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Mutagenic activity of BC and 4CMB in the Salmonella spot test
Mutation Research, 100 (1982) 17-19
17
Elsevier Biomedical Press
MUTAGENIC
ACTIVITY
OF BC AND
4CMB
IN
THE
S A L M O N E L L A
SPOT
TEST
F. H Y L D I G - N I E L S E N l and B. H A R T L E Y - A S P 2
Microbiology Department, H. Lundbeck and Co., Valby, DK-2500 Copenhagen I (Denmark) and Pharmacology Department, AB Leo, S-251 09 Helsingborg2 (Sweden) (Received 16 April 1981) (Accepted 23 June 1981)
The Salmonella
s p o t t e s t is t h e s i m p l e s t w a y t o t e s t s u b s t a n c e s
as several concentrations
sitive than the plate test and therefore should
be considered
for mutagenicity,
c a n b e s c r e e n e d p e r p l a t e . It h a s b e e n r e g a r d e d
as adequate
only a large increase in the number
evidence
as less senof colonies
for mutagenicity.
TABLE 1 M U T A G E N I C A C T I V I T Y OF 4CMB A N D BC AS I N D I C A T E D BY T H E S A L M O N E L L A SPOT TEST Compound
2-Aminoanthracene
Benzylchloride
aConc.
Strain
~g/ml)
TA1535
TA1537
TA1538
+$9
+$9
+$9
-$9
+ + + + ++
--
+ + +
--
+++
--
+ + +
--
+ + +
--
+ + +
--
+ + +
2 20 40 10 1 000 10000
-$9
+ + + + +
-
-$9
+ +
-
TA98
TAI00
+$9
-$9
+$9
} .
.
. . . . .
. . . . .
.
.
.
.
.
-$9
b
.
100 1000 4CMB
1 100 10000 10 1000
.
.
. .
. .
. .
.
. +
. .
.
. . +
a2 or 3 concentrations per plate. b The effect o f the single discs cannot be read (see discussion).
0165-1218/82/0000-0000/$02.75 © Elsevier Biomedical Press
. ? + ? +
.
. + . +
. ? + .
. +
+
+
+
+
.
18 MATERIALS AND METHODS T h e s p o t test was c a r r i e d o u t a c c o r d i n g t o A m e s et al. (1975). B C was d i l u t e d in e t h a n o l a n d 4 C M B d i s s o l v e d a n d d i l u t e d in D M S O . 2 0 / z l o f t h e s o l u t i o n s w e r e i m b i b e d i n t o 6 m m p a p e r discs ( S c h l e i c h e r a n d S c h 0 e l l N o . 2668). T h e c o n c e n t r a t i o n , i n d i c a t e d in the t a b l e s , is t h e c o n c e n t r a t i o n o f the c o m p o u n d in t h e s o l v e n t , i m b i b e d in t h e disc. I n E x p t . 1, 2 o r 3 discs ( o n e disc p e r c o n c e n t r a t i o n ) h a v e b e e n a p p l i e d o n e a c h p e t r i d i s h ( T a b l e 1). In E x p t . 2 o n l y o n e disc was a p p l i e d per p e t r i dish ( T a b l e 2). DISCUSSION O n e o f t h e s u b s t a n c e s in this s t u d y (BC) is n o t s u i t a b l e f o r t e s t i n g in m o r e t h a n o n e c o n c e n t r a t i o n p e r p l a t e w i t h this m e t h o d as t h e r e v e r t a n t c o l o n i e s a r e s p r e a d e v e n l y o v e r t h e w h o l e p l a t e b l u r r i n g t h e i n f l u e n c e f r o m t h e discs o f l o w e r c o n c e n t r a t i o n s . T h i s e f f e c t is p o s s i b l y d u e t o a v a p o u r f r o m t h e s u b s t a n c e s p r e a d b e t w e e n t h e lid a n d t h e s u r f a c e o f t h e a g a r .
TABLE 2 THE NUMBER OF REVERTANTS FOUND AFTER TREATMENT WITH BC AND 4CMB IN THE SALMONELLA SPOT TEST Compound
Conc.
Strain
(~g/ml)
Benzylchloride
4CMB
- $9
23 40 10 100 1 000 10 000 100 000 1
10 100 1 000 10 000 ND, not done. Inhibition of background lawn.
a
TA98
+ $9 Control 2-Aminoanthracene
TA1538
TAIO0
+ $9
- $9
+ $9
- $9
19
52
33
193
136
+ +
12
+ + +
33
+ + +
132
ND ND ND ND 43
ND ND ND ND 14
62 46 54 61 61
23 21 32 28 34
150 152 152 188 1000a
139 145 131 156 452a
34 28 36a 72a 125a
14 20 131a 468a 846a
40 47 59a 90a 145a
28 43 96 350 574a
155 136 185 288a 494a
129 111 162 273 476a
19 CONCLUSIONS 4CMB exhibited rnutagenic activity in TA1538, TA98 and TA100 whereas BC was only active in T A 1 0 0 . 4 C M B was active at a lower concentration than BC in TA100. Thus, 4CMB exhibits a higher mutagenic potential than BC in the Salmonella typhimurium spot test. REFERENCE Ames, B.N., J. McCann and E. Yamasaki (1975) Methods for detecting carcinogensand mutagens with the Salmonella/mammalian-microsomemutagenicity test, Mutation Res., 31,347-364.
