Supplementation of Malate and Yeast in Concentrate Containing High Cassava Chip on Rumen Ecology in Dairy Steers

Abstract

Four, one-year old of dairy steers were randomly assigned according to a 2x2 Factorial arrangement in a 4x4 Latin square design to study supplementation of malate level at 500 vs 1,000 g with yeast (Saccharomyces cerevisiae) at 1,000 vs 2,000 g in concentrate containing high levels of cassava chip. The treatments were as follows: T1 = supplementation of malate at 500 g with yeast at 1,000 g; T2 = supplementation of malate at 500 g with yeast at 2,000 g; T3 = supplementation of malate at 1,000 g with yeast at 1,000 g; T4 = supplementation of malate at 1,000 g with yeast at 2,000 g in concentrate, respectively. The animals were offered the treatment concentrate at 1% BW and ruzi grass was fed ad libitum . The results have revealed that rumen fermentation and blood metabolites were similar for all treatments. The populations of protozoa and fungal zoospores were significantly different as affected by malate level and yeast. In conclusion, the combined use of concentrate containing high level of cassava chip at 70%DM with malate at 1,000 g and yeast at 2,000 g in concentrate with ruzi grass as a roughage could improved rumen ecology in dairy steers.

Full text

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Pakistan Journal of Nutrition 8 (5): 592-596, 2009
ISSN 1680-5194
© Asian Network for Scientific Information, 2009
592
Supplementation of Malate and Yeast in Concentrate Containing High Cassava
Chip on Rumen Ecology in Dairy Steers
Sittisak Khampa , Pala Chaowarat , Rungson Singhalert and Metha Wanapat
Faculty of Agricultural Technology, Faculty of Humanities and Social Sciences,
Rajabhat Mahasarakham University, P.O. Box 44000, Thailand,
Tropical Feed Resources Research and Development Center (TROFREC), Department of Animal Science,
Faculty of Agriculture, Khon Kaen University, P.O. Box 40002, Thailand
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Abstract: Four, one-year old of dairy steers were randomly assigned according to a 2x2 Factorial
arrangement in a 4x4 Latin square design to study supplementation of malate level at 500 vs 1,000 g with
yeast (Saccharomyces cerevisiae) at 1,000 vs 2,000 g in concentrate containing high levels of cassava chip.
The treatments were as follows: T1 = supplementation of malate at 500 g with yeast at 1,000 g; T2 =
supplementation of malate at 500 g with yeast at 2,000 g; T3 = supplementation of malate at 1,000 g with
yeast at 1,000 g; T4 = supplementation of malate at 1,000 g with yeast at 2,000 g in concentrate, respectively.
The animals were offered the treatment concentrate at 1% BW and ruzi grass was fed ad libitum. The results
have revealed that rumen fermentation and blood metabolites were similar for all treatments. The
populations of protozoa and fungal zoospores were significantly different as affected by malate level and
yeast. In conclusion, the combined use of concentrate containing high level of cassava chip at 70%DM with
