in Table 5. There was a positive correlation between moisture and
water loss, and a negative correlation between moisture and tenderness. Moreover,
water loss was negatively correlated with tenderness, which meant that the higher the
moisture, the weaker the WHC and less tender the cooked meat. A significant
relationship was observed between free water and the centrifuge drip (r = -0.785) and
the cooking loss (r = -0.842). The main constituent of fresh meat, water, was mainly
held within the highly organized structures of myofibrillar proteins. (Bertram et al.,
2006) A high amount of intra-myofibrillar water and a low amount of
extra-myofibrillar water might be associated with more tender meat (Pearce et al.,
2011). The distribution and mobility of intra-myofibrillar water were highly
associated with the structural features of the myofibrillar protein network (Wu et al.,
2006). The increased WHC would be consistent with higher protein-water interaction
and/or smaller interstitial water domains (Sánchez-González et al., 2008).
Myofibrillar protein is soluble in salt solutions, and is responsible for much of the
functional characteristics of fresh and processed meat (Xiong, 2018). These results
showed that myofibrillar protein was negatively correlated with water loss and
positively correlated with tenderness, which meant that a high myofibrillar protein
was associated with an improved WHC and tenderness of cooked meat. A significant
relationship was observed between the diameter of the thick microfilaments and the
entrapped water (r = 0.809). The change in the intra-myofibrillar myowater
characteristics was consistent with the increase in protein content/density of the
muscles (Xiong, 2018). However, no significant correlation was observed between the
diameter of the thick microfilaments and meat texture. Based upon the ultrastructure,
the irregular arrangement of the thick microfilaments might be closely related to the
high tenderness of E. sinensis meat. Little research has been done on the dynamic
changes of molecular information about functional groups, bonding types, molecular
conformations, and water distribution. Therefore, a subsequent study of the
protein-protein and water-protein interactions during the cooking would be beneficial.
The characteristics of food raw materials were shown using the histochemical and
textural characteristics of different muscles of E. sinensis. Based on histochemical
properties, it was observed that the breaking rate of muscle fiber after entropy
weighted coefficient method analysis could be quantified to reflect the meat
tenderness of cooked crab, which was very suitable for meat with low-fat, low
connective tissue, and cooked meat which is hard to cut into specific shapes.
Tenderness had a high correlation with moisture, myofibrillar proteins and the water 22
holding capacity. Ultrastructural and physicochemical properties could be the focus of
a follow-up study which would track the textural change of crab meat, analyze
physiological and biochemical changes of the myofibrillar protein with cooking the
reducing tenderness, and further identify proteins indicators of meat tenderness.
Authors acknowledge Professor Peng Yu at the Fudan University (Shanghai,
China) for the help with the TEM. This study was supported by grants from the 2018
National Key R&D Program of China (Grant No.2018YFD0901006), the 2018
National Natural Science Foundation of China (Grant No.31471608), and the 2018
Industrial System Project of Chinese Mitten Crab of Shanghai Municipal Agriculture
Commission (Grant No.D-8004-16-0179).
Conflict of Interest
The authors declare no competing financial interest.
AMSA. (2015). Research Guidelines for Cookery, Sensory Evaluation and Instrumental Tenderness of
Fresh Meat. (2th ed.). Champaign, IL, USA: American Meat Science Association, National
Live Stock and Meat Board. pp 10-85.
496 497 498
AOAC. (2000). Official Methods of Analyses of the Association of Analytical Chemist. (15th ed.). Gaithersburg, MD, USA: Association of Official Analytical Chemists. Barrento, S., Marques, A., Teixeira, B., Mendes, R., Bandarra, N., Vaz-Pires, P., & Nunes, M. L. (2010).
Chemical composition, cholesterol, fatty acid and amino acid in two populations of brown
crab Cancer pagurus: Ecological and human health implications. Journal of Food
Composition and Analysis, 23(7), 716-725.
Bertram, H. C., Kohler, A., Böcker, U., Ofstad, R., & Andersen, H. J. (2006). Heat-induced changes in
myofibrillar protein structures and myowater of two pork qualities. A combined FT-IR
spectroscopy and low-field NMR relaxometry study. Journal of Agricultural & Food
Chemistry, 54(5), 1740-1746.
Benjakul, S., & Sutthipan, N. (2009). Muscle changes in hard and soft shell crabs during frozen storage. LWT - Food Science and Technology, 42(3), 723-729.
Bertram, H. C., Purslow, P. P., & Andersen, H. J. (2002). Relationship between meat structure, water
mobility, and distribution: A low-field nuclear magnetic resonance study. Journal of
Agricultural and Food Chemistry, 50(4), 824-829.
Boleman, S. J., Boleman, S. L., Miller, R. K., Taylor, J. F., Cross, H. R., Wheeler, T. L., Koohmaraie,
M., Shackelford, S. D., Miller, M. F., West, R. L., Johnson, D. D., & Savell, J. W. (1997).
Consumer evaluation of beef of known categories of tenderness. Journal of Animal Science,
Castilho, G. G., Ostrensky, A., Pie, M. R., & Boeger, W. A. (2007). Morphology and histology of the
male reproductive system of the mangrove land crab Ucides cordatus (L.) (Crustacea,
Brachyura, Ocypodidae). Acta Zoologica, 89(2), 157-161.
Çelik, M., Türeli, C., Çelik, M., Yanar, Y., Erdem, Ü., & Küçükgülmez, A. (2004). Fatty acid
composition of the blue crab (Callinectes sapidus Rathbun, 1896) in the north eastern
Mediterranean. Food Chemistry, 88(2), 271-273.
521 522 523 524 525 526 527 528
Cherif, S., Frikha, F., Gargouri, Y., & Miled, N. (2008). Fatty acid composition of green crab (Carcinus mediterraneus) from the Tunisian mediterranean coasts. Food Chemistry, 111(4), 930-933. China Fishery Bureau. (2012). China Fisheries Yearbook. Beijing: Chinese Agriculture Express. pp 36. (in Chinese) China Fishery Bureau. (2017). China Fisheries Yearbook. Beijing: Chinese Agriculture Express. pp 34. (in Chinese) Cohen, M. J., & Hess, A. (1967). Fine structural differences in “fast” and “slow” muscle fibers of the crab. American Journal of Anatomy, 121(2), 285-303.
Dima, J. B., Barón, P. J., & Zaritzky, N. E. (2012). Mathematical modeling of the heat transfer process
and protein denaturation during the thermal treatment of Patagonian marine crabs. Journal of
Food Engineering, 113(4), 623-634.
Fourtner, C. R., & Sherman, R. G. (1972). A light and electron microscopic examination of muscles in
the walking legs of the horseshoe crab, Limulus polyphemus (L.). Canadian Journal of
Zoology, 50(11), 1447-1455.
535 536 537 538
Franzini-Armstrong, C. (1970). Natural variability in the length of thin and thick filaments in single fibres from a crab, Portunus depurator. Journal of Cell Science, 6(2), 559-592. Fukuda, N., Fujiura, M., Kimura, M., Nozawa, H., & Seki, N. (2006). Thermally induced gelation of paramyosin from scallop adductor muscle. Fisheries Science, 72(6), 1261-1268.
Gao, F. F., Wang, Z. G., Peng, Z. Q., Jin, H. G., Wang R. R., Zhang Y. W., & Yao, Y. (2012). Effect of
chopping time on particle size distribution in meat batters. Food Science, 33(5), 74-77.
Guichard, E. (2006). Flavour retention and release from protein solutions. Biotechnology Advances,
Gökoðlu, N., & Yerlikaya, P. (2003). Determinaton of proximate composition and mineral contents of
blue crab (Callinectes sapidus) and swim crab (Portunus pelagicus) caught off the Gulf of
Antalya. Food Chemistry, 80(4), 495-498.
546 547 548 549
Hashimoto, K., Watabe, S., Kono, M., & Shiro, M. (1979). Muscle protein composition of sardine and mackerel. Nippon Suisan Gakkaishi, 45(11), 1435-1441. Hayashi, T., Yamaguchi, K., & Konosu, S. (1981). Sensory analysis of taste-active components in the extract of boiled snow crab meat. Journal of Food Science, 46(2), 479-483.
Hernández-Robledo, V., Martínez Maldonado, M. Á., Uresti-Marín, R. M., Ramírez, J. A., & Velázquez,
G. (2017). Effect of washing treatment and microbial transglutaminase on the gelling
properties of blue crab (Callinectes sapidus) proteins. CyTA - Journal of Food, 15(2),
554 555 556 557 558 559
Hoyle, G. (1978). Distributions of nerve and muscle fibre types in locust jumping muscle. Journal of Experimental Biology, 73(1), 205-233. ISO, I. (2014). Sensory Analysis - General Guidance for the Design of Test Rooms. (ISO 8589: 2007/Amd 1: 2013). Lisboa, Portugal: Instituto Português da Qualidade. Jia, Z. Y., & Zhao, L. (2010). Comprehensive evaluation of power quality based on the model of entropy weight and unascertained measure. Power System Protection & Control, 33-37.
Jiang, H., Yoon, S. C., Zhuang, H., Wang, W., Lawrence, K. C., & Yang, Y. (2018). Tenderness
classification of fresh broiler breast fillets using visible and near-infrared hyperspectral
imaging. Meat Science, 139, 82-90.
KaiĆ, A., & ŽGur, S. (2017). The effect of structural and biochemical changes of muscles during
post-mortem process on meat tenderness. Journal of Central European Agriculture, 18(4),
Karnjanapratum, S., Benjakul, S., Kishimura, H., & Tsai, Y. H. (2013). Chemical compositions and
nutritional value of Asian hard clam (Meretrix lusoria) from the coast of Andaman Sea. Food
Chemistry, 141(4), 4138-4145.
Kaya. Y., Turan. H., & Erdem, M. E. (2009). Determination of nutritional quality of warty crab
(Eriphia verrucosa Forsskal, 1775). Journal of Animal & Veterinary Advances, 8(1), 120-124.
King, D. A., Wheeler, T. L., Shackelford, S. D., & Koohmaraie, M. (2009). Comparison of palatability
characteristics of beef gluteus medius and triceps brachii muscles. Journal of Animal Science,
574 575 576 577
Laemmli, U. K. (1970). Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature, 227(5259), 680-685. Lee, J. S., Lee, Y. G., Park, J. J., & Shin, Y. K. (2012). Microanatomy and ultrastructure of the foot of the infaunal bivalve Tegillarca granosa (Bivalvia: Arcidae). Tissue Cell, 44(5), 316-324.
Li, D., Jia, S., Zhang, L., Li, Q., Pan, J., Zhu, B., Prinyawiwatkul, W., & Luo, Y. (2017). Post-thawing
quality changes of common carp (Cyprinus carpio) cubes treated by high voltage electrostatic
field (HVEF) during chilled storage. Innovative Food Science & Emerging Technologies, 42,
Liu, L. X., Marziano, I., Bentham, A. C., Litster, J. D., White, E. T., & Howes, T. (2008). Effect of
particle properties on the flowability of ibuprofen powders. International Journal of
Pharmaceutics, 362(1-2), 109-117.
Locker, R. H., & Hagyard, C. J. (1963). A cold shortening effect in beef muscles. Journal of the Science of Food & Agriculture, 14(11), 787-793.
Marino, R., Albenzio, M., Caroprese, M., Napolitano, F., Santillo, A., & Braghieri, A. (2011). Effect of
grazing and dietary protein on eating quality of Podolian beef. Journal of Animal Science,
Marino, R., Albenzio, M., Della Malva, A., Santillo, A., Loizzo, P., & Sevi, A. (2013). Proteolytic
pattern of myofibrillar protein and meat tenderness as affected by breed and aging time. Meat
Science, 95(2), 281-287.
Marques, A., Teixeira, B., Barrento, S., Anacleto, P., Carvalho, M. L., & Nunes, M. L. (2010).
Chemical composition of Atlantic spider crab Maja brachydactyla: Human health implications.
Journal of Food Composition and Analysis, 23(3), 230-237.
Martínez, M. A., Robledo, V., Velazquez, G., Ramírez, J. A., Vázquez, M., & Uresti, R. M. (2014).
Effect of precooking temperature and microbial transglutaminase on the gelling properties of
blue crab (Callinectes sapidus) proteins. Food Hydrocolloids, 35, 264-269.
