• TABLE OF CONTENTS
HIDE
 Title Page
 Acknowledgement
 Table of Contents
 List of Tables
 List of Figures
 Abstract
 Introduction
 Literature review
 Effect of energy and protein on...
 Nutritional factors affecting mineral...
 Nutritional factors affecting mineral...
 Effect of variable energy-protein...
 Summary and conclusions
 Appendix
 Bibliography
 Biographical sketch














Title: Nutritional factors affecting mineral status and long term carry-over effects in ruminants /
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Permanent Link: http://ufdc.ufl.edu/UF00097425/00001
 Material Information
Title: Nutritional factors affecting mineral status and long term carry-over effects in ruminants /
Physical Description: xvi, 282 leaves : ill. ; 28 cm.
Language: English
Creator: Rosero, Oswaldo R., 1942-
Publication Date: 1983
Copyright Date: 1983
 Subjects
Subject: Ruminants -- Physiology   ( lcsh )
Sheep -- Feed utilization efficiency   ( lcsh )
Mineral metabolism   ( lcsh )
Minerals in animal nutrition   ( lcsh )
Animal Science thesis Ph. D
Dissertations, Academic -- Animal Science -- UF
Genre: bibliography   ( marcgt )
non-fiction   ( marcgt )
 Notes
Thesis: Thesis (Ph. D.)--University of Florida, 1983.
Bibliography: Bibliography: leaves 261-280.
Additional Physical Form: Also available on World Wide Web
Statement of Responsibility: by Oswaldo R. Rosero.
General Note: Typescript.
General Note: Vita.
 Record Information
Bibliographic ID: UF00097425
Volume ID: VID00001
Source Institution: University of Florida
Holding Location: University of Florida
Rights Management: All rights reserved by the source institution and holding location.
Resource Identifier: alephbibnum - 000366103
oclc - 10033474
notis - ACA4938

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Table of Contents
    Title Page
        Page i
    Acknowledgement
        Page ii
        Page iii
    Table of Contents
        Page iv
        Page v
        Page vi
        Page vii
    List of Tables
        Page viii
        Page ix
        Page x
        Page xi
        Page xii
        Page xiii
    List of Figures
        Page xiv
    Abstract
        Page xv
        Page xvi
    Introduction
        Page 1
        Page 2
    Literature review
        Page 3
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    Effect of energy and protein on calcium, phosphorus and magnesium retention and mineral storage by sheep
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    Nutritional factors affecting mineral status and long-term carry-over effect in sheep. I: Macro elements, animal performance, hemoglobin and hematocrit
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    Nutritional factors affecting mineral status and long-term carry-over effect in sheep. II: Trace minerals
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    Effect of variable energy-protein and mineral concentrations on blood metabolic profiles in sheep
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    Summary and conclusions
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    Appendix
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    Bibliography
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    Biographical sketch
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Full Text














NUTRITIONAL FACTORS AFFECTING MINERAL STATUS AND
LONG TERM CARRY-OVER EFFECTS IN RUMINANTS









BY

OSWALDO R. ROSERO


A DISSERTATION PRESENTED TO THE GRADUATE COUNCIL
OF THE UNIVERSITY OF FLORIDA IN
PARTIAL FULFILLMENT OF THE REQUIREMENTS
FOR THE DEGREE OF DOCTOR OF PHILOSOPHY



UNIVERSITY OF FLORIDA


1983

















ACKNOWLEDGEMENTS

The author wishes to express his sincere thanks to Dr. Lee

McDowell, chairman of his supervisory committee, for his guidance,

encouragement and supervision in planning and conducting the research

and preparation of this dissertation; and also to Dr. C. B. Ammerman,

Dr. J. H. Conrad, Dr. G. 0. Mott and Professor P. E. Loggins for serv-

ing as members of the supervisory committee and for giving suggestions

and for their time and knowledge toward the completion of this work.

Deep respect and appreciation are offered to Mrs. Nancy Wilkinson

and Mrs. Pamela H. Miles for their assistance and help in all phases

of the experiments, particularly in the laboratory work. The author

is very grateful to Dr. F. G. Martin and Dr. T. Thang for assistance

with statistical analyses.

The writer extends thanks to his fellow graduate students and

to the staff and personnel of the nutrition laboratory, who. were

always available for assistance. Special thanks are due to Mr. Jack

Stokes and Mr. Dane Bernis for his assistance and help in the field

work and preparation of the experimental diets. The writer also

wishes to express his gratitude to "Facultad de Ciencias Veterinarias

de la Universidad del Zulia, Maracaibo, Venezuela," which granted

him the scholarship and financial support which made this study pos-

sible. Also, special thanks are due to "Condes of the University of

Zulia" for additional financial support for this research.









Acknowledgement is made to the American Pfizer Company and

Monsanto Company for supplying vitamins A and D and Santhoquin

for the experimental diets of this research.

Thanks are also extended to Miss Sarah McKee and Mrs. Carol Laine

for the preparation of the manuscript.

The author wishes to give special recognition to his wife,

Yully, and children, Romina, Roberto and Roxana for help, patience

and understanding during the execution of this task.


















TABLE OF CONTENTS

Page

ACKNOWLEDGMENTS ........................................... ii

LIST OF TABLES ............................................ viii

LIST OF FIGURES ........................................... xiv

ABSTRACT .................................................. xv

CHAPTER

I INTRODUCTION ....................................... 1

II LITERATURE REVIEW.................................... 3

Mineral Status of Animals............................... 3
Factors Affecting Mineral Status of
Grazing Ruminants ................................... 4
Genetic Variation............................ ... 6
Age Variation ................................... 7
Physiological State and Level of
Production..................................... 7
Biological Availability......................... 8
Other Factors Affecting Mineral Status
of Animals ................................... 10
Essential Major Mineral Elements.................... 13
Calcium and Phosphorus.......................... 13
Metabolism in ruminants .................... 13
Assessment of Ca and P status............... 17
Magnesium...................................... 24
Metabolism in ruminants..................... 24
Assessment of Mg status..................... 27
Sodium, Potassium and Chloride.................. 30
Metabolism in ruminants..................... 30
Assessment of Na and K status............... 33
Essential Trace Mineral Elements.................... 34
Iron........................................... 34
Metabolism in ruminants..................... 34
Assessment of Fe status .................... 35











Page


Copper, Molybdenum and Sulfur ......................
Metabolism in ruminants .......................
Assessment of Cu and Mo status ................
Zinc and Manganese .................................
Metabolism in ruminants ........................
Assessment of Zn and Mn status .................
Cobalt. .............................................
Metabolism in ruminants........................
Assessment of Co status .......................
Selenium. ...........................................
Metabolism in ruminants........................
Assessment of Se status........................
Energy and Protein ............ .........................
Maternal-Fetal Relationships of Trace Elements
in Ruminants ..........................................

III EFFECT OF ENERGY AND PROTEIN ON CALCIUM,
PHOSPHORUS AND MAGNESIUM RETENTION AND MINERAL
STORAGE BY SHEEP ......................................

Introduction ...........................................
Experimental Procedure .......... ........................


Results and Discussion....................
Body Weight and Blood Parameters......
Liver Minerals .......................
Kidney, Heart, Spleen and Muscle
Minerals .............................
Wool Nitrogen and Mineral Concentra-
trations .............................
Bone Mineral Composition ..............
Rib mineral concentrations........
Metacarpal bone mineral concentra-
tions .............................


Calcium, Phosphorus and Magnesium Reten-
tion .......................................
Summary .........................................


....... 94
....... 102


IV NUTRITIONAL FACTORS AFFECTING MINERAL
STATUS AND LONG-TERM CARRY-OVER EFFECT
IN SHEEP. I. MACRO ELEMENTS, ANIMAL
PERFORMANCE, HEMOGLOBIN AND HEMATO-
CRIT ..................................................

Introduction ..........................................
Experimental Procedure.................................
Results and Discussion....................................


105

105
107
111


...........
...........
...........









Page

Ewe Lambs ........................................ 111
Body weight.................................... 111
Hematocrit, hemoglobin and serum
macro elements...................... ........... 111
Rib mineral concentrations...................... 118
Wool nitrogen and mineral concentra-
tions (macro elements)......................... 126
Milk and colostrum mineral concentra-
tions (macro elements) ........................ 128
Newborn and Weaned Lambs........................... 130
Body weights, hematocrit, hemo-
globin and serum macro elements................ 130
Bone tail and metacarpal mineral
concentration.................................. 133
Conclusions............................................. 136
Summary................................................ 139

V NUTRITIONAL FACTORS AFFECTING MINERAL STATUS
AND LONG-TERM CARRY-OVER EFFECTS IN SHEEP.
II. TRACE MINERALS ...................................... 145

Introduction .......................................... 145
Experimental Procedure ................................. 146
Results and Discussion .............................. 148
Trace Elements .................................... 148
Blood parameters ..... ........................ 148
Liver trace elements concentrations............. 151
Wool trace elements........................... 160
Milk and colostrum............................ 162
Newborn and Weaned Lambs........................... 164
Blood trace elements ......................... 164
Conclusions .......................................... 166
Summary................................................. 168

VI EFFECT OF VARIABLE ENERGY-PROTEIN AND MINERAL
CONCENTRATIONS ON BLOOD METABOLIC PROFILES
IN SHEEP .............................................. 173

Introduction........................ ........ ......... 173
Experimental Procedure .................................. 175
Wether Experiment ......... ........................ 176
Ewe Experiment ................................... 177
Results and Discussion ................................ 178
Wether Experiment ................................. 178
Ewe Experiment..................................... 184
Metabolic blood profiles in weaned
lambs........................................... 198
Summary................................................ 200










Page

VII GENERAL SUMMARY AND CONCLUSIONS....................... 204

APPENDIX .................................................... 214

LITERATURE CITED ............................................ 261

BIOGRAPHICAL SKETCH ......................................... 281


















LIST OF TABLES


Table Page

1. COMPOSITION OF EXPERIMENTAL DIETS .................... 74

2. DAILY INTAKE OF TRACE MINERALS ....................... 78

3. EFFECT OF ENERGY-PROTEIN ON BODY WEIGHT, SERUM
MINERALS, HEMOGLOBIN AND HEMATOCRIT IN SHEEP ......... 79

4. EFFECT OF ENERGY-PROTEIN ON LIVER MINERAL COMPO-
SITION IN SHEEP ...................................... 83

5. EFFECT OF ENERGY-PROTEIN ON TISSUE MINERAL COMPO-
SITION IN SHEEP ...................................... 86

6. EFFECT OF ENERGY-PROTEIN ON WOOL NITROGEN AND
MINERAL CONCENTRATIONS IN SHEEP ...................... 88

7. EFFECT OF ENERGY-PROTEIN ON BONE RIB MINERAL
CONCENTRATIONS IN SHEEP .............................. 91

8. EFFECT OF ENERGY-PROTEIN ON BONE METACARPAL
MINERAL CONCENTRATIONS IN SHEEP ...................... 93

9. EFFECT OF ENERGY-PROTEIN ON CALCIUM, PHOSPHORUS
AND MAGNESIUM RETENTION IN SHEEP ..................... 95

10. EFFECT OF ENERGY-PROTEIN ON CALCIUM RETENTION........ 97

11. EFFECT OF ENERGY-PROTEIN ON PHOSPHORUS RETENTION ..... 98

12. EFFECT OF ENERGY-PROTEIN ON MAGNESIUM RETENTION...... 100

13. PAIRED COMPARISONS BETWEEN DIETS 2 VERSUS 1;
AND 4 VERSUS 3 FOR BODY WEIGHT, Ca PERCENT
IN RIB BONE ASH, Zn, Mn, AND Co IN LIVER AND Ca,
P AND Mg RETENTION IN WETHERS ........................ 101

14. MEANS AND STANDARD ERROR OF BODY WEIGHTS IN EWES
FED FOUR EXPERIMENTAL DIETS .......................... 112

15. MEANS AND STANDARD ERRORS OF HEMATOCRIT, HEMO-
GLOBIN AND SERUM Ca, P, Mg, Na AND K IN EWES
FED FOUR EXPERIMENTAL DIETS .......................... 113


viii










Table Page

16. MEANS AND STANDARD ERROR OF BONE RIB MINERAL
CONCENTRATIONS IN EWES FED FOUR EXPERIMENTAL
DIETS ................................................. 119

17. MEANS AND STANDARD ERROR OF BONE RIB MINERAL
CONCENTRATIONS IN EWES FED FOUR EXPERIMENTAL
DIETS ................................................. 120

18. MEANS AND STANDARD ERROR OF BONE RIB MINERAL
CONCENTRATIONS IN EWES FED FOUR EXPERIMENTAL
DIETS ................................................. 121

19. MEANS AND STANDARD ERROR OF BONE RIB MINERAL
CONCENTRATIONS IN EWES FED FOUR EXPERIMENTAL DIETS.... 122

20. MEANS AND STANDARD ERROR OF WOOL NITROGEN, Ca,
P, Mg, Na AND K CONCENTRATIONS IN EWES FED
FOUR EXPERIMENTAL DIETS............................... 127

21. MEANS AND STANDARD ERROR OF MILK AND COLOSTRUM
Ca, P, Mg, Na AND K CONCENTRATIONS IN EWES
FED FOUR EXPERIMENTAL DIETS ........................... 129

22. MEANS, STANDARD ERRORS OF BODY WEIGHTS, HEMATO-
CRIT, HEMOGLOBIN AND SERUM Ca, P, Mg, Na, AND
K IN LAMBS FROM EWES FED FOUR EXPERIMENTAL
DIETS ................................................. 131

23. MEANS AND STANDARD ERROR OF BONE TAIL MINERAL
CONCENTRATIONS IN NEWBORN LAMBS FROM EWES FED
FOUR EXPERIMENTAL DIETS ............................... 134

24. METACARPAL MINERAL CONCENTRATION IN NEWBORN
LAMBS FROM EWES FED FOUR EXPERIMENTAL DIETS............. 135

25. MEANS AND STANDARD ERROR OF BONE TAIL MINERAL
CONCENTRATIONS IN WEANING LAMBS FROM EWES FED
FOUR EXPERIMENTAL DIETS ............................... 137

26. MEANS AND STANDARD ERRORS OF SERUM Fe, Cu, Zn AND
Se IN EWES FED FOUR EXPERIMENTAL DIETS ................ 149

27. MEANS AND STANDARD ERROR OF LIVER MINERAL
CONCENTRATION IN EWES FED FOUR EXPERIMENTAL
DIETS ................................................. 152

28. MEAN AND STANDARD ERROR OF LIVER MINERAL
CONCENTRATION IN EWES FED FOUR EXPERIMENTAL
DIETS ................................................. 154









Table Page

29. MEANS AND STANDARD ERROR OF WOOL Fe, Cu, Zn, Mn,
Mo AND Se CONCENTRATIONS IN EWES FED FOUR
EXPERIMENTAL DIETS...................................... 161

30. MEANS AND STANDARD ERROR OF MILK AND COLOSTRUM
Fe, Cu, Zn, Mn AND Se CONCENTRATION IN EWES
FED FOUR EXPERIMENTAL DIETS ........................... 163

31. MEAN, STANDARD ERRORS OF SERUM Fe, Cu, Zn AND Se
IN LAMBS FROM EWES FED FOUR EXPERIMENTAL DIETS.......... 165

32. EFFECT OF ENERGY AND PROTEIN IN BLOOD METABOLIC
PROFILES (SMAC 25) IN WEATHERS ........................ 179

33. MEANS AND STANDARD ERRORS OF METABOLIC BLOOD
PROFILE (SMAC 25) IN EWES FED FOUR EXPERIMENTAL
DIETS ................................................. 185

34. MEANS, STANDARD ERRORS OF METABOLIC BLOOD
PROFILE (SMAC 25) IN EWES FED FOUR EXPERIMENTAL
DIETS ................................................. 186

35. MEANS, STANDARD DEVIATIONS AND COEFFICIENT OF
VARIATION OF METABOLIC BLOOD PROFILE (SMAC 25)
IN EWES FED FOUR EXPERIMENTAL DIETS BY
PERIODS ............................................... 187

36. ANALYSIS OF VARIANCE-POOLED PERIOD ANALYSIS OF
METABOLIC BLOOD PROFILE (SMAC 25) IN EWES FED
FOUR EXPERIMENTAL DIETS ............................... 193

37. MEANS, STANDARD ERRORS OF BLOOD METABOLIC PROFILES
(SMAC 25) AND SELENIUM IN WEANING LAMBS FROM
EWES FED FOUR EXPERIMENTAL DIETS ...................... 199

38. CHEMICAL COMPOSITION OF EXPERIMENTAL DIETS
(DRY BASIS) ........................................... 214

39. EFFECT OF ENERGY-PROTEIN ON BODY WEIGHT, SERUM
MINERALS, HEMOGLOBIN AND HEMATOCRIT IN SHEEP............ 215

40. EFFECT OF ENERGY-PROTEIN ON TISSUE MINERAL COMPO-
SITION IN SHEEP ....................................... 216

41. EFFECT OF ENERGY-PROTEIN ON LIVER MINERAL
COMPOSITION IN SHEEP.................................... 218

42. EFFECT OF ENERGY-PROTEIN ON BONE METACARPAL
MINERAL CONCENTRATIONS IN SHEEP ....................... 219









Table Page

43. EFFECT OF ENERGY-PROTEIN ON BONE RIB MINERAL
CONCENTRATION IN SHEEP................................ 220

44. EFFECT OF ENERGY-PROTEIN ON WOOL NITROGEN
AND MINERAL CONCENTRATIONS IN SHEEP ................... 221

45. MEANS, STANDARD DEVIATION, AND COEFFICIENT OF
VARIATION OF BODY WEIGHTS IN EWES FED FOUR
EXPERIMENTAL DIETS .................................... 222

46. ANALYSIS OF VARIANCE-MEAN SQUARES FOR BODY
WEIGHTS IN EWES FED FOUR EXPERIMENTAL DIETS............. 223

47. MEANS, STANDARD DEVIATION AND COEFFICIENT OF
VARIATION OF HEMATOCRIT, HEMOGLOBIN AND SERUM
Ca, P, Mg, Na, AND K IN EWES FED FOUR EXPERIMENTAL
DIETS ................................................. 224

48. ANALYSIS OF VARIANCE-MEAN SQUARES FOR HEMATOCRIT,
HEMOGLOBIN AND SERUM Ca, P, Mg, Na AND K IN
EWES FED FOUR EXPERIMENTAL DIETS ...................... 225

49. ANALYSIS OF VARIANCE-POOLED PERIOD ANALYSIS OF
HEMATOCRIT, HEMOGLOBIN AND SERUM Ca, P, Mg, Na
AND K IN EWES FED FOUR EXPERIMENTAL DIETS............... 227

50. MEANS, STANDARD DEVIATION, AND COEFFICIENT OF
VARIATION OF BONE RIB MINERAL CONCENTRATIONS IN
EWES FED FOUR EXPERIMENTAL DIETS ...................... 228

51. ANALYSIS OF VARIANCE-MEAN SQUARES FOR BONE RIB
MINERAL CONCENTRATIONS IN EWES FED FOUR
EXPERIMENTAL DIETS ............ ........................... 229

52. ANALYSIS OF VARIANCE-POOLED PERIOD ANALYSIS OF
RIB BONE MINERAL CONCENTRATIONS IN EWES FED
FOUR EXPERIMENTAL DIETS............................... .. 230

53. MEANS, STANDARD DEVIATION AND COEFFICIENT OF
VARIATION OF WOOL N, Ca, P, Mg, Na AND K CONCEN-
TRATIONS IN EWES FED FOUR EXPERIMENTAL DIETS........... 232

54. ANALYSIS OF VARIANCE-MEAN SQUARES FOR WOOL N,
Ca, P, Mg, Na AND K CONCENTRATIONS IN EWES FED
FOUR EXPERIMENTAL DIETS ............................... 233

