Download chemical composition and protein quality evaluation of high lysine

CHEMICAL COMPOSITION AN D PROTEIN Q U A L I T Y E V A L U A T I O N
OF HIGH LYSINE BARLEYS FED TO GROWING RATS A N D PIGS 1
R. Misir 2 , W. C. Sauer 3 and R. Cichon 4
University of Alberta,
Edmonton; Alberta, Canada T6G 2P5
Summary
Two established high lysine barleys, Hiproly
and Rise 1508 and one experimental high
lysine barley test line (Line 6), all grown in
Alberta under similar soil and environmental
conditions were evaluated as sole protein
sources in one rat and one pig experiment. As
compared with Galt (a normal control barley),
crude protein (N x 6.25) contents were 67, 47
and 16% higher in Hiproly, Line 6 and Rise
1508, respectively. The lysine content (percentage) in the grain and the percentage increase relative to Galt were Hiproly (.72, 80%),
Rise 1508 (.71, 78%) and Line 6 (.58, 45%),
respectively. In the rat trial, relative protein
values (RPV) expressed as percentages of that
of Gah, were 115, 109 and 108 for Line 6,
Hiproly and Riso 1508, respectively. In the pig
trial, mean N retention (as a percentage of N
intake) was similar for Hiproly, Ris~ 1508 and
Line 6 (42.6 to 43.5), but higher (P<.01) than
for Galt (32.5). Apparent biological value
(percentage) was lowest (P<.01) for Gait (50.4)
and highest for Rise 1508 (66.0), but not
significantly different (P>.05) from that of
Hiproly (57.8) or Line 6 (58.7). The high
biological value of Rise 1508, despite the low
true digestibility of N was directly related to
the amount (g) of lysine available for absorp-
tion by the pig. It is concluded that Albertagrown high lysine barleys are nutritionally
superior to a normal control barley. The use of
rats as assay animals may provide valuable data
on protein quality of test barley lines for
growing pigs.
(Key Words: High Lysine Barleys, Relative
Protein Value, Nitrogen Balance, Amino Acid
Digestibilities, Rats, Pigs.)
I ntroduction
The search for cereal grains high in lysine led
to the discovery of high lysine barleys (Munck
et al., 1969), corn (Mertz et al., 1964; Nelson et
al., 1965) and sorghum (Singh and Axtell,
1973). Because the nutritional value of cereal
proteins is limited by the level of lysine (Howe
et al., 1965; Eggum, 1977), the use of high
lysine cereals is expected to reduce protein
supplementation in diets for nonruminant
animals, thus resulting in lower total feed costs.
Protein quality of high lysine barleys was
shown to be superior to that of normal barley
when fed to rats (Munck et al., 1970; Doll et
al., 1974; Balaravi et al., 1976; Stobart, 1977)
and pigs (Mortensen et al., 1975; Thomke and
Widstromer, 1975). The use of high lysine
barley in pig diets resulted in substantial savings
of soybean meal (Thomke and Widstromer,
1975; Newman et al., 1978).
Preliminary information on the protein
quality of experimental barley lines developed
1The authors are grateful for financial support by the plant breeder is usually obtained by
from the Agricultural Research Council of Alberta chemical analysis (Ingversen and Keie, 1973;
(Farming for the Future) and the Alberta Agricultur- Rhodes and Gill, 1980), or both chemical
al Research Trust. Samples of Barley Geneticist, Al- analysis and biological evaluation using rats
berta Agriculture, Lacombe, Canada.
(Doll et al., 1974; Newman et al., 1974; Bansal
2Present Address: Prairie Swine Centre, Dept. of
Anim. and Poul. Sci., Univ. of Saskatchewan, Saskat- et al., 1977; Misir and Sauer, 1982). Such
information is often useful in identifying
chewan, Canada S7N 0WO.
3Dept. of Anita. Sci.;to whom requests for reprints nutritionally superior lines for potential use in
should be addressed.
animal diets. Therefore, the objective of the
4Postdoctoral Fellow, Institute of Food Biotechpresent study was to evaluate, by chemical
nology, Univ. of Agriculture, Olsztyn, Poland.
analyses and in experiments using both growing
Received December 22, 1982.
