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Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

There are no in vitro studies available investigating genotoxicity of butane-1,3 -diol. An Ames Assay with the structurally related read-across substance butane-1,4 -diol revealed negative results, both in the presence and absence of metabolic activation.

Link to relevant study records

Referenceopen allclose all

Endpoint:
in vitro gene mutation study in bacteria
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Justification for type of information:
For details on endpoint specific justification please see read-across report in section 13 or find a link in cross-reference “assessment report”.
Reason / purpose:
read-across source
Reason / purpose:
read-across: supporting information
Key result
Species / strain:
S. typhimurium TA 1535
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
not specified
Remarks:
negative in range finding test with same top dose
Vehicle controls validity:
valid
Untreated negative controls validity:
not examined
Positive controls validity:
valid
Key result
Species / strain:
S. typhimurium TA 1537
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
not specified
Remarks:
negative in range finding test with same top dose
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Key result
Species / strain:
S. typhimurium TA 98
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
not specified
Remarks:
negative in range finding test with same top dose
Key result
Species / strain:
S. typhimurium TA 100
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
not specified
Remarks:
negative in range finding test with same top dose
Vehicle controls validity:
valid
Untreated negative controls validity:
not examined
Positive controls validity:
valid
Key result
Species / strain:
E. coli WP2 uvr A
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
not specified
Remarks:
negative in range finding test with same top dose
Vehicle controls validity:
valid
Untreated negative controls validity:
not examined
Positive controls validity:
valid
Remarks on result:
other:
Remarks:
Without the addition of S9 mix, the result in Test I for the 625 μg/plate showed an increase in revertant colony count which was more than twice that of the solvent control value. However, in the testing of TA1537 with the addition of S9 mix and in the tests with all the other bacteria, there was no increase in revertant colony count more than twice that of the solvent control. The main test was repeated using identical doses. As a result, no increase in revertant colony count more than twice that of the solvent control was noted.
Conclusions:
Under the conditions of this study the test substance was not mutagenic to bacteria with and without metabolic activation.
Executive summary:

The study used as source investigated butane-1,4-diol.The study results of the source compound were considered applicable to the target compound and were used for classification and labelling acc. to REGULATION (EC) No 1272/2008. Justification and applicability of the read-across approach (structural analogue) is outlined in the read-across report in section 13 or find a link in cross reference “assessment report”.

Endpoint:
in vitro gene mutation study in bacteria
Type of information:
experimental study
Adequacy of study:
key study
Study period:
not stated
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Version / remarks:
not stated
Qualifier:
according to
Guideline:
OECD Guideline 472 (Genetic Toxicology: Escherichia coli, Reverse Mutation Assay)
Version / remarks:
not stated
Qualifier:
according to
Guideline:
other: “Methods of Testing New Chemical Substances” (Environmental Protection Bureau notification no. 237, Pharmaceutical Affairs Bureau notification no. 306, Basic Industries Bureau 1987 notification no. 303, dated 31 March, 1987
Version / remarks:
31 March 1987
GLP compliance:
yes
Type of assay:
bacterial reverse mutation assay
Specific details on test material used for the study:
Purity of test material: 99.8 wt% (impurities: 0.06 wt% 1,4-acetoxyhydroxybutane-2, 0.07 wt% 2-(4-hydroxy-butyloxy)tetrahydrofuran)
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Additional strain / cell type characteristics:
not specified
Species / strain / cell type:
E. coli WP2 uvr A
Additional strain / cell type characteristics:
not specified
Metabolic activation:
with and without
Metabolic activation system:
S9 (Kikkoman Co., Ltd., lot no: RAA-333, manufactured 8 September, 1995 and lot no: RAA-338, 15 December, 1995) extracted from oxygen-induced 7-week old Sprague-Dawley male rats administered phenobarbital (PB) and 5,6-benzoflavone (BF
Test concentrations with justification for top dose:
313~5000 μg/plate with a common ratio of 2, top dose according to guideline
Vehicle / solvent:
water for injection use
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: AF2: 2-(2-furyl)-3-(5-nitro-2-furyl)acrylamide; SA: Sodium azide; 9AA: 9-aminoacridine; 2AA: 2-aminoanthracene
Details on test system and experimental conditions:
METHOD OF APPLICATION: in agar (plate incorporation); preincubation

DURATION
- Preincubation period: 20 min at 37 °C
- Exposure duration: 48 h
- Fixation time (start of exposure up to fixation or harvest of cells): 48 h

