1-[(2-butyloctyl)oxy]-3-(3,4-dihydroisoquinolinium-2-yl)propan-2-yl sulfate

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:

from 2006-02-13 to 2006-05-18

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP guideline study

Cross-reference

Reason / purpose for cross-reference:

reference to other study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2006

Report date:

2006

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Version / remarks:

21 July 1997

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

2000/32/EC

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.5100 - Bacterial Reverse Mutation Test (August 1998)

Version / remarks:

August 1998

Deviations:

no

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Reference

Name:

Unnamed

Type:

Constituent

Details on test material:

Sokalan PG B62

Method

Target gene:

His (S. typhimurium)
Trp (E. coli)

Species / strain

Species / strain / cell type:

other: Salmonella typhimurium TA1535, TA100, TA1537, TA98; Escherichia coli WP2 uvrA

Additional strain / cell type characteristics:

not specified

Metabolic activation:

with and without

Metabolic activation system:

S9-Mix from Aroclor 1254 induced rat livers

Test concentrations with justification for top dose:

Standard plate test: 56, 177, 557, 1750, 5500 µg/plate
Preincubation test: 56, 177, 557, 1750, 5500 µg/plate

Vehicle / solvent:

Substances were dissolved in DMSO.

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: With S-9 mix: 2-AA; Without S9-MIx: MNNG, NOPD, AAc, 4-NQO

Details on test system and experimental conditions:

Positive controls
with S-9 mix
- 2-aminoanthracene (2-AA) (SIGMA, A-1381)
2.5 µg/plate, dissolved in DMSO; strains: TA 1535, TA 100, TA 1537, TA 98
60 µg/plate, dissolved in DMSO; strain : Escherichia coli WP2 uvrA
without S-9 mi x
- N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) (FLU144, 68051)
5 µg/plate, dissolved in DMSO
strains : TA 1535, TA 100
- 4-nitro-o-phenylendiamine ( NOPD) (SIGMA, N-9504)
10 µg/plate, dissolved in DMSO
strain: TA 98
- 9-aminoacridine (AAC) (SIGMA, A-1135)
100 µg/plate, dissolved in DMSO
strain: TA 1537
- 4-nitroquinoline-N-oxide (4-NQO) (SIGMA, N-8141)
5 µg/plate, dissolved in DMSO
strain : E. coli WP2 uvrA

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

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

NUMBER OF REPLICATIONS: 3 plates

DETERMINATION OF CYTOTOXICITY
reduction of his- background growth, decrease in the number of his+ revertants

Evaluation criteria:

Generally, the experiment is 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 or above.
The titer of viable bacteria was > 10E08/mL.

Statistics:

no data on statistics

Results and discussion

Test results

Species / strain:

other: Salmonella typhimurium TA1535, TA100, TA1537, TA98; Escherichia coli WP2 uvrA

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

other: A slight decrease in the number of revertants was occasionally observed depending on the strain and test conditions from about 1750 µg/plate onward.

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

An increase in the number of his+ or trp+ revertants was not observed in the standard plate test or in the preincubation test either without S-9 mix or after the addition of a metabolizing system. Precipitation of the test substance was found from about 557 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

Table 1: Number of revertants in the a. standard plate test and b. preincubation test after treatment with control or test substance in the various tester strains.

a.

Strain	control		test substance
	-S9	+S9	-S9	+S9
TA1535	16 ± 1	16 ± 2	16 ± 2 (177 µg)	17 ± 1 (177 µg)
TA100	114 ± 11	123 ± 20	117 ± 9 (56 µg)	144 ± 19 (177 µg)
TA1537	9 ± 2	10 ± 1	8 ± 1 (56 µg)	9 ± 1 (177 µg)
TA98	28 ± 1	41 ± 4	29 ± 2 (56 µg)	39 ± 3 (177 µg)
E.coli wp2 uvra	34 ± 3	36 ± 4	29 ± 4 (177 µg)	52 ± 9 (5500 µg)

b.

Strain	control		test substance
	-S9	+S9	-S9	+S9
TA1535	16 ± 3	17 ± 3	19 ± 2 (1750 µg)	15 ± 2 (557 µg)
TA100	111 ± 7	111 ± 8	115 ± 6 (177 µg)	121 ± 9 (56 µg)
TA1537	10 ± 3	9 ± 2	9 ± 3 (56 µg)	10 ± 2 (557 µg)
TA98	34 ± 5	33 ± 2	32 ± 4 (56 µg)	40 ± 4 (177 µg)
E.coli wp2 uvra	41 ± 4	36 ± 4	54 ± 7 (5500 µg)	35 ± 5 (5500 µg)

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative

The test substance 1-((2-Butyloctyloxymethyl)-2-(3,4-dihydro-isoquinolinium-2-yl)ethyl)sulfate showed no mutagenic effects in a reverse mutation test (Ames test) in bacteria with and without metabolic activation.

Executive summary:

In a GLP conform Ames test according to OECD guideline 471, the substance 1-((2-Butyloctyloxymethyl)-2-(3,4-dihydro-isoquinolinium-2-yl)ethyl)sulfate (purity: 93.0 weight-%) was tested for its mutagenic potential based on the ability to induce point mutations in selected loci of several bacterial strains, i.e. Salmonella typhimurium TA 1535, TA 100, TA 1537, TA 98 and Escherichia coli WP2 uvrA (BASF 2006). Two independent assays were performed, a standard plate test (SPT) and a preincubation test (PIT), both with a dose range of 56 µg - 5500 µg/plate, and both with and without metabolic activation (Aroclor 1254-induced rat liver S-9 mix).

An increase in the number of his+ or trp+ revertants was not observed in the standard plate test or in the preincubation test either without S-9 mix or after the addition of a metabolizing system. Precipitation of the test substance was found from about 557 mg/plate onward. A slight decrease in the number of revertants was occasionally observed depending on the strain and test conditions at doses >= 1750 µg/plate.

According to the results of the present study, the test substance is not mutagenic in the Salmonella typhimurium/ Escherichia coli reverse mutation assay under the experimental conditions chosen here.