55. MEANS, STANDARD DEVIATION AND COEFFICIENT OF
VARIATION OF MILK AND COLOSTRUM Ca, P, Mg, Na AND
K CONCENTRATION IN EWES FED FOUR EXPERIMENTAL
DIETS .................................................. 234









Table Page

56. ANALYSIS OF VARIANCE-MEAN SQUARES FOR MILK AND
COLOSTRUM Ca, P, Mg, Na AND K CONCENTRATION
IN EWES FED FOUR EXPERIMENTAL DIETS.................... 235

57. MEANS, STANDARD DEVIATION, AND COEFFICIENT OF
VARIATION OF BODY WEIGHTS, HEMATOCRIT, HEMOGLOBIN,
AND SERUM Ca, P, Mg, Na AND K IN LAMBS FROM
EWES FED FOUR EXPERIMENTAL DIETS ...................... 236

58. ANALYSIS OF VARIANCE-MEAN SQUARES FOR BODY
WEIGHTS, HEMATOCRIT, AND HEMOGLOBIN IN LAMBS
FROM EWES FED FOUR EXPERIMENTAL DIETS ................. 237

59. ANALYSIS OF VARIANCE-MEAN SQUARES FOR SERUM
Ca, P, Mg, Na AND K IN LAMBS FROM EWES FED
FOUR EXPERIMENTAL DIETS ............................... 239

60. ANALYSIS OF VARIANCE-MEAN SQUARES FOR BONE TAIL
MINERAL CONCENTRATIONS IN NEWBORN LAMBS................ 240

61. MEANS, STANDARD DEVIATION, AND COEFFICIENT OF
VARIATION OF BONE TAIL MINERAL CONCENTRATIONS
IN LAMBS FROM EWES FED FOUR EXPERIMENTAL DIETS.......... 241

62. ANALYSIS OF VARIANCE-MEAN SQUARES FOR BONE TAIL
MINERAL CONCENTRATIONS IN WEANING LAMBS................. 242

63. MEANS, STANDARD DEVIATION AND COEFFICIENT OF
VARIATION OF SERUM Fe, Cu, Zn, AND Se IN EWES
FED FOUR EXPERIMENTAL DIETS ........................... 243

64. ANALYSIS OF VARIANCE-MEAN SQUARES FOR SERUM
Fe, Cu, Zn, AND Se IN EWES FED FOUR EXPERI-
MENTAL DIETS .......................................... 244

65. ANALYSIS OF VARIANCE-POOLED PERIOD ANALYSIS OF
SERUM Fe, Cu AND Zn IN EWES FED FOUR EXPERIMENTAL
DIETS ................................................. 246

66. MEANS, STANDARD DEVIATION AND COEFFICIENT OF
VARIATION OF LIVER MINERALS IN EWES FED FOUR
EXPERIMENTAL DIETS .................................... 247

67. ANALYSIS OF VARIANCE-MEAN SQUARES OF LIVER
MINERAL CONCENTRATIONS IN EWES FED FOUR EXPERIMEN-
TAL DIETS ............................................. 248

68. ANALYSIS OF VARIANCE-POOLED PERIOD ANALYSIS OF
LIVER Fe, Cu, Zn, Mn, Co, AND Mo IN EWES FED
FOUR EXPERIMENTAL DIETS ................................ 249


xii









Table Page

69. MEANS, STANDARD DEVIATION AND COEFFICIENT OF
VARIATION OF WOOL Fe, Cu, Zn, Mn, Mo AND Se
CONCENTRATIONS IN EWES FED FOUR EXPERIMENTAL
DIETS ................................................. 250

70. ANALYSIS OF VARIANCE-MEAN SQUARES FOR WOOL Fe, Cu,
Zn, Mn, Mo AND Se CONCENTRATIONS IN EWES FED
FOUR EXPERIMENTAL DIETS ............................... 251

71. MEANS, STANDARD DEVIATION AND COEFFICIENT OF
VARIATION OF MILK AND COLOSTRUM Fe, Cu, Zn, Mn AND
Se CONCENTRATION IN EWES FED FOUR EXPERIMENTAL
DIETS ................................................. 252

72. ANALYSIS OF VARIANCE-MEAN SQUARES FOR MILK AND
COLOSTRUM Fe, Cu, Zn, Mn AND Se CONCENTRATIONS
IN EWES FED FOUR EXPERIMENTAL DIETS ................... 253

73. MEANS, STANDARD DEVIATION AND COEFFICIENT OF
VARIATION OF SERUM Fe, Cu, Zn AND Se IN LAMBS
FROM EWES FED FOUR EXPERIMENTAL DIETS.................. 254

74. ANALYSIS OF VARIANCE-MEAN SQUARES FOR SERUM Fe,
Cu, Zn AND Se IN LAMBS FROM EWES FED FOUR
EXPERIMENTAL DIETS ........... ......................... 255

75. EFFECT OF ENERGY AND PROTEIN IN BLOOD METABOLIC
PROFILES (SMAC 25) IN WETHERS......................... 256

76. ANALYSIS OF VARIANCE-MEAN SQUARES FOR BLOOD
METABOLIC PROFILES (SMAC 25) IN EWES FED FOUR
EXPERIMENTAL DIETS............. ......... .............. 257

77. ANALYSIS OF VARIANCE, MEAN SQUARES FOR BLOOD
METABOLIC PROFILES (SMAC 25) AND SELENIUM IN
WEANED LAMBS .......................................... 259


xiii

















LIST OF FIGURES


Figure Page

1. Interactions between levels of energy-protein
and minerals in liver Fe, Cu, Zn, Mn and Co
concentrations in ewes.............................. 157

2. Interactions between levels of energy-protein
and levels of minerals in serum creatine, Ca,
ionized Ca, P and triglycerides (SMAC 25) in
ewes fed four experimental diets (period 1)......... 189

3. Interactions between levels of energy-protein
and levels of minerals in serum bilirubin
(SMAC 25) in ewes fed four experimental diets
(period 2) ......................................... .. 192

4. Effect of period in metabolic
blood profiles (SMAC 25)............................ 197
















Abstract of Dissertation Presented to the Graduate Council
of the University of Florida in Partial Fulfillment of the
Requirements for the Degree of Doctor of Philosophy


NUTRITIONAL FACTORS AFFECTING MINERAL STATUS AND
LONG-TERM CARRY-OVER EFFECTS IN RUMINANTS

By

Oswaldo R. Rosero

April, 1983

Chairman: Dr. L. R. McDowell

Major Department: Animal Science

Four experiments were conducted with sheep to study the nutrition-

al factors affecting mineral status and long-term carry-over effects

on animal performance, blood parameters and mineral composition of

selected tissues. In the first experiment, twelve Florida native

crossbred lambs were randomly assigned to two treatment groups, where

two levels of energy-protein (LEP = .8 x maintenance; HEP = 1.8 x

maintenance) and two levels of minerals (LM, approximate requirements;

HM = 2 to 30 times the requirements) were studied as they affected

Ca, P and Mg retention. Wethers fed HEP + HM or LM diets had

greater gains than those receiving LEP diets. Increased dietary

levels of energy-protein did not affect serum mineral concentrations.

Low mineral-HEP diets apparently depressed liver Fe, Cu and Co. In

trial 1 and trial 2, Ca, P and Mg retention was higher in HEP versus

LEP diets.


xv








In the second experiment, effects of two levels of energy-protein

on sheep mineral status (macro elements) and two levels of minerals

on mineral storage and long-term carry-over effects in sheep were

studied. Forty-eight Rambouillet crossbred ewe lambs (28.5 kg

initial body weight) were randomly assigned to four experimental

diets in a 2 x 2 factorial arrangement. Treatments with high miner-

als were administered for only four months of growing period; ewes

were fed LM diets with HEP and LEP levels for the remainder of the

trial. Levels of energy-protein were more important than levels of

minerals in animal performance. No carry-over effects were observed

in hematocrit, Hb, and serum Ca, P, Mg, Na and K. Feeding high

minerals resulted in a carry-over effect of bone rib P concentration,

expressed as percent of dry, fat-free bone, percent of bone ash and

mg/cc until breeding period; but for gestation-parturition period, no

carry-over effect of P was observed.

In the third experiment, effects of two levels of energy-protein

on mineral status and two levels of minerals on mineral storage and

long-term carry-over effects in sheep were studied for trace elements.

No carry-over effects were observed in serum Fe, Cu, Zn and Se in

ewes fed the original experimental diets (HM) for four months. Carry-

over effects were observed in liver Mo concentrations of ewes fed HM

+ LEP diets until the breeding period and in ewes fed HM + HEP diets

until gestation-parturition period. No carry-over effects were

observed in liver Fe, Cu, Zn, Mn and Se.

In the fourth experiment, no carry-over effects were found in

serum metabolic blood profiles (SMAC 25) of ewes fed HM diets for a

four month period.


xvi















CHAPTER I
INTRODUCTION

Low ruminant productivity in the tropics is due to many limitations,

the most apparent being the low feed intake and poor nutrition (protein,

energy, minerals) which consequently result in poor growth and reproduc-

tive rates. In areas of the world where beef and dairy cattle enterpris-

es depend on natural or improved grassland for the supply of nutrients,

animals are likely to be subjected to undernutrition during the dry per-

iod of the year. During those periods, herbage will have a low nutritive

value, the most overriding factor being a shortage of digestible energy

and protein. Cattle grazing such herbage are unable to meet their re-

quirements for maintenance and generally growth, reproduction and lacta-

tion are impaired.

The dietary requirements for minerals are more difficult to accur-

ately define than those for the organic nutrients because many factors

determine the utilization of minerals. Starving cattle were observed

frequently during the long dry season periods in tropical savannas where

native pasture is the only source of feed during the whole year. Low

nutritive value of native pastures causes slow growth rates and low re-

productive performance. Fluctuation of nutrient content of the pasture

results in a particular pattern of growth rate of animals on native

grasses, that is, rapid growth in the rainy season followed by a loss of

body weight during the dry season. A loss of 30 to 50 kg per animal is

not uncommon during a 5- to 6-month dry season. During the dry season,

energy and/or protein deficiencies limit cattle production but during





2


the rainy season, mineral deficiencies may be the major factor which

affects rate of cattle production, because protein and digestible

energy contents of growing grasses are adequate to meet their require-

ments for maintenance and production.

Cattle do, however, survive such periods of undernutrition by

utilizing their energy conservation mechanism. They are able to

mobilize and deplete body tissue reserves during periods of food

scarcity and to replenish the reserves when food is readily available.

The animals which suffered the greater weight loss tended to lose more

body water and protein; during the earlier stages of undernutrition,

the tissue lost is almost entirely body fat. Therefore, a multitude

of factors influence the productivity of cattle in tropical areas,

particularly inadequate nutrition during the long dry period.

The objectives of this research were as follows: 1) To

investigate the effect of different energy and protein levels on

mineral utilization by the animal; 2) To compare two mineral levels

(high and low) on mineral storage and long-term carry-over effects;

and 3) To compare two energy-protein dietary levels on mineral status

and mineral composition of selected tissues.
















CHAPTER 2
LITERATURE REVIEW

Mineral Status of Animals

Minerals play important metabolic roles in animal nutrition; howev-

er, they are not a source of energy and protein but are essential for

biosynthesis of essential nutrients. Mineral deficiency usually involves

several minerals as well as other conditioning factors; however, the de-

ficiency symptoms of one mineral may predominate and affect the perfor-

mance of the ruminant. The numerous mineral interrelationships in nu-

trition are not completely understood. In addition, interrelationships

of minerals with vitamins, protein and energy further complicate the

problem of providing adequate nutrition. Mineral interrelationships

also exist between soil, plants and animals.

Numerous mineral deficiencies, imbalances and toxicities are eco-

nomically important in livestock production throughout the world. For

several decades, a major goal in mineral research has been to discover

and/or develop simple and accurate biochemical measures of the status

of animals for the minerals in which there are important practical prob-

lems (Miller and Stake, 1974). Like soils and plants, animal mineral

status is influenced by many factors. Nevertheless, when appropriate

diagnosis is used, animal tissue concentrations are often better indi-

cators of the mineral status of livestock than either plant or soil con-

centrations (McDowell, 1976).








Many biochemical measures employed in diagnosing mineral problems

involve analysis of tissues and fluids of excreta. The most useful nor-

mally follow discovery of fundamental information on the element; cobalt

and iodine are excellent examples (Mills and Williams, 1971). Estima-

tion of whole blood or plasma mineral concentrations has wide applica-

tion (Underwood, 1971). However, often blood measures are employed where

they are not especially useful. Because of the essential role of many

minerals in enzymes (for example, serum ceruloplasmin and copper), en-

zyme activity has been developed into a useful measure in certain miner-

al deficiencies. Perhaps this approach will have far wider application

in the future (Mills and Stake, 1974).

At present, 22 mineral elements are believed to be essential for

the higher forms of animal life. These minerals comprise seven major or

macronutrient elements, calcium (Ca) phosphorus (P), potassium (K),

sodium (Na), chlorine (Cl), magnesium (Mg) and sulfur (S), and 15 trace

or micronutrient elements, iron (Fe), iodine (I), zinc (Zn), copper (Cu),

manganese (Mn), cobalt (Co), molybdenum (Mo), selenium (Se), chromium

(Cr), tin (Pb), vanadium (Va), fluorine (F), silicon (Si), nickel (Ni)

and arsenic (As). The essentiality of the last six, often referred to

as the "newer" trace elements, is based almost exclusively on growth ef-

fects with animals in highly specialized conditions. In addition to the

22 essential minerals, all plant and animal tissues contain a further 20

to 30 mineral elements, mostly in small and variable concentrations

(Underwood, 1981).

Factors Affecting Mineral Status of Grazing Ruminants

Ideally, animal scientists would like to determine the mineral sta-

tus of an animal by measuring the mineral content of one tissue which is









readily available from a live animal (Conrad, 1978). However, this is

not possible for many reasons. All mineral deficiencies and most ex-

cesses, in their more severe forms, are manifested by characteristic

clinical and pathological disturbances in the animal. When taken in

conjunction with clinical and pathological observations, appropriate

chemical analyses and biological assays of tissues and fluids of animals

are valuable aids in the early detection of mineral abnormalities in

livestock. The choice of tissue or fluid for analysis varies with the

mineral under investigation. Blood, urine, saliva and hair have obvi-

ous advantages because of their accessibility without sacrifice of the

animal. Body tissue sampling presents more difficulties, although sim-

ple liver and tail bone biopsy techniques are now available (Underwood,

1981). Tremendous progress has been made during the past 50 years in

identifying and correcting these mineral deficiencies and toxicities.

However, in the major ruminant livestock-producing areas of the world,

relatively little is known about mineral status of soils, vegetation or

animals. Mineral analyses are complicated and expensive. Therefore, we

are interested in selecting and analyzing the minimum number of tissues

which are most indicative of the mineral status of the animal (Conrad,

1978).

Many research approaches have been suggested for detecting mineral

deficiencies or imbalances in livestock. According to Underwood (1966),

a good diagnosis of mineral disorders in animals should include clinical

signs, pathological examinations and chemical analyses of soils, animal

tissues and feeds. However, subclinical deficiencies or even acute min-

eral deficiencies are difficult to diagnose since many deficiencies have

a common expression as poor performance in animals. The Netherlands'








Committee on Mineral Nutrition (CMN, 1973) has also indicated that clin-

ical signs alone cannot be used as a guide for identifying a given miner-

al deficiency unless it is confirmed by tissue analysis as well as feed

analysis. The mineral status of the animal is influenced by genetic var-

iation, age, physiological state, level of production, biological availa-

bility of the mineral in the diet, parasitism, season and soil ingestion.

Genetic Variation

The effect of breed differences on mineral requirements has often

been observed in livestock (Phillips, 1956; Correa, 1957; Wiener and

Field, 1969). In Brazil, Bos indicus exhibited clinical Co deficiency

when fed forage containing 0.080 ppm Co while Bos taurus were not affect-

ed until the Co level dropped to 0.05 ppm or lower (Correa, 1957). Payne

(1966) suggests the possibility that unacclimatized cattle of temperature

type which sweat profusely and lose saliva and mucus from the mouth may

lose significant quantities of minerals, particularly in the arid tropics.

There is also extensive evidence for marked animal variation within breeds

for the efficiency of absorption of minerals from the diet: 3-35% for Mg

in dairy cows, 40-80% for P and 2-10% for Cu in adult sheep (Field, 1981).

When different breeds of sheep grazed certain pastures in Scotland, one

breed exhibited signs of Cu poisoning whereas another showed signs of Cu

deficiency (Wiener, 1966; Wiener et al., 1977). The most probable cause

for this apparent breed variation in dietary requirements for some micro-

elements could be genetic differences in the efficiency of absorption of

the mineral in the diet (Field, 1981).

Enormous differences exist in the extent of practical problems which

exist among species and classes of animals for a given mineral element (W.

Hiller, 1981). For instance, with Zn, the greatest practical problems









are deficiencies in swine feeding. In contrast, swine are virtually never

deficient in Mn. Without special attention, Mn deficiency can easily be

a serious problem with poultry W-. Miller, 1981). Even the genetic makeup

of a given species of animal can materially affect mineral requirements

and metabolism. The far higher instance of milk fever among Jerseys than

Holsteins reflects such an effect (Miller, 1979). Likewise, Blackface

sheep require and tolerate substantially more copper than Welsh (Wiener

and Field, 1970).

Wiener et al. (1978) found breed differences in the absorption of

dietary Cu by growing lambs and Field and Suttle (1979) found much great-

er variation in the absorption of Mg between monozygotic twin cows. It

is, therefore, suggested that future recommendations of dietary require-

ments, particularly minerals, will take into account breed differences in

the needs of the animal (Field, 1981).

Age Variation

Young animals may be more efficient in metabolizing specific nutri-

ents than mature animals. In mature cows, homeostatic control mechanisms

which regulate the Zn content of tissue are much more effective than in

calves; therefore, mature cows probably are able to tolerate higher con-

centrations of dietary Zn (Kincaid et al., 1976). As indicated by Rook

and Storry (1962), about 30% of skeletal Mg in young animals can be mob-

ilized under conditions of Mg deprivation while in adult animals, only 2%

of bone Mg can be used for physiological needs.

Physiological State and Level of Production

Mineral requirements are highly dependent on the level of productiv-

ity and physiological state of the animal (NRC, 1976). As an example, a

young, pregnant beef cow during her first lactation would have








substantially higher mineral requirements than a mature dry cow. High-

yielding milking cows obviously require much more dietary Ca and P than

low-yielding cows because of the richness of milk in those elements. The

Ca requirements of laying hens tend to follow a similar pattern with in-

creasing egg production but those of P do not. Growing chicks and pigs

consume similar types of diet but chicks require nearly twice the dietary

concentration of Ca and some 20 times the concentration of Mn required by

growing pigs (Underwood, 1981). The minimum Zn requirements for spermato-

genesis and testicular development in young male sheep are significantly

higher than they are for body growth (Underwood and Somers, 1969).

Biological Availability

The relative biological availability of the desired element in a

compound or supplement is one of the major considerations in the selec-

tion of a suitable source of the element (Ammerman and Miller, 1972).

Numerous dietary factors, including protein source and level, interrela-

tionships among the mineral ions and certain chelating agents, influence

the utilization of mineral ions. With some elements, the chemical form

has a major impact on the availability of the element. For instance, Fe

is far more available as ferrous sulfate than as ferric oxide (E. Miller,

1981). Likewise, frequently, ferrous compounds are much more available

than ferric compounds. Generally, monovalent elements such as Na, K and

Cl are highly available. Even so, the effect of the monovalent element,

F, is materially affected by the nature of the monovalent element; sodium

fluoride is more toxic than the same amount of F in phosphate compounds

11 miller, 1981). No element is ever completely absorbed and utilized;

some of it is always lost in the normal digestion and metabolic processes.

