Registration Dossier

Administrative data

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

The substance Isophorone diamine was tested in the Ames Salmonella mutagenicity test for any mutagenic activity according to OECD 471 from 1981. Under the conditions of the study the test item isophorone diamine proved to be non-mutagenic, both in the presence and in the absence of Arochlor-induced liver microsomes, for all test strains used in this study (Hüls AG 1990).

Chinese hamster ovary cells (CHO), treated with Isophorone diamine, were evaluated for chromosome aberrations at three dose levels, in duplicate, together with negative and positive controls. Isophorone diamine is considered to be non-clastogenic to CHO cells (SafePharm 1992).

In this HPRT test with Chinese Hamster ovary (CHO) cells according to OECD TG 476 (1984), Isophorone diamine did not significantly increase the mutant frequency of treated cells. Cytotoxicity was not observed. It is concluded that Isophoron diamine, in the presence as well as in the absence of S9 mix, demonstrates no mutagenic potential in this in vitro mammalian cell assay (Hüls AG 1992).

Link to relevant study records

Referenceopen allclose all

Endpoint:
in vitro gene mutation study in mammalian cells
Remarks:
Type of genotoxicity: gene mutation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1992-01-20 - 1992-03-02
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Guideline study
Qualifier:
according to
Guideline:
OECD Guideline 476 (In Vitro Mammalian Cell Gene Mutation Test)
Version / remarks:
1984
Deviations:
no
GLP compliance:
yes
Type of assay:
mammalian cell gene mutation assay
Target gene:
hypoxanthine-guanine phosphoribosyl-transferase (HPRT) locus
Species / strain / cell type:
Chinese hamster Ovary (CHO)
Details on mammalian cell type (if applicable):
CHO (Chinese hamster ovary) K1 cells
Metabolic activation:
with and without
Metabolic activation system:
Aroclor 1254-induced Wistar rat liver S9
Test concentrations with justification for top dose:
0 - 2 mg/ml
Vehicle / solvent:
HO medium, dimethyl sulfoxide (DMSO)
Untreated negative controls:
yes
Remarks:
= solvent/ vehicle control
Negative solvent / vehicle controls:
yes
Remarks:
Medium
True negative controls:
no
Positive controls:
yes
Positive control substance:
ethylmethanesulphonate
Remarks:
positive control with metabolic activation: 3-Methylcholanthrene Migrated to IUCLID6: without metabolic activation
Details on test system and experimental conditions:
HGPRT assay. ADMINISTRATION:
- Dosing:
preliminary toxicity test:
0; 0.02; 0.03; 0.06; 0.12; 0.2; 0.3; 0.6; 1.2; 2.0 mg/ml
main study; 0; 0.02; 0.06; 0.2; 0.6; 2.0 mg/ml
- Number of replicates: 2
- Application: 2E+05 cells/25 cm2 flask; exposure time 4 h
- Positive and negative control groups and treatment:
positive, without metabolic activation:
300 ug ethyl methanesulfonate (EMS)/ml in HO medium
positive, with metabolic activation:
10 ug 3-methylcholanthrene (MCA)/ml in dimethyl sulfoxide
negative: HO medium (with / without S9 mix)
Evaluation criteria:
statistically significant, dose related increase in mutant frequency at concentrations of the test substance resulting in > 20 % cell survival. Mean
frequency > maximum spontaneous frequency
Statistics:
t-test
Species / strain:
Chinese hamster Ovary (CHO)
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Remarks:
none over test concentration range
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Remarks on result:
other: strain/cell type: CHO (Chinese hamster ovary) K1 cells

no further remarks

Conclusions:
In this HPRT test with Chinese Hamster ovary (CHO) cells according to OECD TG 476 (1984), Isophorone diamine concentrations of
20 - 2,000 mg/l (+/- S9 mix from Aroclor 1254 induced rat livers) did not significantly increase the mutant frequency of treated cells. Cytotoxicity was not observed. It is concluded that Isophoron diamine, in the presence as well as in the absence of S9 mix, demonstrates no mutagenic potential in this in vitro mammalian cell assay at any of the concentrations tested.
Executive summary:

The test item Isophorone diamine was tested for its ability to induce forward mutation at the hypoxanthine-guanine phosphoribosyl-transferase (HPRT) locus in Chinese hamster ovary (CHO) cells in vitro both in the presence and the absence of exogenous metabolic activation(by Arochlor 1254 -induced rat liver S9). The substance was tested in concentrations of 20 mg/l to the solubility limit of 2000 mg/l with and without S9 mix. In this concentration range, Isophorone diamine did not induce any detectable cytotoxicity. Compared to the negative control treatment with Isophorone diamine in both assays (with and without metabolic activation) did not result in any reproducible statistically or biologically significant increase of the mutation frequency of the HPRT locus. It is concluded that Isophoron diamine, in the presence as well as in the absence of S9 mix, demonstrates no mutagenic potential in this in vitro mammalian cell assay.

Endpoint:
in vitro gene mutation study in bacteria
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1990-02-22 - 1990-03-19
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
Guideline study, comparable to OECD guideline 471 with acceptable restrictions (TA 102 or E.coli WP2 were not tested, not required by applied 1983 guideline version)
Qualifier:
according to
Guideline:
EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)
Version / remarks:
Cited as Directive 84/449/EEC, B.14
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Version / remarks:
1983
Deviations:
yes
Remarks:
TA 102 or E.coli WP2 were not tested, not required by applied 1983 guideline version
GLP compliance:
yes
Type of assay:
bacterial reverse mutation assay
Target gene:
mutated gene loci responsible for histidine auxotrophy
Species / strain / cell type:
S. typhimurium, other: TA 98, TA 100, TA 1535, TA 1537, TA 1538
Additional strain / cell type characteristics:
other: histidine auxotroph
Metabolic activation:
with and without
Metabolic activation system:
Aroclor 1254 induced rat S9 liver, male Bor: WISW (SPF/Cpb)
Test concentrations with justification for top dose:
8 - 5000 µg/plate
Vehicle / solvent:
water / dimethyl sulfoxide
Untreated negative controls:
yes
Remarks:
=Solvent/ vehicle control
Negative solvent / vehicle controls:
yes
Remarks:
DMSO; water
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: without metabolic activation: Nitrofluorene (TA 98; TA 1538), Sodium azide (TA 100; TA 1535), Aminoacridine (TA 1537); with metabolic activation: Aminoanthracene (TA 100)
Details on test system and experimental conditions:
Ames test. ADMINISTRATION:
- Dosing:
main test: 8 / 40 / 200 / 1000 / 5000 µg/plate (+/- metabolic activation)
preincubation test: 125/250/500/1000/2000 µg/plate (+/- metabolic activation)
- Number of replicates: 3
- Application:
main test: 50 g/l in water
preincubation test: 40 g/l in water
- Positive and negative control groups and treatment:
positive, TA 98 and TA 1538: nitrofluorene
positive, TA 100 and TA 1535: sodium azide
positive, TA 1537: aminoacridine
negative: solvent
activity of metabolic system: aminoanthracene / TA 100
- Pre-incubation time: 30 min / 30 °C
incubation 96 hours / 37 °C
Evaluation criteria:
mutagenic effects (i.e ratio of revertant rates treated/control >= 2) at <= 5000 µg/plate with generally positive dose-response relationship in any
strain
Statistics:
common mathematical operations (e.g. arithmetic mean / standard deviation)
Species / strain:
S. typhimurium, other: TA 98, TA 100, TA 1535, TA 1537, TA 1538
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
other: 1000 / 5000 µg/plate with / without preincubation
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
PRECIPITATION CONCENTRATION: no precipitation
Remarks on result:
other: other: Salmonella typhimurium

no further remarks

Conclusions:
Under the conditions of the study the test item Isophorone diamine proved to be non-mutagenic in the Ames Salmonella mutagenicity test, both in the presence and in the absence of Arochlor-induced liver microsomes, for all test strains used in this study.
Executive summary:

The substance Isophorone diamine was tested in the Ames Salmonella mutagenicity test for any mutagenic activity. The test organisms were five histidine-auxotrophic Salmonella typhimurium strains (TA 1535; TA 1537; TA 1538; TA 98 and TA 100). Under the conditions of the study the test item isophorone diamine proved to be non-mutagenic, both in the presence and in the absence of Arochlor-induced liver microsomes, for all test strains used in this study.

Endpoint:
in vitro cytogenicity / chromosome aberration study in mammalian cells
Remarks:
Type of genotoxicity: chromosome aberration
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1992-05-26 - 1992-09-11
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Guideline study
Qualifier:
according to
Guideline:
OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)
Version / remarks:
1981
Deviations:
no
GLP compliance:
yes
Type of assay:
in vitro mammalian chromosome aberration test
Target gene:
mammalian cell system (CHO Chinese hamster ovary cell line)
Species / strain / cell type:
Chinese hamster Ovary (CHO)
Details on mammalian cell type (if applicable):
CHO-K1 BH4
cell cycle length of 12 hours
Metabolic activation:
with and without
Metabolic activation system:
male Sprague-Dawley rat liver S9 from Aroclor 1254 induced animals
Test concentrations with justification for top dose:
up to 1375 ug/ml in the main experiments
Vehicle / solvent:
Ham´s F12 medium
Untreated negative controls:
yes
Remarks:
= Solvent/ vehicle control
Negative solvent / vehicle controls:
yes
Remarks:
Ham´s F12 medium
True negative controls:
no
Positive controls:
yes
Positive control substance:
mitomycin C
Remarks:
positive control with S9-mix: Cyclophosphamide Migrated to IUCLID6: without metabolic activation
Details on test system and experimental conditions:
SYSTEM OF TESTING
- No. of metaphases analyzed: first 100 consecutive from each culture if possible
ADMINISTRATION:
- Dosing:
preliminary toxicity test: 0-5000 mg/l
Experiment 1:
312.5-937.5 mg/l without S9; 312.5-1250 mg/l with S9
Experiment 2:
156.25-625 mg/l without S9; 625-1250 mg/l with S9
Confirmatory experiment:
1125, 1250, and 1375 mg/l with S9, both with and without Hepes buffer
- Number of replicates: 2
- Application:
with S9: 4 hours exposure + 8 or 16 h culture period
without S9: continuous for 12 or 20 hours
- Positive and negative control groups and treatment:
negative: solvent
positive with S9: cyclophosphamide, 10 and 5 mg/l
positive without S9: mitomycin C, 0.075 and 0.05 mg/l
concentrations reduced for second experiment
Evaluation criteria:
significant increase in the frequency of aberrations relative to the vehicle control
Statistics:
Fisher's exact test
Species / strain:
Chinese hamster Ovary (CHO)
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
other: >= 2500 ug/ml (+ S9); >= 1250 ug/ml (-S9)
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
GENOTOXIC EFFECTS:
- With metabolic activation: highly statistically significant, but only at maximum dose level and 20 hour treatment. This was demonstrated to be an
artefact caused by the interaction of a high pH value and the S9 metabolic activation system.
- Without metabolic activation: no
- Positive controls: positive
MITOTIC INDEX: mean of 2 counts each; experiment 1 first
- With metabolic activation:
0 mg/l, 12 h: 7.15; 20 h: 9.5
312.5 mg/l, 12 h: 7.8; 20 h: 9.25
625 mg/l, 12 h:10.75; 20 h:10.65
1250 mg/l, 12 h: 4.8; 20 h: 6.15
0 mg/l, 12 h: 9.15; 20 h: 8.8
625 mg/l, 12 h: 4.3; 20 h:11.5
937.5 mg/l, 12 h:10.0; 20 h: 7.35
1250 mg/l, 12 h: 3.4; 20 h: 2.85
- Without metabolic activation:
0 mg/l, 12 h: 7.45; 20 h: 3.8
312.5 mg/l, 12 h: 8.65; 20 h: 3.75
625 mg/l, 12 h: 6.45; 20 h: 4.75
937.5 mg/l, 12 h: 2.0; 20 h: 2.3
0 mg/l, 12 h: 9.55; 20 h: 9.8
156.25mg/l, 12 h: 8.35; 20 h:12.3
312.5 mg/l, 12 h: 7.6; 20 h: 8.7
625 mg/l, 12 h: 6.9; 20 h: 6.8
CHROMOSOMAL ABERRATIONS (cells with aberrations - gaps):
mean of 2 replicates each
- With metabolic activation:
Experiment 1
0 mg/l, 12 h: 0; 20 h: 1
312.5 mg/l, 12 h: 2; 20 h: 0
625 mg/l, 12 h: 2; 20 h: 1
1250 mg/l, 12 h: 4; 20 h: 14
Experiment 2
0 mg/l, 12 h: 1; 20 h: 1
625 mg/l, 12 h: 2; 20 h: 4
937.5 mg/l, 12 h: 3; 20 h: 4
1250 mg/l, 12 h: 2; 20 h: 21
Confirmatory experiment
0 mg/l, 20 h: 4 (with hepes buffer)
0 mg/l, 20 h: 7 (without hepes buffer)
1250 mg/l, 20 h: 6 (with hepes buffer)
1250 mg/l, 20 h: 13 (without hepes buffer)
- Without metabolic activation:
Experiment 1
0 mg/l, 12 h: 2; 20 h: 0
312.5 mg/l, 12 h: 1; 20 h: 4
625 mg/l, 12 h: 1; 20 h: 6
937.5 mg/l, 12 h: 5; 20 h: 1
Experiment 2
0 mg/l, 12 h: 3; 20 h: 3
156.25mg/l, 12 h: 2; 20 h: 2
312.5 mg/l, 12 h: 3; 20 h: 3
625 mg/l, 12 h: 3; 20 h: 6
CYTOTOXIC CONCENTRATION:
- With metabolic activation:
total absence of metaphase cells at >= 2500 mg/l
- Without metabolic activation:
total absence of metaphase cells at >= 1250 mg/l
- A dose-related increase was observed. Since addition of a buffer in the confirmatory experiment reduced toxicity, pH seems to be decisive for cytotoxicity
Remarks on result:
other: strain/cell type: CHO (Chinese hamster ovary) cells

