3 1293 000819320

; , ~ 3 ll'c’lelllllllllllllllll

This is to certify that the

thesis entitled

INFLUENCE OF AMMONIA TREATMENT AND TIME
ENSILING ON PROTEOLYSIS AND FEEDING
VALUE OF CORN SILAGE FOR
DAIRY CATTLE

presented by

Colin O.L.E. Johnson

has been accepted towards fulfillment
of the requirements for

Masters degreein Animal Science

My [KW

/é M /7f/ /

 

 

Date

 

0-7639

 

 

 

RETURNING MATERIALS:
IV1£31_] Place in book drop to
LIBRARIES ‘ remove this checkout from

4...!,...._ your record. FINES will
be charged if book is

 

 

 

returned after the date
stamped below.

 

gr?! 1 r

$
1’" a»

U j} .l.

 

 

INFLUENCE OF AMMONIA TREATMENT AND TIME OF
ENSILING ON PROTEOLYSIS AND FEEDING
VALUE OF CORN SILAGE FOR
DAIRY CATTLE

By

Colin O.L.E. Johnson

A THESIS

Submitted to
Michigan State University
in partial fulfillment of the requirements
for the degree of

MASTER OF SCIENCE

Department of Animal Science

1981

ll 1’ '.' I ’
x... / .x z «

ABSTRACT

INFLUENCE OF AMMONIA TREATMENT AND TIME
OF ENSILING ON PROTEOLYSIS AND FEEDING VALUE
OF CORN SILAGE FOR DAIRY CATTLE
By

Colin O.L.E. Johnson

Whole-chopped corn plants (32-3496 DM) were treated with O, .25, .52, or
1.08% added N as ammonia. About 3 kg of material were placed in 5 mil polythene
bags which were then evacuated and served as experimental silos. Silage samples
were taken on days 0, l, 2, 6, 12 and 51+ and analyzed for dry matter, PH, lactic
acid, total and water insoluble nitrogen, and free amino acids in the water phase.
Added ammonia increased initial pH of silages, but delayed increases in lactic acid.
Increasing levels of ammonia decreased proteolysis. Large increases in alanine
were observed in the water phase of high ammonia-treated silages, increases which
were apparently not related to proteolytic effects (Experiment 1).

In Experiment 2, 2# lactating cows were utilized to determine the relative

feeding value of corn silages treated with Cold-flo (.3696 NH 82% N) or Pro-Sil

3’
(2.4% ammonium solution, 13.6% N). Dry matter and lactic acid were higher for
the Pro-Sil treated silage. Apart from a slightly higher intake of NPN by the Pro-

Sil group, milk yields, intakes, and body weight changes were similar for both

groups.

DEDICATION

To the memory of my beloved mother.

Edna W. Johnson. 1922-1980.

ii

ACKNOWLEDGEMENTS

I would like to express my sincere appreciation to Dr.
J. T. Huber, my major Professor, for accepting me as one of
his graduate students and providing me with invaluable
assistance and guidance throughout my graduate studies.

Also. special thanks are due to the members of my committee.
Dr. W. G. Bergen and Dr. D. Hillman. for their patience and
understanding of my cause.

My thanks also go to all those who aided the cause. par-
ticularly the Department of Dairy Science for financial
assistance during the course of my graduate studies.

Special thanks also go to the Dairy Nutrition Secretary.
Elaine Kibbey, for her generous help throughout the graduate
program.

Finally. I must thank my loving and devoted wife.
Paulette, not only for her remarkable ability to turn none-
too-legible manuscript into beautiful typescript. but also
for her encouragement and patience throughout the graduate

study period.

iii

TABLE OF CONTENTS

LIST OF TABLES . . . . . . . . . .
INTRODUCTION . . . . . . . . . .
LITERATURE REVIEW . . . . . . . . .

Brief History of Ensiling .
Silage Fermentation . . .
Carbohydrate Fermentation .
Protein Degradation . . . .
Non-Protein-Nitrogen Treatment of Corn
Silages . . . . . . . . .
Effect of Ammonia Addition on Silage
Fermentation . . . . . . . .
Feeding Trials . . . . . . . .

MATERIALS AND METHODS . . . . . . .

Experiment 1. Silage Fermentation Study
Experiment 2. Feeding Trial . . . .
RESULTS AND DISCUSSION . . . . . . .
Experiment 1. . . . . . . . . .
Experiment 2. . . . . . . . . .
SUMMARY AND CONCLUSIONS . . . . . . .
BIBLIOGRAPHY . . . . . . . . . .

iv

Page

Cantu m H

N

1

15
19
19
22
2#
2h
as
50
53

LIST OF TABLES

TABLE PAGE
1. Summary of dairy cow performance on corn silage
treated with ammonia at ensiling . . . . 17
2. Silage parameters under investigation . . . 20
3. Analysis of Variance for Split-Plot
Analysis............ 21
h. Ration composition of eXperimental diets. . . 23

5. Effect of ammonia concentration and time of
ensiling on 5 dry matter in corn silage . . 25

6. Effect of ammonia concentration and time of
ensiling on pH in corn silage . . . . . 27

7. Effect of ammonia concentration and time of
ensiling on lactic acid in corn silage . . 28

8. Effect of ammonia concentration and time of
ensiling on total nitrogen in corn
silage . . . . . . . . . . . . . 30

9. Effect of ammonia concentration and time of
ensiling on water insoluble nitrogen in
corn Silage o o o o o o o o o o o 32

10. Effect of ammonia concentration and time of
ensiling on water soluble nitrogen in
corn Silage o o e o o o o o o o o 33

11.1 Effect of ammonia concentration and time of

ensiling on free amino acids in corn

silage water phase . . . . . . . . . 36
11.1 Continued 0 o o o o o o o o o o o o 37

11.1 continu3d o o o o e o o o o o o o o 38

TABLE PAGE

11.2 Effect of ammonia concentration and time of
ensiling on free amino acids in corn
silage water phase . . . . . . . . #0

11.3 Main effect means, standard errors, and
significant differences for free amino
acids in corn silage water phase . . . #1

11.4 Effect of ammonia addition and time of ensil-
ing on free amino acid N in corn silage
water phase . . . . . . . . . . 43

11.5 Effect of ammonia addition and time of
ensiling on total free amino acid N in
corn silage water phase . . . . . . on

12. Dry matter. pH, lactic acid and crude
protein of corn silage composites . . . #6

13. Influence of non protein nitrogen source on
dry matter and protein intakes of dairy #6
cows 0 O O O O O O O O O O 0

1h. Influence of non protein nitrogen source on
milk yield: of dairy cows . . . . . 48

15. Influence of non protein nitrogen source on

milk composition, weight change and feed .
efficiency of dairy cows . . . . . #8

vi

Introduction

It is widely accepted that normal corn silage that is
well eared and preserved is a high-energy source of forage
for cattle, sheep and other ruminants. In addition, it has
the potential of substantially reducing feed costs and in-
creasing beef and milk production per acre of crops fed.
However, these advantages are partially offset by an insuf-
ficiency of protein. a factor which appears to limit volun-
tary intake. Moreover, degradation of protein (or proteoly-
sis) in the resultant silage may influence the quantity of
protein (as Opposed to NPN) which is absorbed and available
for metabolism in the animal's body.'

In addition to ammonia treatment being the preferred
method for making up the protein deficit in corn silage,
ammonia treatment of the whole chapped corn plant at ensiling
has consistently resulted in increased insoluble nitrogen
(Henderson and Bergen. 1972; Huber gt §;.. 1973) and possibly
decreased proteolysis of plant protein in the resultant
silage (Huber gt al.. 1979c). The importance of reduced
proteolysis during ensiling cannot be overemphasized, since
reduced degradation of protein to NPN may increase the supply
of undegraded plant protein to the abomasum and intestines,
and this may enhance intake (Egan and Meir, 1965).

The forms of ammonia most available for silage treatment

are Cold-flo (concentrated anhydrous, 82% nitrogen) and Pro-

Sil ( an ammonium solution containing 13.6% nitrogen). Earlier

studies showed depressed intakes in dairy and beef cattle

fed silage treated with concentrated anhydrous ammonia com-

pared to silage treated with more dilute solutions. More

studies with Cold-flo and ammonia solutions have not directly

compared the forms of treatment in lactating cows. Many

dairymen contemplating ammonia treatment question the relative

value of the different forms of ammonia, but little infor-

mation is available.

Therefore, the objective of this study were two-fold:

1. To characterize the nature of the protein breakdown in
corn silage ensiled with or without ammonia.

2. To determine the relative feeding value of corn silage

treated with two forms of ammonia (Cold-flo vs. Pro-Sil).

Literature Review
Brief History of Ensiling

The process of ensiling is old and has been intensively
studied during the last century (Watson and Nash, 1960). The
object of storing green craps as silage is to preserve the
material with a minimum loss of nutrients. Various ensiling
practices have been used to achieve this end. In 18h3,
Johnston recommended a German method for harvesting and
storing green fodder: the direct cut material was packed into
trenches which were then covered with boards and earth to
facilitate the exclusion cf air. Also. as pointed out by
Jenkins, 188“, the practice of storing grain in pits or under-
ground trenches dates back to the Greeks and Egyptians.

Goffart's work in France had an important influence on
the popularization of the practice in the late 19th century.
He recognized the rapidity with which crop maize ferments
within the silo. He also stressed the importance of reducing
the length of chop so as to facilitate better packing and
exclusion of air from the silage mass. Miles (1918) gave
Reihlen, a German, credit for being the first to preserve the
whole corn plant utilizing a silo.

Corn silage is a basic ingredient in both dairy and beef
cattle rations in many regions of North America and Europe.
Corn harvested for silage in North America has increased
from #7.? million tons in 195h (Waldo and Jorgensen. 1981) to
111.1 million tons in 1980 (U.S. D.A., 1981). Although the

3

ensiling of forage maize as a winter feed for livestock has
been practiced in Europe for at least 100 years (Wilkinson,
1978) the quantity of maize harvested for silage remained
low during the first fifty years of the this century due to
inadequate harvesting machinary. The introduction of the
forage harvester into Europe during the 1960's revolutionized
the technology of the harvesting and ensiling operation
(Wilkinson. 1978).

The current energy crisis and world food shortage with
the resulting emphasis on animal production from forage has
created the proper climate for reevaluating silage production.
Si gge Fermegtation

Once a producer has grown a good, high-yielding crop,
his next objective is to preserve the crap for livestock
feeding. As Barnett (195A) pointed out, the aim of silage
making is ”to achieve within the ensiled mass sufficient
concentration of lactic acid, produced as a result of the
presence of microorganisms within the cut crap, to inhibit
other forms of microbial activity and thus preserve the
material until such time as it is required". Research has
conclusively shown that the process of ensiling green crops
is the most effective and efficient method of storing and
preserving the nutritive value of the whole corn plant. How-
ever, it must be borne in mind, that the feeding value of
silage is largely affected by microbiological, chemical. and
physical factors. The ensiling process is not 100%

efficient and energy losses during the process in the form

of heat and carbon dioxide evolution can be extensive. There-

fore, it becomes extremely important to understand the silage

fermentation process in order to consistently produce high
quality silage.

