Effects of dietary protein source on the digestive enzyme activities

and electrolyte composition in the small intestinal fluid of chickens

L. Q. Ren,* F. Zhao,*1 H. Z. Tan, J. T. Zhao, J. Z. Zhang,* and H. F. Zhang*
*The State Key Laboratory of Animal Nutrition, Institute of Animal Sciences, Chinese Academy of Agricultural
Sciences, Beijing 100193, China; and Wens Foodstuffs Group Corporation Co. Ltd., Guangzhou 527439, China
eral, no significant differences between the 2 dietary
groups were observed for the mean of duodenal, jejuna,
and ileal amylase, trypsin, chymotrypsin, and lipase,
respectively. However, the duodenal amylase activity
was lower in the CMD group than the CSMD group
(P < 0.05), which was probably related to the lower
duodenal pH value that was observed in this group (P
< 0.01). A higher jejunal Mg2+ concentration was observed in chickens that were fed the CMD (P < 0.05),
whereas the differences in the Na+, K+, Cl, and Ca2+
concentrations in the small intestine were not significant between the 2 diets (P > 0.05). In conclusion, the
digestive enzymes and electrolytes in the small intestinal fluid of chickens adapted to the protein source of
the diet, and each segment of the small intestine displayed different modifications.

Key words: chicken, dietary protein source, digestive enzyme activity,

electrolyte composition, small intestinal fluid
2012 Poultry Science 91:16411646
http://dx.doi.org/10.3382/ps.2011-02081

INTRODUCTION
The small intestine is the primary digestion and absorption site of dietary nutrients. The digestive enzymes
and electrolytes in the intestinal fluid that are secreted
by the pancreas, intestinal glands, and mucosal cells are
responsible for the hydrolysis of dietary macronutrients
and play vital roles in regulating products for transport.
Several studies have shown that protease activities in
the intestinal fluid are changed proportionately in response to the amounts of protein in the diet, whereas
the amylase and lipase activities are dependent on their
respective substrate carbohydrate and lipid contents
(Corring, 1980; Valette et al., 1992; Yago et al., 1997;
Zhao et al., 2007). Snook and Meyer (1964) reported
that when rats received whole-egg protein, trypsin and
chymotrypsin activities increased within intestinal con2012 Poultry Science Association Inc.
Received December 7, 2011.
Accepted March 16, 2012.
1 Corresponding author: zsummit@163.com

tents when compared with a casein diet, which indicated that the types of ingested protein sources could
affect digestive enzyme secretion. As a component of
intestinal fluid, electrolytes in the intestinal tract are
required for proper digestive enzyme functionality and
the absorption of dietary hydrolysates (Clemens and
Maloiy, 1978; Knarreborg et al., 2003). Sullivan et al.
(1974) reported that magnesium and calcium could
bind protein and were correlated with the amylase and
protease activities in the pancreatic juice of pigs, which
meant dietary composition could affect the ion concentration of intestinal fluid. In the practical feed formulation for Chinese yellow chickens, corn is the main
source of cereal grain, whereas the proteinaceous ingredient varies greatly. To reduce the feed costs, soybean
meal is usually substituted with relative low-quality
protein feed, such as cottonseed meal, peanut meal, or
rapeseed meal. However, the antinutritional ingredients
and imbalanced digestible amino acids in the diet may
have related influences on the activities of intestinal
digestive enzymes and the concentration of ion in intestinal fluid. Few data were reported on the relationship

1641

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ABSTRACT An experiment was conducted to investigate the effects of dietary protein source on the digestive enzymes and electrolyte composition of digesta fluid in the duodenum, jejunum, and ileum of chickens. A
2 3 factorial and completely randomized design that
used 2 types of diets that differed only in their protein
source [a corn-soybean meal-miscellaneous meal diet
(CSMD) and a corn-miscellaneous meal diet (CMD)]
and 3 types of cannulated roosters (duodenal, jejunal,
and ileal cannulations) was adopted. The experiments
included 5 replicates for each of the 6 treatments, and
each replicate involved 3 cannulated chickens. The digesta samples were collected for 1 h every 4 h between
09:30 and 18:30 h on d 31, 33, and 35 of the experiment. The amylase, lipase, trypsin, and chymotrypsin
activities and the electrolyte composition in the duodenal, jejunal, and ileal fluid were determined. In gen-

1642

Ren et al.
Table 1. Composition and nutrient content of the pre-experimental and experimental diets on DM
basis
Ingredient (%)