malate at 1,000 g and yeast at 2,000 g in concentrate with ruzi grass as a roughage could improved rumen
ecology in dairy steers.
Key words: Malate, yeast, Saccharomyces cerevisiae, cassava chip, rumen ecology, dairy steers
INTRODUCTION
Cassava (Manihot esculenta, Crantz) production in
tropical areas has a potential use in ruminant livestock
nutrition and feeding. Cassava root contains high levels
of energy and has been used as a source of readily
fermentable energy in ruminant rations (Wanapat, 2003;
Kiyothong and Wanapat, 2004; Promkot and Wanapat,
2005). One strategy for using high degradable
carbohydrates is to use in combination with readily
available NPN sources such as urea. Urea is commonly
used as N source when highly soluble carbohydrates
are fed and maintained (Wohlt et al., 1978). However,
efficient utilization of protein and Non-protein Nitrogen
(NPN) in ruminants depends upon knowledge of the
basic principles underlying ruminal microbial N
metabolism (Fernandez et al., 1997). Moreover, ruminal
pH has great impact on rumen fermentation efficiency
(Wanapat, 2003).
Some strictly anaerobic bacteria use a reductive or
reverse citric acid cycle known as the succinate-
propionate pathway to synthesize succinate and (or)
propionate. Both malate and fumalate are key
intermediates in the succinate propionate pathway and
S. ruminantium uses this pathway (Gottschalk, 1986).
The fact dicarboxylic acids, especially malate and
fumalate, stimulate lactate utilization is consistent with
the presence of this pathway in this ruminal anaerobe
(Callaway and Martin, 1996). Previous studies by
Sanson and Stallcup
MATERIALS AND METHODS
Animals, diets and experimental design: Four, one-year
old of dairy steers weighing at 150±10 kg. Cows were
randomly assigned according to a 2x2 Factorial
arrangement in a 4x4 Latin square design to study two
levels malate at 500 vs 1,000 g with yeast
(Saccharomyces cerevisiae) at 1,000 vs 2,000 g in
concentrates supplementation on ruminal fermentation(1984) reported that
supplementation of malate in ruminant diets has been
shown to increase nitrogen retention in sheep and
steers and to improve average daily gain and feed
efficiency in bull calves. In addition, supplementing diets
with yeast (Saccharomyces cerevisiae) increases milk
production of dairy cows and weight gain of growing
cattle (Brossard et al., 2006). Production responses
attributed to yeast are usually related to stimulation of
cellulolytic and lactate-utilizing bacteria in the rumen,
increased fiber digestion and increasedflow of microbial
protein from the rumen which may be beneficial for
feedlot cattle fed high-grain diets (Guedes et al., 2007).
However, the use of malate and yeast in cassava based-
diets has not yet been investigated. Therefore, the
objective of this experiment was to investigate the
supplementation of malate
supplementation in concentrates containing high level of
cassava chip with ruzi grass as a basal roughage on
ruminal fermentation and digestibility of nutrients in dairy
steers.