Although there is no clear-cut division for the various
phases of the fermentation process, Barnett (1954) originally
desribed the process as consisting basically of four phases
and possibly a fifth phase depending on the course of fermen-
tation. These phases are:

Phase 1. Plant cells continue to respire, utilizing available
intermediary metabolites and simple sugars, result-
ing in the production of 002 and heat.

Phase 2. A relatively short phase of acetic acid production
by coliform bacteria (and others).

Phase 3. The conversion of soluble carbohydrates to lactic
acid by lactobacilli and streptococci, with most
silages reaching this stage two to three days after
filling.

Phase 4. Stage of quiescence of silage mass. In the absence
of buffer additions, lactic acid production contin-
ues and reaches a peak at 1 to 3 percent of the
fresh weight for corn silage. The pH of the ensiled
material declines to below 4.0 when the lactic acid
producing bacteria cease fermentative activity. The
ensiled material, if undistured, is now quite stable

for prolonged storage.

Phase 5. Proliferation of butyric acid producers. This
normally occurs in silages where the acidity is not
high enough to prevent the conversion of residual
carbohydrates and lactic acid to butyric acid by
clostridial bacteria. Ammonia from deamination of
protein, carbon dioxide and heat often accompany
this phase, particularly when air is introduced into
the silo mass (Huber, 1979a).

As mentioned before, losses due to the ensiling process can

be very extensive, and a large share of these losses can be

attributed to the action of plant and microbial cell respira-

tion (Barnett, 1954; Kempton, 1958; Watson and Nash, 1960).

When the plant material is ensiled, the living cells continue

to reSpire until all the oxygen is used up. The production

of lactic acid is an anaerobic process, and therefore it is

essential that anearobic conditions become established as

soon as possible within the ensiled mass. Previous research
has shown that delays in establishing anaerobic conditions
can be detrimental to the silage fermentation (Miller 23 gl..

1961; Langston gt a;., 1962: Ohyama gt g;., 1970). Obligate

anaerobes. such as clostridia and yeasts have been known to

compete with facultative lactic acid microbes for available
water soluble carbohydrates under aerobic conditions

(Takahashi. 1970). Utilizing sterilized corn inoculated with

bacteria.to make silage, Peterson gt gl..(1925), demonstrated

that plant cell respiration was nonessential and that

bacteria are mainly reSponsible for the production of acids

from sugar and starches.

Two major compositional changes occur in the corn plant
during ensiling: namely, the conversion of water-soluble
carbohydrates to short-chain organic acids and the degrada-
tion of plant protein to non-protein nitrogenous compounds
(Johnson.gt,gl., 1967, Demarquilly and Andrieu, 1973; Bergen
23 g;.. 1974).

' Carbohydrate Fermentation

In silage making. the green chop is preserved by lactic
acid formed in the fermentation process. The production of
lactic acid requires comparatively large amounts of ferment-
able carbohydrates in the ensiled material. Thus, the amount
of fermentable carbohydrates in the green chop becomes of
prime importance in producing the desired type of fermenta-
tion (Melvin, 1965, 0hyama.gt gl., 1973; Ohyama and Masaki,
1974). The carbohydrates available for fermentation are
primarily those within the water soluble carbohydrate frac-
tion (Zimmer, 1971).

It is generally accepted that as the corn plant matures.
soluble carbohydrates contained in the leaves and stems are
converted to insoluble starch in the kernels: thus, the total
level of soluble carbohydrates decreases, and in turn reduces
fermentation as well as the level of lactic acid produced.
However, McCullough (1973) summarized the soluble carbohydrate
content of nine creps (including maize), and reported that
under most ensiling conditions, the crops contained a suffi-

cient amount of available carbohydrate (about 6 to 8% of the

dry matter content). Ensiling the whole maize plant at 30 to
35% of dry matter has also been shown to supply adequate sol-
uble carbohydrates for fermentation (Hillman, 1977).

Prolonged cell respiration under aerobic conditions can
significantly decrease the quantity of available carbohy-
drates remaining for the anaerobic fermentation phase (Zimmer,
1971: McCllough, 1973). However, the soluble carbohydrates
surviving aerobic metabolism are fermentable by a variety of
micro-organisms, of which lactic acid bacteria are the most
important (Whittenbury 23 g;., 1967). Since the desired
fermentation is a rapid production of lactic acid, the
desired changes would be a low pH with a minimum loss of
carbohydrates. Johnson 33 gl., (1966) reported decreases in
corn plant soluble carbohydrate ranging from 39 to 83%, and
concurrent conversion to lactic acid. Both lactic and acetic
acids contribute towards avoiding undesirable fermentations.
However. lactic acid is the more important, it being present
in larger amounts than acetic acid and exerting a greater
effect on pH (Greenhill, 1964).

In summary, the most striking feature of carbohydrate
fermentation is the production of organic acids of which
lactic acid is the most important. Although there are many
factors that can influence the extent of lactic acid produc-
tion, the chief determining factor will be the amount of
water soluble carbohydrates available for fermentation.

Protein Degradation

The degradation of plant protein to non-protein

nitrogenous compounds during ensiling has long been a subject
of discussion. It is noteworthy that even in well-preserved
silages about 50% net breakdown of the protein may take

place (McDonald and Whittenbury. 1973). Various factors have
been known to affect the course and extent of the breakdown.
Some of those implicated are. dry matter or moisture content
of the crap at the time of ensiling, pH of the silage mass,
lactic acid content. level of oxygen. temperature, and most
of all enzymatic activity.

The importance of dry matter content of the whole plant
material at the time of ensiling cannot be overemphasized. it
being the most decisive factor influencing the extent of
protein degradation (Bergen 23 g;., 1974). Geasler (1970).
working with corn silage material, found that material
ensiled at 48 and 60% dry matter contained 30 and 26% of
total nitrogen in water soluble form. reSpectively. A
similar phenomenon was observed by Bergen gt g;.. (1974)
using corn silage material ranging from 32 to 85% dry matter.
and by Hawkins (1969) using alfalfa silage material. Immature
silage (low dry matter) tend to be higher in protein content,
but a larger portion is degraded to non-protein nitrogen
during fermentation (Hillman and Fox. 1977). According to
Andrieu and Demarquilly (1974), protein degradation is
inversely related to the dry matter content at ensiling.

The pH value is the most universal measure used in
assessing silage quality. In general, pH values associated

with well-preserved silages are between the ranges of 3.8 to

10

4.2 (Greenhill. 1964; Bergen. 1980). while that of poor
quality silages are above 4.2 (Greenhill. 1964). McCullough
(1961), is of the opinion that the high pH usually associated
with undesirable silage fermentation is a result of the
apparent protein breakdown with the resulting effect of
increased buffering capacity. Later. McDonald and Henderson
(1962) suggested that the contribution from the protein
fraction resulted from protein degradation to decarboxylated
amino acid bases. De Vuyst gt gl.. (1971) have made a
comprehensive study of the changes in amino acid composition
of grass and alfalfa during ensilage. They found that in
silages ranging in pH from 4.9 to 5.7. considerable degrada-
tion of amino acids occurred. eSpecially the basic amino
acids (lysine. histidine and arginine). In the same experi-
ment, these workers also observed that the amount of alanine
almost doubled in some silages. Hughes (1971) in a study of
high pH silages observed a decline in amino acid nitrogen
with a concomitant increase in volatile nitrogen which was
mostly ammonia.

The preservation of plant material as silage is depend-
ent on the accumulation of lactic acid and a correSponding
decrease in pH. However. even if a rapid drOp in pH is
achieved and the growth of proteolytic bacteria is inhibited.
some ammonia is still formed in the silo (Mo and Fyrileiv.
1979). Some lactic acid producing bacteria are also capable
of both deaminating and decarboxylating amino acids

(Whittenbury gt g;.. 1967). Brady(1966) isolated lactic

11

acid bacteria from farm silages and showed that Lactobacil-
lus plantarum and Pediococcus gp. deaminated serine to
pyruvate and arginine to ornithine. Ammonia nitrogen levels
in high lactate silages are frequently of the order of less
than 10% of total nitrogen (McDonald and Edwards, 1976).

In addition to deterioration of silage during the
utilization period, high amounts of oxygen during ensiling
have been shown to result in the formation of large quanti-
ties of volatile nitrogen from the deamination of amino acids

(Ruxton and McDonald, 1974).

Temperature is one of the factors affecting the success
of silage making. since the lactic acid bacteria have an
optimum temperature for growth. However. according to Nilsson
33 §;.. (1956), the Optimum temperature of 80 to 100°F for
lactic acid producing bacteria is also within the tempera-
ture range of proteolytic bacteria. Thus, it was not
surprising when Wieringa, (1960) reported that the ammonia
fraction in silages stored at 68°F was almost as high as at
95°F. Nilsson.gt,gl.. (1956) further stressed that tempera-
ture becomes an important factor in silage fermentation when
the crop is of high-protein and low-sugar content.

Both plant and microbial enzymes have also been impli-
cated as affecting the course and extent of protein degrada-
tion during the ensiling process. The general consensus is
that protein degradation is a two phase process; the first
involving the rapid breakdown of plant protein into peptides

and amino acids, primarily by endogenous plant proteases

12

(Barnett, 1954; Kemble. 1956: Watson and Nash, 1960:
McDonald and Whittenbury, 1973: Bergen g3,gl.. 1974), and
the second involving the subsequent degradation of amino
acids to volatile nitrogen and other nitrogenous compounds
by clostridial activity (McDonald and Whittenbury, 1973;
Oshima and McDonald, 1978).

 

Non-Protein-Nitrogen Treatment of Corn Silgges

In periods of high cost of oilseed meals. simple
nitrogen-containing substances such as urea. ammonia or
ammonium salts are added to cattle rations to replace plant
protein supplements. These compounds are referred to as a
group as non-protein nitrogen (NPN).

The notable advantages of corn silage are partially off-
set by its well known nutritional insufficiency with reSpect
to protein. Nevertheless. being high in energy and organic
acids, fermented silage make an ideal carrier of NPN in
ruminant rations. Urea has traditionally been the most widely
utilized from of NPN for treating silages. However. since
anhydrous ammonia is a cheaper source of NPN. researchers
became interested in its application to corn silage at
ensiling. This program was initiated at the Michigan State
University in the years 1967 and 1968. Evolving from the
original use of anhydrous ammonia mixed with water (aqueous
ammonia) were aqua ammonia (21-23%N), ammonia-molasses
mineral-solution (now bearing the trade name Pro-Sil) and
Cold-flo ammonia. These three sources of ammonia have been

hound to give results superior to urea, and allow for higher

13

intakes of NPN without decreasing milk production (Huber gt
g;.. 1980).

This section of the paper will consider data available
on the effects of using ammonia as an additive to the chopped
corn plant material at ensiling. and subsequent effects in
feeding trials.

