Fd Cosmet. Toxicol. Vol. 3, pp. 547-551. Pergamon Press 1965. Printed in Great Britain

SUMMARY OF TOXICOLOGICAL DATA

DISPOSITION OF B U T Y L A T E D H Y D R O X Y T O L U E N E ( B H T ) I N T H E RAT

R. TYE, J. D. ENCEL and I. RAPIEN The Kettering Laboratory, University of Cincinnati, Cincinnati, Ohio, USA

(Summary* o f unpublished report dated 2 October 1962)

Introduction

The present investigation was undertaken with the primary objective of comparing the characteristics of the absorption, distribution, storage and excretion of butylated hydroxy- toluene (BHT; 3,5-di-tert-butyl-4-hydroxytoluene) by rats, with those of a related com- pound. The 14C isotope of BHT labelled in the r-posit ion of the tert-butyl groups was used throughout the present experiments.

Absorption and excretion

Oral route. Pairs of young, adult, albino rats of the Nelson strain (1 male and 1 female per pair) were given 1-5 oral doses of about 0.2 mmoles [~4C]BHT/kg, i.e. about 44 mg/kg, on alternate days commencing day 0 and ending on day 9. Daily elimination of 14C in the urine and faeces was followed until the pairs of rats were killed off on days 1, 3, 6 and 8, i.e. 24 hr after the administration of the respective terminal doses. However, after the fifth and final dose, the last remaining pair of rats was kept alive for a further period of 8 days (until day 17), during which the daily excretion of 14C was also recorded.

Table 1. Fate of repeated oral doses of [14C]BHT (0"2 mmoles/kg) in the rat

Percentage of total dose in Percentage of

Gastro-intestinal total dose Faeces Urine tract Body accounted for

No. of doses M F M F M F M F M F

1 57.8 22-6 2-8 18.6 27.5 35.5 4.4 15.9 92-0 92.6 2 69.6 30.9 6.2 36.5 23.2 17.5 4.5 8.8 103.5 93-7 3 79-5 45.9 4.0 25.4 12.9 18.5 2.3 6.8 98.7 98.6 4 82.5 NR 3.9 NR 10.0 NR 2.5 NR 98-9 NR 5 76.9 50.6 15.1 43.1 0.4 3.7 0.3 0.6 92.7 98.0

M=Male F=Female NR=No result due to escape of animal

F rom Table 1 it can be seen that substantial amounts of 14C were eliminated f rom the body via the urine and faeces. A negligible amount (0.05%) of the dose was exhaled f rom the lungs of 2 males in 24 hr following the administration of a dose of 0-2 mmoles/kg. Moderate amounts of 14C were still present in the gastro-intestinal tract, but comparatively

*Prepared by BIBRA and published with the permission of the responsible authorities.

M 547

Tab

le 2

. T

empo

ral c

ours

e o

f eli

min

atio

n o

f 14

C in

faec

es a

nd u

rine

of r

ats

give

n 1-

5 or

al d

oses

of

[x4C

]BH

T (0

.2 m

mol

es/k

g)

Per

cent

age

of

do

se e

xcre

ted

on

day

s A

ver

age*

pe

rcen

tage

1-

2 3-

5 6-

7 8-

9 10

-12

13-1

4 •

15-1

6 17

o

f to

tal

No

. o

f d

ose

do

ses

F U

F

U

F U

F

U

F U

F

U

F U

F

U

excr

eted

Mal

es

I 84

"8*

2"8

88

(27.

5)

2 67

.1

7.4

93.1

* 4.

9 86

(4

6.3)

3

63-0

4-

3 92

.0

5.9

122.

4*

1.8

63

(38-

8)

4 72

.6

3.9

86.3

6.

1 93

-8

3.7

117.

6"

1.8

97

(40-

1)

5 66

-8

8.1

75.9

18

.7

67.9

9.

7 76

.3

12.3

67

.8

15.6

24

.6

6.7

3.4

2.4

3.2*

1.

9 92

(1

"8)

Av

erag

et

71

5 87

9

95

5 97

7

85

Fem

ales

1

58"1

* 18

"6

77

(35"

5)

2 33

"6

34'4

6

3"2

*

38"6

85

(3

5"0)

3

32"6

24

"9

63"6

35

"4

96"8

* 16

"1

90

(55"

5)

4 N

R

13"9

N

R

NR

N

R

NR

N

R

NR

N

R

NR

N

R

NR

N

R

NR

N

R

NR

N

R

5 41

-1

26.4

47

.1

54.9

36

-6

31.0

53

.3

31.3

52

.5

50.1

16

.5

14.0

2-

5 5.

3 22

.0*

2.4

97

(18.

6)

Av

erag

et

43

24

58

43

67

24

87

O 8 >.

F=

Fae

ces

U=

Uri

ne

N

R=

No

re

sult

due

to

esca

pe o

f an

imal

. *T

otal

val

ue,

incl

udin

g 14

C c

on

ten

t o

f ga

stro

-int

esti

nal

trac

t 24

hr

afte

r d

osi

ng

; th

is a

mo

un

t is

sh

ow

n i

n br

acke

ts.

tAv

erag

e va

lues

are

exp

ress

ed t

o ne

ares

t w

hole

nu

mb

er.

SUMMARY OF TOXICOLOGICAL DATA 549

small amounts were retained elsewhere in the body. Quantitative recoveries of the total ~4C administered were obtained. It is noteworthy that the fate of BHT in the body was the same, irrespective of the number of doses administered.

