TABLE 3. TRITIUM CONCENTRATION IN BLOOD SERUM OF DAIRY COWS FOLLOWING
A SINGLE ORAL ADMINISTRATION OF 100 mCi OF TRITIATED WATER

Time H-21 H-90 H-171 J-175
h yci/1 uci/1 pei/1 uci/1
1 124 250 136 112
2 232 318 326 374
3 259 303 410 406
4 330 426 414
5 250 298 412 417
6 238 295 430 411
7 242 305 428 440
8 248 294 427 441
9 335 291 437 410
10 229 279 404 423

TABLE 4. TRITIUM CONCENTRATION IN BLOOD SERUM OF DAIRY COWS FOLLOWING
A SINGLE ORAL ADMINISTRATION OF 200 mCi OF TRITIATED WATER

Time H-116 H-132 H-135 J-175
d uci/1 uci/1 uci/1 uci/1
1/24 173 240 150 272
2/24 313 320 197 425
3/24 308 355 306 505
4/24 354 403 334 437
5/24 378 391 340 507
6/24 329 418 394 495
7/24 369 406 359 488
8/24 356 397 359 491
9/24 372 384 362 457

10/24 247 365 340 449

11/24 331 377 362 507

12/24 343 401 375 492
2 38.2 58.3 87.6

16 2.98 16.2 9.93 8.70

24 0.629 2.87 2.74 1.52

10


lactating and nonlactating animals, three Hereford steers were each adminis-
tered 65 mCi of tritiated water in the same manner as the dairy cows. Blood
was collected from all three groups of cows and the serum portion analyzed by
direct liquid scintillation counting.

The data were analyzed by performing a nonlinear least squares regression
on a sequence of successively more complex mathematical models in the class of
sums of exponential terms functions. The best of the regression functions was
chosen using the general linear hypothesis test and half-lives were calculated
from the exponential coefficients of the best-fit regression. .

RESULTS

The individual values for whole milk are shown in Table 1 for the four
cows receiving 100 mCi of tritiated water. The half-lives for tritium transfer
to whole milk are presented graphically in Figure 1. This figure represents
the mean of the whole milk concentration values listed in Table 1. Tritium in
the whole milk of cows appears to decrease with a 3.04 + 0.09-day half-life
during the 20 days following oral administration of 100 mCi of tritiated water.
The transfer of tritium to milk then changed to exhibit a slope with a half-
time of 11.1 + 2.58 days followed by a much longer half-time of >120 days.

In order to determine the biological half-time of tritium in the different
milk components, a second experiment was conducted in which each of the dairy
cows received 200 mCi of tritiated water. The results of this experiment are
shown in Table 2 and Figure 2. The initial half-times shown by the milk com-
ponents indicate little difference between the milk serum and the butterfat
with half-times of 2.93 + 0.14 days and 3.28 + 0.35 days, respectively. The
protein fraction of the milk exhibited a considerably shorter half-time of
2.08 + 0.56 days. The longer half-time portion of the curves showed the
normal variance of results with time and lower activity. The shortest half-
time was indicated by the tritium activity in the serum portion of the milk.
This was found to be 43.7 + 4.28 days. The results of the protein analysis
showed that a relatively constant incorporation of tritium into protein oc-
curred during the 2nd and 3rd months after administration, followed by a half-
time of >50 days for the remainder of the study. The incorporation of tritium
into butterfat decreased with a longer half-time of 60.7 + 43.0 days and then
increased during the latter part of the study. .

The blood levels of tritium were similar for both the lactating dairy cows
and the beef animals during the first 12 hours after administration as shown in
Tables 3 and 4 and Figure 3. The tritium levels in blood serum from beef
cattle shown in Table 5 were averaged and the half-lives determined to be
4.05 + 0.21 days and 40.36 + 9.82 days as shown in Figure 4.


DISCUSSION

Results of this study show that the turnover rate of tritium in the body-
water pool of lactating dairy cows is not appreciably different from nonlac-
tating beef animals at 3 days and 4 days, respectively. The tritium turnover
rate 60 days after ingestion showed even less difference, 44 days for lactating
dairy cows and 40 days for beef animals. This result is similar to that noted
by Black et al., (1964) who found the average half-life for body water in
lactating cows to be 3.54 + 0.10 days and that in nonlactating cattle to be
3.40 + 0.18 days over the course of a 15-day study following intravenously
administered tritiated water to dairy cows and steers. The greater differences
in T, noted by Aschbacher et al.,(1965) 3 days for nonlactating and 5 days for
lactating cows may have been due to the use of fat, nonlactating, nonpregnant
cows, whereas the present study and Black's study utilized steers. There may
be differences in the rate of tritium turnover between the two experimental
groups due to the differences in formation and turnover of muscle by steers
versus adipose tissue by nonlactating cows.

