1,4,6,10-Dodecatetraen-3-ol,3,7,11-trimethyl-,(4E,6E)-

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Remarks:

Type of genotoxicity: gene mutation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1999-03-17 to 2000-04-17

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP and guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2000

Report date:

2000

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Version / remarks:

1997

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)

Version / remarks:

1992

Deviations:

no

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

1,4,6,10-Dodecatetraen-3-ol,3,7,11-trimethyl-,(4E,6E)-

EC Number:

611-799-6

Cas Number:

59121-99-2

Molecular formula:

C15H24O

IUPAC Name:

1,4,6,10-Dodecatetraen-3-ol,3,7,11-trimethyl-,(4E,6E)-

Test material form:

liquid: viscous

Method

Species / strain

Species / strain 1

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98 and TA 100

Species / strain 2

Species / strain / cell type:

E. coli WP2 uvr A

Metabolic activation:

with and without

Metabolic activation system:

Liver homogenates (S9 fraction) from male Sprague-Dawly rats, induced with 500 mg/kg bw Aroclor 1254 (single injection intraperitoneal)

Test concentrations with justification for top dose:

All concentrations in µg/plate:
Experiment 1:
0; 20; 100; 500; 2,500; 5,000
Experiment 2:
0 ; 20; 100; 500; 2,500; 5,000
Experiment 3:
Salmonella strains: 0; 0.8; 4; 20; 100; 500
E. coli: 0; 4; 20; 100; 500; 1,000
Experiment 4:
Salmonella strains: 0; 3.125; 6.25; 12.5; 25; 50
E. coli: 0; 4; 20; 100; 500; 1,000

Vehicle / solvent:

- Vehicle/solvent used: DMSO
- Justification for choice of solvent/vehicle: Due to the limited solubility of the test substance in water, DMSO was selected as the vehicle, which had been demonstrated to be suitable in bacterial reverse mutation tests and for which historical control data are available.

Controls

Controls 1

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: 2-aminoanthracene (2-AA)

Remarks:

With metabolic activation; strains: TA 1535, TA 100, TA 1537, TA 98, E. coli WP2 uvrA

Controls 2

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

N-ethyl-N-nitro-N-nitrosoguanidine

Remarks:

Without metabolic activation; strains: TA 1535, TA 100

Controls 3

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: 4-nitro-o-phenylendiamine (NOPD)

Remarks:

Without metabolic activation; strains: Ta 98

Controls 4

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

9-aminoacridine

Remarks:

Without metabolic activation; strains: TA 1537

Controls 5

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

4-nitroquinoline-N-oxide

Remarks:

Without metabolic activation; strains: E. coli WP2 urvA

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation); preincubation

DURATION
- Preincubation period: 20 min
- Exposure duration: 48-72 hours

NUMBER OF REPLICATIONS: 3

DETERMINATION OF CYTOTOXICITY
- decrease in the number of revertants
- clearing or diminution of the background lawn (= reduced his- or trp- background growth)
- reduction in the titer

Evaluation criteria:

Acceptance criteria:
Generally, the experiment is to be considered valid if the following criteria are met:
- The number of revertant colonies in the negative controls was within the normal range of the historical control data for each tester strain.
- The sterility controls revealed no indication of bacterial contamination.
- The positive control articles both with and without S-9 mix induced a significant increase in the number of revertant colonies within the range of the historical control data.
- The titer of viable bacteria was > 10^9 / mL.

Evaluation criteria:
The test item is considered positive in this assay if:
- A dose-related and reproducible increase in the number of revertant colonies, i.e. about doubling of the spontaneous mutation rate in at least one tester strain either without S-9 mix or after adding a metabolizing system.

A test substance is generally considered nonmutagenic in this test if:
- The number of revertants for all tester strains were within the historical negative control range under all experimental conditions in two experiments carried out independently of each other.

Results and discussion

Test results

Test results 1

Species / strain:

S. typhimurium TA 1535, TA 1537, TA 98 and TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

depending on the strain and conditions from about 20 - 500 µg/plate onward

Vehicle controls validity:

not examined

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 2

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

depending on the strain and conditions from about 20 - 500 µg/plate onward

Vehicle controls validity:

not examined

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Water solubility: There was a limited solubility of the test substance in water.
- Precipitation: No test substance precipitation was found.

ADDITIONAL INFORMATION ON CYTOTOXICITY: A bacteriotoxic effect (reduced background growth, decrease in the number of revertants, reduction in the titer) was observed depending on the strain and test conditions from about 20 µg - 500 µg/plate onward.