CSMD1

CMD1

72.16
17.20

2.80
3.00
0.90
1.62
1.00
0.36
0.28
0.38
0.15
0.05
0.10

2,900
17.05
3.80
2.77

0.94
0.54

63.65
7.97
3.00
3.00
3.00
4.00
4.60
3.00

3.00
1.34
1.10
1.00
0.60
0.29
0.15
0.12
0.08
0.10

3,150
19.15
6.96
3.72
12.36
3.94
50.35
0.89
0.63

64.37

6.00
5.17
4.00
5.00
4.47
3.00

3.00
1.28
1.14
1.00
0.78
0.29
0.16
0.12
0.12
0.10

3,150
18.66
7.22
3.40
13.42
4.16
48.14
0.89
0.63

1CSMD

= corn-soybean meal-miscellaneous meal diet, CMD = corn-miscellaneous meal diet.

per kilogram of diet: vitamin A (retinyl acetate), 2,700 IU; vitamin D3 (cholecalciferol), 400 IU;
vitamin E (dl--tocopheryl acetate), 10 IU; vitamin K3 activity, 0.5 mg; thiamine, 2.0 mg; riboflavin, 5.0 mg;
pantothenic acid, 10.0 mg; niacin, 30 mg; pyridoxine, 3.0 mg; choline, 750 mg; folic acid, 0.5 mg; biotin, 120 g;
vitamin B12, 10 g; manganese (MnSO4H2O), 80 mg; zinc (ZnSO4), 80 mg; copper (CuSO45H2O), 8 mg; iron
(FeSO47H2O), 80 mg; iodine (KI), 0.7; and selenium (Na2SeO3), 0.3 mg.
3The values are calculated according to the AME values of feedstuffs for chickens (Ministry of Agriculture of
China, 2004).
4The values are determined values. ADF = acid detergent fiber; NDF = neutral detergent fiber.
2Supplied

between dietary nutrient content and digestive enzyme

activities, ion concentration in intestinal fluid of duodenum, jejunum, and ileum of chicken, respectively. This
information is important to understand how to improve
the utilization of dietary nutrients formulated with lowquality protein ingredients. Recently, our laboratory
has designed intestinal cannula to repeatedly collect a
great volume of digesta from intestine of live birds for
analyzing the composition of intestinal fluid (Zhao et
al., 2007). With this method, the current study was
conducted to determine the effects of dietary protein
source on the digestive enzyme activities and electrolyte composition of digesta fluid in the duodenum, jejunum, and ileum of chickens.

MATERIALS AND METHODS

Birds and Diets
This experiment was conducted at Guangdong Wens
Foodstuffs Group Co. Ltd. and was approved according to the animal care and handling procedures at the
Institute of Animal Sciences, Chinese Academy of Agricultural Sciences, Beijing. A 3 2 factorial, completely

randomized design was used for the experiment. One

hundred and twenty 15-wk-old Chinese yellow roosters
were selected by weight (2.3 to 2.5 kg) and randomly
divided into 3 groups with 40 birds per group. All of the
birds were placed into individual cages (0.50 m length
0.42 m width 0.55 m height) in a temperaturecontrolled room (25C) under 12 h of light per day. The
birds from 1 group were fitted with cannulas in the
duodenum, the remaining 2 groups were cannulated in
the jejunum or ileum according to procedures described
by Zhao et al. (2007). The duodenal cannula was fitted
on the U-shaped corner of the duodenum, whereas the
jejunal cannula was fitted 1.5 to 2.0 cm from Meckels
diverticulum of the jejunum, and the ileal cannula was
placed parallel to the end of a pair of ceca. After the intestinal cannulation, feed was withdrawn and the birds
were tube-fed 20 mL of a dextrose solution (20 g/100
g of water) 3 times daily for 3 d. Then, birds were fed
a pre-experimental diet (Table 1) from d 4 to 15 of the
experiment. On d 16, 30 of the 40 cannulated chickens
in each group were randomly selected and divided into
2 subgroups. There were 5 replicates with 3 cannulated
chickens each. Two subgroups of each cannulated chicken group were submitted to a 15-d adaptation period