Effectg of Ammonia Addition on Silage Fermentation

Although ammonia is added principally as a nutritive
supplement to furnish crude protein to cattle. it also exerts
a profound effect on silage fermentation. It was noted
earlier. that under natural ensiling conditions energy losses
in the form of heat and carbon dioxide evolution can be
extensive. Treating the chopped corn plant at ensiling
with ammonia has shown to reduce such losses when compared
to the untreated material (Juengst gt gt.. 1975: Huber gt g;.,
1979b). Juengst gt g;.. (1975) reported that the untreated
silage produced more carbon dioxide during the first 24 hours
than the ammonia (Pro-Sil) treated silage produced during
the entire 11 days of fermentation.

Initial increases in pH and higher lactic acid concen-
trations have also been reported for ammonia-treated corn
silages (Cash 1972: Henderson and Bergen 1972; Huber gt g;..
1973: Juengst gt g;.. 1975). This phenomenon has been
attributed to the increased buffering capacity resulting
from ammonia treatment. thus enabling the lactic acid-
producing bacteria to survive longer (Britt and Huber. 1975).

However, this phenomenon has not been found to be operative

14

at all levels of ammonia additions. Levels of ammonia
addition above 1% of silage dry matter have been found to
decrease lactic acid production (Huber gt g;.. 1979), and
total inhibition is achieved at 2.5% of the dry matter (Britt
and Huber. 1975).

Also worthy of note. is the increased water insoluble
nitrogen concentrations resulting from ammonia treatment
(Henderson and Bergen. 1972: Huber gt g;.. 1973). Bergen
gt g;.. (1974). suggested that this increased water insoluble
nitrogen fraction might be related to decreased proteolysis
of the original plant protein. More recent studies (Huber
gt g;.. 1979c. 1980a) revealed that 40% of the added ammonia
was recovered as ammonia in the water insoluble nitrogen
fraction in unfermented silage. In the fermented silage. 28%
of the added ammonia was recovered in the insoluble nitrogen
fraction. as revealed by 15N analyses (Huber gt g;.. 1979c):
this accounted for only 59% of the total increase in the
insoluble nitrogen fraction. The remainder was presumably
due to decresed breakdown of original plant protein as
suggested by Bergen gt g;.. (1974).

Apart from its influence on the silage fermentation
process, ammonia treated silages have also been shown to be
more stable during the utilization period (Britt and Huber.
1975: Soper and Owen. 1977; Buchanan-Smith. 1980). Britt
(1973). has attributed this increased stability to the

antifungal action of ammonia and ammonium salts.

15

Feedigg Trials

The ability of the ruminant to utilize NPN as a partial
replacement for the preformed protein in the ration is well
documented. However. it must be borne in mind that with
increasing levels of dietary nitrogen, rumen ammonia increases
more rapidly with NPN than with natural protein (National
Research Council, 1976). The ammonia nitrogen released from
NPN compounds as the result of rumen microbial enzyme action
may be utilized in the synthesis of microbial protein (Reid,
1953), but the extent to which this can be done will depend
on the energy concentration in the ration (Satter and Roffler
1975: Chalupa. 1978). A deficiency in intake of metabolize
energy may result in decreased animal performance (Wilkinson.
1978). However. as pointed out by Huber and Hung (1981).
inclusion of NPN‘in dairy cattle rations is profitable as
long as production is not diminished and utilization for
microbial protein synthesis is assured.

For the past 13 years, several investigations have been
made by workers at the Michigan State University to determine
the relative feeding value of corn silage treated with the
various forms of nitrogen-containing compounds at ensiling.
Many of the studies have revealed that milk production
(Huber and Santana, 1972; Huber et al., 1973; Boman, 1980;
Huber gt g;.. 1980b) and body weight gains (Henderson and
Bergen. 1972: Cook and Fox. 1976) in cattle fed silages
treated with ammonia solutions prior to ensiling. were

comparable or higher than those fed untreated or urea-treated

16

silages. Other workers (Honig and Zimmer. 1975: Buchanan-
Smith, 1980) have reported similar results in beef cattle
studies.

However. a closer look at the relative feeding value.
particularly in dairy cows. of corn silage treated at
ensiling with the different forms of anhydrous ammonia
(gaseous. aqueous, and ammonia-molasses-mineral solution)
reveals that the apparent differences are within the realm
of experimental error. A look at the summary in Table 1. of
some data on dairy cow performance on ammonia treated corn
silage will tend to support this view. Nevertheless. the
data in Table 1 reveals that the forms of ammonia can be
safely added at ensiling to corn silages varying in dry
matter content from 30 to 42% without depressing milk yields;
ammonia (Pro-Sil) addition to 53% dry matter depressed milk
yields as shown in experiment 2 on Table 1. Urea addition to
high dry matter silages (above 42%) have consistently
resulted in lower milk yields of lactating dairy cows than
addition to lower dry matter silages (Huber gt g;.. 1968:
Van Horn gt g;.. 1968).

Also worthy of note in Table 1, experiment 3, is the
significant (P<,05) depression in intake of dry matter with
the more concentrated ammonia (gaseous NH3 - 82%N).
According to the reported literature. silage protein was
lower for the gaseous ammonia than for the Pro-Sil treatment
(12.1 vs. 14.2% of dry matter). but the total dietary

protein for the gaseous ammonia treatment was 13.9% which

17

.oo .so .cN .w: .m .a .so msoa .Noc: Nm.m .mmmwsooe NN.mm .Amzz mac 2 Ne.MN waoeamocou monoxae <

.naNmN use MNmN ..Ns no page: "mucmcmcam

.z Nam uoawmu:00s
.H mom

.2 NNN emcoaoeocm

 

 

 

 

 

 

 

 

 

m~.H oq.~ mo.H m.mm q.o~ o.q~ o.q ¢.Nm

NN.H oq.m om.H ~.<w q.o~ H.¢~ o.N N.~m meoum

NH.~ we.~ oc.~ m.mw m.om m.m~ mm.o “.mm

EN.N os.~ mN.N N.Nm m.oN o.e~ mm.o s.~m mzz anommso
NDHDNmsou m.mNmNcs .omwm

mN.N mm.~ mm.~ o.~m m.qm m.c~ o.~ w.Hm meoom

me.N MN.N s~.N o.mm m.s~ N.NN mNm.o s.sm umzz snowman
ADLDNmaoo m.~NmNcm .oamw

o~.~ om.~ Hq.H e.qw o.- ~.mm o.q mm

om.o No.m mo.~ N.mm c.wa 5.6N e.m mm

a~.~ mo.~ <m.~ ¢.om o.mm ~.m~ o.~ Om demon;
NDLDNmacu N.onNcN .oaxm

mm.~ mw.~ «N.H ~.om o.- w.m~ m.~ Ne

cm.N mm.~ m~.H ¢.Hm m.- o.om o.~ Hm oflamoum

mm.~ Hm.~ n~.H m.mw m.o~ m.om o.H Hm quz m=c<
NDLDNmaou m.asaooo .oaxm

we Dawes: Neon N N we we N
umuum: hum Hmuoe owMNNm mucoumwmuom .Huaxm .Humxmioum compo Dwayne
xawz oxoucH Houumz Mun :0fiuooooum Jaw: N San mucoaumoue

 

mafiawmcm um mNCOEEm sag: woumouu owmaww once so mocmeuomuma Sou swamp mo huoseom .H m4m<e

18

was not low enough to depress intake (Huber gt g;.. 1979b).
However. the occurrence was not evident in eXperiment 4,
Table 1: highest intakes were recorded for the group
receiving the .77% gaseous ammonia treated silage (Huber gt

21.. 1979b).

Materials and Methods

Experiment 1, Silgge Fermentation Study

Whole corn plants (32-34% dry matter) were field chopped
and brought to the Beef Cattle Research Center where 30 kg
portions were treated with 0. .25, .52, or 1.08% added N as
ammonia (40% N). This required the addition of 0. .63. 1.3
or 2.7% ammonia (of silage dry matter).

To ensure complete mixing and uniform application of
treatments, each 30 kg portion of the chopped material was
weighed. then placed into a small feed mixer where the‘
Specified treatments were applied as the material rotated.
Immediately after treatment, duplicate samples of the freshly
chopped material. of about 3 kg each. were hand packed into
double layered polythene bags. The bags were evacuated to
remove air. tied securely with string. then placed in a
container packed with dry ice. These constituted day 0
samples after harvest. The same procedure was repeated for
the other samples, except that they were placed in 200 liter
covered plastic barrels to ensile for 1, 2. 6, 12 and 54 days
in a protected area at about 25°C. Total time elapsed
between harvesting and sealing of the bags was about 2 to 3
hours. Bagged samples were removed at the various times and
frozen at -20°C until analyzed. Parameters under investiga-
tion are shown in Table 2.

Silgge Analyses

Silage samples were removed from the freezer and allowed

19

20

to thaw at room temperature. Thereafter. dry matter was
determined on 50 gram portions by drying in a forced-air
oven at 60°C for 48 hours. The remaining portion of the

silage sample was comminuted by a commerical food cutter.

Table 2. Silage Parameters under Investigation

 

Dry matter

pH

Lactic acid

Total nitrogen

H20 insoluble nitrogen

H20 soluble nitrogen

Individual free amino acids (AA)
Essential and non-essential AA
Branched chain AA

Total free AA

 

Twenty-five gram.portions of the comminuted sample
were then homogenised in 100 ml double-distilled water for
2 minutes in an 0mmi-mixer. The homogenizer cup was immersed
in ice during the process. The pH was determined on the
homogenate. and 12 grams were removed and centrifuged at
20,000 x g for 20 minutes. The residue was washed with
double-distilled water and recentrifuged. This was repeated
three times. The residue was then analyzed for insoluble
nitrogen by macro-Kjeldahl. Total nitrogen was also deter-
mined by macro-Kjeldahl on 12 grams of the homogenate.
Soluble nitrogen was determined by difference between the

total and insoluble nitrogen. Fifteen ml of the supernatant

21

was deproteinized with 50% sulfosalicyclic acid (1 part SSA
to 10 parts supernatant) by centrifugation at 15,000 x g for
20 minutes. and analyzed for free amino acids as described
by Bergen gt g;.. (1974) and for lactic acid according to
Barker and Summerson (1941).
Statistical Analysis

To facilitate studying the effects of treatment and time
on the parameters studied (Table 2). a Split-plot repeat
measure design was utilised (Table 3). Error ”a“ tested
treatment effects, and error "b" tested both the time (day)
effect and interaction of time with treatment. Reaponse to

treatment was measured using orthogonal polynomial contrasts.

Table 3. Analysis of Variance for Split-Plot Analysis

 

Sgugce of Vggiance Degrees of Frgedom‘
Treatmentsa 3
Batches/treatment (Error a) 4

Days 5(2)b
Treatment x Days 15(6)
Batches x Days (Error b) 20(8)

 

3 Treatment = Control (no ammoniajj .25, .52, and 1.08%
ammonia.

b Numbers in parenthesis represent degrees of freedom for
analysis of amino acids (only 3 days were analyzed).

Dunnett's test as described by Gill (1978) was used to
compare ammonia treatments with the control, and other days

With day_0.

22

Ex er'me t 2. Feedi T al

Whole-chopped corn plant (32 to 37% dry matter) was
harvested for silage and placed in two 20 x 60 ft. silos
located at the Michigan State University Dairy Cattle Center.
Before ensiling, the chopped material (i inch) was treated
with either 2.“% Pro-Sil (13.6% N) or .36% anhydrous
ammonia (82% N) by the Cold-flo process.