NUMBER OF REPLICATIONS:main test and repeatability test, 3 plates were used for both control groups and for each dose

DETERMINATION OF CYTOTOXICITY
not stated
Evaluation criteria:
If, for 1 or more of the 5 strains of bacteria used, under conditions without S9 mix and with S9 mix, the mean value of the revertant colony count on the surface of plates containing the test substance was 2 or more times that of the solvent control, and, if that increase could be repeated or dose-dependency was noted, then the test substance in this series of tests would be determined to be mutagenic (positive). However, if the dose showed that the mean value of the revertant colony count at the second time of testing was more than 2 times that of the solvent control, but the solvent control value was less than 10 and a dose-dependent increase in the revertant colony count was not noted, then the determination would be negative.
Statistics:
not performed
Key result
Species / strain:
S. typhimurium TA 1535
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
not specified
Remarks:
negative in range finding test with same top dose
Vehicle controls validity:
valid
Untreated negative controls validity:
not examined
Positive controls validity:
valid
Key result
Species / strain:
S. typhimurium TA 1537
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
not specified
Remarks:
negative in range finding test with same top dose
Vehicle controls validity:
valid
Untreated negative controls validity:
not examined
Positive controls validity:
valid
Key result
Species / strain:
S. typhimurium TA 98
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
not specified
Remarks:
negative in range finding test with same top dose
Vehicle controls validity:
valid
Untreated negative controls validity:
not examined
Positive controls validity:
valid
Key result
Species / strain:
S. typhimurium TA 100
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
not specified
Remarks:
negative in range finding test with same top dose
Vehicle controls validity:
valid
Untreated negative controls validity:
not examined
Positive controls validity:
valid
Key result
Species / strain:
E. coli WP2 uvr A
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
not specified
Remarks:
negative in range finding test with same top dose
Vehicle controls validity:
valid
Untreated negative controls validity:
not examined
Positive controls validity:
valid
Remarks on result:
other:
Remarks:
Without the addition of S9 mix, the result in Test I for the 625 μg/plate showed an increase in revertant colony count which was more than twice that of the solvent control value. However, in the testing of TA1537 with the addition of S9 mix and in the tests with all the other bacteria, there was no increase in revertant colony count more than twice that of the solvent control. The main test was repeated using identical doses. As a result, no increase in revertant colony count more than twice that of the solvent control was noted.
Conclusions:
Under the conditions of this study the test substance was not mutagenic to bacteria with and without metabolic activation.
Executive summary:

A bacterial mutagenicity test (according to OECD guidelines 471 and 472 and GLP-compliant) was performed in Salmonella typhimurium TA1535, TA1537, TA98 and TA100, and in Escherichia coli WP2 uvrA as preincubation test, with and without metabolic activation.

Tests with and without S9 mix were conducted twice, within the range of 313 -5000 μg/plate with a common ratio of 2.

For the test of TA1537, without the addition of S9 mix, the result in Test I for the 625 μg/plate showed an increase in revertant colony count which was more than twice that of the solvent control value. However, in the testing of TA1537 with the addition of S9 mix and in the tests with all the other bacteria, there was no increase in revertant colony count more than twice that of the solvent control.

For TA1537, since the results without S9 mix were different for Test I and Test II, the main test was repeated using identical doses. As a result, no increase in revertant colony count more than twice that of the solvent control was noted.

In all of the tests conducted, the positive control group showed an increase in revertant colony count for all of the test bacteria, which meant that, along with the values measured for the solvent control group, the revertant colony count was within the range of historical values, so the efficacy of this test series was confirmed.

It is concluded, based on the above results that the test material is not mutagenic (negative) for the test strains used.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (negative)

Genetic toxicity in vivo

Description of key information

Negative in an in vivo chromosome aberration test and dominant lethal test.

Based on these in vivo studies the substance is not considered to be genotoxic.

Link to relevant study records

Referenceopen allclose all

Endpoint:
in vivo mammalian germ cell study: cytogenicity / chromosome aberration
Remarks:
Type of genotoxicity: chromosome aberration
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1981
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: well performed study (cytogenetic study over 3 generations!) with some shortcomings in study conception and documentation
Reason / purpose:
reference to same study
Reason / purpose:
reference to same study
Reason / purpose:
reference to same study
Principles of method if other than guideline:
genotoxicity test in vivo after subchronic oral exposure over 3 generations
GLP compliance:
not specified
Type of assay:
chromosome aberration assay
Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Age at study initiation: 14-15 weeks
- Housing: individually
- Diet: ad libitum
- Water: ad libitum

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/- 2 °C
- Photoperiod: 12 h dark / 12 h light
Route of administration:
oral: feed
Details on exposure:
SEMIPURIFIED DIET
20% casein
8% refined corn oil
4% salt mix
1% vitamin mix
33.5% corn starch
33.5% dextrose

DIET PREPARATION
- test diets were prepared by substituting 1,3-butanediol for equal amounts by weight of corn starch and dextrose
Duration of treatment / exposure:
Rats were treated 4 weeks before the mating period. Female rats of the F0 were fed diets containing 1,3-butanediol throughout the mating, gestation and lactating period of the F1A generation. Pubs of the F1A were reared to maturity. After 11 weeks of feeding, 25 males and 25 females from each dosage group of F1A animals were randomly selected and paired to produce the F2A generation and the F2A litter was mated to produce the F3A generation.
Frequency of treatment:
daily
Post exposure period:
none
Remarks:
Doses / Concentrations:
5, 10, 24%
Basis:
nominal in diet
No. of animals per sex per dose:
At least two animals per sex and dose from the F1A, F2A and F3A generation were examined.
Control animals:
yes, plain diet
Positive control(s):
none
Tissues and cell types examined:
femur bone marrow
Details of tissue and slide preparation:
DETAILS OF SLIDE PREPARATION:
Animals were treated with 1 mg/kg bw colchicine intraperitoneally 3-4 h prior to examination (exact time point of examination not stated). The bone marrow was washed with 5 ml of hank's balanced salt solution (HBSS). The isolated cells were washed with HBSS repeatedly, suspended in hypotonic fetal calf serum and incubated for 20 min at 37 °C. Fixation of the cells was performed in methanol/glacial acetic acid (3:1 mixture) overnight at 4 °C and stained on coverslips with 2% aceto-orcein.

METHOD OF ANALYSIS:
100-250 metaphase cells per dose group were examined for chromosomal aberrations at 900x magnification by phase-contrast microscopy.


OTHER:
Statistics:
Statistical analysis was not stated.
Key result
Sex:
male/female
Genotoxicity:
negative
Toxicity:
no effects
Remarks:
only slight depression of body weight gain
Vehicle controls validity:
valid
Negative controls validity:
not examined
Positive controls validity:
not examined
Additional information on results:
Dietary concentrations of 5, 10 and 24% correspond with body doses of 2000, 4000 and 9600 mg/kg bw for males and 2500, 5000 and 12000 mg/kg bw for females (based on a daily food consumption of 40 and 50 g/kg bw for males and females, respectively, according to the Guidance on Information Requirements R.8).

The number of abnormal cells was not increased with respect to the normal range of aberrant cells in untreated F1A, F2A and F3A animals. No specific abnormalities were observed in the treated animals and no dose-related effects were noted.

Conclusions:
The test substance did not induce chromosomal aberrations after subchronic oral exposure of rats over 3 generations with dietary concentrations of up to 24%.
Executive summary:

Rats were fed butane-1,3-diol in concentrations up to 24% of the diet and paired to produce F1A, F2A and F3A litters. Analysis of the femur bone marrow of at least two animals per sex and dose of these litters revealed no increase in chromosomal aberrations. This study was performed with doses high enough to cause a reduced body weight gain (Hess et al., 1981).

.

Endpoint:
in vivo mammalian germ cell study: cytogenicity / chromosome aberration
Remarks:
Type of genotoxicity: other: chromosome aberration, gene mutation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1981
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: well performed study with some shortcomings in documentation
Reason / purpose:
reference to same study
Reason / purpose:
reference to same study
Reason / purpose:
reference to same study
Principles of method if other than guideline:
genotoxicity test in vivo after subchronic oral exposure
GLP compliance:
not specified
Type of assay:
rodent dominant lethal assay
Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Age at study initiation: 14-15 weeks
- Housing: individually
- Diet: ad libitum
- Water: ad libitum

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/- 2 °C
- Photoperiod: 12 h dark / 12 h light
Route of administration:
oral: feed
Details on exposure:
SEMIPURIFIED DIET
20% casein
8% refined corn oil
4% salt mix
1% vitamin mix
33.5% corn starch
33.5% dextrose