Downloaded from http://ps.oxfordjournals.org/ by guest on October 12, 2016

Corn
Soybean meal, 43% CP
Cottonseed meal, 42% CP
Peanut meal, 45.5% CP
Rapeseed meal, 36% CP
Corn gluten meal, 60% CP
Soybean oil
Wheat middings
Wheat bran
Expanded soybean
Calcium hydrophosphate
Limestone
Premix2
l-Lys (65%)
Sodium chloride
dl-Met (solid)
Sodium bicarbonate
l-Thr
Choline chloride (50%)
Nutrient content
ME3 (kcal/kg)
CP4 (%)
Crude fat4 (%)
Crude fiber4 (%)
ADF4 (%)
NDF4 (%)
Starch4 (%)
Calcium (%)
Total P (%)

Pre-experimental
diet

PROTEIN SOURCE ON DIGESTIVE ENZYME ACTIVITIES

1643

Sample Collection and Chemical Assay

Statistical Analysis

Digesta were collected for 1 h every 4 h from 09:30

to 18:30 h on d 31, 33, and 35 of the experiment, and
the collection method was similar to the procedure described by Zhao et al. (2007). The birds had free access
to feed and water during the digesta collection. The
small intestinal fluid was made by centrifuging digesta
samples for 10 min at 1,250 g at 4C according to
the method that was described by Furuya et al. (1979),
and then the supernatant was stored at 20C. After
the digesta collection, the frozen sample was thawed in
4C. One milliliter of digesta fluid from each of the 3
collection times and 3 collection days was pooled and
vortexed for the analysis of enzyme activities and electrolyte composition.
Amylase activity was measured with soluble starch
as a substrate at pH 6.9 and using the method that
was described by Dahlqvist (1962). Trypsin activity
was assayed with Na-p-toluolsulfonyl-l-arginine methyl
ester hydrochloride (TAME) as a substrate at pH 8.1
according to procedures described by Wirnt (1974a,b).
A similar method was used for the chymotrypsin determination, but TAME replaced N-benzoyl-l-tyrosine
ethyl ester (BTEE) at pH 7.8; the TAME and BTEE
were obtained from Sigma-Aldrich (St. Louis, MO). Lipase activity was determined using a Diasys reagent
box (Lipase DC FS, DiaSys Diagnostic Systems GmbH,
Holzheim, Germany). All of the digestive enzyme activities were expressed in U/mL of the intestinal fluid.
The pH value of the intestinal fluid was measured with
a pH meter at 39C (PB-10, Sartorius, Germany). The
Na+, K+, and Cl concentrations were determined us-

The data were tested for homogeneity of variance using the Levene test (Milliken and Johnson, 1984), data
were then analyzed by ANOVA according to a completely randomized design with treatments arranged in
a 2 3 factorial arrangement using the GLM procedures of SAS version 8 (SAS Institute, 1990). An interaction was included in the statistical model as Y = diet
effect + intestinal effect + diet intestine effect + error. Treatment means were separated using orthogonal
contrasts with P < 0.05 considered significant.

Item
Essential amino acid (%)
Arg
His
Ile
Leu
Lys
Met
Phe
Thr
Val
Nonessential amino acid (%)
Ala
Asp
Cys
Glu
Gly
Pro
Ser
Tyr
Total amino acid

CSMD1

CMD1

1.14
0.54
0.69
1.95
1.10
0.37
0.94
0.78
0.85

1.08
1.59
0.36
3.54
0.76
1.16
0.95
0.71
18.51

1.06
0.52
0.64
1.99
1.14
0.36
0.93
0.80
0.83

1.10
1.47
0.34
3.44
0.73
1.16
0.94
0.73
18.18

1CSMD = corn-soybean meal-miscellaneous meal diet; CMD = cornmiscellaneous meal diet.

RESULTS AND DISCUSSION

The effect of dietary protein source on the digestive
enzyme activities of the small intestine are shown in
Table 3. No differences between the 2 dietary groups
were observed for the mean of duodenal, jejuna, and
ileal amylase, trypsin, chymotrypsin, and lipase, respectively. However the duodenal amylase activity was lower in the corn-miscellaneous meal diet (CMD) group
than the corn-soybean meal-miscellaneous meal diet
(CSMD) group (P < 0.05). These results showed that
the protease activity in the small intestinal fluid did
not change in response to the type of dietary protein
consumed. Similar results were obtained by Low (1982)
and Partridge et al. (1982) in studies with pigs that were
fitted with cannulas in the pancreatic ducts. However,
Snook and Meyer (1964) and Valette et al. (1992) demonstrated that the consumption of diets with different
protein sources significantly modified trypsin and chymotrypsin activity in the pancreatic juice of pigs and