Before a required nutrient can be of nutritional value, it must be in a








form that can be digested, absorbed and transported to the part of the

body where it is utilized for its essential function (Peeler, 1972).

Other constituents of the diet often have a major impact on the

amount of minerals needed and tolerated. For instance, the Cu require-

ment and tolerance are very closely related to the Mo in the diet. As

the Mo increases, the need and tolerance for Cu also increases. Even

the form of the Mo seems to have an influence, with that in natural

forages having more impact than added as inorganic Mo (Miller, 1979).

In many respects, the dietary requirements for minerals are more

difficult to accurately define than those for the organic nutrients be-

cause many factors determine the utilization of minerals. For example,

interrelationships among minerals or relationships between minerals and

organic fractions may result in enhanced or decreased mineral utilization.

Numerous mineral interrelationships which affect requirements and mineral

status of the animal include Ca-P, Ca-Zn, Cu-Mo sulfate, Cu-Fe, Se-Ar,

Se-S, Fe-P, Na-K and Mg-K. The organic constituents of the diet can

have a major impact on the amounts of different mineral elements needed

and tolerated. A good illustration is the relationship between vitamin

E and Se, and vitamin B12 and Co; the effect of vitamin D on Ca and P

metabolism is also well known. Goitrogenic substances and chelates such

as oxalic acid and phytic acid each influence specific mineral require-

ments (McDowell, 1976). A good example of these mineral interactions

was reported by Suttle and Mills (1966); signs of Cu toxicity in pigs

fed 250 ppm Cu or Cu sulfate as a growth stimulant appear when the con-

comitant dietary intakes of Zn and Fe are "normal"; i.e., adequate in

the absence of supplementary Cu sulfate; but no such signs of Cu toxicity

appear when additional Fe and Zn are supplied at the rate of 150 ppm Fe








and 150 ppm Zn. In fact, this level of supplementation with Fe and Zn

afforded protection against Cu given at the extremely high level of 450

ppm (Underwood, 1979).

In conclusion, many factors, such as age and physiological status

of the animal (growth, lactation), levels of various dietary components,

duration and route of exposure, and biological availability of the com-

pound, influence the level at which a mineral element causes an adverse

effect and consequently affects mineral status of the animal (NRC, 1980).

Other Factors Affecting Mineral Status of Animals

Parasites affect the mineral status of animals; according to McDowell

(1976), parasitism can produce Fe deficiency in grazing animals. Strong-

ylus has been shown to reduce liver Cu concentration in ruminants (Boya-

zoglu, 1973).

Adequate intake of forages by cattle is essential in meeting miner-

al requirements. Factors which greatly reduce forage intake, such as

low protein (< 7.0%) content and increased maturity, lignification and

stem-leaf ratios all reduce the total mineral consumed (McDowell, 1976).

Another important factor to take into consideration in tropical

areas is the effect of season on the mineral status of the animal, par-

ticularly in areas where pasture is the only source of feed for the cat-

tle during the whole year. The mineral composition of forages varies

according to many factors; among these are the age of the plant, the

soil and fertilization, differences among species and varieties, seasons

of the year and the cutting intervals (Gomide, 1978). Seasonal factors

like light, temperature and rain could justify certain variations in the

chemical composition of forage during the year. The marked decline of P

and K as plants mature is not paralleled by comparable declines in trace








minerals. Whole plant concentrations of trace minerals may increase, de-

crease or show no consistent change with stage of growth, plant species

or soil and seasonal conditions. Changes in trace mineral concentrations

of forages related to season and stage of growth are of greater signifi-

cance to the animals in areas in which marginal levels are present (Con-

rad, 1978). On the basis of low tropical forage mineral concentrations

during the dry season, it is logical to assume that cattle would most

likely suffer mineral inadequacies during this time. However, grazing

cattle were more prone to develop Co or P deficiencies and the clinical

signs were most severe after the rains when pastures were green and plen-

tiful. Increased incidences of mineral deficiencies during the wet sea-

son are less related to forage mineral concentrations than to the greatly

increased requirements for these elements by the grazing animal (Correa,

1957; Van Niekerk, 1974; McDowell, 1976). During the wet season, live-

stock gain rapidly since energy and protein supplies are adequate. As-

sociated with rapid growth during the wet season, mineral requirements

are high while during the dry season, inadequate protein and energy re-

sult in the animal losing weight, thereby greatly limiting mineral re-

quirements (McDowell, 1976).

The grazing animal obtains its intake of microelements from a vari-

ety of sources: from different plants in a mixed sward, from different

parts of the same plant and from the soil (Field, 1981). In view of the

importance of soil ingestion as a source of mineral elements to the graz-

ing animal, Healy (1973) suggested not only the usual sequence, soil-

plant-animal, be considered but also a direct soil-animal effect. Graz-

ing livestock obtain part of their mineral supply from sources other than

forage, particularly from water and soil. Peak soil ingestion is favored








by soils with a weak structure and poor drainage, by high stocking rates,

by high earthworm populations and during the dry season when pasture

growth is poor (McDowell and Conrad, 1977). In New Zealand, animal in-

gestion of soil can reach 75 kg for sheep and 600 kg for dairy animals

(Healy, 1974). These amounts would represent daily intakes of approxi-

mately 200 and 1600 g, respectively, for sheep and cattle. Mayland et

al. (1975) determined soil ingestion by cattle under semi-arid conditions

in southern Idaho and found values ranging from 3 to 30% of soil in fecal

dry matter, with an average of 14%. Soil and dust contamination of herb-

age can at times provide a further significant source of minerals to graz-

ing animals, especially when grazing intensity is high or when pasture

availability is low. With elements such as Co and I which occur in soils

in concentrations usually much higher than those of the plants growing on

them, soil ingestion can be beneficial to the animal (Underwood, 1981).

By contrast, the Cu antagonists, Mo and Zn, are biologically available

in soils and their ingestion from soil contamination of herbage may be a

factor in the etiology of hypocuprosis in cattle and swaybackk" (Cu defi-

ciency) in sheep (Suttle et al., 1975).

Rosa (1980) studied the effect of soil ingestion in sheep and re-

ported that inclusion of 10% Costa Rican soil decreased body weight, in-

creased unabsorbed P and decreased apparent and true P absorption. Also,

metabolic fecal P and P retention values were numerically lower. In a

recent publication, Fries and Marrow (1982) reported a study of soil in-

gestion by dairy cattle. Fecal samples were obtained from animals of

various management groups from nine dairy herds. Titanium of feces was

the indicator of soil ingestion which was calculated for 60% digestibil-

ity of the total ration dry matter. The authors found mean soil ingestion








as percent of dry matter intake by groups of yearling heifers and dry

cows ranged from .52 .11 to .81 .10 for those confined to concrete;

from .25 .04 to 2.41 .26 for those with access to unpaved lots with

no vegetation; from 1.56 .21 to 3.77 1.50 for those with access to

unpaved lots with sparse vegetation; and from 1.38 .33 to 2.46 .50

for those on pasture but receiving supplemental feed as well. The aver-

age soil intake by herds on pasture ranged from 4 to 8% of dry matter in-

take when the cows received no other feed (Healy, 1968). Ingestion of

soil by grazing sheep (Healy and Ludwing, 1965) and beef cattle on range

(Mayland et al., 1975) was similar relative to dry matter intake to that

for dairy cows. Soil ingestion by both cattle and sheep could be reduced

markedly when pasture was supplemented by other feeds (Healy, 1968; Healy

et al., 1967; Healy and Drew, 1970).

Essential Major Mineral Elements

Calcium and Phosphorus

Metabolism in ruminants

Calcium and P have been recognized as important essential mineral

elements. They are major constituents of bones and teeth, and their

roles in numerous other physiological and biochemical processes are cru-

cial to the well-being of all animals. Calcium and P depend on vitamin

D and Mg for proper utilization. Naturally occurring deficiencies of Ca

and P in domestic animals usually develop in quite different circumstan-

ces and a dual deficiency, in which the two minerals are equally limiting,

is rare. Phosphorus deficiency is predominantly a condition of grazing

ruminants, especially cattle, whereas Ca deficiency is more a problem of

hand-fed animals, especially pigs and poultry (Underwood, 1981).








Due to highly regulated control mechanisms, most animals will de-

plete a considerable portion of bone Ca reserves before most other func-

tions are impaired by lack of Ca. In contrast, several functions and

performance measures, including growth, reproduction and milk synthesis,

can be impaired very quickly by inadequate dietary P (Miller, 1981).

Calcium is required in large quantities for skeletal development

and approximately 99.5% of the body Ca is found in the bones. There-

fore, Ca requirements are highest during periods of rapid skeletal de-

velopment and during periods of lactation since milk contains approxi-

mately .12% Ca (Conrad, 1978). Although osteogenesis constitutes the

major demand for Ca, it also has several regulatory functions.

Phosphorus is a factor in the metabolism of almost all nutrients

because of its role in enzyme activity. Approximately 80% of P is found

in the osseous tissue where it constitutes about 16.5% of the bone ash.

The remaining 20%, found in soft tissue, has vital metabolic roles in

carbohydrate metabolism in the formation of hexosephosphate, creatine

phosphate and adenylic acid, and in protein metabolism where it is pres-

ent in nucleoproteins and phosphoproteins (Conrad, 1978).

Normal metabolism of Ca and P requires an adequate supply of vita-

min D which is especially involved in intestinal absorption of these min-

erals. Severe deficiency of Ca and P causes inadequate calcification of

the growing skeleton and other deviations in bone development, of which

the best known is rickets in young cattle, also often caused by deficien-

cy of vitamin D. In mature cattle, excessive demineralization of the

skeleton, called osteoporosis, may result (CMN, 1973).

The most sensitive and earliest biochemical measure of P deficiency

is a reduction in serum inorganic P (Underwood, 1966). Values









consistently below 4.5 mg/100 ml in poultry, cattle and sheep or below

6.0 in swine are indicative of P deficiency. However, the CMN (1973)

did not consider serum inorganic P to be sufficiently sensitive to rec-

ommend it in diagnosing problems with cattle as forage analyses give ear-

lier and more detailed information. Even though serum Ca does decline

with deficiency, especially in some species and ages of animals, the ho-

meostatic or physiological mechanisms regulating it are more effective

than for P or most other minerals (Underwood, 1966; Miller, 1970).

Concentration of Ca in blood plasma is subject to hormonal control.

Normal concentration is 9 to 12 mg/100 ml. Lowered values occur in new-

ly calved, highly productive cows. In cows with milk fever, values fall

markedly even if the ration contains sufficient Ca; with rations severely

deficient in Ca, deficiency is particularly likely in young cattle (CMN,

1973). Calcium deficiency is not common among grazing cattle, except for

high milk-producing cows or those kept on pasture produced by acid and

sandy soils of humid areas with no legumes (Underwood, 1966).

Parturient hypocalcemia is a metabolic disease associated with par-

turition and the initiation of lactation. It is characterized by hypo-

calcemia, hypophosphatemia and hypomagnesemia. This failure of the Ca

homeostatic mechanism has been associated with many factors (Jorgensen,

1974; Hibbs, 1950). Recent theories as to the cause of parturient par-

esis are related to the metabolism of vitamin D. In normal metabolism,

vitamin D is converted to 25-hydroxycholecalciferol (25-OHD) and then to

1,25 (OH) 2D in the kidney prior to exerting its potent Ca mobilization

action on bones and intestines (DeLuca, 1974). A failure of the kidneys

to biosynthesize sufficient 1,25(OH) 2D was suggested as a possible ex-

planation for parturient hypocalcemia (Fraser and Kodicek, 1970).








However, recent studies have demonstrated that the kidney is responsive

to this hypocalcemia (Horst et al., 1977; Horst et al., 1979). It has

therefore been proposed that parturient hypocalcemia results from an un-

explained target organ resistance to one of the Ca-mobilizing hormones

(Yarrington et al., 1977; DeLuca, 1977).

Excessive dietary Ca interferes with the metabolism of several oth-

er mineral elements and some organic constituents of the diet. The im-

portance of Ca:P ratios is widely recognized but this ratio is less crit-

ical with ruminant animals than in simple stomach animals. Phosphorus

contributes about 1% of the total animal body weight but unlike Ca, only

80% of the total quantity is found in the bones and teeth. The remaining

20% is distributed throughout the body in every living cell, being in-

volved in almost all metabolic reactions (Church, 1971). About 10% is

combined with protein, lipids and carbohydrates and in other compounds

in blood and muscle. The remaining 10% is widely distributed in various

chemical compounds (Harper et al., 1979).

Range cattle depend heavily on native pasture for a large share of

their P and Ca needs. Most pasture and range forage contains adequate

amounts of Ca. Legumes are an excellent source and usually supply the

animals' needs when included in the diet (NRC, 1975). In plants, P is

an important constituent of a number of biologically important organic

compounds such as nucleoproteins, sugar phosphate, ATP, etc., and defi-

ciency symptoms including chlorosis and death of older leaves, greatly

reduced plant size and purple coloration due to anthocyanin production

(Whiteman, 1980).

Sousa (1978), in northern Mato Grosso, Brazil, reported the highest

mean forage P concentrations as .2% during the wet season compared to








.08% P during the dry season. Calcium, however, was higher during the

dry season (.67%) than during the wet season (.34%).

Assessment of Ca and P status

Blood plasma. The concentration of Ca in blood plasma is influenced

only by severe deficiency whereas that of inorganic P cannot be used as a

practical criterion at all (CMN, 1973). Underwood (1966) considered Ca

levels in blood plasma as a good indicator of the Ca status of grazing

animals and suggested 9-11 mg Ca/100 ml blood plasma as the normal level.

Cunha et al. (1964) reported the value of 10-12 mg Ca/100 ml blood serum

as the normal level for healthy cattle, with deficiency occurring when

these values fall below 8 mg/100 ml. Wise et al. (1963) found that be-

cause of the homeostatic mechanism, cattle tended to resist the depletion

of plasma Ca. Underwood (1966), on the other hand, indicated that heif-

ers and young cows, having a more efficient mechanism for mobilizing Ca

from the bone, were more resistant to milk fever than older cows. In

sheep, blood levels of Ca below 9 mg/100 ml serum indicate a Ca deficien-

cy (hypocalcemia) according to the NRC (1975). Lebdosoekojo (1977) re-

ported that serum Ca levels were not affected by complete mineral supple-

mentation but seasonal fluctuations were noted.

Duncan (1958) reported that P was less readily mobilized from bone

than Ca. Thus low serum inorganic P concentration is the first indica-

tion of a dietary P deficiency. Cunha et al. (1964) considered cows with

concentrations lower than 5 mg P/100 ml serum as deficient. Similarly,

Underwood (1966) indicated that plasma inorganic P was a satisfactory

criterion for assessing the P status of animals. Underwood (1966) re-

ported that in P-deficient animals, there is a negative correlation be-

tween plasma P and plasma Ca. As plasmaP decreases during P deficiency,








plasma Ca increases until values of 13 to 14 mg% are reached. Chicco et

al. (1973) reported that dietary P supplementation increased plasma P

levels but depressed plasma Ca concentration; conversely, high dietary

Ca tended to reduce plasma P levels. Reed et al. (1974) reported that

serum inorganic P levels were increased by supplementing cattle with bone

meal but Ca and Mg levels were depressed.

Inorganic P contents of plasma differed between animals in a herd

and between months within animals (Blosser et al., 1951). There was a

tendency for inorganic P concentration in plasma to decrease just prior

to parturition as did Ca concentration, with the lowest point at the time

of calving (Wilson et al., 1977). McAdam and O'Dell (1982) studied min-

eral profiles of blood plasma of lactating dairy cows and found that at

parturition, plasma P concentration decreased for all animals except

young cows fed plain salt. Phosphorus concentration remained fairly

constant for animals on each treatment (salt versus minerals) throughout

lactation, with only minor elevations and depressions.

Nursing cows grazing P-deficient pasture in Florida had an average

serum P value of 2.55 mg/100 ml of serum (Becker et al., 1933). This P

level was raised to 4.02% after the cows had been supplemented with bone

meal. Increasing inorganic P in the plasma results in the formation of

a colloidal Ca-phosphate complex that is rapidly removed from the circu-

lation (Irving, 1973). Kitchenham et al. (1975) found the mean blood

serum inorganic P of the rapidly growing calves to be higher (P < .01)

than in conventionally reared heifer calves. Growth rate was signifi-

cantly correlated (P < .05) with concentration of serum inorganic P in

calves reared by the conventional system but not with the rapid growth

system. Plasma percentages of inorganic P were highest in first








lactation cows and declined with subsequent lactations. Season of calv-

ing had a significant effect on plasma inorganic P, with cows calving in

November to December having the highest concentrations. The season of

calving effect on plasma inorganic P cannot be attributed to diet since

ration components and proportion of grain to concentrate remained con-

stant during the trial. Also, time of sampling should be standardized

to reduce the effect of diurnal variation in plasma concentration of in-

organic P (Forar et al., 1982).

The CMN (1973) considers that the concentration of inorganic P in

blood plasma varies widely because of factors not well understood.

Little et al. (1971) reported that the rise in inorganic P of blood plas-

ma, which occurs with time, is due to hydrolysis of organically combined

P; in this case, ATP from the cellular fraction. Inorganic P in blood

is markedly affected by recent dietary P intake in cattle (Little, 1968,

cited by Little, 1972). Another experiment conducted by Cohen (1974) re-

ported that blood plasma P concentration in cattle was significantly re-

lated to P intake (P < .05) but the relationship varied (P < .05) depend-

ing on time of the day at which samples were collected. In the work done

by Gartner et al. (1965, cited by Little et al., 1971), the blood plasma

inorganic P was substantially increased by excitement and exercise.

Levels of P in blood plasma of cattle from two different regions of

Costa Rica were reported by Kiatoko (1976) to be 3.3 .6 and 3.2 .5

mg/100 ml of plasma. The same author studied the mineral status of beef

cattle herds from four soil order regions of Florida and found that mean

plasma P in all regions was above the critical level of 4.5 mg/100 ml re-

ported by Underwood (1966) during the wet season, while during the dry

season, concentrations in the southeast region (Histosol soil order) were








below this level. Of all the animals studied, 13% had low plasma P dur-

ing the dry season (Kiatoko et al., 1982).

Lebdosoekojo et al. (1980) studied the mineral nutrition of beef

cattle grazing native pastures on the eastern plains of Colombia and re-

ported the concentrations of serum minerals in cows given supplemental

NaCl; only serum P was below the deficient level during the rainy season

but during the dry season, P content of the forage increased. However,

forage intake was low because of limited availability. Therefore, an

increase in serum P concentration was apparently due to a decrease in P

demand at the tissue level or an increase in availability of P in the

forage.

Bone Ca and P. About 99% of the Ca and 80% of the P are found in

the bones and teeth so that bone formation and maintenance are quantita-

tively their most important functions (Underwood, 1981). Abnormalities

of bones and teeth can occur at any age. Rickets is the term used to de-

note the skeletal changes which result from defective calcification of

the growing bone in young animals. Osteomalacia is used to describe the

condition in which excessive mobilization of minerals, particularly Ca

and P, has occurred in adult bone. Withdrawal of Ca and P from the bones

occurs normally and regularly in dairy cows at the height of lactation

and in hens during intensive egg-laying, even when intakes are otherwise

adequate. Withdrawal of minerals during periods of inadequate intake

does not take place equally from different parts of the skeleton. The

spongy bones, ribs, vertebrae and sternum, which are the lowest in ash,

are the first to be affected. The compact shafts of the long bones such

as humerus, femur and tibia and of the small bones of the extremities are

the last reserves to be used. In each case, the essential change is a








reduction in the total mineral content of the bones, with little alter-

ation in the proportions of the minerals in the remaining ash (Underwood,

1981). In a longer experiment (14-18 months) with growing sheep fed a

moderately P-deficient but otherwise adequate diet, the total ash concen-

tration of the ribs and vertebrae were some 20% lower and that of the

long bones over 8% lower than those of similar bones from sheep on the

same diet supplemented with phosphate (Stewart, 1934-35, cited by Under-

wood, 1981).

Bone ash consists almost entirely of Ca and P salts and the relative

amounts of these elements show little variation; consequently, the ash

content of bone is commonly used as a measure of the state of Ca and P

nutrition (Maynard and Loosli, 1971). The nature of the diet can affect

the mineral relationships in bone, even though the ash content is not

appreciably changed. In mammals, the bone is made of approximately 36%

Ca, 17% P and .8% Mg, based on dry fat-free bone, as reported by Maynard

and Loosli (1971). Bone is not static structure; there is an active me-

tabolism. Isotope studies have shown that there is a continuous inter-

change of Ca and P between the blood and bone and between various parts

of the bones. Therefore, Ca and P in the body are in dynamic state,

similar to the situation for fat and protein, and the net result of the

interchange determines the nutritional status with respect to a given

physiological need (Maynard et al., 1979).

Little (1972), in Australia, reported that cattle fed a P-deficient

ration (8% crude protein and .08% P, dry matter basis) for 6 weeks showed

levels of 66.8 2.7 and 61.8 1.5% rib ash, 24.5 .5 and 23.8 1.7%

Ca and 11.5 .5 and 11.1 .4% P on dry fat-free bone basis at the be-

ginning and end of this period, respectively.








Ammerman et al. (1974) conducted an investigation of mineral com-

position of tissues from beef cattle under grazing conditions in Panamg

and reported levels of 60.5 to 67.7% for bone ash, 37.6 to 38.2% for Ca

and 17.6 to 18.1% for P./ Kirk et al. (1970) reported that cows on phos-

phate fertilized pastures had metacarpal and metatarsal bones of greater

density than cows on unfertilized pastures; the respective values were
3 3
2.00 to 2.05 g/cm as compared to 1.96 g/cm3. Kiatoko (1976) obtained a

value of 48.2 to 55.6% bone ash on dry, fat-free bone basis in cattle

grazing on Costa Rican farms which had less than .33% P in the forages.

Mendes (1977) found bone ash Ca means ranged from 36.96 to 38.45% and P

ash from 15.13 to 15.54%. When he expressed these means as percent dry, fat-

free bone, the Ca means ranged from 22.7 to 24.8% while P ranged from

9.38 to 10.1% in cattle.

Cohen (1973) correlated the P concentration in pasture, blood, hair

and bone samples. There was a significant correlation (r = .97) between

P in the pasture and in the dry, fat-free ribs. Bone density, bone ash

and mineral concentrations of whole metacarpal bones were lower during

the rainy season for bulls receiving NaCI only (Lebdosoekojo et al.,

1980). This finding suggests that the animals had mobilized bone

reserves during the rainy season to sustain body needs and it is in

agreement with the serum data.

Pond et al. (1975) reported that pigs fed high Ca:P ratios (1.2:1)

had a higher bone ash content than the pigs fed a normal Ca:P ratio

(.5:.4) in the diet. In another experiment conducted with pigs, Bayley

et al. (1975) reported bone ash and breaking strength were increased by

P supplementation at levels of .32 and .48%. As reported by Benzie et

al. (1959) ribs of ewes were more readily rehabilitated and resorbed









than long bones; consequently, status of certain minerals is better de-

tected by ribs. However, Blincoe et al. (1973) found no difference in

Ca and Mg concentrations in caudal vertebrae, right rib and femur.

Campo and Tourtellote (1967) reported Ca and P contents in spongy parts

of long bones of calves of 36.5 to 37.1% and 16.4 to 18.7%, respectively.

Sousa et al. (1979) reported mineral data from six farms in north-

ern Mato Grosso, Brazil. Rib bone ash means were 37.7 2.5 and 15.5

.06% for Ca and P, respectively, during the dry season and 37.6 2.7

and 15.0 .7% for Ca and P, respectively, during the wet season. The

levels of rib bone ash Ca are considered normal but P is below the nor-

mal level and considered as borderline to deficient. Rib bone ash P

levels in this study were based on non-productive animals. In the same

study, they observed that mean rib ash P during the dry season was high-

er than during the wet season, even when forage P levels were low, sug-

gesting a higher requirement during the wet season compared with the dry

season.

An experiment conducted by Peducasse (1982) in tropical areas of

Bolivia to determine the mineral status of beef cattle during the dry

season found deficient levels of Ca (< 37.6%) and P (< 17.6%) in bones

(as % bone ash). Ninety-six percent of the bone samples had Ca levels

lower than 34% bone ash and 45% of the samples were less than 17% P.

Rosa (1980) studied bone ash concentration as affected by P levels

in the diet and found that added dietary P produced a 3.9% increase (P

< .01) in bone ash. Conversely, excess dietary Al affected bone ash by

causing a 2.3% reduction (P < .01) in that parameter. Phosphorus con-

tent in bone ash was also affected by dietary P, with additional P in-

creasing (P < .01) bone P from 16.8 to 17.2%. He also reported no major








effects of dietary P, Fe or Al on bone Ca. Valdivia (1977) also observed

decreased bone ash percentages in sheep fed 2,000 ppm dietary Al. Recent

studies by Hidiroglou et al. (1982) in relation to the chemical composi-

tion of sheep bones as influenced by Mo supplementation reported greater

concentration in the latter ossification portion of the bone in wethers

was unaffected by dietary Mo. The compact shaft contained more ash, Ca,

P and Mg than the proximal and distal parts of the bone.

Some research groups consider serum inorganic P to be sufficiently

sensitive and recommends it for diagnosing P deficiency. However, it is

significant that the CMN (1973) recommends forage analyses because stud-

ies have shown that earlier and more detailed information can be obtained.

It is apparent that both serum inorganic P and forage P analyses are ef-

fective in determining the P status of grazing ruminants and that either

or both can be used (Conrad, 1978).

Magnesium

Metabolism in ruminants

The most important practical Mg deficiency problem in farm animals

is the condition, largely confined to lactating cows, known as grass

tetany (Miller et al., 1974; Underwood, 1966). The clinical signs of

tetany are caused by inadequate Mg in serum and other extracellular flu-

ids; however, often low serum Mg does not result in tetany.

Magnesium is intimately associated with Ca and P through distribu-

tion and metabolism. Magnesium is the fourth most abundant cation in

the body. All tissues of the higher animals contain Mg, usually in con-

siderable quantities; Na, K and Ca are also major constituents. The oc-

currence of Mg can only be understood when these four major electrolytes

are considered together. Approximately 70% of the body Mg supply is in








the skeleton, the remainder being found widely distributed in the various

fluids and other soft tissues. One-third of the supply in the bones is

subject to mobilization for soft tissue when the intake is inadequate.

As indicated by Rook and Storry (1962), about 30% of skeletal Mg in young

animals can be mobilized under conditions of Mg deprivation while in

adult animals, only 2% of bone Mg can be used for physiological needs.

Cardiac muscle, skeletal muscle and neural tissue depend on a proper

balance between Ca and Mg ions.

According to Walker and Vallee (1964), Mg takes part in about eighty

enzymatic reactions. Magnesium is an active component of several enzyme

systems in which thiamine pyrophosphate (TPP) is a co-factor. Oxidative

phosphorylation is greatly reduced in the absence of Mg. It is also an

essential activator for the phosphate-transferring enzymes myokinase,

diphosphopyridine nucleotide kinase and creatine kinase. It also acti-

vates pyruvic acid carboxylase, pyruvic acid oxidase and the condensing

enzymes for the reactions in the Krebs cycle (Swenson, 1970). Magnesium

is vitally involved in the metabolism of carbohydrates and lipids as a

catalyst of a wide array of enzymes which require these elements for op-

timum activity (Walker, 1969). In the light of these functions, it is

not surprising that Mg deficiency in animals is manifested clinically in

a wide range of disorders, which include retarded growth, hyperirrita-

bility and tetany, peripheral vasodilation, anorexia, muscular incoor-

dination and convulsions (Underwood, 1981).

Although the adult cow contains about 250 g Mg, hardly any of it

can be released into circulation when intake is insufficient. In con-

trast, young cattle can mobilize Mg more efficiently from body stores

and develop clinical signs only after a longer interval. When Mg supply








and inevitable metabolic losses are in balance and supply is thus just

adequate, about 2.5 g Mg is excreted in urine daily. When the supply is

more than adequate, excess is excreted in urine and the concentration in

blood plasma remains in the normal range. When the amount of Mg absorbed

is inadequate, daily excretion in urine drops sharply, sometimes to less

than 0.1 g (CMN, 1973).

In ruminants, a physiological Mg deficiency frequently occurs in

adult animals when turned into fresh pasture during spring and autumn

in certain areas of the world. The pastures on which the animals de-

velop the grass tetany are not usually low in Mg; therefore, there must

be other factors related to Mg utilization in ruminants: 1) High K in

the herbage is perhaps the most frequent factor implicated in grass tet-

any. Experiments concerning the effects of K fertilization of pastures

on the incidence of hypomagnesemic tetany have yielded inconclusive re-

sults (Bartlett et al., 1954; Sims and Crookshank, 1956; Ritchie and

Hemingway, 1963). Fontenot et al. (1960) found that high K, high pro-

tein diets typical in this respect of lush, young grass depressed Mg re-

tention in sheep. In another experiment, Newton et al. (1972) found

that feeding a high K ration (4.9% K) to lambs resulted in a 46% decrease

in apparent Mg absorption. Metson et al. (1966), in New Zealand, report-

ed that K depresses the uptake of Mg, Na and Ca. 2) High N fertiliza-

tion of forages has been shown to increase crude protein content of the

forage and has adversely affected Mg utilization in cattle grazing the

herbage. Metson et al. (1966) studied hypomagnesemic tetany in New Zea-

land and found that high protein content of grass is a causal factor in

the development of this disease. Head and Rook (1955) postulated that

high ruminal ammonia N levels might result in the formation of a complex









between N and Mg and interfere with Mg absorption. Rosero (1975) stud-

ied the effects of species, stage of maturity and level of N fertiliza-

tion on the Mg availability for ruminants and found that Mg retention

was lower (P < .05) with early stages of maturity (high N content). Also,

fertilization lowered the Mg intake, % absorption (P < .10) and Mg bal-

ance (P < .05). 3) Excesses of citric acid on trans-aconitic acid ap-

parently depress blood Mg enough to cause tetany when cattle are under

stress. Scotto et al. (1971) indicated that administration by drenching

of KC1 and citrate or transaconitase would produce a tetany in a high

percentage of experimental cattle. 4) Low soluble carbohydrates, low

Ca, high P and low Na in the pastures are involved in the metabolism of

Mg (Matsen et al., 1976; House and Mayland, 1976). 5) Changes in

amounts of soluble Al may also be an important route through which cli-

matic and agronomic factors alter incidence of grass tetany (Allen et

al., 1980). Serum Mg levels in Al-treated steers (4,000 ppm Al) dropped

within 24 hours after treatment began and declined 32% by the end of 4

days (P < .01). After treatments were discontinued, serum Mg levels re-

turned to normal. Also, the authors found that the rumen contents from

tetanous animals averaged 2373 ppm Al, more than 5 times the level found

in normal fistulated animals.

Assessment of Mg status

Blood plasma. According to the CMN (1973), the approximate normal

level of serum Mg in cattle is 2.0 to 3.5 mg/100 ml. Below the level of

2.0 mg Mg, deficiency begins, with 1.0 mg considered an extreme deficien-

cy. However, Cunha et al. (1964) suggested 2.5 mg of Mg/100 ml of serum

as a normal level in cattle and reported that calves showing clinical

signs of tetany contained levels as low as .1 mg Mg/100 ml serum. On








the other hand, Underwood (1966) considered the normal level of serum Mg

in cattle to be 1.8 to 3.2 mg/100 ml.

Kiatoko (1976) reported blood plasma Mg levels of 1.8 .2 and 1.9

.3 mg/100 ml in cattle from two different regions of Costa Rica.

Claypool (1976) studied the factors affecting Ca, P and Mg status of

dairy cattle on the Oregon coast and found a large source of variation

in plasma Mg was due to herd difference (35%), with means of 2.2 to 2.6

mg/100 ml of serum.

O'Kelley and Fontenot (1969) reported that levels of .18, .19 and

.16% Mg in the rations of lactating beef cattle on a dry matter basis

in the first, second and third phases of lactation are necessary to main-

tain a level of 2 mg/100 ml serum Mg. Likewise, levels of .12, .10 and

.13% Mg on a dry matter basis for beef cows at 145, 200 and 255 days of

gestation, respectively, were needed to maintain that level of 2 mg/100

ml serum Mg (O'Kelley and Fontenot, 1973). Jerez (1982) studied the

mineral status of grazing beef cattle in the eastern region of the Do-

minican Republic, consisting of Romana Red, Criollo and Brahman breeds.

Mean serum Mg concentrations for regions 1, 2 and 3 were 2.1, 2.3 and

2.7 mg/100 ml, respectively. Of the 73 serum samples analyzed for Mg,

only 19% exhibited concentrations below the critical level of 2 mg/100

ml (McDowell and Conrad, 1977). Most forage concentrations (67%) were

adequate in Mg. Kemp (1960) reported a positive correlation between the

Mn content in forage and serum. Chicco et al. (1973) reported that high

dietary Ca depressed Mg content in bone and plasma and decreased Mg

utilization. Conversely, high Mg in the diet reduced plasma Ca. Diet-

ary P had little effect on Mg utilization. A positive correlation was

found between dietary Mg and either plasma or bone Mg.








Bone ash Mg. Harrington (1975) found a significant negative corre-

lation between bone ash concentration of Mg (rib, metacarpal) and the

length of time foals were fed a Mg-deficient diet. The normal Mg con-

centrations in rib bone of cattle range from .67 to .70% (Blaxter and

Sharman, 1955). Magnesium levels of .19 to .35% rib ash from cattle

with hypomagnesemia were reported. Loaiza (1968) reported levels of Mg

in metacarpal bone ash from grazing cattle from .56 to .61%.

Mendes (1977) reported that percent bone Mg calculated on dry, fat-

free bone basis in the rainy season from six ranches in the northern

part of Mato Grosso, Brazil, varied from .46 to .49%. Lebdosoekojo

(1977) reported Mg levels of .60 and .73% in rib ash during the rainy

and dry seasons, respectively, when bulls were kept on native pasture

with only salt as a supplement. For those receiving a complete mineral

supplement, Mg levels were .71 to .69% for the rainy and dry seasons,

respectively. Sousa (1978), in Brazil, reported rib bone Mg levels from

all six farms studied were slightly below normal levels but they were

higher than in animals with grass tetany. Rib bone ash Mg levels during

the wet and dry seasons were .46 and .49%, respectively. The lower lev-

el of rib bone ash Mg during the wet season is probably due to higher

requirements when animals are gaining weight rapidly, lactating and per-

forming other production functions.

Urinary excretion of Mg. According to the CMN (1973), daily urin-

ary excretion is a better criterion of Mg supply than plasma concentra-

tion. Even Mg concentration in a random sample of urine at any time of

day gives a good indication of supply. Tentative criteria are as fol-

lows: more than 100 mg/l, adequate to liberal; 20-100 mg/l, inadequate;

and less than 20 mg/l, severely deficient, danger of tetany. Within the








usual practical range, the percentage of Mg absorbed is not materially

affected by the amount consumed (Miller et al., 1974). Likewise, gen-

erally Mg content of tissues is not elevated with excess intake. Rather,

homeostasis is maintained by the excretion of excess Mg via urine; thus

urinary excretion of Mg is a threshold phenomenon. Substantially more

than a trace of Mg in urine indicates adequate absorbed Mg to meet the

animal's needs. On the other hand, very low urine Mg indicates either a

deficient or only barely adequate intake (Miller and Stake, 1974).

Magnesium conservation by the kidneys is believed to be mediated in

part by the parathyroid hormone (PTH). Losses of kidney function and

responsiveness to parathyroid hormone in cows consuming diets high in K

and organic acids could quickly lead to a mineral imbalance (Deetz et

al., 1981a. Magnesium excretion is believed to be mediated by glomer-

ular filtration and tubular resorption. Organic acids such as citric

acid may interfere with Mg and Ca resorption by chelating the ions, thus

forming complexes that would be poorly resorbed by the renal tubule

(Deetz et al., 1981b).

Sodium, Potassium and Chloride

Metabolism in ruminants

Sodium, K and Cl are of great importance in the metabolic processes

of all animals, with deficiencies causing rapid impairment of growth and/

or production. Sodium and Cl, along with K, function in maintaining os-

motic pressure, regulating acid-base equilibrium and controlling water

metabolism in body tissues. It is convenient to consider Na, K and Cl

together because of the broad similarities in their functions and re-

quirements in the animal body and their interactions with each other,

and because Na and Cl are associated in the form of common salt (Under-

wood, 1981).









Animals need to receive a regular supply of NaCl because there is

limited body storage capacity and any excess consumed is rapidly excret-

ed in the urine. When the animal is deprived of NaCl, it is able to

conserve the limited body reserves by largely eliminating urinary losses.

Even after prolonged severe deficiency, neither blood levels of NaCl nor

the amounts secreted in the milk decrease. Thus, lactating animals suf-

fer most from the lack of salt in the diet (Loosli, 1978). Sodium con-

stitutes about .2% of the body and occurs primarily in the extracellular

fluids, playing an important function in the regulation of neutrality in

the blood (Maynard and Loosli, 1971). It not only functions in the reg-

ulation of the balance of body fluids but also plays major roles in nerve

impulse transmission, the rhythmic maintenance of heart action and in

metabolism of carbohydrates and proteins (Fenner, 1980). On the other

hand, Cl is found both in intra- and extracellular fluids, making up

two-thirds of the acid ions in the body (Maynard and Loosli, 1971).

Thus, it also serves an important function in the maintenance of acid-

base balance in the body in addition to being a constituent of HC1 in

the gastric juice secreted into the stomach. The normal routes of Na

excretion are the urine, sweat and feces. Sodium losses from the bowel

are usually very low while those from sweat are higher in hot, tropical

climates (Pearson and Wolzak, 1982). Urinary excretion is the most im-

portant mechanism of Na elimination and is determined by the balance be-

tween Na filtered from blood in the glomerulus and that actively reab-

sorbed from the renal tubules (Pearson and Wolzak, 1982).

The total salt requirement of growing lambs approximates .40% of

dietary dry matter. The Na requirement is .04 to .10% of dietary dry

matter (NRC, 1975). The requirement for Cl is unknown. Range operators








commonly provide 220-340 g of salt per ewe per month. Some drylot tests

show that lambs consume about 9 g daily; mature sheep in drylot may con-

sume more.

The metabolism of NaCl is altered in hypertensive patients. One of

the major changes observed is an expansion of the extracellular fluid

volume and, in particular, an increase in the blood volume (Pearson and

Wolzak, 1982). The renin-argiotensin-aldosterone (RAA) system is known

to adjust distal tubular Na reabsorption in the kidney and hence excre-

tion to balance the Na needs of the body (Collings and Spangenberg, 1980,

cited by Pearson and Wolzak, 1982). Kolata (1981) found that high extra-

cellular Na ion concentrations have been noted in a variety of cell

types from humans and animals suffering from hypertension.

Potassium is the third most abundant element in the animal body,

surpassed only by Ca and P. In contrast to Na, which is the main elec-

trolyte in the plasma and extra-cellular fluids, K is present primarily

inside the cells. The blood cells, or erythrocytes, contain approxi-

mately 25 times as much K as is present in the plasma. Muscle and nerve

cells are also very high in K, containing over 20 times as much as that

present in the interstitial fluids (Thompson, 1978). Over two-thirds of

the body K is found in the muscle and skin. A dressed carcass will con-

tain about 75% of the body K.

Potassium requirements of beef cattle have been reported to be .6

to .8% of dietary dry matter (NRC, 1976). Requirements at this level

would be amply met with high forage diets which usually contain several

times the amount present in high grain diets. Gomide et al. (1969) stud-

ied the mineral composition of six tropical grasses and noted that the

average K content at 4 weeks was 1.42% vs .30% at 36 weeks of age.








These workers noted that a low forage K content may be a critical factor

in the poor performance of cattle, particularly during the dry season.

Loosli (1978) reported that Na deficiency is most likely to occur in

animals grazing pastures heavily fertilized with K which depresses Na

uptake by grasses. He added that since Na secretion into milk is high

even during Na deficiency, lactating animals suffer from lack of salt in

the diet.

Assessment of Na and K status

Because of its rapid reaction to deficiency long before clinical

signs appear, the best criterion is concentration of Na and K in saliva.

Sodium deficiency causes a fall in Na and a rise in K. Normal values in

saliva are 3.3 .3 Na and .3 .1 g/liter K. Less than 1 g/liter for

Na and more than 2.5 g/liter for K signal marked deficiencies, sometimes

even clinical signs (CMN, 1973). According to Kemp (1964, 1966), if less

than 3 g/liter Na is excreted per day, a dietary deficiency is implied.

Normal saliva values are given in Meq/liter as 145 for Na and 7 for K,

with a Na:K ratio of 20. The comparable values for the deficient animals

were 40 to 90, giving a Na:K ratio of .45. This adaptive change in the

Na:K ratio of parotid saliva is sufficiently sensitive to have been used

to estimate the Na requirements of lactating ewes (Morris and Peterson,

1975, cited by Underwood, 1981).

Due to large reserves of mobilizable body Na, plasma Na is not a

sensitive indicator of Na status. Fecal excretion of Na is variable

(Kemp, 1964) and not a reliable index of Na status. Milk Na is only

slightly decreased with low Na intake. Depressed plasma K is character-

istic in K deficiency. Reduced milk K and increased hematocrit readings

are observed during K depletion in lactating dairy cows (Pradhan and

Hemken, 1968).








Essential Trace Mineral Elements

Iron

Metabolism in ruminants

For many years, nutritional interest in Fe was focused on its role

in hemoglobin formation and oxygen transport. Iron is important in

electron transport mechanism in cells and as a component of several heme

enzyme systems (Church and Pond, 1975). Iron deficiency is one of the

most commonly occurring deficiency diseases of swine and humans. It is

rarely of practical concern in cattle, sheep and poultry except where

precipitated by chronic blood loss due to other conditions. Approxi-

mately 25% of total body Fe is stored as ferritin and hemosiderin in

the liver, spleen and other tissues (Thomas, 1970). According to

MIcDowell et al. (1978), Fe deficiency is unlikely to occur in ruminants

except in circumstances involving blood loss such as parasitic infesta-

tion or disease.

The great majority of Fe is contained in hemoglobin located in the

erythrocytes. Hemoglobin composes approximately one-third of the red

blood cell mass or about 1% of the body weight of man and domestic ani-

mals. There is also a considerable amount of Fe stored as ferritin and

hemosiderin in the liver and spleen. Hemoglobin contains about .34% Fe

but ferritin may contain up to 20% Fe and hemosiderin up to 35% (E. Miller,

1981). There is much less Fe in myoglobin, which is similar to hemoglo-

bin in composition and has much the same function in muscle as that of

hemoglobin in the erythrocyte. While transport Fe transferring ) and Fe

enzymes contain a very small percentage of the total body Fe, these

forms of Fe are vital. In all species, Fe deficiency results in a hypo-

chronic, microcytic anemia with low serum Fe, increased total serum Fe








binding capacity (TIBC) and a decreased transferring saturation (Underwood,

1971). In humans, 35-40% saturation is normal (Bothwell and Finch, 1962).

Less than 18% transferring saturation indicates impaired erythropoiesis.

Iron deficiency anemia develops in the young domestic animal during the

suckling period because of the low level of Fe contained in milk. It de-

velops more rapidly in suckling pigs than in the lamb or calf because of

the much greater growth rate of the pia during the suckling period (E. Miller,

1981).

Assessment of Fe status

Ferritin is the main storage compound of the body and its concentra-

tion in the tissues, together with that of hemosiderin, reflects the Fe

status of the animal. A high positive correlation between serum ferri-

tin concentrations and body Fe stores exists in man (Walters et al.,

1973, cited by Underwood, 1981) so that its estimation is a useful diag-

nostic tool. The principal carrier of Fe in the blood is another non-

heme protein compound, transferring or siderophilin, which is present in

the blood of all vertebrate species. Serum Fe and Fe binding capacity,

the basis for calculating % transferring saturation, can be determined

simultaneously and by a completely automated procedure (Friedman and

Cheek, 1971, cited by Miller and Stake, 1974). However, the limited

number of determinations reported for livestock and unknown range of

normal variations restrict the interpretation which can be made for

this biochemical means of diagnosing Fe deficiency. The body has a

limited ability to excrete Fe. Most of the Fe eliminated from the body

is via the feces, but this consists primarily of unabsorbed food Fe.

Losses from urine and sweat are minor. Even the loss of Fe in milk by

high-producing dairy cows is not great because of the low level of Fe

in milk, .5 ppm ,E. Miller, 1981).








Extensive research on the biological availability of various Fe

sources has been conducted in recent years (Ammerman and Miller, 1972).

The availability of Fe in different compounds varies enormously (Miller,

1979). Generally, Fe in soluble compounds such as ferrous sulfate and

ferric citrate is much more available to cattle than in ferric oxide or

Fe phytate (Ammerman et al., 1967; Bremner and Dalgarno, 1973). Levels

of dietary Fe exceeding 1000 ppm for growing cattle have generally re-

duced growth rate and plasma inorganic P levels (E. Miller, 1981).

Liver Fe. The liver is the center of mineral metabolism in the

animal body and is a useful organ for estimating the Fe, Cu, Zn, Mn and

Co status of animals (Boyazoglu et al., 1972). Hartley et al. (1959)

reported that the approximate normal level of Fe in cattle liver ranges

from 180 to 340 ppm on a dry matter basis. Cunha et al. (1964) indicat-

ed 200 to 300 ppm as the normal level of Fe in the cattle liver. Ammer-

man (1970) found liver levels of 100 to 300 ppm Fe in cattle in Florida

in an adequate state of Cu nutrition; the animals did not show deficien-

cy signs consistently until the Cu levels decreased to 25 ppm.

Among the organs and tissues of the body, the liver and spleem usu-

ally carry the highest Fe concentrations, followed by the kidney, heart,

skeletal muscle, pancreas and brain. Variation among species can be very

high in the liver, kidney and spleen. In some species, notably the rat,

rabbit, sheep and man, but not in the dog, the liver has a high storage

capacity of Fe (Underwood, 1966). Lebdosoekojo (1977) found that the Fe

levels in the liver of young bulls on native pasture with and without

complete mineral supplementation were 351 and 408 ppm dry matter, re-

spectively. Watson et al. (1973) found a decrease in liver Fe from 1055

to 678 ppm (P < .05) when dietary Mn was increased from 30 to 4030 ppm








in diets for wether lambs. In the liver, the decrease in Fe concentra-

tion from 178 to 99 ppm was associated with an increase in the concen-

tration of Cu from 622 to 2521 ppm. Rosa (1980) studied Cu, Zn and Fe

interrelationships in sheep and found high dietary Fe produced an ele-

vation (P < .01) in Fe accumulation in liver (212 to 788 ppm). High di-

etary Zn antagonized this effect (P < .05) by reducing Fe storage result-

ing from excess dietary Fe. Standish et al. (1971) reported a signifi-

cant Fe x P interaction effect in liver Fe values. The authors found

that greater intake of P did not significantly affect liver Fe when added

to the low Fe diet but with the high Fe diet, it reduced (P < .05) liver

Fe.

Boyazoglu (1973) and Cunha et al. (1964) observed an inverse rela-

tionship between Cu and Fe in the liver. Ott et al. (1966c) observed

increased liver Fe in lambs fed a diet containing high levels of Zn.

Heinrich (1971, cited by Miller and Stake, 1974) proposed that Fe depots

can be determined by measuring "diffuse storage Fe" in the cytoplasm of

the bone marrow reticuloendothelial cells. The procedure detects pre-

latent Fe deficiency but is unsuited for routine use in either man or

animal.

Coleman and Matrone (1969, cited by Davis, 1980) presented evidence

that in high Zn-fed rats, the amount of ferritin was about one-third

that found in rats fed a normal Zn diet. The excess dietary Zn resulted

in the formation of an "Fe-poor ferritin." Zinc toxicity did not appear

to interfere with the incorporation of amino acids into the ferritin.

There was some suggestion that the turnover rate of ferritin Fe and fer-

ritin protein in the Zn-fed rats may have been faster than in rats fed

the control diet (Davis, 1980).








Many consider percent saturation to be the most practical means of

detecting Fe deficiency in its early states (Underwood, 1971). The Fe

content of transferring reflects available circulating Fe at the hemoglo-

bin synthesis site.

Copper, Molybdenum and Sulfur

Metabolism in ruminants

There are advantages in considering Cu, Mo and S together because

of their nutritional interrelationships and their profound metabolic in-

teractions.

In many areas of the world, Cu deficiency is a major problem for

grazing cattle (Underwood, 1981; CMN, 1973). Except for milk-fed ani-

mals, most naturally occurring Cu deficiencies are conditioned by diet-

ary factors (high Mo, sulfide, sulfate or sulfur-containing amino acids)

that interfere with Cu utilization. Young and growing animals have a

higher Cu requirement and therefore a higher deficiency incidence than

mature ones. Young sheep are very susceptible to Cu deficiency (Under-

wood, 1966), with important genetic differences among breeds (Wiener and

Field, 1970). Copper deficiency in livestock results in a wide variety

of disorders. In addition to depressed growth, common signs include ane-

mia, osteoporosis, cardiovascular connective tissue defects and demyeli-

nation of the central nervous system.

In some areas, the deficiency of Cu results from vegetation low in

Cu, but in many locations, it may result from one of several factors in

the forage; e.g., the presence of high concentrations of Mo in the for-

age is one of the predominant factors interfering with Cu metabolism.

Forages grown on organic soils of Florida, for example, are high in Mo

and their consumption can result in molybdenosis or hypocuprosis in graz-

ing cattle (Becker et al., 1965). Studies of the nutritional physiology








of Cu were given a further impetus by Australian investigations of chron-

ic Cu poisoning in sheep. Copper retention was shown to be dependent

upon the Mo status of the diet, and the limiting effect of Mo was shown,

in turn, to depend upon the inorganic sulfate status of the diet and of

the animal (Dick, 1956, cited by Underwood, 1971). In relation to the

distribution of Cu in body tissues and fluids, the adult human body was

calculated to contain 110--120 mg of total Cu. Newborn and very young

animals are normally much richer in Cu per unit of body weight than

adults of the same species. The newborn levels are largely maintained

throughout the suckling period, followed by'a steady fall during growth

from weaning when adult levels are reached (Underwood, 1971).

The fact that cytochrome oxidase contains Cu immediately establish-

es an essential metabolic role for this microelement. Field conditions

actually exist in which sheep grazing pastures of the same or comparable

levels of herbage Cu suffer from either Cu deficiency or chronic Cu poi-

soning. There is a three-way interaction between Cu-Mo and inorganic

sulfate, with the primary site in the gut: reduction in the rumen of

sulfate to sulfide, reaction of this sulfide to form thiomolybdate, and

reaction of the thiomolybdate-CuMoSO4 (Dick et al., 1975, and Suttle, 1975,

cited by Underwood, 1979). Adverse effects of Mo on Cu utilization in

the tissues have also been demonstrated but the three-way interaction is

not yet fully understood and several studies with pigs have failed to

show significant reductions in tissue Cu levels from high dietary intakes

of Mo and S (Dale et al., 1973, and Kline et al., 1973, cited by Under-

wood, 1979). Underwood (1979) also adds that Mo can interfere with Cu

metabolism at well below 5 ppm Mo or more that have generally been used

under experimental conditions or that occur in molybdenosis areas. For

example, Suttle (1974) repleted a group of hypocupremic ewes with a







semi-purified diet containing 8 mg Cu/kg and one of four dietary Mo lev-

els, 0.5, 2.5, 4.5 and 8.5 mg/kg. Using rate of recovery in plasma Cu

as a measure of the efficiency of Cu utilization, the successive incre-

ments in dietary Mo decreased that efficiency by 40, 80 and 40%, re-

spectively. These results indicate that differences of 1 ppm in dietary

Mo are of significance with respect to Cu utilization by ruminants. Es-

sentially similar observations were made with guinea pigs. Where diet-

ary Cu intakes are high, more Mo is required to induce hypocupremia.

This has drawn attention to the importance of the dietary Cu:Mo ratio.

Miltimore and Mason (1971, cited by Underwood, 1979), on the basis of

Canadian experiments with cattle, reported that the critical Cu:Mo ratio

in animal feeds is 2:1 and that feeds or pastures with lower ratios than

2:1 would be expected to result in a conditioned Cu deficiency. Alloway

(1973) reported the results of a study of English pastures which also

reveal the importance of the Cu:Mo ratio to the incidence of hypocuprosis

in sheep and suggest that the critical ratio is even higher than 2:1,

perhaps nearer 4:1.

The first indication of an essential role for Mo in animals came

from the discovery that the flavoprotein enzyme, xanthine oxidase, con-

tains Mo and that its activity depends on the presence of this metal.

Also, aldehyde oxidase and sulfite oxidase are Mo-containing metalloen-

zymes (Underwood, 1981). A primary Mo deficiency in commercially fed

farm animals or in grazing livestock has never been reported. Such a

deficiency seems unlikely because of the low Mo requirements of animals,

despite the fact that large areas of Mo-deficient soils exist in which

yield responses to applications of Mo occur in crops and pastures (Under-

wood, 1981). Herbage in the Netherlands contains up to 5 mg Mo per kg;

at prevalent contents of Cu in herbage, such values can be taken to be








harmless for cattle. Higher values may be met on alkaline soils where

the pasture is rich in clover and on pasture polluted from industry (CMN,

1973).

Assessment of Cu and Mo status

The determination of Cu in the diet or pasture has limited diagnos-

tic value and can, in fact, be seriously misleading unless other elements

with which Cu interacts, particularly Mo and S, are determined also. The

criteria most widely used for Cu deficiency are the concentrations of Cu

in the liver and the blood (Underwood, 1981). Plasma Cu can indicate a

deficiency but does not reflect higher "marginal safety" liver storage

(Hartmans, 1973, cited by Miller and Stake, 1974). The Cu status of

plasma from cattle, sheep and swine can be readily ascertained from serum

ceruloplasmin activity. Ceruloplasmin, a true oxidase enzyme (ferroxi-

dase) synthesized in liver, contains up to 8 Cu atoms/mole, depending

upon species. In sheep, cattle and swine, a high percentage of the

plasma Cu exists as ceruloplasmin, with high correlations between serum

Cu and ceruloplasmin activity.

The normal range of blood plasma Cu for sheep and cattle is 0.6 to

1.5 pg/ml (Underwood, 1971). Values consistently below 0.6 pg/ml indi-

cate Cu deficiency in ruminants. Anemia is a common expression of Cu

deficiency in all species where the deficiency is severe or prolonged.

In these circumstances, the blood Cu in mammals falls as low as .1-.2

Pg/ml where normal hematopoiesis cannot be sustained. Copper does not

appear to be involved in the heme biosynthetic pathway but is essential

for the absorption of Fe from the intestinal mucosa, the mobilization of

Fe from the tissues and its utilization in hemoglobin synthesis. These

functions are accomplished by ceruloplasmin. This enzyme is necessary








for the formation of Fe (III) transferring, the transport vehicle of Cu

(Underwood, 1981).

Other methods such as histological examination of the tibia (Suttle

et al., 1972) or cardiovascular tissues can be sensitive indicators of

the Cu status of cattle, sheep and swine. However, in large animals,

they are not very useful in detecting early Cu deficiency stages as ani-

mals must be biopsied or sacrificed.

According to Cunha et al. (1964), normal Cu concentration of whole

blood in the healthy mature bovine is 75 to 100 pg/100 ml. The CMN

(1973) indicates that plasma Cu concentrations of 60 to 75 pg/100 ml are

considered slightly deficient while levels below 40 pg/100 ml are clear-

ly deficient. For blood serum, the critical Cu values are to be multi-

plied by .85 to .9. Claypool et al. (1975) found a positive relationship

between liver and plasma Cu concentrations. They suggested that liver Cu

on the order of 40 ppm was necessary to maintain plasma Cu of 91 pg/100

ml. Plasma Cu levels below 50 pg/100 ml were suggestive of low liver Cu

concentrations.

Liver Cu and Mo. The liver is the main storage organ of the body

for Cu so that liver Cu concentrations would be expected to provide a

useful index of the Cu status of the animal. Liver Cu values vary great-

ly with the species and age of the animal and in certain disease states,

and also with the nature of the diet. Among domestic livestock, liver

Cu values are consistently high in healthy sheep, cattle and ducks, with

a normal range of 100-400 pg/g on the dry basis, with a high proportion

of values lying between 200 and 300 pg/g. In sheep and cattle, liver Cu

concentrations vary only slightly from birth to maturity whereas in pigs,

they decline with age (Underwood, 1981). Liver Cu concentrations reflect








the dietary status but they are influenced by the dietary proportions of

Mo and S, by high intakes of Zn and CaCO3 and other dietary components

as well. Animals are able to store large reserves of Cu in the liver.

The Cu, upon reaching the liver, which is the principal organ involved

in the metabolism of this element, is incorporated in the mitochondria,

microsomes, nuclei and soluble fraction of the parenchymal cells in pro-

portions that vary with age, strain and Cu status of the animal. The

CMN (1973) reported that the approximate normal level of Cu in cattle

liver is 200 ppm on a dry matter basis. Levels below 50 ppm indicate a

deficiency and below 10 ppm, extreme deficiency. The Cu levels in the

livers of young bulls on native pasture, with and without complete min-

eral supplementation, were 342 and 232 ppm on a dry matter basis, respec-

tively, as reported by Lebdosoekojo (1977). Underwood (1979) reported

that a forage Cu level of 8 to 10 ppm (dry matter basis) can produce

chronic Cu toxicity in sheep and cattle when the concurrent Mo plus S

dietary levels are abnormally low (.2 ppm Mo or less). Generally, liver

Cu concentrations in the majority of animal species decrease as the ani-

mal matures; however, in cattle there is little variation between young

and mature animals, although sometimes young animals may have higher Cu

concentrations than adults (Church, 1971).

In sheep and cattle, liver Cu concentrations are influenced by var-

ious dietary factors and can be reduced by increasing Mo and S dietary

levels. Standish et al. (1971) reported that high dietary Fe depressed

absorption of Cu. Sheep fed a basal diet containing 77 ppm Cu supple-

mented with 0, 400 or 1600 ppm Fe in the form of ferrous sulfate showed

Fe concentrations in the liver of 185, 269 and 605 ppm, respectively,

while the corresponding liver Cu levels were 260, 145 and 44 ppm. Cattle








grazing on acid soils had liver Cu contents of 6 ppm (dry matter basis),

yet none showed signs of ill health, as reported by Bingley and Anderson

(1972). The same authors reported levels of 2 ppm Cu in pastures grazed

by calves with falling disease in Australia and found liver Cu levels of

1.6 to 6.7 ppm (dry matter basis) in dead animals. Rosa (1980) studied

the interrelationship between Cu, Zn and Fe using Florida native mature

wethers and found that there was a main effect of high dietary Cu on liv-

er Cu concentration represented by an increase (P < .01) of hepatic Cu

from 341 to 850 ppm. Liver Cu was also affected by an interaction (P <

.05) of Fe by Zn. Both high dietary Zn and Fe increased liver Cu concen-

tration in the presence of low levels of the other element.

The tolerance of farm animals to high dietary Mo intake varies with

the species, the amount and chemical form of the ingested Mo, the Cu

status of the animal and the diet, and the S content of the diet and its

content of substances such as protein, methionine and cystine capable of

oxidation to sulfate in the body. Cattle are by far the least tolerant

species, followed by sheep, while horses and pigs are the most tolerant

of domestic livestock. In normal diets, the level of Mo in the liver is

of the same order, namely 2-4 ppm, in several species of widely differing

dietary habits. Similar concentrations occur in the livers of newborn

lambs, indicating that this element is not normally stored in the fetal

liver during pregnancy. However, Mo concentrations 3 to 10 times the

normal level were observed in the livers of newborn lambs from ewes re-

ceiving a high Mo diet (Underwood, 1971). This author also suggests that

Mo readily passes the placental barrier in this species. Adult sheep and

cattle retain Mo concentrations in their livers of 25 to 30 ppm as long

as they are ingesting large or moderately large amounts of Mo. The








levels rapidly return to normal when the administration of the extra Mo

ceases.

Underwood (1971) reported that about one-half to three-fourths of

the total body Mo of sheep is situated in the skeleton, with the next

largest proportions in the skin, wool and muscles and only about 1% of

the total in the liver. This contrasts markedly with the distribution

of total body Cu in which a high proportion occurs in the liver in sheep

and very little in the skeleton. The author concluded that the Mo level

in the liver of an animal, therefore, gives little indication of its di-

etary Mo status and is of limited diagnostic value for this reason, un-

less the sulfate and protein status of the diet of the animal is also

known.

Hidiroglou et al. (1982) reported bone abnormalities in sheep and

cattle from Mo toxicity. Bones of the Mo-supplemented animals contained

more Mo than those from nonsupplemented animals. The author also found

that the Ca and P contents of sheep bones were unaffected by Mo supple-

mentation.

Cattle are much more subject to Mo toxicosis than are sheep. Signs

of molybdenosis include diarrhea and rapid loss of weight. The disease

usually occurs on pasture containing 5 to 20 ppm Mo on a dry matter basis

but when dietary Cu intake is abnormally low or dietary sulfate intake is

high, Mo intake as low as 1 to 2 ppm may be toxic (Pope, 1975). Ward

(1978) has reviewed studies that indicated that in the liver, Mo inhib-

ited the oxidation of sulfides to sulfate, resulting in the accumulation

of sulfides in the liver and their precipitation as Cu sulfides.

McDowell et al. (1982), studying cattle in Florida, found that mean

liver Mo contents were 2.8 and 3.0 ppm during the wet and dry seasons,







respectively. These values are in agreement with approximate normal lev-

els of 2 to 4 ppm indicated by Underwood (1977) and suggest that Mo was

not present in high enough concentrations to interfere greatly with Cu

metabolism. In a tropical region of South America (Bolivia), Peducasse

(1982), working with grazing Zebu-Criollo cattle, found that mean liver

Mo content was 4.3 ppm, with a range of 3 to 6.3 ppm. Lebdosoekojo (1977)

reported liver Mo levels in young bulls on native pastures with and with-

out mineral supplementation as 3.9 and 4.5 ppm, respectively.

Sousa (1978) evaluated the mineral status of beef cattle in northern

Mato Grosso, Brazil, and found that mean liver Mo contents were 2.5 .8

and 2.7 1.0 during the dry and wet seasons, respectively. There was a

trend to have more than the normal range of liver Mo during the wet sea-

son. This increase in liver Mo was due to a moderate increase in forage

Mo during the wet season (2.3 ppm vs 1.6 ppm). Levels of Mo up to 40 mg/

d tend to increase hepatic Cu levels while dietary Mo levels beyond 40

mg/d may alter the hepatic levels very little (Ammerman and Miller, 1975).

Toxicosis is the major concern in Mo nutrition. Suttle (1980) sug-

gested that values even less than 10 ppm dietary Mo in ruminants affect

Cu metabolism. Clinical signs of Mo toxicosis are similar to those of

/ Cu deficiency. Molybdenum-toxic areas characteristically occur on poor-

ly drained neutral or alkaline soils. According to Underwood (1977), Mo

toxicity occurs in cattle grazing pastures with 20 to 100 ppm Mo but not

in cattle grazing normal pasture with 3 to 5 ppm Mo or less.

Chronic Cu poisoning is the result of continual ingestion of Cu in

concentrations that exceed the maximum safe level of 80 ppm. During the

time of accumulation of Cu in the liver, clinical signs are absent but

later a hemolytic crisis may occur. This crisis is characterized by the








sudden onset of severe hemolysis and hemoglobinemia associated with se-

vere jaundice, liver and kidney damage and rapid death (Br akley et al.,

1982).

Zinc and Manganese

Metabolism in ruminants

Zinc had been established as essential for laboratory animals in the

1930's but even in 1956, relatively little was known about its nutrition

and metabolism in cattle (W. Miller, 1981). Since 1960, Zn nutrition and

metabolism in cattle has been the subject of considerable research. Be-

fore its cause was shown to be inadequate Zn, parakeratosis was a major

practical problem in swine production. According to the CMN (1973), Zn

deficiency has been established as a practical problem in mature cattle

only in tropical South America. Signs were poor growth, parchment-like

thickening of the skin called elephant skin or parakeratosis especially

on the muzzle and limbs but also around the base of the tail, on the

neck and flanks, and the skin extremely susceptible to wounds and infec-

tions. In the Netherlands, such signs have been observed sporadically

in calves.

Zinc is widely distributed throughout the body and plays an essen-

tial role in many body processes. It is present in many enzyme systems

involved with the metabolism of feed constituents (Cunha, 1981). Zinc

metalloenzymes include carbonic anhydrase, alcohol dehydrogenase, alka-

line phosphatase, carboxypeptidase, RNA and DNA polymerases, thymidine

kinase and others whose structure and functions have been critically re-

viewed by Riordan and Valle (1976, cited by Underwood, 1981). Zinc was

found to play a vital role in DNA synthesis and nucleic acid and protein

metabolism so that all systems of the body suffer in Zn deficiency,








particularly when the cells of particular systems are rapidly dividing,

growing or synthesizing. For these reasons, growth and reproduction es-

pecially are affected by lack of Zn (Underwood, 1981).

The estimated Zn requirement for dairy cattle is 40 ppm in the diet.

There may be certain conditions or an interrelationship with other nutri-

ents which might increase Zn needs. For example, a small percentage of

Dutch-Friesian calves are born with an apparently inherited defect that

causes a very severe Zn deficiency which can be temporarily corrected by

high amounts of Zn (Cunha, 1981). Animals are quite tolerant of exces-

sive dietary Zn. The first effects are lower feed consumption and re-

duced weight gains. Excessive Zn interferes with the metabolism of some

other trace elements, especially Cu and Fe. Except where massive amounts

of Zn are fed either intentionally or accidentally, it appears unlikely

that too much Zn should be a practical problem (W. Miller, 1981).

Studies on excess Zn levels indicate that lactating dairy cows fed

1279 ppm of Zn in the diet did not experience reduced performance. Grow-

ing cattle fed 900 ppm Zn exhibited decreased weight gains and feed effi-

ciency (Cunha, 1981). On the basis of these studies, the 1978 NRC publi-

cation on nutrient requirements of dairy cattle gives an estimated safe

level of 500 ppm Zn in young cattle and 1000 ppm in older cattle. The

Zn requirement for sheep is 35-50 ppm in the diet; the toxic level is

1000 ppm (NRC, 1975). The major homeostatic control route of Zn is a

variable percentage of absorption. Variable endogenous fecal excretion,

but not urinary excretion, also contributes to Zn homeostasis (W. Miller,

1981). Zinc levels of 1.7 g/kg of diet and higher caused reduced feed

consumption and depraved appetite characterized by excessive salt and

other mineral consumption and wood chewing in beef cattle (Ott et al.,








1966b) while Zn consumption above 1.5 g/kg of diet caused depressed feed

consumption in lambs (Ott et al., 1966a). The author also reported that

water consumption was also suppressed by force feeding 4.0 to 6.0 g Zn

daily. No other external symptoms of the toxicity were observed but pro-

longed consumption of high levels of Zn caused death.

Manganese was first recognized as an essential mineral element for

animals in 1931 when it was shown to be required by rats and mice for

growth and reproduction (Perry, 1981). The enzymes that are activated

by Mn are numerous and include kinases, hydrolases, transferases and de-

carboxylases. Activation was found usually to be shared with other bi-

valent cations, notably Mg (Valle and Coleman, 1964, cited by Underwood,

1981). A specific function for Mn in the synthesis of the mucopolysac-

charide has been demonstrated. It appears that Mn functions in the met-

abolism of carbohydrates and lipids are very important. The mitochon-

dria normally contain high levels of the element as do pyruvate carboxy-

lase and superoxide dismutase (Perry, 1981).

Manganese is poorly absorbed and excreted mainly in the feces, with

absorbed Mn also appearing in the feces, mostly via the bile and pancre-

atic juices. High dietary intakes of Ca and Fe reduce Mn absorption and

different mineral sources of the element vary greatly in their availabil-

ity. The animal body has only a limited capacity to store mobilizable

reserves of Mn. The bones, liver and kidneys normally carry higher con-

centrations of Mn than the blood or muscles and the former can be raised

or lowered by substantially increasing or decreasing the Mn intake of

the animal (Hidiroglou, 1979).

Although progress has been substantial, current information on nu-

trition and metabolism of Mn for dairy cattle is still incomplete (W. Miller,








1981). The author also reported that relatively large amounts of Mn are

present in most soils and in most feed ingredients. The percentage of

dietary Mn absorbed is low (typically around 3 to 4%) and variable, de-

pending on the dietary content. Manganese deficiency had not been pro-

duced in sheep or goats until 1968 when early weaned lambs receiving a

purified diet containing less than 1 ppm Mn over a 5-month period exhib-

ited bone changes similar to those seen in other Mn-deficient animals.

The exact requirements of sheep for Mn are not known (NRC, 1975).

General symptoms of Mn deficiency include impaired growth, skeletal

abnormalities, disturbed or depressed reproduction and abnormalities (in-

cluding ataxia) of the newborn (Underwood, 1971). In cattle, the Mn re-

quirement is substantially higher for reproduction and birth of normal

calves than for growth (NRC, 1978). Rojas et al. (1965) found in one

experiment that all calves born from cows fed 16-17 ppm dietary Mn for a

12-month period had neonatal deformities. Heifers and cows fed low Mn

diets are slower to exhibit estrus, are more likely to have "silent

heats" and have lower conception rates (NRC, 1978).

Assessment of Zn and Mn status

In feeding trials with Zn-deficient diets, signs appear rapidly,

sometimes within a week, in young cattle. Before they appear, concen-

tration of Zn in blood plasma falls. Young cattle seem to be directly

dependent on supplies in the ration and not to hold any significant mo-

bilizable reserve. Zinc concentration in plasma of healthy cows is .60

to 1.40 mg/liter. Immediately after calving, values may fall to about

.50 mg/liter. For clinical signs, values are usually less than .40.

Repeatedly low concentrations in plasma are a reasonable criterion for

determining Zn status of the animal but values can fluctuate rapidly and

are greatly affected by infection or poor food intake (CMN, 1973).








Zinc is present in the blood plasma, erythrocytes, leucocytes and

platelets. Almost all of the Zn in erythrocytes occurs as carbonic an-

hydrase. Subnormal carbonic anhydrase activity occurs in the blood of

Zn-deficient calves. A decline in plasma or serum Zn has been observed

in deficient animals of all species studied (Underwood, 1981). Mills et

al. (1967) reported a fall from normal values of .8-1.2 pg Zn/ml to be-

low .4 pg/ml in the blood serum of severely deficient lambs and calves.

Under experimental conditions, many biochemical changes have been

identified in severely Zn-deficient animals. Those with most promising

diagnositc value are plasma (or serum) Zn, hair Zn, bone Zn and alkaline

phosphatase content of plasma or other tissues (Miller and Stake, 1974).

These authors suggested that alkaline phosphatase is substantially af-

fected by numerous other factors, with large individual variability with-

in herds and large mean difference in similar herds. The Zn content in

male sex organs and secretions, which are also normally high, similarly

reflect the status of the animal. Values of 105 44 and 74 5 ppm Zn

(dry matter basis) were reported for the testes of normal and Zn-deficient

rams, respectively (Underwood and Somers, 1969, cited by Underwood, 1981).

Plasma Zn has been reported to decrease during parturition in the cow

(Prior, 1976; Dufty et al., 1977) and to decrease more in cows with dys-

tocia than in normal cows (Dufty et al., 1977). In contrast to plasma

Zn, which tends to reflect dietary changes, hair Zn is reduced when a

Zn-deficient diet is fed over a period of time (Miller et al., 1965b).

However, hair Zn is affected by many other factors; thus, under practical

conditions, its diagnostic value is severely limited.

According to the CMN (1973), there is as yet no practical criterion

for assessing the Mn status of animals. Liver biopsy seems. the most








promising. Analysis of hair is useless since results are difficult to

interpret. Decreased bone and blood alkaline phosphatase have been ob-

served in Mn deficiency (Underwood, 1966). However, as discussed with

Zn, alkaline phosphatase is affected by many conditions and thus is not

a good biochemical measurement of Mn deficiency (Miller and Stake, 1974).

Blood Mn values are extremely variable, reflecting both individual vari-

ability and analytical inadequacies. Whole blood concentration substan-

tially below .02 pg Mn/ml, nevertheless, suggest the possibility of a di-

etary deficiency in sheep and cattle, according to Hidiroglou (1979).

The Mn contents in wool and feathers apparently reflect the dietary stat-

us of the animals but their diagnostic value is doubtful, at least at

marginal intakes. However, the wool of lambs fed a low Mn diet for 22

weeks had an average of only 6.1 ppm Mn compared with 18.7 ppm in the

wool of control lambs (Lassiter and Morton, 1968, cited by Underwood,

1981). Manganese is distributed throughout the body and is found in

higher concentrations in bone, liver, kidney and pancreas (McDowell et

al., 1978).

The Mn concentration in the whole diets remains the most useful

means of detecting possible deficiency in animals. According to the NRC

(1976), most of the roughages contain more than 30 ppm Mn. Thomas (1970)

found interrelationship of Mn with other dietary factors such as organic

compounds, Ca, P, Mg and Fe. Protein concentrates of animal origin

such as meat meal and fish meal are poorer sources of Mn (5--15 ppm)

than the usual protein supplements of plant origin such as soybean

meal (30--50 ppm). Milk and milk products are even lower in this metal

due to the generally very low content of Mn in cow's milk (20--40 mg/

liter), according to Underwood (1981).








Liver Zn and Mn. Zinc content of tissues other than plasma de-

creases quite slowly, if at all, when a Zn-deficient diet is fed (Miller,

1969). Zinc content in a number of calf tissues, including liver, kid-

ney and pancreas, was increased several fold when 600 ppm supplemental

Zn was fed and before any symptoms of toxicity appeared (Miller et al.,

1970). Factors which reduce Zn absorption in the gastrointestinal tract

have also been indicated to reduce Zn concentration in the liver.

Standish et al. (1971) reported that Fe fed to cattle at high levels of

400 to 1600 ppm tended to decrease liver Zn concentrations; moreover, Cd,

Ca, Mg, P and Cu, as well as chelating agents such as EDTA, vitamin D and

phytic acid, influence Zn absorption and metabolism (Miller, 1972).

Underwood (1962) suggested that liver Zn values above 125 ppm should be

considered as the normal value for cattle. Neathery et al. (1973) re-

ported tissue Zn concentration as affected by dietary Zn of 16.6 and 39.5

ppm in the dry diet and found liver Zn levels of 109 and 119 ppm, respec-

tively. Watson et al. (1973) also found lowered Zn concentrations in the

liver of sheep fed high dietary Mn levels. The binding pattern of liver

Zn, like the total Zn concentration, was unaltered in Zn deficiency;

therefore, for detecting mineral status of animals, Zn in liver is not a

good indicator. Other tissues such as bones, pancreas, the male sex

glands, hair and blood plasma are better indicators of the mineral stat-

us in cases of Zn deficiency.

As far as Mn is concerned, liver can be used as a criterion to dif-

ferentiate between deficient and sufficient supply of the element. The

approximate normal level of Mn in cattle liver is 8 to 10 ppm (dry mat-

ter basis); below 8 ppm Mn indicates deficiency (Underwood, 1977).

McDowell et al. (1978) suggests that a Mn deficiency can best be detected








by the combination of liver (less than 6 ppm Mn) and dietary (less than

20 to 40 ppm) analyses. The bones, liver, kidney, pancrease and pitu-

itary gland normally carry higher Mn concentrations (1-3 ppm on fresh

basis) than do other organs. The skeletal muscles are among the lowest

in Mn (.1-.2 ppm) of the tissues of the body. The levels in the bones

can be raised or lowered by substantially varying the Mn intake of the

animal. The storage capacity of the liver for Mn is limited, compared

with the great capacity of this organ to accumulate Fe and Cu (Underwood,

1971).

Kiatoko (1979) found that hair Mn was negatively correlated with

liver Mn, indicating that hair is not a good indicator of Mn status.

Rojas et al. (1965) fed low Mn (15.8 to 16.9 ppm) diets to 6 to 8-year

old cows. All calves born to deficient dams exhibited clinical signs

of deficiency of Mn. Liver Mn concentrations in control and deficient

calves were 11.84 and 6.94 ppm on a dry matter basis, respectively.

Under such conditions, 25 ppm Mn in the diet was considered marginal and

liver Mn levels of 9 ppm by dry weight indicated a borderline deficiency.

Finally, Watson et al. (1973) reported an increased liver Mn concentra-

tion in sheep from 9.9 to 44.2 ppm as the dietary Mn increased from 30

to 4030 ppm.

Cobalt

Metabolism in ruminants

Cobalt was first shown to be an essential nutrient for sheep and

cattle as an outcome of Australian investigations of two naturally oc-

curing diseases, "coast disease" of sheep and "wasting disease" or en-

zootic marasmus of cattle (Underwood, 1981). Progress in understanding

the mode of action of Co in the animal organism was slow until 1948 when








two groups of workers independently discovered that the antipernicious

anemia factor, subsequently designated as vitamin B 2, is a Co compound

containing 4% of the metal. Cobalt is an essential component of vita-

min BI2 which is synthesized by rumen microorganisms. Cobalt should be

fed since injected Co is completely effective in alleviating Co defi-

ciency symptoms.

The minimum Co requirement of dairy cattle is about 0.10 ppm of the

dry ration (Ammerman, 1970; Underwood, 1971). Since the required level

is more than the amount contained in many forages and some concentrates,

supplemental Co is needed under many practical situations. The main

source of energy to ruminants is not glucose but acetic and propionic

acids and smaller amounts of butyric and other fatty acids produced by

fermentation in the rumen. Any breakdown in the utilization of these

acids involving vitamin BI2 would therefore seriously affect the Co-

deficient ruminant. A breakdown in propionate metabolism at the point

in the metabolic pathway where methylmalonyl-CoA is converted to succinyl-

CoA, a reaction catalyzed by methylmalonyl-CoA isomerase, a vitamin B -

requiring enzyme, has been shown to be a primary defect in the Co-

deficient sheep (Marston et al., 1961, cited by Underwood, 1981). A se-

vere Co deficiency causes appetite failure which is due, at least in

part, to the animal's inability to metabolize propionate. This is fol-

lowed by the onset of anemia and eventually, extreme emaciation. Vitamin

B12 has also been shown to exert a potent influence on the recycling of

methionine and via methionine, on folate metabolism. This occurs through

the activity of a second vitamin B 2-containing enzyme, 5-methyltetrahy-

drofolate homocysteine methyltransferase, which catalyzes the reformation

of methionine from homocysteine. Therefore, the activity of this








methyltransferase is depressed in the liver of vitamin B l2-deficient

sheep, with possible impaired nitrogen retention (Gawthorne and Smith,

1974, cited by Underwood, 1981).

