PHARMACOKINETICS OF AROCLOR 1254

COMPONENTS AFTER INTRAVENOUS

ADMINISTRATION TO SWINE AND SHEEP
R. E. BORCHARD, L. G. HANSEN, W. G. HUBER,
R. L. METCALF, and M. E. WELBORN
Department o f Physiology and Pharmacology
College o f Veterinary Medicine
University o f Illinois
Urbana, Illinois 61801

A method for quantitating individual PCB components as well as total PCB has
been utilized in determining 2-compartment pharmacokinetic parameters of
Aroclor 1254 after an i.v. dose in sheep and pigs. The resultant parameters show
that PCB components are not eliminated from blood of these species at the same
rates. The methods used provide a realistic means for assessing the time course of
PCB in the living animal.

In recent years, polychlorinated biphenyls (PCB) have been found in animal tissues
and food and water supplies throughout the world (Fishbein 1972, Maugh 1972) and
they are known to accumulate in the food chain of man and domestic animals (Nisbet
and Sarofim 1972). Although the industrial uses o f PCB's have recently been largely re-
placed by other substances (Broadhurst 1972), the environmental accumulation over the
past 30 years will present a real hazard to man and animals for years to come.

With this in mind, it becomes necessary to monitor exposure of animals to PCB's and
to predict the fate o f chemicals in the body. A very useful method o f monitoring PCB
exposure in the animal is through detection o f the substance in blood.

Since the commercial PCB products are composed of a mixture o f isomers containing
one to ten chlorine atoms each, it is necessary to monitor not only the total PCB, but
also the various components o f the mixture. This becomes more evident when one con-
siders that the bio-accumulation, metabolism and toxicity of these compounds vary ac-
cording to percentage and positions o f chlorination.

Animal exposure to PCB's may occur through many sources and more than one com-
mercial product may be present to complicate the exposure pattern. The resulting mix-
ture o f residues in the animal tissues may not resemble that o f any of the commercially
distributed mixtures, but one may still be able to make reasonable predictions o f its fate

Dr. Borchard's current address is College of Veterinary Medicine, Pullman, Wash. 99163; Dr.
Huber's, Hoffman-LaRoche, Nutley, N.J. 07110. Any correspondence should be addressed to Dr.
Hansen.

Archives of Environmental Contamination 179

and Toxicology, Vol. 2, No. 2, 1974, © 1974
by Springer-Verlag New York Inc.
180 R.E. Borchard et al.

in the animal by considering its component parts. This paper presents a preliminary study
of the pharmacokinetics of the component parts of a PCB mixture1 (Aroclor 1254) in
sheep and swine after an i.v. dose.

Materials and m e t h o d s
Three cross-bred ewes and three cross-bred pigs were used in this study. The three
sheep were 5- to 8-year-old western ewes weighing 70, 58, and 81 kg. The three pigs were
10 to 12 weeks of age, weighed 29, 29, and 38 kg, and consisted of two gilts and one
barrow of Hampshire-Yorkshire breeding.

The dosage of Aroclor 1254 in each animal was three mg/kg, given as an i.v. bolus of
a 2.0 percent solution of the Aroclor in acetone and propylene glycol. Blood samples
(6 to 8 ml) were collected for 24 hours after injection. In the sheep, injections were made
into and samples drawn from the jugular veins with needle and syringe. In the pigs, jugu-
lar and femoral cannulas were used for injection and sampling. The cannulas were placed
in the pigs while under pentobarbital anesthesia at least 24 hours before the in-
jections were made. Control blood samples were taken from each animal prior to injec-
tion of Aroclor.

Blood samples were extracted with hexane saturated acetonitrile, partitioned into
hexane, cleaned up on alumina columns and analyzed by electron capture gas chroma-
tography as described previously (Welborn et al. 1974).

Individual PCB components (Figure 1) were quantitated on the basis of the relative
amounts normally present in the commercial mixture (i.e. component concentration is
given as 1.0 ppm at the concentration present in 1.0 ppm of Aroclor 1254). Components
were also quantitated on the basis of actual component concentration by utilizing the
percentage contributed to the commercial mixture by that component (Hirwe et at.
1973). The actual amount of each component present in the injected solution can also be
determined from the percent component (Table I). The average i.v. dose of Aroclor 1254
in the sheep was 208 mg and in the pigs was 96 mg.

The i.v. data were fitted to a two-compartment open model (Riegelman et al. 1968) by
a high speed computer program for solution of the general least squares problem (Moore
and Zeigler 1959).

The general two-compartment open model:

A K1 B
Central [ " K2 " l Peripheral [
IK3
elimination,
metabolism, etc.

1Monsanto Electrical Grade, Lot No. KB-05-612.

 Pharmacokinetics of Aroclor 1254 Components 181

The mathematical two-compartment open model:

dA d__BB = K1 A - KzB
d--t- = K2B - K1A - K3A and
dt

The equation for the open two-compartment system, derived by integrating the model
differential equations:

CA- Dose ( a - K s ) e -at + Dose ( K 2 - / 3 ) e - ~ t

VA ( a - t~) VA (~ --'~)
or CA = A e - a t + Be-# t

where CA = Blood concentration at time t.

A = Blood concentration contributed by c o m p a r t m e n t A at t ; o.
B = Blood concentration contributed by compartment B at t = o.
= First order rate constant of elimination from compartmer~t A.
= First order rate constant o f elimination from compartment B.
VA -- Volume of the central compartment.

The following relationships were used to calculate pharmacokinetic parameters for an

open two-compartment system:

Co (blood concentration at t = o) = A + B
.693
t½ A (central compartment) -

B c tl I= j

HI
G K
O
/M
P

o
Fig. 1. Gas chromatography tracings of Aroclor 1242 (top) and Aroclor 1254 (bottom)
illustrating peak designation by relative retention time.
182 R . E . Borchard et aL

.693
t ~ B (peripheral compartment) -

K 1 =a+3-K 2 -K 3

K2 = ,a3
K3
Co
K 3 =
A+B
3
Dose
VA = m
A+B
Dose
Vd (apparent volume of distribution) = B

Dose
TBC (Total b o d y clearance) = /3 B

The values for A, B, a, and/3 were generated by the computer program.

Table I. Amounts of Aroelor 1254 Components Present in the Average i.v.

Dose in Sheep and ~'gs

Percent found Mg in average dose

Component in Aroclor 1254 Sheep Pigs

E ~ 0.2 0.4 0.2

F !
G 2.0 4.2 1.9
H 1.0 2.1 1.0
I 0.8 1.7 0.8
J 16.1 33.5 15.5
K 16.4 34.1 15.7

L ~ 20.2 42.0 19.4

M!
N 16.7 34.7 16.0
O 13.7 28.5 13.2
P 8.2 17.1 7.9
Q 2.1 4.4 2.0
R 1.9 4.0 1.8
S 0.6 1.2 0.6
T - - -

Totals 99.9 208.0 96.0

 Pharmacokinetics of Aroclor 1254 Components 183

The component values obtained for the rate constant, K3 and total body clearance,
TBC were compared statistically for possible differences occuring between species and
within each species. A paired t-test was used to compare individual components
between species and the students t-test was used to compare individual components
within species (Huntsberger 1967).

Results
Analysis o f samptes taken for 24 hours after an i.v. injection o f three mg/kg of
Aroclor 1254 provided data (Tables II and III) to plot the disappearance o f individual
PCB components from the blood o f sheep and pigs (Figures 2, 3, 4, 5). The earlier
sampling times (i.e., three through 30 min) represent a distribution phase and were not
included in the rate calculations; the data were necessary, however, to confirm even dis-
tribution and animals exhibiting irregular distribution patterns were excluded from the
studies.

The data were then fitted to the 2-compartment model and pharmacokinetic para-
meters were obtained which were used to plot fitted curves for the components and to
calculate rate constants and volumes described by the model (Table IV). Fitted curves
were drawn for the components in terms o f concentration relative to Aroclor 1254
(Figures 6 and 8), as well as in terms of actual concentration of each component
(Figures 7 and 9). The slopes of the curves, as well as all calculated constants remain the
same, whether the relative or actual values are used. However, if one uses the actual

Table II. Concentrations of Individual Components of Aroclor 1254

in Sheep Blood After 3. 0 mg/kg i.v.

Mean ppb for component Total

PCB
Time (hr) J K L M N 0 P Q R (ppm)

1 72 90 44 36 94 79 54 16 17 .502

2 48 66 27 19 69 60 41 1t 12 .353

3 27 45 14 12 45 40 35 8 11 .237

4 24 41 13 - 45 39 30 7 10 .209

6 12 25 6 - 27 28 21 5 - .124

8 7 20 6 - 20 24 17 3 - .097

12 5 15 4 - 14 t6 11 1 - .066

24 - 11 2 - 8 11 6 - -- .038
184 R.E. Borchard et al.

values for blood concentration, he must also use the actual value for the amount of com-
ponent injected.

The curves drawn using concentration values relative to Aroclor 1254 have different
positions on the graph than those drawn using actual component concentrations because
some component concentrations were high, relative to the amount injected, but were
still comparatively low in actual concentration. For example, at 24 hours in sheep, there
is more of components O and P relative to the amounts injected, but there is more of
component K in absolute quantity.

Pharmacokinetic parameters describing the 2-compartment model were obtained for

components J through R in pigs and components J, K, L, N, O, and P in sheep. Com-
ponent M disappeared very rapidly in sheep; it was not detected in the blood after three
hours following injection; the half-life was estimated to be 1.2 hours. Components Q and
R, relatively small fractions of Aroclor 1254, were not detected with enough consistency
in sheep blood to permit proper data analysis. Parameters for total PCB were calculated
for sheep and pigs, by using the sum of all component concentrations at each sampling
time.

The calculated values obtained for K 3 and TCB were compared for components J, K,
L, N, O, P, and total PCB with a paired t-test (for comparisons between species) and the
student's t-test (for comparisons within species). The results of these comparisons are
found in Table V.

Table III. Concentrations of Individual Components of Aroclor 1254

in Pig Blood After 3.0 mg/kg i.v.

Mean ppb for component Total

PCB
Time (hr) J K L M N O P Q R (ppm)

1 125 165 89 91 185 t52 102 27 32 .968

2 69 110 57 54 127 104 72 17 20 .630

3 48 83 40 39 87 73 60 14 16 .460

4 34 66 32 30 70 58 49 13 14 .366

6 26 45 20 20 46 47 34 9 11 .258

8 17 33 I1 8 32 32 30 6 8 .177

12 17 22 10 5 27 31 29 6 7 .154

24 8 15 5 4 13 16 16 4 3 .084
 Table IV. Estimated Pharmacokinetic Parameters for Aroclor 1254 Components After an i.v.
InJection in Sheep and Pigs.

A ~ B /3 t 21 A t 21 B K1 K2 K3 VA Vd TBC
ppm as ppm as
Component 1254 hr.-1 1254 hr.-1 hr. hr. hr.-1 hr.-1 hr.-1 Li. Li. Li/hr.

Sheep c~
O
J .647 .584 .103 .107 1.188 6.492 .158 .171 .362 277.33 2019.42 216.08
K .664 .455 .129 .028 1.524 25.106 .253 .099 .131 262.30 1612.42 45.15
L .794 .731 .077 .050 .948 13.767 .338 .111 .332 238.81 270t.30 135.07
N .612 .394 .t34 .045 1.758 15.399 .165 .109 .165 278.82 1552.24 69.85 O
O .628 .494 .202 .045 1.404 16.400 .246 .154 .136 250.60 1029.70 43.25
P .638 .253 .142 .029 2.733 23.970 .110 .067 .105 266.67 1464.79 42.48
TotN PCB .616 .465 .115 .047 1.490 14.745 .205 .113 .194 284.54 1808.70 85.01

Pigs
J 1.284 .768 .189 .058 .903 11.991 .377 .150 .299 65.24 508.47 24.49
K 1.136 .400 .188 .029 1.732 23.561 .202 .085 .t42 72.58 511.17 14.82
L 1.123 .425 .110 .033 1.630 21.178 .184 .068 .206 77.94 873.64 28.83
M . . . . . . . .111 .164 .342 71.08 - -
N 1.438 .534 .287 .054 1.298 12.731 .238 .135 .215 55.71 334.84 18.08
O 1.408 .583 .344 .044 1.189 15.886 .304 .152 .171 54.85 279.36 12.29
P 1.309 .521 .489 .037 1.329 18.582 .277 .167 .114 53.45 196.52 7.27
Q 1.230 .402 .370 .028 1.725 24.517 .220 .112 .098 60.06 259.73 7.27
R 1.818 .805 .806 .072 .861 9.668 .385 .297 .195 36.62 119.23 8.58
TotN PCB 1.213 .537 .252 .046 1.291 15.065 .262 .132 .189 65.60 348.100 17.54

Oo
ta~
 Table V. Comparison of K 3 and Total Body Clearance Values.

K3 TBC K3 TBC K3 TBC

Component Sheep vs Pig Sheep vs Pig Sheep Sheep Pig Pig

J NSD Sheep > Pig > K,N,O,P > K,N,O,P Total PCB > K,O,P Total PCB >P
K NSD NSD NSD NSD NSD NSD
L Sheep > Pig Sheep > Pig > K,O,P NSD NSD > K,O,P
N NSD Sheep > Pig K,O,P NSD NSD NSD
O NSD NSD NSD NSD NSD NSD
P NSD NSD NSD NSD NSD NSD
Total PCB NSD NSD NSD NSD NSD NSD
All values NSD Sheep > Pig NSD NSD NSD NSD

NSD = No Significant Difference at P < .05.

 Pharmacokinetics of Aroclor 1254 Components 187

Discussion
Pharmacokinetic parameters for the elimination of Aroclor 1254 components from
the blood of sheep and pigs after an i.v. dose have been determined. It is apparent that
some PCB components are handled differently by the animal body, especially the lower
chlorinated components J, L, and M (Hirwe et al. 1974). Based on preliminary results of
further studies underway in our laboratory even more differences among the components
and between species can be expected, especially with longer studies that may lead to
expansion of the pharmacokinetic model. We are not accounting for all of the PCB in
the animal at any given time; hence, the open model. Elimination may be accounted for
by metabolism, excretion, or further distribution.