The difference in the rate of incorporation of tritium into proteins as
compared to its incorporation into non-exchangeable portions to form stable
carbon~tritium lipid bonds is indicated by the shorter T_ of 2.08 and about 50
days for protein in milk as compared to T, of 3.28 and 6? days for butterfat.
The concentration of tritium in fat was noted by Schirch and Mason (1963) while
investigating concentrations in humans chronically exposed to tritium.

Moyer and DuVigneaud (1972) found the appearance of tritium in protein was
maximal after 1 day following intravenous administration to rats, but fell to
about 20 percent of the peak level by day 5. Tritium incorporated into serum
fatty acids became maximal during day 1 and remained relatively constant until
the 12th day, whereas, the appearance of tritium in serum cholesterol increased
over a period of 12 days. It would appear that the complexity of the chemical
transformations of tritium (Smith and Taylor, 1969) does not allow a simple
conclusion to be drawn concerning the fate of ingested tritium.

The relatively long half-time of >120 days for the transfer of tritium to
whole milk indicates that milk may be the major contributor to the radiation
dose of humans consuming tritium-contaminated dairy products. However, al-
though milk solids constitute only 1/10 of the total volume of milk, the
radiation dose resulting from consumption of butterfat and cottage cheese may
exceed that from whole milk due to the longer biological half-lives and in-
corporation of tritium in fat, protein, and carbohydrates. It will be neces-
sary to establish the turnover of tritiated milk solids in humans before it
will be possible to accurately estimate the radiation dose resulting from the
consumption of tritium-contaminated dairy products.

14


FOREWORD

Protection of the environment requires effective regulatory actions which
are based on sound technical and scientific information. This information must
include the quantitative description and linking of pollutant sources, trans~
port mechanisms, interactions, and resulting effects on man and his environ-
ment. Because of the complexities involved, assessment of specified pollutants
in the environment requires a total systems approach which transcends the media
of air, water, and land. The Environmental Monitoring and Support Laboratory-
Las Vegas contributes to the formation and enhancement of a sound integrated
monitoring ‘data base through multidisciplinary, multimedia programs designed
to:

. develop and optimize sustems and strategies for moni-
toring pollutants and their impact on the environment

. demonstrate new monitoring systems and technologies by
applying them to fulfill special monitoring needs of
the Agency's operating programs

This report presents the results of an investigation designed to evaluate
the short- and long-term behavior of tritium in beef animals and dairy cows
with emphasis on the resultant hazard to humans consuming meat and dairy
products. It is hoped that this information will be of use to those individ-
uals who must assess hazards from accidental release of tritium, designers of
nuclear waste processing facilities and those responsible for nuclear reactor
site selection. Additional information regarding this study may be obtained by
contacting the Exposure Dose Assessment Branch of the Monitoring Systems

Research and Development Division.

Geor§e BW. Morgan
Director
Environmental Monitoring and Support Laboratory
Las Vegas

iii


(100mCi)

(200mCi)

uCi/ liter of BLOOD SERUM

HOURS

Figure 3. Average concentration of tritium in blood serum following a single
oral administration of tritiated water to dairy cows and beef
animals.

1l


TECHNICAL REPORT DATA
(Please read Instructions on the reverse before completing)

1. REPORT NO. 2. 3. RECIPIENT'S ACCESSION NO,
EPA-600/3-77-076
4, TITLE AND SUBTITLE iS. REPORT DATE
June 1977

TRITIUM RETENTION BY COWS AND STEERS
AND TRANSFER TO MILK

6. PERFORMING ORGANIZATION CODE

7. AUTHOR(S) 8. PERFORMING ORGANIZATION REPORT NO.
A. L. Mullen, A. A. Moghissi, J. C. Wawerna,
B. A. Mitchell, E. W. Bretthauer and R. E. Stanley 7

9. PERFORMING ORGANIZATION NAME AND ADDRESS 10. PROGRAM ELEMENT NO.
Environmental Monitoring and Support Laboratory 1PA628
Office of Research and Development Ti CONTRACTIGHANT NO-

U.S. Environmental Protection Agency
Las Vegas, NV 89114

12. SPONSORING AGENCY NAME AND ADDRESS 13, TYPE OF REPORT AND PERIOD COVERED
U.S. Environmental Protection Agency-Las Vegas, NV FINAL FY70~-71
Office of Research and Development 14. SPONSORING AGENCY CODE
Environmental Monitoring and Support Laboratory EPA/600/07

Las Vegas, NV_ 89114
15. SUPPLEMENTARY NOTES.

