EFFECTS OF DIETARY POTASSIUM CARBONATE AND FAT CONCENTRATION IN HIGH
DISTILLER GRAIN DIETS FED TO DAIRY COWS
THESIS
Presented in Partial Fulfillment of the Requirements for the Degree Master of Science in
the Graduate School of The Ohio State University
By
Kathryn Cara Lamar, B.S.
Graduate Program in Animal Sciences
The Ohio State University
2013
Master’s Examination Committee
Dr. William P. Weiss, Advisor
Dr. Steven C. Loerch
Dr. Kristy M. Daniels
Copyright by
Kathryn Cara Lamar
2013
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ABSTRACT
Distiller grains with solubles (DGS) can induce milk fat depression when included
in dairy cow diets at greater than 20% DM. In vitro experiments have found that
potassium (K) supplementation with potassium carbonate (K2CO3) decreased
concentrations of biohydrogenation intermediates associated with milk fat depression
(MFD), such as trans-10, cis-12 conjugated linoleic acid (CLA). These intermediates are
often produced when diets are fed to cows with high concentrations of polyunsaturated
fatty acids, like those in DGS. We hypothesized that there would be an interaction
between level of K and level of fat. We hypothesized that adding K2CO3 to a high fat diet
based on DGS would alleviate MFD. We also hypothesized that the addition of K2CO3 to
a low fat diet based on DGS would have no effect on milk fat percent because these
diets would not cause MFD. Sixteen Holstein cows averaging 157 days in milk were
placed into 4 blocks; each block comprised a 4x4 Latin square with 21 d periods and a
2x2 factorial arrangement of treatments. The basal diet (no added K or fat) contained
27% DGS, 47% corn silage, 22% starch, 32% NDF, 4.2% long chain fatty acid, and 1.2% K
(DM basis). Treatments were 0 or 2.3% added fat from corn oil (in high fat diets, DGS +
corn gluten meal + corn oil = 27%) with 0 or 1% added K. Diets with added K had
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supplemental K in the form of K2CO3 from DCAD Plus® (Church & Dwight Co., Inc.,
Princeton, NJ). DCAD is a measure of the balance between cations and anions in the
diet. DCAD over 20 meq/100 g DM improves performance for lactating cows, while low
or negative DCAD benefit dry cows. Diets with added K had a DCAD of approximately 30,
while diets without added K had a DCAD of 2. This low DCAD may have limited
performance for cows fed diets without added K. Dry matter intake (DMI) decreased
with added fat (21.0 vs. 22.5 kg/d; P<0.01) and tended to decrease with added K (21.4
vs. 20.1 kg/d; P<0.06). Milk yield decreased with added fat (30.5 vs. 32.3 kg/d; P<0.01),
which may have been due to decreased DMI. No fat x K interaction was observed
perhaps because MFD occurred with all diets. Milk fat percent increased with added K
(2.82% vs. 2.56%; P<0.01) and decreased with added fat (2.51% vs. 2.89%; P<0.01). Milk
fat yield was affected similarly and tended to increase with added K (0.87 vs. 0.82 kg/d;
P<0.10) and decreased with added fat (0.76 vs. 0.93 kg/d; P<0.01). Trans-10, cis-12 CLA
in milk decreased with added K (P<0.02) indicating that the additional K 2CO3 was
decreasing incomplete biohydrogenation. Trans-10, cis-12 CLA increased with added fat
(P<0.01) because excess unsaturated fatty acids in the diet results in increased
incomplete biohydrogenation. Supplemental K2CO3 led to an increase in milk fat for both
high fat and low fat diets indicating that it could be used to alleviate MFD, though values
did not return to levels of a typical Holstein cow.
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ACKNOWLEDGEMENTS
Big thanks to Dr. Weiss for giving me a shot. Thanks for all the time you spent
reading over my proposal, work plan, and thesis, watching me practice presentations,
and teaching me how to be an all around better scientist and researcher. Thanks to Dr.
Loerch for being there when I needed help stringing words together for applications and
cover letters and for all of his help and support. Thanks to Dr. Kristy Daniels for all of
her advising while I was here and also for her help and support, as well.
Donna Wyatt, thank you so much for teaching me how to be in a lab. All of my
techniques and knowledge are because of you and I can’t thank you enough for that. I
will miss our talks greatly.
To Kevin and his farm crew, thank you for all the work you guys did for my
experiment.
To my family and friends, I can’t even begin to express how invaluable your love
and support were during this time. There’s no way I could have made it through all of
this without you.
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VITA
September 28, 1990………………………………………Born – Columbus, Ohio
May 2008………………………………………………………Gahanna Lincoln High School
2011………………………………………………………………B.S. Agriculture, The Ohio State
University
2011 to present………………………………………………Graduate Research, Animal Sciences,
The Ohio State University
FIELDS OF STUDY
Major Field: Animal Sciences
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TABLE OF CONTENTS
Abstract ................................................................................................................................ii
Vita .......................................................................................................................................v
Table of Contents ................................................................................................................vi
List of Tables ...................................................................................................................... vii
List of Figures .................................................................................................................... viii
Chapter 1: Literature Review .............................................................................................. 1
Introduction..................................................................................................................... 1
Sodium............................................................................................................................. 2
Sodium Absorption ...................................................................................................... 3
Sodium Deficiency ....................................................................................................... 4
Sodium Toxicity............................................................................................................ 5
Potassium ........................................................................................................................ 5
Potassium Absorption ................................................................................................. 7
Potassium Deficiency ................................................................................................... 8
Potassium Toxicity ....................................................................................................... 8
Chloride ........................................................................................................................... 9
Chloride Absorption..................................................................................................... 9
Chloride Deficiency .................................................................................................... 10
Chloride Toxicity ........................................................................................................ 11
Sulfur ............................................................................................................................. 11
Sulfur Absorption ....................................................................................................... 12
Sulfur Toxicity ............................................................................................................ 13
Dietary Cation Anion Difference ................................................................................... 13
Hypocalcemia ................................................................................................................ 14
Summary ....................................................................................................................... 17
Chapter 2: Introduction .................................................................................................... 19
Chapter 3: Materials and Methods ................................................................................... 22
Chapter 4: Results and Discussion .................................................................................... 28
References ........................................................................................................................ 63
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LIST OF TABLES
Table 1: Ingredient composition of diets, DM basis ......................................................... 39
Table 2: Nutrient composition of diets ............................................................................. 40
Table 3: Nutrient composition of corn silage ................................................................... 41
Table 4: Nutrient composition of distiller grains .............................................................. 42
Table 5: Effects of treatment on DMI, milk production, and milk composition, all 3 wk of
treatment .......................................................................................................................... 43
Table 6: Effects of treatment on DMI, milk production, and milk composition, wk 3 ..... 44
Table 7: Effects of treatment on milk fatty acid concentrations, all 3 wk ........................ 45
Table 8: Effects of treatment on milk fatty acid concentrations, 30 h into treatment .... 47
Table 9: Effects of treatment on milk fatty acid concentrations, d 21 of treatment ....... 49
Table 10: Effect of treatment on proportion of C 18:0 and C 18:2 relative to total
concentration of C 18 fatty acids ...................................................................................... 51
Table 11: Effect of treatment on estimated urine excretion (L/d) ................................... 52
Table 12: Effect of treatment on mineral intake and excretion (g/day) ......................... 53
Table 13: Effect of treatment on urine mineral excretion/mineral intake....................... 55
Table 14: Effect of treatment on mineral milk concentration, g/kg................................. 56
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LIST OF FIGURES
Figure 1: Effect of level of dietary fat and K on DMI by week within period. .................. 57
Figure 2: Effect of level of dietary fat and K on DMI over the entire period. ................... 58
Figure 3: Effect of level of dietary fat and K on milk production over the entire period . 59
Figure 4: Effect of level of dietary fat and K on milk fat yield over the entire period ...... 60
Figure 5: Effect of level of dietary fat and K on milk fat percent over the entire period . 61
Figure 6: Correlation of milk fat percent to trans-10 cis-12 concentration ..................... 62
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CHAPTER 1: LITERATURE REVIEW
Introduction
Dietary cation anion difference (DCAD) is the measure of the difference between
cations, sodium (Na) and potassium (K), and anions, chloride (Cl) and sulfur (S), in dairy
cows diets. These ions have the greatest effect on acid-base balance in the body. Adding
cations increases DCAD value and adding anions decreases the DCAD value. DCAD levels
in diets can affect many things including feed intake, acid-base levels in the body, blood
and urine pH. The DCAD is generally expressed as milliequivalents (mEq) of [(Na+ + K+) –
(Cl– + S2–)]/100 g of dietary DM. It can also be expressed as mEq/kg. The DCAD equation
DCAD = (Na+ + K+) - (Cl- + S2-) is the most commonly used form of the equation in dairy
cattle nutrition (Ender et al., 1962; Block, 1984). Lean et al., (2006) concluded that this
equation was the best for predicting milk fever incidence in dairy cows. Horst et al.
(1997) recommended that other anions and cations be included in the equation. He
proposed the equation DCAD = (0.38 Ca2+ + 0.3 Mg2+ + Na+ + K+) - (Cl- + S2-). Goff (2000)
proposed a variation of this equation. His equation took into account the capacity of
different salts to acidify urine and recommended DCAD = (0.15 Ca2+ + 0.15 Mg2+ + Na+ +
K+) - (Cl- + 0.25 S2- + 0.5 P3-). Tucker et al. (1991) suggested that the DCAD equation
should be DCAD= (0.38 Ca2+ + 0.3 Mg2+ + Na+ + K+) - (Cl- + 0.6 S2- + 0.5 P3-) based on the
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research of Spears et al. (1985). The DCAD equation used in this thesis will be DCAD =
(Na+ + K+) - (Cl- + S2-).
Sodium
Sodium is the primary extracellular cation (Aitken, 1976). Heart function and
nerve impulse conduction and transmission are dependent on the proper balance of Na
and K. Sodium is also involved in the sodium-potassium pump, which enables transport
of glucose, amino acids, and phosphate into cells, and hydrogen, calcium, bicarbonate,
K, and Cl ions out of cells (Lechene, 1988). Sodium is a major component of salts in
saliva, which buffers acid from ruminal fermentation (Blair-West et al., 1970).
Milk production can increase with addition of sodium bicarbonate (NaHCO3).
Sodium bicarbonate produced small increases in DMI and milk yield (MY) (Canale and
Stokes, 1988). Sodium bicarbonate increased ruminal pH and MY response was maximal
with 0.70% Na and 1.58% K (Stokes and Bull, 1986). The addition of Na2CO3 at 0.78%
increased milk fat percent, age and yield, and 4% fat corrected MY (Belibasakis and
Triantos, 1991). Though, in one commercial herd where diets were based on alfalfa hay,
0.8% NaHCO3 reduced MY in second lactation and older animals (Canale and Stokes,
1988). Dairy cattle absorb dietary Na very efficiently, but only very small amounts are
stored in a form that is readily available for metabolism. Concentration in milk is
between 25 and 30 mEq/L. Sodium concentration increases during mastitis when serum
leaks into milk, but is not significantly affected by dietary Na content (Kemp, 1964)
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Typical concentrations of Na in blood plasma are 150 mEq/L and 160 to 180 mEq/L in
saliva.
Empirical modeling of data from 15 experiments with lactating cows conducted
in either cool or warm seasons showed that DMI and MY were improved by dietary
concentrations of Na well above those needed to meet requirements (Sanchez et al.,
1994a,b). Dry matter intake (DMI) and MY responses over a range of dietary Na
concentrations from 0.11 to 1.20% DM were curvilinear, with maximum performance at
0.70 to 0.80% DM. In hot weather, MY and DMI increased when Na was supplemented
from 0.18% Na to 0.55% total dietary Na with either NaCl or NaHCO3 (Schneider et al.,
1986).