The Pro-Sil was metered on to the chopped material as
it moved from self-unloading wagons into the blower to be
conveyed into the silo. The gaseous ammonia was also metered
on at the blower: however. it was first recondensed to a
liquid at -28°F in a simple cooling chamber before it was
metered on to the chopped material. The composition of Pro-
Sil is as shown in the footnotes of Table 1.

For 14 days prior to the feeding trial, 24 lactating
Holstein cows were fed a pretreatment ration (14% crude
protein) of corn silage. .33 kg concentrate per kg milk and
4.5 kg of alfalfa haylage. At the end of the pretreatment
period. cows were paired for milk yield. stage of lactation.
breeding groups and age. Within each pair. the two experi-
mental diets (Table 4) were randomly assigned. resulting in
a total of 12 cows per diet. Diets were fed at 10% in excess
of appetite during the treatment period which lasted for 12
weeks.

Silage samples were collected three times per week

during the feeding period and composited into three-week

23

Table 4. Ration composition of experimental diets

 

 

 

Feedstuff Cold-flo Pro-Sil

Dry Crude Dry Crude

Mattera Proteinb Mattera Proteinb
Corn silage 49.2 6.41 50.1 6.61
Alfalfa hay 9.9 1.37 10.0 1.38
Ground ear corn 30.4 3.01 31.0 3.07
42% Soy supplement0 7.4 3.49 7.4 3.49
Dicalcium phDSphate 1.0 .... .2 ....
Limestone .1 .... .3 ....
Calcium sulfate 1.3 .... .7 ....
Trace mineral saltd .7 .... .3 ....
Total composition 100.0 14.28 100.0 14.55
'E7%-3ffitotal .

h % contributed

9 Guaranteed chemical analysis: 42% protein (11.9% NPN),
2% Ether extract. 8.5% CF. 3% Ca, 1.05% P, 2.06% NaCl.
1.15% K. .9% Mg. and .6% S. Each Kg soy supplement
supplemented with 4400 IU vit. A and 2000 IU vit. D.

d Guaranteed to contain: .35% Zn. .2% Mn. .2% Fe. .03% Cu.
.005% Co. .007% I. and 96% salt.

24

composites. Feed intakes and milk weights were recorded
daily. and an AM and PM composite milk sample was taken
biweekly. Cows were weighed on two consecutive days. seven
days after beginning and at the end of treatment.

Silages were analyzed for dry matter. pH. lactic acid
and crude protein as in Experiment 1. Milk fat was determined
according to Babcock, milk protein by micro-Kjeldahl and
milk solids by AOAC (1965).

The paired T test as described by Gill (1978) was used

to statisically analyze the measured parameters.

Results and Discussion
Experimegt 1, Silgge Fermentation Study
Results from this study are presented in Tables 5
through 11.5. Analysis of variance of data for Tables 5
through 10. 11.2. and 11.3 did not reveal significant inter-
actions between treatment and time. so main effect means are
presented in addition to treatment combination means. In
Table 11.1. only treatment combination means are presented
since interactions were significant for the data.
Dry Matter: Initial dry matter content of silages was not
significantly altered by treatments or time of ensiling
(Table 5). This observation is consistent with other studies
(Britt and Huber. 1975: Huber gt g;.. 1979c: Boman. 1980).
However. a tendency towards a decrease over time is evident,
and this appears to be greater in the controls than the

treated silages. More water production coupled with a greater

5
2

.mm._H ma same has has No House unaccepm o

.No.a.H ma some vsospsopp and we Nopao cascadem n

.oaam Non

mcoapmcfisaovoc 039 new; madam Hopcosfiuoaxo 03p Scum mommno>m Poomoumou mosas> amaopmmtm

 

 

 

 

om.om mN.Nm s«.~m N~.~m om.mm mm.mm oases sac
om.Nm mn.on ms.Nm s~.mm oo.~m ~m.~m mm.~m mo.”
om.Nm Nw.om aa.am mo.mm o~.~m ~m.mm m .N« mm.o
om.am m¢.om ms.on mo.~m m:.~m om.fim om.~m mm.o eacoes<
nam.am ma.m~ No.0m mo.Nm N~.Nm mm.~m om.mn o Hoopeoo
I so m
some on NH 0 m m I o mmz mm
psoSpmomm NMQMAAMQMIHMINHMHWH z oooo< psoSPMowm

 

shoe :N Novena hum R :o

ommHNm

wcaaamso mo mafia use sowpmppcoocoo macossm mo voommm .m mnm<a

26

carbon dioxide evolution in controls than ammonia treated
silages might explain the higher decreases observed for the
controls. This parameter was not under investigation in the
present study. but Juengst gt,g;.. (1975) reported that
untreated silage produced more carbon dioxide during the
first 24 hours than the ammonia treated silage had produced
during 11 days of fermentation.

In our study, percent dry matter decreases from 0 to
54 days were 11.6. 7.4, 6.0 and 6.3% for the control. low.
intermediate and high ammonia additions. repectively. Smaller
percentage decreases for the ammonia-treated silages suggest
a lower energy loss during ensiling. and higher conservation
of the energy originally present in the chopped corn plant.
pg; Initially. ammonia treatment of silages resulted in
higher (P<301) pH values than controls (Table 6). This
observation is consistent with those of other studies
(Henderson and Bergen. 1972: Britt and Huber. 1975: Huber
gt,gl.. 1979b). and is thought to be associated with a
longer fermentation period and higher lactic acid. However.
by day 12 of ensiling pH values of all silages were similar
.and appeared to plateau about 4.0. In general, pH increased
with added levels of ammonia but decreased with time of
ensiling.
Lacttc Acid: Ammonia additions tended to delay increases in
lactic acid during the early stages of ensiling. but as

ensiling progressed increases were greater for ammonia-treated

.mm. + ma some seasonanaoo vcospsopv has no heaps unaccepm

.om._H ma some zoo mom mo house oumocmpm

.m:._H ma some psoEpmohp has no Nouns oumocmvm

.ANo.vac Hopscoo ease seamen saucaoaeacmam

.Ano.vmv accuses can» sense: sapeaoNcNemNm

.NNo.vmv o sac ease nosed sopcaoacocwam

.Amo.vmv o see can» Nazca sapcaoNcNewNm

.Aaoevmv oesoamoa Hammad PcmoachwNm

.oaam pea

mcofipmswsampoc 039 new: madam Hmwcosfiaomxo esp Seam momsuo>m vsommuama mooam> nuances u

Dove’s-I easel

 

sma.s ewa.= omm.: mm.o oN.N so.s scams sac

am.m ow.e mm.: mm.m mm.m ma.m mm.m mo.N

mo.e eso.s woe.s wme.m osm.s wo~.m Mae.m mm.o
no.3 woo.= c:o.: ems. mo~.: som.m omN.N m~.o «Noose<
mmo.m emm.m som.m ms.m m~.6 md.m Nms.m o Noupnoo

. Bel

memos an «N o N a o mmz as

a osvmoufi I; z @6004 usesvmomm

 

emsaum choc ca :n so wcwaamce no asap and soapmuvcoosoo macosss mo poogwm .0 mqm<s

8
2

.N0. + ma some soapm:H9500 escapees» has me Noose camocmpm
.om..H ma some use has me Noose ouwocspm

.om._fl ma some psesasouv has no Nahum whosoevm

.Amo.vmv Houasoo Eon“ vcmpommoo havsmoamdswwm

.Naoxvav 0 sea page seams: saacaoaoocmam

.AmoAvmv omcoamoa nsocfia pseudmficwam

.oaam Non

QC'OQ‘HOO

mcoavmcasheuou Moog 29M: seams HaemosHNoaxm 039 seam mommao>m snowshoes mosam> umasnmmln

.

 

 

 

 

 

oom.n oma.o omo.e 0:.N we. no. cases see

am.N so.m 62.: Nm.N no. No. no. mo.N

NN.m amm~.N wem.e ewes. we“. no. o. mm.o

sN.m comm.o oms.e sou. on~.~ oN.N o. m~.o «Noose<
omo.n om~.e omm.o can.s omm.~ me. one. o Noopcoo

--------uw----------umpmss sue.melm.-- ........ --- 2min

came an NH m m N o «:2 ma
Mmmswmoual, m, m 6 we an 2 oouc< psoavmwmm

owmadm

:Noo ca macs cavoma so wcaawmso mo made use coupon»:oocoo swcossm no voommm .u mqm<a

29

than control silages (Table 7). This differential response
to treatment was probably due to the increased buffering
capacity resulting from ammonia treatment. thus requiring
more lactic acid to reduce the pH to a low of 4.0. At day

54, lactic acid content of silages was higher (P<301) for
the low and intermediate levels of added ammonia. but
similar to controls for the high ammonia treatment (Table 7).

A lower lactic acid content throughout the ensiling
period for silages treated with the high ammonia (Table 7)
suggests that lactic acid production is inhibited at this
level of added ammonia. Other researchers have reported
similar results for ammonia-treated silages (Britt and Huber.
1975: Huber gt gin 1979c: Roma. 1980).

t t o e : Tetal nitrogen (% of dry matter) increased
over time in controls and treated silages (Table 8). This
was probably due to the carbon dioxide loss during fermenta-
tion and lower nitrogen retention by unfermented silages. as
suggested by Huber gt,g;.. (1979c). However. total nitrogen
for subsequent days was not different (P<310) from that at
day 0.

Comparing mean values of ammonia treated silages with
those of the controls. the highest level of added ammonia
resulted in higher (P<;05) nitrogen (1.32 vs 2.45% of dry
matter) . Similar results have been reported previously
(Huber gt,g;.. 1973. 1979b.c. 1980b).

‘Ingglnt1g_flitygggn: Water insoluble nitrogen (% of dry matter)

.mo. H.m« some use has no nouns camocovm o
.3”. H ma sees acosvmeup has no mouse unaccepm o
.Amoqvmv Houpsoo one can» Momma: zapsmoamacwum o
.Auo.vmv omcoomma seesaa pcmoauficmNm n

. .oaam hem

mcoavmcasuovoo 039 new: uoamm Hmwcosauonxo 039 Sean mowmuo>o ucomeuaou meoam> seasons m

 

 

 

 

 

0
3
NA.N oo.N mN.N mN.N mN.N mo.N «case see
oms.~ aw.~ an.~ as.~ a:.~ Hm.~ nm.~ mo.N
mm.“ ~o.~ no.“ mm.« 5.“ mm.“ mm.” ~n.o
sm.« am.N um.u am.« m.H on.“ as.” m~.o manosS<
owm.H «m.a on.” on.” mm.a an.d mu.a o Houpsoo
-------n-----------uoppas sue «0.x, ............. =o.s
asses . HM NH N H o mmz mm
a osvmommr mquaquM=M¢2mewi z covo< psoawmwmm
ommdam

caoo ma somehow: Hope» :0 wcaaamce me made and coapmhpsoocoo sacosss mo poommm .m mam<a

31

increased linearly (P<;01) with added ammonia (Table 9). but
tended to decrease over time. Water insoluble nitrogen was
maximized at the highest level of added ammonia. and the
corresponding mean values were higher (P<;05) than the
controls (1.15 yg. .87% of dry matter).