16. ABSTRACT

Eight lactating dairy cows and three steers received a single oral ad-
ministration of tritiated water. Milk and blood from the lactating cows and
blood from the steers were periodically collected and analyzed for tritium
content.

The tritium content of whole milk decreased with time giving a curve
expressed as a three-component exponential which yielded half-times of 3.04 +
0.09, 11.1 + 2.58, and >120 days. Tritium in the blood serum of steers de-
creased with half-lives of 4.05 + 0.21 and 40.4 + 9.82 days.

Additional study of the milk to assess tritium incorporation in the
various fractions showed half-times of 2.93 + 0.14 and 43.7 + 4.28 days for
milk serum; 2.08 + 0.56 and >50 days for milk protein; and 3.28 + 0.35 and
60.7 + 43 days for butterfat.

ICENTIFIERS/OPEN ENDEC TE

Beef Dairy Products

Tritium retention
Biochemistry Isotopes j Tritium transfer
Cattle Radiation chemistry «Nevada Test Site
Dairy Cows Radiobiology :
Tritium i
i 18B ’
es See a | — i
19. SECURITY CLASS (This Report) 27, NO. OF PAGES
RELEASE TO PUBLIC UNCLASSIFIED 28
20. SECURITY CLASS (This page) 22, PRICE
UNCLASSIFIED

EPA Form 2220-1 (9-73)


€T

ras 10 -
im
iv
s
=
5
z
ir]
7)
a
re)
°
g
a
. °
2 10 F
=
u
=e

Figure 4.

Tp74.0520.21d

1740.36 +982 d

i 1 _ = a n 1
2 4 6 8 10 «(12 20 40 60 80 100 120
HOURS DAYS

Average concentration of tritium in blood serum

following a single oral administration of
65 mCi of tritiated water to three beef cattle.


TABLE 2.

TRITIUM CONCENTRATION IN yCi/1 OF WATER OR ITS EQUIVALENT IN MILK COMPONENTS FOLLOWING A
SINGLE ORAL ADMINISTRATION OF 200 mCi OF TRITIATED WATER (Continued)

Time Serum Protein Fat

d He-16 | H-132 | H-135 J J**-175 H-16 | H-132 J H-135 {| J-175 H-16 |_H-132 J H-135 J J-175
128 0.002 0.008 0.041 0.011 0.022

135 0.006 0.004 0.057 0.101 | 0.142 0.028 0.022 | 0.034
137 0.004 0.003 0.054 0.101 0.048 0.034 | 0.036
142 0.017 0.003 0.047 0.088 | 0.094 0.022 0.026 | 0.021
144 0.011 0.002 0.041 0.066 | 0.071 0.021 0.031 | 0.024
149 0.010 0.006 0.033 0.035 | 0.034 0.022 0.026 | 0.017
156 0.010 0.004 0.017 0.026 | 0.032 0.020 0.019

158 0.002 0.005 0.013 0.035 0.017 0.014 0.013 0.013
163 0.005 0.003 0.016 0.051 | 0.026

177 0.003 0.002 0.039 0.054 0.015 0.009
185 0.002 0.002 0.029 0.026 0.012 | 0.009
191 0.003 0.003 0.024 0.041 | 0.022 0.016 0.045 | 0.014
199 0.005 0.030 0.023 | 0.032 0.021 0.023
205 0.004 0.039 0.0297} 0.043 0.016 0.016 | 0.028
215 0.002 0.0278 0.0315} 0.017

226 0.001 0.0207 0.026 0.024 0.024 0.023
233 0.015 0.099 | 0.014
240 0.013 0.035
247 0.017 0.057
254 0.016 0.064
261 0.045
269 0.036
275 0.018
282 0.017

*H = Holstein

wKJ = Je

rsey



ACKNOWLEDGMENTS

The authors wish to express their appreciation to S. R. Lloyd, B. Johnson,
L. Dana, J. Titus, R. Hope, and the other personnel of the U.S. Environmental
Protection Agency Experimental Farm for their assistance during the experi-
mental phase of this study, and to R. R. Kinnison and P. Fort for the sta-
tistical analysis of the data.

viii


FOREWORD
ABSTRACT

LIST OF FIGURES
LIST OF TABLES
ACKNOWLEDGMENTS
INTRODUCTION
CONCLUSIONS
RECOMMENDATIONS
METHODS

RESULTS
DISCUSSION

REFERENCES

CONTENTS

Page
iii
iv
vi
vii

viii

14

15


ABSTRACT

Eight lactating dairy cows and three steers received a single oral ad-
ministration of tritiated water. Milk and blood from the lactating cows and
blood from the steers were periodically collected and analyzed for tritium
content.

The tritium content of whole milk decreased with time giving a curve
expressed as a three component exponential which yielded half-times of 3.04 +
0.09, 11.1 + 2.58, and >120 days. Tritium in the blood serum of steers de-
creased with half-lives of 4.05 + 0.21 and 40.4 + 9.82 days.

Additional study of the milk to assess tritium incorporation in the
various fractions showed half-times of 2.93 + 0.14 and 43.7 + 4.28 days for
milk serum; 2.08 + 0.56 and >50 days for milk protein; and 3.28 + 0.35 and
60.7 + 43 days for butterfat.

iv


© MILK SERUM: Ty, =2.9320.14 d
© PROTEIN: Ty= 2.080.56d

10 @BUTTERFAT: T)=3.28+0.35d
1 y TZ
10 b PROTEIN: T,->50d
eBUTTERFAT: Tp= 60.692 43.04 d
7 © MILK SERUM: T2=43.69+ 4,28 d
= 0
= 10-F
a)
8
= -1
Uo 10F
2
-2
10 fF
3
10
1 1 A 1 r 1 1 1 . 1 1 4
20 40 60 80 100 120 140 160 180 200 220
DAYS
Figure 2.

Average concentration of tritium in milk components following a single oral administration
of 200 mCi of tritiated water to four dairy cows.


Number

LIST OF TABLES

Tritium Concentration in uCi/l of Water Or Its
Equivalent in Whole Milk and Components Following
A Single Oral Administration of 100 millicurie of
Tritiated water

Tritium Concentration in yCi/l of Water Or Its Equiv-
alent in Milk Components Following A Single Oral
Administration of 200 millicurie of Tritiated

Water

Tritium Concentration in Blood Serum of Dairy Cows
Following A Single Oral Administration of 100
millicurie of Tritiated Water

Tritium Concentration in Blood Serum of Dairy Cows
Following A Single Oral Administration of 200
millicurie of Tritiated Water

Tritium Concentration in Blood Serum of Beef Cattle

Following A Single Oral Administration of 65
millicurie of Tritiated Water

vii

10

10

12


REFERENCES

Aschbacher, P. W., T. H. Kamal and R. G. Cragle. 1965. "Total body water
estimations in dairy cattle using tritiated water." J. Animal Sci.
24:430.

Black, A. L., N. F. Baker, J. C. Bartley, T. E. Chapman and R. W. Philips.
1964. "Water turnover in cattle." Sci. 144:876.

Glascock, R. F., and E. W. Wright. 1962. "Some Examples of the Use of Radio-
isotopes in Biochemistry." Uses of Radioisotopes In Animal Biology and
the Medical Sciences. Vol. 1. Academic Press, New York. p. 185.

Hausler, W. J., Jr., ed. 1972. Standard Methods for the Examination of Dairy
Products. Am. Pub. Health. Assoc. Washington, D.C. p. 225.

Horowitz, W., ed. 1970. Official Methods of Analysis of the Association of
Official Analytical Chemists. Assoc. of Official Anal. Chem., Washington,
D.C. 1015 pp.

Kirchmann, R., A. Lafontaine, J. Van den Hoek and G. Koch. 1969. "Transfer
and distribution of tritium among the main components of milk produced by
cows having ingested contaminated water." Comp. Rend. Soc. Biol. 63:1459.

Kirchmann, R., J. Van den Hoek and A. Lafontaine. 1971. "Transfer et incor-
poration du tritium dans les constituants de l'herbe et du lait en condi-
tions naturelles." Health Phys. 21:61. July, 1971.

Lieberman, R., and A. A. Moghissi. 1970. "Low-level counting by liquid scin-
tillation. II. Applications of emulsions in tritium counting." Int. J.

Appl. Radiat. Isotopes. 21:319.

Lubran, M., and G. Corsini. 1960. Mineiva Nucl. 4:130.

Moghissi, A. A., and M. W. Carter. 1968. "International standard with identi-
cal system properties for determination of liquid scintillation counting
efficiency." Anal. Chem. 40:812.

Moghissi, A. A., and M. W. Carter. 1973. Tritium. Messenger Graphics,
Phoenix, Arizona. p. 3277.

Moghissi, A. A., E. W. Bretthauer and E. H. Compton. 1973. "Separation of

water from biological and environmental samples for tritium analysis."
Anal. Chem. 45:1565.