DIET PREPARATION
- test diets were prepared by substituting 1,3-butanediol for equal amounts by weight of corn starch and dextrose
Duration of treatment / exposure:
Rats were treated 4 weeks before the mating period. Female rats of the F0 were fed diets containing 1,3-butanediol throughout the mating, gestation and lactating period of the F1A generation. At 1-2 weeks after weaning of the F1A litter, F0 females were mated with different males and the F1B generation was produced. Ten males per dose group were used for the dominant lethal test. They were housed individually in mating cages and fed the same diet concentrations as the F0 generation. For 8 consecutive weeks, 2 virgin females (100 days old) were introduced each week and remained for 7 days. Afterwards the females were kept individually for another 7 days and then examined.
Frequency of treatment:
daily
Post exposure period:
none
Remarks:
Doses / Concentrations:
5, 10, 24%
Basis:
nominal in diet
No. of animals per sex per dose:
ten males of the F1B generation
Control animals:
yes, plain diet
Positive control(s):
none
Tissues and cell types examined:
The reproductive tract of the mated females was examined with respect to the number of implantates, resorption sites, viable and dead fetuses
Evaluation criteria:
The mutagenic index (% resorptions/implantation sites) was calculated according to a method of Epstein and Schaffer (reference stated).
Statistics:
Statistical analysis was performed, but not stated in detail.
Key result
Sex:
male/female
Genotoxicity:
negative
Toxicity:
no effects
Remarks:
only slight depression of body weight gain
Vehicle controls validity:
valid
Negative controls validity:
not examined
Positive controls validity:
not examined
Additional information on results:
Dietary concentrations of 5, 10 and 24% correspond with body doses of 2000, 4000 and 9600 mg/kg bw for males and 2500, 5000 and 12000 mg/kg bw for females (based on a daily food consumption of 40 and 50 g/kg bw for males and females, respectively, according to the Guidance on Information Requirements R.8).

The percentage of pregnancies as well as the percentage of viable fetuses per implantation site were not significantly different between treatment and control groups. The mutagenic index did not show a trend with increasing doses.

  control  5%  10%  24%
No. pregnancies total  106  97  130  117 
% Pregnancies (20 matings)  66.3  60.6  81.3  73.1 
Implant sites  1165  1024  1452  1310 
Viable fetuses total  1101  962  1389  1269 
% Viable fetuses/implant sites  94.5  94.0  95.7  96.9 
Resorptions total  64  62  63  41 
% Resorptions/implant sites*  5.5  6.1  4.3  3.1 

*:mutagenic index

Conclusions:
The test substance did not induce dominant lethal effects after subchronic oral exposure of rats with dietary concentrations of up to 24%.
Executive summary:

Rats were fed butane-1,3-diol in concentrations up to 24% of the diet and paired to produce F1A and F1B litters. Males of the F1B generation were used to examine dominant lethal effects after mating them with virgin females. The exposure did not cause a significant effect with respect to fertility, viable fetuses per implantation sites and percentage of resorption per implantation sites (mutagenic index). A dose-related trend was not evident. This study was performed at high doses, which produced reduced body weight gain (Hess et al., 1981).

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (negative)

Mode of Action Analysis / Human Relevance Framework

In the absence of information on species specific effects or metabolism the results are regarded as relevant for humans.

Additional information

There are no studies available investigating genotoxicity of butane-1,3 -diol in vitro. However, in a reliable Ames Assay with the structurally closely related substance butane-1,4 -diol negative effects were reported, both in the presence and absence of metabolic activation. Additionally, negative results were obtained with butane-1,3 -diol in two in vivo genotoxicity assays, a chromosome aberration test and a dominant lethal test (Hess et al., 1981). Both test systems cover chromosomal aberrations and the latter gene mutational effects as well. Based on these data it is concluded that butane-1,3 -diol is not genotoxic.

Further, butane-1,3 -diol is rapidly metabolised to gamma-hydroxybutyrate (Mehlmann et al., 1971; Tate et al., 1971; Tobin et al., 1978), which is an endogenous product of mammalian metabolism and not considered to be genotoxic. Negative results in two carcinogenicity studies in rats and dogs with butane-1,3 -diol further support the conclusion that butane-1,3 -diol is not genotoxic.





Justification for classification or non-classification

Based on the negative effects (no genotoxicity observed) in in vivo assays with butane-1,3 -diol and an Ames Assay with the structurally related read-across substance butane-1,4 -diol no classification for mutagenicity is required according to Regulation (EC) No 1272/2008.