The Professional Animal Scientist 30 (2014):150–179

©2014 American Registry of Professional Animal Scientists

InutritionR of ruminants—Current
NVITED : Applied protein
EVIEW

status and future directions1

F. N. Owens,*2 PAS, S. Qi,* and D. A. Sapienza†
*DuPont Pioneer, Johnston, IA 50131; and †Sapienza Analytica, Slater, IA 50244

ABSTRACT
The metabolizable protein (MP) sys-
tem initially outlined by Burroughs et
al. (1975) and expanded in later NRC
publications separates the need for N for
ruminal microbes within the rumen from
the postruminal need for amino acids
for growth and maintenance of the host
ruminant. Compared with the CP sys-
tem, the MP system represents a clearer
understanding of the complexity of the
protein metabolism of ruminants. Using
compiled data sets from recent publica-
tions, the effect of dietary CP concentra-
tion on performance of feedlot and dairy
cattle was reevaluated. Maximum per-
formance (rate of gain; milk production)
required higher CP concentrations than
routinely are being fed, with the added
performance being due at least partially
to greater DMI of diets containing more
CP. Precision of the NRC (2000) MP
model 1 was evaluated by comparing its
predicted values with measurements of
duodenal flow determined with growing-
finishing cattle fed 118 different concen-
trate-rich diets. Though duodenal supply
1
Presented at the American Registry of
Professional Animal Scientists (ARPAS)
Symposium: Applied Nutrition of
Ruminants—Current Status and Future
Directions, July 10, 3013, ADSA/ASAS Joint
Annual Meeting, Indianapolis, Indiana.
2
Corrresponding author: fred.owens@
pioneer.com
of microbial protein increased with
intake of TDN, diet TDN concentration
was not correlated (R2 = 0.00) with the
extent of OM fermented in the rumen.
Although undegraded intake protein
and MP increased with DMI, precision
of predicting degraded intake protein,
undegraded intake protein, and MP per
kilogram of diet was poor, reflecting im-
precision of estimates or equations and
failure of prediction equations to match
in vivo measurements. The paucity of
data supporting values suggested for
individual feeds and inclusion of numer-
ous theoretical but unverified equations
within current MP models severely limits
their precision and usefulness for field
application.
Key words: protein, microbial pro-
tein synthesis, rumen-escape protein,
requirement, microbiome
INTRODUCTION
For decades nutritionists have
formulated diets simply to maintain a
minimum CP (N × 6.25) concentra-
tion in DM. Even today, ruminant
nutritionists and producers cringe if
they notice that the CP concentra-
tion of a formulated diet falls below
the mean concentration commonly fed
in the industry (e.g., 13% of DM for
feedlot cattle or 17% of diet DM for
lactating dairy cows). The marked di-
vergence in dietary protein concentra-
tions of diets routinely fed to feedlot
cattle versus lactating dairy cows
illustrates that despite the physiologi-
cal similarity in digestive function of
all ruminants, dietary requirements
differ among classes due largely to
differences in composition and yield of
products.
For nonruminants, diet formula-
tion involves combining finely ground
maize grain with soybean meal and
adding a dab of minerals, vitamins,
and perhaps an amino acid or 2
depending on estimated prececal
availability of amino acids from these
2 feedstuffs. In contrast, ruminant
nutritionists must contend with a very
wide diversity of feed ingredients and
supplements that differ in composi-
tion. This large number of feed ingre-
dients when combined with processing
methods that alter the site and extent
of digestion provides ruminant nu-
tritionists and consultants an almost
endless array of components to tweak
when attempting to improve produc-
tivity or efficiency, a task that helps
ensure long-term job security.
The goal of this paper was to ap-
praise protein requirements both from
a traditional CP and one metaboliz-
able protein (MP) approach (NRC,
2000) by comparing measured with
predicted quantities of microbial pro-
tein, degraded intake protein (DIP),
undegraded intake protein (UIP),
and MP and to discuss several aspects

of amino acid nutrition gleaned from
trials with nonruminants. No live-
stock were used or harmed during
preparation of this treatise, although
the senior author intermittently was
stressed by the panel chair regard-
ing tardiness in preparation of this
manuscript.
MATERIALS AND METHODS
For examining the relationship
of performance to dietary protein
concentration and the ability of the
NRC (2000) MP system to predict
the duodenal flow of microbial and of
dietary protein that escaped ruminal
digestion, 3 data sets were assembled.
The first consisted of results from
19 recent publications with growing-
finishing cattle where 43 specific
diets were supplemented with vari-
ous amounts of CP for a total of 155
diet-protein comparisons. The second
included similar data from recently
published trials with lactating cows
fed 25 specific diets from 13 publica-
tions for a total of 75 diet-protein
combinations. Data from these 2 sets
were analyzed by regressing perfor-
mance measurements against dietary
CP concentration and testing for
significance of linear, quadratic, and
cubic responses to CP within diet by
including diet as a fixed factor. When
significant responses were detected,
the intercept of the regression line
was adjusted so that the plotted re-
gression line passed through the mean
production rate at the mean protein
concentration. Regression lines are
plotted together with absolute values
reported from individual studies. The
third data set consisted of 37 different
basal diets from 21 publications for a
total of 118 different diet-supplement
combinations where duodenal flow
by cattle had been measured so that
extent of ruminal OM digestion and
duodenal flow of microbial protein
and undigested dietary protein were
known based on digestion and micro-
bial markers. Mean values and ranges
within this data set are illustrated in
Table 1. Measured flows were used
to calculate the supply of microbial
CP. Values for microbial CP supply
Protein nutrition of ruminants
(MPS), UIP, DIP, and MP predicted
from TDN, UIP, and DIP tabular
estimates for the compiled diets
from model 1 of NRC (2000) were
compared with the MPS, UIP, DIP,
and MP measurements within these
experiments. The relationship of the
predicted to observed values was ap-
praised by linear regression; indi-
vidual values and the linear regression
line relating predicted with observed
values are plotted and statistical esti-
mates are provided.
Classical CP Requirements
Diets for domestic animals often are
formulated to provide an amount of
each required dietary nutrient that
will meet or exceed the minimum
requirement for that nutrient. Mini-
mum requirements in turn have been
based on avoiding either specific signs
or symptoms characteristic of a nutri-
ent deficiency or consistent reductions
in productivity. Dietary requirements
traditionally have been appraised by
supplying various amounts of the test
nutrient and determining the mini-
mum quantity needed to achieve a
maximum or a plateau in rate of gain
or efficiency of production. Although
in vivo performance measurements
have served as the gold standard for
quantifying requirements, performance
responses can prove misleading.
Increasing the supply of a nutrient
in the diet may increase performance
not only through meeting a deficiency
for that nutrient but also indirectly
through increasing DMI or digestibil-
ity of energy. Similarly, substituting
one ingredient for another may alter
DMI or digestibility, and supplemen-
tal nutrients can have overlooked or
unrecognized physiological effects.
Generally, a surplus amount (an
allowance) of a nutrient is included
in commercially formulated feeds to
compensate for variability in nutrient
content of individual diet ingredients
and imprecision in diet compilation as
well as to ensure that nutrient supply
for all animals within a group is ad-
equate despite differences among the
individual animals in DMI, weight,
nutritional history, and genetic merit.
151
Efficiency of use of land, water, or
feed resources is of widespread con-
cern, yet sustainability of an individ-
ual business or enterprise relies on an
economic return on a current or short-
term basis. Maximum rate or efficien-
cy of production is not always con-
gruent with least cost performance,
with maximum longevity of animals,
with efficiency of nutrient use, or with
minimizing adverse environmental
effects. Certain least-cost sources of
energy may contain excesses of certain
nutrients (e.g., protein and phospho-
rus in distillers products). In con-
trast, when considering economics of
production, some degree of deficiency
and sacrifice in animal performance
can be justified when no economic
benefit or only a limited performance
response from supplementing a mar-
ginally deficient nutrient is apparent.
Though diets could be customized for
specific groups of animals and altered
with stage of production, logisti-
cal difficulties in properly compiling
and delivering diets formulated for
separate animal groups often proves
cumbersome. Consequently, numerous
factors beyond minimum requirements
for nutrients as defined experimental-
ly will continue to dictate the nutrient
content of diets formulated by skilled
nutrition consultants.
Whether diets are being formulated
to meet minimum requirements by
the traditional CP system or through
using an MP model, the basic prem-
ise behind a requirement is that the
needs of an animal for maintenance,
production, and health should be met.
Consequently, both systems must
consider the deposition, secretion,
and inevitable losses of protein from
the body. Protein expenditures differ
markedly between feedlot cattle and
lactating dairy cows, so these 2 classes
will be discussed separately.
CP for Growing-Finishing
Cattle. The amounts of protein
deposited or lost daily by typical
growing-finishing feedlot cattle at
various shrunk BW are illustrated in
Figure 1. Because expenditures are
inevitable, these losses of N must be
replaced by either protein or NPN
provided in the diet.
152 Owens et al.

Table 1. Statistics for measurements from individual diets from summarized trials and values predicted from
various sources

Variable1 n Mean SD Minimum Maximum

Cattle weight 118 305.8 94.5 106.0 493.0
Grain, % of diet DM 118 72.3 7.4 41.9 85.0
DMI, kg/d 118 5.74 1.38 2.73 10.76
DMI, % BW/d 118 1.95 0.31 1.18 2.58
Concentrate intake, kg/d 118 4.92 1.22 2.33 9.44
Forage intake, kg/d 118 0.63 0.24 0.00 1.39
TDN, % of DM (NRC, 2000) 118 83.87 3.34 75.81 89.37
CP, % of DM 118 12.67 1.53 10.29 19.23
NDF, % of DM (NRC, 2000) 118 16.44 4.65 11.36 36.89
DIP, % of DM (NRC, 2000) 118 8.24 1.41 5.91 11.98
UIP, % of DM (NRC, 2000) 118 4.43 0.97 1.89 8.41
DIP, % of CP (NRC, 2000) 118 64.9 6.6 49.3 85.6
UIP, % of CP (NRC, 2000) 118 35.1 6.6 14.4 50.7
Diet ME, Mcal/kg of DM (NRC, 2000) 118 3.03 0.12 2.74 3.22
Diet NEm, Mcal/kg of DM (NRC, 2000) 118 2.05 0.10 1.81 2.21
Diet NEg, Mcal/kg of DM (NRC, 2000) 118 1.39 0.09 1.18 1.53
OM intake, g/d 118 5,435 1,323 2,541 10,222
Duodenal OM flow, g/d 118 2,786 807 1,274 6,102
Fecal OM output, g/d 118 1,045 350 442 2,515
OM digested in rumen, % of OM intake 118 48.6 7.5 26.2 61.7
OM digested in total tract, % of OM intake 118 80.7 4.4 66.9 88.1
Starch intake, g/d 118 2,813 1,033 1,221 6,440
Duodenal starch flow, g/d 118 506 387 83 1,840
Fecal starch output, g/d 118 74 108 3 721
Ruminal starch disappearance, % of starch intake 118 81.9 11.0 44.5 94.3
Total-tract starch digestion, % of starch intake 118 97.7 3.4 79.4 99.8
NDF intake, g/d 32 830 247 560 1,360
Duodenal NDF flow, g/d 32 552 177 335 1,080
Fecal NDF output, g/d 32 470 144 291 940
NDF digested in rumen, % of NDF intake 32 32.7 11.6 13.0 53.6
NDF digested in total tract, % of NDF intake 32 42.9 7.4 25.0 59.2
CP intake, g/d 118 732.6 217.7 333.1 1,471.3
Duodenal CP flow, g/d 118 754.0 205.6 395.6 1,273.1
Microbial CP flow to duodenum, g/d 118 440.3 127.3 231.9 872.5
Microbial efficiency, g of microbial protein/ruminally truly fermented OM × 100 105 22.2 4.2 10.0 33.9
Microbial efficiency, g of microbial protein/ruminally fermented OM × 100 118 17.2 4.2 7.4 29.5
Duodenal flow of nonmicrobial CP, g/d 118 313.7 133.4 50.0 680.6
Total-tract CP digested, % of CP intake 115 71.5 4.5 56.9 81.6
Ruminal pH 95 5.93 0.25 5.35 6.54
Ruminal acetate, % of VFA 101 52.0 6.4 39.3 67.0
Ruminal propionate, % of VFA 101 33.4 7.7 14.2 51.6
Ruminal butyrate, % of VFA 101 11.4 2.0 6.7 16.9
Metabolizable protein supply, g/d (calculated from duodenal CP flow) 118 533 149 276 924
Values predicted by various models of ruminal protein metabolism          
DIP intake, g/d (NRC, 2000) 118 475 146 208 917
Microbial CP flow to duodenum, g/d (NRC, 2000) 118 557 128 255 966
Microbial CP flow to duodenum, g/d (NRC, 2001) 118 600 185 175 1,274
Microbial CP flow to duodenum, g/d (Burroughs et al., 1975) 118 625 146 290 1,156
Microbial CP flow to duodenum, g/d (Valadares Filho et al., 2010) 118 579 135 269 1,071
Microbial CP flow to duodenum, g/d (Clark et al., 1992) 118 636 121 370 1,076
Microbial CP flow to duodenum, g/d (NRC, 1985, for nonlactating cattle) 118 565 192 199 1,332
UIP, g/d (NRC, 2000) 118 258 93 90 554
Metabolizable protein supply, g/d (NRC, 2000) 118 563 148 263 1,062
1
DIP = degraded intake protein; UIP = undegraded intake protein.