15


EPA-600/3-77-076
June 1977 Ecological Research Series



into the environment and because of the lack of consistent information on the
behavior of tritium in components of man's food chain, this study was under-
taken in dairy cows and beef animals to evaluate both the short- and long-term
behavior in one of the more significant sources of exposure.

CONCLUSIONS

1. Little significant difference was noted in the rate of tritium in
body water turnover in lactating dairy cows versus steers.

2. Although whole milk is the major route of exposure of humans consum-
ing tritiated dairy products, additional information on the turnover rate of
tritium in milk solids is necessary before an accurate determination of radi-
ation dose can be made.

RECOMMENDATIONS

In the event of contamination of agricultural areas with tritium, the
potential hazard to humans ingesting tritiated dairy products must be carefully
evaluated. As more knowledge is gained of the metabolism of specific tritiated
compounds, it may be found that incorporation of tritium into nucleic acid, via
deoxyribonucleic acid (DNA), may influence all aspects of biological activity
and have greater effect than previously ascribed. Further studies with a
biological receptor are necessary to accurately estimate exposure/dose result-
ing from the consumption of tritium.

METHODS

These studies were carried out at the Nevada Test Site on the experimental
farm operated by the U.S. Environmental Protection Agency for the U.S. Energy
Research and Development Administration. The dairy animals selected for the
study were either Holstein or Jersey cows 3 to 9 years of age, were in their
30th to 100th day of lactation, and were producing 20 to 31 kilograms (kg) of
milk per day. The beef animals were 4- to 6-year-old Hereford steers. All the
animals were allowed free access to water and alfalfa hay. In addition, the
dairy cows received a pelleted commercial dairy feed containing 16% protein.

The cows were milked with a milking machine, and blood samples from the
cows and steers were taken by jugular venipuncture. Tritiated water was
administered to each animal following collection of milk and/or biood for
background activity determinations. The tritiated water was placed in gelatin
capsules containing starch and administered orally using a balling gun. Milk
samples were collected twice daily from the dairy animals, and blood samples


INTRODUCTION

As an isotope of hydrogen, tritium is incorporated into essentially all
components of biological systems. Tritium is produced as a result of nuclear
fusion and, to a certain extent, nuclear fission explosives, including those
used in the nuclear stimulation of natural gas formations (Moghissi and Carter,
1973). Tritium is also produced in all nuclear power reactors both as fission
and activation products. Due to the presence of large quantities of tritium in
the initial stages of operation, it is expected that tritium production in
fusion reactors will be substantially higher than in fission reactors. Because
of the problems associated with separation and disposal of tritium from nuclear
waste, dilution is presently used as the disposal method for this radionuclide.
However, it is expected that in the foreseeable future tritium will be sepa-
rated and contained.

Cow's milk is recognized as one of the major routes of the intake of
radioactive pollutants into the human body. Attempts to study tritium metab-
olism in bovines have been numerous. Black et al., (1964) studied the turnover
rate of tritium in a total of 17 lactating and nonlactating cattle. They
observed no difference in the turnover rate of tritium in body water (T,) in
nonlactating as compared to lactating animals. Their T, values ranged from 2.8
to 4.1 days with an average of 3.5 days. However, aschBacher et al., (1965)
studied a number of physiological parameters in dairy cows using four lactating
and two nonlactating animals. They observed T, values for tritium of 3.0 days
for lactating and 5.2 days for nonlactating cows.

The incorporation of tritium into the organic fraction of milk subsequent
to the intake of tritiated food has also been repeatedly studied. Lubran and
Corsini (1960) and Glascock and Wright (1962) studied tritium transfer into
certain components of milk subsequent to the intake of tritiated triglycerides.
These authors clearly showed the importance of considering the chemical form of
tritium intake when determining the composition of tritiated compounds in milk.
In a series of papers Kirchmann et al., studied tritium behavior in dairy cows
(Kirchmann et al., 1969; Kirchmann et al., 1971; and Van den Hoek and Kirch-
mann, 1971). These authors observed values of 3 to 5 days for the T. of
tritium in lactating cows. They also observed an increase of tritium concen-
tration in milk solids if tritiated grass was fed to the cows when compared to
intake as tritiated water. Values for the biological half-life of tritium in
milk solids were compared to those in body water.

Potter et al.,(1972) also studied tritium behavior using one dairy cow.
They observed a two-component exponential excretion rate corresponding to half-
lives of 3.1 and 40 days, respectively. They also showed a substantial incor-
poration of tritium into the organic component of the milk.

Due to concern over the continued release of large quantities of tritium


LIST OF FIGURES

Average concentration of tritium in whole milk
following a single oral administration of 100 mCi
of tritiated water to four dairy cows.