Accepted April 2, 1984.
rats and pigs, the protein quality of high lysine
1011
JOURNAL OF ANIMAL SCIENCE, Vnl. 59, No. 4, 1984
1012
MISIR ET AL.
barley test lines grown in the province of
Alberta, Canada.
Experimental Procedure
All the barleys were grown in Alberta under
similar environmental and soil conditions. Line
6 was developed at the Alberta Agricultural
Research Station, Lacombe, Canada. The
barleys used in both experiments were harvested from the same field during the 1978
crop year. Protein content (N X 6.25) and
partial amino acid (AA) analyses of the barleys
are presented in table 1.
Exp. 1. Three-week old Sprague-Dawley
weanling rats were assigned on the basis of sex
into weight groups from which they were
allocated randomly to five treatment groups,
each consisting of 12 males and 12 females. The
mean initial weight + SE for rats of both sexes
was 52 -+ 1 g. The rats were individually housed
in metabolic cages (25 x 18 cm) with 1-cm
mesh floors. The room was equipped with an
s Sheffield Chemical Co., Norwich, NY.
Christy and Norris, Chelmsford, England.
automatic 12-h light-dark cycle (0700 to 1900
h) and maintained at a temperature of 24 + 1 C.
The rats were fed ad libitum cornstarch-based
diets (table 2), formulated to contain 2, 5 and
8% crude protein from each of the barleys: Gait
(a normal commercial barley serving as the
control), Hiproly (a high protein, high lysine
barley; Munck et al., 1969), Ris~ 1508 (a high
lysine mutant barley; Ingversen et al., 1973)
and Line 6 (an experimental high lysine barley
test line). The ANRC casein s was used as the
reference protein. Before being mixed with the
other dietary ingredients, all grain samples were
finely ground in a laboratory mill6 equipped
with a .75-ram screen. For each rat, the diet
was weighed into a separate plastic cup fitted
with a tight lid. Daily dietary allowances were
taken from these cups and placed into food
jars, each equipped with a circular screen (1-cm
pore size) and a metal cover (with a circular opening, 2.5 cm diameter) to minimize
spillage. Water was provided ad libitum. The
rats were put on a 14-d experimental period
that consisted of a 9-d adaptation period
followed by a 5-d period for total collection of
feces. Immediately thereafter, the rats were fed
a protein-free diet (table 2) for a similar exper-
TABLE 1. PROTEIN CONTENT AND PARTIAL AMINO ACID ANALYSES OF THE PROTEIN
OF THE BARLEY SAMPLES
Barleys
Gait
Hiproly
Ris# 1508
Line 6
Protein, % dry matter
Amino acids, g/16 g N
10.75
18.00
12.50
15.81
Indispensable
Arginine
Histidine
Isoleucine
Leucine
Lysine
Methionine
Phenylaianine
Threonine
Valine
Dispensable
Alanine
Aspartic acid
Cysteine
Glutamic acid
Glycine
Proline
Serine
Tyrosine
4.91 (.53) a
2.17 (.23)
3.48 (.37)
6.98 (.75)
3.74 (.40)
1.22 (.13)
4.74 (.51)
3.29 (.35)
4.22 (.45)
3.87 (.42)
6.25 (.67)
1.96 (.22)
22.13 (2.38)
3.81 (.41)
10.04 (1.08)
4.15 (.45)
2.23 (.24)
4.59 (.83)
2.13 (.38)
3.65 (.66)
7.01 (1.26)
4.01 (.72)
1.37 (.25)
5.72 (1.03)
3.34 (.60)
4.19 (.75)
6.73 (.84)
2.94 (.37)
3.67 (.46)
7.32 (.91)
5.71 (.71)
1.51 (.19)
4.22 (.53)
4.32 (.54)
4.93 (.62)
4.67 (.74)
2.04 (.32)
3.53 (.56)
6.82 (1.08)
3.67 (.58)
1.21 (.19)
5.01 (.79)
3.27 (.52)
4.11 (.65)
4.13 (.74)
6.50 (1.17)
1.09 (.20)
23.36 (4.20)
3.78 (.68)
11.05 (1.99)
4.17 (.75)
2.19 (.39)
5.35 (.67)
9.54 (1.19)
1.96 (.24)
17.48 (2.18)
5.57 (.69)
7.04 (.88)
4.76 (.59)
2.42 (.30)
3.86 (.61)
6.14 (.97)
1.51 (.24)
23.29 (3.68)
3.68 (.58)
10.60 (1.67)
4.14 (.65)
2.07 (.33)
avalues in parentheses refer to amino acid content expressed as a percentage of the grain (dry matter basis).