level and yeast

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Pak. J. Nutr., 8 (5): 592-596, 2009
593
efficiency and digestibility of nutrients. The dietary
treatments were as follows: T1 = supplementation of
malate at 500 g with yeast at 1,000 g; T2 =
supplementation of malate at 500 g with yeast at 2,000
g; T3 = supplementation of malate at 1,000 g with yeast
at 1,000 g; T4 = supplementation of malate at 1,000 g
with yeast at 2,000 g in concentrate, respectively. The
composition of dietary treatments and ruzi grass used
are shown in Table 1, 2.
Cows were housed in individual pens and individually
fed concentrate at 1%BW. All cows were fed ad libitum
of ruzi grass with water and a mineral-salt block. Feed
intake of concentrate and roughage were measured
separately and refusals recorded. The experiment was
run in four periods, each experimental period lasted for
21 days, the first 14 days for treatment adaptation and for
feed intake measurements whist the last 7 days were for
sample collections of rumen fluid and faeces. Body
weights were measured daily during the sampling
period prior to feeding.
square design with 2x2 Factorial arrangement of
treatments using the General Linear Models (GLM)
procedures of the Statistical Analysis System Institute
(SAS, 1998). Treatment means were compared by
Duncan’s New Multiple Range Test (DMRT) (Steel and
Torrie, 1980).
Data collection and sampling procedures: Roughage
and concentrate were sampled daily during the
collection period and were composted by period prior to
analyses. Composites samples were dried at 60 C and
ground (1 mm screen using Cyclotech Mill, Tecator,
Sweden) and then analyzed for DM, ether extract, ash
and CP content (AOAC, 1985), NDF, ADF and ADL
(Goering and Van Soest, 1970).
Rumen fluid samples were collected at 0, 2 and 4 h
post-feeding. Approximately 200 ml of rumen fluid was
taken from the middle part of the rumen by a stomach
tube connected with a vacuum pump at each time at the
end of each period. Rumen fluid was immediately
measured for pH and temperature using (HANNA
instruments HI 8424 microcomputer) after withdrawal.
Rumen fluid samples were then filtered through four
layers of cheesecloth. Samples were divided into two
portions. One portion was used for NH -N analyses
where 5 ml of H SO solution (1 M) was added to 50 ml
24
of rumen fluid. The mixture was centrifuged at 16,000 g
for 15 min and the supernatant stored at -20 C prior to
NH -N analysis using the micro Kjeldahl methods
3
(AOAC, 1985). Another portion was fixed with 10%
formalin solution in normal saline (Galyean, 1989).
The total count of bacteria, protozoa and fungal
zoospores were made using the methods of Galyean
(1989) based on the use of a haematocytometer
(Boeco). A blood sample (about 10 ml) was drawn from
the jugular vein at the same time as rumen fluid
sampling, separated by centrifugation at 5,000 g for 10
minutes and stored at -20 C until analysis of Blood Urea
Nitrogen (BUN) according to the method of Crocker
(1967).
o
3
o
o
Statistical analysis: All data obtained from the
experiment were subjected to ANOVA for a 4x4 Latin
RESULTS AND DISCUSSION
Chemical composition of feeds and feed intake: The
chemical compositions of roughage and concentrate
diets fed in dairy cows are presented in Table 2.
Concentrate diets contained similar concentrations of
DM, OM, CP, NDF, ADF and TDN. Diets containing high
levels of cassava chip based diets had a slightly higher
Non-structural Carbohydrate (NSC) and lower NDF due
to increased level of cassava chip in the diets.
Furthermore, the chemical composition of ruzi grass is
presented in Table 2.
The effects of malate level with yeast (Saccharomyces
cerevisiae) on feed-intake of dairy steers are presented
in Table 2. Feed intake were non-significantly different
Table 1: Ingredients of concentrate used in the experiment (%
DM basis)
Dietary treatments1
-----------------------------------------------------------
Conc. I Conc. II
Ingredient (%DM)
Cassava chip70
Palm meal3
Soybean meal 10
Molasses5
Coconut oil4
Urea3.5
Sulfur1
Salt1
Limestone1
Mineral mix 1.5
Malate (g) 500 500
Yeast (g)1,000 2,000
Conc. = concentrate;
Items Conc. IIIConc. IV
70
3.5
10
5
4
3
1
1
1
1.5
70
3
10
5
4
3.5
1
1
1
1.5
70
3.5
10
5
4
3
1
1
1
1.5
1,000
1,000
1,000
2,000
1
Table 2:Chemical composition of concentrates and ruzi grass
used in the experiment
Dietary treatments1
------------------------------------------------------------
Chemical
compositions
(%)
DM
OM
CP
NDF
ADF
TDN
ME, Mcal/kg (DM)
Feed cost (US$/kg)
DM = dry matter, CP = crude protein, OM = organic matter, NDF
= neutral detergent fiber, ADF = acid detergent fiber, TDN = total
digestible of nutrients, ME = metabolizable energy. Conc. =
concentrate
Conc.
I
88.7
91.1
16.2
13.7
8.8
79.5
2.9
0.25
Conc.
II
89.4
91.2
16.1
12.9
7.9
79.7
2.9
0.28
Conc.
III
88.7
91.1
16.2
13.7
8.8
79.5
2.9
0.28
Conc.
IV
89.4
91.2
16.1
12.9
7.9
79.7
2.9
0.30
Ruzi
gras
42.3
87.6
8.1
35.6
26.8
55.1
1.9
0.06