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to one of 2 experimental diets that were isonitrogenous

(165 g of CP/kg) and isoenergetic (3.15 Mcal of ME/
kg) but differed only in their protein sources (Table 1).

ing an electrolyte analyzer (Medica Easylyte, Bedford,

UK). The Ca2+ and Mg2+ concentrations were assayed
by the method described by Barnett et al. (1973) and
Mann and Yoe (1957), respectively. All of the ion concentrations were expressed as millimoles per milliliter
(mmol/mL) of intestinal fluid. The DM contents of the
diets were determined by oven drying at 105C for 5 h.
The nitrogen determination was according to the combustion method, using an FP2000 nitrogen analyzer
(Leco Corp., St. Joseph, MI). The diets were also analyzed for crude fiber (method 962.09) and ether extract
(method 920.39) according to the procedures of the
Association of Official Analytical Chemists (AOAC,
1990). The acid detergent fiber and neutral detergent
fiber contents were determined according to the procedure described by Van Soest et al. (1991). Starch
content was analyzed by the enzyme digestion method
(method 979.10; AOAC, 2002). Amino acid concentrations in the experimental diets were determined by
HPLC and shown in Table 2 (L-8900 Amino Acid Analyzer, Hitachi, Tokyo, Japan). The samples were hydrolyzed for 24 h at 110C with 6 N HCl before analysis.
Sulfur-containing amino acids were analyzed after cold
performic acid oxidation for 16 h before acid hydrolysis.
All analyses were performed in duplicate.

Table 2. Analyzed amino acids composition of the experimental

diets on a DM basis

1644

Ren et al.

Table 3. Effect of dietary protein quality on digestive enzymes in intestinal fluid of chickens
Item

Duodenum
Duodenum
Jejunum
Jejunum
Ileum
Ileum

Duodenum
Jejunum
Ileum

Amylase,1
U/mL

Lipase,1
U/mL

Trypsin,1
U/mL

Chymotrypsin,1
U/mL

106.1
5.73
430.7
430.3
183.3
232.6
27.96

55.9c
430.5a
207.5b

249.6
238

<0.001
0.617
0.049
0.028
0.992
0.217

1.52
1.27
0.41
0.42
0.05
0.05
0.11

1.40a
0.41b
0.05c

0.66
0.62

<0.001
0.389
0.433
0.123
0.95
0.991

13.4
14.2
49.8
50.5
19.2
20.6
3.34

13.8b
50.2a
19.9b

27.5
28.4

<0.001
0.727
0.992
0.877
0.881
0.764

8.00
6.97
14.5
13.3
4.77
4.46
0.95

7.48b
13.7a
4.58c

9.09
8.25

<0.001
0.291
0.888
0.453
0.39
0.821

acMeans

within a column with no common superscripts differ significantly (P < 0.05).

represents 5 observations of 9 pooled samples collected for 1 h every 4 h in 12 h in d 31, 33, and 35 of the experiment.
2CSMD = corn-soybean meal-miscellaneous meal diet; CMD = corn-miscellaneous meal diet.
1Mean

the intestinal fluid of rats, respectively. It is difficult to

compare the present work to previous studies because
our study used 2 practical diets; in comparison, Snook
and Meyer (1964) examined 2 semipurified diets that
contained casein or rapeseed as protein sources, and
Valette et al. (1992) investigated 2 purified diets that
contained 150 g of casein or whole-egg protein per kilogram of diet. Because the substrate that was used was
crucial in the determination of enzyme activity (Nicholson et al., 1974; Corring, 1980; Swanson et al., 2000),
these inconsistencies may have been caused because the
physical or chemical differences in our 2 experimental
diets were too small to be practically significant (Tables
1 and 2). In the present study, duodenal amylase activity was less in the CMD diet group. The percentage
of starch, acid detergent fiber, or neutral detergent fiber in the 2 experimental diets were similar (Table 1);
therefore, the amylase activity modification may just
slightly be influenced by dietary ingredients. Previous
studies have shown that amylase activity is sensitive
to the pH value of digesta, and the optimum pH of
pancreatic amylase in chickens was 7.5 (Osman, 1982;
Ao et al., 2008). In the current study, the duodenal pH
value of the CMD group was lower than that of the
CSMD group (4.78 vs. 5.17; Table 3); correspondingly,
the amylase activity of the CMD group was much lower
than that of the CSMD group (5.73 vs. 106.06 U/mL).
These results indicated that the lower duodenal pH in
chickens in the CMD group may be another factor that
was responsible for the decrease in duodenal amylase
activity. In the current study, the effects of the dietary
protein source on lipase activities in the small intestinal
fluid were not significant.