HIGH LYSINE
BARLEYS
imental period. Feed and water were removed
from the cages 3 h before the rats were reweighed at the end of both adaptation and
collection periods. During the test periods, feed
not consumed plus feed spillage were measured
for each rat. The feces from each ragewere
collected daily and stored at - 2 0 C until
required for sample preparation and analysis.
Exp. 2. Eight Yorkshire x Lacombe barrows,
ranging in initial live weight from 43 to 47 kg,
were individually confined to metabolic cages
that permitted separate collection of feces and
urine. The barn was provided with continuous
lighting and maintained at a temperature and
relative humidity of 20 to 22 C and 70 to 75%,
respectively. Before being put on this experiment, the pigs were fed a 16% crude protein
diet based on barley (50%), wheat (31.5%),
soybean meal (15.0%) and adequately fortified
with vitamins and minerals (NRC, 1979).
The four experimental diets (table 3) were
formulated to be isonitrogenous and isofibrous
b y the addition of purified cellulose 7 and
cornstarch. Minerals and vitamins were added
to meet or exceed NRC (1979) specifications.
The barleys were ground in a Wiley mill s with a
3-mm mesh screen before being mixed with the
other ingredients.
Each of the four experimental periods lasted
10 d, during which each pig was given a 900-g
meal twice daily at 0700 and 1900 h. Two pigs
were fed each diet in each of the four periods
making a total of eight observations per diet.
Between the experimental periods, the pigs
were fed ad libitum the previously described
16% crude protein grower diet for 4 d. Water
was supplied ad libitum. Total daily output of
feces and urine were collected during the last 5
d of each period.
Chemical Analyses. Before analyses, the
samples were ground in a Wiley mill 7 with a .75
mm mesh screen. Grain, casein, diets and
composite fecal samples (Exp. 1 and 2) were
analyzed for dry matter and Kjeldahl N (AOAC,
1970). Amino acid analyses were conducted on
grain samples, pig diets and feces (pooled
among days for each period), as described by
Sarwar and Bowland (1975).
Calculations and Statistical Analysis. Methods
VAlphafloc,
supplied
by Lee Chemicals,
Yonge Street, Toronto, Ontario, Canada.
s Arthur M. Thomas, Philadelphia, PA.
1119
FOR RATS AND PIGS
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MISIR ET AL.
TABLE 3. PERCENTAGE INGREDIENT COMPOSITION OF THE PIG DIETS (EXP. 2)
Diet
Ingredient
Gait
Hiproly
Rise
Line 6
Barley (IFN 4-00-549)
Cornstarch (IFN 4-02-889)
Alpha floc
Soybean oil (IFN 4-07-983)
Calcium carbonate (IFN 6-02-632)
Calcium phosphate (IFN 6-01-081)
Mineral m i x a
Vitamin mixb
Chemical analyses
Crude protein
Dry matter
94.8
3.0
1.2
.5
57.4
33.9
3.0
3.0
.9
1.3
82.5
9.7
2.5
3.0
1.1
.7
64.7
27.4
2.4
3.0
.9
1.1
.3
.3
.3
.3
.2
.2
.2
.2
9.22
89.44
9.26
89.63
9.24
89.77
9.15
89.60
acontributed the following nutrients per kilogram of diet: Na 1.9 g; CI, 2.9 g; Co, .16 rag; Cu, 10 mg; I, .23
mg; Fe, 65 mg; Mn, 50 mg; Se, .10 mg.
bcontributed the following vitamins per kilogram of diet: vitamin A, 4,500 IU; vitamin D, 1,400 IU; alphatocopherol, 35 IU; menadione, 75 ug; choline 1.25 g; folic acid, 25 mg; niacin, 30 mg; pantothenic acid, 10 mg;
riboflavin, 5 rag; thiamin, 5 mg; vitamin B6,8 mg; vitamin B~, 50 ug.
used for calculating apparent digestibilities
(AD) were those described by Kuiken and
Lyman (1948). In Exp. 1, metabolic fecal N
values (table 4, footnote 6) for computing true
digestibilities (TD) of N were obtained when
the rats were fed the protein-free diet (table 2).