Sodium Absorption
Agricultural Research Council (1980) estimated that 91% of Na consumed by
cattle was absorbed. Apparent absorption of Na by dairy cows fed fresh forages ranges
from 77 to 95%, with an average of 85% (Kemp, 1964). Sodium chloride (NaCl) is most
often used and the Na in NaCl is essentially 100% available. Sodium absorption occurs
throughout the digestive tract by an active transport process in the reticulorumen,
abomasum, omasum, and duodenum. Passive absorption occurs through the intestinal
wall. Substantial active absorption against a sizable concentration gradient also occurs
in the lower small intestine and large intestine (Renkema et al., 1962). Sodium
concentrations in blood and tissues are maintained principally via reabsorption and
excretion by the kidneys. There is close synchrony between the excretion of Na, K, and
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Cl. Sodium is the central effecter of ion excretion and changes in renal resorption
determine Na excretion. Endocrine control via tissue receptors and the reninangiotensin system, aldosterone, and atrial natriuretic factor monitor and modulate Na
concentrations in various tissues, which consequently control fluid volume, blood
pressure, K concentrations, and renal processing of other ions. Kidneys are efficient in
reabsorbing Na when dietary Na is deficient.
Sodium Deficiency
When Na is deficient, it is decreased in saliva and is reabsorbed in the kidneys.
Mallonee et al. (1982a) found when feeding a diet without supplemental NaCl (0.16%
Na) feed intake and MY began to decline within 1 to 2 wk. Pica and drinking of urine of
other cows were also observed (Mallonee et al., 1982a). Although dietary Cl
concentration was not measured in the study, potassium chloride (KCl) was
supplemented (1.0% DM KCl), so Cl deficiency was not the cause of the condition.
Babcock (1905) fed a diet very low in Na to dairy cows and described intense craving for
salt and general pica. Other deficiency signs include loss of appetite, rapid loss of body
weight, haggard appearance, lusterless eyes, and rough hair coat (Underwood, 1981).
More extreme signs of deficiency include loss of coordination, shivering, weakness,
dehydration, and cardiac arrhythmia leading to death. Feeding lactating dairy cows a
diet with no supplemental NaCl (0.16% Na DM) resulted in marked depressions in DMI
and MY after just 1 to 2 wk of feeding (Mallonee et al., 1982a).
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Sodium Toxicity
The National Research Council (2001) states that the maximum tolerable dietary
concentration of NaCl is 4% for lactating cattle, which equals about 1.6% Na. (National
Research Council, 2005). High intakes of NaCl can lead to an increase of incidence and
severity of udder edema (Randall et al., 1974). Toxicity signs included severe anorexia,
reduced water intake, dehydration, weight loss, and ultimately physical collapse.
Feeding diets with 0.88% Na from NaCl or NaHCO3 to mid-lactation Holstein cows did
not cause toxicity or reduce feed intake and MY compared with 0.55 % Na (Schneider et
al., 1986). With an adequate supply of clean drinking water, cattle can tolerate large
quantities of dietary NaCl. Jaster (1978) provided drinking water with 0 or 2.5 g/L NaCl
for a 28-d period to lactating cows and MY declined and water consumption increased.
Cattle drinking water that contained 0.7 to 1.5 mEq/L NaCl suffered from toxicosis
(Weeth et al., 1960; Weeth and Haverland, 1961).
Potassium
Potassium is the main intracellular cation and is the third most abundant mineral
element in the body. It must be supplied daily in the diet because there is little storage
in the body and the animal’s requirement for K is highest of all the mineral element
cations. Milk contains about 38.5 mEq/L K. Saliva typically contains 10 mEq/L, whereas
concentrations in ruminal fluid range from 40 to 100 mEq/L. Blood plasma contains 5 to
10 mEq/L K per liter. The majority of K in blood is located within red blood cells (Aitken,
1976; Hemken, 1983). Early research indicated that 0.70 to 0.75% dietary K was
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sufficient to meet requirements of early and mid- to late-lactation cows (Dennis et al.,
1976; Dennis and Hemken, 1978; Erdman et al., 1980), though the requirement is now
set at 1.0% DM (National Resource Council, 2001).
Potassium is involved in osmotic pressure and membrane potential, acid-base
regulation, water balance, nerve impulse transmission, muscle contraction, oxygen and
carbon dioxide transport; phosphorylation of creatine, pyruvate kinase activity, as an
activator or co-factor in many enzymatic reactions, cellular uptake of amino acids and
synthesis of protein, carbohydrate metabolism, and in maintenance of normal cardiac
and renal tissue (National Research Council, 2001; Stewart, 1981; Hemken, 1983).
Forages are generally higher in K than grains (National Research Council, 2001).
In an empirical modeling of data with 1,444 cow-period observations, DMI and
MY responses over a range of dietary K concentrations from 0.66 to 1.96 %, results were
curvilinear, with maximum performance when diets contained 1.50% K in the cool
season. In the warm season, DMI and MY increased as K% DM increased (Sanchez et al.,
1994a,b). Mallonee (1984) found no benefit of increasing dietary K from 1.07 to 1.58%
on feed intake or lactational performance of mid-lactation Holstein cows. Feed intake
and MY were reduced with the 4.6% K, and water intake, urinary excretion, and total K
excretion were increased with increasing concentrations of K in a study by Fisher et al.
(1994). Many studies have found that feeding higher concentrations of dietary K than
needed to meet National Research Council (2001) recommendations of lactating cows in
thermoneutral environments increased feed intake and MY compared with cows fed
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lower dietary concentrations (Beede et al., 1983; Schneider et al., 1984; Mallonee et al.,
1985; Schneider et al., 1986; West et al., 1987; Sanchez, 1994a). A dietary K
concentration of 1.5% DM during heat stress maximized lactational performance (Beede
and Shearer, 1991). Scheider et al. (1986) showed greater production from heatstressed cows if dietary K levels were above NRC recommended levels.
Potassium Absorption
Potassium in feeds exists as simple ions, which are readily available for
absorption (Emanuele and Staples, 1990, 1991; Ledoux and Martz, 1990). Hemken
(1983) indicated that K is almost completely absorbed with a true absorption of 95% or
greater for most feedstuffs. Average apparent absorption of K in eight forages fed to
cattle and sheep was 85% (Miller, 1995). An absorption coefficient value of 90% for K is
used for all types of feedstuffs and mineral sources of K (National Research Council,
2001). Potassium is mainly absorbed from the duodenum by simple diffusion, though
some absorption also occurs in the jejunum, ileum, and large intestine. The main
excretory route of excess absorbed K is via the kidneys. This route is lower
concentrations of K in plasma and milk, higher blood hematocrit reading, and overall
primarily under regulation by aldosterone, which increases Na resorption in the kidney
with the concomitant excretion of K. Blood acid-base status also affects urinary
excretion of K (McGuirk and Butler, 1980). With the onset of an alkalotic condition,
intracellular protons are exchanged with K in blood plasma as part of the regulatory
mechanisms to maintain acid-base equilibrium and blood pH, reducing K in blood. A
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large gradient exists between intracellular renal tubule concentrations of K and that of
urine. This gradient affects the passage of K from the tubular cells into urine.
Potassium Deficiency
Signs of K deficiency include a decrease in feed and water intake, reduced body
weight and MY, pica, loss of hair glossiness, decreased pliability of the hide, muscle
weakness. Rate and severity of deficiency appeared to be related to milk production,
where higher producing cows were affected more quickly and severely than lower
producing cows. Signs of severe K deficiency were manifested in lactating dairy cattle
fed diets with less than 0.15 % K (Pradhan and Hemken, 1968; Mallonee et al., 1982b).
In a trial with mid-lactation cows, 0.42% dietary K reduced DMI and MY; however, no
differences in DMI or MY were noted for cows consuming diets with 0.69 or 0.97 % K
DM (Dennis and Hemken, 1978). Severe K deficiency can occur in diets with 0.06 to
0.15% K (Pradhan and Hemken, 1968; Mallonee et al., 1982b).
Potassium Toxicity
Absorbed K in excess of requirements is excreted mainly in urine. Feeding dietary
K above requirement can reduce magnesium absorption and may cause udder edema.
The maximum tolerable concentration in the diet for dairy cattle is 3.0% of DMI
(National Research Council, 2001). When 4.6% dietary K via supplemental potassium
carbonate (K2CO3) was fed to cows during early lactation, DMI and MY were reduced,
and water intake, urinary excretion, and total K excretion were increased (Fisher et al.,
1994).
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Chloride
Depending on MY, the dietary requirement for Cl is between 0.25%-0.30% of
dietary DM. Typical concentrations of Cl in blood plasma are between 90 and 110 mEq/L
and 10 to 30 mEq/L in ruminal fluid. During lactation, Cl concentration is highest in
colostrum, declines to normal levels, then rises again at the end of lactation and is
present in milk at 32.5g/L on average (Agricultural Research Council, 1965). Chloride is
the most common anion in extracellular body fluids of mammals (Fettman et al, 1983),
making up more than 60 % of the total anion equivalents in extracellular fluid. As such,
Cl is also the major anion in gastric secretions in the form of hydrochloric acid (HCl),
which is also known as gastric acid and is needed for digestion. Gastric acid is necessary
for the activation of pepsin, which is required for protein digestion. Chloride is also
essential for the activation of pancreatic amylase and is found in large concentration in
bile and other intestinal juices (Phillipson, 1977). About 80 % of the Cl in the digestive
tract arises from digestive secretions in saliva, gastric fluid, bile, and pancreatic juice.
Chloride Absorption
Chloride is absorbed throughout the digestive tract. The absorption coefficient
for Cl from both feedstuffs and mineral sources is approximately 90% for dairy cattle
(Henry, 1995c). The Cl from HCl is absorbed in the small intestine by passive diffusion
along an electric gradient by exchange with bicarbonate (Tucker et al., 1987). Chloride is
transported across the ruminal wall to blood against a wide concentration gradient
(Sperber and Hyden, 1952). Chloride is co-transported actively with Na across the rumen
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wall (Martens and Blume, 1987). Excess Cl is excreted mainly in urine and feces as NaCl
or KCl (Sanchez et al., 1994b). Regulation of the concentration of Cl in extracellular fluid
and its homeostasis is coupled with Na. Chloride excretion is also influenced by the
bicarbonate ion. If blood bicarbonate rises, a similar amount of Cl is excreted by the
kidneys to maintain systemic acid-base balance.
Chloride Deficiency
Babcock (1905) offered free choice KCl instead of NaCl to a cow fed a diet with
no supplemental NaCl and the cow ate a considerable amount of it, suggesting that the
deficiency was for Cl, not NaCl. Though, it could also suggest that cattle preferred the
taste of KCl to NaCl. Experiments that fed diets with 0.18% Cl (i.e, less than the current
NRC requirement) found that cows conserved Cl by dramatically reducing excretion of Cl
in urine, feces, and milk (Coppock et al., 1979; Fettman et al., 1984). Coppock et al.,
(1979) also found that cows fed a low Cl diet consumed more salt block than cows fed
0.40% Cl. Fettman et al. (1984) found that cows fed 0.10 % Cl rapidly exhibited clinical
signs of deficiency and poor performance compared with those fed medium and high
concentrations of dietary Cl. Clinical signs of Cl deficiency are reduced appetite, weight
loss, lethargy, emaciation, decreased lactation, constipation, cardiovascular depression,
excessive thirst, and excessive urination. In advanced stages, cows can suffer from
severe eye defects, reduced respiration rates, and blood and mucus in feces.
Metabolically, Cl deficiency resulted in severe metabolic alkalosis and low blood Cl
which can result in low blood Na and K.
