Note: Descriptions are shown in the official language in which they were submitted.
This inven-tion relates to a method for improving the
production of propionic acid and thus increasing the propionate:
acetate ratio in the course of the microbiological conversion of
cellulose and starch containing feedstuffs to short chain
volatile fatty acids in the ruminant animal. This invention
further relates to the use of antibiotic AV290, antibiotic
AV290 sulfate~ an antibiotic AV290-syntan complex, an antibiotic
AV290~alkyl sulfate complex, and/or an antibiotic AV290 alkylated
derivative for increasing the production of propionic acid and
suppressing the production of acetic acid during the micro-
biological digestive processes taking place in the animal rumen.
This invention also relates to novel compositions of
matter useful in favorably altering the acetate:propionate
ratio in the course of the microbiological digestive processes
taking place in the rumen of animals such as cattle, sheep and
goats. More particularly, it relates to compositions in oral
dosage unit form comprising from about 10 mg. to about 1 gram
per daily dosage unit of one or more of the following active
ingredients tin any proportions) which are useful for enhancing
the production of propionates in said animals rumen while the
animals feed is digested:
(1) antibiotic A~290 whose preparation and properties
are disclosed in United States Patent 3,338,786;
(2) antibiotic AV290 sulfate which is disclosed in
United States Patent 3,855,410;
(3) an antibiotic AV290-syntan complex prepared as
described in United States Patent 3,832,462;
(4) an antibiotic AV290-alkyl sulfate complex derived
by treatment of the antibiotic with an alkali metal alkyl
sulfate as set forth in United States Patent 3,856,937;
(5) an antibiotic AV290 alkylated derivative derived
by treatment of the antibiotic with a lower alkyl halide as
.'', ~
fiS~a~)
defined and described in United States Patent 3,954,973.
The invention also rela-tes to a stable premix com
position for the treatment and prevention of ketosis in
ruminants comprising a feed premix containing from about 1% to
about 90% by weight of an active ingredient selected from the
group consisting of antibiotic AV290, antibiotic AV290 sulfate,
an antibiotic AV290-syntan complex, and antibiotic AV290-alkyl
sulfate complex, an antibiotic AV290 alkylated derivative and
mixtures thereof in any proportion.
The major nutritive portion of the ruminants diet
consists primarily of polysaccharides such as cellulose and
starches. These primary sources of said animals energy
requirements~ and of basic intermediates for tissue building
processes, are hydrolized in said animals rumen by microbio-
logical processes to monosaccharides, primarily to glucose.
The thus formed glucose is then further degraded by enzymatic
processes to pyruvic acid and derivatives thereof. These in
turn are further converted through various enzymatic processes
to acetic acid and propionic acid and derivatives thereof.
Simultaneously, and also in the rumen, some of the acetic acid
is converted to butyric acid. Although butyric acid is the
component most efficiently metabolized by ruminants while acetic
acid is the least efficiently utilized product of the above
referred-to digestive processes, the formation of butyric acid
in the course of said microbiological processes is energetically
not very favorable. As stated above, butyric acid is formed
from acetic acid, and since one mole of glucose yields two moles
of acetic acid and two moles of methane and/or carbon dioxide
gas, and since two moles of acetic acid are consumed in the for-
mation of one mole of butyric acid, therefore, for every mole ofbutyric acid formed, two moles of methane and/or carbon dioxide
gas are generated, representing a considerable energy loss.
Additionally, it is known that production of acetic acid (and/or
acetates) in larger than normal amounts may lead to the produc-
tion of ketone bodies (acetoacetate, acetone and ~-hydroxy
butyrate) which can cause ketosis in ruminants, especially if
said animals are under stress. It has been found that the
supplementary feeding of propionic acid (and/or propionates) is
beneficial for minimizing the effects of ketosis. A further
advantage of feeding a diet high in propionic acid to ruminants
such as cattle, sheep and goats is to lower the incidence of
ketosis.
Thus it would be of advantage both to the animal grower
and feedlot operator if the microbiological processes of the
rumen could be altered so that the production of propionic acid
from carbohydrates is enhanced while that of acetic acid is
suppressed.
The novel method of the present invention comprises
orally administering to ruminants such as cattle, sheep and
goats antibiotic AV290 or the above-identified salts, complexes,
and derivatives thereof in amounts sufficient to promote the
formation of propionates while suppressing the formation of
acetates.
Advantageously, we have found that by orally administer-
ing to said ruminants antibiotic AV290 or the above-identified
salts, complexes, or derivatives thereof (and/or mixtures thereof)
in amounts of from about 10 mg. to about one gram per head per
day or from about 0.05 mg. to about 50.0 mg./kg. of body weight
per day, the acetate-propionate ratio is altered in favor of the
propionates in the course of the microbiological digestive
processes taking place in the rumen of said animals. Thus, by
promoting the formation of propionates and suppressing the
formation of aceta-tes, the incidence of ketosis among said
.
rUmlnantS lS mlnlmlZed.
~Y'`'),
ii2~
1rrhe active compounds of this invention, an-tibiotic
AV290 and the salts, complexes, and derivatives thereof, can
be conveniently adminis-tered to ca-t-tle, s~leep and goats as
feed addltives in amounts ranging from about 8 grams to about
one kilogram per ton of feed to provide 10 mg. to one gram
per head per day or 0.05 mg. to 50.0 mg./kg. of body weight
per day of said compound. Should it be desired, antibiotic
AV290 and its pharmaceutically acceptable salts, complexes,
and derivatives may also be formulated for oral administra-
tion as feed premixes~ concentrates, tablets, pills and bol-
uses, which may be prepared by accepted and well known meth-
ods, using pharmaceutically acceptable carriers, diluents,
binding and lubricating agents.
The present invention is further illustrated by
the set forth below:
Example 1
Evaluation of the in vitro efficacy of the antibiotic AV2~0
for altering the acetic acid:propionic acid ration during
fermentation in rumen fluid~
.
The steers used in the e~periment have surgically
installed fistulas that open into the rumen. The steers are
offered water and a daily ration of alfalfa hay ad libitum
as well as 10 lbs. of a standard 15% protein grain ration
of the following composition:
25 Component Percent by Weight
Corn32.7
Barley10.0
Oats 7 5
Molasses 10.0
30 Soybean Oil Meal (48~ protein) 13.8
52~
1 seet Pulp 2.5
Corn Gluten Feed 12.5
Distillers Grain 7.5
Trace Mineral Mix 0.05
5 Salt 1.0
Dicalcium Phosphate 2.0
Method
A sample of rumen fluid (500 ml.) is aspirated
into a flask and strained -through four layers of cheesecloth
to remove coarse feed particles. Ten ml. of the clarified
rumen fluid is introduced into an incubation flask contain-
ing 200 mg. of a substra-te of the following composition:
Component Percent by Weight
Corn Starch 68
Soybean Oil Meal 15
Alpha Cellulose 17
The drug to be tested ~AV290) is mixed into this
feed substrate at levels of from 40 to 5,000 ppm. These
levels are equivalen-t to 8 to 100 ug of drug per flask or
0.4 to 50 ug per ml. of final incubation fluid. To the
above mixture of drug/feed substrate and rumen rluid, 10 ml.
of a buffer of the following composition is added:
Component grams/liter
Sodium bicarbonate 19.6
Disodium hydrogen phosphate
heptahydrate 14.0
Potassium chloride 1.14
Sodium chloride 0.94
Magnesium sulfate
heptahydrate 0.24
2~
l Calcium chlori~e 0.08
The pH of the thus prepared fermentation mixture
is 7.2. Nex-t, each flask is flushed with carbon dioxide
to produce an anaerobic atmosphere and the fermentation mix-
tures are incubated for 24 hours in a 37C. constant temper-
ature bath while the flasks are shaken. Simultaneously, un-
treated control samples are also incubated~ Additionally
several samples are taken at the start of the incubation per-
iod and acidified with 6N hydrochloric acid to pH 2.0 to serve
as zero-time control samples. After 24 hours, the fermenta-
tion in the treated and control samples is stopped by the
addition of enough 6N hydrochloric acid to lower the pH of
the fermentation mixtures to 2Ø A portion of each sample
is then centrifuged at 2,000 x g for 15 minutes and the clear
supernatant removed for analysis by gas chromatography to
determine concentrations of acetic and propionic acid, and
butyric acid in the samples. The data obtained are summar-
ized in Table I and show the molar percent of each volatile
fatty acid (VFA) produced during fermentation in the presence
of antibiotic AV290 as well as the ratio, acetic to propionic
acid; while in Table II the same data are expressed as per-
cent change from the control.
~l l - - - -
O u ~ ~ ,-~ ~ ~ ~ ~r ~ ~ I_ o
,~ -~ O ~r oo ~D Cl~ CO ~ I_ ~ CO ~
~l . . . ~ ~ . . . ~ .
~; a) R (`1 ~ ~ ~1 ~1 ~I ~ ~1 ~ ~`I
~¢~
_ _ ___ _ __ _ __ _ _
_ .
,1 r- ~ Ln ~ ~r ,~ ~r ~D 0
S~ . . . . . . . . .
~ ~O ~D ~ ~ ~ O~) ~ ~ ~ 00
:~ ~1 r-l ~1 ~ ~ ~1 r-l r~l ~ ,_J
~ _ _ _ _ _ _
O
J~ O
~r co ~ ~r ~D ~ ~ ~1 ~ 00
~ O . . , . . . . . . .
O ~1 ~ a. ~ Cl~ O Lr) ~ ~ O ~r
)-I 0~ ~`1 ~ t~ ~ ~ ~ ~ ~ ~ ~
H 3
~ _ _ _ _ _ _ _ I
E~ o
.~ a~ ~n ~ ~ o ~D ~ ~ 00 ~1
~ 00 ~ U) ~D ~D ~D u~ U~ ~D ~
Y _ u~ _ ~ 11'~ 4~ u~ u~ A __
.~
~:1) Q
h .
. a ~ O O O O O O O O O O
~ O O O O ~-:Y O O O ~
0~ O O t~ ~1 Ll~ O ~J
_. _ __ _ _ _. _ __
O O ~ O ~
__E~ O ~ ~ ~ I
-- 7 --
;i2~
.. _ _ _ _ __ _ _ ___
h O
~ h ~ ~1 fr~ ~ ~ ~ ~r ~D
5:: ~ ~ ~ ~ o r- 1_ ~D t~l
U
~ l l l l l l
~U~
__ _ __ _ _ _ _ _ _
.~ ~r ~ o ~r ~ ~D ~ ~r
~ ~ ~ ~ ~o a~ 03 ~ oo
o m ~ ~ ,~ + ~ ~ ~ +
~o U _ _ _ __ _
U o .~,
~ ~ ~ n ~ I_ u~ ~ ~D
~ o . . . . . . . .
O ~ ~ ~ ~D O ~ 00 ~` ~r ~1
H ~1 ~: P. ~ ~ ~ + ~`I (`~ ~ +
H 4_1 C O ~ + + + +
R U P~
E~ ~ ~ _ _~ _ _ __ _ _
O
.~ u~ r~) t~ a~ ~ ~1 ~D ,_
a) CO ~D ~r ~ ~D ~D ~) ~
Q) V l l l l I
¢
a) O O O o o o o o
O O O ~ O O O er
O O~ O ~ O O ~
,_~U~ _ _ _ _ _ _
~::
O ~ 0
a~
~ ~ 4
a
___~ _~ ~ ..__ _ _
-- 8 --
1 The above data clearly show that antibiotic AV290 prepara-
tions are effective in increasing the propionic acid produc-
tion while simultaneously suppressing ace-tic acid production
during fermentation in rumen fluid.
Example 2
Evaluation of the in vivo efficacy of antibiotic AV290 for
.
altering the acetic acid:propionic acid ratio during fermen-
tation in the rumen of sheep.
.Twenty-four whether lambs, weighing approximately
40 kg. each, are randomly allotted to four groups of 6 lambs
each. One group serves as unmedicated control and receives
water and the following diet ad libitum:
Component Percent by Weight
Ground corn cob 35.0
Dehydrated alfalfa meal 20~0
Ground corn lg.4
Cane molasses 12.0
Soybean oil meal (48% protein) 12.0
Dicalcium phosphate 1.0
Iodized salt 0.5
Trace mineral mix 0.1
Vitamin A and D3 premix 0.02
Corn oil 0.01
The other three groups receive water and the same
diet ad libitum, except that 20, 50 or 100 ppm. of AV290
-
lauryl sulfate is incorporated into the diet of the respec-
tive groups. After 19 days on the diet, the sheep are sac-
rificed and samples of rumen fluid are obtained and analyzed
as in Example 1. The data obtained are summarized in Table
III and show the molar percent of each volatile fatty acid
1 (VFA) produced in the presence of AV290 as well as the ratio
of acetic to propionic acid; while in Table IV the same data
are expressed as percent change from the control.
-- 10 --
6~
~0~'~ O In~'~ __ _. 0~'~--O _
r~O ~ CO ~ CO r~rlO . . .
O ~ ~. ~ . . ~ ~ ~1 In r~
r1 aJ Q ~ ~1 ~ ~1 t~ ~ ~4 ~ ':r u~
O p:; t) O
; Q~ ~ ~ Q
!
_ _ _ _ ~o _ _ _
. ~ Lr) ~ a~ ~ . co ~o a~
u~ ~ O cn
m ~ ~1 ~1 ~ ~ ~ ~ ~
h
~ _ _ _ O~ _ _
a) ~
h 5:~
P~ .~ O ~
h O 1~ 1~ a~ o ~ O t~l ~) co
1~ ~ U~ ~I ~D ~ a) .,~ ~ ,-i
,~ ~ ~i ~`I ~ ~ C) Q ~r I~ o
H ¦ O O H ~ O ~ +
(U _ _ _ _ a) _ _ _
O O
.~ _l co r- ~1 .~ ~I ~ a~
~ al ~ ~ 0 Q~ ~D ~ U~
. ~ ~, ~o ~ U~ ~ l l l
t ~!
o o o o ~.~ o o o
o ~a ~, , o O ~ N n o
~1~ ~
~ _ _ _ ~--1 _ _
. ~ ~ o~ ~ ~ .
~ ~3 1~ ~1 0 N ~
) 1 O ~ ) h ~ 1
_ U ~ E~ ~
5~
Th~ above data, especicllly those found in Table IV,
clearly show that AV290 lauryl sulfate is very effective in
increasing the propionic acid production while suppressing
the acetic acid production in vivo, when orally administered
to sheep.
Example 3
Evaluation of the efficacy of AV290 for altering the acetic
acid:propionic acid ratio during fermentation in the rumen
,
of cattle.
Eighteen feedlot cattle, weighing approximately
350 kg. each, are randomly allotted to three groups of 6
animals each. Water and the ration described in Example 2
are offered to the animals ad libitum. One group serves as
unmedicated control, while groups two and three each receive
lS in their daily diet 33 ppm. or 99 ppm. of AV290 lauryl sulfate,
respectively.
The feeding trial is conducted for a period of
56 days, with rumen fluid samples being obtained from the
animals on days 14, 28 and 56 aft~r the beginning of the
2d trial. The rumen fluid samples are handled and analyzed
for volatile fatty acids (VFA) by the method of Example l.
The thus obtained data are summarized in Table V, while in
Table VI the same data are expressed as percent change from
the controls.
5Z~
\,~; _ _ _ _ _ ._ _ o _ oo
~ O a~ r~ ~ a~ ~ ~o ~ o o
o ~-~ . . . . . . ~ . .
~ O ~ ~ ~ ~ ~ ~ ~ f~ ~
~; ~'C h
rl ~r a) o~ ~ t~ N cr r-l cn
m ,~ r-l ~ r-l r l r l O~ r-l
~: _ __ _ _ _ _ __
h O ~ ~D ~ o u~ ~ ~ o~ ~
tr~ rl r-- ) ~ ~ N ~) 00 co ~!0
r l O r l r-l r l ~1 t~l N ~1 ~1 N
_ _ _ _ _ __ _
rl ~ ~D 00 t-- 00 u) ~ ~ cD
(I) ~ a~ u) u~ ~ ~_i ~i O oo
~ ~ ~ U~ ~D ~D ~D ~ ~D ~S~ n
~1 _ _ __ _ _ __ __ _
~J O r-l C10 ~ r-l ~ u~ r-l ~ u~
~ ~1
__ __ _ _ _ _ _
~ Q
4~ .~ o ~ a~
~ ______
3 r-l Orl ~)
~ N ~ ~
O ~ ~ ~
-- 13 --
~2~1
_ ... __ _ _ _ _
~o
~ a~ o co I~ ~ ~
o ~ o . . . . . .
.~ ~ r1 ~r ~ ~ ~r ~r co
~ ~U ~ r~ ~I N ~ ~r r7
~ ~ o I l l l l l
U7 _ _ _ _
~ t~
h r~l ~ 1-- ~ r~ oo
O >1 rx~ 1~ r.~l U~ U) r~
+ + + ~ +
_ _ _ __
,o
o u: ~ ~r c~
U O . . . . . .
~ ~D ~1 a~ r~3 u~ U~
4 N r~J -1 ~9 It~
~ O ~ + f ~ + +
h
_ _
H ~) r l Cl~ r~) Ir~ 11 ) ~
~> ~1 Ll'~ ~D ~ O r~l O
~ _ ~ l l _1 ~1 ~~1 ~
tl~ ~
~ ~J
~ u~ ~r ro ~D ~r r~o ~D
~8 1 r~l ~ ~ ~ u~
n ~
_ _
o ~ r~ rJ~
r) a~
.,~
_
a)
~ o ~
rd r~
~ ~ ~1
E~ ~ ~d
... _ .... . . . ~
-- 14 --
It can be clearly seen from Table V and especially
from Table Vl that the oral adrninistration of AV290 lauryl
sulfate to cattle via their diet very effectively alters the
VFA production during fe~nentation in the r~nen of said
animals in favor of propionic acid.
- 15 -