In Exp. 2, metabolic fecal values for calculating
TD of N and A A by pigs were those previously
determined by Anderson et al. (1981). In Exp.
1, regression slope and correlation coefficients
(body weight changes on N consumed) were
calculated for casein and the barleys (Snedecor
and Cochran, 1967). Relative protein values
(RPV) were determined using the regression
slope coefficients of the test protein, expressed
as a fraction of that of casein (FAO-WHO,
1975). In Exp. 2, apparent biological value
was taken as the percentage of the apparently
absorbed N that was retained.
The data for the four barleys were subjected
to one-way analysis of variance (Exp. 1), or
analyzed using a double 4 x 4 Latin square
arrangement of barleys x barrows (Exp. 2). In
both experiments, significance among treatment means were established by the NewmanKeuls' multiple range test (Snedecor and
Cochran, 1967).
Results and Discussion
Lysine content was highest in Rise 1508
followed by Hiproly, Gait and Line 6 (table 1).
For all the barleys, the level of threonine, the
second limiting A A in barley (Aw-Yong and
Beames, 1975; Eggum, 1977) was lower than
corresponding lysine levels, but followed the
same trend. In addition, lowest levels o f glutamic acid and proline were observed for Ris~
1508. Relative to Galt, increases in the grain
levels of lysine and threonine were 80 and 71%
for Hiproly, 78 and 54% for Rise 1508 and 45
and 49% for Line 6, respectively. A previous
study in Denmark, which compared Rise 1508
to its parent Bomi (Ingversen et al., 1973),
reported similar increases for lysine and threonine, in addition to decreases in glutamic acid
and proline. The present results show that Rise
1508 also expressed its genetic potential for
improved A A content in a Western Canada
environment.
The value for the regression slope coefficient
was highest (P<. 10) for casein (table 4). Among
the barleys, the value for Line 6 was higher
(P<.10) than that for Gait, but was not different (P>.10) from those for Hiproly or Rise.
Differences among regression slope coefficients
were reflected in the RPV, which indicate their
relative capacities to support tissue growth
(Sammonds and Hegsted, 1976). The RPV for
the barleys, expressed as a percentage of that of
Gait, were 109.3 for Hiproly, 108.2 for Ris~
1508 and 115.8 for Line 6, i.e., an average
HIGH LYSINE BARLEYS FOR RATS A N D PIGS
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MISIR ET AL.
t h e t h r e e d i f f e r e n t levels f e d t o t h e r a t s . T h e r e
w a s n o s i g n i f i c a n t d i f f e r e n c e in p r o t e i n d i g e s t i bilities (AD or TD) among the barleys for rats.
These values were lower (P<.01) than the AD
or TD values for casein (table 4).
i m p r o v e m e n t o f 1 1 % in t h e p r o t e i n q u a l i t y o f
these high lysine barleys compared with Gait
for growing rats. The high correlation coeffic i e n t s s u g g e s t e d t h a t t h e p r o t e i n in t h e b a r l e y s
tested was utilized with the same efficiency at
T A B L E 5. N I T R O G E N BALANCE A N D A P P A R E N T BIOLOGICAL V A L U E OF HIGH LYSINE
B A R L E Y DIETS (EXP. 2)
Diet
Item
Gait
Hiproly
Ris~ 1508
Line 6
Intake N, g/5 d
Fecal N, % intake N
Urinary N, % intake N
Retained N, % intake N
Apparent biological value, %
127.9
35.4 b
32.0 b
32.5 x
50.4 x
130.2
25.3 c
31.5 b
43.2Y
57.8 x y
129.6
32.2 b
23.1 c
44.7Y
66.0Y
128.4
27.8 c
29.6 b
42.6Y
58.7 x y
SE a
.2
1.3
1.7
2.1
2.4
a s t a n d a r d error o f the mean.
b'CMeans within a row with different superscripts differ (P<.01).
x'YMeans within a row with different superscripts differ (P<.05).