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Chloride Toxicity
Maximum tolerable dietary concentration of Cl is 4.5% for non-lactating cows
and 3.0% for lactating cows (National Research Council, 2005). Negative effects of
excess dietary Cl include decreased DMI and MY, with effects being more pronounced in
summer weather than in winter (Sanchez et al., 1994a). Empirical models with 1,444
cow-period means showed that increasing dietary Cl from 0.15 to 1.62% decreased DMI
and MY of mid-lactation cows (Sanchez et al., 1994a). Negative effects of increased
dietary Cl were more dramatic in summer weather than in winter (Sanchez et al.,
1994a). This is consistent with the results of Escobosa et al. (1984) showing profound
exacerbating effects of high dietary Cl on acid-base balance and milk production during
heat stress.
Sulfur
The S requirement for dairy cows is 0.20% of dietary DM. The dietary
requirement of S provides enough substrate to ensure maximal microbial protein
synthesis (Bouchard and Conrad, 1973 a, b). Sulfur is found in the amino acids
methionine, cysteine, homocysteine, and taurine (National Research Council, 2001). It is
also present in the B-vitamins, thiamin and biotin. Non-protein nitrogen, such as urea,
added to diets cannot be incorporated into microbial protein unless adequate S is
present for cysteine and methionine formation. The strong reducing environment within
the rumen can reduce dietary sulfate, sulfite, and thiosulfate to sulfide (Lewis, 1954).
High concentrations of dietary S can decrease feed intake and overall performance of
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ruminants. While in the digestive tract, hydrogen sulfide gas, hydrogen sulfide in
solution, sulfur dioxide gas, and pentathionic acid are produced (Lis, 1983). Sulfide
absorbed from the rumen can be detoxified by oxygenated hemoglobin in the blood
(Evans, 1967) and the liver through the sulfide oxidase system (Anderson, 1956).
Not all bacteria in the rumen utilize all forms of S (Emery et al., 1957a).
Elemental S is not well utilized by many ruminal bacteria (Ishimoto et al., 1954). Emery
(1957a) and Emery et al. (1957b) reported that ruminal microbes produce twice as much
cysteine as methionine from inorganic sulfate. Bryant (1973) found that the
predominant ruminal cellulolytic bacteria, Fibrobacter succinogenes, could utilize sulfide
or cysteine but not sulfate. Many strains of Ruminococcus grew in media containing only
sulfide or sulfate-sulfur (Bryant, 1973).
Sulfur Absorption
Sulfur incorporated into microbial protein is absorbed from the small intestine as
cysteine and methionine. Some dietary S is absorbed as the sulfate or sulfide anion.
Sulfate-sulfur is absorbed more efficiently in the small intestine than other sources of S
(Bird and Moir, 1971). Elemental S is much less available, probably because it is not very
soluble (Fron et al., 1990). Lignin sulfonate is also a poorly utilized source of S (Bouchard
and Conrad, 1973a). The sulfur-containing amino acids provide a major dietary source of
S for the ruminal microbes. Protection of proteins and amino acids from ruminal
degradation could result in less S being available for microbial protein synthesis in the
rumen, but will help the cow obtain amino acids required for her tissues. Methionine,
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methionine analogs, and sulfate salts are utilized equally well in meeting the dietary S
requirements of the cow and ruminal microbes (Bouchard and Conrad, 1973a, b; Bull
and Vandersall, 1973; Thomas et al., 1951).
Sulfur Toxicity
Studies by Beke and Hironak (1991) and Mcallister et al. (1997) found that
polioencephalomalacia, a neurological disease associated with thiamine deficiency,
occurs in beef cattle fed greater than 0.5% S. For dairy cattle, the NRC set maximum
dietary S levels at 0.3% for high starch diets and 0.5% for high forage diets (National
Research Council, 2005). Symptoms of high concentration of dietary S are severe watery
diarrhea, respiratory distress, muscle twitching, severe dehydration, congested lungs,
acute enteritis, abdominal pain, and strong odor of hydrogen sulfide on the breath (Lis,
1983). Neurotoxic effects of sulfide are caused by eructation of hydrogen sulfide along
with other gasses from the rumen. These gasses are then absorbed through the lungs
(Bird, 1972). Sulfates are less toxic, though they can cause an osmotic diarrhea and
excess sulfate added to rations can reduce feed intake and performance (Kandylis,
1984).
Dietary Cation Anion Difference
Tucker et al. (1988) found that cows fed DCAD 20 mEq/100 g of DM yielded 9%
more milk than those fed a DCAD of -10 mEq/100 g of DM. West et al. (1991) found in
both hot and cool environments, increasing DCAD from -12 to 31 mEq/100 g of DM
increased DM, MY, 4% FCM, and milk protein. Delaquis and Block (1995) reported that
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increasing DCAD from 6 to 26 mEq/100 g of DM increased milk production in early and
mid-lactation, but not in late lactation. Wildman et al. (2007) fed diets providing DCAD
of 25 or 50 mEq and higher DCAD improved DMI, MY, and concentrations of milk fat and
protein. Erdman (2011) and Hu and Murphy (2004) found that maximal MY and DMI
occurred at DCAD of 34-40 mEq/100 g of DM. In a study by Harrison, et al. (2012),
increased milk production was achieved at a DCAD of 53 mEq/100 g of DM versus a
DCAD of 32 mEq/100 g of DM.
Hypocalcemia
The transition period in dairy cow, as defined by Grummer (1995), is the three
wk before and after parturition. The transition period is considered the most difficult
time for a dairy cow, determining the cow’s health, production, and reproduction in the
subsequent lactation (Keady et al., 2001). During the transition period, the clearance of
Ca to the placenta ceases, but the lactational Ca demand increases rapidly (Roche et al.,
2003b). Though milk fever affects only a small percentage of cows, nearly all cows
experience some decrease in blood calcium during the first days after calving, while
their intestines and bones adapt to the calcium demands of lactation (Ender et al. 1971;
Ramberg, 1974).
Nearly all high producing dairy cows will undergo some degree of hypocalcemia
within the first 2 d of parturition and subsequent onset of lactation (Ramberg et al.,
1984). Daily body turnover of Ca changes from approximately 10 g in non-lactating cows
to greater than 30 g in lactating cows (Horst et al., 1997). In cows with hypocalcemia,
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the calcium homeostatic mechanisms, which normally maintain blood calcium
concentration between 9 and 10 mg/dL, fail and the lactational drain of calcium causes
blood calcium concentration to fall below 5 mg/dL. Normal physiological levels of
plasma calcium range from 8.5 to 11.5 mg/dL and a normal decrease in plasma calcium
of 2 mg/dL is expected at calving (Niedermeier et al., 1949). Hypocalcemia may be
classified as subclinical, with levels from 7.5 to 8.5 mg/dL, or clinical, with levels from 5.0
to 6.0 mg/dL (Jorgensen, 1974). During the dry period calcium requirements are minimal
and the mechanisms in place to replace calcium lost from the plasma pool are relatively
inactive (Ramberg et al., 1984).
Hypocalcemia can impair muscle and nerve function enough so that cows are
unable to rise. Cows that have had milk fever are more susceptible to other disorders
such as lack of appetite, mastitis, displaced abomasum, retained placenta, and ketosis
(Curtis et al., 1985). Hypocalcemia at calving is a predisposing factor for dystocia,
prolapsed uterus, retained placenta, and early metritis (Erb and Grohn, 1988; Grohn et
al., 1989). Animals suffering from clinical hypocalcemia exhibit symptoms such as
decreased appetite, tetany, inhibition of urination and defecation, lateral recumbency,
and eventual coma and death if untreated (Horst et al., 1997). Milk production may
suffer long after the transition period has passed (Block, 1984).
The adaptation to the onset of lactation during the critical first days of lactation
is accomplished by release of parathyroid hormone (PTH), which reduces urinary
calcium losses, stimulates bone calcium resorption, and increases 1,25-dihydroxyvitamin
15
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D synthesis to enhance active intestinal transport of calcium. All three must be
operational if hypocalcemia is to be minimized. Milk fever risk factors reduce the
efficiency of one or more of these homeostatic mechanisms. By carefully regulating the
amount of 1,25-dihydroxyvitamin D produced, the amount of dietary calcium absorbed
can be adjusted to maintain a constant concentration of extracellular calcium (DeLuca,
1979; Bronner, 1987).
Metabolic alkalosis impairs the physiologic activity of PTH so that bone
resorption and production of 1,25-dihydroxyvitamin D are impaired reducing the ability
to successfully adjust to the calcium demands of lactation (Block, 1984; Block, 1994;
Gaynor et al., 1989; Goff et al., 1991; Phillippo et al., 1994). Evidence suggests that
metabolic alkalosis induces conformational changes in the PTH receptor, which prevents
tight binding of PTH to its receptor.
Prepartum DCAD dramatically affects Ca metabolism at parturition (Oetzel et al.,
1988; Goff et al., 1991). Diets with a low DCAD alter Ca homeostasis and increase
plasma Ca concentration at parturition, which helps prevent hypocalcemia. Goff et al.
(1991) observed that Jersey cows fed anionic diets had an increased 1, 25-(OH) 2 D
response per unit of decline in Ca in serum. Anionic diets increase the efficiency of Ca
absorption from the gastrointestinal tract (Lomba et al., 1978). Studies demonstrated
that addition of anions to the prepartal diet could prevent milk fever (Ender et al., 1971;
Block, 1984), though Hu and Murphy (2004) found no evidence of a direct relationship
between blood Ca concentration and DCAD postpartum. Higher blood Ca concentrations
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were reported for cows fed anionic diets (Block, 1984; Joyce et al., 1997) as acidogenic
diets increase serum Ca by increasing Ca mobilization from bone.
Summary
Dietary DCAD can have a large effect on dairy cows. For lactating cows, high
levels of DCAD, from 30-40 mEq/100 g DM, can lead to an increase in DMI, milk
production, and yield of milk components. For dry cows, a low or negative DCAD can
decrease the risk of hypocalcemia, or milk fever, which decreases the risk of other
significant illnesses as well. DCAD affects overall body acid-base balance, which affects
the overall health of a dairy cow and balance of other minerals in the body, as well.
Understanding of the effects of the balance between the cations and anions in dairy
cows can be beneficial for dairy farmers.
Potassium is one of the cations used to calculate DCAD. In dairy cows, it has
recently been reported that addition of K in the form of K2CO3 can increase milk fat
production by decreasing the incomplete ruminal biohydrogenation caused by
unsaturated fat in the diet. We hypothesized that supplementing K 2CO3 to a diet with a
high concentration of fat from DGS would alleviate MFD, but would have no effect when
supplemented to a diet with lower concentrations of fat because these lower fat diets
would likely not induce MFD. Further, we hypothesized that the positive effect of K2CO3
would be caused by reducing the production of trans-10, cis-12 CLA caused by
incomplete biohydrogenation of the dietary PUFA. The objective of this study was to
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determine whether supplemental K2CO3 could alleviate the decrease in milk fat percent
caused by an increase of unsaturated fat in a diet with DGS.
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CHAPTER 2: INTRODUCTION
Dried distiller grains with solubles (DGS) are a byproduct of the ethanol industry
and are traditionally fed as a protein supplement. Distiller grains with solubles are also
an excellent source of energy due to their high concentration of digestible NDF and fat,
specifically C 18:2 (Schingoethe et al., 2009). Griinari et al. (1998) demonstrated severe
MFD when unsaturated fat from corn oil, which is similar to the fat in DGS, was added to
diets that contained 14.8 vs. 32.1% NDF. Leonardi et al. (2005) found slight, linear
decreases in milk fat content as DGS increased from 0 to 15% of diet DM. In a study by
Kalscheur (2005), milk fat content was lower only in DGS diets that contained less than
50% forage and 22% forage NDF. Experiments by Abdelqader et al., (2008) and Leonardi
et al. (2005) indicated that the effects of adding corn oil to diets is similar to those when
fat from DGS is added, when levels of dietary fat are similar. Diets containing high
concentrations of polyunsaturated fatty acids (PUFA), such as that found in DGS, can
depress milk fat content. The predominant PUFA in dairy cow diets are linolenic, C 18:3,
and linoleic, C 18:2, acids in plant lipids. Milk fat depression (MFD) occurs when milk fat
yield is reduced, but milk volume and other components are not affected (Peterson, et
al. 2003). Milk fat depression caused by abomasal infusion of trans-10, cis-12 CLA can
decrease milk fat percent within 10 hours (Harvatine and Bauman, 2007b). Diet-induced
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MFD develops over 7 to 18 days with the lowest point of milk fat depression occurring 18
days into treatment (Shingfield et al., 2006b).