Figure 1. Estimated daily CP expenditures and maintenance requirements of growing-
finishing feedlot cattle at various empty BW. Color version available in the online PDF.
Within this figure, estimates of rate
of gain, DMI, protein composition of
gain, scurf loss, and metabolic fecal
loss were derived or calculated from
NRC (2000) performance estimates.
Endogenous urinary CP was added
to the requirement list based on the
quantity of nonurea N excreted in
urine by lactating cows fed diets
across a range of protein contents
(Colmenero and Broderick, 2006)
adjusted for the difference in DMI
between growing and lactating cattle.
These inevitable urinary N losses
would include excess microbial nucleic
acids, allantoin, uric acid, creatinine,
and possibly some glutamine; these
should be proportional to the amount
of microbial protein synthesized in the
rumen, to release of nitrogenous com-
pounds during tissue turnover, and
to acid–base balance of the animal.
Nucleic acids synthesized by rumi-
nal microbes are not fully excreted
because certain nucleic acids are
degraded to N that can be recycled to
the rumen as urea.
Retained CP decreases from 30 to
15% of total expenditures as cattle
increase in weight because deposited
lipid comprises a larger portion of
Protein nutrition of ruminants
weight gained. Relating protein gain
to daily energy deposition at various
rates of gain based on the equation
employed by NRC (2000) matched
measured values for body protein
mass of cattle at various weights
(NRC, 2000) very closely; this sup-
ports the concept that equations re-
lating protein gain to energy gain by
growing-finishing cattle is valid. Meta-
bolic fecal CP and endogenous uri-
nary CP, the primary components of
maintenance, commonly are presumed
to increase in proportion to DMI,
whereas scurf CP is considered to be
proportional to surface area (NRC,
2000). Nevertheless, the estimate of N
required for maintenance employed by
NRC (2000) shown in Figure 1 is cal-
culated from metabolic body size. The
sum of inevitable fecal, urinary, and
scurf CP losses exceeded the NRC
(2000) estimated CP requirement for
maintenance of 440-kg cattle by only
4%, indicating that these independent
estimates agree very closely. When
related to the expected DMI for a diet
rich in concentrate, the total of these
CP expenditures decreases from 6.4
to 5.2% of DMI as empty BW in-
creases, a decrease of 19% associated
153
with maturation. Hence, the need for
dietary protein should decrease with
maturation unless efficiency of CP
conversion to these expenditures also
decreases. Nevertheless, these expen-
ditures still total less than 50% of the
protein concentration of diets typi-
cally fed to finishing cattle, reflecting
an inefficiency of converting dietary
N to deposited protein and to replace
these inevitable losses by growing
ruminants.
Administration of certain hormones
(e.g., androgens) or metabolically ac-
tive compounds (e.g., β-agonists) can
improve G:F markedly. Compounds
that reduce the rate of protein turn-
over or degradation by animals should
reduce those N losses and the energy
requirements associated with protein
turnover (endogenous urinary N and
perhaps metabolic fecal N). Admin-
istration of an androgen reduced
the loss of BW by cattle maintained
under drought conditions (Houston
et al., 1992); androgens might prove
useful to reduce both protein and en-
ergy requirements of ruminants being
maintained at a constant weight. Dur-
ing growth, compounds that increase
protein retention by reducing protein
turnover and associated losses of N by
the body should enhance efficiency of
N and energy use without increasing
the dietary requirement for N.
In contrast, estrogenic compounds
that increase mature weight of rumi-
nants would be expected to increase
the grams of protein required daily at
a given BW and rate of gain. How-
ever, estrogenic implants typically
increase DMI as well and should
increase MPS; this could dampen or
cancel their effect on dietary protein
requirements. Based on the review
of DiCostanzo and Zehnder (1999),
the MP requirement for steers not
implanted or implanted with a moder-
ate or an aggressive implant is 14.1,
9.1, and 11.8 g/kg of metabolic size,
respectively. Those estimates can be
compared with expenditures noted in
Figure 2. Assuming that cattle from
which these expenditures were derived
had received moderate implants, total
protein expended daily ranged from
51 to 63% of the estimated metabo-
154
Figure 2. Comparison of daily CP expenditures with predicted need for metabolizable
protein (MP) for growing-finishing feedlot cattle at various empty BW. EUN-CP =
endogenous urinary CP. Color version available in the online PDF.
lizable protein requirement. This ef-
ficiency with which dietary CP or MP
can replace expenditures would not be
expected to be a constant due to dif-
ferences in the potential for NPN to
replace certain losses and differences
in the amino acid composition of MP
relative to the amino acids ratios
present in tissue or in N being lost by
ruminants.
Performance responses to concentra-
tions of dietary CP were estimated
from the compiled data set from 19
published trials involving 43 diets
with feedlot cattle where the supply
of protein was altered so that re-
sponse within a diet and a specific set
of cattle of similar genetic and envi-
ronmental history could be evaluated
and merged across diets. Performance
within specific time periods was sepa-
rated when interim information was
provided. Based on regression within
these studies, rate of gain increased
linearly (P < 0.01) and quadratically
(P < 0.04) as concentration of protein
in the diet increased (Figure 3). Based
on this data set, ADG should reach a
peak at 15.2% dietary CP. Although
Owens et al.
ADG peaked at a CP concentration
higher than typically fed to growing-
finishing feedlot cattle, the expected
ADG at 13% CP remained at 98% of
peak ADG.
As has been observed often, DMI
also tended to increase linearly (P =
0.08) with dietary CP concentration
(Figure 4) with no indication of a
plateau. Consequently, the increased
ADG associated with higher dietary
CP concentrations may be attribut-
able partially to an increased intake
of energy.
Can the increase in DMI fully
explain the increase in rate of gain
noted with addition of CP to the
diet? Should CP requirements be
based on maximum rate of gain, G:F,
or efficiency of energy use? The rela-
tionship of G:F to dietary CP within
these same experiments is shown in
Figure 5.
The G:F increased linearly (P =
0.02) with concentration of CP in the
diet. These increases in gain and G:F
should be of direct economic interest
to livestock producers. However, some
of this increase in G:F still could be
explained by increased dilution of
the maintenance energy requirement
associated with higher DMI. Conse-
quently, the relationship of observed
ME of the diet, based on DMI, rate
of gain, and mean BW, to the ME of
content of the diet calculated from
ME of the individual diet ingredients
was determined. Results are shown in
Figure 6.
This ratio of observed to calculated
ME, though surprisingly variable
among studies, was not significantly
altered by diet CP, indicating that
dilution of maintenance theoretically
alone might explain the increased G:F
observed with diets containing more
CP. Alternatively, the additional CP
may be providing some other dietary
ingredient (e.g., a macro or trace
mineral, a ruminal or blood buffer, a
nutrient that alters ruminal retention
time) that similarly could result in an
increased DMI.
Age and degree of maturity certain-
ly influence protein expenditures and
requirements (Figure 1). Crude pro-
tein concentrations above 13% of DM
often increase performance and feed
net energy values for cattle weighing
less than 300 kg, likely due to provid-
ing some limiting amino acid(s) as
discussed by Zinn et al. (2007). How-
ever, protein levels above 11% seldom
increase the rate or efficiency of gain
for feedlot cattle weighing more than
450 kg except possibly with cattle fed
diets composed of steam-flaked corn.
Although intakes of DM and energy
were reduced when the dietary pro-
tein concentrations decreased below
13% very late in a feeding trial where
steam-flaked corn was fed, N retention
was not increased when additional
urea was provided (Cole et al., 2006).
This indicates that the additional
urea may have acted through some
mechanism beyond simply meeting
a metabolic demand for N or amino
acids. Within the rumen, supplemen-
tal true protein can alter the sup-
ply of various N sources (ammonia,
peptides, certain amino acids, nucleic
acids) that in turn may alter micro-
bial population or its diversity as well
as the composition and efficiency of
growth of bacteria. Such compounds

Figure 3. Daily gain of growing-finishing feedlot cattle fed 36 different diets
supplemented with various amounts of CP from 16 different published articles. Color
version available in the online PDF.
may be either essential or stimulatory
nutrients for certain microbial species
based on pure culture studies. Urea
supplementation has been reported to
decrease the incidence of liver ab-
Figure 4. Daily DMI of growing-finishing feedlot cattle fed 36 different diets
supplemented with various amounts of CP from 16 different published articles. Color
version available in the online PDF.
Protein nutrition of ruminants
scesses (Zinn et al., 2003; Wagner et
al., 2010), perhaps because ruminal
ammonia serves as a transitory rumi-
nal buffer helping cattle cope with the
acid load from large meals of rapidly
155
fermented grains. Serving as a weak
base with a high pH, ammonia should
increase ruminal pH (Trenkle, 1979)
and thereby would enhance activity
of cellulolytic bacteria. For the host
animal, absorbed ammonia also can
alter blood base excess, provide the
host animal with added N for biosyn-
thesis of nonessential amino acids,
and through elevating blood urea,
enhance cycling of urea to the large
intestine. Increased entry of urea into
the large intestine, through increas-
ing pH at that site and in feces, may
increase extent of compensatory
digestion of OM in the large intestine.
Thereby, supplemental DIP might
alter not only the extent of ruminal
but also of postruminal digestion.
Low blood osmolality also can reduce
salivary flow, ruminal buffering, and
the liquid dilution rate of ruminal
contents. Behavior measurements also
indicate that urea supplementation
may decrease meal size and increase
meal frequency and thereby stabilize
ruminal digestion.
A concentration of plasma urea N
of finishing cattle below 5 to 8 mg/
dL was proposed by Johnson and
Preston (1995) to reflect inadequacy
of dietary N; plasma urea N values
above 12 mg/dL in contrast may
reflect an excess and waste of N.
Numerous factors can influence blood
and milk urea concentrations (Brod-
erick and Clayton, 1997; Hof et al.,
1997). Nevertheless, blood or milk
urea concentrations should mirror
metabolic protein status and prove
useful to detect excesses, a factor of
concern in waste management. If in
vitro systems and proposed increases
in microbial protein cannot read-
ily explain performance responses to
supplemental N, physiological altera-
tions beyond the rumen deserve close
scrutiny. In contrast, if amounts of N
from degraded DIP plus N recycled
to the rumen are insufficient to meet
DIP requirements, as reflected by low
ruminal ammonia and low blood urea
concentrations, and if performance
responses are greater from supple-
mented DIP than UIP, then correc-
tion of a DIP deficiency and improved
N status probably are responsible for
156
Figure 5. G:F of growing finishing cattle fed 36 different diets supplemented with
various amounts of CP. Color version available in the online PDF.
performance responses to additional
CP. Monitoring both N balance and
performance responses is necessary
to differentiate between direct and
Figure 6. Ratio of performance-calculated ME to diet ME content for growing-
finishing cattle fed 36 different diets supplemented with various amounts of CP. Color
version available in the online PDF.
Owens et al.
indirect performance responses to ad-
dition of N to the diet.
Ammonia requirements for micro-
bial growth were clearly illustrated
by trials with the continuous flow
fermentation system by Satter and
Slyter (1974). Ammonia-N concen-
trations above 5 mg/dL failed to
increase microbial protein yield with
either fibrous or concentrate feeds,
whereas concentrations below this
point, particularly below 2 mg/dL,
reduced microbial protein yield. These
values often are cited as benchmarks
to judge whether or not the supply
of ruminally degraded N or NPN is
adequate for bacterial fermentation
within the rumen. In sharp contrast
with these values, Bach et al. (2005)
found no correlation between effi-
ciency of microbial growth and am-
monia concentrations in feeding trials.
Unfortunately, ammonia N concentra-
tions fell below 5 mg/dL in only 5 of
their 285 studies, and the within-trial
responses to various ammonia concen-
trations were not quantified. Several
studies cited by Kertz (2010) in his
review of urea feeding indicated that
ammonia concentrations up to 24 mg/
dL increased nonammonia N flow or
in situ DM disappearance in several
studies. Postprandial changes in am-
monia concentrations and differences
in ammonia concentrations at vari-
ous sampling sites (e.g., usually lower
within the ruminal raft than in free
liquid) or within microenvironments
(lower concentrations internal to
plant cells or within a glycocalyx than
in free liquid) could be responsible
for the response to higher ammonia
concentrations. Difficulty in obtaining
rumen samples with animals that are
not equipped with ruminal cannulas
complicates the routine use of ruminal
ammonia concentration as a measure-
ment of ruminal status, but direct
measurements are needed if ruminal
ammonia is of concern. You cannot
manage what you do not measure.
Precisely why a low ruminal am-
monia concentration reduces micro-
bial yield has been debated. At low
ruminal ammonia concentrations,
ammonia uptake by ruminal microbes
appears dependent on glutamine
synthetase, an energy-requiring reac-
tion, whereas at higher ammonia
concentrations, its uptake by glutamic
acid dehydrogenase does not require

energy. Consequently, efficiency of
microbial growth per unit of OM
fermented should be slightly lower
when ruminal ammonia concentra-
tions are low. Furthermore, when
the supply of ammonia is inadequate
or conversely when energy supply is
excessive, ruminal bacteria store more
glycogen-like carbohydrate. Carbohy-
drate storage is inefficient for bacteria
considering that a substantial frac-
tion of the energy ultimately available
from metabolism of carbohydrate is
expended simply for importing simple
sugars. Other energy-spilling reactions
have been associated with ruminal N
deficiency or an energy excess (Russell
and Cook, 1995; Hackmann, 2014).
Nevertheless, research trials evaluat-
ing the need for ruminally degraded N
in vivo always should measure and re-
port ruminal ammonia concentrations
for comparison with in vitro estimates
of the ammonia requirement.
Phase feeding (e.g., reducing the
protein supply for more mature
feedlot cattle or lactating cows
later in lactation in parallel with
the reduced need for protein reten-
tion or secretion) should reduce feed
cost, improve efficiency of N use, and
reduce N waste without depressing
gain or efficiency (Martin et al., 1976;
Cole et al., 2006; Vasconcelos et al.,
2006; Zinn et al., 2007; Erickson and
Klopfenstein, 2010). By formulating 2
finishing feedlot rations, one high and
a second lower in protein, appropri-
ate mixtures of these 2 rations could
readily match the projected need for
dietary protein while yet allowing
dietary protein levels to be gradually
reduced for pens of feedlot cattle or
lactating cows throughout a given
time period. Currently, consultants
routinely target protein concentra-
tions for feedlot cattle at all stages of
finish at an average of 13.5% of DM
(Vasconcelos and Galyean, 2007).
Because microbial protein when com-
bined with escape protein from most
grain-rich diets should satisfy the MP
or postruminal amino acid needs for
heavier feedlot cattle, simply provid-
ing an adequate concentration of
ammonia or ammonia precursors from
NPN to maintain ruminal metabolism
Protein nutrition of ruminants
should be sufficient for maximum
rate and efficiency of gain for heavier
feedlot cattle. Although fluctuating
protein levels might be expected to
decrease ruminal stability and in-
crease the incidence of metabolic dis-
orders, studies with oscillating protein
levels to date have not detected any
adverse effects on animal performance
or health (Cole, 1999; Cole et al.,
2003; Archibeque et al., 2007).
The numerical linkage between
quantity of OM fermented and the
quantitative ammonia need for syn-
thesis of microbial protein also has
been extrapolated to apply chrono-
logically. This led some researchers to
suggest that rate of fermentation and
rate of ammonia release in the rumen
should be synchronized. Orchestrat-
ing for a slowed or attenuated release
of ammonia from NPN thereby was
proposed to improve or maintain
activity of those ruminal microbes
that ferment feed components slowly
(e.g., cellulose). Some in vitro studies
supported the concept that synchrony
between fermentation and ammonia
release rates can increase either the
quantity of microbial protein synthe-
sized or the extent of OM digestion.
In contrast, benefits from slow-release
compounds have never been apparent
in animal trials where both positive
(urea) and negative (unsupplemented)
diets have been included. In a sum-
mary of trials, Reynolds and Kris-
tensen (2008) found no evidence of a
performance benefit from synchroniz-
ing the rates of N and carbohydrate
fermentation. Failure of an in vivo
response to match that predicted in
vitro presumably reflects differences
in either physiology or metabolism
between these 2 systems. Such fac-
tors could include observations that
1) productive ruminants are fed or
voluntarily consume meals at frequent
intervals so that ruminal fermenta-
tion is semi-continuous and thereby
dissimilar to the single or intermit-
tent batch cultures often employed in
vitro, 2) the rumen has a large ballast
so that a single meal represents a rel-
atively small addition to total ruminal
contents, 3) some consumed feeds are
flushed from the rumen before being
157
fully mixed with rumen contents,
4) N in the form of urea is recycled
continuously to the rumen both in
saliva and by diffusion through the
ruminal wall, and 5) microbes may
adapt to asynchronous availability
of N. In studies with steers, Miz-
wicki et al. (1980) tested effects of
supplementing prairie hay (2.6% CP)
with urea. Added urea increased N
retention equally well whether the
urea was included within just one or
with each of the 24 meals throughout
the day, showing no benefit from a
simulated attenuated ammonia release
rate. Likewise, the benefit from a DIP
supplement over a negative control
diet proved to be equal when provided
as infrequently as every sixth day as
when provided daily for grazing beef
cows (Bohnert et al., 2002). How-
ever, they observed that when UIP
was supplemented every sixth day,
N balance was lower than when UIP
was supplemented every day or every
third day. Obviously, attenuated am-
monia release from NPN sources helps
avoid ammonia intoxication. However,
certain NPN compounds with attenu-
ated release such as biuret require an
extended adaptation time, whereas
other complexes or compounds are
not fully digested in the rumen but
simply are excreted or defecated
unused. To increase ruminal escape
of amino acids or other nutrients,
anthelmintics, or antibiotics, specific
payloads are coated with or imbed-
ded into spherical particles that resist
degradation within the rumen. Even
though ruminal escape has been the
primary objective for developing
such materials, release of the payload
within the small intestine for absorp-
tion often has limited the nutritional
value of such compounds. Both rumi-
nal escape and intestinal availability
must be achieved before a payload
can have value nutritionally for the
host ruminant.
Lactating Cows. Protein (N ×
6.25) losses that must be replaced
daily for lactating cows at various
points in a lactation curve as estimat-
ed from production rates, DMI, and
BW from production curves compiled
by NRC (2001) are graphed in Figure
158
7. Within this figure, milk protein
yield follows milk production, scurf
loss was calculated from metabolic
body size, fetal protein deposition
begins only late in lactation, and fecal
protein loss was calculated from DMI.
Endogenous urinary loss was calculat-
ed from nonurea N excretion in urine
by lactating cows fed diets differing in
protein content (Colmenero and Brod-
erick, 2006). Such urinary N losses
would include purines, uric acid,
allantoin, and creatinine that should
parallel microbial protein synthesis
and tissue turnover plus additional
glutamine lost during metabolic aci-
dosis. As a proportion of total protein
expenditures, milk CP accounted for
60% early in lactation but less than
40% late in lactation. The protein re-
quirement for maintenance employed
by NRC (2001), the highest of the 3
estimates of MP studied by Chizzotti
et al. (2008) per unit of metabolic
size, is below the estimate of metabol-
ic fecal loss alone. Although recycling
of N helps to conserve and exchange
N within various pools of the body, all
these expenditures except fetal growth
involve N losses by lactating cows
that cannot be reduced by N recycling
unless animals practice coprophagy.
Metabolic fecal loss is calculated to
increase from 33 to 40% of total body
expenditures as lactation proceeds.
The total of these 5 sources of protein
loss adjusted for tissue mobilization
early in lactation was the equivalent
of 11.6, 10.1, and 8.9% of diet DMI
for wk 1, 2, and 3 of lactation, re-
spectively; thereafter this percentage
decreased steadily to 7.5% at 48 wk.
If conversion of dietary protein to MP
remained constant throughout lacta-
tion, the dietary protein need simply
to replace lost N would decrease by
18% from wk 6 to 48. This supports
the suggestion of Wu and Satter
(2000) that need for dietary CP
decreases as milk production declines
and days in milk increases. With
a dietary protein concentration of
17%, the mean efficiency of convert-
ing dietary protein to protein in milk
and other tissues would decrease from
slightly over 31% early in lactation to
below 17% near the end of lactation
Owens et al.
for a weighted average across the full
lactation of 25.4%, a value slightly
less than the 25.5% estimated for a
17% CP diet calculated by regress-
ing milk N efficiency against diet CP
concentrations across recent lactation
trials as is discussed below.
Because maintenance makes a sub-
stantial contribution to total loss of N
by ruminants, it deserves detailed at-
tention. As outlined by NRC (2001),
inevitable losses include the amino
acids lost as scurf as well as N lost as
metabolic fecal protein and as nucleic
acids and other compounds inevitably
excreted in urine. Amino acids provid-
ed in excess of the immediate needs of
the animal must either be oxidized or
placed into protein reserves for either
a short (plasma proteins) or a longer
term (protein depots). Surprisingly,
changes in concentrations of short-
term storage proteins have not been
monitored routinely in N balance
trials. Metabolic fecal protein loss has
been estimated by numerous research-
ers (NRC, 1985) to be about 30 g of
CP/kg of DMI. Estimates of the total
MP requirement for maintenance all
have been based on metabolic size
(g of MP × kg−0.75 shrunk BW): 3.52
(Smuts, 1935); 3.25 (INRA, 1988);
3.8 (NRC, 2000); 2.3 (Chizzotti et
al., 2008). To what degree metabolic
fecal protein fully represents a loss of
tissue amino acids versus microbial
matter synthesized within the diges-
tive tract is not yet clear. Because a
substantial portion of fecal N consists
of microbial residues, metabolic fecal
loss often has been attributed fully to
microbial matter synthesized within
digestive tract, especially the large
intestine. If true, this N expense could
be charged against ammonia or non-
specific N and simply replaced with
NPN or DIP. But if ileal N is derived
partly from body secretions or debris
sloughed from the mucosa of the
digestive tract, some metabolic fecal
N must be derived from body pro-
tein. Flow measurements at the ileum
indicate that the amount of N that
enters the large intestine from the
ileum normally exceeds the amount
of N defecated. So even though urea
diffuses into the large intestine from
blood, the net balance for the large
intestine and cecum is toward absorp-
tion of N. Consequently, metabolic
fecal N may be an underestimate of
the quantitative loss of protein from
the upper digestive tract that enters
the large intestine. Irrespective of
its origin, N losses associated with
maintenance of both lactating cows
(Figure 7) and finishing cattle (Figure
1) comprise a substantial proportion
of the total loss of N from the body
that must be replaced. The quantita-
tive and qualitative origins of meta-
bolic fecal protein (e.g., secretions,
enzymes, cell debris) need to be delin-
eated, ideally with isotope techniques,
to fully comprehend their metabolic
cost in terms of both amino acids and
energy (Lapierre et al., 2014). Indeed,
the high oxygen use of liver and the
digestive tract likely can be attributed
partially to replenishment of both
enzymes and sloughed surface tissues
of the digestive tract that are secreted
into or eroded from the gut during
the process of digestion.
If metabolic fecal loss is attrib-
uted even partially to erosion of the
digestive tract by passage of feed and
fibrous particles, increases in pas-
sage, associated either with increases
in DMI or diet indigestibility, should
increase metabolic fecal N loss. The
NRC (1985) proposed that metabolic
fecal N could be estimated either
as 30 g of CP/kg of DMI or 90 g of
CP/kg of fecal DM output for dairy
cows based on a presumed diet DM
digestibility of 67%. If metabolic fecal
loss is attributed primarily to DM
and coarse particle NDF leaving the
abomasum and eroding the intestinal
tract, its estimation from fecal output
rather than DMI or metabolic size
would seem more appropriate. Rea-
soning further, metabolic fecal loss
of N theoretically could be reduced
either by restricting DMI or by
decreasing fecal output (e.g., decreas-
ing dietary NDF or increasing the
extent that NDF is digested within
the rumen). Reducing metabolic fecal
N loss should spare intestinal tissue
and could reduce the need for mobi-
lization of protein reserves early in
lactation and thereby improve protein