T A B L E 6. A P P A R E N T DIGESTIBILITIES (PERCENTAGE) OF C R U D E PROTEIN
A N D AMINO ACIDS IN THE BARLEY DIETS BY PIGS (EXP. 2)
Barley
Item
Gait
Hiproly
Riso 1508
Line 6
Dry m a t t e r
80.0 b
87.7 c
78.1 b
85.6 c
Crude protein
64.6 d
74.7 c
67.8 bd
72.2 be
1.3
82.0xY
83.0Y
73.1 c
77.6 c
67.1 c
61.7Y
80.6 c
70.1Y
84.0Y
84.6Y
64.8 b
72.2 bc
67.2 c
57.5 x y
69.2 b
68.2 sy
82.6 x y
82.9Y
70.7 c
75.4 be
62.2 e
53.1 x
77.6 c
68.6 x y
.7
.6
1.3
1.0
1.8
2.0
1.1
1.2
66.7 c
69.3 c
73.1 x
86.1 c
71.8 c
86.7 c
67.0Y
63.5 c
70.4 c
78.0 x y
80.6 b
71.9 c
79.8 d
64.6Y
62.4 e
65.9 c
84.2Y
85.5 c
69.0 c
86.8 c
66.7Y
1.8
1.4
1.3
.7
1.3
.7
1.2
SE a
.9
A m i n o acids
Indispensable
Arginine
Histidine
Isoleucine
Leucine
Lysine
Methionine
Phenylalanine
Threonine
Dispensable
AIanine
Aspartic acid
Cysteine
Glutamic acid
Glycine
Proline
Tyrosine
78.3 x
81.0Y
61.6 b
69.4 b
50.6 b
, 53.6 x
71.1 b
62.0 x
48.1 b
58.7 b
84.6Y
81,9 b
60,2 b
83.3 b
60.7xY
a s t a n d a r d error o f the mean.
b'C'dMeans within a row with different superscripts differ (P<.01).
x'YMeans within a row with different superscripts differ (P<.05).
1017
HIGH LYSINE BARLEYS FOR RATS AND PIGS
Fecal N excretion (as a percentage of intake
N) was similar for pigs fed Hiproly or Line 6
but lower (P<.05) for those fed Galt or Ris#
1508 (table 5). Urinary N loss was lowest
(P<.05) for pigs fed Rise 1508 as compared
with Gait, Hiproly or Line 6. Retained N, a
reflection of the balance of indispensable A A
available for protein synthesis, was lowest
(P<.05) in pigs fed Gait; however, the value for
Line 6 was not different (P>.05) from corresponding values for Hiproly and Ris~ 1508,
indicating that Line 6 might possess nutritional
characteristics similar to the two established
high lysine barleys.
For pigs, both AD and TD of protein and
dispensable AA were similar or higher than
corresponding values for Gait (tables 6 and 7).
The AD of crude protein was highest (P<.01) in
Hiproly, lowest in Gait and intermediate in
Line 6 and Rise 1508. Among the indispensable
AA, the AD of lysine and threonine, were
higher (P<.01) in Hiproly, Ris# 1508 and
Line 6 than in Galt. The AD of methionine was
not different (P>.05) in Hiproly or Riso 1508
but was higher (P<.05) than in Line 6 or Gait.
The TD of protein, indispensable and dispensable AA followed the same trend, but were
higher than corresponding AD values.
The A A availability estimates determined by
aid of the fecal analysis method (Kuiken and
Lyman, 1948) should be interpreted with
caution. Experiments with pigs showed that the
ileal analysis method provides more accurate
estimates (Zebrowska, 1973; Sauer, 1976).