Diets that induce MFD are known to alter ruminal lipid metabolism, leading to
an increased production of specific trans and conjugated fatty acid isomers that are
absorbed in the lower gastrointestinal tract and inhibit milk fat synthesis in lactating
dairy cows (Shingfield and Griinari, 2007; Shingfield et al., 2010; Maxin et al., 2011).
Under some conditions of diet-induced MFD, rumen production, and milk fat content of
trans-10, cis-12 conjugated linoleic acid (CLA) increases (Harvatine et al., 2009;
Shingfield et al., 2010; Maxin et al., 2011).
The addition of potassium carbonate (K2CO3) may alleviate the negative effects
of the unsaturated fats in corn oil. Jenkins et al. (2011) found that the addition of K2CO3
to cultures of mixed ruminal microorganisms decreased production of trans-C18:1 and
trans-10, cis-12 CLA (Jenkins et al., 2011). An in vitro study by Morris et al. (2012)
showed that incomplete biohydrogenation induced by unsaturated fatty acid was
alleviated by addition of 3% potassium (K) in the form of K2CO3, but not by addition of
3% K in the form of KCl, showing that K2CO3 was better at reducing incomplete
biohydrogenation than KCl. Harrison et al. (2012) evaluated the dietary K requirements
using K2CO3. Diets included K at levels of approximately 1.3% and 2.1% DM. The study
found that diets with 2.1% K had increased dry matter intake (DMI), milk fat percentage,
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milk yield (MY), and efficiency of milk production per unit of DMI when compared to
cows fed the diet with 1.3% K. Though, MFD did not occur in this experiment.
We hypothesized that supplementing K2CO3 to a diet with a high concentration
of fat from DGS would alleviate MFD, but would have no effect when supplemented to a
diet with lower concentrations of fat because the low fat diets would not induce MFD.
Further, we hypothesized that the positive effect of K2CO3 would be caused by reducing
the production of trans-10, cis-12 CLA caused by incomplete biohydrogenation of the
dietary PUFA.
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CHAPTER 3: MATERIALS AND METHODS
Animals and Diets
All procedures involving animals were approved by the Institutional Animal Care
and Use Committee of The Ohio State University. Twelve multiparous (157+8 days in
milk (DIM), 37+2 kg/d MY) and four primiparous (156+4 DIM, 29+1 kg/d MY) Holstein
cows were used. Multiparous cows were blocked by MY. Each block comprised a 4x4
Latin square with 21 d periods and a 2x2 factorial arrangement of treatments. Cows
were fed one of four diets (Table 1). Treatments were 0 or 1.9% added K2CO3 (DCAD
Plus®; Church & Dwight Co., Inc., Princeton, NJ) with 4.2 or 5.8% long chain fatty acids
(LCFA) (Table 2). All diets contained 47% corn silage (Table 3) and 27 % DGS (Table 4) or
its equivalent from a mix of DGS, corn gluten meal, and corn oil. The mixture of DGS,
corn gluten meal, and corn oil was used rather than a source of DGS with a higher fatty
acid concentration because it minimized differences in nutrient composition between
treatments that may have occurred if we had simply purchased a higher fat DGS to use
for our high fat diets. Treatments were designated as low fat without added K (LF-K),
low fat with added K (LF+K), high fat without added K (HF-K), and high fat with added K
(HF+K). The inclusion rate for DGS in this experiment was chosen because previous
research found decreased milk fat percentage and yield when cows were fed diets with
20% DGS (Hippen et al., 2004). For our experiment, K2CO3 included at 1.9% increased
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dietary K by 1%. When K2CO3 was added to the diet, soybean hulls were decreased
because this had the most minimal effect on differences of NE L between treatments.
Diets were formulated to meet nutrient requirements for 650 kg lactating dairy
cows producing 40 kg milk/d (National Research Council, 2001). Cows were housed in tie
stalls and fed once daily (0400h). Diets were offered ad libitum (feed refusals averaged
9.2%, as fed basis) as mixed rations. Amount of feed offered and refused were
measured daily and were used to determine DMI. Diets were adjusted weekly to
account for changes in corn silage DM concentration. Cows were milked twice daily
(0200 and 1300 h) and milk weights were recorded electronically at each milking. Cows
were body condition scored (1= emaciated; 5=obese) by two independent people
(averaged) at the beginning of the experiment and on d 21 of each period. Cows were
weighed at the beginning and end of the experiment and on d 21 of each period at 0800
h. Body weight change was calculated as the difference between the end of the period
weight and the end of the previous period weight.
Sampling Collection and Analysis
Samples of fresh feeds were collected weekly and composited by period. Orts
(10% of wet weight) were taken once a wk, stored in the freezer for up to 2 wk, and
composited within cow and period. Samples of DGS were collected from the feed facility
once per period. Weekly samples of silage and refusals were analyzed for DM (100°C for
48 h). A composite of corn silage and each refusal sample were dried at 55°C and ground
in a Wiley mill (Arthur H. Thomas Co., Philadelphia, PA) through a 1-mm screen before
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nutrient analyses. Grains were not ground or oven-dried. Samples were analyzed for
DM (100°C oven for 24 h), ash (AOAC, 2000), starch (Weiss and Wyatt, 2000), NDF
(Ankom200 Fiber Analyzer, Ankom Technology, Fairport, NY) with sodium sulfite and
amylase (Ankom #FAA), CP (Kjeldahl N x 6.25, (AOAC 984.13.4.2.09, 2000), and LCFA
(Weiss and Wyatt, 2003).
Initial analysis of concentrate samples with added K resulted in starch levels
higher than expected based on formulation and the concentrations in the other
concentrate mixes. The pH of the assay solutions containing samples of the K
supplemented mixes after the 60 min water bath incubation at 90°C were 3 pH units
higher than samples without added K (Table 1). This higher pH was a result of the
additional K and was outside the optimal pH range for the alpha amylase. Several pH
comparisons found that an addition of 50 uL 2M HCl lowered the pH so that proper
starch analysis could occur.
Silage and refusal samples were dry-ashed and concentrate samples were
perchloric acid digested. After digestion, mineral analyses were conducted using an
inductively coupled plasma spectrograph (Service Testing and Research [STAR]
Laboratory, Ohio Agricultural Research and Development Center, Wooster, OH).
Chloride concentrations were analyzed by extraction with 0.5% nitric acid followed by
potentiometric titration with silver nitrate using a Brinkman Metrohm 848 Titrino Plus.
(Brinkmann Instruments Inc., Westbury, NY) by Cumberland Valley Analytical Services
(CVAS; Hagerstown, MD). Samples of the four total mixed rations (TMR) were
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constructed from dried samples of forages and concentrates and were analyzed for 30-h
in vitro NDF digestibility (Goering and Van Soest, 1970) by CVAS. Corn silage particle size
distribution was measured on wet period composite samples using a Penn State Particle
Separator (PSPS) (Lammers et al., 1996). Particle size distributions of concentrate mix
samples were measured by dry sieving using a vertical oscillating sieve shaker
(Analysette 3; Fritsch, Oberstein, Germany) equipped with a stack of sieves (W. S. Tyler,
Inc., Mentor, OH) arranged in descending mesh size. Sieve mesh sizes were 1.18, 0.6,
0.3, and 0.15 mm. The DCAD value of feeds was determined using the equation DCAD =
[(%Na × 43.5 + %K × 25.6) − (%Cl × 28.2 + %S × 62.5)] (Ender et al., 1962). Drinking water
was collected from the farm once per period and was analyzed for mineral content
using an inductively coupled plasma spectrograph (STAR Laboratory).
Milk samples (a.m. and p.m. milkings) were collected on d 3, 7, 10, 14, 17, and 21
of each period for determination of milk, fat, protein, lactose, (B2000 Infrared Analyzer
(Bentley Instruments, Chaska, MN) and MUN concentrations (Skalar SAN Plus
segmented flow analyzer; Skalar Inc., Norcross, GA) by DHI Cooperative, Inc. (Columbus,
OH). Milk yields from d 3, 7, 10, 14, 17 and 21 were used to calculate yields of milk
components. The energy concentration of milk was calculated from milk fat, protein,
and lactose (NRC, 2001).
Milk samples (p.m.) were collected 30 h after the start of each period, and d 7,
14, and 21. Samples were stored at 4°C up to 24 hours until milk fat removal. Samples
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were centrifuged at 20,000xg for 20 min at 4°C. The fat layer was skimmed off and
stored at -20°C until milk fat analysis by gas chromatography.
Milk samples (a.m.) were collected on d 21 for mineral analysis. The samples
were stored at 4°C up to 1 wk until analysis. Samples were warmed by heating at 37°C
for 15 minutes and homogenized by repeated (10x) pouring into a beaker. A 2-mL
aliquot was digested with nitric acid and analyzed for mineral content using an
inductively coupled plasma spectrograph (STAR Laboratory).
Urine samples (~100 mL) were collected by vulva stimulation on days 16 and 17
from each cow. Individual samples were analyzed and then averaged by cow by period.
Samples were stored at -20°C until analysis. Thawed samples were warmed by heating
at 37°C for 15 min and homogenized by repeated (10x) pouring into a beaker. A 5-mL
aliquot was nitric acid digested and analyzed for minerals using an inductively coupled
plasma spectrograph (STAR Laboratory). Samples were also analyzed for creatinine
(Cayman Chemical Item Number 500701, Ann Arbor, MI) and Kjeldahl N (AOAC
984.13.4.2.09, 2000). Mineral excretion was determined using mineral analysis and
calculated urine excretion. Urine excretion was estimated using creatinine as urine
excretion (L/d) = (29 mg daily creatinine excretion/kg BW)/ (analyzed creatinine in
sample) (Valaderes et al., 1999). Urine excretion was also estimated as urine excretion
(L/d) = (0.0259*kg BW * MUN (mg/dL))/ (analyzed N in sample) (Kauffman and St-Pierre,
2001).
Statistical Analysis
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Data were analyzed with mixed models using the MIXED procedure of SAS
(version 9.3, SAS Institute, Inc., Cary, NC). Denominator degrees of freedom for all tests
were adjusted using the Kenward-Rogers option.
One model included the effects of cow within square (random, df= 12), square
(random, df= 3), period (random, df= 3), dietary fat (fixed, df = 1), dietary K (fixed, df =
1), K x fat interaction (fixed, df= 1), and the residual error (random, df = 42). This model
was used to analyze urine, milk minerals, and results that included data from week 3
only that had been averaged within cow within period.
Daily milk yield and DMI were averaged by week within cow and period and
weekly means were analyzed using a model that included the effect of cow within
square (random, df= 12), square (random, df= 3), period (random, df= 3), dietary fat
(fixed, df = 1), dietary K (fixed, df = 1), K x fat interaction (fixed, df= 1), effect of wk
(fixed, df=2), K x wk (fixed, df=2), fat x wk (fixed, df=2), and fat x K x wk (fixed, df=2), and
the residual error. Week was a repeated effect. The model for milk composition was the
same except day (df =5 for composition and 3 for milk fatty acids) replaced the wk terms
in the model. The SLICE option of LSMEANS was used if a K by time or fat by time
interaction occurred. If a time by treatment effect occurred for a specific dependent
variable, then data from wk 3 of each period were averaged within cow and were
analyzed using the above model.
