Directory UMM :Data Elmu:jurnal:A:Animal Feed Science and Technology:Vol86.Issue3-4.Aug2000:
Animal Feed Science and Technology
86 (2000) 177±190
Phosphate rather than surfactant accounts for the
main contribution to enhanced ®bre digestibility
resulting from treatment with boiling
neutral detergent
P.M. Kennedy, J.B. Lowry*, L.L. Conlan
CSIRO Tropical Agriculture, PMB 3, Indooroopilly, Qld, Australia 4068
Received 6 December 1999; received in revised form 31 March 2000; accepted 8 June 2000
Abstract
It is known that extraction of some grasses with neutral detergent (ND) increases the in vitro
digestibility [Kennedy, P.M., Lowry, J.B., Conlan, L.L., 1999. J. Sci. Food Agric. 79, 544]. Here, we
report experiments which de®ned the contribution of ND components to digestibility increases.
Substrates were prepared from spear grass (Heteropogon contortus) by boiling at neutral pH in solutions
of 0.05 M disodium EDTA, 0.018 M sodium borate, and 0.03 M sodium phosphate, and a mixture of the
three solutions. Phosphate was the most effective single component in increasing ND ®bre (NDF)
digestibility in vitro, after 120 h of fermentation, from 472 to 522 g/kg NDF, equivalent to 68% of the
increase found with boiling ND. NDF digestibility of bagasse at 120 h increased from 162 to 186, 230
and 277 g/kg NDF after boiling for 1 h in water, neutral phosphate and ND, respectively.
Phosphate treatment of bagasse produced a 44% increase in NDF digestibility, compared to increases
of 5, 8, 14, and 16 % for rhodes grass (Chloris gayana), spear grass, angleton grass (Dicanthium
aristatum) and carpet grass (Axonopus af®nis), respectively. Increases of cumulative gas production
against incubation time indicated that most of the increased fermentation had occurred by 24 h of
fermentation for the grasses, whereas 72 h was needed for bagasse. There were also improvements in
NDF digestion with neutral phosphate treatment of spear grass at temperatures of 75, 85 or 958C, with
prolonged treatment times required at lower temperatures. These treatments reduced the content of acid
insoluble ash in NDF and increased the rate of production of gas during fermentation. Removal of
minerals from the cell wall matrix appeared to be responsible for the increase in NDF digestibility
caused by phosphate treatment. The possible commercial use of phosphate on-farm to upgrade
nutritional quality of straws is discussed. # 2000 Elsevier Science B.V. All rights reserved.
Keywords: Pretreatment; Rumen microbes; Digestibility; Neutral detergent; Phosphate
*
Corresponding author. Tel.: 61-7-32142840; fax: 61-7-32142882.
E-mail address: brian.lowry@tag.csiro.au (J.B. Lowry).
0377-8401/00/$ ± see front matter # 2000 Elsevier Science B.V. All rights reserved.
PII: S 0 3 7 7 - 8 4 0 1 ( 0 0 ) 0 0 1 6 4 - 4
178
P.M. Kennedy et al. / Animal Feed Science and Technology 86 (2000) 177±190
1. Introduction
Increasing digestibility of plant ®bre for use by ruminants is desirable both in European
countries, where excess crop residues are a disposal problem, and in tropical countries
where there is dry-season loss of pasture quality and optimal utilisation of ®brous
residues is essential for animal production. Much research has been directed towards
chemical treatment of crop residues and improvements have been achieved by treatment
with alkali or urea or oxidative methods (e.g. alkaline hydrogen peroxide) which increase
accessibility to cell wall polysaccharides by microbial enzymes. However, alkali and
peroxide treatments have attendant safety and environmental concerns and treatment
with urea solution requires several weeks for the evolved ammonia to react with the
roughage.
In contrast to the partial solubilisation of hemicellulose that occurs during alkali
treatment, boiling of feedstuffs in neutral detergent (ND) was developed to provide a
laboratory assessment of ®bre by removing cell solubles (Van Soest et al., 1991).
Kennedy et al. (1999) demonstrated that treatment of some mature grasses with boiling
ND solution for 1 h increased extent and rate of digestion by rumen microbes measured in
vitro. This enhancement of microbial digestion was substantial for spear grass
(Heteropogon contortus), with digestibility of ND ®bre (NDF) increasing by 50% after
48 h fermentation by rumen microbes. In those experiments, we did not assess whether a
milder treatment requiring fewer chemicals or lower temperatures would produce a
similar improvement in digestion.
Accordingly, the aim of the present investigation was to de®ne which component(s)
(i.e. sodium dodecyl sulphate, sodium phosphate, sodium EDTA or sodium borate) of the
ND salts mixture was responsible for the increased digestibility, and whether similar
effects could be achieved using non-ionic detergents and treatment temperatures lower
than 1008C. Initially, we suspected that the effect was due to surfactant dislodging a
lignin fraction, as indeed occurs to a much larger extent during acid detergent treatment
(Lowry et al., 1994). Alternatively, EDTA, by chelation and removal of calcium ions from
the pectic components of the cell wall matrix, or borate, because of its well known ability
to complex with hydroxyl groups in certain con®gurations (Sinner et al., 1975), might
contribute to the activity shown by the complete ND. A subsidiary aim of this study was
to determine whether less rigorous conditions that are more applicable for farm-scale
applications, would result in economically signi®cant increases in ®bre digestion.