Average concentration of tritium in milk components
following a single oral administration of 200 mCi
of tritiated water to four dairy cows.

Average concentration of tritium in blood serum
following a single oral administration of triti-
ated water to dairy cows and beef animals.

Average concentration of tritium in blood serum

following a single oral administration of 65 mCi of
tritiated water to three beef cattle.

vi

1l

13


TABLE 5. TRITIUM CONCENTRATION IN BLOOD SERUM OF BEEF CATTLE FOLLOWING
A SINGLE ORAL ADMINISTRATION OF 65 mCi OF TRITIATED WATER

Time Cow #662 Cow #669 Cow #675
d pci/1 uci/1 pci/1
1/24 -—- --- 72
2/24 127 118 194
3/24 165 126 165
4/24 174 191 184
5/24 181 219 194
6/24 215 223 202
7/24 218 256 216
8/24 203 239 206
9/24 184 239 207
10/24 206 255 212
11/24 202 240 214
12/24 210 265 215
1 208 221 216
2 135 122 173
8 110 144 105
16 28.3 37.3 ---
23 11.4 12.1 11.2
28 5.45 5.51 4.84
30 1.92 2.09 2.21
65 0.071 0.096 -—-
72 0.064 0.063 0.065
79 0.056 0.076 0.075
86 0.044 0.039 0.047
93 0.028 0.078 0.031
106 0.065 0.067 0.030
114 0.029 0.023 0.030
121 0.038 0.030 0.020
128 0.028 0.021 0.018
135 0.023 0.018 0.025
139 0.015 0.018 0.017
142 --- 0.018 0.022
144 0.018 0.017 0.045
149 0.020 0.022 0.084
156 0.009 0.011 0.012
158 0.015 0.021 0.010
163 0.010 0.009 0.009

12


DISCLAIMER

This report has been reviewed by the Environmental Monitoring and Support
Laboratory-Las Vegas, U.S. Environmental Protection Agency, and approved for
publication. Mention of trade names or commercial products does not constitute
endorsement or recommendation for use.


TABLE 1. TRITIUM CONCENTRATION IN yCi/l OF WATER OR ITS EQUIVALENT IN WHOLE MILK AND COMPONENTS FOLLOWING
A SINGLE ORAL ADMINISTRATION OF 100 mCi OF TRITIATED WATER

MILK
Day-Post H-21 H-90 H-171 J-175 Day-Post H-21 H-90 H-171 J-175
te) 251 283 344 428 26 0.929 1.62 1.64 1.71
1 209 277 376 405 28 0.792 1.10 1.10 1.05
2 178 . 218 332 314 31 0.378 0.601 0.590 0.48
3 156 189 274 276 34 0.202 0.333 0.376 0.330
4 128 158 244 232 37 0.132 0.223 0.294 0.210
5 98.1 124 176 182 40 0.888 0.122 0.158 0.120
6 80.2 105 138 142 43 0.105 0.139 0.181 0,160
7 63.2 86.5 116 116 49 0.040 0.058 0.068 0.076
8 51.6 69.4 98.8 103 56 0.030 0.040 0.065 0.057
9 41.2 49.1 69.2 76.8 . 63 0.038 0.042 0.075 0.071
lo 33.2 48.1 57.6 63.2 70 0.028 0.038 0.888 0.063
11 28.0 40.7 51.9 54.6 77 0.029 0.045 0.045
12 21.9 31.3 39.0 41.0 84 0.027 0.043 0.043
13 16.6 25.0 30.9 35.2 91 0.030 0.033 0.047
14 13.2 20.4 23.6 27.1 98 0.030 0.038
16 9.33 13.3 16.2 17.6 105 0.024 0.033
1g 5.30 8.64 9.94 10.4 112 0.029 0.030
20 3.76 6.00 6.46 6.76 119 0.025 0.030
22 1.78 2.90 3.50 3.20 126 0.051 0.060
24 1.40 2.45 2.00 2.29 154 0.069 0.069

161 0.054 0.029


TABLE 2. TRITIUM CONCENTRATION IN uCi/1 OF WATER OR ITS EQUIVALENT IN MILK COMPONENTS FOLLOWING A
SINGLE ORAL ADMINISTRATION OF 200 mCi OF TRITIATED WATER