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Pak. J. Nutr., 8 (5): 592-596, 2009
594
Table 3: Effects of malate level and yeast (Saccharomyces cerevisiae) on feed-intake and rumen ecology in dairy steers
Treatments
-------------------------------------------------------------
ItemsT1T2
DM feed intake (%BW)
Roughage 1.61.7
Concentrate 1.01.0
Total 2.62.7
ADG (g/d)279.7 288.1 297.5
Rumen fermention
Temperature ( C) 40.2 39.8
Ruminal pH6.6 6.8
NH -N (mg/dl)17.1 18.3
3
Values on the same row with different superscripts differ (p<0.05). T1 = malate at 500 g with yeast at 1,000 g. T2 = malate at 500
g with yeast at 2,000 g. T3 = malate at 1,000 g with yeast at 1,000 g. T4 = malate at 1,000 g with yeast at 2,000 g. Probability of
main effects of level malate (M) in concentrates (500 vs 1,000 g), levels of yeast (Y) (1,000 vs 2,000 g), or the MxY interaction.
* = p<0.05, NS = p>0.05.
Contrast
------------------------------------------------------------------
SEMM
12
T3T4Y MxY
1.7
1.0
2.7
1.8
1.0
2.8
0.18
-
0.31
2.47
NS
NS
NS
*
NS
NS
NS
*
NS
NS
NS
NS 303.7
abcc
39.1
6.7
17.9
40.1
6.9
19.7
2.35
0.07
2.14
NS
NS
NS
NS
NS
NS
NS
NS
NS
o
a,b,c
1
2
Table 4: Effects of malate level and yeast on ruminal microorganisms in dairy steers
Treatments
Total direct--------------------------------------------------------------
counts (cell/ml)T1
Bacteria (x10 ) 6.1
Protozoa
Holotric (x10 ) 3.1
Entodiniomorph (x10 ) 10.3
Fungal zoospores (x10 )2.4
Values on the same row with different superscripts differ (p<0.05). T1 = malate at 500 g with yeast at 1,000 g. T2 = malate at 500
g with yeast at 2,000 g. T3 = malate at 1,000 g with yeast at 1,000 g. T4 = malate at 1,000 g with yeast at 2,000 g. Probability of
main effects of level malate (M) in concentrates (500 vs 1,000 g), levels of yeast (Y) (1,000 vs 2,000 g), or the MxY interaction.
* = p<0.05, NS = p>0.05.
Contrast
----------------------------------------------------------------
SEMM
1.22*
12
T2
7.2
T3
8.9
T4
10.9
Y
NS
MxY
NS
11
a ababb
3.0
7.8
3.6
2.5
4.1
5.5
2.1
3.4
7.0
0.26
0.71
0.51
*
*
*
NS
*
*
NS
NS
NS
4aa abb
5abcc
4

aabb
a,b,c

1
2
among treatments and was higher in dairy steers
receiving T4 than T3, T2, T1 (2.7, 2.6, 2.6 and 2.5% BW,
respectively). This data indicated that malate level with
yeast supplementation had no effect on feed-intake in
dairy steers. This result was in agreement with earlier
work by (Sommart et al., 2000 and Khampa et al., 2006)
which reported that inclusion of cassava chip in diets
resulted in satisfactory animal performance and had no
negative effects on animal health in finishing beef cattle
and lactating dairy cows.
2004) for increasing microbial protein synthesis, feed
digestibility and voluntary feed intake in ruminant fed on
low-quality roughages.
Characteristics of ruminal fermentation and blood
metabolism: Rumen ecology
measured for temperature, pH and NH -N (Table 3).
Rumen pH at 0, 2 and 4 h post-feeding were unchanged
by dietary treatments and the values were quite stable at
6.6-6.9, but all treatment means were within the normal
range which has been reported as optimal for microbial
digestion of fiber and also digestion of protein (6.0-7.0)
(Hoover, 1986).
Ruminal NH -N concentrations was not altered by
3
malate level and yeast supplement in diets containing
high cassava-based diets. As NH -N is regarded as the
most important nitrogen source for microbial protein
synthesis in the rumen. In addition, the result obtained
was closer to optimal ruminal NH -N between at 15-30
mg% (Wanapat and Pimpa, 1999; Chanjula et al., 2003,
parameters were
3
3
3
Rumen microorganisms populations: Table 4 presents
rumen microorganism populations. The populations of
fungal zoospores, protozoa and total bacteria direct
counts were significantly different and populations of
bacteria had higher numbers in heifer receiving diets T4
than T3, T2 and T1. In contrast, the present number of
protozoa in the rumen was decreased by malate level
and yeast supplementation in high cassava-based
diets. In the experiment by Newbold et al. (1996) has
shown that feeding 100 mg of malate per day in sheep
caused an increase in the number of total bacteria and
tended to increase the population of cellulolytic bacteria.
In agreement with these observations, Lopez et al.
(1999) reported that fumalate (another intermediate in
the succinate to propionate pathway) increased the
number of cellulolytic bacteria almost three-fold during
fermentation in the RUSITEC system. In addition
Guedes et al. (2007) reported that yeast are usually
related to stimulation of cellulolytic and lactate-utilizing
bacteria in the rumen, increased fiber digestion and
increasedflow of microbial protein from the rumen which
may be beneficial for feedlot cattle fed high-grain diets.
As cassava chip can be readily degraded in the rumen
and ruminal pH was decreased, malate could stimulate