17
Elsevier Biomedical Press
MUTAGENIC
ACTIVITY
OF BC AND
4CMB
IN
THE
S A L M O N E L L A
SPOT
TEST
F. H Y L D I G - N I E L S E N l and B. H A R T L E Y - A S P 2
Microbiology Department, H. Lundbeck and Co., Valby, DK-2500 Copenhagen I (Denmark) and Pharmacology Department, AB Leo, S-251 09 Helsingborg2 (Sweden) (Received 16 April 1981) (Accepted 23 June 1981)
The Salmonella
s p o t t e s t is t h e s i m p l e s t w a y t o t e s t s u b s t a n c e s
as several concentrations
sitive than the plate test and therefore should
be considered
for mutagenicity,
c a n b e s c r e e n e d p e r p l a t e . It h a s b e e n r e g a r d e d
as adequate
only a large increase in the number
evidence
as less senof colonies
for mutagenicity.
TABLE 1 M U T A G E N I C A C T I V I T Y OF 4CMB A N D BC AS I N D I C A T E D BY T H E S A L M O N E L L A SPOT TEST Compound
2-Aminoanthracene
Benzylchloride
aConc.
Strain
~g/ml)
TA1535
TA1537
TA1538
+$9
+$9
+$9
-$9
+ + + + ++
--
+ + +
--
+++
--
+ + +
--
+ + +
--
+ + +
--
+ + +
2 20 40 10 1 000 10000
-$9
+ + + + +
-
-$9
+ +
-
TA98
TAI00
+$9
-$9
+$9
} .
.
. . . . .
. . . . .
.
.
.
.
.
-$9
b
.
100 1000 4CMB
1 100 10000 10 1000
.
.
. .
. .
. .
.
. +
. .
.
. . +
a2 or 3 concentrations per plate. b The effect o f the single discs cannot be read (see discussion).
0165-1218/82/0000-0000/$02.75 © Elsevier Biomedical Press
. ? + ? +
.
. + . +
. ? + .
. +
+
+
+
+
.
18 MATERIALS AND METHODS T h e s p o t test was c a r r i e d o u t a c c o r d i n g t o A m e s et al. (1975). B C was d i l u t e d in e t h a n o l a n d 4 C M B d i s s o l v e d a n d d i l u t e d in D M S O . 2 0 / z l o f t h e s o l u t i o n s w e r e i m b i b e d i n t o 6 m m p a p e r discs ( S c h l e i c h e r a n d S c h 0 e l l N o . 2668). T h e c o n c e n t r a t i o n , i n d i c a t e d in the t a b l e s , is t h e c o n c e n t r a t i o n o f the c o m p o u n d in t h e s o l v e n t , i m b i b e d in t h e disc. I n E x p t . 1, 2 o r 3 discs ( o n e disc p e r c o n c e n t r a t i o n ) h a v e b e e n a p p l i e d o n e a c h p e t r i d i s h ( T a b l e 1). In E x p t . 2 o n l y o n e disc was a p p l i e d per p e t r i dish ( T a b l e 2). DISCUSSION O n e o f t h e s u b s t a n c e s in this s t u d y (BC) is n o t s u i t a b l e f o r t e s t i n g in m o r e t h a n o n e c o n c e n t r a t i o n p e r p l a t e w i t h this m e t h o d as t h e r e v e r t a n t c o l o n i e s a r e s p r e a d e v e n l y o v e r t h e w h o l e p l a t e b l u r r i n g t h e i n f l u e n c e f r o m t h e discs o f l o w e r c o n c e n t r a t i o n s . T h i s e f f e c t is p o s s i b l y d u e t o a v a p o u r f r o m t h e s u b s t a n c e s p r e a d b e t w e e n t h e lid a n d t h e s u r f a c e o f t h e a g a r .
TABLE 2 THE NUMBER OF REVERTANTS FOUND AFTER TREATMENT WITH BC AND 4CMB IN THE SALMONELLA SPOT TEST Compound
Conc.
Strain
(~g/ml)
Benzylchloride
4CMB
- $9
23 40 10 100 1 000 10 000 100 000 1
10 100 1 000 10 000 ND, not done. Inhibition of background lawn.
a
TA98
+ $9 Control 2-Aminoanthracene
TA1538
TAIO0
+ $9
- $9
+ $9
- $9
19
52
33
193
136
+ +
12
+ + +
33
+ + +
132
ND ND ND ND 43
ND ND ND ND 14
62 46 54 61 61
23 21 32 28 34
150 152 152 188 1000a
139 145 131 156 452a
34 28 36a 72a 125a
14 20 131a 468a 846a
40 47 59a 90a 145a
28 43 96 350 574a
155 136 185 288a 494a
129 111 162 273 476a
19 CONCLUSIONS 4CMB exhibited rnutagenic activity in TA1538, TA98 and TA100 whereas BC was only active in T A 1 0 0 . 4 C M B was active at a lower concentration than BC in TA100. Thus, 4CMB exhibits a higher mutagenic potential than BC in the Salmonella typhimurium spot test. REFERENCE Ames, B.N., J. McCann and E. Yamasaki (1975) Methods for detecting carcinogensand mutagens with the Salmonella/mammalian-microsomemutagenicity test, Mutation Res., 31,347-364.