Assessment of Co status

Young, growing sheep are the most sensitive of all animals to Co

deficiency; next are mature sheep, calves between 6 and 18 months of age

and mature cattle. Cobalt deficiency occurs in many regions of Latin

America and mostly, but not exclusively, is restricted to grazing rumin-

ants which have little or no access to concentrates (McDowell and Conrad,

1977). According to the CMN (1973), the Co concentration in tissues is

too low for easy estimation as a criterion of Co status. Because of this

and the lack of specific clinical signs, little is known about the effect

of nutrition and environmental conditions on Co status. The best method

of tracing deficiency is to give cattle a supplement of Co salts and to

see how they react.

Effective biochemical diagnostic measures for Co deficiency have

been developed (Underwood, 1966, 1971). Since this is an area problem,

knowledge of the soil (Co, pH, etc.) is very useful. The Dutch consider

soil Co of greatest diagnostic value. When Co is extracted with acetic

acid (2.5% v/v), more than .3 ppm indicates adequate Co whereas less

than .1 ppm is low. Under most conditions, mean pasture values of .10

ppm Co or more supply sufficient Co to prevent deficiency. However, if

the herbage consistently contains less than .08 ppm Co, a deficiency can

be predicted with a relatively high degree of confidence (Conrad, 1978).

In Queensland, Australia, Winter et al. (1977) reported that the

occurrence of Co deficiency in grazing animals was related to the pres-

ence of sandy soils low in Co content and to the seasonal variation of








Co concentration in forages; during the wet season, they found forages

with .126 ppm Co while in the dry season, levels decreased to .005 ppm.

Sousa (1978) reported that most of the native forages in the lowlands

of Mato Grosso, Brazil,had mean levels in the range of .04 to .14 ppm Co.

Research reviewed by Ammerman (1970) suggested that hair Co levels might

also have some value in predicting the animal's Co status. Serum methyl-

malonate and fleece growth rates are more reliable indicators of Co defi-

ciency in sheep than urinary methylmalonate (Judson et al., 1981). The

authors also reported that liveweight and wool growth responses to Co

pelleted therapy in weaner sheep of South Australia were useful.

Liver Co and vitamin Bl2. Cattle and sheep with normal stores of

vitamin B12 in their livers can consume a Co-deficient diet for months

without showing any signs of a vitamin B12 deficiency. Reduced ruminant

liver stores of Co and vitamin B12 are indicative of a dietary Co defi-

ciency and storage levels are frequently used to determine the Co status

of ruminants (Ammerman, 1981). Based on a study in New Zealand, Under-

wood (1971) has indicated that Co concentrations of .04 to .06 ppm or

less in the liver of cattle and sheep indicate Co deficiency. Cunha et

al. (1964) suggested a liver Co level of .04 ppm indicated extreme defi-

ciency. Most studies indicate that cattle and sheep liver Co levels be-

low .1 ppm on a dry basis are low to deficient while levels between .15

and .30 ppm are normal for healthy animals (Conrad, 1978). Liver vitamin

B12 concentration is a more sensitive and reliable criterion than liver

Co concentration.

Andrews (1960, cited by Miller and Stake, 1974) adopted the follow-

ing criteria for determining the Co status of sheep: values of vitamin

B12 in fresh liver below .07 ppm are considered to indicate severe Co








deficiency; between .07 and .10 ppm B 2, moderate Co deficiency; between

.11 and .19 ppm B12, mild deficiency; and values over .19 ppm B12 consid-

ered adequate.

Cobalt is known to interact with Fe, Cu, Se and Mo. An interrela-

tionship has been suggested between Co and Se. Cobalt-deficient sheep

are more susceptible to Se toxicity than sheep fed adequate Co (Ammerman,

1981). It is suggested that the Co-Se relationship may involve vitamin

BI2 in the metabolism of dimethyl selenide. The NRC (1976) has recom-

mended a level of .05 to .10 ppm Co in the diet. Mendes (1977) reported

mean liver Co values ranging from .040 to .691 ppm in the wet season and

.124 to .730 ppm in the dry season in 5 classes of cattle in northern

Mato Grosso, Brazil. Kiatoko (1979), in cattle of Florida, found that

liver Co concentration was 1.06 ppm in cows and .88 ppm in heifers where

forage Co levels varied from .09 to .12 ppm in the wet season and from

.12 to .26 ppm during the dry season. Peducass4 (1982), studying cattle

in Bolivia, found that mean liver Co concentrations were .57 and .30 ppm

in the two regions sampled; none of the liver samples contained levels

below the .05 ppm cited by McDowell and Conrad (1977) as borderline to

deficient for grazing cattle. Jerez (1982) studied the mineral status

of grazing cattle in three regions of the Dominican Republic and found

that liver Co concentrations for regions 1, 2 and 3 were .39, .42 and

.65 ppm, respectively. Mtimuni (1982), working in Malawi, reported that

Co was not deficient in the liver samples analyzed.

Selenium

Metabolism in ruminants

Selenium was first recognized as an essential trace mineral in 1957

when it was found to prevent liver necrosis in rats. Subsequently, Se








deficiency affecting a number of systems and producing a variety of le-

sions has been observed in swine, poultry, horses, sheep and cattle

(Hidiroglou, 1980). In cattle, white muscle disease is the most common-

ly recognized problem. For many years, biological interest in Se was

confined to its toxic effects on animals. Two naturally occurring dis-

eases of livestock, "blind staggers" and "alkali disease," observed in

parts of the Great Plains of North America, were identified as manifes-

tations of acute and chronic Se poisoning, respectively. These discov-

eries gave a stimulus to investigation of Se in soils, plants and animal

tissues with a view to determining minimum toxic intake and developing

practical means of prevention and control (Underwood, 1981).

Selenium is necessary for growth and fertility in animals and for

the prevention of a variety of disease conditions which show a variable

response to vitamin E. A metabolic interrelationship between Se, vita-

min E and S-containing amino acids exists at the cellular level. Sele-

nium functions in the cytosol through glutathione peroxidase (GSH-Px).

Glutathione peroxidase uses glutathione, a tripeptide with a S, to re-

duce hydrogen peroxide and organic hydroperoxides to less harmful prod-

ucts (Hidiroglou, 1980). Also, glutathione peroxidase, through its role

in the metabolism of hydroperoxides, may be involved in the synthesis of

various prostaglandin derivatives. Vitamin E acts as a lipid soluble

antioxidant in the cell membrane.

The metabolism of absorbed Se appears to be similar for ruminants

and nonruminants. A portion of dietary Se becomes incorporated into mi-

crobial material. The major excretory pathway for oral Se is fecal in

ruminants and urinary in nonruminants under most conditions (Martin and

Gerlach, 1972). In the younger preruminant lamb or calf, Se deficiency








exerts damaging effects more frequently than in the older animal. With

development of the rumen, affected animals may recover from NMD (Whanger,

1970, cited by Ammerman et al., 1978). The effects of vitamin E and Se

deficiency have been postulated to result from loss of membrane integ-

rity which leads to cell death. Addition of polyunsaturated fatty acids

to the diet tends to exacerbate these deficiency defects whereas synthet-

ic antioxidants, in many cases, will alleviate the signs of vitamin E

and Se deficiency. Selenium was also classified as an antioxidant due

to its ability to prevent a number of vitamin E deficiency diseases

(Sunde and Hoekstra, 1980). The authors also suggested that in a typic-

al animal cell, lipid-soluble cx-tocopherol scavenges free radicals and

possibly quenches singlet oxygen in the membranes. GSH-Px and superoxide

dismutase react with peroxides and superoxide, respectively, in the cy-

tosol and mitochondrial matrix space and catalase destroys H202 in the

peroxisomes. The relative concentration and the importance of these

protective species vary from tissue to tissue and from specie to specie

and result in the variety of Se and/or vitamin E deficiency signs ob-

served in different species.

Vitamin E and Se seem to have an additive effect on the reduction

of serum glutamic oxalacetic transaminase (SGOT) activity, increasing

survival time of the lambs and decreasing the ratio of urinary creatine

to creatine excretion in lambs less than 8 weeks old. Thus, the need

for vitamin E in the diets of nursing lambs is related to Se in the diet

and vice versa (Pope, 1975).

Failure in reproductive function and a high incidence of retained

placentas have been associated with Se-deficient rations. In some stud-

ies, supplementation with Se, Se-vitamin E or Se and increased protein,








significantly reduced the incidence of retained placenta (Hidiroglou,

1980). The dietary requirement of Se by most species is considered to

be about .1 ppm. In grazing animals, three distinct Se deficiency syn-

dromes have been described: "white muscle disease" (WMD) in newborn or

young lambs and calves; unthriftiness, with poor growth rates which may

occur in the absence of any other recognizable disease; and infertility

(McDowell, 1978).

Assessment of Se status

The most widely used assessment of Se status is blood Se concentra-

tion. Low blood Se is always found in Se-deficient conditions. A di-

rect relationship between blood GSH-Px activity and Se concentrations

has been established (Hidiroglou, 1980). The author also reported that

Se is incorporated into the erythrocyte GSH-Px at the time of erythro-

poiesis; thus GSH-Px levels are less affected by daily variations in the

dietary level. In two experiments reported by Kuchel and Buckley (1969,

cited by Underwood, 1971), the concentration of Se in the whole blood of

sheep grazing pastures of normal Se status ranged from .06-.20 (mean .10)

yg/ml in one study and from .04-.08 (mean .06) ig/ml in the second exper-

iment. The administration of Se pellets induced a rapid rise in blood

Se to levels as high as .15-.25 pg/ml, depending on the amount of Se in

the pellets. Segerson and Johnson (1980) studied the effect of Se and

reproductive function in yearling Angus bulls and found pooled Se at 21-

day intervals averaged .01 and .08 (P < .001) for control and Se-supple-

mented bulls, respectively. They concluded that injections of supple-

mental Se increased both serum and tissue concentrations of this element.

No overt clinical signs of Se deficiency were observed in control bulls

even though serum Se concentrations were as low as .01 ppm. In contrast,








the serum Se concentration (.08 ppm) for treated bulls was comparable to

serum levels considered adequate for various aspects of reproductive per-

formance in beef and dairy cows. In this study, Se in serum was corre-

lated (P < .05) with Se in kidney, liver, seminal vesicle and testis (P <

.10) tissues.

Since Se is deposited in all the tissues of the body, except the fat,

of animals consuming seleniferous feeds, high concentrations of the ele-

ment provide indisputable evidence of an excessive intake. The Se con-

tent in urine, blood and hair similarly reflect dietary intakes but are

highly variable. Hair from normal cows generally contains 1-4 ppm Se

compared with 10-30 ppm for cattle on seleniferous range (Underwood,

1981). The concentration of Se in milk varies greatly with the Se in-

take of the animal. Perry et al. (1977) observed milk Se concentrations

ranging from 7 to 33 ppb when cows were supplemented with linseed meal.

Conrad and Moxon (1979) found that 4.8% of supplemental Se was trans-

ferred to milk when animals were fed a Se-deficient diet but that only

.9% of added Se was transferred to the milk of cows consuming diets ade-

quate in Se. Ammerman et al. (1980) found that milk Se concentrations,

which ranged from 7 to 20 ppb, were higher (P < .01) for cows on the

linseed meal plus Se treatment than for those receiving no Se supplemen-

tation. Sousa and Moxon (1982) studied the serum Se levels in cattle

from Brazil and found serum Se deficiency (< .02 ppm) in grazing dairy

cows. One calf was observed with white muscle disease (WMD) with low

serum Se levels (.005 ppm). Mans et al. (1980) reported that plasma Se

adjusted slowly and latently when increased Se was fed and that Se ac-

commodation lasted for 7 weeks or more.








Liver and kidney Se. The kidney and the liver are the most sensi-

tive indicators of the Se status of the animal and the Se concentrations

in these organs can provide valuable diagnostic criteria. Andrews et al.

(1968, cited by Underwood, 1971) suggested that Se levels of less than

0.25 ppm are indicative of marked Se deficiency in sheep. Concentrations

greater than 1.0 ppm Se in the kidney cortex and .1 ppm in the liver are

considered normal and one-half the quantity of these levels indicates a

marginal degree of Se-responsive unthriftiness. McDowell et al. (1978)

indicated that normal Se levels in the liver of cattle are usually above

.25 to .50 ppm (dry matter basis) while values on the order of 5 to 15

ppm (dry matter basis) are suggestive of an excessive Se intake. Sele-

nium (ppm) in kidney and liver tissues of control and Se-treated bulls,

respectively, was .84 and 1.27 (P < .005) and .1 and .37 (P < .001)

(Segerson and Johnson, 198:J). Ammerman et al. (1980) found that liver

concentrations were higher (P < .05) for calves from cows fed linseed

meal calculated to provide a natural Se adequate level than they were

for calves from cows fed the natural, Se-deficient soybean meal supple-

ment.

Kiatoko (1979), in Florida, reported liver Se concentrations were

below critical levels (< .25 ppm) in 32.2 and 38.8% of the samples in

the wet and dry seasons, respectively. Like forage and soil Se, liver

and hair Se concentrations were deficient. Peducass6 (1982) reported a

mean of .70 ppm in cattle from tropical areas of Bolivia. McDowell et

al. (1982) studied trace mineral status of cattle in Florida and conclud-

ed that the most pronounced finding from the analysis of samples in all

regions was the low Se status of pastures, soils and animal tissues. No

effect of Se administered in mineral supplements existed as samples were








collected before the Food and Drug Administration approved dietary addi-

tions of this element. They suggested the need for increasing dietary

intake of Se because of persistent reports of white muscle disease in

Florida.

Much of the Se in tissues is highly labile and transfer of animals

from seleniferous to nonseleniferous diets is followed by rapid and then

slow loss of Se from the tissues via bile, urine and/or expired air.

Concentration in tissues tends to reflect dietary Se concentrations,

particularly when provided by natural dietary ingredients as compared to

selenate or selenite (NRC, 1980). Ku et al. (1980, cited by NRC, 1980)

found the Se concentration of swine skeletal muscle (.034-.521 ppm, wet

basis) was highly correlated (r = .95) with that in natural swine diets

(.027-.493 ppm, air dry) from 13 different U.S. locations. When these

workers added sufficient Se (.4 ppm) from sodium selenite to raise a low

Se (.04 ppm) swine diet to the level found in a South Dakota swine diet

(.44 ppm from natural sources), respective skeletal muscle Se concentra-

tions were .12 and .48 ppm, wet basis. Corresponding liver Se concentra-

tions were .61 and .84 ppm, wet basis. A similar pattern of tissue Se

concentrations has been found in cattle and sheep (Ullrey et al., 1977).

McDowell et al. (1977) reported that supplementation of a basal ration

with .10 ppm Se as sodium selenite resulted in an eightfold increase in

hepatic Se, nearly a fivefold increase in renal cortical Se and a 25-fold

increase in blood Se concentrations compared to pigs fed the unsupplement-

ed basal ration. Also, supplementation of the basal ration with 100 ppm

vitamin E resulted in increased renal cortical (P < .01) and blood (P <

.05) Se concentrations. The authors finally suggested that hepatic, re-

nal cortical and blood Se concentrations of .25, 2.5 and .1 ppm (dry *








basis), respectively, were determined to be the critical levels below

which clinical illness, death or lesions of Se-vitamin E deficiency

could be expected.

Although Se concentration in plasma and liver provide the best in-

dicator of current dietary Se intake in cattle, erythrocyte GSH-Px activ-

ity is suitable as an alternative test for the routine diagnosis of Se

deficiency. Also, there is a clear relationship (r = .97) between blood

Se concentration and erythrocyte GSH-Px activity in samples tested from

50 mixed age Friesian/Jersey cattle in New Zealand (Thompson et al.,

1981).

Energy and Protein

Insufficient energy probably limits performance of sheep more than

other nutritional deficiencies and may result from inadequate amounts of

feed or from feed of low quality (NRC, 1975). Energy, the principal di-

etary constituent, generally represents between 70 and 90% of the daily

dry matter intake. In young animals, an insufficient supply of energy

results in retarded growth and delay in the onset of puberty; in lactat-

ing dairy cattle, it results in a decline in milk yield and loss of body

weight. Severe and prolonged energy deficiency depresses reproductive

function. There is a close relationship between energy and minerals in

the metabolism of the animal body. For example, during glycolysis, the

anaerobic degradation of glucose to yield lactic acid, there are differ-

ent metabolic pathways, in which different enzyme systems prevail. Phos-

phorus, part of the high-energy ATP, is the first utilized during the two

priming steps of glycolysis. The phosphorylation of D-glucose at the 6

position by ATP to yield D-glucose 6-phosphate is catalyzed by two types

of enzymes, hexokinase and glucokinase, which differ in their sugar





66

specificity and affinity of D-glucose. Both hexokinase and glucokinase

require a divalent cation (Mg or Mn ) (Lehninger, 1975).

Energy requirements for breeding cattle are 1.9 Mcal metabo-

lizable energy (ME) or 2.3 Mcal digestible energy (DE) per kg dry

matter (NRC, 1976). For grazing livestock in tropical areas, forages

are the major source of the essential nutrients of energy, protein,

vitamins and minerals. Butterworth (1964) reported the values of

digestible energy of 21 tropical grasses to range between 2.23 and

3.20 Mcal per kg. The same author (Butterworth, 1967) also reported

that the availability of energy and protein as measured by their

apparent digestion by sheep and cattle declines rapidly with advanc-

ing maturity. Minson (1980) worked with tropical forages in

Australia and reported that voluntary intake of digestible organic matter

(DOM) is an acceptable expression of forage quality because it is close-

ly related to digestible energy (DE) intake and to animal performance.

Voluntary intake and nutrient digestibility must be considered separate-

ly because they are often not closely related across species. In a study

of 41 southern forages (Moore et al., 1980), the correlation (r) between

dry matter intake and digestibility was .69. As they advance beyond a

few weeks growth, most tropical forages have high lignin content which

influences digestibility and feed intake. Moore and Mott (1973) report-

ed that the crude protein percentages should be examined first when an

explanation of an unexplained low production is observed before looking

for other limitations such as those related to forage structure, other

nutrients and toxic effects. Also, the authors suggest that lignin must

be considered as the primary structural inhibitor of quality in tropical

grasses within a given species. Using percentage of crude protein (CP)

and in vivo digestible organic matter, Golding (1976) developed the








following equation to predict digestible energy (DE) concentration of

warm seasonal grasses:

DE = (4.15 DOM + 1.200 CP 4.59) x 1/100

Supplementation of low protein forages (less than 7% crude protein,

dry matter basis) with protein may increase voluntary intake (Ventura et

al., 1975). With respect to supplemental energy, the interaction is even

more complex. There are two extreme effects, substitutive or additive

(Moore and Mott, 1973). With high quality forage, increasing levels of

supplemental grain may result in a decreased intake of forage due to a

substitution of grain DE for forage DE. With low quality forage, howev-

er, increasing grain intake may have little effect on forage intake and

grain DE and forage DE are additive.

Crude protein is often the main limiting nutrient for livestock in

the tropics with approximately 7% as the minimum level required for pos-

itive nitrogen balance in mature grazing animals (Milford and Haydock,

1965). In ruminating cattle, the amino acids required may be obtained

from dietary protein and some non-protein compounds. Protein is required

for maintenance, growth, reproduction and lactation. Protein is espe-

cially important for the lactating cow because milk solids contain about

27% protein. Cattle store some protein in the blood, liver and muscle.

These reserves may be used over a short-term period of protein deficien-

cy, especially to maintain gestation and lactation (NRC, 1978). Irregu-

lar or delayed estrus is the major sign of protein shortage in diets for

breeding females. Little (1975), in Australia, studied the effect of

supplemental protein-P and P alone on pregnant cows grazing native pas-

tures during the late dry season. The results showed that protein plus








P supplementation reduced greatly the interval from calving to first

postpartum estrus. During the wet season in tropical areas, livestock

gain weight rapidly since energy and protein supplies are adequate and

thus the mineral requirements are high (McDowell, 1976). Van Niekerk

(1974) reported that the beneficial effect of P was primarily during

the wet season, although the P content in the grass was at its highest.

In the ruminant, the addition of fermentable carbohydrate to the

diet increases the digestibility of dietary Mg. Also, absorption of

Mg occurs mainly before reaching the duodenum. Volatile fatty acids

(VFA) are the principal end products of microbial digestion of carbo-

hydrate in the rumen and its absorption appears to have a striking in-

fluence on the transport of water and minerals, including Mg. Addition

of lactose resulted in a decrease in acetate and an increase in propic-

nate; rumen ammonia also decreased to a very low level (Rayssiguier and

Poncet, 1980). The authors suggested that extra fermentable carbohy-

drates might be beneficial both in increasing absorption of Mg, Ca and

P, in addition to energy.

Byers and Moxon (1980) conducted a study with Hereford steers to

assess the relationship between Se adequacy and protein requirements to

growing and finishing cattle. They found that Se levels are most criti-

cal during early stages of growth and when cattle are fed diets marginal

or deficient in protein. Zinc deficiency also is reported to be a prom-

inent feature associated with severe protein-energy malnutrition (PEM)

(Underwood, 1981). Osteoporosis has been demonstrated in experimental

animals fed a high protein diet. Also, it has been well established

that increased consumption of protein in humans and animals results in

increased urinary Ca excretion. Alkaline and acid phosphatase activity








in bone increased 2.5 and 2.3 times, respectively, reflecting increased

matrix turnover induced by the high protein availability (Weiss et al.,

1981).

Maternal-Fetal Relationships of Trace Elements in Ruminants

Many studies have demonstrated a variety of alterations in the fe-

tus and in newborns when excesses or deficiencies of several mineral el-

ements were offered to pregnant and lactating animals. Only in the past

two decades have techniques for studying transfer and metabolism in vi-

tro been defined (Hidiroglou and Knipfel, 1981). Trace elements enter

the body of the fetus from very early in gestation. In order to do this,

they must cross the placenta but how they achieve this is not entirely

clear. Many of them circulate in the serum combined with protein in the

fetal serum. The fetus is completely dependent upon the dam for its sup-

ply of minerals. Different types of placenta have shown varying degrees

of permeability to minerals, carbohydrates, fats and proteins. In spe-

cies in which placental barriers are strong, such as swine and ruminants,

the fetus is partly supplied by absorption with the fetal placenta of the

secretion from the uterine glands, i.e., the embryotropic route (Palludan

et al., 1969).

In cattle, effects of Cu deficiency usually are postnatal while in

sheep and goats, symptoms of Cu deficiency often occur in utero (Hidiro-

glou and Knipfel, 1981). High Cu content in most newborn animals has

suggested placental transfer and storage before birth (Prior, 1964).

However, little or no quantitive tissue studies on Cu placental trans-

fer have been reported for ruminant animals. The daily amounts of Cu

deposited in the total products of conception of the ewe during the

first, second and third trimester averaged 15, 85 and 186 mg/day,








respectively (Moss et al., 1974). These data indicate a concentration

barrier for Cu between the ewe and the fetus in the syndesmochorial type

placenta. Lesions in the brain and spinal chord characteristic of enzo-

otic ataxia could be detected as early as 99 days post-conception in fe-

tal lambs where enzootic ataxia occurred (Smith et al., 1977). Seaman

and Hartley (1981) studied congenital Cu deficiency in goats. They found

Cu levels in the serum of kids and their dams and in the livers of the

kids below normal. Williams and Bremner (1976) reported that Cu concen-

trations in liver increased towards the end of the gestation in ewes;

liver Cu begins to decrease soon after birth, presumably from mobiliza-

tion to meet the needs of other tissues of the growing animal. The preg-

nant ewe appears to be equipped poorly to protect her lamb against the

effect of a dietary deficiency of Cu (Hidiroglou and Knipfel, 1981).

The mammalian newborn does not consistently carry higher total body

Zn concentrations than mature animals of the same species (Underwood,

1977). There is a little fetal Zn storage; lactation represents a major

homeostatic demand for Zn (Stake et al., 1975). In mature cows, homeo-

static control mechanisms which regulate the Zn content of tissue are

much more effective than in calves. Liver Zn concentration, initially

very high, declined rapidly as pregnancy advanced (Williams and Bremner,

1976). An adequate Zn intake for gestation in the goat resulted in se-

vere deficiency during lactation. Zinc is retained during pregnancy

primarily in the placenta; transport varies with gestation, age and fe-

tal placenta exchanged Zn with blood plasma four times faster than mater-

nal placenta (NRC, 1979).

Studies in ruminants have indicated that Mn deficiency during ges-

tation has deleterious effects on the developing embryo. Inadequate









dietary Mn induces an abnormal development of the epiphysical fetal car-

tilage (Hidiroglou and Knipfel, 1981). The concentration of Mn in the

liver of newborn lambs appears to be useful for assessing the Mn status

of the dam (Hidiroglou, 1979). Data presented by Hansard (1972, cited

by Hidirouglou and Knipfel, 1981) suggest that Mn was transferred readi-

ly and comparatively rapidly from ewe to fetus. Following administration

of 54Mn to pregnant ewes, the concentration of radioactivity in the pla-

centa peaked at 12 h post-injection, with the placental concentration

representing more than 50% of the total 54Mn concentration in the fetal

compartment; after 168 h, more than half of the 54Mn had accumulated in

the fetus, with placental concentration decreasing to about 25% of the

total fetal compartment. These data suggest that Mn was transferred

rapidly from ewe to fetus. Neonatal calves born to dams on low Mn diets

exhibited reduced Mn in liver and kidney (Rojas et al., 1965).

Perry et al. (1978) studied the effect of supplemental Se (0, 1, 2

or 5 mg per cow daily starting 90 days prepartum and extending through

6 to 7 months of lactation) on Se levels in blood serum of cows and their

calves. The authors found that 5 mg/day levels increased calf serum Se

over that of calves whose dams were fed the 1 and 2 mg levels. Also,

calf liver and kidney Se levels were higher (P < .05) for calves from 5

mg Se-supplemented cows than for those from the control group. These

data indicated that dietary Se of the cows was able to cross the placen-

tal membrane.

















CHAPTER III
EFFECT OF ENERGY AND PROTEIN ON CALCIUM, PHOSPHORUS
AND MAGNESIUM RETENTION AND MINERAL STORAGE BY SHEEP

Introduction

A proper supply of all essential nutrients is required for maxi-

mum animal performance under any type of environment. Inadequate

supply of any given nutrient may adversely affect animal performance

in different ways. Dietary requirements for minerals are more

difficult to accurately define than those for the organic nutrients

(energy and protein) because many factors affect mineral utilization.

A major problem for ruminant animals dependent on tropical grass-

lands is the prolonged dry season which lasts four to six months.

During this period, grasses mature and become dry; consequently, the

nutritional value of native pastures is low in protein, energy, vita-

min A and specific minerals, particularly P. Animals gain weight during

the rainy season when there is a plentiful supply of high quality forage

which provides adequate protein and digesitble energy (Mtimuni, 1982).

As forage quality declines during the dry season, animals may lose as

much as 30% of their peak weight gained during the rainy season (Van

Niekerk, 1974).

Insufficient energy probably limits performance of sheep more

than other nutritional deficiencies and may result from inadequate

amounts of feed or from feed of low quality. Poorly digested low-

quality forages also lead to reduced feed intake. Forage may also be

so high in water that energy intake is limited (N.R.C., 1975).









Insufficient protein intake also results in reduced appetite,

lowered feed intake and lowered feed efficiency. Minson (1971) esti-

mated the critical level of crude protein in pasture to be between 6.0

and 8.5% while Van Niekerk (1974) states that the major limiting

nutrient from pasture is low protein content of grasses which usually

falls below 7%. Protein supplementation without concurrent energy

supplementation did not show response for beef cattle grazing natural

pastures during the dry season in Zambia, Africa (Walker, 1957).

Energy and protein are closely related with the metabolism of Ca,

P and Mg in the animal body. The principal objective of this experi-

ment was to investigate the effects of two levels of energy-protein

(low and high) and two levels of minerals (low and high) on Ca, P and

Mg retention by the animal, and the effects of diets on mineral

storage, blood parameters and mineral composition of selected animal

tissues.

Experimental Procedure

Twelve Florida native crossbred wether lambs averaging 50 kg

initial weight were randomly assigned to two treatment groups, where

two levels of energy and protein and two levels of minerals were

studied as they affected Ca, P and Mg retention.

In the first trial, the animals were fed a semi-purified diet

(table 1) high in minerals (2 to 30 times maintenance requirements)

with two levels of energy-protein (low = .8 x maintenance; high = 1.8

x maintenance). The animals were fed the two experimental diets for

three months, housed in two pens (6 animals per pen) where water

was available ad libitum, before being placed in metabolism cages for














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retention studies. This balance trial consisted of an 8-d adjustment

period and a subsequent 8-d total collection period (feces and urine).

In the second trial, dietary mineral concentrations were reduced

and the same animals were fed again for another three months with a

semi-purified diet (table 1), low in minerals with two levels of

energy-protein (high and low) and housed in the same pens as previously.

Following the 3-month period, animals were placed in individual meta-

bolism cages for a retention study. At the end of this second trial,

the lambs were sacrificed by exsanguination and liver, spleen, kidney,

heart, muscle (longissimus dorsi) and metacarpal bones were removed

and weighed and frozen for subsequent analysis.

Lambs were initially weighed, wormed (Levamisole) and allowed to

adjust to experimental diets in metabolism cages for 8 days. The two

balance trials involved an 8-d preliminary period followed by an 8-d

total fecal and urinary collection period in which a constant daily

feed intake of 1000 g per head was maintained. Water was supplied ad

libitum. The rations were fed in the morning. During the 8-d collec-

tion period, urine was collected every 24 hours. Feces were

collected each day into collection pans and aliquots of 10% of the

total weight taken. Aliquots per period (24 hours) per lamb were

mixed and stored at 0 C. Ration samples were collected each day in

paper bags and at the end of the experiment were ground into 1 mm

particle size in a Wiley mill. Feces were dried for 48 hours in a

force-draft oven at 600 C. The dry fecal material was weighed and

ground to pass a 1 mm sieve. Fecal and ration samples were analyzed

for Ca, P and Mg using methods described by Fick et al., 1979.









All lambs were weighed monthly and blood samples collected at

the end of each trial. Feed consumption, orts, fecal and urinary

excretions and animal performance was recorded daily. Blood serum

samples were deproteinized with 10% trichloracetic acid (TCA) and then

analyzed for mineral content according to methods described by Fick

et al. (1979). Calcium, Mg, Fe, Cu, Zn and Mn were analyzed by atomic

absorption spectrophotometry (Perkin-Elmer 306) according to procedures

recommended by the manufacturers (Anonymous, 1973). Phosphorus was

determined by the colorimetric technique described by Harris and

Popat (1954). Hemoglobin (Hb) was determined by the modification of

the colorimetric method of Martinek (1970) and hematocrit by the

microhematocrit method. Molybdenum concentrations were analyzed by

flameless atomic absorption spectrophotometry using a Perkin-Elmer

503 according to procedures recommended by the manufacturer (Anonymous,

1974).

Ration and tissue samples were processed and analyzed for mineral

content according to methods described by Fick et al. (1979). Liver

and bone biopsies were taken at the end of each trial for mineral

analysis. The sampling for liver biopsy was carried out as described

by Fick et al. (1979) and bone biopsy in sheep as described by Little

(1972).

Serum, wool and tissue Se were determined by a modification of

the fluorimetric method (Whetter and Ullrey, 1978). Wool and dietary

crude protein determination has been described by Technicon Industrial

Systems (1978).

Specific gravity was determined for bone samples according to

the procedure of Little (1972) as modified by Mtimuni (1982) and was









conducted as follows: Air-dried bone was weighed in air on an

analytical balance. A soft piece of wire about 15 cm long with a

noose at one end was weighed in air and later in water kept at 4 C

in a 200 ml beaker. The wire was suspended from the balance by a

hook and immersed into the water at the same level samples were to be

immersed in water. The samples were dried and ether extracted

following procedures outlined by Fick et al. (1979) and subsequently

analyzed for Ca, P and Mg.

The biological utilization of Ca, P and Mg was measured as

apparent absorption (total intake fecal) and net retention (total

intake (fecal + urinary)). For example:



(Total Mg intake) (Total fecal Mg) 00
Apparent Mg absorption = (Total Mg intake)x 100




Net Mg Retention = (Total Mg intake) (Total fecal Mg Total

urinary Mg)

Data were analyzed by the General Linear Model procedure of the

Statistical Analysis System (Barr et al., 1976) utilizing the facilities

of the Northeast Regional Data Center located on the campus of the

University of Florida, Gainesville. Means and standard deviations

were calculated; t-test was applied to find the significant difference

among diets.

Results and Discussion

Body Weight and Blood Parameters

Daily intake of trace minerals in wether lambs are shown in table

2. Effect of energy-protein treatments on body weight, serum minerals,

hemoglobin and hematocrit of lambs are presented in table 3. In









TABLE 2. DAILY INTAKE OF TRACE MINERALS

EXPERIMENT I, TRIALS 1 AND 2 *

TRIAL 1 (HM) TRIAL 2 (LM)

Item Diet 2b Diet 4d Diet ia Diet 3c
LEP HEP LEP HEP


Fe, ppm 137 139 72 103

Cu, ppm 12 13 6 7

Zn, ppm 235 281 81 65

Mn, ppm 80 84 26 19

Co, ppm 1.2 1.0 .4 .8

Se, ppm .19 1.2 .12 .51




* Daily intake of trace minerals based on feed intake and composition
of experimental diets.

a Low mineral + low energy-protein.

High mineral + low energy-protein.

c Low mineral + high energy-protein.

High mineral + high energy-protein.
















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trial 1, no differences (P > .05) were found between diets 2 and

4 (high in minerals, with low and high energy-protein, respectively)

in relation to monthly gain (kg). In trial 2, differences (P < .05)

were found in lambs fed diets 1 and 3 (low in minerals) in both the

second and third weighing (50.90 vs 64.10 kg and 50.50 vs 65.50 kg,

respectively). Serum P and Se were higher (P < .01) in diet 4 (HEP +

HM) versus diet 2 (LEP + HM): 8.5 vs 5.6 mg/100 ml P and 0.21 vs 0.16

pg/ml Se, respectively. No differences in serum Ca, Mg, Na, K, Fe,

Cu, Zn, Hb and hematocrit were found (P < .05) as a result of different

energy-protein concentrations between diets 2 and 4. There were also

no differences in serum minerals, Hb and hematocrit concentrations

for animals fed low minerals with different energy-protein concentra-

tions (diets 1 and 3). Paired comparisons between diets 1 and 2 (LEP

+ LM vs LEP + HM) and 3 and 4 (HEP + LM vs HEP + HM) were made for

hematocrit, Hb and serum minerals with no differences found (P > .05).

Wethers fed high energy-protein diets with either high or low

mineral concentrations had greater gains than diets low in energy-

protein. During this 7-month experiment, animals fed low energy-

protein diets with high and low minerals increased only 2.60 kg body

weight (47.8 kg initial weight vs 50.4 final weight), as compared with

animals fed high energy-protein diets with high and low minerals

which increased 22.3 kg body weight (43.10 kg initial weight vs 65.40

kg final weight). High energy-protein diets (1.8 x maintenance

requirements) increased wethers' average daily gain 106.2 g/animal,

while wethers fed low energy and protein diets (.8 x maintenance

requirements) averaged only 12.4 g/d/animal.









Rosa (1980) studied the P, Al and Fe interrelationships in sheep

and reported that the addition of 0.25% P to the basal diet improved

(P < .01) average daily gain from 105 to 148 g/animal; also, excess

of dietary Fe (800 ppm) decreased (P < .01) average daily gain from

165 to 97 g/animal.

The high energy-protein diet with high minerals (diet 4) increased

only serum P and Se. Rosa (1980) found that excess dietary Fe (800

ppm) increased (P < .01) inorganic serum P (6.3 to 7.6 mg/100 ml).

This author also found that blood Hb and hematocrit levels increased

when dietary Fe was increased (11.4 to 16.3 g/100 ml Hb and 33.9 to

46.6% hematocrit, respectively).

Levels of energy-protein and increased levels of dietary minerals

did not affect serum mineral concentrations; it seems through homeo-

static mechanisms that the animal mobilizes minerals from body re-

serves to maintain normal serum concentrations.

Liver Minerals

None of the six minerals analyzed in liver (table 4) in trial 1

were affected (P > .05) by dietary treatment. In trial 2, however,

the animals receiving low mineral diets with high energy-protein (diet

3) had lower (P < .05) concentrations of Fe, Cu and Co with no dif-

ferences (P > .05) found in Zn, Mn, Mo and Se concentrations. High

dietary energy-protein concentrations in the presence of low minerals

apparently depressed liver Fe, Cu and Co.

Paired comparisons between diets 1 and 2 (LEP + LM vs LEP + HM)

and 3 and 4 (HEP + LM and HEP + HM) were also made which would include

both time and treatment effects. No differences (P > .05) were found















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in Fe, Cu, Zn, Mn, Co and Mo liver concentrations between diets 3

and 4. However, liver Mn and Co concentrations were higher (P < .05)

for diet 1 vs 2, respectively, as follows: 9.97 vs 4.38 ppm Mn, and

0.59 vs 0.24 ppm Co. Wethers fed diet 2 (LEP + HM) also had higher

(P < .1) liver Zn concentrations than animals fed diet 1 (LEP + LM)

(241.0 vs 79.0 ppm, respectively).

Wethers fed high mineral diets had higher concentrations of

liver Mn, Co and Zn. Obviously reliable comparisons cannot be made

between two trials because of the time difference but trends can be

observed. Mean liver Fe concentrations in wethers fed the four experi-

mental diets were as follows: 497 ppm, Diet 4; 356 ppm, Diet 2;

339 ppm, Diet 1; and 151 ppm, Diet 3. McDowell et al. (1980) reported

the critical Fe levels to be 180 ppm in cattle. Based on this critical

level, wether mean liver Fe concentrations were adequate except in

animals fed diet 3 (HEP + LM) in which they were below the critical

level. Ammerman et al. (1967) found mean liver Fe concentrations of

169 ppm in calves fed Fe-deficient diets.

Liver Cu is reported to be the best criterion for assessing the

Cu status of cattle (CMN, 1973). McDowell et al. (1980) reported the

critical level for Cu to be between 25 and 75 ppm in cattle. In the

four experimental diets, wether liver Cu concentrations are higher

than these critical levels. Miller and Miller (1962) reported a

range of 84 to 132 ppm liver Zn concentrations for apparently healthy

cattle. Although not significant (P > .03), high mineral diets tended

to increase liver Zn concentrations in animals (241.0 and 198.7 vs

79.0 and 83.0 ppm for diets 2, 4, 1 and 3, respectively). Liver Mn

concentrations were above the critical concentration of 6 ppm




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