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Pak. J. Nutr., 8 (5): 592-596, 2009
595
lactate utilization by S. ruminantium and could improve
pH in the rumen. It is possible that supplementation of
malate with yeast may play an important role in
increasing bacterial populations. Moreover, Martin et al.
(1999) reported that increasing dietary concentrations of
malate might help to reduce problems associated with
ruminal acidosis by stimulating lactate utilization by S.
ruminantium.
Crocker, C.L., 1967. Rapid determination of urea
nitrogen in serum
deproteinzation. Am. J. Med. Technol., 33: 361-365.
Fernandez, J.M., T. Sahulu, C. Lu, D. Ivey and M.J.
Potchoiba, 1997. Production and metabolic aspects
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Galyean, M., 1989. Laboratory Procedure in Animal
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Goering, H.K. and P.J. Van Soest, 1970. Forage Fiber
Analysis (apparatus, reagent, procedures and
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USDA, Washington, D.C.
Gottschalk, G., 1986. Bateria metabolism (2nd Ed.).
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Guedes, C.M., D.M. Goncalves, A.M. Rodrigues and A.
Dias-da-Silva, 2007. Effects of a Saccharomyces
cerevisiae yeast on ruminal fermentation and fibre
degradation of maize silages in cows. Anim. Feed
Sci. Technol., 145: 27-40.
Hoover, W.H., 1986. Chemical factors involved in
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2766.
Khampa, S., M. Wanapat, C. Wachirapakorn, N. Nontaso
and M. Wattiaux, 2006. Effect of levels of sodium dl-
malate supplementation on ruminal fermentation
efficiency in concentrates containing high levels of
cassava chip in dairy steers. Aisan-Aust. J. Anim.
Sci., 19: 368-375.
Kiyothong, K. and M. Wanapat, 2004. Growth hay yield
and chemical composition of cassava and Stylo 184
grown under intercropping. Aisan-Aust. J. Anim. Sci.,
17: 799-807.
Lopez, S., C. Newbold and R.J. Wallace, 1999. Influence
of sodium fumarate addition on rumen fermentation
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Williams, 1999. Effect of DL-malate on ruminal
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Promkot, C. and M. Wanapat, 2005. Effect of level of
crude protein and use of cottonseed meal in diets
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dairy cows. Asian-Aust. J. Anim. Sci., 18: 502-511.
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SAS, 1998. SAS/STAT User’s Guide. Version 6.12. SAS
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Conclusions: Based on this experiment, it could be
concluded that supplementation of malate level with
yeast (Saccharomyces cerevisiae) in concentrate
containing high level of cassava chip maintained could
improved ruminal fermentation efficiency. Moreover, high
level of cassava chip in diet resulted increase
populations of bacteria, but decreased protozoal
populations. These results suggest that the combined
use of concentrates containing high level of cassava
chip at 70% DM with malate at 1,000 g and yeast at
2,000 g in concentrate could highest improved rumen
ecology in dairy steers.
ACKNOWLEDGEMENTS
The authors would like to express their most sincere
gratitude and appreciation
Mahasarakham University, Tropical Feed Resources
Research and Development Center (TROFREC) and
The National Research Council of Thailand for their
financial support of research and the use of research
facilities.
to the Rajabhat
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