When treatment combination means were expressed as a
percentage of total nitrogen as shown in Table-9. decreases
over time greater for the controls than treated silages.
Decreases in percent of total nitrogen as water insoluble
nitrogen over time were 27, 16. 8. and 3 for the control.
low, intermediate and high ammonia. respectively (Table 9).

Similar results have been reported by others (Henderson
and Bergen. 1972: Huber gt g;.. 1979c. 1980b). and it was
suggested that the increased insoluble nitrogen concentra-
tions resulting from ammonia treatment was due to decreased
proteolysis (Bergen.gt,g;.. 1974). increased synthesis of
microbial protein (Juengst gt,gl .. 1975). or possibly a
binding of ammonia by insoluble compounds in silage (Huber
gt 31.. 1980b). A higher water insoluble nitrogen fraction
has been found to be associated with increased milk yields
of cows on ammonia compared to urea-treated silages. particu-
larly when they are fed large amounts of non-protein
nitrogen (Huber gt.g;.. 1980a).

Wate S ub N'tro e : The water soluble nitrogen content of
silages was not determined directly. but by difference
between the total (Table 8) and water insoluble nitrogen
fraction (Table 9).

32

.:0Npomnw pomp pom sowouwas Hope» moIR Pcomoaaou memosNCosma :N memosoz
.eo. + ma some has has no Noose pudendum
.eo. + ma some possesses has me mouse upmocmpm
.Nmo.vmv Nospcoo as» can» canon: saocaoNaNcmNm o

.Aaojvmv omsoomoa smegma scooamficmflm

m
e

U

.oaam you n

 

mCONpmcHsNouoo 039 29a: moaflm amazosauoaxolozp Souk mewono>m vsomoumoh mesam> usasnmaww

 

 

 

so. oo.N so. so.N so.N oo.N scams sac
omH.N Amsvss.a AneV3N.N “meand.e_ Amsvom.a Ammvnm.a No Vac.” mo.N
mo.a mongoo.a Ammvmo.a fiancee. Ammvfio.a Lemons." Am Vao.N ~m.o
so. Romano. Amwvom. Asovmm. Amovao.N Nemcao.a Awmvsm. m~.o macossa
saw. fiancee. .movmm. Asovnm. Aesvms. Income. masavmm. o Hopscoo
-i--------.-:--------ca»pas sec «min--- ...... a ........ spin
modes on «a N a o m=z mm
» sapwomb mrm, «m o o m on z.ceoo< asoapsowm

 

 

 

ommawm shoe :N

cowouads eaosaomcw News: :0 wcaaamco me saw» one sodaoNoSoosoo eacossm no woeumm .m mqm<s

33

.coapomhm vmzv New Comohpac Hopofiik Psomouamu monogamoama ca massacz

.no. + ma some how and mo Noose opmocmpm

.oH..H ma some Naospmoup has no Nouns ohmosmvm

.Nao.vmc o see can» canoes sapeaoNcoawNm

.Amo.vav o sun can» seamen sapeaoacacwom

.Nao.vmv Hopscoo as» can» seems: sapcaoNcacmNm

.Nmo.vmv Hopscoo as» can» seems: sapcaoacocmnm

.Auouvmv omconmoa unocaa vamonchHm

. .cmmaupN: No masses

cansaomcfi News: and Am manmmpaavov somzpon oocouommac on» voomouaou mooam> seduces u

no'omfi-IbDS-a

 

 

 

cmo.a com. com. NA. on. as. cases sea
son. Newcom.a Ammvoe.N Namvam.a Aamv «.N Naacmo.o Newcom.a mo.N
one. Now No.N Am one. Lemons. Ammw a. Aoavos. Am See. ~m.o
om. Non mm. As can. as com. Am an. Asmcam. A: can. m~.o ancoss<
was. Iowans. Ammvmm. Ammvms. Aomvmn. Ammvms. Nammvsm. o Hoopeoo
---------------------noppae see colmun-------- ....... -- an N .
cams Mm «N. a N o mmz as
PmoSpseum .mcw . c o m owl z oooo< pcoEpmomm

 

emsawm choc a“
sewage“: mansaom News; :0 wcwaamso we saw» one soaemupseocoo dwsosas no vacuum .oH mam<9

. 34

Water soluble nitrogen (% of dry matter) increased with
added ammonia (P<.01) and time of ensiling. Both intermediate
and highest level of added ammonia resulted in higher (P<.05
and .01. reSpectively) mean values than controls (Table 10).

Means for days 6. 12. and 54 were higher (P<.05 and .01)
than day 0. A strong negative correlation (-.85) between day
means for pH (Table 6) and water soluble nitrogen (Table 10)
suggests that the apparent increase in water soluble nitrogen
over time was inversely related to the rate of pH fall. There-
fore. it would appear that the more rapid the decline in pH.
the less protein breakdown. Evidence of this has been obtained
where crop material was acidified with formic acid during
harvesting (McDonald and Edwards. 1976).

From the percentage values for water soluble nitrogen
(% of total nitrogen) given in Table 10 for day 0 samples. it
would appear that extensive proteolysis occurs from the time
the crop is harvested. and is accentuated during the ensiling
process. Although higher mean values were recorded for ammonia
treated silages. it is apparent that increases are higher for
controls than treated silages.
Pyge amtgg-acids in water phase: Aggregate concentrations of
non essential amino acids (Table 11.1) contributed to a
greater preportion of the total free amino acids than essen-
tial amino acids (Tables 11.2 and 11.3). Of particular import-
ance. was the increase noted for alanine (Table 11.1) on high
ammonia treatment between days 0 and 54: alanine concentra—

tion increased approximately eight-fold above initial values.

35

In terms of % distribution this value would account for 70%
of total free non essential amino acids (Table 11.1) and 49%
of total free amino acids in the water phase (Table 11.2).
The decrease in aSpartic acid (Table 11.1) suggests some
conversion to alanine. since aspartic acid can be decarboxy-
lated by aSpartate-l-decarboxylase to alanine (Kemble. 1956).
However. the relatively small loss in aspartic acid compared
to the large increase in alanine discounts aspartic acid as
the major contributor (Table 11.1). A more plausable explana-
tion for the increased alanine seems through pyruvate. since
pyruvate can be reduced to lactic acid. or transaminated to
alanine. From the low lactic acid levels (Table 7) recorded
for the high ammonia treatment, it seems possible that some
of the pyruvate produced was diverted to alanine. Similar
observations have been made in continuous culture studies
(Erfle gt g;.. 1977) where it was shown that alanine concen-
tration rises rapidly when the ammonia concentration is high
in the rumen. These workers suggested that the alanine in

the rumen was probably acting as a reservoir for ammonia.

36

 

 

 

 

 

 

 

oomo.mmm oma.:m~ omN.osN oaa.:o~ am
ems. o Hm.mm ama.esa cmn.n:N N
mm.mN No.ms m«.os oo.oa mo.mm o acacaa<
mm.~m mm.- mm.an mo.Ns em
sose.m scam.a .ms.NH mme.as H
mm.~ so.m c~.m mm. ca.o o maaosau
mo.mo mo.NN ea.aoa N~.omN an
scam.~m oomm.mm eoa:.N omm.mm N
NE.@ w~.mn mm.a~ mo.N Nm.~m o meadows
os.sm :N.ea as.mm No.mm an
moa.s~ ons~.s~ was.~c can.sm a
em.s mm.Nm om.m~ mm.a~ ea.mm o ocaemm
Na.om s~.~s as.“ so.mN am
eN~.NN sous.m~ soam.o mao.ss a
mo.n mo.- =2.N~ mm.o~ an.om o acacomone
am.mm NH.N~N mo.aaa so.NNN am
eH~.ms pas.NN onm.ss omm.aa a
no.~« N~.ma nm.os mo.mm am.es 0 case
:oavamam<
:::::::::::::ommnnlbmr.Hs\w:::uw::::::::
exam mo.H mm. Tl, mm. o mcaaamno menus
kfivmz 3 2 e83 - .3 was 2:5

2H space

omega News: ommaam shoe

Ofidfim OOH.“ GO wfidfldmfiw H0 05%». USN SOHPdHPQQOSOO “H3055 MO POOMMH Hofid mgm<9

37

 

 

 

 

 

 

enso.mn comm.om oon. cs ooa.n~N an
fim one. N ems. Nu mm.mm N
an.: N no. N ems. mN No. mm o chcaNm
-Nscoam
«N.NN «.mn one. N: as. an em
am.nN o.~N new. NN as. o: N
o~.m m~.e~ mm.mN ma. Nu me. an o acNmousa
moos.es ooam.aNN scam. NNN 0mm. mmm .sm
a~.mm mN.om NN. an osmfi N
as.~N :m.a Nm.m~ me. ea c ocNossN
m mm 0 .mm Saw. as can. on an
some. .mN momm.mN Boa. oN Mom. an N
n~.a ma.mN ma. :N omo. AN 0 acNosaNuomN
as. Nu am.n~ N6.N~ can. an an
mooo. c oooo.o eooo.o coo. o N
N~.N oo. o oo.o oo.o oo. o o ochoNaNos
Bee. mm Ema. we so. No mm. mNN am
Sun. Nu Ema. Nn use. an cam. as N
66.6 :T om mn.o~ on. em NN. mm o ocNNa>
Illlllltlllllltmmdnn ON: HE\&3IIII:III:IIIIII
exam mouN o chacho moNos
L I we mamml enas<
Neoschcoov N.NN mqm<a

NNo.vmv Nnmz-ov Nonpcoo
Amo.vmv Ammz-ov Noupcoo
No.vmv o ado
mo.vmv o amc

scum NamNmNNNc NNpsaoNNNchm a
scam pcoumNNNu NNNcNoNNNcmNm o
scum NamNmNNNu NanmoNNNcmNm o
Scum pamNaNNNc NszmoNNNchm n
cams mo Nahum vhwccmpm u 2mm

 

38

 

 

 

wo.mmN mm.NmN om.mma am
oomm.mm mo oNN.mmN N
mm.mN mo.~a m:.wm o :cho
cmnocmnm
Nn.mmm we own mm Nam ow.mNN an
on.NmN o o omo.No N
wo.an mm.mmN hm.No o NaNpcommmuco:
NNNoe
Nm.mN mn.o: an
o:m.m oNN.m N
mm.m a .m N=.NN o chuNNmN:
uuuunnnuxunmmacm owfllwfikwsnuu aaaaa nun:
msmm tmo.N o chHNmno chom
11 mo vamp och<

 

 

 

 

Avozzavcoov N.NN mqm<a

39.

Concentrations of methionine and phenylalanine (Table
11.1) were relatively low in the water phase, being less
than 8% of the total free amino acids (Table911.2 and 11.3).

Branched chain amino acid (isoleucine. leucine and valine)
concentrations in the water phase also increased over time.
Mean values at day 5b were higher (P<.Ol) than at day 0
(Table 11.1). Also. mean values for the ammonia treated
silages were lower (P<.01) than controls.