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CHAPTER 4: RESULTS AND DISCUSSION
Diet
Diet composition and analyses are shown in Tables 1 and 2. The DCAD was 2
mEq/100 g DM for diets without additional K and about 30 mEq/100 g DM for added K
diets. Many studies have found that a DCAD of 20 mEq/100 g DM or higher result in
higher milk production and DMI for lactating dairy cows (Tucker et al., 1988; Delaquis
and Block, 1995; Wildman et al., 2007). In an analysis of 16 studies, Hu et al. (2007a)
found that DMI, milk yield, and milk component production were maximized at 47
meq/kg. Dry cows benefit from a low or negative DCAD, which helps prevent
hypocalcemia (NRC, 2001). The diets with added K had DCAD values that have been
found to be adequate for lactating cows, but it is possible that the low DCAD values
could have limited performance for the cows fed the diets without added K.
The total amount of dietary fat increased with the high fat diets, but the fatty
acid profile of diets was similar between low and high fat diets. It is important to note
that about 46% of the fatty acid present in the study was from C 18:2. The sulfur in the
diets was high at 0.44% DM (National Research Council, 2001), which was likely due to
the high inclusion of DGS (Table 4).
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Corn silage 30 h NDF digestibility was determined because low corn silage
digestibility can decrease DMI. Based on other studies, corn silage digestibility should
have been adequate and should not have led to a decrease in DMI (Oba and Allen,
1999b). In vitro TMR digestibility and concentrate mix particle size were similar between
diets, indicating that diets should have been digested similarly. A study by Rémond et al.
(2004) found that grain should be finely ground (particle size <1 mm) to prevent a
decrease in total tract starch digestibility. In this study, grain particle size averaged less
than 0.70 mm for all concentrate mixes, so starch degradability should have been
adequate. Saliva is necessary for buffering of the rumen and saliva production is
determined by chewing time. Chewing time was not assessed in this study, though
based on studies by Lammers et al. (1996) and Kononoff et al. (2003), corn silage
particle size should have been adequate for proper chewing and saliva production.
Cattle had free access to water which contained 99 mg/L Cl, 0.24 mg/L P, 1.7 mg/L K, 67
mg/L Ca, 30 mg/L Mg, and 20 mg/L S. These concentrations were typical and should
have had little to no effect on DCAD or treatment (Solomon et al., 1995; National
Research Council, 2001; Beede, 2005; Castillo et al., 2013). The DGS contained 4.1%
starch, 28% NDF, 30.8% protein, 7.8% LCFA, 0.98% S, and 1.27% K (Table 4), which is
similar to typical DGS, though they were slightly lower in fat, as they were a reduced fat
DGS (Anderson et al., 2006).
Dry Matter Intake, Milk Production, and Milk Composition
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Fat by week and K by week interactions were observed for DMI (Table 5; Figure
1). For cows fed high fat diets, DMI was low and constant over the 3 wk, but increased
by the end of the period for cows fed low fat diets. Intake by cows fed diets with
supplement K were affected by treatment (P<0.01), whereas cows fed diets without
supplemental K was constant and higher over the 3 wk period (P<0.17). Figure 2 shows
this K by day and fat by day interaction in more detail. Low fat diets were affected by wk
(P<0.01), but high fat diets were not (P<0.92). Fat by wk interactions were also observed
for MY (Table 5; Figure 3). Milk yield for cows fed high fat diets were affected by
treatment over time (P<0.01), but MY for cows fed low fat diets were unchanged
(P<0.24).
Fat by day interactions were observed for both milk fat percent and milk fat yield
(Table 5). As shown in Figures 4 and 5, these measurements followed similar patterns,
with a lowest milk fat percent around day 18. Milk fat percent and fat yield were both
unaffected by K by time (P<0.95 and P<0.67, respectively). Milk fat yield with low fat
diets were unaffected by time (P<0.33), but the high fat treatment affected milk fat
yield by the end of the period (P<0.01). The changes over time for production and milk
composition data were indicative of the cows adjusting to treatments. Because of this
assumed adjustment, we decided to look at solely week 3 to compare treatment effects.
Table 6 shows the production and milk composition results for the final wk of the
periods. DMI decreased with added fat (P<0.01). This decrease in DMI with added fat
was expected. Many studies have found that dietary unsaturated fats decrease DMI
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(Pantoja et al., 1994; Pantoja et al, 1996; Firkins and Eastridge, 1994). Extensive
biohydrogenation of FA occurs in the reticulorumen (Viviani, 1970) and should reduce
the hypophagic effects of some sources of unsaturated fat, but biohydrogenation is
reduced when the amount of unsaturated fat in the diet increases (Christensen et al.,
1998), so DMI is still reduced.
Diets with added K also tended to decrease DMI (P<0.06). This is contrary to a
study by Harrison et al. (2012) that found the addition of K2CO3 at 2.1% DM versus 1.3%
increased DMI. There were many differences between this study and Harrison et al.
(2012), but the biggest difference was the length of time that diets were fed. Harrison et
al. (2012) fed his cattle the same diet, either 2.1% K or 1.3% K, for 12 wk, but 2.1% K
diets did not increase DMI until 3 or 4 wk into the experiment. As the cows in this
experiment were only on treatments for 3 wk, it is possible we would have seen this
increase in DMI with added K diets we fed out treatments for a long period of time.
Milk NEL (Mcal/d) is a measure of the energy in daily milk excretion. It is
calculated using yields of milk fat, milk protein, and milk lactose (National Research
Council, 2001). Milk NEL had a K by fat interaction (P<0.03) (Table 5) where adding fat
did not statistically affect diets with added K, but did statistically affect diets with added
K. This difference in milk NEL between diets without added K was likely caused because
milk fat yield increased with added K, leading to a decrease in NE L for diets with added K.
Yields of milk components for diets with added K were similar, resulting in similar NE L.
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The HF-K diet did not have similar yields to the LF-K diet because it did not have the
added K to increase in milk fat yield to values near the low fat diet.
As shown in table 6, MY decreased for high fat diets (P<0.01), which may have
been a result of decreased DMI. Milk protein percent and yield decreased with added K
(P<0.01). This was not expected and we are not certain why this occurred. Milk protein
percent increased with added fat, though milk protein yield was unaffected (P<0.12).
Because MY decreased with added fat and milk protein yield was unaffected, this
increase in milk protein percent was likely due to a change in concentration of milk
protein yield relative to MY.
Milk urea nitrogen (MUN) decreased with added K (P<0.01). As shown in Table
11, urine excretion increase with added K (P<0.01). This increase in urine is likely
associated with an increase in water intake. MUN is directly related to the concentration
of blood urea nitrogen (BUN), which is affected by the amount of water in the body.
Increasing water intake increases body water, thus diluting BUN and, ultimately, MUN.
Water intake was not measured in this study, but urine volume was estimated. Feeding
excess K is associated with increased water intake and urine excretion (Fisher et al.,
1994). Table 11 shows that estimated urine excretion increased with added K (P<0.01),
which may be indicative of an increase in water intake, which would increase body
water and decrease BUN and MUN concentration.
Net energy of lactation (Mcal/kg milk) increased with added K (P<0.04) and
decreased with added dietary fat (P<0.01). This is indicative of the decrease in milk fat
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with added fat, which decreases the energy in milk, and the increase in milk fat with
added K, which increases the energy in milk. Body condition scores, body weights, and
body weight changes were not affected by treatment (Table 4).
Milk fat percent increased with added K (P<0.01) and decreased with added fat
(P<0.01). Milk fat yield was affected similarly and tended to increase with added K
(P<0.10) and decreased with added fat (P<0.01). Adding K to the high fat diet resulted in
increased milk fat as hypothesized, but we did not expect that this would also occur for
the low fat diets. We expected added K to increase milk fat percent by decreasing
incomplete biohydrogenation, which results in decreased milk production. We did not
expect incomplete biohydrogenation to occur for the low fat diets, which is why we did
not expect an increase in milk fat percent for the low fat diets. All diets caused milk fat
depression. This may have occurred because of the high level of S in the diet. The high
amount of S from the DGS could have decreased rumen pH (Felix and Loerch, 2011). A
low rumen pH with the high amount of unsaturated fatty acid in both the low and high
fat diets could have caused the milk fat depression with all diets. Ivancic and Weiss
(2004) found a decrease in milk fat percent when 0.4% S was fed vs. 0.2% S.
Milk Fatty Acids
Many of the milk fatty acids were affected by fat by time or K by time
interactions (Table 7). We assumed that this may have been due to the cows adjusting
to new diets at 30 h into each period, but when only data from d 7, 14, and 21 were
analyzed, time interactions were still observed, which indicates that cows may still have
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been adjusting to diets on days 7 and 14. Because of these suspected adjustments, we
looked at data from d 21 (Table 9), which follows along with the previous analysis of
solely wk 3 of production and milk composition data.
Table 8 shows the milk fatty acid analysis of samples collected at 30 h into
treatment. This 30 h collection represents the initial adjustments to the new diets and is
especially essential for estimating changes in ruminal environment to the new diets.
Branched chain fatty acids have been found to have the potential to predict molar
proportions of volatile fatty acids (VFA) in the rumen (Vlaeminck et al., 2006). Often
when MFD occurs, the molar proportion of ruminal acetate decreases and proportion of
ruminal propionate increases. Concentrations of iso C14:0 and iso C15:0 in milk fat are
positively related with rumen proportions of acetate and negatively related with molar
proportions of propionate (Vlaeminck et al., 2006). Iso C 14:0 increased with added K
(P<0.01) and decreased with added fat (P<0.01). Iso C 15:0 behaved similarly. It also
increased with added K (P<0.03) and decreased with added fat (P<0.03). These results
may indicate that ruminal concentrations of acetate were increasing and proportion of
propionate was decreasing with added K. This may also indicate that proportion of
acetate decreased with added fat and concentration of propionate increased with
added fat.
Milk fat depression results in a decrease in proportion of short chain fatty acids
(SCFA) and an increase in the proportion of LCFA (Peterson et al., 2003). Short chain
fatty acids, those less than 16 carbons, were increased with added K (P<0.01) and
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decreased with added fat (P<0.01) (Table 9). The C 16 fatty acids were increased with
added K (P<0.02) and decreased with added fat (P<0.01). Long chain fatty acids, those
greater than 16 carbons, decreased with added K (P<0.01) and increased with added fat
(P<0.01). The increase in LCFA with added fat may have been caused by the overall
increase of LCFA in the diets. Trans isomers, which are included in the LCFA, lead to a
decrease in SCFA, as well.
Many of the trans isomers, including trans-6+8 18:1, trans-9 18:1, trans-10 18:1
and trans-12 18:1 increased with added fat (P<0.01) and decreased with added K
(P<0.01) (Table 9). Trans-10, cis-12 CLA is a biohydrogenation intermediate that
increases in concentration when incomplete biohydrogenation occurs. Table 9 shows
that trans-10, cis-12 CLA in milk decreased with added K (P<0.02) indicating that the
additional K is increasing milk fat production by decreasing incomplete
biohydrogenation. Trans-10, cis-12 increased with added fat (P<0.01) because excess
unsaturated fatty acids in the diets results in increased incomplete biohydrogenation. As
Figure 6 shows, there was a linear relationship between milk fat percent and trans-10,
cis-12.
Biohydrogenation is the process of changing dietary unsaturated fatty acids, such
as C 18:2, into saturated fatty acids, such as C18:0. Table 10 shows the effect of
treatment on proportion of C 18:0 and C 18:2 relative to total concentration of C 18
fatty acids. Both C 18:0 and C 18:2 concentration were unaffected by added fat, though
35
`
C 18:0 increased with added K (P<0.01) and C 18:2 decreased with added K. This may
indicate that adding K increased biohydrogenation of C 18:2 to C 18:0.
Mineral Excretion
Mineral intake and excretion was calculated for wk 3 because that is the week
urine samples and milk samples for mineral analysis were taken. As shown in Table 11,
urine excretion (L/d) estimated using both creatinine and MUN equations did not differ
statistically (P<0.29). Estimated urine excretion increased with added K (P<0.01) and
with added fat (P<0.01). The increase with added K was expected as excess K is
associated with increased water intake and urine excretion (Fisher et al., 1994). For the
creatinine method, fat had no significant effect on estimated urine excretion (P<0.16),
but urine excretion increased with added fat for the MUN method (P<0.05).
Urine excretion of K increased with added K (P<0.01) as was expected, as an
increase in K intake would lead to an increase in K in urine. Magnesium excretion in
urine decreased with added K for both treatments (P<0.01) and added fat (creatinine,
P<0.01; MUN, P<0.02). Urinary mineral excretion was measured to evaluate the effects
of K on Mg absorption. Assuming typical diets and DMI, diets with approximately 0.2%
Mg will usually meet Mg requirements for lactating cows (NRC, 2001); though adding K
decreases Mg absorption (Newton et al., 1972). There are two equations suggesting
additional Mg is required when excess K is fed. Weiss (2004) suggested an additional
0.08% Mg for each additional 1% dietary K, which calculates to a maximum of 0.30% Mg
for our added K treatments. Schonewille et al., (2008) suggested an additional 0.02% for
36
`
each additional 1% dietary K, which calculates to 0.23% Mg. We planned to meet
requirements by preparing diets that supplied 0.30% Mg, though analysis shows that
high fat without added K only contained 0.25% Mg. Though, for Weiss (2004) and
Schonewille et al., (2008), a maximum of 0.22% Mg would suffice for the without added
K treatments. Also, Mg being present in the urine is a clear indication that enough Mg
was fed.
The decrease of Mg in urine with added fat may have been due to the decrease
in intake or increased number of free fatty acids in the rumen. Increased dietary fat
leads to a decrease in Mg absorption (Ramirez and Zinn, 2000). Magnesium binds to free
fatty acids. The fatty acids are dissociated in the abomasum, which frees up the Mg, but
Mg is absorbed in the rumen only. Urinary S excretion decreased with added K (P<0.01)
for both treatments and with added fat for the creatinine method. The decrease with
added fat may have been due to the decrease in S intake, though the difference
between S intakes with added K was not significant and does not explain this decrease
in urine S excretion. Mineral values in urine are reasonable based on papers by Tucker
and Hogue (1990) and Nennich et al., (2006). Table 13 shows the effect of treatment on
the percentage of urine mineral excretion/mineral intake. Magnesium urine
excretion/intake also decreased with added K, which was likely a result of K blocking Mg
absorption.
Milk mineral excretion were affected by treatment, but concentrations of
minerals in milk (Table 14) were normal (National Research Council, 2001). Though
37
`
dietary fat did decrease concentrations of P, Ca, Mg, S, and Na in milk, which may have
been caused by the decrease in mineral intake of all of these minerals with added fat
diets (P<0.01).
Conclusion
Diets based on DGS that contained high concentrations of PUFA led to MFD. The
addition of K2CO3 to the high fat diets increased milk fat percent and milk fat yield.
Though all diets led to milk fat depression, which may have been a result of the high
amount of S in the diets due to the high S in the DGS. Milk fat analysis suggested that
additional K2CO3 decreased incomplete biohydrogenation as trans-10, cis-12 CLA
decreased with added K treatments. An increase in proportion of C 18:0 along with a
decrease in C 18:2 may indicate that adding K led to an increase in biohydrogenation of
C 18:2 to C 18:0.
38
`
Table 1: Ingredient composition of diets, DM basis
39
Ingredient
Corn Silage, %
Concentrate Mix, %
Dried distiller grains with solubles2, %
Corn Oil, %
Corn gluten meal, %
DCAD Plus3, %
Corn (ground), %
Soybean meal 48% CP, %
Soybean hulls, %
Calcium carbonate, %
Magnesium oxide, %
Trace mineral salt, %
Mineral and vitamin premix4, %
LF-K
47.0
53.0
27.0
8.5
6.5
6.9
2.78
0.13
0.64
0.55
Treatment1
LF+K HF-K
47.0
47.0
53.0
53.0
27.0
22.0
2.3
2.7
1.9
8.5
8.5
6.5
6.5
5.0
6.9
2.78
2.78
0.13
0.13
0.64
0.64
0.55
0.55
1
HF+K
47.0
53.0
22.0
2.3
2.7
1.9
8.5
6.5
5.0
2.78
0.13
0.64
0.55
Treatments were low fat without added K (LF-K), low fat with added K (LF+K), high fat without added K (HF-K), and high fat
with added k (HF+K).
2
Dakota Gold® Dried Distiller Grains with Solubles (Poet Nutrition, Sioux Falls, SD).
3
DCAD Plus®; Church & Dwight Co., Inc., Princeton, NJ
4
Contained 71.4 % biotin (220mg/kg) premix (DSM Nutritional Products, Inc., Parsippany, NJ), 11.3% Selenium premix
(200mg/kg), 0.4% copper sulfate, 9.4% Vitamin E premix (44 IU/g), 5.6% Vitamin D premix (3,000 IU/g), and 1.9% Vitamin A
premix (30,000 IU/g).
39
`
Table 2: Nutrient composition of diets
Nutrient
Long chain FA (LCFA), %DM
C 16:0, g/100 g of LCFA
C 18:0, g/100 g of LCFA
C 18:1, g/100 g of LCFA
C 18:2, g/100 g of LCFA
LF-K
4.2
7.4
2.7
22.4
46.0
Other and unidentified, g/100 g of LCFA 21.4
Starch, %DM
23.4
NDF, %DM
32.1
Forage NDF, %DM
17.9
CP, %DM
17.2
Ash, %DM
5.6
K, %DM
1.24
Na, %DM
0.21
Cl, %DM
0.48
S, %DM
0.40
Mg, %DM
0.32
Ca, %DM
0.92
P, %DM
0.46
2
DCAD, (mEq/100 g DM)
2
30 hour NDF Digestibility, %NDF
67.9
Concentrate mix mean particle size, mm
0.60
Concentrate mix pH
4.56
NEL, Mcal/kg3
1.63
MP allowable milk, kg/d
33.5
1
Treatment1
LF+K HF-K HF+K
4.3
5.8
5.9
7.4
6.9
7.0
2.6
2.5
2.4
22.1 23.2 23.3
45.1
22.7
24.9
30.2
17.9
17.3
7.9
2.29
0.24
0.47
0.40
0.32
0.89
0.45
31
69.8
0.59
7.42
1.59
32.4
47.3
20.1
23.3
31.0
17.9
17.4
5.4
1.19
0.20
0.50
0.36
0.25
0.69
0.44
2
67.3
0.62
4.60
1.70
34.0
47.2
20.1
25.3
30.0
17.9
17.5
7.5
2.20
0.22
0.47
0.37
0.29
0.86
0.46
29
68.6
0.65
7.54
1.68
34.2
Treatments were low fat without added K (LF-K), low fat with added K (LF+K), high fat
without added K (HF-K), and high fat with added k (HF+K).
2
DCAD, mEq/100 g DM = [(%Na × 43.5 + %K × 25.6) − (%Cl × 28.2 + %S × 62.5)].
3
NRC (2001)
40
`
Table 3: Nutrient composition of corn silage (n=4)
Item
Mean SD
DM %
41.2 1.0
Starch, %DM
33.5 2.2
NDF, %DM
37.8 2.7
CP, %DM
7.7 0.5
Ash, %DM
3.57 0.52
30 hour NDF Digestibility, %NDF
54.7 0.29
1
Particle size, as fed
Top Screen, % as fed
6.3 2.7
Middle Screen, % as fed
69.7 1.8
Pan, % as fed
24.0 2.2
1
Penn State Particle Separator (Lammers et al., 1996)
41
`
Table 4: Nutrient composition of distiller grains (n=4)
Item
Starch, %DM
NDF, %DM
CP, %DM
LCFA, %DM
C 16:0, g/100 g of LCFA
C 18:0, g/100 g of LCFA
C 18:1, g/100 g of LCFA
C 18:2, g/100 g of LCFA
Other and unidentified, g/100 g of LCFA
Ash, %DM
K, %DM
Na, %DM
Cl, %DM
S, %DM
Mg, %DM
Ca, %DM
P, %DM
Grain pH
42
Mean
4.1
28.0
30.8
7.8
13.1
2.4
23.9
50.5
10.1
5.38
1.27
0.23
0.21
0.97
0.41
0.03
0.89
3.59
SD
0.62
0.51
0.64
0.39
0.40
0.07
0.17
1.34
1.70
0.10
0.04
0.02
0.02
0.02
0.01
0.002
0.02
0.04
`
Table 5: Effects of treatment on DMI, milk production, and milk composition, all 3 wk of treatment (n=16 cow period per
treatment)
43
Item
DMI, kg/d
Milk yield, kg/d
Milk NEL3, Mcal/kg
Milk NEL, Mcal/d
Milk fat, %
Milk fat, kg/d
Milk protein, %
Milk protein, kg/d
Lactose, %
Lactose, kg/d
MUN, mg/dL
LF-K
23.0
32.8
0.64
21.4
2.86
0.97
3.39
1.12
4.83
1.60
15.7
Treatment1
P<
LF+K HF-K HF+K SEM
K
Fat KxFat KxTime2 FatxTime KxFatxTime
21.3 21.6 20.5 1.04 0.01 0.01 0.34
0.02
0.01
0.59
31.1 31.5 31.1 3.40 0.09 0.27 0.25
0.76
0.01
0.97
0.66 0.62 0.65 0.018 0.01 0.01 0.28
0.87
0.01
0.92
20.5 19.5 20.3 2.17 0.82 0.01 0.03
0.33
0.01
0.93
3.08 2.63 2.93 0.19 0.01 0.01 0.47
0.96
0.01
0.96
0.96 0.84 0.92 0.12 0.07 0.01 0.05
0.68
0.01
0.64
3.33 3.48 3.37 0.092 0.03 0.06 0.43
0.08
0.03
0.68
1.03 1.08 1.05 0.096 0.01 0.57 0.22
0.75
0.32
0.56
4.78 4.82 4.85 0.079 0.85 0.17 0.06
0.73
0.76
0.52
1.49 1.52 1.52 0.16 0.08 0.32 0.08
0.75
0.13
0.74
13.7 16.4 14.6 0.45 0.01 0.01 0.53
0.24
0.16
0.28
1
Treatments were low fat without added K (LF-K), low fat with added K (LF+K), high fat without added K (HF-K), and high fat
with added k (HF+K).
2
DMI and milk yield samples were collected weekly. Milk composition data was collected on d 3, 7, 10, 14, 17, and 21.