Time Serum Protein Fat

d *H*-16 H-132 H-135 JRR-175 H-16 H-132 H-135 J-175 H-16 H-132 H-135 J-175

0.3 181 144 -- --- 73 112 245 98.8 24.0 |-=- 17.9 11.9

1 354 -o- 322 159 94.4 138 135 --- 110 83.8 131

1.4 261 403 300 426 145 --= -—- 283 --- 7 ial 114

2 206 284 235 358 123 151 154 177 78.9 | 100 -- 72.1

2.4 188 290 206 315 114 152 70.9 76.3

3 142 242 210 314 95.1 4119 106 118 72.0 93.5 75.6 68.5

4 160 267 237 310 69.6 102 87.6 65.2 56.3 60.5

5 139 244 203 248 50.6 54.5 75.5 53.8

6 91.8 169 147 180 44.7 47.0 37.7 53.1 42.0 39.9

7 72.7 153 124 145 35.3 94.5 26.5 39.0 33.5 37.2

9 39.0 95.0 75.0 --- 15.2 43.0 14.7 28.7 22.6 23.8
ll 22.0 54.0 43.4 26.2 4.38 10.37 25.0 6.86 | 16.6 20.2
14 6.98 27.0 22.5 20.7 7.03 | 16.1 9.78 4.32 12.0 9.39 8.99
16 2.97 17.2 16.0 8.00 3.27 | 12.1 8.29 2.42 6.79 5.96 4.40
22 0. 385 4.45 2.19 1.44 2.72 4.42 3.52 3.26 1.66 2.58 1.66 1.04
24 0.225 3.04 1.85 0.770 0.956) 2.56 4.95 2,52 0.429] 1.60 0.955 0.661
29 0.217 1.27 0.750 --- 1.04 0.717 1.43 2.55 0.336 0.510 0.398
31 0.098 0.720 0.450 0.300 1.29 0.664 1.49 0.189)
37 oo --- --- -— 0.407 1.14 1.32 1.05 0.151] 0.687 0.162 0.673}
45 0.027 0.091 0.053 0.042 0.426] 1.10 0.270 0.910 0.085} 0.178 0.093 0.078
52 0.020 0.019 0.039 0.030 0.584] 0.684 0.588 1.82 0.060) 0.066 0.054
59 0.034 0.016 0.023 0.019 0.678) 0.626 0.653 2.06 0.061} 0.086 0.052 0.055
65 0.067 0.039 0.090 0.025 0.243) 1.09 0.481 0.822 0.065) 0.153 0.066 0.070
72 0.013 0.023 0.026 0.013 0.499] 0.866 0.452 1.05 0.033] 0.059 0.051 0.039
79 0.011 0.022 0.016 0.015 0.154 1.60 0.884 1.08 0.032 0.052 0.034 0.043}
86 0.010 0.006 0.013 0.009 0.119 0.583 0.596 0.030) 0.227 0.033 0.048]
93 0.010 0.004 0.010 0.008 0.103] 0.480 0.118 0.099 0.026] 0.258 0.038 0.053)
100 0.006 0.003 0.014 0.007 0.043] 0.252 0.057 0.056 0.047) 0.396 0.154 0.111
107 0.003 0.057 0.019 0.003 0.072] 0.444 0.086 0.082 0.040) 0.421 0.149 0.080)
114 0.004 0.063 0.005 0.005 0.060 0.132 0.076 0.090 0.038 0.150]
120 0.009 | --- 0.009 0.010 0.048 0.114 0.023 0.025 0.037



EPA-600/3-77-076
dune 1977

TRITIUM RETENTION BY COWS AND STEERS
AND TRANSFER TO MILK

by

A. L. Mullen, A. A. Moghissi} J. Ce Wawerna?

B. A. Mitchell, E. W. Bretthauer, and R. E. Stanley
Environmental Monitoring and Support Laboratory
Las Vegas, Nevada 89114

lpresent Address
Office of Interdisciplinary Programs
Georgia Institute of Technology
Atlanta, Georgia 30332

*present Address
Criminalistics Laboratory
Las Vegas Metropolitan Police Department
Las Vegas, Nevada 89106

ENVIRONMENTAL MONITORING AND SUPPORT LABORATORY
OFFICE OF RESEARCH AND DEVELOPMENT
U.S. ENVIRONMENTAL PROTECTION AGENCY
LAS VEGAS, NEVADA 89114


Moghissi, A. A., E. W. Bretthauer, E. L. Whittaker and D. N. McNelis. 1975.
“Oxygen bomb combustion of environmental and biological samples for tritum
analysis." Inter. J. Appl. Isotopes.

Moyer, A. W., and V. DuVigneaud. 1972. J. Biol. Chem. 143:373.

Potter, G. D., G. M. Vattuone and D. R. McIntyre. 1972. "Metabolism of triti-
ated water in the dairy cow." Health Phys. 22:405.