Figure 7. Estimated daily CP expenditures and maintenance requirements of Holstein
dairy cows at various stages of lactation. Color version available in the online PDF.
status and productivity through a full
lactation. Decreased metabolic fecal N
loss could be responsible for a portion
of the performance benefits reported
from limit feeding growing cattle,
from selecting and feeding high-quali-
ty forages that contain less undigested
NDF, and from increased rates of
ruminal NDF digestion (e.g., with
brown midrib forages; Oba and Allen,
1999; Stone et al., 2012). Because
protein status is linked to site and
extent of digestion of DM and NDF,
selection of higher-quality, lower-NDF
forages with a higher rate of NDF
digestion in the rumen should prove
useful not only to increase the sup-
ply of energy available from the diet
and to improve N status by reducing
metabolic loss but also to reduce the
amount of energy required for replace-
ment of eroded tissues.
The degree that body protein is
mobilized concurrently with body
energy reserves early in lactation
when energy balance is negative is
uncertain. Nitrogen metabolism and
retention data were compiled from
8 trials within the data set where
supplemental CP was supplied to
lactating cows in trials starting at dif-
ferent stages of lactation. Calculated
Protein nutrition of ruminants
as N intake minus the sum of milk,
urinary, and fecal protein, nitrogen
balance often was negative early in
lactation (circled values in Figure 8)
despite the very high amounts of di-
etary protein being supplied in these
studies. The negative N balance could
be interpreted to indicate that both
protein and energy reserves were be-
ing mobilized early in lactation when
energy balance was negative. Note
that N balance also was negative at
other points in lactation when the CP
supply was near or below the project-
ed milk protein yield line.
Performance responses to dietary
CP concentrations have been mea-
sured in numerous trials with lac-
tating dairy cows. The regression
equation developed from 82 studies
by NRC (2001) [milk yield = 0.8 ×
DMI + 2.3 × CP − 0.05 × CP2 −
9.8] indicated that milk production
should reach its maximum when diet
DM contains 23% CP. This target
CP concentration is achieved by few
dairy producers unless a feedstuff
that is rich in protein also is a least-
cost source of energy (e.g., distillers
grains). Performance, diet, and excre-
tion data were compiled from 14 more
recent studies with 60 different diets.
159
Based on regression within these stud-
ies, FCM yield reached a plateau with
20.6% dietary CP (Figure 9).
Being curvilinear in both the NRC
(2001) equation and this summary,
milk yield increased at a decreas-
ing rate as dietary CP increased.
Based on averages from these 2
equations, decreasing dietary protein
from 18% to 17, 16, and 15% of diet
DM (sequentially reducing the daily
dietary protein supply by about 0.5
kg per step) would be expected to
decrease expected daily FCM yield by
an average of 1.4, 3.0, and 5.0 kg/d
below FCM production with 18%
CP, changes that may prove statisti-
cally nonsignificant in single studies
but yet important economically. In
addition, milk production and thereby
the need for CP decreases as lacta-
tion continues as noted in Figure 7.
Wu and Satter (2000) proposed that
from 7 to 16 wk of lactation, 19% CP
gave maximum yields, 17% CP was
adequate before and after this point,
and 16% CP was adequate after 30
wk of lactation with their diets and
milk production their cows achieved.
However, even late in lactation when
production was much lower in trials
by Kalscheur et al. (1999), milk yield
often increased when more supple-
mental CP was provided. Neverthe-
less, altering dietary protein content
to match the need for N within
specific segments of lactation rather
than targeting a single CP for an
entire lactation seems like a logical
approach to conserve N. Compared
with decreasing the dietary CP con-
centrations early in or throughout the
lactation period, reducing CP later in
lactation is less likely to reduce FCM
yields for the total lactation.
As noted in trials with feedlot cattle
(Figure 2), DMI tended to increased
linearly (P < 0.06) and quadrati-
cally (P = 0.12) as the concentration
of dietary CP increased, reaching a
maximum at 19.4% CP (Figure 10).
This again reflects a synergy between
dietary CP concentration and energy
intake. Can the increase in DMI fully
explain the increase in milk produc-
tion from addition of CP to these
diets? Milk efficiency, calculated as
160 Owens et al.

FCM per kilogram of DMI from these

experiments, is shown in Figure 11.
Milk efficiency increased slightly but
linearly (P < 0.03) with addition of
dietary CP to the diet. Whether this
truly represents an increase in meta-
bolic efficiency or simply greater dilu-
tion of maintenance remains uncertain
though response appeared greater
when efficiency was greatest (early
in lactation). This increase in milk
efficiency with added protein should
increase economic return to producers
unless the cost of adding protein is
substantial.
Total-tract digestibility of DM and
NDF tended to peak at somewhat
lower dietary CP concentrations (17.4
and 17.9% CP, respectively; data
not shown) than needed to maximize
milk yield or DMI. This suggests
that the DMI stimulation associated
with added CP may be independent
Figure 8. Daily milk protein yields and nitrogen balance of lactating cows fed 8 of gut fill. Milk fat and milk protein
different diets supplemented with various amounts of supplemental CP at various percentages reached plateaus at 19.4
stages of lactation. Circled values represent cases where nitrogen balance was negative and 16.9% CP, respectively (data not
(Neg). Color version available in the online PDF.
shown). If added protein were in-
creasing the supply of limiting amino
acids, one would expect that milk
true protein, an index often used to
monitor protein status of lactating
cows, should plateau at a point near
that needed for maximum FCM yield,
not a lower point. Certainly, milk
urea concentrations should continue
to increase as protein intake increases.
When providing supplemental protein
or rumen-escape methionine, Leonardi
et al. (2003) observed that milk pro-
tein percentage declined when higher-
CP diets were fed even though milk
protein yield tended to increase with
both the higher and lower dietary CP
concentrations when supplemental ru-
men bypass methionine was fed.
Efficiency of converting dietary N
to milk N decreased consistently as
dietary CP concentration increased
(Figure 12). This is attributable
primarily to large increases in urinary
N loss (Figure 13) but small increases
in fecal N loss (Figure 14). Increased
Figure 9. Daily FCM yields of lactating cows fed 25 different diets supplemented with urinary N loss in turn is associated
various amounts of supplemental CP from 13 different published articles. The best- with an increase in daily urine volume
fitting regression line based on newly calculated data are illustrated by the solid line, and presumably in daily water intake.
whereas values predicted from the NRC (2001) are shown in the dashed line. Color Greater water intake, through in-
version available in the online PDF. creasing rumen liquid dilution rate,

Figure 10. Daily DMI by lactating cows fed 25 different diets supplemented with
various amounts of supplemental CP from 13 different published articles. Color version
available in the online PDF.
might affect ruminal fill, DMI, and
MPS. Concerned about the adverse
environmental effects of secreted N
and the high cost of most sources of
Figure 11. Milk efficiency, calculated as FCM divided by DMI, with 25 different diets
supplemented with various amounts of CP. Color version available in the online PDF.
Protein nutrition of ruminants
supplemental protein, Chase et al.
(2009, 2012) illustrated that decreas-
ing the dietary CP concentrations to
16% or below can improve efficiency
161
of converting feed N to milk N, reduce
the acreage needed for waste applica-
tion, and potentially reduce the cost
of purchased feed off the farm while
maintaining milk production. Com-
pared with urinary plus fecal N loss
daily with an 18% CP diet, decreas-
ing CP content of the diet to 17, 16,
and 15% CP should decrease N in
these waste products by 6.2, 12.2,
and 18.3%. Through reducing urinary
excretion, efficiency of N use is im-
proved when the concentration of di-
etary protein is decreased (Figure 12).
Although few dairies formulate diets
to the 20 or 23% CP diets needed to
maximize milk production, any sacri-
fice in milk yield below this maximum
must be balanced against the reduced
cost for supplemental dietary protein
to calculate economic return. In some
instances, reducing the protein and
phosphorus content of dairy rations
will increase cost of milk production
when a feed ingredient rich in pro-
tein and phosphorus has a low cost
(Stewart et al., 2012). Targeting lower
dietary CP concentrations for groups
of cows later in lactation certainly
appears justified to decrease N waste
and the environmental footprint of ex-
cess N, but potential carryover effects
of marginal CP levels early in lacta-
tion deserve further attention.
The MP System
The “metabolizable protein feeding
standard” initially was outlined and
described clearly in an infrequently
cited paper from 1975 by Burroughs
et al. Some of the constants and equa-
tions proposed initially continue to be
used in updated models (NRC, 1985,
2000, 2001). When thinking about
protein requirements for ruminants,
nutritionists use schizophrenic or at
least bipolar reasoning. All animals
require amino acids (MP) for growth
or production and maintenance.
This need is met by a supply of true
protein digested to amino acids in the
intestines and absorbed. For rumi-
nants, this MP supply is derived from
microbial protein synthesized in and
flowing to the small intestine plus
dietary protein that escapes ruminal
162 Owens et al.

digestion (UIP). Only the fractions of

these components that are digested to
amino acids within the small intestine
and are absorbed will contribute to
the MP supply. For MPS, the DIP
supply for ruminal microbes must
be adequate to meet the need of
microbes for MPS if the maximum
supply of MP is desired. Postrumi-
nal degradation of specific sources of
dietary protein that escape ruminal
digestion and availability of individual
amino acids within that protein must
be estimated if requirements are to be
based on the need for specific amino
acids. Finally, from the MP require-
ment standpoint, the quantitative
need for supplemental amino acids
for both maintenance and production
must be predicted despite wide fluc-
tuations in production as well as tem-
porary mobilization or storage of pro-
tein reserves by the host animal. This
logical yet systematic approach can
Figure 12. Efficiency of converting dietary CP to milk protein by lactating cows fed
25 different diets supplemented with various amounts of supplemental CP from 13 be simulated readily through descrip-
different published articles. Color version available in the online PDF. tive models despite the fact that most
of the quantitative estimates and
equations within such models have
proven difficult to ascertain directly
or reliably. Ideally, knowledge of MP
as well as its amino acid content and
availability, when combined with
estimates of requirements, would al-
low feed formulators to predict which
amino acids are deficient and add
appropriate protein sources or rumen-
escape compounds to compensate for
deficiencies. From a point as distant
as the diet, balancing supply with
requirements seems as unwieldy as at-
tempting to tune a television set with
a 3.05-m pole. Various remote control
devices (nutrition models) have been
developed, but their field applica-
tion and evidence of benefits seems
hazy at best. The ultimate goal in
amino acid nutrition with ruminants
is the same as with nonruminants—to
provide an adequate but not excessive
amount of essential and nonessential
N that will precisely meet the need
of an animal for a specified or desired
rate of growth or production.
Individual variables involved in
Figure 13. Daily excretion of urinary nitrogen by lactating cows fed 15 different diets the supply side of any MP system
supplemented with various amounts of supplemental CP from 8 different published need to be evaluated independently
articles. Color version available in the online PDF. to appraise the accuracy of current