The proper pH value of the intestinal tract is critical

for proper digestive enzyme functionality, and pH values that are outside the normal ranges can result in decreased digestion and absorption, which can eventually
reduce growth performance (Shih and Hsu, 2006). In
the present study, the lower duodenal pH value in the
CMD group indicated that it had a weaker buffer capacity than the CSMD group. This result was supported
by Berot and Briffaud (1983) and O Hare et al. (1984),
who demonstrated that the buffer capacity varied from
one protein feed to another. The pH value varied over
a large range in the duodenum, but it varied very little
in the jejunum and ileum. The mean variation coefficients of the pH value in duodenum, jejunum, and
ileum were 4.72, 1.27, and 0.80%, respectively (Table
4). Braude et al. (1976) also found that the pH value
in the duodenum was the highest after feeding, and it
decreased with increased time after feeding; however,
the pH varied over a much smaller range in the jejunum
and ileum, which indicated that the digesta were more
effectively buffered by intestinal secretions in the jejunum and ileum. In our study, a large flow-rate in the
duodenum was observed; it was approximately 3- and
4-fold the amount of the flows in the jejunum and ileum, respectively. Because the strong acid chymus flowing from the gizzard stayed such a short time in the
duodenum for buffering, the duodenal fluid was more
sensitive to changes in dietary macronutrients than the
fluid in the jejunum or ileum. In the current study, an
increasing distance from the stomach was correlated
with an increase in the pH values; the mean pH values
of the duodenum, jejunum, and ileum were 4.98, 8.12,
and 8.77, respectively. Pang and Applegate (2007) ob-

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Diet2
CSMD
CMD
CSMD
CMD
CSMD
CMD
SEM
Main effect
Intestinal section
Duodenum
Jejunum
Ileum
Diet
CSMD
CMD
Source of variation, P-value
Intestinal section
Diet
Intestinal section diet
CSMD vs. CMD
CSMD vs. CMD
CSMD vs. CMD

Intestinal
fluid

1645

PROTEIN SOURCE ON DIGESTIVE ENZYME ACTIVITIES

Table 4. Effect of dietary protein quality on the electrolyte composition in intestinal fluid of chickens
Item

pH,1 mean
(CV, %)

Na+,1
mmol/mL

K+,1
mmol/mL

Cl,1
mmol/mL

Ca2+,1
mmol/mL

Mg2+,1
mmol/mL

Duodenum
Duodenum
Jejunum
Jejunum
Ileum
Ileum

5.17 (3.84)
4.78 (5.60)
8.11 (1.15)
8.12 (1.39)
8.74 (0.90)
8.79 (0.70)
0.069

4.98c
8.12b
8.77a

7.34
7.23

<0.001
0.060
0.007
0.001
0.951
0.627

12.0
16.4
37.8
34.0
58.8
54.6
3.74

14.2c
35.9b
56.7a

36.2
35.0

<0.001
0.698
0.444
0.414
0.48
0.435

13.3
13.2
11.5
11.4
14.7
14.4
0.34

13.3b
11.5c
14.6a

13.2
13.0

<0.001
0.517
0.945
0.85
0.767
0.524

109.4
115.2
102.8
104.0
15.0
16.6
2.16

112.3a
103.4b
15.8c

75.7
78.6

<0.001
0.117
0.509
0.07
0.698
0.605

13.6
15.9
11.6
13.4
4.89
4.91
1.36

14.8a
12.5a
4.90b

10.0
11.4

<0.001
0.228
0.683
0.241
0.363
0.99

8.15
9.41
17.4
21.4
14.3
16.5
0.77

8.78c
19.4a
15.4b

13.3b
15.8a

<0.001
0.001
0.221
0.258
0.001
0.057

Duodenum
Jejunum
Ileum

acMeans

within a column with no common superscripts differ significantly (P < 0.05).