Schirch, L. V., and M. Mason. 1963, J, Biol. Chem. 238:1032,
Smith, T. E., and R. T. Taylor. 1969. "Incorporation of tritium from triti-
ated water into carbohydrates, lipids, and nucleic acids."

UCRL-50781. 24 p.

Van den Hoek, J., and R. Kirchmann. 1971. Radioecology Applied to the Pro-
tection of Man and His Environment. Comm. of European Communities. p. 1.

16 ‘8U, S. GOVERNMENT PRINTING OFFICE: 1977—784-860


RESEARCH REPORTING SERIES

Research reports of the Office of Research and Development, U.S.
Environmental Protection Agency, have been grouped into five series. These
five broad categories were established to facilitate further development and
application of environmental technology. Elimination of traditional grouping
was consciously planned to foster technology transfer and a maximum
interface in related fields. The five series are:

Environmental Health Effects Research
Environmental Protection Technology
Ecological Research

Pens

Environmental Monitoring
5. Socioeconomic Environmental Studies

This report has been assigned to the ECOLOGICAL RESEARCH series. This
series describes research on the effects of pollution on humans, plant and
animal species, and materials. Problems are assessed for their long- and
short-term influences. Investigations include formation, transport, and
pathway studies to determine the fate of pollutants and their effects. This work
provides the technical basis for setting standards to minimize undesirable
changes in living organisms in the aquatic, terrestrial, and atmospheric environments.

This document is available to the public through the National Technical
Information Service, Springfield, Virginia 22161.


were collected from all animals hourly for the first 12 hours and then at
longer intervals during the remainder of the project. Milk samples were stored
in 4-liter polyethylene containers to which 10 milliliters (ml) of formaldehyde
was added to retard spoilage. Blood samples were centrifuged and the serum
portion was removed for tritium analysis. All samples were processed for
analysis within 24 hours.

This study was conducted as a series of three consecutive experiments.
The milk transfer portion of the study utilized two groups of four lactating
dairy cows (three Holsteins and one Jersey) in each group. Each of the cows
was given a single 100-millicurie(mCi) dose of tritiated water. The milk from
this group was analyzed for tritium activity in the whole milk.

The total activity in the milk was determined directly by liquid scin-
tillation counting utilizing an internal standard. The scintillation liquid
consisted of p-xylene containing 7 grams (g) of 2,5-diphenyloxazole (PPO) and
1.5 g of bis-(o-methylstyrylbenzene) (bis-MSB) per liter mixed with Triton N101
(Rohm and -Haas, Philadelphia, Pennsylvania) in a volume ratio of 2:1. This
mixture could incorporate up to 10 ml of water or milk in a 25-ml vial (Lieber-
man and Moghissi, 1970). Counting efficiency was determined by using an inter-
nal standard with identical system properties (Moghissi and Carter, 1968). The
whole milk was mixed thoroughly and two aliquots of each sample were prepared
with one of them having a known quantity of tritium added.

The second group of dairy cows received 200 mCi of encapsulated tritiated
water each. Milk from these cows was separated into milk, serum, butterfat,
and protein fractions and the tritium activity of each fraction was determined.

Milk serum was separated by the addition of trichloracetic acid (15
g/100 m1). The presence of about 100 milligrams (mg) of hydrogen ions per gram
in the acid was regarded as acceptable and within expected errors. The mixture
was allowed to stand for about 30 minutes and filtered, and the filtrate was
distilled. The tritium activity in the distillate was determined by liquid
scintillation counting.

Cream was separated from the milk by cryogenic centrifugation. Fat was
separated from cream using heat and 100 ml of Triton and 25 grams of hexameta-
phosphate per liter of cream as described by Horowitz (1970). The purity of
the fat was checked using the Babcock test (Hausler, 1972).

The proteins were separated from the milk serum and dried for several days
in an oven at a temperature of 40° C. The dried protein was weighed and the
organically bound tritium separated by combustion (oxidation) in a Parr bomb.
After the water of combustion was collected, its tritium content was determined
by liquid scintillation counting (Moghissi et al., 1975).

After installation of a large scale azeotropic distillation system (Mo-
ghissi et al., 1973), the protein and serum separation was repeated using this
procedure and the results indicated a reasonable agreement between the two
methods,

In order to determine the difference in blood tritium levels between


leg 04:
10 14 73.04*0.09d

UCi/ liter of MILK
3S

2 +
Tpr1112=2.58d

Tp->120d

20 40 60 80 100

DAYS

Figure 1. Average concentration of tritium in whole milk following a single

oral administration of 100 mCi of tritiated water to four dairy
cows.