Figure 14. Daily excretion of fecal nitrogen by lactating cows fed 15 different diets
supplemented with various amounts of supplemental CP from 8 different published
articles. Color version available in the online PDF.
estimates and to update specific equa-
tions. Individual variables of interest
include 1) MPS; 2) DIP, the supply of
intake protein degraded within the ru-
men and its counterpart; 3) UIP, the
quantity of protein that escapes rumi-
nal digestion; 4) the quantity of N re-
cycled to the rumen that supplements
dietary DIP; 5) the fraction of DIP
that is available for use by ruminal
microbes; and 6) the combination of
MPS and UIP appropriately adjusted
to calculate MP. Two additional com-
ponents related to the use of supplied
MP are 7) intestinal availability of N
or amino acids from microbial protein
and UIP and 8) the quantity of N
or of individual amino acids needed
by the animal for maintenance plus
production. The first 5 of these items
were evaluated using a data set com-
piled from the literature where duo-
denal flows of microbial protein and
rumen-escape protein were measured
with growing-finishing cattle equipped
with intestinal cannulas. Although
data from growing or lactating cattle
were used during development of vari-
ous MP systems (NRC, 1985, 2001),
Protein nutrition of ruminants
much of the subsequent evaluation of
models has been based instead on at-
tempts to relate observed production
rates of sets of cattle to the presumed
supply and availability of MP and net
energy.
Microbial Protein Synthesis
Within the Rumen. Extensive
reviews of ruminal N metabolism
have been published (Bach et al.,
2005; Valadares Filho et al., 2010).
Reviews by Firkins et al. (2007) and
Krause et al. (2013) provide further
details regarding the composition and
activities of the microbial biome of
the rumen. Because microbial protein
typically comprises 50 to 80% of the
CP supply that reaches the small
intestine, maximizing the quantity of
microbial protein flowing from the ru-
men to the omasum or duodenum has
been of primary concern for ruminant
nutritionists. Microbial yield or mass,
microbial flow, and microbial efficien-
cy are separate terms; yield or MPS
is of primary interest for the host
ruminant. The amount of microbial
protein synthesized within the rumen
exceeds the quantity than leaves the
163
rumen (MPS) because a proportion
of the ruminal microbes die and lyse
or are consumed and digested by
protozoa within the rumen. Because
degraded microbes are never counted
as being synthesized, gross or total
synthesis should not be confused with
net yield or net synthesis (MPS).
That output of microbial protein from
the rumen is reduced by protozoa is
supported by the results of Leng et
al. (1989), who found that averaged
across 6 studies, bacterial output from
the rumen was 15% lower for faunated
than defaunated animals. When based
on N isotopes and rumen sampling,
intraruminal cycling of N provides
estimates of gross rather than net
synthesis. Gross synthesis of microbial
protein yield can be limited by avail-
ability of energy (NEl, TDN, OM), of
ammonia or other N sources, or other
nutrients (minerals, vitamins, growth
factors). In contrast, ruminal outflow
as a proportion of gross synthesis will
be reduced when either time for rumi-
nal degradation or protozoal activity
is increased. Based on past literature
summaries, ruminal output of micro-
bial protein with cattle fed typical
diets appears roughly proportional to
the amount of OM that disappears
from the rumen, although MPS from
carbohydrate fermentation is much
greater than from ruminally ferment-
ed protein or lipid. Microbial protein
output from the rumen or MPS is
estimated in vivo by combining the
quantity of microbes within a duode-
nal or omasal sample with the daily
flow of the sample based on some in-
digestible or undigested flow markers.
Flow can be measured directly from
continuous flow fermenters. Valadares
Filho et al. (2010) has discussed and
scrutinized various markers and meth-
ods in detail.
Several different estimates of energy
availability for MPS have been em-
ployed by various researchers. Micro-
biologists typically calculate yields
from laboratory fermenters as Yatp
(i.e., microbial dry weight per mole of
ATP) or yield per mole of substrate
or hexose fermented. Calculated ATP
yields from OM digested could be
164
used to estimate microbial yields in
vitro or even in vivo if ATP yields
and production of specific end prod-
ucts were certain. Apparent digestion
of OM in the rumen (the amount of
OM that disappears during passage
through the rumen) is calculated as
the difference between OM intake
and duodenal OM flow, a value often
called fermented OM. However, OM
entering the duodenum includes not
only undigested dietary OM but also
OM present within ruminal microbes
that consists of microbial mass that
is synthesized or assembled OM from
the diet. Consequently, OM present at
the duodenum includes some synthe-
sized microbes. Thereby, efficiency of
microbial growth often is calculated
as yield per unit of OM truly ferment-
ed, which is OM intake minus duode-
nal flow that is above and beyond the
microbial mass. Calculations based
on fermented OM often are called
efficiency of microbial protein syn-
thesis or microbial growth efficiency,
whereas efficiency values based on
OM truly fermented are called true
efficiency of microbial protein synthe-
sis with the acronym MOEFF. Having
several different bases for estimating
yield of microbial protein has resulted
in confusion among readers, individu-
als compiling data, and researchers.
Microbial yield must not be con-
fused with efficiency of microbial
protein synthesis any more than
fuel mileage, an efficiency estimate,
is confused with the total fuel use
or distance traveled. Increasing the
quantity of carbohydrate fermented
increases microbial output, but re-
sponses are not strictly linear. Nev-
ertheless, under most conditions, the
supply of energy appears to be the
factor that first limits ruminal mi-
crobial growth within the rumen. For
field application of an MP system,
available energy must be based on the
supply of some feed analyte that can
be measured or predicted. As an esti-
mate of energy available for microbial
growth, Burroughs et al. (1975) and
many researchers subsequently have
used TDN intake as an estimate of
digestible energy intake based on the
Owens et al.
assumption that TDN intake should
be correlated with the amount of OM
that is fermented within the rumen.
For evaluation of the MP model 1
system (NRC, 2000), data were com-
piled from 118 high-concentrate diets
that had been fed to growing-finish-
ing cattle. Measurements recorded
included DM and N intake, diet
composition, duodenal flow of N and
OM, microbial CP at the duodenum,
and ruminal escape of dietary pro-
tein. To derive estimates for the MP
model 1 of NRC (2000), TDN, NDF,
DIP, and UIP values were calculated
for each dietary ingredient based on
tabular values from NRC (2000) and
compiled. As indicated previously,
microbial protein yield from the ru-
men was expected to be proportional
to the amount of OM or carbohydrate
fermented in the rumen. Yield of
microbial protein is plotted against
intake of TDN for each of these 118
diets in Figure 15.
Microbial N yield increased linearly
as intake of TDN increased. The in-
tercept was not significantly different
from zero; regression of values forced
through the origin would indicate that
91 g of microbial CP was generated
for each kilogram of TDN consumed.
In previous publications, various
equations have been used to relate
yields of microbial CP to intake of
TDN or NEl as noted below.
Burroughs et al. (1975); NRC (2001):
MPS, g of CP =
TDN intake in kg × 130.
NRC (1985): for dairy, MPS,
g of CP = 6.25 × (TDN intake
in kg × 26.13 − 31.9).
NRC (1985): for beef cattle, MPS,
g of CP = 6.25 × [TDN intake in kg
× (8.63 + 14.6 × forage intake in kg
− 5.16 × forage intake in kg squared
+ 0.595 × concentrate intake in kg)].
NRC (2000): MPS, g of CP =
(TDN intake in kg × 130) reduced
by 2.2% for each 1% decrease in the
dietary concentration of effective
NDF below 20% of DM.
NRC (2001): MPS, g of CP =
6.25 × (11.45 × NEl intake in kg
− 30.93). Assuming that only 85%
of DIP can be captured by ruminal
microbes, the amount of DIP
required is 1.18 times MPS.
Clark et al. (1992): MPS, g of CP =
14.69 × OM intake in kg + 21.94.
Valadares Filho et al. (2010): MPS, g
of CP = TDN intake in kg × 120.38.
Values from these equations are com-
pared with MPS measured from these
118 diets in Figure 16.
Four equations (Burroughs et al.,
1975; NRC, 1985, for dairy; NRC,
2001; Valadares Filho et al., 2010) are
strictly linear functions of TDN or
NEl intake, whereas other equations
(Clark et al., 1992; NRC, 1985; NRC,
2000) are based on the digestibility
or intake of forage and concentrate
separately or had intercepts not
strictly related to TDN intake and
therefore are presented as individual
points in Figure 16. The MPS values
from the compiled measurements from
the data set of feedlot cattle fed high
concentrate diets fell 24 to 30% below
other estimates (91 vs. 120 to 130 g of
microbial CP per kg of TDN con-
sumed). Based on a summary of 997
diets or measurements from Brazil,
Valadares Filho et al. (2010) indicated
that microbial yield per unit of TDN
averaged 7% more for dairy than beef
cattle though the concentrate levels
fed to beef cattle in their trials was
not specified. The NRC (1985) based
microbial yield estimates on complete-
ly different equations for beef versus
dairy cattle.
 Protein nutrition of ruminants 165

for microbes that ferment starch.

The greater prevalence of starch than
NDF fermenting ruminal bacteria
within the rumen of cattle fed diets
rich in concentrate should lead to a
lower efficiency of microbial growth.
On this basis, Russell et al. (1992)
proposed and NRC (2000) adopted
the concept of reducing the proposed
efficiency with which dietary TDN is
converted into microbial protein when
diets contain less than 20% effective
NDF and ruminal pH is low. Rate
of liquid dilution rate also may be
involved and similarly should be al-
tered by effective NDF. Like animals,
ruminal microbes expend energy for
both growth and maintenance, and
the amount of energy expended for
maintenance is time dependent. As
intake of DM and NDF increase, both
of which are greater with diets for lac-
tating cows than for growing-finishing
Figure 15. Daily duodenal flow of CP for intestinally cannulated growing-finishing feedlot cattle, retention time for liquid
cattle fed 118 different diets at various intakes of TDN in 37 trials from 28 different and unattached microbes within the
published articles. Color version available in the online PDF. rumen decreases (Seo et al., 2009).
A shorter ruminal residence time for
Why should efficiency of microbial suggested that 0.05 to 0.15 g of carbo- microbes, similar to a faster growth
protein production be less with diets hydrate is expended by each gram of rate for cattle, reduces the proportion
that contain more grain or concen- bacteria for each hour of maintenance, of energy expended for maintenance.
trate? Russell and Baldwin (1981) with this value being 3-times greater Consequently, microbial efficiency
increases as ruminal residence time
decreases. Although more small-parti-
cle feed components are flushed from
the rumen when ruminal residence
time is reduced, this increased yield
of microbial protein with higher DMI
and with diets rich in NDF could be
attributed partially to a shortened
residence time for microbes within
the rumen. This matches with the
observation of Bach et al. (2005), who
found that microbial protein yield de-
creased as extent of ruminal digestion
of OM increased, probably the result
of a higher amount or more exten-
sive processing of grain in the diet
or a reduced intake of DM, both of
which would increase ruminal reten-
tion time. Certainly, the efficiency of
microbial growth increases markedly
as dilution rate increases (Isaacson et
al., 1975).
The factor most closely related to
Figure 16. Literature estimates of predicted duodenal flow of microbial protein microbial yield should be the amount
flow from 6 different sources for comparison with measured duodenal flow of CP for
of energy fermented in the rumen that
growing-finishing cattle fed 118 different diets at various intakes of TDN. Color version
available in the online PDF. yields energy for maintenance and
166
Figure 17. Ruminal disappearance of OM with 118 different feedlot diets that
contained various grain sources and concentrations of TDN. Color version available in
the online PDF.
growth of ruminal microbes. Conse-
quently, ruminal disappearance of OM
or carbohydrate should be of primary
Figure 18. Duodenal microbial protein supply of growing-finishing cattle versus daily
DMI. Color version available in the online PDF.
Owens et al.
interest. How was ruminal disappear-
ance of OM related to TDN content
of the diet within these studies where
several sources of grain processed by
various methods were fed? This rela-
tionship is shown in Figure 17.
Clearly, the measured extent of
disappearance of OM was poorly cor-
related (R2 = 0.00) with TDN con-
tent across these 118 diets. That the
proportion of dietary carbohydrate
fermented within the rumen can dif-
fer markedly among feeds and within
feed constituents and can be altered
by feed source or grain processing
should come as no surprise (Firkins
et al., 2001). To what degree can the
differences in DMI alone among these
trials explain the observed increases
in microbial protein yield? Microbial
protein yield is regressed against DMI
in Figure 18.
Within these 118 diets, TDN ranged
from 76 to 89% of DM, whereas daily
DMI ranged from 2.7 to 10.8 kg or,
as a percentage of BW, 1.2 to 2.6%/d
(Table 1). An ideal MP model should
predict supply and demand equally
well for cattle that differ in size, con-
dition, and production even though
equations inherent in such models
often were derived from experiments
in which diversity has been limited.
For example, with low intakes of feed
and water, ruminal residence time
would increase, which in turn should
lower the efficiency of microbial
growth as discussed above. Indeed,
regressions indicated that DMI alone
could explain 47% of the variation (R2
= 0.4715) in MPS yield; the combi-
nation of DMI with TDN content of
the diet explained 48% (R2 = 0.4846)
of the total variation. Addition of
dietary TDN concentration to the
regression equation failed to improve
data fit. Whether regressions devel-
oped in the past that related micro-
bial yield to intake of TDN or energy
(NRC, 1985, 2001) have appropriately
accounted for independent effects of
DMI and TDN is uncertain. How was
microbial yield related to TDN con-
tent of the diets among these studies?
These regressions without or with a
zero intercept are shown in Figure 19.
The relationship of MPS to TDN
concentration of the diet, an index
of total-tract energy digestion, was
nonexistent (R2 = 0.01). Because the

Figure 19. Duodenal microbial protein supply of growing-finishing cattle versus TDN
concentration of the diet being fed. Color version available in the online PDF.
range in DMI was much greater than
the range in TDN content of diets in
these studies, one would expect that
TDN intake should be related more
closely to DMI than to TDN content
of the diet. If extent of OM disap-
pearance in the rumen limits micro-
bial growth, then the preferred index
for predicting microbial yield should
be ruminal OM disappearance, not
TDN. Because TDN is imprecise as
a predictor of ruminal OM digestion
(Figure 17), some base other than
TDN (e.g., in situ, in vitro, or gas
yield measurements) should be prefer-
able for predicting extent of ruminal
digestion of consumed carbohydrate.
To date, TDN intake has served as
the inherent component for predicting
microbial protein synthesis in most
MP systems developed in the United
States. To determine how factors
beyond the quantity of OM fermented
in the rumen related to DMI are driv-
ing microbial protein yield, greater
research attention should be paid to
such factors. Within this data set,
multiple regression indicated that
efficiency of MPS (g of MPS/g of
ruminally fermented OM) was related
negatively (P < 0.01) to grams of
Protein nutrition of ruminants
ruminally digested starch and grams
of ruminally digested nonstarch OM,
as had been observed previously by
Bach et al. (2005), but related posi-
tively to diet CP concentration (P
< 0.01) and DMI as a percentage of
BW (P < 0.05). Unfortunately, these
4 factors still could account for only
45% of the total variation in micro-
bial efficiency. Note that the latter 2
factors could reflect MPS responses
to ruminal residence time of liquids
and particles. Residence time can be
altered by ruminal input of both DM,
NDF, and fluid (imbibed water and
saliva produced during eating and
rumination) as well as urine excretion
(associated with intakes of salt and N,
antidiuretic hormone, and diuretics)
that drives water consumption.
UIP. Estimates for DIP and UIP
for individual feeds are listed in NRC
(2000) tables with the sum of these
2 being total dietary protein. These
estimates have been derived primar-
ily from laboratory measurements
associated with N solubility in certain
solvents as modified by additional in
situ, in vitro, or in vivo measurements
for specific feeds.
167
Simultaneous with microbial growth
and protein synthesis in the rumen,
ruminal microbes readily degrade
most sources of dietary N and recy-
cled N. Proteins in the diet, in saliva,
and from nonviable ruminal microbes
are degraded partially to peptides,
amino acids, and ammonia. Extent
of protein degradation varies with
both the accessibility and susceptibil-
ity of a protein source to microbial
attack or engulfment as well as time
for attack and the activity of proteo-
lytic microbes and protozoa within
the rumen. Susceptibility to attack
usually is correlated with solubility,
but solubility varies with ruminal pH
and feed processing. Consequently,
extent of ruminal protein degradation
and UIP varies with ruminal pH and
perhaps with microbial adaptation to
a given source of protein. In contrast,
protein accessibility typically is tied
to encapsulation or close association
with fiber; reducing particle size by
extensive feed processing will expose
more plant protein for attack by ru-
minal microbes.
Though most ruminal bacteria,
particularly those that ferment cel-
lulose, thrive with ammonia as their
sole source of N, bacteria that ferment
starch and sugars often exhibit in-
creased growth rates and yields when
provided with peptides and amino ac-
ids. Although free amino acids can be
absorbed from the rumen (Liebholz,
1971) and from 9 to 16% of total flow
of amino acid N in omasal samples
consisted of free amino acids (Reynal
et al., 2007), free amino acids as well
as peptides are rapidly degraded by
ruminal microbes (Wallace, 1996),
leaving only very low concentrations
within the rumen. Ruminal microbes
will adapt to degrade specific amino
acids; this makes estimates of amino
acid degradation based on incuba-
tion with rumen fluid from unadapted
animals questionable. Ruminal escape
of dietary protein also can differ
with adaptation or other dietary
constituents that alter ruminal pH
and protozoal activity (Loerch et al.,
1983). Branch-chained fatty acids or
amino acids are required for growth of
several prominent cellulose-digesting
168
bacterial species and pure-culture and
in vitro studies have illustrated the
nutritional benefits from supplemental
peptides and branch-chained fatty ac-
ids. However, dietary supplementation
with peptides or branch-chained fatty
acids seldom has proven beneficial in
vivo, likely because ruminal concen-
trations of such compounds already
are adequate due to degradation of
dietary protein or to crossfeeding and
lysis of microbes within the rumen.
Nevertheless, the fact that peptides
can be detected within the rumen in-
dicates that peptide degradation can
be a rate-limiting step in proteolysis
(Wallace, 1996).
In vitro benefits from providing
peptides or proteins rather than
amino acids may reflect an energetic
advantage for bacterial uptake of
polymers. The energy required for
activating one mole of amino acid for
uptake by bacteria may equal that
required to activate a long peptide
or protein. Because the amount of
energy required by bacteria for uptake
of monomers (sugars, amino acids,
nucleic acids) from the media can be
substantial and because the concen-
tration and timely availability of
monomers may be limited, benefits in
rate or efficiency of microbial growth
from providing monomers rather than
polymers in vitro have been detected
very rarely.
Reducing the prevalence of protozoa
present in the rumen decreases the
ruminal ammonia concentration by
reducing the extent to which proto-
zoa consume and subsequently digest
small feed particles and ruminal
bacteria. Certain microbial species
(i.e., Prevotella, Ruminobacter, Bu-
tyrivibrio, Selenomonas) are actively
proteolytic. Through altering the
populations of proteolytic and pep-
tidolytic bacteria within the rumen,
certain ionophores and antibiotics can
reduce degradation of protein or pep-
tides within the rumen as summarized
by Walker et al. (2005). Compounds
that can inhibit proteolytic enzymes
directly or reduce the prevalence or
activity of certain bacterial, protozoal,
or fungal species, particularly hyper-
ammonia producers, within the rumen
Owens et al.
should increase UIP values of feeds
(Wallace, 1996). Some research efforts
have been directed toward preventing
proteolysis of feed components before
feeding. However, preventing proteoly-
sis within a feed before feeding may
not increase the postruminal supply
of protein if that protein remains
readily degraded within the rumen.
However degradation of protein to
ammonia before feeding (e.g., during
fermentation of crops or feeds) would
deprive the ruminal microbes of spe-
cific nitrogenous polymers that might
enhance microbial growth.
Time for ruminal digestion is a criti-
cal variable that controls the extent
of ruminal degradation of protein
components. As outlined by Seo et al.
(2009), an increase in DMI or NDF
will decrease ruminal retention time
of liquids and particles. Currently,
most models of ruminal digestion
are based on the assumption that all
consumed liquids and solids enter the
rumen, mix thoroughly with similar
components within the rumen, and
are metered from the rumen at a
stable and constant rate. Yet, turn-
over of liquids always is greater than
for solid particles found in the rumen.
Ruminal stratification, both horizon-
tally and associated with the ruminal
raft, and vertically, as reflected by
the high moisture content of reticu-
lar contents, indicate that mixing
of components within the rumen is
incomplete; ruminal contents are not
homogeneous. Following a meal, ru-
minants invariably consume available
water. Such water can sluice reticu-
lar contents to the omasum. Indeed,
consumed feed particles will appear in
abomasal contents immediately after
a meal, particularly when the rumen
is fully occupied. Ruminal retention
also may change diurnally. Weight of
ruminal contents of grazing ruminants
is greater after an evening than after
a morning grazing bout, perhaps an
adaptation for retaining forage for
nocturnal rumination (Gregorini et
al., 2008). If passage rate changes di-
urnally with ruminants, the extent of
ruminal escape of fed materials could
be altered by providing specific feed
components at specific times or fre-
quencies. Considering that biological
clocks regulate organ metabolism in
other species (Grens, 2013), it would
not be surprising to find circadian
cyclicity within the digestive tract of
ruminants. These physical as well as
chemical aspects of rumen function
deserve greater research attention.
Escape of dietary protein with these
118 diets was calculated as total
duodenal flow of N minus microbial N
and ammonia N and thereby includes
all endogenous N that is present.
Predicted ruminal escape of dietary
N was calculated from DMI, diet com-
position, and tabular UIP values for
individual feed components provided
by NRC (2000). These values were
compared with measured duodenal
flow of N that was not ammonia or
microbial protein. On the average,
tabular values indicated that UIP
comprised 4.4% of diet DM (range
of 1.8 to 8.4%), an equivalent of
35.1% of dietary protein (range of
14 to 51%). Predicted UIP is plot-
ted against daily flow of dietary plus
endogenous protein across these 118
diets. Measured UIP plus endogenous
protein averaged 258 g daily (range of
90 to 554 g) as plotted in Figure 20.
Measured rumen-escape protein
tended to increase as predicted UIP
increased, but the relationship was
weak. Because ruminal residence time
also can be altered by DMI, these
values were recalculated per unit of
DMI. Results are shown in Figure 21.
This relationship was much weaker,
presumably because higher DMI is
associated with a shorter ruminal resi-
dence time for degradation of dietary
protein sources. Based on their effect
on daily gain of cattle, Tedeschi et al.
(2005) concluded that tabular UIP
values were inconsistent and proposed
that both TDN and UIP were altered
by level of DMI.
Because processing of oilseeds and
grains can influence accessibility
and susceptibility of protein sources
to proteolysis, some standardized
procedure for estimating or predicting
in vivo UIP is needed. In addition,
adjustments for ruminal pH and NDF
digestion should prove helpful. Clas-
sically, UIP is simply calculated as
 Protein nutrition of ruminants 169

CP minus DIP. Though numerically

valid, all UIP is not equally digestible.
Although regression may indicate that
true digestibility of dietary CP by
ruminants averages 90%, studies with
nonruminants indicate that intestinal
digestibility of UIP can fall below
70% (Boucher et al., 2009). Similar to
the ‘c’ fraction in kinetic analyses and
indigestible NDF, indigestible UIP is
present in substantial amounts within
certain feed ingredients (e.g., feath-
ers, hair, or leather that has not been
hydrolyzed; feeds including distillers
products; heat-damaged feeds). Pro-
tein sources indigestible in pepsin or
by chicks or indelibly bound to NDF
could be subtracted from CP before
formulating diets or calculating UIP
within MP systems. Adjusting the
protein content for UIP for individual
feeds for expected differences in intes-
tinal digestion to derive an available
Figure 20. Undegraded intake protein (UIP) in grams per day predicted by NRC
UIP value would avoid the need to
(2000) equations versus UIP recovered at the duodenum by growing-finishing cattle
fed 118 different feedlot diets. The steeper line represents a perfect fit of predicted to
include an additional layer of adjust-
observed values. Color version available in the online PDF. ment factors within an MP system.
DIP. Estimates of DIP, calculated
as the difference between CP intake
and bypassed protein, comprised an
average of 8.7% of diet DM (with a
range of 4.9 to 12.5%) for diets in this
summary, an equivalent to 65% of di-
etary CP. Predicted DIP values for di-
ets based on tabular values from NRC
(2000) were compared with measured
DIP values as plotted in Figure 22.
Measured DIP generally increased
with estimates of dietary DIP, prob-
ably because a substantial portion
of dietary DIP in these diets often
was provided by urea. Because UIP
estimates differed with level of DMI,
measured DIP also was plotted
against available DIP per kilogram of
diet (Figure 23).
Precision of predicting DIP was re-
duced when the effect of level of DMI
was removed. This again indicates
that the magnitude of the regression
predicting UIP and thereby DIP was
being driven partially by differences
in DMI.
Extent of microbial protein synthe-
Figure 21. Undegraded intake protein (UIP) in grams per kilogram of diet predicted sis can be limited by availability of
by NRC (2000) equations versus UIP recovered at the duodenum by growing-finishing ammonia or DIP. The NRC (2001)
cattle fed 118 different feedlot diets per kilogram of diet fed. Color version available in has proposed that only 85% of DIP
the online PDF. is available for synthesis of microbial
170 Owens et al.

protein. To determine the relationship

of microbial yield to the available DIP
value from NRC (2000), data from
these 118 feedlot diets were plotted
(Figure 24).
Among these diets, only 4 had
microbial protein yields that exceeded
the amount of available DIP based on
the DIP estimates for diet ingredients
from NRC (2000). This would suggest
that microbial protein yield was not
being limited by supply of available
DIP. However, because predicted DIP
differed from measured DIP, the rela-
tionship of measured DIP, calculated
as CP intake minus measured UIP, to
MPS also was calculated as shown in
Figure 25.
Based on the measured DIP sup-
ply, the supply of DIP from the diet
alone would have been insufficient
for synthesis of microbial protein
synthesis within the rumen with 67%
Figure 22. Ruminally available degraded intake protein (DIP) in grams per day of these diets. This indicates that
predicted by NRC (2000) equations versus dietary protein degraded in the rumen by synthesis of microbial protein po-
growing-finishing cattle fed 118 different feedlot diets. Color version available in the
tentially could be limited by supply
online PDF.
of DIP from the diet. The need for
DIP will increase as grain is more
extensively processed and extent of
carbohydrate digestion within the
rumen increases. Therefore, DIP sup-
ply is more likely to limit supply of
MP or extent of ruminal digestion if
grains are processed more extensively.
Certainly, performance often increases
when additional supplemental NPN
is supplied, particularly with more
extensively processed grains as dis-
cussed by Klopfenstein and Erickson
(2002). However, the extent to which
the observed increase in performance
from added DIP is attributable to an
indirect effect on DMI and not to an
increase N retention as illustrated by
Cole et al. (2006) remains unclear.
Urea that is recycled to the rumen
supplies additional DIP for rumi-
nal microbes. Extent to which N is
recycled to the rumen may differ with
diet, being lower with feedlot diets
with low NDF concentrations due to
minimal rumination of such diets.
Duodenal output of CP was plotted
Figure 23. Ruminally available degraded intake protein (DIP) in grams per kilogram
of diet predicted by NRC (2000) equations versus dietary protein degraded in the
against dietary CP intake in Figure
rumen per kilogram of diet fed by growing-finishing cattle fed 118 different feedlot 26 to examine the degree to which
diets. Color version available in the online PDF. duodenal output of microbial protein
 Protein nutrition of ruminants 171

might exceed DIP available from the

diet alone.
Duodenal flow of CP often exceeded
CP intake, with the difference tending
to be slightly greater with lower CP
intakes. On the average, duodenal CP
flow exceeded CP intake by 19 g daily
though as much as 363 g (38% of CP
intake) was being lost and as much as
231 g (39% of CP intake) was being
gained during passage through the ru-
men. These values likely are underes-
timates of total recycling of N to the
rumen because only the recycled DIP
captured as microbial protein would
increase the duodenal CP supply.
Suspicion should be raised whenever
estimates of duodenal UIP plus micro-
bial protein differ markedly from CP
supply in any MP system.
Metabolizable Protein Supply.
Of the duodenal CP supply, only a
Figure 24. Ruminally available degraded intake protein (DIP) in grams per day portion is available to the host rumi-
predicted by NRC (2000) equations versus measured microbial protein entering the nant as amino acids. The NRC (1985,
duodenum daily of growing-finishing cattle fed 118 different feedlot diets. Color version 2000, 2001) estimated that 80% of
available in the online PDF. microbial CP is true protein and that
small intestinal digestibility of true
protein from both microbial CP and
UIP is 80%. Using these estimates,
MP values were calculated based on
duodenal supply of microbial protein
and UIP. The NRC (2000) tabular
values for MP based on predicted
MP and UIP for feed components are
compared with estimates of MP sup-
ply based on duodenal flow in Figure
27.
Predicted MP supply increased as
observed MP supply increased though
many values deviated from the ideal
regression line, with predicted val-
ues explaining 69% of the observed
variation even though neither MPS
nor ruminal escape of dietary protein
had been predicted with this degree
of accuracy. This indicates that the
underestimate of MPS of NRC (2000)
of Figure 16 was being compensated
partially by the greater-than-expected
escape of dietary protein (Figure 21).
Again considering the wide range in
DMI among these diets, the expected
supply was plotted against predicted
Figure 25. Ruminally available degraded intake protein (DIP) in grams per day supply per kilogram of diet in an
predicted by NRC (2000) equations versus measured microbial protein entering the attempt to remove differences that
duodenum in grams per day for growing-finishing cattle fed 118 different feedlot diets. could be driven by or associated with
UIP = undegraded intake protein. Color version available in the online PDF. differences in DMI (Figure 28).
172
Figure 26. Crude protein entering the duodenum in grams per day versus CP
consumed daily by growing-finishing cattle consuming feedlot diets. Color version
available in the online PDF.
The relationship between supply of
MP based on duodenal CP compo-
nents and MP supply predicted from
NRC (2000) equations fell apart when
these values were adjusted for dif-
Figure 27. Metabolizable protein (MP) supply in grams per day predicted by NRC
(2000) versus MP supply estimated from daily duodenal flow of microbial protein plus
protein that escaped ruminal digestion. Color version available in the online PDF.
Owens et al.
ferences in DMI. This supports the
suggestion that duodenal supply of
microbial protein and of UIP were be-
ing driven strongly by DMI. Failure of
predicting values per kilogram of diet
would represent the sum of impreci-
sions for predicting microbial protein
synthesis and UIP per kilogram of
diet.
Although the MP system seems
sound physiologically, these direct
comparisons of duodenal supply of
microbial protein and rumen-escape
protein based on 118 different feed-
lot diets indicate that the equations
involved with predicting MP supply
require further testing and refinement.
Any model system must accurately
predict the duodenal supply of CP
and its components before attempting
to match this supply with the more
nebulous requirements for amino acids
for growth and maintenance. Mea-
surements at the duodenum, an in-
termediate point in digestion, should
be useful to appraise the veracity and
modify estimates of the supply of
protein or amino acids. Attempting to
short-circuit the science and develop
equations and models based on a
black-box mentality and simply relate
performance or production to mea-
sured dietary constituents is unlikely
to prove fruitful.
Amino Acid Requirements
of Ruminants
Several studies have been con-
ducted in which an increased supply
of certain amino acids has increased
rate and efficiency of gain of light-
weight cattle. Addition of a lysine-rich
supplement (fish meal) to a diet rich
in steam-flaked corn increased rate
and efficiency of gain early in a feed-
ing trial with steers started on feed at
122 kg (Zinn et al., 2000). Similarly,
Sindt et al. (1993) detected improve-
ments in the G:F from supplementing
a urea-based diet with a mixture of
blood meal and feather meal early
during a feeding trial with 260-kg
calves. In a calf-finishing trial with
237-kg steers fed a 12% CP urea-
supplemented diet, Klemesrud et al.
(2000) detected a quadratic response
in ADG during the first 56 d of a 161-
d feeding study from supplementing
the diet with rumen-escape methio-
nine plus lysine. Plasma lysine in-
creases with supplementation support

Figure 28. Metabolizable protein (MP) supply in grams per kilogram of diet predicted
by NRC (2000) versus MP supply estimated from daily duodenal flow of microbial
protein plus protein that escaped ruminal digestion per kilogram of diet. Color version
available in the online PDF.
the contention that lysine supply was
increased, but the fact that plasma
concentrations increased with the
first level of supplementation implies
that the lysine requirement already
had been exceeded. Based on swine
studies, corn grain is notably low in
lysine and tryptophan and 92% of the
diet fed in their trial consisted of corn
products (corn-gluten feed, high-mois-
ture corn, dry-rolled corn, and corn si-
lage). In contrast, Hussein and Berger
(1995) detected no performance
response to supplementation of a 13%
CP diet based on whole corn with this
same rumen-escape lysine–methionine
mixture with 183-kg steers. Unfor-
tunately, without knowledge of the
basal supply of indispensible amino
acids at the duodenum as determined
with intestinally cannulated animals,
responses from supplementation yield
little information about the total
amount of an individual amino acid
that might be required for growth
and maintenance. Measured intesti-
nal amino acid flow can be compared
against various indices of amino acid
Protein nutrition of ruminants
requirements derived from other spe-
cies as illustrated by Zinn and Shen
(1998), but the applicability of those
requirement estimates to ruminants
is questionable. Nevertheless, when
performance responses are detected
to rumen-escape amino acids, results
have commercial application. Ex-
trapolating such responses to other
conditions where diets differ in com-
position or to animals that differ in
maturity, production rate, and DMI is
foolhardy.
Because the requirements for MP
and essential amino acids decrease
as feedlot cattle mature, it would be
helpful for those attempting to com-
pile and interpret data sets if authors
presented interim intake and per-
formance data in published articles.
Also, when multiple protein levels and
sources are fed and interactions are
not significant, authors often present
only the means averaged across other
variables. Without individual treat-
ment data, the published information
is useless for compiling into data sets.
Publications also should list the UIP
173
and DIP estimates that authors have
used for individual feeds and for total
diets in their reports.
As with energy, requirements for
amino acids represent the sum of 2
factors—one portion being used for
production and a second portion
being used for maintenance. Only
through subdividing amino acid re-
quirements can one logically appraise
the total need at various levels of pro-
duction that occur in practice (e.g.,
from growing heifers or dry cows to
lactating cows; from feedlot steers to
stocker cattle or wintering beef cows).
Amino Acid Requirements for
Production. The 8 to 12 amino acids
that cannot be synthesized by tissues
of animals or birds at a sufficient rate
to meet the demands for mainte-
nance and production are considered
essential amino acids. By synthesiz-
ing essential amino acids, microbial
protein will partially or can com-
pletely displace the need for essential
amino acids in the diet. However, for
maximum rates of growth of younger,
growing ruminants and for lactat-
ing ruminants, supplementation of
most diets with protein sources that
provide essential amino acids will
increase production. Both ruminants
and nonruminants synthesize nones-
sential amino acids when a source of
nonspecific N (e.g., NPN) is provided.
Luckily, the ratio of essential to non-
essential amino acids in microbial pro-
tein is very high, almost 50% of total
N, a fact that is surprising consider-
ing that a substantial portion of total
N consists of nucleic acids. Though
certain amino acids have additional
fates that are irreversible (synthesis of
cystine, tyrosine, niacin, thyroxin, car-
nitine, glutathione, methyl-histidine),
the primary use of amino acids is for
either production (milk, lean tissue,
wool, mohair) or maintenance. The
quantities of amino acids retained or
secreted by an animal can be esti-
mated from the amino acid content
of products and the rate of produc-
tion. Because production rates differ
with stage of lactation or growth and
because body protein reserves are
mobilized when the need for protein
174
(or energy) exceeds the supply, amino
acid requirements for production will
vary with stage and rate of produc-
tion. Changes in amino acid require-
ments with level of milk production
when combined with the potential to
raid protein reserves as well as car-
ryover effects markedly complicate
the interpretation of results from both
crossover and full-length lactation
experiments. Within either growth
or lactation trials, providing interim
data can help delineate the specific
time intervals when responses dif-
fered over time as shown in trials by
Wu and Satter (2000) and Zinn et al.
(2000). Without such interim data, it
is impossible to delineate when ben-
efits are present. Although the drain
on amino acids for proteins secreted
in milk or deposited can be measured
with a high degree of precision, amino
acid requirements for maintenance
have proven much more difficult to
appraise both quantitatively and
qualitatively.
Quantitative N Requirements.
Several aspects of amino acid nutri-
tion obvious from experiments with
nonruminants as gleaned from vari-
ous sources need to be borne in mind
when appraising amino acid nutrition
of ruminants. These presumptions and
their application to N metabolism in
ruminants are outlined below.
Presumption 1. Daily amino acid
requirements and supplies should be
expressed as grams, not percentages.
A diet that contains 20% protein
with 2.5% lysine will meet the lysine
requirement as well as a diet that
contains 10% protein of which 5% is
lysine. The lysine percentage of the
diet is irrelevant. Yet, because perfor-
mance of ruminants often is limited
by the supply of available energy, the
amino acid requirements for main-
tenance and production ultimately
could be expressed relative to the
amounts of energy dedicated to main-
tenance versus production.
Presumption 2. An amino acid
is required because its α-keto acid
cannot be synthesized in adequate
amounts to meet the need of an ani-
mal. Most α-keto acids, whether es-
sential or not, are routinely transami-
Owens et al.
nated. For example, branched-chained
amino acids are found in bacterial
strains that require branched-chain
fatty acids. Though this interchange
of N should not negate the use of
labeled N as a marker for microbial
flow, it can confound estimates of N
turnover estimates and interchange of
N among ruminal microbes.
Presumption 3. Certain amino ac-
ids are converted by the body to oth-
ers (methionine to cystine and phenyl-
alanine to tyrosine). These reactions
are not reversible. All 4 of these
amino acids are used for synthesis
of protein though the relative ratios
required can differ. When evaluating
the available supply for an animal for
these amino acids, the 2 sulfur amino
acids should be considered as a single
unit and the 2 phenolic amino acids
should be considered as a unit be-
cause the requirement for methionine
or phenylalanine can be met partially
by cystine and tyrosine, respectively.
Supplementation with methionine can
meet a shortage of either methionine
or cystine, so a performance response
to methionine supplementation does
not determine whether methionine or
cystine was deficient. Consequently,
an increase in the postruminal supply
of dietary cystine or cysteine, as from
feather meal, might prove as useful as
a rumen-bypassed methionine supple-
ment.
Presumption 4. Amino acids
interact. Excesses of lysine and of
a branched-chain amino acid will
accelerate oxidative degradation of
arginine and of other branched-chain
amino acids, respectively. Supplemen-
tation of the second limiting amino
acid, through causing an amino acid
imbalance, will depress DMI and
N retention of growing ruminants
(Papas et al., 1974). Presumably,
supplementation with amino acids in
amounts above the quantity required
can generate an imbalance and de-
press performance.
Presumption 5. Amino acids in
excess are deaminated and the carbon
skeleton is oxidized. Degradation ap-
pears proportional to the amino acid
concentration in blood plasma. Both
oxidation and plasma concentrations
will increase above a baseline value
when the supply of an amino acid ex-
ceeds its need. Consequently, plasma
amino acid concentrations and amino
acid oxidation are useful tools to
determine the break-point when the
supply exceeds the requirement. How-
ever, these break-points merely rep-
resent the point at which some other
factor (another amino acid or nutri-
ent or energy) limits production or
performance. If and when this second
ceiling is raised, the requirement for
the first nutrient increases. The clas-
sical relationship of performance to
supply of a limiting amino acid is il-
lustrated in Figure 29. Note also that
efficiency of use of the limiting amino
acid for production never plateaus but
changes with degree of deficiency or
excess. Consequently, efficiency of use
of an added amino acid is useless as
an index of a requirement. However,
efficiency of use always increases with
supplementation of an amino acid
when that amino acid is deficient.
Presumption 6. When degraded,
amino acids yield N for synthesis of
nonessential amino acids. Methionine
and cystine also yield sulfur, a nutri-
ent used in and recycled to the rumen
and occasionally deficient for NDF
digestion. Certain amino acids are
active metabolically (synthesis of thy-
roxin, glutathione, niacin, carnitine);
others can alter hormone secretions
(growth hormone effects of arginine),
and others may alter motility of the
digestive tract and passage rate (and
potentially ruminal dilution rate).
Consequently, a performance response
to a supplemented amino acid does
not necessarily involve correction of
an amino acid deficiency. An increase
in nitrogen balance when combined
with an increase in product secretion
per unit of N intake is the ultimate
test for determining that an amino
acid deficiency was being met by
supplementation.
Presumption 7. Supplementation
with an amino acid should enhance
protein synthesis and N retention
only when that amino acid is the
first limiting nutrient. Because an
individual amino acid is only a small
fragment of depot or secreted pro-

Figure 29. Theoretical performance (solid line) and efficiency (dashed line) responses
to supplementing a diet with an amino acid below and above the amount required to
maximize performance. Color version available in the online PDF.
teins, N retention or secretion should
increase to a much greater degree
than the amount of added N being
supplied if the deficient amino acid
is being supplemented at the proper
rate. If diets already are formulated
to provide an adequate quantity of
protein to maximize production, no
production response to amino acid
supplementation should be expected.
Supplemental amino acids are costly,
and their supplementation to diets
for nonruminants is justified economi-
cally only with reduced-protein diets.
Supplementation of such diets can
restore production to the rates seen
with a higher-protein diet. By displac-
ing dietary protein with a balanced
mixture of limiting amino acids, effi-
ciency of N use is increased and waste
of N is reduced. An excess of N will
increase synthesis of urea and loss of
energy in urine, but true proteins con-
sistently have a greater gross content
than carbohydrates. This can fully
compensate for the additional loss of
energy in urea. An excess supply of
NPN similarly will increase urinary
energy loss, but because NPN sources
do not provide additional gross energy
like true proteins, this energy loss is
not compensated. So in addition to
avoiding ammonia intoxication, avoid-
ing excessive amounts of NPN should
Protein nutrition of ruminants
reduce urinary loss of energy associ-
ated with urea synthesis and secre-
tion. Because production economics
typically dictates diet formulation,
environmental effects, though of ethi-
cal concern, often are ignored.
Presumption 8. Amino acid con-
tent of protein sources differs widely
and amino acid requirements differ
with performance level and product.
The ratio among amino acids released
from mobilized body reserves differs
from the ratios used for maintenance
and production. This reduces the ef-
ficiency of transfer of N from storage
to production. Similarly, responses to
supplementation with a specific amino
acid should not be expected across
various stages of production or when
the dietary protein sources or supplies
of rumen-escape protein differ.
Presumption 9. Both a negative
control (the unsupplemented diet)
and a positive control diet (where
supply of all nutrients including N
and S are supplied) are required in
studies designed to quantify responses
to amino acids or N. Ideally, some
nonessential amino acids should
supply additional N in the control
diet. In several trials, postruminal
supplementation with glutamic acid, a
nonessential amino acid, increased N
retention by growing sheep (Schelling
175
and Hatfield, 1968; Nimrick et al.,
1970), indicating that this nonessen-
tial N source somehow was beneficial.
Increases in plasma concentrations or
oxidation rates can detect when an
excess has been supplied.
Amino Acid Requirements for
Maintenance. That the pattern
of amino acids required for mainte-
nance reflects the pattern of amino
acids present in mixed protein of the
body was suggested by Young and
El-Khoury (1995). This assumption is
inherent whenever the amino acid re-
quirements for producing animals are
suggested to be proportional to body
composition (Ainslie et al., 1993). If
amino acid requirements for growth
and maintenance were proportional
to the amino acid composition of the
body, thousands of experiments de-
signed to quantify amino acid require-
ments were unnecessary. Although
this concept has been integrated into
some models employed to predict ami-
no acid requirements for ruminants, it
has no scientific basis. Trials sum-
marized by Coon (2000) indicate that
maintenance requirements for both
growing and adult birds are related
more closely to the amino acid com-
position of feathers than to the amino
acid content of body tissue. Likewise,
the pattern of amino acids required
by sows differs from that of growing
pigs (Moehn et al., 2011), and the
ratios among amino acids required for
adult rats and adult humans differ
from those needed by growing rats
(Kim et al., 1997a,b,c) and humans
(Munroe, 2005), respectively. The
Munroe (2005) paper nicely outlines
a wide variety of aspects of amino
acid requirements and interactions.
Amino acid requirements for main-
tenance have been estimated from
the amounts of individual amino acid
required simply to maintain weight of
immature animals (Owens and Pet-
tigrew, 1989). Based on such measure-
ments with laboratory animals, the
pattern of amino acids required for
maintenance was related closely (r
= 0.97) to the amino acid composi-
tion of keratin tissues. In contrast,
the pattern of amino acids required
for growth was correlated with the
176
Figure 30. Amino acid composition of cow milk, muscle tissue, ruminal bacteria, and
keratin tissue as a percentage of CP. S-AA = total of sulfur-containing amino acids
(methionine plus cystine). Color version available in the online PDF.
amino acid composition of lean tissue.
With ruminants, rate of growth and
production typically is low relative
to that for the smaller, nonruminant
species. Because maintenance require-
ments are related to metabolic size, as
suggested from studies with nonru-
minants, the effect of maintenance
on total amino acid requirements is
greater for larger animals and the
proportion of amino acids devoted to
maintenance is highest when produc-
tion per unit of weight is lowest. It
seems ironic that the inefficiency of
converting dietary energy and protein
to animal products is criticized widely
by adult human males, the least ef-
ficient animals on our entire planet.
On a qualitative basis, amino acid
compositions of milk, muscle, and
keratin tissue differ markedly. Con-
version of one protein to another to
maintain essential tissues or produce
milk is inefficient because proportions
of amino acids among proteins differ.
Owens et al.
Note that keratin is very rich in sulfur
amino acids (cystine and methionine)
but low in lysine and histidine relative
to either milk or muscle tissue (Figure
30) and low in histidine relative to
bacterial protein.
Presumably, the priority order of
amino acid use is first for mainte-
nance (keratin synthesis), second for
milk, and third for building muscle
reserves. Efficiency of conversion of
one protein to another depends on the
degree that their amino acid composi-
tions match. For synthesis of keratin,
bacterial protein is markedly deficient
in sulfur amino acids (primarily cys-
tine), whereas muscle is short of histi-
dine. In contrast, conversion of muscle
tissue reserves to milk should be lim-
ited by threonine supply. Any excess
amino acids mobilized from tissues
are catabolized with N being excreted
in urine. Depending on specific con-
versions involved at a given stage of
growth or lactation, supplementation
with specific rumen-escape protein
sources or with rumen-escape amino
acids may improve N balance and re-
duce urinary N loss. But until amino
acids requirements and the supply of
absorbed amino acids are understood
quantitatively with greater certainty,
production responses to amino acid
supplement are likely to be haphazard
and inconsistent (Robinson, 2010) de-
spite frequent indications that rumen-
protected lysine (Polan et al., 1991)
or ruminally protected methionine can
increase milk protein concentrations.
Ideally, N-balance procedures should
be employed to ensure that responses
are increasing efficiency of protein
use. Some of the variation in response
to supplementation with rumen-pro-
tected amino acids in the past can be
attributed to technological problems
in achieving both ruminal escape and
intestinal amino acid release. The
change in the amino acid require-
ments at various stages of lactation
also has added to the variability in
response.
A Unifying Theory for Protein
Metabolism of Ruminants
Increasing the duodenal N sup-
ply through supplementation often
increases DMI and ruminant produc-
tion or performance. Although this
DMI response could be attributed to
recycling of supplemental N to the
rumen and to an increase in the rate
or extent of DM ruminal digestion,
protein content of duodenal liquid
(wet-matter basis) has proven to be
surprisingly constant based on sev-
eral studies in which diets divergent
in forage and protein content have
been fed (Bocheva et al., 1980; Zinn
and Owens, 1982). This could indi-
cate that duodenal flow and thereby
rumen liquid dilution rate may be
regulated by the quantity of soluble
protein present in chyme reaching the
small intestine. Ruminal dilution rate
in turn could be altered by regula-
tion of the reticular-omasal orifice
or by fluidity of ruminal contents.
Abomasal infusion of phenylalanine
increased ruminal NDF digestion of
a high-concentrate diet, presumably

through decreasing ruminal outflow
and passage of chyme to the small in-
testine (Vite et al., 2013). Regulation
of ruminal outflow could be advan-
tageous energetically for survival of
grazing ruminants. Slowly fermented,
high-NDF forages typically are low in
protein content; an extended time for
ruminal fermentation would increase
time for and extent to which OM is
fermented in the rumen and poten-
tially increase the postruminal supply
of microbial protein. Indeed, with
very-low-quality grazed winter range
grass, no free liquid could be detected
in the rumen of cows (John Wagner,
2013, Colorado State University, Fort
Collins, personal communication). In
contrast, rapidly fermented, grazed
forages typically are rich in protein
and low in NDF so that extending the
time for ruminal fermentation should
not be necessary to increase energy
availability for a grazing ruminant.
Cold stress, presumably via elevated
thyroid gland activity, will increase
DMI and rate of digesta passage.
As reviewed by Young (1981), the
increased DMI with cold stress often
more than compensates for decreased
digestibility associated with faster
passage through the digestive tract.
Consequently, regulation of ruminal
dilution via protein content of duo-
denal contents could increase com-
petitiveness of evolving ruminants.
In addition to other DMI regulation
schemes (gastro-intestinal fill; hepatic
oxidation; organoleptic compounds;
chemostasis and other satiety signals),
ruminants, through sensing protein
content of intestinal chyme, may
somehow impose feedback control on
time for fermentation of consumed
feeds. This unique capacity would
help ruminants use both low- and
high-quality feed components, con-
verting both into products desired by
humankind.
IMPLICATIONS
Increasing dietary CP concentration
curvilinearly increased milk produc-
tion and daily gain largely due to
increases in DMI. However, efficiency
Protein nutrition of ruminants
of conversion of dietary N to product
N decreases linearly as CP increases.
Microbial protein flow to the duode-
num increased with TDN intake, but
dietary TDN concentration was not
correlated with ruminal OM disap-
pearance. Per unit of DMI, duodenal-
ly measured DIP, UIP, and MP failed
to match estimates from the MP
system tested, presumably because
the effects of extent of ruminal OM
digestion, ruminal residence time, pro-
tozoal activity, and ruminal pH and
NDF digestibility are ignored. Specific
rumen-escape amino acids may simpli-
fy detection of deficiencies, but wide
diversity in composition of ruminant
diets prohibits broad extrapolation of
results. Unless combined with duode-
nal amino acid passage measurements,
responses to individually supple-
mented rumen-escape amino acids
are useless for quantifying amino acid
requirements for maintenance and
production of ruminants.
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