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Any other information on results incl. tables

Experiment 1 & 2 (standard plate test)

Dose (µg/plate)	Mean number of revertant colonies of 3 replicates (± S.D.) for different strains of S. typhimurium and E. coli
	TA 100	TA 98	TA 1535	TA 1537	WP2 uvrA
	Results without S9
DMSO	142 ± 8	27 ± 3	21 ± 4	10 ± 2	30 ± 5
20	120 ± 10	23 ± 3	16 ± 3	10 ± 1	28 ± 6
100	115 ± 10	19 ± 3	16 ± 3	8 ± 3	27 ± 1
500	64 ± 13	13 ± 3	12 ± 2	5 ± 2	24 ± 4
2500	0	0	0	0	24 ± 3
5000	0	0	0	0	13 ± 3
MNNG (5)	684 ± 145		602 ± 27
NOPD (10)		926 ± 27
AAC (100)				818 ± 81
4-NQO (5)					991 ± 61
	Results with S9 (1:9)
DMSO	133 ± 3	34 ± 1	20 ± 3	11 ± 2	37 ± 4
20	142 ± 16	20 ± 3	16 ± 1	8 ± 2	37 ± 3
100	117 ± 36	17 ± 3	16 ± 3	9 ± 1	35 ± 4
500	23 ± 2	16 ± 2	9 ± 4	6 ± 3	28 ± 2
2500	0	0	9 ± 2	0	27 ± 8
5000	0	0	0	0	16 ± 3
2- AA (2.5)	1087 ± 90	1038 ± 23	168 ± 21	145 ± 14	241 ± 11

Experiment 3 (standard plate test)

Dose (µg/plate)	Mean number of revertant colonies of 3 replicates (± S.D.) for different strains of S. typhimurium and E. coli
	TA 100	TA 98	TA 1535	TA 1537	WP2 uvrA
	Results without S9
DMSO	110 ± 9	27 ± 3	20 ± 3	12 ± 3	25 ± 1
0.8	97 ± 6	27 ± 2	15 ± 3	9 ± 2	-
4	90 ± 9	22 ± 2	16 ± 3	10 ± 2	23 ± 4
20	99 ± 6	17 ± 4	16 ± 3	8 ± 2	22 ± 4
100	52 ± 7	10 ± 1	12 ± 2	3 ± 2	28 ± 3
500	6 ± 3	8 ± 2	11 ± 2	1 ± 0	21 ± 2
1000	-	-	-	-	22 ± 3
MNNG (5)	697 ± 72		842 ± 89
NOPD (10)		799 ± 60
AAC (100)				845 ± 41
4-NQO (5)					822 ± 101
	Results with S9 (1:9)
DMSO	112 ± 10	36 ± 3	19 ± 3	12 ± 2	34 ± 3
0.8	98 ± 8	39 ± 2	16 ± 3	10 ± 1	-
4	90 ± 2	28 ± 2	16 ± 4	9 ± 1	31 ± 2
20	77 ± 3	36 ± 2	13 ± 2	9 ± 2	30 ± 1
100	32 ± 19	24 ± 4	8 ± 3	6 ± 1	31 ± 3
500	8 ± 4	13 ± 2	7 ± 2	2 ± 2	30 ± 3
1000	-	-	-	-	26 ± 4
2- AA (2.5)	1100 ± 59	922 ± 15	202 ± 22	113 ± 10	22 ± 19

Experiment 4 (Preincubation test)

Dose (µg/plate)	Mean number of revertant colonies of 3 replicates (± S.D.) for different strains of S. typhimurium
	TA 100	TA 98	TA 1535	TA 1537
	Results without S9
DMSO	112 ± 7	23 ± 4	18 ± 2	9 ± 2
3.125	108 ± 14	21 ± 3	14 ± 5	6 ± 3
6.25	100 ± 9	23 ± 6	15 ± 2	8 ± 1
12.5	101 ± 14	20 ± 2	17 ± 2	8 ± 2
25	89 ± 14	20 ± 3	15 ± 1	8 ± 2
50	90 ± 8	14 ± 2	5 ± 1	0
MNNG (5)	1177 ± 22		1027 ± 43
NOPD (10)		1158 ± 40
AAC (100)				717 ± 11
	Results with S9 (1:9)
DMSO	109 ± 8	32 ± 8	17 ± 2	8 ± 2
3.125	117 ± 18	30 ± 2	16 ± 4	11 ± 2
6.25	105 ± 16	30 ± 2	14 ± 4	10 ± 2
12.5	113 ± 10	33 ± 3	15 ± 1	9 ± 2
25	116 ± 35	28 ± 5	9 ± 4	7 ± 1
50	94 ± 7	30 ± 4	14 ± 3	8 ± 2
2- AA (2.5)	644 ± 19	687 ± 38	138 ± 20	106 ± 11

Dose (µg/plate)	Mean number of revertant colonies of 3 replicates (± S.D.) for E. coli
	WP2 uvrA
	Results without S9
DMSO	24 ± 4
4	24 ± 5
20	18 ± 3
100	19 ± 3
500	18 ± 7
1000	16 ± 2
4-NQO (5)	746 ± 39
	Results with S9 (1:9)
DMSO	33 ± 4
4	27 ± 4
20	24 ± 11
100	20 ± 5
500	16 ± 7
1000	16 ± 1
2- AA (2.5)	221 ± 27

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative