SWRHL-34r
CALCIUM IN HOCK JOINTS OF WILDLIFE RUMINANTS
IN SELECTED AREAS OF THE UNITED STATES
by
Raymond A. Brechbill,
Ronald E. Engel and
Robert C. Kramp
Bioenvironmental Research Program
Southwestern Radiological Health Laboratory
U. S. Public Health Service
Department of Health, Education, and Welfare
Las Vegas, Nevada
July 21, 1967
This study performed under a Memorandum of
Understanding (No. SF 54 373)
for the
U. S. ATOMIC ENERGY COMMISSION
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LEGAL NOTICE
This report was prepared as an account of Government sponsored
work. Neither the United States, nor the Atomic Energy Commission,
nor any person acting on behalf of the Commission:
A. Makes any warranty or representation, expressed or implied,
with respect to the accuracy, completeness, or usefulness of the in-
formation contained in this report, or that the use of any information,
apparatus, method, or process disclosed in this report may not in-
fringe privately owned rights; or
B. Assumes any liabilities with respect to the use of, or for damages
resulting from the use of any information, apparatus, method, or pro-
cess disclosed in this report.
As used in the above, "person acting on behalf of the Commission" in-
cludes any employee or contractor of the Commission, or employee
of such contractor, to the extent that such employee or contractor of
the Commission, or employee of such contractor prepares, dissemin-
ates, or provides access to, any information pursuant to his employ-
ment or contract with the Commission, or his employment with such
contractor.
0'78
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SWRHL-34r
CALCIUM IN HOCK JOINTS OF WILDLIFE RUMINANTS
IN SELECTED AREAS OF THE UNITED STATES
by
Raymond A. Brechbill,
Ronald E. Engel and
Robert C. Kramp
Bioenvironmental Research Program
Southwestern Radiological Health Laboratory
U. S. Public Health Service
Department of Health, Education, and Welfare
Las Vegas, Nevada
July 21, 1967
This study performed under a Memorandum of
Understanding (No. SF 54 373)
for the
U. S. ATOMIC ENERGY COMMISSION
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ACKNOWLEDGEMENTS
The authors are grateful to the following conservation agencies and
individuals for their generous cooperation in furnishing the wildlife
ruminant hock joints which were utilized for this study: U. S. Fish
and Wildlife Service, Fort Niobrara Wildlife Refuge; Kofa Game
Range; National Bison Range; National Elk Range; Wichita Wild-
life Refuge; San Andres National Wildlife Refuge; Desert Game
Range; U.S. Forest Service, San Bernardino National Forest;
Nebraska State Game Forestation and Parks Commission; Idaho
Fish and Game Department; Nevada Fish and Game Commission;
Oregon State Game Commission; Minnesota Department of Conser-
vation - Section of Game; Utah Fish and Game; Kentucky - Depart-
ment of Fish and Wildlife Resources; and Mr. Patrick J. Ray,
Westwood, Massachusetts.
This work was done as part of a U. S. Public Health Service radi-
ation surveillance study of wildlife ruminants in the Nevada Test
Site environs under a Memorandum of Understanding, SF 54 373,
between the Atomic Energy Commission and the U. S. Public
Health Service, Las Vegas, Nevada.
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ABSTRACT
The percent calcium of bone ash found in hock joints of 63 mule
deer (Odocoileus hemionus), 14 white-tailed deer (Odocoileus
virgianus), 14 elk (Cervus canadensis), 13 desert bighorn sheep
(Ovis canadensis nelsoni), 9 antelope (Antilocarpa americana),
and 7 buffalo (Bison bison) is reported. The average calcium con-
tent of hock joints in the 6 species from 18 different areas through-
out the United States was 37. 6% of bone ash. Differences between
species, regardless of geographic locations, were small when
present.
11
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TABLE OF CONTENTS
ACKNOWLEDGEMENTS i
ABSTRACT ii
TABLE OF CONTENTS iii
LIST OF TABLES iv
INTRODUCTION 1
MATERIALS AND METHODS 2
RESULTS AND DISCUSSION 4
SUMMARY 7
REFERENCES 8
111
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LIST OF TABLES
Table 1. Percent calcium of ashed bone reported in literature. 1
Table 2. Percent calcium in bone ash by species and geograph-
ical location. 4
Table 3. Percent calcium/bone ash in various species regard-
less of location. 5
Table 4. Percent calcium in bone ash found in hock joints of
mule deer by sex. 6
Table 5. Percent calcium in bone ash found in hock joints of
mule deer by age class. 6
IV
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INTRODUCTION
The assessment of 9°Sr levels in bones of mammals has been expressed
in various units, e.g., pCi 90Sr/gm calcium, pCi 9°Sr/gmbone ash,
pCi 90Sr/gm wet bone (fresh bone), pCi 9°Sr/gm dried bone. A great
many of the earlier reports list 9°Sr levels in bone as pCi of 90Sr/gm
of calcium (strontium units). However, when reported in pCi 9°Sr/gm
bone ash, calcium content is not normally determined. In comparing
data presented in strontium units with that in pCi of 9°Sr/gm bone ash
where the percent calcium is not given, one must use an assumed
constant percent (Ca/gm bone ash) to convert either unit to the other.
Values reported in the literature for wildlife ruminants indicate that
bone ash contains from 37.4% to 38. 8% calcium depending on what bone
of the skeleton is analyzed and from what species the bone is collected.
(Table 1).
Table 1. Percent calcium of ash bone reported in literature.
_ . Number of Bone % Ca in
Species ci c -DAT. Reference
Samples Specimen Bone Ash
Deer Not given Hock Joint 38. 7 Lindberg
Animals* Not given Not given 38.8_+.50** Holtzman '
White-tailed deer 18 Mandible 37-4 +. 16*** Shultz
White-tailed deer 18 Antler 36. 2 +. 28*** Shultz
*Species not given. **Explanation of plus or minus value not reported
. , , _ , , , , in original reference.
i-vvOne standard error of the mean.
The primary objective of this study was to determine the calcium con-
tent in hock joints of various wildlife species and to determine if these
values were species or geographically dependent or both. This infor-
mation is necessary to aid in determining whether our 9^5r values are
a reflection of true fallout, species differences, analytical error, or
calcium deficiency and to enable comparison of our findings with those
reported in the literature as strontium units.
1
Note: pCi = pico curie.
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MATERIALS AND METHODS
During the fall and winter hunting season of 1965, 120 wildlife ruminant
hock joints were received from various conservation agencies throughout
the United States as well as from interested individuals. The samples
were obtained from the Northeastern United States, Oklahoma, Montana,
Nebraska, Kentucky, Nevada, Utah, Minnesota, Oregon, Idaho, Kansas,
Arizona, and California.
The samples were taken from the right or left leg of the animal by
cutting the bone at a point four inches above and below the hock joint.
All skin and attached muscular tissues were removed prior to analysis.
Calcium analysis was performed by the United States Public Health
Service, Southwestern Radiological Health Laboratory, Las Vegas,
(4)
Nevada. The technique developed by Johns is included in detail
because it is a modification of the classic calcium oxalate technique .
The bone sample is ashed at 500°C in an electric furnace. A 1. 00 gram
portion of the ground bone ash is then dissolved in 6N HC1. Fifty ml
of saturated oxalic acid solution is added and the pH adjusted to 3. 0
with 6N ammonium hydroxide to precipitate calcium oxalate. The
solution is allowed to stand for 12-24 hours to permit the precipitate
to settle. The supernatant is removed by filtration and the calcium
oxalate precipitate washed several times with 0. 5% ammonium oxalate.
The precipitate and filter paper are placed in a muffle furnace and
heated to 500°C for conversion of calcium oxalate to the calcium oxide.
The ash is then dissolved in dilute nitric acid and diluted to 250ml with
deionized water. A 5ml subsample is diluted to approximately 50ml, to
which 5ml 6N potassium hydroxide is added, and the solution allowed
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to stand for at least 3 minutes. One-tenth gm (Cal-Red Indicator
Dilute*) is then added, and the solution titrated with 0. 1M
ethylenediaminetetraacetate disodium salt (EDTA Salt). The EDTA
Salt solution is standardized against a CaCO3 primary standard using
the same buffering and titration procedure.
Percent calcium in bone ash is calculated according to the following
formula:
ml EDTA Salt x M EDTA Salt x 40. 1 x 100
% calcium in bone ash =
Sample weight in grams x 1000
*Registered Trade Mark, U. S. Patent Office, Washington, D. C.
3
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RESULTS AND DISCUSSION
The average calcium content of six species from 18 different geo-
graphic areas are arranged in descending order (Table 2). The high-
est and lowest mean value, a difference of 2. 2%, consisted of only
one sample each. If these two are disregarded, the difference be-
tween the buffalo (Montana) and the mule deer(Southeast Oregon) is
0.9%. A statistical test of the difference between mean values
failed to show a significant difference among species from various
locations.
Table 2. Percent calcium in bone ash by species and geographical
location.
Species
Buffalo*
Buffalo
Elk
White-tailed deer
White-tailed deer
Elk
Antelope
Elk*
White-tailed deer
Mule deer
Mule deer
Mule deer
Mule deer
Elk*
Antelope
Desert bighorn sheep
Mule deer
Mule deer
Mule deer
Mule deer
Mule deer
Mule deer
Desert bighorn sheep*
Mule deer*
Location
Oklahoma
Montana
Montana
Nebraska
Kentucky
Oklahoma
Montana
Southern Nevada
New England states
Utah
Nebraska
Southern Nevada
Minnesota
Northeastern Nevada
Northern California
Arizona
Idaho
New Mexico
Southern California
Northern Nevada
Northwest Oregon
Southeast Oregon
Southern Nevada
Northeast California
X% Ca in No. of
Bone Ash Samples
38.8
38. 1
38. 1
38. 0
38. 0
38.0
38. 0
37.8
37. 7
37. 7
37.6
37. 5
37. 5
37. 5
37. 5
37.4
37.4
37.4
37.4
37.4
37.4
37.2
37.2
36.6
37. 5-38. 8
37. 7-38.4
37. 3-39. 0
37. 7-38. 2
36. 9-39. 3
37. 7-38.4
37. 3-38.2
37. 3-38.4
36. 9-38. 1
36. 7-38. 1
36.4-38.4
36.9-37.9
37. 3-37. 7
37. 1-38.4
36. 6-37.9
36.9-37. 7
36.9-38. 2
36.9-38.4
36. 7-38.0
35.8-37.9
36. 7-37. 7
36. 2-38.0
1
6
4
6
4
6
5
2
4
6
3
12
6
2
4
5
6
6
4
8
6
5
2
1
*Inadequate number of samples to be included in statistical analysis,
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The percent calcium in bone ash in various species regardless of location
are listed in Table 3. The buffalo, white-tailed deer and elk have the
highest values while the antelope, mule deer and desert bighorn sheep
have the lowest. It is interesting that, in general, the buffalo, white -
tailed deer and elk are animals from higher rainfall regions. Generally
it is assumed that areas of calcium deficiency occur in areas of high
rainfall where the calcium carbonates have been leached from the soil.
Our data show that the calcium content in the hock joints of animals
browsing in these areas do not have lower values than those animals
grazing in areas of low rainfall.
Table 3. Percent calcium/bone ash in various species regardless of
location.
Species
Desert bighorn sheep
Mule deer
Antelope
Elk
White-tailed deer
Buffalo
X% Ca in Bone Ash
37. 3
37.4
37,8
37.8
37. 9
38.2
s*
0. 536
0. 529
0.488
0.441
0.476
0.453
No. of
Samples
13
63
9
14
14
7
'"standard deviation
The average calcium content in the mule deer hock joints is identical
for both male and females (Table 4). Perhaps this is a reflection of
the calcium drain on the body of the male mule deer during antler
formation, which under certain conditions may be equal to the calcium
drain of the lactating female mule deer. Females show a greater
variance (Table 4) than do the males probably because both lactating
and non-lactating does were sampled.
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Table 4. Percent calcium in bone ash found in hock joints of
mule deer by sex.
Sex
Male
Female
X % Ca in Bone Ash
37.
37.
4
4
36
35
Range
. 7-38.4
. 8-38. 1
0.
0.
s*
504
579
No. of
Samples
27
31
Total 58**
* standard deviation
**total hock joints that were identified by sex.
The calcium content in bone ash of mule deer hock joints is arranged
by age class in Table 5. Examination of the mean values indicates
an apparent increase in calcium content from the 0. 5 year age class
(fawns) to the 3. 5 year age class (maturity), then a decrease in
animals above that age class. However, it was not possible to verify
the increase statistically due to the large variation of % Ca values
within each age group (as indicated by the ranges shown). The factors
contributing to this variation, in order of importance, are the group-
ing of animals into half-year age groups, the natural animal to animal
variation and the error of the method of chemical analysis.
Table 5. Percent calcium in bone ash found in hock joints of mule
deer by age class.
Age X%Ca
0. 5 years
1.5 years
2.5 years
3. 5 years
4. 5 years*
in Bone Ash
37. 0
37. 2
37.6
37. 7
37.4
Range
35.8-37. 9
36.4-38. 1
36. 7-38.2
36.9-38.4
36.6-38. 1
No. of Samples
7
12
7
7
22
Total 55**
* 4. 5 years or greater.
**total hock joints that were identified by age.
It is apparent that by increasing the number of samples of certain species
from several locations, little would be gained as no significant differences
were noted in calcium content among species, location or sexes where there
was an adequate sample size for statistical analysis. The data obtained
during this study will serve as background as well as an aid in com-
paring our data with that of others.
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SUMMARY
The average calcium content of hock joints in six species of wild-
life ruminants from 18 different areas was 37. 6% of bone ash.
Differences between species, regardless of geographic locations,
were small when present.
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REFERENCES
( Lindberg, R.G. and J.G. Olafson, 1958. Preliminary Report --The
Occurrence of 9°Sr in Nevada Deer Herds. University of California at
Los Angeles, U.S. AEC Report, unpublished.
Holtzman, R. B. , 1963. Desirability of Expressing Concentrations of
Mineral-seeking Constituents of Bone as a Function of Ash Weight.
Health Physics, 8(3): 315-319.
Schultz, V. , 1965. Comparison of Strontium -- 90 Levels Between
Antler and Mandible of White Tail Deer. J. Wild. MGMT 29(1) 33-38.
(4\
Johns, F. B. , et al, 1962. A Rapid Determination of Calcium Using
Ethylenediaminetetraacetate (Di-sodium salt) as a Titrant. U.S. Public
Health Service, Southwestern Radiological Health Laboratory, Las
Vegas, Nevada, unpublished.
'Scott, W.W., 1955. Standard Methods of Chemical Analysis. Volume I,
D. Van Nostrand Company, Inc. pp. 211-12.
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DISTRIBUTION
1 - 15 SWRHL, Las Vegas, Nevada
16 James E. Reeves, Manager, NVOO/AEC, Las Vegas, Nevada
17 Robert H. Thalgott, NVOO/AEC, Las Vegas, Nevada
18 Chief, NOB, DASA, NVOO/AEC, Las Vegas, Nevada
19 D. H. Edwards, Safety Evaluation Div. , NVOO/AEC, Las Vegas, Nev.
20 R. C. Emens, NTS Support Office, AEC, Mercury, Nevada
21 Martin B. Biles, DOS, USAEC, Washington, D. C.
22 JohnS. Kelly, DPNE, USAEC, Washington, D. C.
23 P. A. Allen, ARFRO, ESSA, NVOO/AEC, Las Vegas, Nevada
24 G. D. Ferber, ARL, ESSA, Washington, D. C.
25 Ernest C. Anderson, NCRH, PHS, Rockville, Maryland
26 James G. Terrill, Jr. , Director, NCRH, PHS, Rockville, Maryland
27 Donald J. Nelson, NCRH, PHS, Rockville, Maryland
28 Bernd Kahn, NCRH, RATSEC, Cincinnati, Ohio
29 Arve H. Dahl, NCRH, PHS, Rockville, Maryland
30 Charles L. Weaver, NCRH, PHS, Rockville, Maryland
31 R. T. Moore, NCRH, PHS, Rockville, Maryland
32 Northeastern Radiological Health Lab. , Winchester, Mass.
33 Southeastern Radiological Health Lab. , Montgomery, Alabama
34 D. W. Hendricks, Safety Evaluation Div. , NVOO/AEC, Las Vegas, Nev.
35 Howard L. McMartin, SAB, NCRH, USPHS, Rockville, Maryland
36 Arthur Wolff, Research Branch, NCRH, USPHS, Rockville, Maryland
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37 Mail & Records, NVOO/AEC, Las Vegas, Nevada
38 Paul T. Tueller, University of Nevada, Reno, Nevada
39 Charles Hanson, U S Fish and Wildlife Service, Las Vegas, Nev.
40 V. R. Bohman, University of Nevada, Reno, Nevada
41 Bruce Browning, California Fish & Game Dept. , Food Habit
Laboratory, Sacramento, California
42 George Welsh, Arizona Fish and Game Dept. , Kingman, Arizona
43 Al Jonez, U S Bureau of Reclamation, Boulder City, Nevada
44 Fred Isbell, U S Forest Service, Las Vegas, Nevada
45 District Supervisor, Nevada Fish & Game Commission, Las Vegas, Nev.
46 Director, Nevada Fish & Game Commission, Reno, Nevada
47 Director, National Park Service, Boulder City; Nevada
48 District Manager, U S Bureau of Land Management, Las Vegas, Nev.
49 District Manager, U S Fish and Wildlife Service, Las Vegas, Nev.
50 University of Nevada Library, Reno and Las Vegas, Nevada
51 & 52 USAEC Div. of Tech. Inf. Extension, Oak Ridge, Tenn. (2)
53 R. S. Davidson, Battelle Memorial Institute, Columbus, Ohio
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