3
Calculated from milk fat, protein, and lactose yield (NRC, 2001)
43
`
Table 6: Effects of treatment on DMI, milk production, and milk composition, wk 3 of treatment (n=16 cow period per
treatment)
44
Item
DMI, kg/d
Milk yield, kg/d
Milk NEL2, Mcal/kg
Milk NEL, Mcal/d
Milk fat, %
Milk fat, kg/d
Milk protein, %
Milk protein, kg/d
Lactose, %
Lactose, kg/d
MUN, mg/dL
BCS
BW, kg
BW Change, kg/d
LF-K
22.9
33.0
0.63
21.0
2.74
0.92
3.41
1.12
4.83
1.60
16.0
3.0
671
0.68
Treatment1
P<
LF+K HF-K HF+K SEM
K
Fat KxFat
22.1 21.3 20.7 0.98 0.06 0.01 0.88
31.6 30.6 30.4
3.6 0.24 0.01 0.34
0.65 0.59 0.62 0.020 0.04 0.01 0.43
20.2 17.8 18.9 2.08 0.75 0.01 0.11
2.99 2.39 2.64 0.13 0.01 0.01 1.00
0.94 0.72 0.80 0.98 0.10 0.01 0.27
3.33 3.56 3.41 0.085 0.01 0.01 0.33
1.03 1.07 1.03 0.10 0.01 0.12 0.25
4.77 4.84 4.86 0.086 0.69 0.17 0.28
1.49 1.47 1.48 0.16 0.10 0.02 0.05
13.9 16.0 14.6 0.63 0.01 0.24 0.22
3.2
3.2
3.1 0.20 0.43 0.47 0.08
673 676
671 16.45 0.75 0.62 0.32
0.45 0.22 0.38 0.21 0.88 0.22 0.37
1
Treatments were low fat without added K (LF-K), low fat with added K (LF+K), high fat without added K (HF-K), and high fat
with added k (HF+K).
2
Calculated from milk fat, protein, and lactose yield (NRC, 2001)
44
`
Table 7: Effects of treatment on milk fatty acid concentrations, all 3 wk (n=16 cow period per treatment)
45
Fatty Acid, g/100g fatty acids2 LF-K
4:0
2.86
6:0
1.77
8:0
0.89
10:0
2.01
12:0
2.55
iso 13:0
0.03
anteiso 13:0
0.04
13:0
0.11
iso 14:0
0.10
14:0
9.83
iso15:0
0.21
anteiso 15:0
0.51
14:1
1.27
15:0
1.06
iso 16:0
0.27
16:0
24.95
iso 17:0
0.45
C16:1 + anteiso 17:0
2.21
17:0
0.61
17:1
0.23
18:0
10.90
trans-6+8 18:1
0.73
trans-9 18:1
0.63
trans-10 18:1
2.26
trans-11 18:1
0.69
trans-12 18:1
0.41
Treatment1
P<
LF+K HF-K HF+K SEM
K
Fat KxFat KxDay3 FatxDay KxFatxDay
2.94 2.71 2.82 0.129 0.10 0.02 0.84
0.34
0.02
0.72
1.81 1.54 1.63 0.11 0.06 0.01 0.48
0.01
0.01
0.67
0.90 0.73 0.78 0.06 0.13 0.01 0.32
0.01
0.01
0.52
1.97 1.58 1.68 0.14 0.50 0.01 0.10
0.01
0.01
0.53
2.41 2.08 2.11 0.15 0.21 0.01 0.05
0.01
0.01
0.61
0.03 0.02 0.02 0.002 0.76 0.01 0.73
0.39
0.07
0.78
0.03 0.05 0.04 0.006 0.01 0.01 0.52
0.49
0.23
0.24
0.10 0.10 0.12 0.01 0.69 0.63 0.14
0.89
0.20
0.56
0.12 0.08 0.10 0.007 0.01 0.01 0.64
0.11
0.67
0.17
9.48 8.53 8.59 0.33 0.20 0.01 0.06
0.04
0.15
0.04
0.22 0.17 0.19 0.006 0.01 0.01 0.21
0.97
0.01
0.27
0.51 0.42 0.45 0.01 0.02 0.01 0.06
0.31
0.01
0.31
1.12 1.12 1.03 0.09 0.01 0.01 0.29
0.45
0.08
0.86
0.98 0.91 0.88 0.05 0.01 0.01 0.21
0.01
0.01
0.92
0.32 0.25 0.29 0.03 0.01 0.01 0.42
0.36
0.99
0.63
25.36 23.32 23.16 0.56 0.65 0.01 0.29
0.43
0.02
0.47
0.45 0.43 0.43 0.01 0.91 0.01 0.54
0.08
0.03
0.65
2.14 2.10 2.01 0.11 0.04 0.01 0.86
0.54
0.03
0.80
0.60 0.52 0.54 0.02 0.37 0.01 0.04
0.18
0.01
0.66
0.23 0.21 0.22 0.01 0.94 0.01 0.60
0.10
0.16
0.86
11.55 11.92 12.77 0.50 0.01 0.01 0.54
0.73
0.01
0.50
0.59 1.01 0.82 0.05 0.01 0.01 0.21
0.40
0.89
0.94
0.50 0.95 0.70 0.05 0.01 0.01 0.02
0.07
0.35
0.96
1.72 3.31 2.50 0.20 0.01 0.01 0.11
0.24
0.12
0.89
0.78 0.71 0.82 0.10 0.02 0.52 0.75
0.02
0.01
0.23
0.35 0.54 0.88 0.22 0.53 0.14 0.36
0.41
0.44
0.37
45
`
cis-9 18:1
cis-11 18:1
18:2
20:0
20:1
18:3
cis-9, trans-11 CLA
CLA (other)
trans-10, cis-12 CLA
25.77 26.56 27.19 27.66 0.99 0.02 0.01
0.77 0.72 0.90 0.80 0.04 0.01 0.01
4.62 4.24 5.22 4.62 0.15 0.01 0.01
0.15 0.16 0.16 0.18 0.007 0.01 0.01
0.14 0.14 0.14 0.14 0.005 0.01 0.23
0.30 0.27 0.31 0.28 0.008 0.01 0.01
0.58 0.61 0.64 0.67 0.04 0.22 0.01
0.05 0.04 0.06 0.05 0.004 0.01 0.01
0.04 0.04 0.07 0.05 0.007 0.01 0.01
1
0.55
0.09
0.05
0.52
0.62
0.62
0.91
0.06
0.01
0.61
0.04
0.01
0.67
0.55
0.01
0.01
0.22
0.19
0.01
0.01
0.01
0.30
0.13
0.15
0.01
0.03
0.12
0.85
0.95
0.48
0.27
0.91
0.72
0.34
0.89
0.78
Treatments were low fat without added K (LF-K), low fat with added K (LF+K), high fat without added K (HF-K), and high fat
with added k (HF+K).
2
Number of carbons: number of double bonds.
3
Data were collected at 30 h, d 7, 14, and 21.
46
46
`
Table 8: Effects of treatment on milk fatty acid concentrations, 30 h after diets were first fed (n=16 cow period per
treatment)
47
Fatty Acid, g/100g fatty acids
4:0
6:0
8:0
10:0
12:0
iso 13:0
anteiso 13:0
13:0
iso 14:0
14:0
Iso 15:0
anteiso 15:0
14:1
15:0
iso 16:0
16:0
iso 17:0
C16:1 + anteiso 17:0
17:0
17:1
18:0
trans-6+8 18:1
trans-9 18:1
trans-10 18:1
2
LF-K
2.79
1.70
0.85
1.91
2.42
0.03
0.03
0.10
0.10
9.35
0.20
0.51
1.25
0.98
0.28
24.0
0.45
2.20
0.56
0.22
11.4
0.76
0.65
2.43
Treatment1
P<
LF+K HF-K HF+K SEM
K
Fat K*Fat
2.79 2.93 2.88 0.17 0.80 0.25 0.82
1.59 1.69 1.60 0.10 0.16 0.93 0.87
0.78 0.83 0.76 0.062 0.13 0.62 1.00
1.66 1.78 1.66 0.13 0.04 0.47 0.50
2.13 2.25 2.13 0.15 0.02 0.28 0.29
0.03 0.03 0.03 0.002 0.09 0.54 0.57
0.03 0.06 0.03 0.008 0.01 0.01 0.02
0.09 0.10 0.10 0.01 0.43 0.66 0.45
0.11 0.08 0.09 0.01 0.01 0.01 0.57
9.00 8.73 8.57 0.41 0.18 0.01 0.61
0.21 0.18 0.18 0.01 0.03 0.03 0.63
0.48 0.45 0.47 0.02 0.85 0.01 0.10
1.12 1.13 1.03 0.12 0.03 0.06 0.77
0.93 0.94 0.92 0.05 0.24 0.32 0.56
0.31 0.24 0.28 0.03 0.07 0.10 0.69
24.5 23.5 23.2 0.49 0.86 0.01 0.11
0.47 0.45 0.46 0.02 0.21 0.91 0.58
2.26 2.14 2.07 0.14 0.97 0.07 0.36
0.58 0.54 0.57 0.02 0.15 0.36 0.51
0.24 0.22 0.23 0.01 0.11 0.39 0.79
11.8 11.7 12.6 0.63 0.01 0.05 0.29
0.66 1.01 0.87 0.05 0.01 0.01 0.62
0.61 0.92 0.76 0.05 0.03 0.01 0.18
2.08 3.02 2.64 0.19 0.02 0.01 0.91
47
`
trans-11 18:1
trans-12 18:1
cis-9 18:1
cis-11 18:1
18:2
20:0
20:1
18:3
cis-9, trans-11 CLA
CLA (other)
trans-10, cis-12 CLA
0.59
0.42
27.0
0.79
4.78
0.16
0.14
0.30
0.56
0.05
0.05
0.52
0.35
26.5
0.82
4.66
0.16
0.14
0.28
0.49
0.05
0.07
0.90
0.56
28.0
0.82
4.90
0.16
0.13
0.30
0.74
0.06
0.06
0.75 0.12 0.25 0.01
0.45 0.02 0.01 0.01
27.8 0.80 0.01 0.45
0.80 0.05 0.86 0.99
4.73 0.15 0.14 0.30
0.20 0.02 0.20 0.12
0.14 0.005 0.31 0.06
0.29 0.008 0.22 0.73
0.62 0.05 0.02 0.01
0.05 0.004 0.14 0.30
0.05 0.02 0.86 0.92
1
0.68
0.37
0.76
0.25
0.77
0.29
0.86
0.69
0.59
0.76
0.24
Treatments were low fat without added K (LF-K), low fat with added K (LF+K), high fat without added K (HF-K), and high fat
with added k (HF+K).
2
Number of carbons: number of double bonds.