Table 1 also reveals a striking sex difference in the main pathways for the elimination of BHT. Whereas, females excreted considerable quantities in their urine (19--43 ~o of the dose), males put out as much as 58-83 ~o of the dose in the faeces but only 3-15 ~o in the urine. Despite the greater absorption of BHT by females, both sexes readily eliminated the anti- oxidant from the body.

Table 3. Excretion of x4C in urine and faeces of rats given single subcutaneous injections of [a4C]BHT

Percentage of dose excreted in

Faeces Urine Percentage of Dose total doso

(mmoles/kg) 0-2 days 2-4 days 0-2 days 2-4 days excreted

0-0003 40 37 11 3 91 0.0022 26 12 7 3 48 0.025 24 6 4 2 37 0.22 20 9 5 2 36

Details of the daily faecal and urinary output of BHT are shown in Table 2. As already indicated in Table 1, there was no tendency on the part of either sex for the total output of z4C to drop with increasing number of doses. After the fifth and concluding dose, the expected decline in the amount of excreted ~4C ensued. By day 17, up to 92 ~o by males, and 97 ~ by females, of the total BHT administered, had been eliminated from the body.

Subcutaneous route. In view of the high faecal output of the antioxidant, the subcu- taneous route was employed to seek the main excretory pathway of that part of the dose that had been absorbed into the body. The study was limited to 4 female rats of the Charles River CD strain to which were given single, subcutaneous injections of 0.0003, 0.0022, 0.025 and 0.22 mmoles/kg in peanut oil. Urinary levels of 14C were estimated 2 and 4 days after the administration of the material. In direct contrast to the oral route, females ex- creted the majority of the dose (29-77 ~) in the faeces and small amounts (6-14~) in the urine (Table 3). Increasing the dose resulted in a lowering of the relative rate of excretion of ~4C, an effect which was reflected equally in the faeces and urine.

Distribution and storage

Employing the same schedule of oral dosage as described above, the distribution and storage of [~4C]BHT were determined in the tissues of the rat (Table 4). The highest concen- trations of ~4C were detected in liver, kidneys, lungs and body fat. Notably low levels were present in the brain, blood and testes. In general, females retained more 14C in the tissues than males, a difference which was especially marked in the gonads. Concentrations of 14C in tissues were not related to the number of doses administered, and hence there was no accumulation of ~4C in the tissues on repeated administration. In fact a similar pattern of the distribution of ~4C in the tissues emerged the day after each dose was given. With the excep- tion of the skirt and body fat, the majority of the tissues of rats, killed 8 days after the fifth and final dose, contained less 14C than the tissues of rats killed only 24 hr after this dose had

S.A

Tab

le 4

. P

erce

ntag

e dis

trib

utio

n o

f 1~

C in

tis

sues

foll

owin

g re

peat

ed o

ral d

oses

of

[t'C

]BH

T (

0.2

mm

oles

/kg)

Per

cent

age

of s

ingl

e do

se p

er g

tis

sue

No.

of

i do

ses

Liv

er

Kid

neys

B

rain

L

ungs

H

eart

G

onad

s A

dren

als

Spl

een

Blo

od

Ski

n M

uscl

e F

at

Mal

es

,.¢

1 0'

050

0'01

2 0.

0017

0'

013

0.00

6 0-

004

0.02

1 0.

005

0.01

0 0.

017

0.00

3 0-

023

2 0.

I00

0-02

5 0.

0034

0'

011

0.01

6 0.

007

0.01

6 0.

009

0.01

5 0.

025

0.00

9 0-

069

3 0'

086

0.01

4 0.

0017

0.

015

0-01

0 0.

005

0'03

3 0.

008

0.01

5 0"

019

0.00

4 0.

044

d 4

0"08

8 0.

023

0.00

57

0.02

3 0.

013

0.00

6 0.

022

0-00

9 0"

017

0.04

2 0-

005

0"06

5 [~

5*

0-

008

0.00

3 0.

0004

0.

004

0.00

3 0.

0003

0-

012

0.00

6 0.

012

0-00

4 0.

001

0.00

9 0 O

Fe

mal

es'l"

I

0-13

7 0"

039

0-00

95

0'05

9 0.

039

0"05

6 0'

065

0'01

5 0.

051

0.12

0 0.

020

0-16

1 2

0.16

2 0.

042

0.00

91

0.04

5 0.

040

0"15

0 0"

089

0.01

6 0.

066

0.11

7 0'

033

0.12

3 3

0"13

5 0"

038

0.00

95

0"06

7 0'

033

0"07

7 0-

196

0.01

6 0.

052

0"18

5 0-

019

0.24

3 5*

0.

011

0.00

7 0-

0010

0.

013

0.00

5 0.

016

0.01

2 0'

010

0'02

1 0.

018

0.00

2 0'

054

*Ani

mal

s w

ere

kill

ed o

ff 8

day

s af

ter

fift

h do

se,

othe

r an

imal

s w

ere

kill

ed 2

4 hr

aft

er r

ecei

ving

thei

r fi

nal

dose

. fN

o r

esul

ts f

or f

emal

es o

n 4

dose

s be

caus

e of

esc

ape.

SUMMARY OF TOXICOLOGICAL DATA 551

been administered. This observation may be attributed to the continued excretion of gradually declining amounts of 14C in the 8-day period after the fifth dose, as shown in Table 2.

Summary and conclusion Repeated oral doses of [14C]BHT did not influence the rapid rate of elimination of 14C

from the bodies of male and female rats. The main excretory pathway was that of the faeces but the urinary route was far more important in females than in males. The greater absorp-

14 tion of [ C]BHT by f~males was reflected in the higher levels of 14C in their tissues, as compared with males.frhere was no evidence of accumulation of [14C]BHT in the body under the conditions 6f repeated dosage employed in these experiments