We have not attempted to identify the compartments of the model, although it is

obvious that at least a portion of the central compartment is the blood-vascular system
(sampling compartment) and that the peripheral compartment should include the fatty
tissues of the body.

This study has utilized a method for quantitation of individual PCB components as
well as total PCB, based on the characterization of the PCB mixture used for the study.

2.0

1.0

0,5
LO

o
<

I3.
t~

,10
Swine

.05 Sheep

12 24
Time (hr)

Fig. 2. Elimination of component N of Aroclor 1254.

 188 R . E . Borchard et al.

1.0--

0.5
I.o

L
<

Swine
.10

~ Sheep

i L t
6 12 24
Time (hr)

Fig. 3. Elimination of component O of Aroclor 1254.

2.0 --

1,0 --

0.5--

10 Sh p

O5

L 1 1
6 12 24
Time (hr)
Fig. 4. E l i m i n a t i o n of c o m p o n e n t P of A r o c ] o r 1254.
 Pharmacokinetics of Aroclor 1254 Components 189

1.0 -~

0.5

Swine
o_ \\ ~ . ~ 2 . 3
o. .05
•\ ~ Sheep
\
\\ --%/2. 3
\
\
\
I \, I ........1
6 12 24
Time (hr)

Fig. 5. Elimination of total PCB illustrating computer-determined values for A, c~, B, and

1"0 I

Sheep
0.5

Lf)

o
<
.10

Q.

.05
PCB

I I
.02 ; 12 24
Time (hr)

Fig. 6. Computer-generated elimination rates for Aroclor 1254 components from sheep
blood. Relative values.
190 R . E . Borchard et al.

100 -

50
Sheep

E
O
Q.

en
O.
e~

2 I f ""--.q L
6 12 24
Time (hr)
Fig. 7. Computer-generated elimination rates for Aroclor 1254 c o m p o n e n t s from sheep
blood. Absolute values.

5.0

Swine

1.0

0.5

.10
!otal PCB
Q.

.05

oli 6 12
Time (hr)
24

Fig. 8. Computer-generated elimination rates for Aroclor 1254 c o m p o n e n t s from swine

blood. Relative values.
 Pharmacokinetics of Aroclor 1254 Components 191

500 ~-

Swine

100

50
O
r~
E
0
O
=0
m
10
ct.

5.0 ~ - L

t t I
6 12 24
Time (hr)

Fig. 9. Computer-generated elimination rates for Aroclor 1254 components from swine
blood. Absolute values.

Acknowledgments
This study was supported by research contract FDA 72-116, Department of Health,
Education and Welfare, Food & Drug Administration, Bureau of Veterinary Medicine.

The authors wish to thank Mr. Walter Crackel for his assistance with computer
methods, Ms. Joan Czachorski for her technical assistance, and Monsanto Chemical
Company for kindly furnishing the Aroclor 1254.

References
Broadhurst, M. G.: Use and Replaceability of Polychlorinated Biphenyls: Environ.
Health Perspec. 2, 81 (1972).
Fishbein, L.: Chromatographic and Biological Aspects of Polychlorinated Biphenyls. J.
of Chromatography 68, 345 (1972).
Hirwe, S. N., R. E. Borchard, L. G. Hansen, R. L. Metcalf: Gas-Liquid Chromato-
graphic and Mass Spectrometric Characterization of Aroclor 1242 and 1254 Com-
ponents. Bull. Environ. Contain. Toxicol. Accepted for publication (1974).
Huntsberger, D. V.: Elements of Statistical Inference. 2 ed. Boston: Allyn and Bacon
(1967).
192 R.E. Borchard et al.

Maugh, T. H.: Polychlorinated Biphenyls: Still Prevalent, but Less of a Problem.

Science 178, 388 (1972).
Moore, R. H., and R. K. Zeigler: The Solution of the General Least Squares Problem
with Special Reference to High-speed Computers. Los Alamos Scientific Laboratory
Publication LA-2367. (1959).
Nisbet, C. T., and A. F. Sarofim: Rates and Routes of Transport of PCB's in the
Environment. Environmental Health Perspectives 1,21 (t 972).
Riegetman, S., Loo, J. C. K., and M. Rowland: Shortcomings in Pharmacokinetic Analysis
by Conceiving the Body to Exhibit Properties of a Single Compartment. J. of
Pharmaceut. Sci. 57, 117 (1968).
Welborn, M. E., R. E. Borchard, S. N. Hirwe, L. G. Hansen, and R. L. Metcalf: Extraction
and analysis of Aroclor Components in Blood. J. Ass. Offic. Anal. Chem. Submitted
for publication (1974).

Manuscript received September 12, 1973; accepted January 8, 1974