2. Materials and methods
2.1. Substrates and experiments
The focus of experiments 1 and 2 was identi®cation of the active components in ND
responsible for increasing ND ®bre digestibility; that for experiments 3, 4 and 5 was
comparison of the effects on spear grass with other grasses and the effect of boiling time,
while that for experiments 6 and 7 was the ef®cacy of treatment temperatures below
1008C, and the importance of substrate particle size.
P.M. Kennedy et al. / Animal Feed Science and Technology 86 (2000) 177±190
179
Table 1
Content in substrates prepared from the same batch of spear grass, of neutral detergent ®bre (NDF) and organic
matter (OM), resulting from treatment by various surfactants and salts, together with the increase in NDF
digestibility after 120 h of in vitro fermentation
Substrate
NDF content
(g/kg DM)
OM
Temperature
(g/kg NDF)
1
2
3
4
5
6
7
8
9
10
11
12
13
750
928
943
938
924
908
896
912
930
929
910
922
902
947
982
977
971
977
968
968
975
982
970
979
986
962
boiling
boiling
boiling
boiling
boiling
boiling
boiling
boiling
boiling
boiling
boiling
208C
Surfactant
SDS
SDS
SDS
Triton X-100
Triton X-100
Tween 20
Tween 20
SDS
Saltsa
P, B, E
bis-Tris
P, B, E
bis-Tris
bis-Tris
P,B,E
E
B
P
P, B, E
Increase in
digestibility (g/kg)b
0
32
73
41
ÿ5
ÿ19
34
36
46
30
31
50
11
a
E: 0.05 M Na2EDTA; B: 0.018 M Na2B4O7; P: 0.015 M NaH2PO40.015 M Na2HPO4, all adjusted to
pH 7.0.
b
Increase in NDF digestibility compared to treatment 1. All differences from treatment 1 were signi®cant
(P0.05, Table 1). Among those treatments involving single components of the ND
solution, 0.03 M PO43ÿ (treatment 12) was the most effective, resulting in increased
digestibility of 50 g/kg NDF, equivalent to 68% of the increase obtained with the
boiling ND solution (difference between treatments 3 and 12, P
86 (2000) 177±190
Phosphate rather than surfactant accounts for the
main contribution to enhanced ®bre digestibility
resulting from treatment with boiling
neutral detergent
P.M. Kennedy, J.B. Lowry*, L.L. Conlan
CSIRO Tropical Agriculture, PMB 3, Indooroopilly, Qld, Australia 4068
Received 6 December 1999; received in revised form 31 March 2000; accepted 8 June 2000
Abstract
It is known that extraction of some grasses with neutral detergent (ND) increases the in vitro
digestibility [Kennedy, P.M., Lowry, J.B., Conlan, L.L., 1999. J. Sci. Food Agric. 79, 544]. Here, we
report experiments which de®ned the contribution of ND components to digestibility increases.
Substrates were prepared from spear grass (Heteropogon contortus) by boiling at neutral pH in solutions
of 0.05 M disodium EDTA, 0.018 M sodium borate, and 0.03 M sodium phosphate, and a mixture of the
three solutions. Phosphate was the most effective single component in increasing ND ®bre (NDF)
digestibility in vitro, after 120 h of fermentation, from 472 to 522 g/kg NDF, equivalent to 68% of the
increase found with boiling ND. NDF digestibility of bagasse at 120 h increased from 162 to 186, 230
and 277 g/kg NDF after boiling for 1 h in water, neutral phosphate and ND, respectively.
Phosphate treatment of bagasse produced a 44% increase in NDF digestibility, compared to increases
of 5, 8, 14, and 16 % for rhodes grass (Chloris gayana), spear grass, angleton grass (Dicanthium
aristatum) and carpet grass (Axonopus af®nis), respectively. Increases of cumulative gas production
against incubation time indicated that most of the increased fermentation had occurred by 24 h of
fermentation for the grasses, whereas 72 h was needed for bagasse. There were also improvements in
NDF digestion with neutral phosphate treatment of spear grass at temperatures of 75, 85 or 958C, with
prolonged treatment times required at lower temperatures. These treatments reduced the content of acid
insoluble ash in NDF and increased the rate of production of gas during fermentation. Removal of
minerals from the cell wall matrix appeared to be responsible for the increase in NDF digestibility
caused by phosphate treatment. The possible commercial use of phosphate on-farm to upgrade
nutritional quality of straws is discussed. # 2000 Elsevier Science B.V. All rights reserved.
Keywords: Pretreatment; Rumen microbes; Digestibility; Neutral detergent; Phosphate
*
Corresponding author. Tel.: 61-7-32142840; fax: 61-7-32142882.
E-mail address: brian.lowry@tag.csiro.au (J.B. Lowry).
0377-8401/00/$ ± see front matter # 2000 Elsevier Science B.V. All rights reserved.
PII: S 0 3 7 7 - 8 4 0 1 ( 0 0 ) 0 0 1 6 4 - 4
178
P.M. Kennedy et al. / Animal Feed Science and Technology 86 (2000) 177±190
1. Introduction
Increasing digestibility of plant ®bre for use by ruminants is desirable both in European
countries, where excess crop residues are a disposal problem, and in tropical countries
where there is dry-season loss of pasture quality and optimal utilisation of ®brous
residues is essential for animal production. Much research has been directed towards
chemical treatment of crop residues and improvements have been achieved by treatment
with alkali or urea or oxidative methods (e.g. alkaline hydrogen peroxide) which increase
accessibility to cell wall polysaccharides by microbial enzymes. However, alkali and
peroxide treatments have attendant safety and environmental concerns and treatment
with urea solution requires several weeks for the evolved ammonia to react with the
roughage.
In contrast to the partial solubilisation of hemicellulose that occurs during alkali
treatment, boiling of feedstuffs in neutral detergent (ND) was developed to provide a
laboratory assessment of ®bre by removing cell solubles (Van Soest et al., 1991).
Kennedy et al. (1999) demonstrated that treatment of some mature grasses with boiling
ND solution for 1 h increased extent and rate of digestion by rumen microbes measured in
vitro. This enhancement of microbial digestion was substantial for spear grass
(Heteropogon contortus), with digestibility of ND ®bre (NDF) increasing by 50% after
48 h fermentation by rumen microbes. In those experiments, we did not assess whether a
milder treatment requiring fewer chemicals or lower temperatures would produce a
similar improvement in digestion.
Accordingly, the aim of the present investigation was to de®ne which component(s)
(i.e. sodium dodecyl sulphate, sodium phosphate, sodium EDTA or sodium borate) of the
ND salts mixture was responsible for the increased digestibility, and whether similar
effects could be achieved using non-ionic detergents and treatment temperatures lower
than 1008C. Initially, we suspected that the effect was due to surfactant dislodging a
lignin fraction, as indeed occurs to a much larger extent during acid detergent treatment
(Lowry et al., 1994). Alternatively, EDTA, by chelation and removal of calcium ions from
the pectic components of the cell wall matrix, or borate, because of its well known ability
to complex with hydroxyl groups in certain con®gurations (Sinner et al., 1975), might
contribute to the activity shown by the complete ND. A subsidiary aim of this study was
to determine whether less rigorous conditions that are more applicable for farm-scale
applications, would result in economically signi®cant increases in ®bre digestion.
2. Materials and methods
2.1. Substrates and experiments
The focus of experiments 1 and 2 was identi®cation of the active components in ND
responsible for increasing ND ®bre digestibility; that for experiments 3, 4 and 5 was
comparison of the effects on spear grass with other grasses and the effect of boiling time,
while that for experiments 6 and 7 was the ef®cacy of treatment temperatures below
1008C, and the importance of substrate particle size.
P.M. Kennedy et al. / Animal Feed Science and Technology 86 (2000) 177±190
179
Table 1
Content in substrates prepared from the same batch of spear grass, of neutral detergent ®bre (NDF) and organic
matter (OM), resulting from treatment by various surfactants and salts, together with the increase in NDF
digestibility after 120 h of in vitro fermentation
Substrate
NDF content
(g/kg DM)
OM
Temperature
(g/kg NDF)
1
2
3
4
5
6
7
8
9
10
11
12
13
750
928
943
938
924
908
896
912
930
929
910
922
902
947
982
977
971
977
968
968
975
982
970
979
986
962
boiling
boiling
boiling
boiling
boiling
boiling
boiling
boiling
boiling
boiling
boiling
208C
Surfactant
SDS
SDS
SDS
Triton X-100
Triton X-100
Tween 20
Tween 20
SDS
Saltsa
P, B, E
bis-Tris
P, B, E
bis-Tris
bis-Tris
P,B,E
E
B
P
P, B, E
Increase in
digestibility (g/kg)b
0
32
73
41
ÿ5
ÿ19
34
36
46
30
31
50
11
a
E: 0.05 M Na2EDTA; B: 0.018 M Na2B4O7; P: 0.015 M NaH2PO40.015 M Na2HPO4, all adjusted to
pH 7.0.
b
Increase in NDF digestibility compared to treatment 1. All differences from treatment 1 were signi®cant
(P0.05, Table 1). Among those treatments involving single components of the ND
solution, 0.03 M PO43ÿ (treatment 12) was the most effective, resulting in increased
digestibility of 50 g/kg NDF, equivalent to 68% of the increase obtained with the
boiling ND solution (difference between treatments 3 and 12, P