In general. concentrations of free amino acids in corn
silage water phase increased over time (Tables 11.1 to 11.3).
Mean values for total free amino acids at day 5h were higher
(P<.Ol) than at day 0. but tended to be lower (P<.01) with
ammonia-treated than control silages (Table 11.3). This was
probably due to a decrease in proteolytic activity in the
ammonia-treated silages as suggested by Huber gt al.. 1973
and confirmed by Bergen.g§.§l.. 197h and Huber 23 g;.. 1979c.

ee ’ acid it 0 en as e cent of o i ' si a e N 2
Table 11.h shows the effect of ammonia addition and time of
ensiling on free amino acid N in corn silage water phase.
Nitrogen values are expressed as a percentage of total basal
(total N minus added N) nitrogen (Table 8) and provide
another way for comparing ammonia treatments with controls.

Total free amino acid N in corn silage water phase
increased over time in both controls and treated silages.
However. increases were greater for the controls than treated
silages. In day 0 samples, total free amino acid N (Table 11.“)
in the water phase accounted for 10.32, 8.16, 8.1h and 8.11%

#0

 

 

 

 

 

 

oa.omaa mn.omoa mm.meaa m«.aaea an
am.~o: ma.mma .mmm om.mna a
mm.ma: mm.am: .Hm: 53.5oc o fleece
no.5mn om.mo= mo.aom om.mmm an
ca.:oa a~.mma m~.ma~ n:.mmm H
m:.~oa o~.mo~ ~n.oma om.a- o Humveemme
fleece
~:.aa mn.s~ pa.mm m~.~n an
mo.na «3.5a o.o~ ma.e~ a
~e.m~ e~.m~ o.- oa.nn o ecaemwe<
05.nm ma.mm mm.ooa oo.m~« am
:~.m~ mm.on aa.o: mn.m~ a
mo.w: mo.o: H~.oa ma.au o madman
ma.oa ms.mm Hm.om an.mm an
wo.a: as. m ma.mm ma.oa H
an.me so.me ee.ne :~.me e ~mz-=ao a sac
u:-unuauuunnunaoum:n cum H£\msuununnnnuuuuan ;
mo." . . o maaaaeeo eeaoe
no mmmm, ocaa<

 

.ommsn nova: mwmaam :hoo

:« madam ocaam comm :o waaadmco no made and :oavmueCoocoo munoasm mo voommm N.HH mam<a

1.
hv

.Aao.vmv Anzzuow flamenco ease eezoa aapceoaeaemam a
mz o

.55.: Am

Houvcoo can» aosoa thGMOHmacwqm m
.Mao.vmv o awe use» eeewae aaeceoaeacmam e
. moevmv o see can» seem“: sauceeaeacmam o
.cmma Ho hoaho cumvcmvm n 2mm 9

.Wwwa mammmlaa zzonm mosam> sacs op mccommouaom m

 

 

 

 

 

us.am omw.amm www.mmc oma.umu mm.mmm .mm.mn mum.amma co.oam $0.:«m Hmpoa
em.om woo.mm~ mma.mo~ onm.mmm u~.aam mo.:w eea.umm ao.a- Ho.m- <<ma
o¢.~ oo.o~ mm.N~ mm.m~ 0:.on «n.m oo.n~ mo.m« ww.n~ wu<
an.n no.5: e~.am me.oe a~.ea on.“ oa~.mo m~.mn o:.~m mag
mm.e om.«w mm.mm Hm.am ma.wm u~.: m~.mw om.am «:.mo mzzsac a 5H0
92mm o. 92mm HM‘ iIdH o modem

6 uv< . mhwml ocaad

among nova: ommawm choc :a modem

cages moan you moocohoumau vcmodHH:Mam and .mmouuo pudendum .mcmma vacuum Cam: m.HH mnm<a

#2

of total basal nitrogen for controls. low, intermediate and
high ammonia treatments. reapectively. By day 54, deamination
(as measured by the increase in free amino acids) accounted
for 9.3. 7.1, 5.7 and 8.2% of the basal nitrogen for controls,
low, intermediate and high ammonia treatments. However. if
alanine increases on high ammonia were assumed equal to those
of the control silages. deamination between days 0 and 5h
would only account for 2.39% of total basal nitrogen (Table
11.5).

Though increases were recorded for threonine. methionine.
isoleucine. tyrosine, phenylalanine and arginine. they
appeared to be least affected by ensiling: their individual
contributions to the total free amino acid N in the water
phase (Table 11.“) accounted for less than 1% of total basal
nitrogen. I

In general, a lower amino acid N content in the water
phase of treated silages suggests that there was less deamina-
tion in treated than control silages. Moreover. increasing
ammonia decreased deamination. These data are the first to
show large increases in alanine in the water phase of corn
silages treated with high ammonia. increases which were

apparently not related to proteolytic effects.

“3

.Houvsoo op Amado possmmm ommouosa ocfisma< 9
.a2 poops wands z advop .o.«v amp was pcoEpanp vmsv no“ sowoupfl: Amman Havov+m.

 

 

 

 

 

 

Am.oHv

pam.ea om.ma H~.ma no.ma eo.m mm.a mw.m as.oa "a.m :H.o ma.m mm.oa fleece
aw.a mn.~ an.~ as.: «a. «a. do.” mm.a mm. mm. mm. ~H.H «um
mm.aa no.5 ma.a mo.m mm.n ~a.: mn.m mo.m 35.: 35.: mm.s ma.m <<mze
wm.s ma.e ~a.a mm.oa mm.~ Ho.~ an.m «3.: am.m ae.m ma.m ma.m <<me
@ 0 WW. om. HR. #3. #3. NW. 00. NC. 0W. 3Q. M0. WH<
we.” no.“ mm.a ma.~ we. mu. am. am. No.” oo.a om. «m.a use
mm.“ am. ma.a so.“ am. ma.a :a.H ma. an.“ an.” as.” as.« mm: ”mu
m u
mm. mm. as. co.“ as. «N. am. ma. am. am. as. mm. was
mm. «s. cm. ma. ed. «a. ma. w mm. as. 0”. ma. mm. one
ma. mm. «n. mm. «a. o«. as. mm. mm. ma. om. an. use
an. N~.H an.” n~.~ mm. :m. an. no. NH. mm. an. as. sea
mm. an. a. nu. ma. ma. as. an. mo. 5“. ma. em. eHH
as. am. N. am. o o o o o o o o pm:
He. we. no. 0 .a an. on. me. an. an. on. m. an. Hu>
m:.m ~m.n ow.~ a .m a~.H mm.“ ms.~ o .m ~m.~ on.“ e.” mm.a ea<
an. as. an. «5. am. as. mm. m . as. ma. cu. «m. use
ma. co. _:~.a no.“ as. am. am. am. as. as. me. 0:. cum
on. em. an. mo.” mm. a . ma. :m. m . me. a. No. gem
an. as. am. am. am. am. on. an. a . am. am. .as. use
am. mm.” as.“ so.“ om. mm. ma. mo.” aa. «a. we. «a. nm<

-----------------w------------mm.Have» me m-------------------------------

. nmz

o Podw mm. mm. 0 mo.“ mm. .mu. c me 2 eeee<
« o mmmm

 

 

omega have; owmadm

shoe :4 2 ages cause «can so msaaauso_mo cad» c:s.:ouaacum «access no Poomhm :.HH mqm<a

an

.Az venom mssfle 2 Have» .o.a~ h¢u_usm pcmmpmmup asap son sowohvas Amman Hmpov R,m

 

 

 

am.~ ee.m we.a mm.e emceee
om.oa oo.ma H~.ma me.aa em sen
«a.m aa.m ma.o ~n.oH o awn
--------------u-----mzfleece we mn-----------------
mp.“ an. mm. {x o
.2o :2 we 2 emcee . seem

 

 

 

omega amps: mwmawm :uoo

CM z odes ocasd «mum Haven :0 wcdamem mo med» cam coavnucm masosfia Ho voommm m.HH mam<e

45
Experiment 11: Feedigg Trial
Silage Analyses: Table 12 shows the dry matter, pH, lactic

acid and crude protein of silage composites collected during
the 12 weeks of treatment.

Dry matter content of silages was higher (P<.05) for the
Pro-Sil than Cold-flo treated (36.h# Kg. 3h.61%). This was
probably due to a higher dry matter of corn plant material
entering the Pro-Sil silo. or the added minerals and molasses
in Pro-Sil. since there was little (1_.5 units) or no
variation from mean values (Table 12) in either silage
throughout the entire feeding period.

The lactic acid content of the Pro-Sil treated silage
was higher (P<.02) than the Cold-flo (7.67 zg. 6.30% of dry
matter)3 suggesting that minerals in Pro-Sil were affecting
the silage fermentation separate frOm the ammonia. Evidence
of this was obtained by Lomas g1,§;.. (1978) who found 16%
higher lactic acid in corn silage treated at ensiling with
anhydrous ammonia (Cold-flo) plus minerals compared to the
anhydrous ammonia alone. A lower lactic acid content of
Cold-flo than Pro-Sil treated silages was also reported by
Fox and Cook. (1977).

Differences in pH and crude protein of silages were
small (P>.10).

Aging; Respogge: Response of lactating dairy cows to the two

NPN treated silages is summarized in Tables 13 through 15.
‘Dry matter intakes from silage (based on a fixed % of

total ration as shown in Table h) as Kg/day or % of live

#6

TABLE 12. Dry matter. pH, lactic acid and crude protein of
corn silage compositesa

 

r

b

 

ggem Cold-flo Pro-Sil ssm Effggt
Dry matter (fl) 3n.61 36.na .33 P<.05
pH u.uu n.37 .12 NSC
Lactic acid ($0M) 6.30 7.67 .15 P<.02
Crude protein ($0M) 13.03 13.20 .16 NS

 

E"Mean values represent averages for 3-four week composites

b of the silages fed during the 12 weeks treatment period.
SEM 3 standard error of mean difference.

° NS = non significant (P;.10).

TABLE 13. Influence of non protein nitrogen source on dry
matter and protein intakes of dairy cowsa

 

 

‘NBN source
Itgm Cold—f;0 Pro-Sil SEMb Effect
Silage dry matter (Kg/day) 8. 56 8.75 .3“ NSc
(% BW) 1. he 1.u9 .05 NS

Total dry matter (Kg/day) 17.39 17.47 .68 NS
(5 BW) 3.01 2.95 .10 NS

Total crude protein (Kg/day) 2.51 2.57 .11 NS

Crude protein from NPN
(% of total GP) 16.05 16.75 .08 P<.001

 

a’Me'an values represent 12 cows per treatment for 12 weeks.
SEM a standard error of mean difference.
° NS a non significant (P>.10).

a?
weight were similar for both groups (Table 13). Previous
work (Huber 23 §;.. 1979b) showed depressed intakes by dairy
cattle of silage treated at ensiling with .375% gaseous
ammonia (82% N). However. this effect was not found to be
consistent (Huber 23 g;.. 1979b). Intakes of total dry matter
as Kg/day or % of live weight were also similar for both
silages (Table 13). Intakes of total crude protein tended
to be higher for the Pro-Sil group. but differences were not
significant. However, intakes of crude protein from NPN (% of
total crude protein) were higher (P‘.001) for the Pro-Sil
than Cold-flo group (Table 13). A higher intake of silage
dry matter by the Pro-Sil group. though not significant might
explain the higher NPN intake (Table 13).