Mizuta, S., Yoshinaka, R., Sato, M., & Sakaguchi, M. (1994). Characterization of collagen in the
muscle of several crustacean species in association with raw meat texture. Fisheries Science,
Moholisa, E., Hugo, A., Strydom, P. E., & van Heerden, I. (2017). The effects of animal age, feeding
regime and a dietary beta-agonist on tenderness of three beef muscles. Journal of the Science
of Food and Agriculture, 97(8), 2375-2381.
MØller, A. J., Vestergaard, T., & Wismer-Pedersen, J. (1973). Myofibril fragmentation in bovine
longissimus dorsi as an index of tenderness. Journal of Food Science, 38(5), 824-825.
Pearce, K. L., Rosenvold, K., Andersen, H. J., & Hopkins, D. L. (2011). Water distribution and
mobility in meat during the conversion of muscle to meat and ageing and the impacts on fresh
meat quality attributes - A review. Meat Science, 89(2), 111-124.
Pool, M. F., & Klose, A. A. (1969). The relation of force to sample dimensions in objective
measurement of tenderness of poultry meat. Journal of Food Science, 34(6), 524-526.
Rhee, M. S., Wheeler, T. L., Shackelford, S. D., & Koohmaraie, M. (2004). Variation in palatability and
biochemical traits within and among eleven beef muscles. Journal of Animal Science, 82(2),
Shao, J. H., Deng, Y. M., Jia, N., Li, R. R., Cao, J. X., Liu, D. Y., & Li, J. R. (2016). Low-field NMR
determination of water distribution in meat batters with NaCl and polyphosphate addition.
Food Chemistry, 200, 308-314.
Shelud’ko, N. S., Tuturova, K. F., Permyakova, T. V., Plotnikov, S. V., & Orlova, A. A. (1999). A novel
thick filament protein in smooth muscles of bivalve molluscs. Comparative Biochemistry &
Physiology B: Biochemistry and Molecular Biology, 122(3), 277-285.
Silva, D. R., Torres Filho, R. A., Cazedey, H. P., Fontes, P. R., Ramos, A. L., & Ramos, E. M. (2015).
Comparison of Warner-Bratzler shear force values between round and square cross-section
cores from cooked beef and pork Longissimus muscle. Meat Science, 103, 1-6.
Skonberg, D. I., & Perkins, B. L. (2002). Nutrient composition of green crab (Carcinus maenus) leg meat and claw meat. Food Chemistry, 77(4), 401-404.
Squire, J. (1981). Comparative ultrastructures of diverse muscle types. In Squire, J. (Ed.) The
Structural Basis of Muscular Contraction. Boston, MA, USA: Springer. pp 381-469.
Strasburg, G. M., & Xiong, Y. L. (2017). Physiology and chemistry of edible muscle tissues. In
Damodaran, S., & Parkin, K. L. (Eds). Fennema's Food Chemistry. Boca Raton, FL, USA:
CRC Press. pp 955-1015.
Sánchez-González, I., Carmona, P., Moreno, P., Borderías, J., Sánchez-Alonso, I., Rodríguez-Casado,
A., & Careche, M. (2008). Protein and water structural changes in fish surimi during gelation
as revealed by isotopic H/D exchange and Raman spectroscopy. Food Chemistry, 106(1),
Wu, Z., Bertram, H. C., Kohler, A., Böcker, U., Ofstad, R., & Andersen, H. J. (2006). Influence of
aging and salting on protein secondary structures and water distribution in uncooked and
cooked pork. A combined FT-IR microspectroscopy and 1H NMR relaxometry study. Journal
of Agricultural & Food Chemistry, 54(22), 8589-8597.
Xiong, Y. L. (2018). Muscle proteins. In Yada, R. Y. (Ed.). Proteins in Food Processing. Philadelphia, PA, USA: Woodhead. pp 127-148.
Zhen, G., Chen, H., Tsai, S. Y., Zhang, J., Chen, T., & Jia, X. (2018). Long-term feasibility and
biocompatibility of directly microsurgically implanted intrafascicular electrodes in free
roaming rabbits. Journal of Biomedical Materials Research Part B: Applied Biomaterials,
Zhong, F., Hou, M., He, B., & Chen, I. (2017). Assessment on the coupling effects of drip irrigation
and organic fertilization based on entropy weight coefficient model. PeerJ, 5, e3855.