represents 5 observations of 9 pooled samples collected for 1 h every 4 h in 12 h in d 31, 33, and 35 of the experiment.
2CSMD = corn-soybean meal-miscellaneous meal diet; CMD = corn-miscellaneous meal diet.
1Mean

served a mean pH of 6.22 in the duodenum-jejunum

and a pH of 6.26 in the ileum in a slaughter trial with
3-wk-old broilers. The same method was used by Heller
(1936), who observed that the pH values of the duodenum, upper ileum, and lower ileum of 8-wk-old chickens
were 6.52, 6.47, and 8.29, respectively. Although the
age and species of the animals and the diets used in the
previously mentioned studies may have contributed to
the variation in the measurements, the digesta collection methods may primarily account for the differences
between our study and the aforementioned reports.
Intestinal electrolytes are primarily derived from the
secretions of the bile, pancreatic, and intestinal glands,
and they play an important role in digestive enzyme
activities and dietary hydrolysate transportation (Pinheiro et al., 2004). In the current study, the CMD decreased the jejunal Mg2+ concentration, but it did not
significantly influence the Na+, K+, Cl, and Ca2+
concentrations in each segment of the small intestine
(Table 4). This result was partially in agreement with
the study by Hendrix and Bayless(1970), who found
that the concentrations of major ions in the intestine
were not easily affected by dietary nutrients because the
major ions could be rapidly absorbed by enterocytes.
The Mg2+ ion can inhibit the net water absorption in
the jejunum and influence the flow rate of digesta in
the intestinal tract (Malagelada et al., 1978; Partridge
et al., 1982). As a result, the higher jejunal Mg2+ concentration in the CMD group might be correlated with
a greater digestive water requirement in this group.
The mechanism of the intestinal Mg2+ concentration
regulation is not very clear, and further investigation
is needed. The current study showed that Na+ was the
major cation in the small intestine tract, whereas Cl

was the major anion in the duodenum and jejunum.

The apparent large anion deficit in the ileum may be
partially explained by the unmeasured bicarbonate
ions. The observed pH values in the ileum, which were
as high as 8.77, strongly suggest that bicarbonate was
the major anion that increased in concentration as the
chloride concentration decreased (Fordtran and Locklear, 1966; Clemens and Maloiy, 1978). The Ca2+ and
Mg2+ ions only slightly contributed to the total cation
concentration; the same phenomenon was observed by
Partridge et al. (1982) in pig.
The length of the small intestine in adult chickens
is approximately 1.1 to 1.2 m. Because the intestinal
morphology, chyme retention time, and the influences
of the pancreatic, bile, or intestinal gland secretions
on each intestinal segment are diverse, the digestive
enzymes and electrolyte composition of digesta from
each intestinal segment may be quite different. In the
present study, amylase, trypsin, and chymotrypsin activity in the jejunal fluid were significantly higher than
in the duodenal and ileal fluid (P < 0.01), whereas the
lipase activity in the duodenal fluid was higher than in
that of the other 2 intestinal segments (P < 0.05). This
result was confirmed by the report that dietary lipids are primarily digested in the duodenum (Krogdahl,
1985), while carbohydrates and some polypeptides were
mainly hydrolyzed and absorbed in the jejunum (Sklan
and Hurwitz, 1980; Osman, 1982; Wiseman, 2006). The
electrolyte composition of the duodenal, jejunal, and
ileal fluid were significantly different, which was likely
because the electrolyte composition of the duodenal
fluid was primarily influenced by pancreatic and bile
secretions, whereas that of the jejunal and ileal fluid
were mainly subject to intestinal gland secretions.

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Diet2
CSMD
CMD
CSMD
CMD
CSMD
CMD
SEM
Main effect
Intestinal section
Duodenum
Jejunum
Ileum
Diet
CSMD
CMD
Source of variation, P-value
Intestinal section
Diet
Intestinal section diet
CSMD vs. CMD
CSMD vs. CMD
CSMD vs. CMD

Intestinal
fluid

1646

Ren et al.

In conclusion, the CMD may have influenced amylase

activity by changing the pH value of digesta fluid, but
it did not significantly affect protease activity. With
the exception of magnesium in the hindgut, the electrolytes in the small intestinal fluid were not easily affected by dietary nutrients. The digestive enzymes and
electrolyte composition differed among each segment
of the small intestine to maintain proper physiological
function.

ACKNOWLEDGMENTS

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The authors thank the National Natural Science

Foundation of China (30901037), State Innovation
Method Project (2009IM033100), and GWFG for their
financial support. We also thank S. J. Liu, Y. G. Yin,
Z. K. Liu, J. Z. Tan, B. M. Mi, and F. Yan (Institute
of Animal Sciences, Chinese Academy of Agricultural
Sciences, Beijing) for the cannulation operations and
sample collection.

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