Milk yields of cows averaged about 28 Kg/day during pre-
treatment (Table lh). The decrease in milk yields from pre-
treatment to the end of the trial was probably due to cows
being put on trial in mid lactation, averaging about 125 days
postpartum when treatment commenced. Milk yields and persis-
tencies during treatment were slightly higher for cows on
Cold-flo, whereas fat-corrected and solids-corrected milk
were slightly higher for those on Pro-Sil, but differences
were not significant.

Milk components were all within the normal range, and
did not differ significantly between groups (Table 15). Both
groups gained .29 kg body weight during treatment, typical of
cows after peak lactation. Efficiency of conversion of feed

to milk was also similar for both groups (Table 15).

#8

TABLE 1n. Influence of non protein nitrogen source on milk
yields of dairy cowsa

 

 

NPN source . b
tem Cold-flo Pro-Si; SEM Effect
Pretreatment (Kg/day) 28.07 28.32
Treatment (Kg/day) 29.88 2h.25 1.32 NSC
Persistencyif)d I 88.96 85.40 2.9h NS
Fat corrected milk (Kg/day) 21.89 22.15 1.22 NS
Solid corrected milk (Kg/day) 22.21 22.27 1.20 NS

 

fMean values other than pretreatment represent 12 cows per
b treatment.
SEM = standard error of mean difference.
° NS = non significant (P>.10).
d Persistency (%) = (treatment milk/pretreatment milk) x 100.

TABLE 15. Influence of non protein nitrogen source on milk
composition, weight change and‘feed effeciency of
dairy cows

 

 

 

Itgm_ - Cold-flo Pro-Sil SEEP Effect
Milk composition c
Protein (fi) 3.20 3.06 .11 NS
Fat (%) 3.20 3.ua .13 NS
Solids not fat ($) 8.76 8.75 .13 NS
Weight change (Kg/day) .29 .29 .08 NS
Feed efficiencyd 1.u3 1.u1 .08 NS

 

a'iiVIean values represent 12 cows per treatment for 12 weeks.
b SEM I standard error of mean difference.
3 NS a non significant (P>.10).

Feed efficiency a treatment milk yield/total dry matter
intake.

“9
Although there have been no previous comparisons between
the feeding value of corn silages treated with Cold-flo and
Pro-Sil for dairy cattle. these results compare favorably
with those of Donaldson and Thomas, (1978) for Cold-flo
treated corn silage, and of Huber 23 al.. (1979b) for Pro-
Sil treated corn silage.

SUMMARY AND CONCLUSIONS

This study consisted of two experiments. Experiment 1
determined the nature of the protein break-down occuring in
corn silages treated with or without ammonia. Whole chopped
corn plants (32-34% DM) were treated with 0. .25. .52. or
1.08% added N as ammonia (of silage dry matter). Immediately
after treatment. about 3 kg material were placed in 5 mil
polythene bags which were then evacuated and served as
experimental silos. Silage samples were taken on days 0. 1.
2, 6, 12 and Sh and analyzed for dry matter. pH, lactic acid,
total and water-insoluble nitrogen, and free amino acids in
the water phase. The fermentation patterns for dry matter.
pH. lactic acid, total and water insoluble nitrogen were
similar to those reported by other researchers (Britt and
Huber. 1975: Huber gt_gl.. 1980b) for ammonia treated corn
silage. Alanine concentrations increased substantially over
time in the water phase of high ammonia treated silages, a
phenomenon which is.apparently not related to proteolytic
effects. It was suggested that the increased alanine was
probably through pyruvate. Increased total amino acid N
between days 0 and 5h on high ammonia treatment would account
for 2.h% of total basal (original) nitrogen if alanine

increase assumed equal to controls. Apart from the large

50

51
increases in alanine. these data support earlier studies
that suggest that ammonia inhibits plant protein breakdown.
Further eXperimentation is needed to more clearly define the
nature of alanine increases in ammonia-treated silages. and
possible nutritional significance in ruminant nutrition.

In Experiment 2, 29 lactating Holstein cows were employed
to determine the relative feeding value of corn silages
treated with Pro-Sil or anhydrous ammonia (by Cold-flo
process). Prior to going on trial, cows received a standard-
ization ration (1h% crude protein) for 1h days. Thereafter.
cows were paired based for milk yields. stage of lactation.
breeding group and age. Within each pair. experimental diets
(49 to 50% of either treated silage) were randomly assigned.
The feeding period lasted for 12 weeks. during which time
silage samples were collected and analyzed for dry matter.
pH. lactic acid and crude protein. Daily dry matter intakes
and milk weights were recorded, and biweekly milk samples
analyzed for milk fat, protein and total solids. Cows were
weighed on two consecutive days, seven days after beginning
and at the end of treatment. Dry matter content of silages
was higher for the Pro-Sil than Cold-flo treated, which was
apparently due to a higher dry matter of corn plant material
entering the Pro-Sil silo, or the added minerals and molasses
in Pro-Sil. Lactic acid was also higher for the Pro-Sil silage.
but differences in pH and crude protein were negligible.

Dry matter and total crude protein intakes were similar for

52
both silages. However, the slightly higher intake of silage

(based on a fixed % of total ration) by the Pro-Sil group

was reflected in a higher intake of crude protein from

NPN. No major differences in milk yields, persistencies.

milk components or feed efficiencies were observed between
groups during treatment. Both groups showed similar weight
gains. These data suggest that Cold-flo treated corn silages
can be used as efficienty as Pro-Sil treated silages in
rations for lactating dairy cows. However. rations containing
Cold-flo treated corn silage would need greater mineral

supplementation than those with Pro-Sil silage.

Bibliography

Andrieu. J. and C. Demarquilly. 1974. Feeding value of maize
forage 111. Influence of composition and fermentation
characteristics on digestibility and voluntary intake
of maize silage. Ann. Zootech. 23:27.

AOAC. 1965. Official Methods of Analysis (10th Ed.) Associa-
tion of Official Analytical Chemists. Washington. D.C.

Balch. C. C. 1967. Problems in predicting the value of non-
protein nitrogen as a substitute for protein in rations
for farm ruminants. World Review of Animal Production.

3:84.

Barker. S. B. and W. H. Summerson. 1941. The colorimetric
determination of lactic acid in biological material.
Jo B1010 Chemo 13835350

Barnett. A. J. G. 1954. Silage Fermentation. Academic Press.
Inc. New York.

Bentley. 0. 6.. E. W. Klosterman and P. Engle. 1955. The use
of urea to increase the crude protein content of corn
silage ior fattening steers. Ohio Agr. Expt. Sta. Res.
3111. 77 e

Bergen. W. G. 1975. The influence of silage fermentation on
nitrogen utilisation. Proc. 2nd Annu. Silage Symp..
Ann Arbor. MI.

Bergen. W. G. 1980. The ensiling process of corn silage. In.
Peed Management. A Watt Publn. Dec. p.22.

Bergen. W. G.. E.M. Cash and H. E. Henderson. 1974. Changes
in nitrogenous compounds of the whole corn plant during
ensiling and subsequent effects on dry matter intake by
sheep. J. Anim. Sci. 39:629.

Boman. R. L. 1980. Influence of source and level of non-

protein nitrogen additions on the nutritive value of
corn silage. Ph.D. Thesis. Mich. State Univ.. E. Lansing.

53

54

Bothast. R. J.. E. B. Lancaster and C. W. Hesseltine. 1973.
Agmogia kills Spoilage molds in corn. J. Dairy Sci.
5 :2 1. .

Brady. C. J. 1966. The redistribution of nitrogen in silage
by lactic-acid-producing bacteria. Aust. J. Biol. Sci.
1931230

Britt. D. G. 1973. Effect of organic acids and non-protein
nitrogen on fungal growth. nutritive value. fermenta-
tion and refermentation of corn silage and high
moisture corn. Ph.D. Thesis. Mich. State Univ.. E.
Lansing.

Britt. D. G. and J. T. Huber. 1975. Fungal growth during
fermentation and refermentation of non-protein nitrogen
treated corn silage. J. Dairy Sci. 58:1666.

Buchanan-Smith. J. G. 1980. cold-flo ammonia treatment on
corn for silage at two dry matter levels and its
preservation and feeding value for yearling steers.
72nd Ann. Mtg. Am. Soc. Anim. Sci. p.349 (Abstr.).

Chalupa. W. V. 1978. Advances in Experimental Medicine and
Biology. Production of animal protein from non-protein
nitrogen chemicals. Mendel Friedman ed.. Plenum Press.
NeYe 1058h730

Cook. R. J. and D. a. Fox. 1976. Anhydrous ammonia addition
to corn silage. J. Anim. Sci. 43:317 (Abstr.).

Coppock. C. E. and J. B. Stone. 1968. Corn silage in the
ration of dairy cattle: A review. New York State
Collegg of Agriculture-Cornell University Misc:
B1111 e‘ 9 e

Demarquilly. C. and J. Andrieu. 1973. Nutritive value and
utilisation by cattle of green. ensiled and dehydrated
whole maize plant. Proc. 24th Annu. Meeting Eur. Ass.
Anim. Prod.. Vienna. Austria. Sept. 1973.

DeVuyst. A.. W. Vervack. M. Vanbelle. and V. Jadin. 1971. Z.
fur Tierphysiol. Tierernahr. und Futtermittelk. 27:82.

Donaldson. B. M. and J. W. Thomas. 1978. Personal communica-
tion.

Egan. A. R. and R. J. Moir. 1965. Nutritional status and
intake regulation in sheep 1. Effects of duodenally
infused single doses of casein. urea and propionate
upon voluntary intake of low protein roughage by sheep.
Aust. J. Agric. Res. 16:437.

55

Erfle. J. D.. F. D. Sauer. and S. Mahadevan. 1977. Effect of
ammonia concentration on activity of enzymes of ammonia
assimilation and on synthesis of amino acids by mixed
gumenéBacteria in continuous culture. J. Dairy Sci.

0310 0

Fox. D. G. and R. J. Cook. 1977. Performance of steer calves
fed corn silage treated with three sources of anhydrous
amgogia. Mich. State Univ.. Agr. Expt. Sta.. Res. Rep.
32 : 9.

Geasler. M. R. 1970. The effect of corn silage maturity.
harvesting techniques and storage factors on fermenta-
tion parameters and cattle performance. Ph.D. Thesis.
Mich. State Univ.. E. Lansing.

Gill. J. L. 1978. Design and analysis of experiments in the
animal and medical sciences: Vol. 1. Iowa State Univ.
Press. Ames.

Goffart. M. A. 1877. The Ensiling of Maize and Other Green
Fodder Crops. Trans. and Publ.. by J. B. Brown.

Greenhill. W. L. 1964. The buffering capacity of pasture
’plants with special reference to ensilage. Aust. J.
Agric. Res. 15:511.

Hawkins. D. R. 1969. The effect of dry matter levels of
alfalfa silage on intake and metabolism in the rumi-
nant. Ph.D. Thesis. Mich. State Univ.. E. Lansing.

Henderson. H. E. and W. G. Bergen. 1972. Treating corn silage
with gaseous ammonia for feed lot cattle. Mich. State
Univ.. Agr. Expt., Sta. R68. Rep. 17u'180

Hillman. D. and D. G. Fox. 1977. Nutrient content of corn
silage. In. Corn Silage. Mich. State Univ. Ext. Bull.
E-1139 o

Honig. H. and E. Zimmer. 1975. Experiments on use of Pro-Sil
as a NPN additive for maize silages. Proc. 2nd Ann.
Silage Symp. Ann Arbor. MI.

Huber. J. T. 1975. Protein and non-protein nitrogen utilisa-
tion in practical dairy rations. J. Anim. Sci. 41:954.

Huber. J. T. 1979a. Making good silage is an art and a
science. In. Hoards Dairyman. August Publication.

p. 1059.

56

Huber. J. T.. H. F. Bucholtz and R. L. Boman. 1980a. Ammonia
versus urea-treated silages with varying urea in
concentrates. J. Dairy Sci. 63:76.

Huber. J. T.. J. Foldager and N. E. Smith. 1979c. Nitrogen
distribution in corn silage treated with varying levels
of ammonia. J. Anim. Sci. 48:1509.

Huber. J. T. and L. Kung Jr. 1981. Protein and non protein
gitrogen utilisation in dairy cattle. J. Dairy Sci.
:1170.

Huber. J. T.. R. E. Lichtenwalner. R. E. Ledebuhr and C. M.
Hansen. 1979b. Gaseous ammonia treatment of corn silage
for dairy cows. J. Dairy Sci. 62:965.

Huber. J. T.. R. E. Lichtenwalner and J. W. Thomas. 1973.
Factors affecting response of lactating cows to ammonia-
treated corn silages. J. Dairy Sci. 56:1283.

Huber. J. T. and 0. P. Santana. 1972. Ammonia treated corn
silage for dairy cattle. J. Dairy Sci. 55:489.

Huber. J. T. N. E. Smith and J. Stiles. 1980b. Influence of
time after ensiling on distribution of nitrogen in
corn silage treated with ammonia. J. Anim.Sci. 51:1387.

Huber. J. T. and J. W. Thomas. 1971. Urea-treated corn silage
in low protein rations for lactating cows J. Dairy
8310 54:224.

Huber. J. T.. J. W. Thomas and R. S. Emery. 1968. Response
of lactating cows fed urea-treated corn silage
harvgsged at varying stages of maturity. J. Dairy Sci.
5131 0 0

Hughes. A. D. 1970. The non-protein nitrogen composition of
grass silages 11. The changes occurring during the
storage of silage. J. Agric Sci.‘Camb. 75:421.

Hughes. A. D. 1971. The non-protein nitrogen composition of
grass silage 111. The composition of spoilt silages.
J. Agric. SCiep Cambe 76.329.

Jenkins. H. M. 1884. Report on the practise of ensilage at
home and abroad. Royal Agr. Soc. J. 20:126.

Johnson. R. R.. K. E. McClure. L. J. Johnson. E. W. Klosterman
and G. B. Triplett. 1966. Corn plant maturity. I.
Changes in dry matter and protein distribution in corn
plants. Agron. J. 58:151. '

57

Johnson. R. R.. K. E. McClure. E. W. Klosterman and L. J.
Johnson. 1967. Corn plant maturity. 3. Distribution of
nitrogen in corn silage treated with limestone. urea
and diammonium phosphate. J. Anim. Sci. 26:394.

Johnston. J. F. W. 1843. The feeding qualities of the natural
and artificial grasses in different States of dryness.
Agr. Soc. Scotland. 1:57(Trans.).

Juengst. F.. H. E. Henderson. J. Walters. J. Pieper. K.
Kramer and J. Dika. 1975. Effect of Pro-Sil treatment
on the microbiologiCal and chemical characteristics of
corn silage during primary and secondary fermentation.
Proc. 2nd Ann. Silage Symp. Ann Arbor. MI.

Kemble. A. R. 1956. Studies on the nitrogen metabolism of the
ensilage process. J. Sci. Food Agric. 7:125.

Kempton. A. G. 1958. Bacterial. biochemical and environmental
interrelations in fresh and ensiled forages. Ph.D.
Thesis. Mich. State Univ.. E. Lansing.

Langston. C. W.. C. Bouma and R. M. Conner. 1962b. Chemical
and bacteriological changes in grass silage during the
early stages of fermentation. 11. Bacteriological
changes. J. Dairy Sci. 45:618.

Langston. C. W.. H. G. Wiseman. C. H. Gordon. W. C. Jacobson.
C. G. Melin. L. A. Moore and J. R. McCalmont. 1962a.
Chemical and bacteriological changes in grass silage
during the early stages of fermentation. I. Chemical ,
changes. J. Dairy Sci. 45:396.

Lomas. L. W.. D. G. Fox. W. G. Bergen and J. R. Black. 1978.
The effect of anhydrous ammonia treated corn silage and
protein supplementation strategy on the performance of
growing and finishing steers Mich. State Univ.. Agr.
Expt. Sta.. Res. Rep. 353:143.

Mabbit. L. A. 1951. The role of plant cells in the ensilage
process: an approach to the problem. Proc. Soc. appl.
Bact. 14:147.

Mac Pherson. H. T. and P. Violante. 1966. Ornithine putrescine
and cadaverine in farm silage. J. Sci. Fd. Agric. 17:124.

Mc Cullough. M. E. 1961. A study of factors associated with
silage fermentation and dry matter intake by dairy
cows. J. Anim. Sci. 20:288.

Mc Cullough. M. E. 1973. Optimum feeding of dairy animals
for meat and milk. Univ. of Georgia Press. Athens.

58

McDonald. P. and R. A. Edwards. 1976. The influence of
conservation methods on digestion and utilisation of
forages by ruminants. Proc. Nutr. Soc. 35:201.

McDonald. P. J. and A. R. Henderson. 1962. Buffering capacity
of herbage samples as a factor in ensilage. J. Sci.
FOOd Agric. 1333950

McDonald. P. and R. Whittenbury. 1973. The ensilage process.
In. Chemistry and Biochemistry of Herbage. Vol. 3.
(G. W. Butler and R. W. Bailey eds.) Academic Press:
New York and London. “

Melvin. J. F. 1965. Variations in the carbohydrate content
of lucerene and the effect on ensilage. Aus. J. Agric.
Res. 1639510

Miles. M. 1918. Silos. ensilage and silage. A practical
treatise on the ensilage of fodder corn. Orange Judd.
Co. N.Y.

Miller. W. J.. H. L. Dalton and J. K. Miller. 1961. Sudan
grass silage at two stages of maturity versus ryegrass

and crimson clover with two filling procedures. J.
Dairy Sci. 44:1921.

Mo. M. and E. Fyrileiv. 1979. Methods of estimating ensiling
losses. Acta Agriculturea Scandinavica 29:49.

NRC. 1976. Urea and other non protein nitrogen compounds in
animal nutrition. Nat. Acad. Sci. Washington. D.C.

Nilsson. R.. L. Toth and C. Rydin. 1956. Studies on fermenta-
tion processes in silage. The role of temperature.
Arch. fur Mikrobiologic. 23:366.

Ohshima. M. 1970. Studies on the fate of free amino acids
in silage 1. The fate of monoamino dicarboxylic acids
in reg clover juice during incubation. Jap. J. Zootech
$01. 1397.

Ohshima. M. and P. McDonald. 1978. A review of the changes
in nitrogenous compounds of herbage during ensilage.
J. Sci. Fd. Agric. 29:497.

Ohyama. Y. and S. Masaki. 1974. Effects of ensilage tempera-
ture on silage fermentation with Special reference to
the stage of the temperature treatment and soluble
carbohydrate content of the material. Jap. J. Zootech.
SCis u58h190

59

Ohyama. Y.. S. Masaki and T. Morichi. 1970 Studies on various
factors affecting silage fermentation§V111. Changes in
microflora and organic acid composition during ensilage
as affected by the introduction of air after ensiling.
Jap. J. Zootech. Sci. 41:625.

Ohyama. T.. S. Masaki and T. Morichi. 1973. Effects of temper-
ature and glucose addition on the process of silage
fermentation. Jap. J. Zootech Sci. 44:59.

Reid. J. T. 1953. Urea as protein replacement for ruminants.
' A review. J. Dairy Sci. 36:955.

Ruxton. I. B. and P. J. McDonald. 1974. The influence of
oxygen on ensilage 1. Laboratory studies. J. Sci. Food
Agric. 253107.

Satter. L. D. and R. E. Roffler. 1975. Nitrogen requirement
and utilisation in dairy cattle. J. Dairy Sci. 58:1219.

Smith. R. H.. D. N. Salter and K. Daneshvar. 1977. Utilisa-
tion of nitrogenous nutrients and particularly NPN by
the bovine. J. Nuclear agric. biol. 6:8.

Soper. I. G. and F. G. Owen. 1977. Improving silage preserva-
tion and stability with an ammonia-molasses-mineral
solution. J. Dairy Sci. 60:1077.

Takahashi. M. 1970. Influence of level of initial air inclu-
sion in ensiling on.quality of silage. 7. Relation of
the level of initial air inclusion and the seepage cf
piang juice to the quality. J. Jap. Soc. Grassland Sci.
1 :9 .

U.S.D.A. 1981. Crop production-Annual Summary. p. A-3.

Van Horn. H. H.. C. F. Foreman and J. E. Rodriguez. 1968.
Effect of high-urea supplementation on feed intake and
milk production of dairy cows. J. Dairy Sci. 50:709.

Voss. N. 1966. Amines and ammonia as products of protein
decomposition in silage. Proc. 10th Int. Grassland
Congress (Helsinki). p. 540.

Waldo. D. R. 1968. Symposium. Nitrogen utilisation by the
ruminant. Nitrogen metabolism in the ruminant. J.
Dairy Sci. 51:2 5.

Waldo. D. R. and N. A. Jorgensen. 1981. Forages for high
animal production: Nutritional factors and effects of
conservation. J. Dairy Sci. 64:1207.

60

Watson. S. J. and M. J. Nash. 1960. The Conservation of Grass
and Forage Crops. Oliver and Boyd. London.

Whittenbury. R.. P. McDonald and D. G. Bryan Jones. 1967. A
short review of some biochemical and microbiological
aSpects of ensilage. J. Sci. Fd-Agric. 18:441.

Wieringa. G. W. 1960. Some factors affecting silage fermenta-
tion. Proc. 8th Inter. Grassland Congress. p.497.

Wilkinson. J. M. 1978. The ensiling of forage maize: effects
on composition and nutritive value. In. Forage Maize:
Production and Utilisation. E. S. Bunting g: §;(eds.).
London . ARC .

Zimmer. E. 1969. Biochemische Grundlagen der Einsauerung.
Proc. 3rd Gen. Meet. Grassld. Fed. Braunschweig. p.113.

Zimmer. L. 1971. Factors affecting silage fermentation in
silo. Inter. Silage and Res. Conf.