However, several studies (Sauer et al., 1977;
Just et al., 1980; Sauer et al., 1981; Taverner and Farrell, 1981) have shown that as far as
lysine and methionine in barley are concerned,
it may be justifiable to use the fecal analysis
method. In most instances, differences between
ileal and fecal availabilities for these amino
acids were nonsignificant and o f a small magnitude. Of the other more important AA, the
availability of threonine should be interpreted with care, because as was shown in the
studies referred to for this AA, ileal availabilities
TABLE 7. TRUE DIGESTIBILITIES (PERCENTAGE) OF CRUDE PROTEIN AND AMINO ACIDS
IN THE BARLEY DIETS BY PIGS (EXP. 2)
Barley
Item
Gait
Hiproly
Riso 1508
Line 6
SEa
Crude protein
72.5 b
82.5 c
75.6 bd
80.1 cd
1.3
Indispensable
Arginine
Histidine
Isoleucine
Leucine
Lysine
Methionine
Phenylalanine
Threonine
Valine
84.9 x
87.7 x
74.0 d
78.0 c
62.9 d
63.0 x
78.6 d
75.5 x
78.4Y
89.0xy
89.8xy
84.9 c
86.1 e
78.4 c
78.7Y
86.7 c
83.2Y
85.8Y
89.1xy
89.9xY
77.1 cd
80.9 cd
75.9
73.8 y
78.1 d
79.0xy
81.1Y
90.4Y
91.0Y
84.4 c
85.3 c
76.3 c
75.0Y
85.5 c
83.7Y
81.5Y
.7
.6
1.4
1.0
1.7
1.9
1.1
1.3
1.1
Dispensable
Alanine
Aspartic acid
Cysteine
Glutamic acid
Glycine
Proline
Serine
Tyrosine
60.8 d
71.8 x
95.7 d
86.1 x
70.8 d
86.7'cd
80.3 x
71.7 x
78.5 c
81.7Y
92.8 bd
90.1Y
82.4 c
89.8 c
86.7Y
78.1Y
73.4 c
79.5Y
89.3 b
86.3 x
79.7 c
85.1d
82.7xy
75.4xy
76.6 c
80.8Y
100.3 c
90.1Y
81.3 c
90.4 c
86.7Y
80.0Y
1.8
1.4
1.0
.6
1.4
.6
1.0
1.3
Amino acid
astandard error of the mean.
b'c'dMeans within a row with different superscripts differ (P<.01).
x'YMeans within a row with different superscripts differ (P<.05).
1018
MISIR ET AL.
were a p p r o x i m a t e l y 10 percentage units lower
than the corresponding fecal availabilities.
A comparison of the A A c o m p o s i t i o n (table
1) with the A A requirements (NRC, 1979)
suggests that Hiproly or Rise 1508 w h e n used
as the sole source of dietary protein could m e e t
the total A A requirements for the 60 to 100 kg
pig; however, available AA, calculated using
data on barley A A c o n t e n t (table 1) and T D
values (table 7), indicated that all the barleys might be low in m e t h i o n i n e plus cysteine.
Only Hiproly and Rise 1508 seemed a d e q u a t e
in lysine. Also, Rise 1508 might be low in
isoleucine; and Gait low in isoleucine and
threonine.
The TD of crude protein, with the e x c e p t i o n
of Gait, c o m p a r e d reasonably well in pigs and
rats and were 83.4 vs 82.5%, 78.4 vs 75.6% and
81.1 vs 80.1% for Hiproly, Risr 1508 and Line
6, respectively (tables 4 and 7). The T D of
crude protein in G a h was considerably higher in
rats (81.3%) than in pigs (72.5%), the reasons
for which are n o t clear. Studies by Eggum
(1973) on one barley sample showed similar T D
of protein and A A in barley for pigs and rats.
The i m p r o v e m e n t in protein quality o f the
high lysine barleys (as c o m p a r e d with Gait)
ranged f r o m 8.2 to 15.8% w h e n measured by
the RPV m e t h o d with rats (table 3). When
measured by the N balance m e t h o d with pigs,
the i m p r o v e m e n t was larger and ranged f r o m
32.9 to 36.6%. The differences in i m p r o v e m e n t
in protein quality m a y be related to the different m e t h o d s (i.e., RPV vs N balance), and also
to the lower TD of protein in Gait (and presumably also for the amino acids) in pigs
(72.5%) than in rats (81.3, tables 4 and 7).
A l t h o u g h the responses in i m p r o v e m e n t of
protein quality were smaller with the RPV
m e t h o d , this m e t h o d discriminates b e t w e e n
normal and high lysine barleys and m a y be
useful for initial studies on e x p e r i m e n t a l high
lysine barley test lines as was also shown in
previous studies (Misir and Sauer, 1982).
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