48
48
`
Table 9: Effects of treatment on milk fatty acid concentrations, d 21 of treatment (n=16 cow period per treatment)
49
Fatty Acid, g/100g fatty acids
SCFA
4:0
6:0
8:0
10:0
12:0
iso 13:0
anteiso 13:0
13:0
iso 14:0
14:0
iso15:0
anteiso 15:0
14:1
15:0
iso 16:0
16:0
LCFA
iso 17:0
C16:1 + anteiso 17:0
17:0
17:1
18:0
trans-6+8 18:1
2
LF-K
23.5
2.82
1.76
0.91
2.06
2.60
0.03
0.06
0.12
0.10
9.92
0.21
0.51
1.31
1.08
0.26
24.6
51.7
0.47
2.23
0.62
0.23
10.7
0.77
Treatment1
P<
LF+K HF-K HF+K SEM
K
Fat KxFat
24.0 18.6 20.4 1.09 0.01 0.01 0.06
3.01 2.53 2.75 0.16 0.01 0.01 0.84
1.94 1.35 1.61 0.11 0.01 0.01 0.25
1.00 0.62 0.78 0.07 0.01 0.01 0.14
2.22 1.35 1.69 0.17 0.01 0.01 0.13
2.67 1.85 2.12 0.18 0.01 0.01 0.09
0.03 0.03 0.02 0.001 0.01 0.01 0.08
0.05 0.05 0.05 0.011 0.40 0.83 0.55
0.12 0.09 0.09 0.014 0.92 0.01 0.72
0.12 0.08 0.09 0.010 0.02 0.01 0.21
9.92 8.06 8.62 0.43 0.06 0.01 0.06
0.22 0.16 0.18 0.009 0.01 0.01 0.88
0.53 0.41 0.43 0.02 0.07 0.01 0.99
1.14 1.15 1.11 0.10 0.02 0.03 0.10
1.06 0.85 0.86 0.06 0.77 0.01 0.61
0.31 0.26 0.26 0.03 0.19 0.21 0.20
25.2 22.8 23.7 0.49 0.02 0.01 0.54
50.5 58.4 55.6 1.51 0.01 0.01 0.12
0.45 0.42 0.40 0.02 0.18 0.01 0.93
2.06 2.22 2.11 0.14 0.06 0.79 0.65
0.62 0.49 0.52 0.02 0.06 0.01 0.26
0.22 0.22 0.22 0.01 0.33 0.35 0.33
11.5 12.0 12.3 0.58 0.05 0.01 0.41
0.59 1.07 0.79 0.06 0.01 0.01 0.30
49
`
trans-9 18:1
trans-10 18:1
trans-11 18:1
trans-12 18:1
cis-9 18:1
cis-11 18:1
18:2
20:0
20:1
18:3
cis-9, trans-11 CLA
CLA (other)
trans-10, cis-12 CLA
0.67
2.46
0.63
0.41
25.7
0.79
4.79
0.15
0.14
0.30
0.57
0.05
0.04
0.47
1.75
0.84
0.36
25.6
0.69
4.11
0.17
0.14
0.26
0.65
0.04
0.04
1.02
3.52
0.53
0.55
28.5
0.94
5.53
0.16
0.14
0.32
0.55
0.07
0.09
0.66
2.52
0.64
0.44
28.3
0.83
4.56
0.16
0.14
0.27
0.58
0.05
0.05
0.08
0.25
0.07
0.03
0.98
0.06
0.13
0.008
0.006
0.009
0.04
0.005
0.01
50
1
0.01
0.01
0.01
0.01
0.69
0.01
0.01
0.03
0.35
0.01
0.16
0.01
0.02
0.01
0.01
0.01
0.01
0.01
0.01
0.01
0.11
0.09
0.01
0.30
0.01
0.01
0.28
0.46
0.31
0.15
0.88
0.84
0.14
0.07
0.20
0.98
0.53
0.50
0.12
Treatments were low fat without added K (LF-K), low fat with added K (LF+K), high fat without added K (HF-K), and high fat
with added k (HF+K).
2
Number of carbons: number of double bonds.
50
`
Table 10: Effect of treatment on proportion of C 18:0 and C 18:2 relative to total
concentration of C 18 fatty acids (n=16 cow period per treatment)
Fatty Acid, g/100g C 18 FA2
18:0
18:2
Treatment1
P<
LF-K LF+K HF-K HF+K SEM
K
Fat KxFat
22.4 24.6 21.9 23.6 0.75 0.01 0.17 0.68
10.1
8.8 10.1
8.8 0.30 0.01 0.93 0.86
1
Treatments were low fat without added K (LF-K), low fat with added K (LF+K), high fat
without added K (HF-K), and high fat with added k (HF+K).
2
Number of carbons: number of double bonds.
51
`
Table 11: Effect of treatment on estimated urine excretion (L/d) (n=16 per treatment)
Liters
Average
Creatinine method2
MUN method3
LF-K
24.2
23.4
24.5
Treatment1
P<
LF+K HF-K HF+K SEM
K
Fat KxFat Method KxMethod FatxMethod KxFatxMethod
32.5 24.9 37.5 2.45 0.01 0.01 0.05
0.29
0.68
0.27
0.38
33.2 24.0 36.0 2.67 0.01 0.16 0.36
31.7 25.6 38.8 2.62 0.01 0.05 0.14
1
Treatments were low fat without added K (LF-K), low fat with added K (LF+K), high fat without added K (HF-K), and high fat
with added k (HF+K).
2
Urine volume calculated as (29 mg daily creatinine excretion/kg BW)/ (analyzed creatinine in urine sample)
3
Urine volume calculated as (0.0259*kg BW * MUN (mg/dL))/ (analyzed N in urine sample)
52
52
`
Table 12: Effect of treatment on mineral intake and excretion (g/day) (n=16 per treatment)
53
Mineral
Intake2
Phosphorus
Potassium
Calcium
Magnesium
Sulfur3
Sodium
Nitrogen
Milk Excretion
Phosphorus
Potassium
Calcium
Magnesium
Sulfur
Sodium
Urine Excretion
Creatinine method4
Phosphorus
Potassium
Calcium
Magnesium
Sulfur
Sodium
Nitrogen
Treatment1
LF-K LF+K HF-K HF+K SEM
K
P<
Fat
KxFat
103
282
191
96
91
48
629
94
488
505
94
89
53
609
91
254
150
90
77
43
586
91
438
407
88
77
46
569
4.8
17
13
3.8
3.8
2.1
27
0.01
0.01
0.01
0.04
0.27
0.01
0.06
0.01
0.01
0.01
0.01
0.01
0.01
0.01
0.01
0.12
0.01
0.93
0.32
0.14
0.88
28
49
35
3.5
9.6
12
26
47
33
3.3
9.2
11
27
46
35
3.6
9.3
13
26
45
34
3.4
8.9
11
2.7
5.5
3.4
0.4
0.9
1.8
0.01
0.13
0.02
0.07
0.02
0.19
0.18
0.09
0.56
0.46
0.07
0.83
0.53
0.53
0.82
0.95
0.92
0.84
2.1
185
4.5
11
59
37
264
2.1
351
0.5
7.0
45
20
253
2.5
167
3.8
8.3
49
35
253
1.9 0.9 0.57 0.88
355 23.5 0.01 0.51
0.4 0.7 0.01 0.41
5.3 0.9 0.01 0.01
41 3.2 0.01 0.01
33 4.3 0.01 0.09
235 10.7 0.17 0.16
0.48
0.34
0.48
0.18
0.09
0.03
0.70
53
`
MUN method5
Phosphorus
Potassium
Calcium
Magnesium
Sulfur
Sodium
Nitrogen
2.2
194
4.9
12
63
40
273
2.0
336
0.4
6.6
42
19
236
2.7
182
4.4
9.1
53
38
268
2.0 0.9
383
24
0.4 0.9
5.9 1.0
44 4.0
36 4.7
256 12.6
0.40
0.01
0.01
0.01
0.01
0.01
0.02
0.67
0.35
0.74
0.02
0.33
0.06
0.41
1
0.63
0.12
0.69
0.13
0.12
0.03
0.17
Treatments were low fat without added K (LF-K), low fat with added K (LF+K), high fat without added K (HF-K), and high fat
with added k (HF+K).
2
Calculated as grams of mineral in DM fed – grams mineral of mineral in DM refusal
3
Calculated as % in diet x DMI
4
Urine volume calculated as (29 mg daily creatinine excretion/kg BW)/ (analyzed creatinine in urine sample)
5
Urine volume calculated as (0.0259*kg BW * MUN (mg/dL))/ (analyzed N in urine sample)
54
54
`
Table 13: Effect of treatment on urine mineral excretion/mineral intake (n=16 per
treatment)
Mineral
Creatinine method2
Phosphorus
Potassium
Calcium
Magnesium
Sulfur
Sodium
Nitrogen
MUN method3
Phosphorus
Potassium
Calcium
Magnesium
Sulfur
Sodium
Nitrogen
Treatment1
LF-K LF+K HF-K HF+K SEM K
P<
Fat
KxFat
1.91
64.9
2.35
11.6
72.9
80.7
34.6
2.23
71.4
0.10
7.42
55.8
44.4
49.8
2.94
65.2
2.42
9.22
60.3
76.9
38.2
2.01
80.9
0.11
5.90
51.0
69.6
57.8
0.95
3.44
0.38
0.81
5.47
9.65
3.47
0.54
0.01
0.01
0.01
0.01
0.01
0.01
0.41
0.02
0.88
0.01
0.01
0.16
0.01
0.20
0.02
0.93
0.43
0.16
0.06
0.21
2.04
68.6
2.54
12.2
78.3
88.7
36.3
2.10
68.2
0.08
6.90
52.1
40.8
47.5
3.15
72.0
2.87
10.2
66.0
84.1
40.9
2.15
88.7
0.11
6.61
54.3
74.5
62.9
1.00
5.20
0.56
1.04
6.11
10.4
3.80
0.38
0.10
0.01
0.01
0.01
0.01
0.01
0.28
0.02
0.69
0.15
0.28
0.12
0.01
0.32
0.08
0.74
0.28
0.12
0.04
0.08
1
Treatments were low fat without added K (LF-K), low fat with added K (LF+K), high fat
without added K (HF-K), and high fat with added k (HF+K).
2
Calculated as (29 mg daily creatinine excretion/kg BW) / (analyzed creatinine in sample)
3
Calculated as (0.0259*kg BW * MUN (mg/dL)) / (analyzed N in sample)
55
`
Table 14: Effect of treatment on mineral milk concentration, g/kg (n=16 per treatment)
Mineral
Phosphorus
Potassium
Calcium
Magnesium
Sulfur
Sodium
LF-K
0.85
1.48
1.06
0.11
0.29
0.38
Treatment1
LF+K HF-K
0.83 0.89
1.47 1.51
1.05 1.15
0.11 0.12
0.29 0.30
0.36 0.41
HF+K
0.86
1.50
1.13
0.11
0.30
0.38
1
SEM
0.02
0.03
0.04
0.003
0.007
0.04
K
0.05
0.43
0.32
0.19
0.26
0.22
P<
Fat
0.01
0.08
0.01
0.01
0.02
0.30
KxFat
0.55
0.77
0.69
0.40
0.23
0.82
Treatments were low fat without added K (LF-K), low fat with added K (LF+K), high fat
without added K (HF-K), and high fat with added k (HF+K).
56
`
Figure 1: Effect of level of dietary fat and K on DMI by week within period (n=16 per
treatment). Low Fat=4.2% LCFA; High Fat=5.2% LCFA; K-=1.2% K; K+=2.2% K.
25
24
23
DMI (kg)
22
Low Fat
High Fat
K-
21
K+
20
19
18
1
2
Week Within Period
57
3
`
Figure 2: Effect of level of dietary fat and K on daily DMI over the period (n=16 per
treatment). Low Fat=4.2% LCFA; High Fat=5.2% LCFA; K-=1.2% K; K+=2.2% K.
24
23.5
23
22.5
DMI (kg)
22
Low Fat
21.5
High Fat
K-
21
K+
20.5
20
19.5
19
0
5
10
Day Within Period
58
15
20
`
Figure 3: Effect of level of dietary fat and K on milk production over the period (n=16 per
treatment). Low Fat=4.2% LCFA; High Fat=5.2% LCFA; K-=1.2% K; K+=2.2% K.
37
36
35
Milk Production (kg)
34
33
Low Fat
32
High Fat
K-
31
K+
30
29
28
27
1
2
Week Within Period
59
3
`
Figure 4: Effect of level of dietary fat and K on milk fat yield over the period (n=16 per
treatment). Low Fat=4.2% LCFA; High Fat=5.2% LCFA; K-=1.2% K; K+=2.2% K.
1.2
1.1
Milk Fat Yield (kg)
1
Low Fat
0.9
High Fat
KK+
0.8
0.7
0.6
0
5
10
15
Day Within Period
60
20
`
Figure 5: Effect of level of dietary fat and K on milk fat percent over the period (n=16 per
treatment). Low Fat=4.2% LCFA; High Fat=5.2% LCFA; K-=1.2% K; K+=2.2% K.
3.4
3.2
Milk Fat %
3
Low Fat
High Fat
2.8
KK+
2.6
2.4
2.2
0
5
10
15
Day Within Period
61
20
`
Figure 6: Correlation of milk fat percent to trans-10 cis-12 concentration (n=16 per
treatment)
Trans-10, cis-12 CLA=0.0928 – 0.014 Milk Fat %
0.6
Trans-10, cis-12 CLA
0.5
P<0.07
0.4
R² = 0.05
0.3
0.2
0.1
0
0
1
2
3
Milk Fat %
62
4
5
6
`
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