Table 1 The biometric data of edible tissues from E. sinensis Group
648 649 650
Composition (g/100 g wet weight) Abdomen Leg Gonad
Data are mean ± standard deviation (n = 6). Different lowercase letters in the same row indicate significant difference (p < 0.05). EC: Edible contribution.
Table 2 Proximate compositions of claw, abdomen, and leg muscles from E. sinensis Tissue Claw Abdomen Leg
652 653 654
Moisture 78±1 c
Composition (%) Crude Protein Fat
(90±2) A 17±0.4 b
(1.3±0.3) A 0.13±0.01 b
(10±1) A 1.9±0.1 b
Data are mean ± standard deviation (n = 3). Values in the parenthesis represent the content based on dry weight. Different lowercase or uppercase letters in the same column indicate significant difference (p < 0.05). ES: E. sinensis.
Table 3 Nitrogen distribution in claw, abdomen and, leg muscles from E. sinensis Nitrogen distribution (g/100 g wet weight) Tissue
656 657 658
Data are mean ± standard deviation (n = 3). Values in the parenthesis represent the content based on total protein. Different lowercase or uppercase letters in the same column indicate significant difference (p < 0.05).
660 661 662
Table 4 The absolute value (µm/s) of the linear slope equation and the corresponding weighted coefficients (%) with different characteristic parameters, and instrument tenderness value (µm/s) of different aquatic animals Tissue
Data are mean ± standard deviation (n = 3). Different lowercase letters in the same column indicate significant difference (p < 0.05).
Table 5 The correlation analysis between various histochemical and textural indices
of E. sinensis.
668 669 670 671 672
EW: Entrapped water, FW: Free water, MF: Myofibrillar protein, DTF: Diameters of thick microfilaments, CD: Centrifuge drip, CL: Cooking loss, IT: Instrumental tenderness, ST: Sensory tenderness, * Significant at the 5% level, ** Significant at the 1% level.
Fig. 1. SDS–PAGE patterns of different fractions of claw, abdomen, and leg muscles
from E. sinensis. W, S, M, A, T denotes whole portion, sarcoplasmic, myofibrillar,
alkali-soluble, and stroma fractions, respectively.
Fig. 2. Light microscopical microscopy (A) and transmission electron microscopy (B)
of the claw, abdomen, and leg muscles from E. sinensis. MB: muscle bundle.
Fig. 3. Water distributions, centrifuge drip, and cooking loss of the claw, abdomen,
and leg muscles from E. sinensis. BW, EW, FW, CD and CL denotes bound water,
entrapped water, free water, centrifuge drip, and cooking loss, respectively. Different
letters indicate significant differences (p < 0.05) between muscles.
Fig. 4. The variation of particle size with different characterization parameters.
Fig. 5. The distribution of particle size of different aquatic animals with different
homogenization times. ES, PT, RP, PC, and CA denoted E. sinensis, P. trituberculatus,
R. philippinarum, P. clarkii, C. auratus, respectively. Different letters indicate
significant differences (p < 0.05) between times.
Fig. 6. The correlation analysis between instrumental measurement and sensory
evaluation of 7 muscle tissues from different aquatic animals. Different letters
indicate significant differences (p < 0.05) between aquatic animals.
692 693 694
696 697 698 699
80 BW EW FW CD CL
Content ( g/100g wet weight )
700 701 702
704 705 706
c d d
Partical Size (µ m)
Partical Size (µ m)
b a b
a b b
d b c
Partical Size (µ m)
Partical Size (µ m)
b a a
c b b
c d c c
a a 100
b Partical Size (µ m)
a a a a
b b b
b b b
c c bc b
bc cd d c c
d e d
ES-claw ES-abdomen ES-leg PT-abdomen RP-adductor PC-abdomen CA-dorsal
711 712 713
Highlights The breaking rate of muscle fiber based on particle size analysis can be quantified to reflect the tenderness of different muscle tissues from Eriocheir sinensis. This method is very suitable for cooked meat with low-fat, low connective tissue, and meat which is hard to cut into specific shapes. Higher myofibrillar protein suggested a stronger water holding capacity, and more tender meat.