no further remarks

Conclusions:
The test substance Isophorone diamine is considered to be non-clastogenic to CHO cells in vitro under conditions of this study.
Executive summary:

Chinese hamster ovary cells (CHO), treated with Isophorone diamine, were evaluated for chromosome aberrations at three dose levels, in duplicate, together with negative and positive controls. Three treatment conditions were used. Concentrations of 156.25 – 1,375 mg/l (+/-S9 mix from Aroclor 1254-induced rat livers) were employed. The negative controls gave frequencies of aberrations within the range expected for the CHO cell line. Both of the positive control chemicals gave significant increases in the frequency of aberrations indicating the satisfactory performance of the test and of the activity of the metabolsing system. Isophorone diamine demonstrated a highly statistically significant increase in the frequency of cells with aberrations at the maximum dose level in the 20 -hour with matabolic activation treatment only. However this was demonstrated to be an artefact caused by the interaction of a high pH value and the S9 metabolic activation system. Isophorone diamine is considered to be non-clastogenic to CHO

cells in vitro under conditions of this study.

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

Genetic toxicity in vivo

Description of key information

In this micronucleus assay with NMRI mice according to OECD TG 474, 5 male/female animals per group were orally administered single doses of 50, 150, or 500 mg/kg bw isophorone diamine. The highest dose was considered the maximum tolerable dose (MTD) based on the induction of toxic effects without major effects on survival within 72 hours of test substance administration. Isophorone diamine treatment did not result in an increase in the number of micronucleated polychromatic erythrocytes (PCE), nor did it negatively affect the PCE/NCE ratio. Therefore, isophorone diamine is considered to be non-mutagenic under the conditions of this micronucleus assay (Cytotest Cell Research 1990).

Link to relevant study records
Reference
Endpoint:
in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus
Remarks:
Type of genotoxicity: chromosome aberration
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1990-04-17 - 1990-05-17
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Guideline study
Qualifier:
according to
Guideline:
OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)
Version / remarks:
1983
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.12 (Mutagenicity - In Vivo Mammalian Erythrocyte Micronucleus Test)
Version / remarks:
1984
Deviations:
no
GLP compliance:
yes
Type of assay:
micronucleus assay
Species:
mouse
Strain:
NMRI
Sex:
male/female
Details on test animals and environmental conditions:
TEST ORGANISMS:
- Supplier: BRL Tierfarm Füllinsdorf (Switzerland)
- Age: minimum 10 weeks + 5 days acclimatization
- Weight at study initiation: approximately 30 g
- Fasting period before study: 18 hours
- Housing: single
- Diet: ALTROMIN standard diet, ad libitum):
- Water: tap water ad libitum
- Acclimation period:
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 +/- 3 °C
- Humidity (%): 30 - 70 %
- Photoperiod (hrs dark / hrs light): 12h / 12h
Route of administration:
oral: unspecified
Vehicle:
aqua dest.
Details on exposure:
ADMINISTRATION:
- in a pre-experiment 500 mg/kg bw was estimated to be the maximum tolerated dose, cytotoxic reactions were not observed
- Sampling times and number of samples: 24, 48, or 72 hours after treatment
- Control groups and treatment:
negative: vehicle
positive: cyclophosphamide, dissolved in physiol. saline, 40 mg/kg bw
Duration of treatment / exposure:
single dose
Frequency of treatment:
single dose
Post exposure period:
24, 48 or 72 hours
Remarks:
Doses / Concentrations:
50, 150, or 500 mg/kg, dissolved in 10 ml/kg bw dose volume
Basis:
actual ingested
No. of animals per sex per dose:
6 per dosage group and sex with 3 cases of post-treatment duration for negative control and treated groups totals 4 x 3 + 1 groups x 6 animals x 2 sexes = 156 animals; only 5 animals per group and sex were evaluated
Control animals:
yes, concurrent vehicle
Positive control(s):
cyclophosphamide, dissolved in physiol. saline, 40 mg/kg bw
Tissues and cell types examined:
femora, bone marrow
Details of tissue and slide preparation:
- 1000 PCE (polychromatic erythrocytes) per animal were analysed for micronuclei
Evaluation criteria:
- Criteria for evaluating results:
either a statistically significant dose related increase in the number of micronucleated polychromatic erythrocytes,
or a reproducible statistically significant positive response for at least one of the test points
- Criteria for selection of M.T.D.: toxic reactions without major effects on survival within 72 hours
Statistics:
Mann-Whitney test
Sex:
male/female
Genotoxicity:
negative
Toxicity:
yes
Remarks:
Maximum tolerated dose determined in pre-experiment
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
EFFECT ON PCE/NCE RATIO: not affected (PCE = polychromatic erythrocytes, NCE = normochromatic erythrocytes)
GENOTOXIC EFFECTS:
For the positive control a significant and clear increase in the frequency of micronucleated polychromatic erythrocytes was observed.
The mean frequencies of micronucleated polychromatic erythrocytes in the groups treated with the test item were in the same range as those of the vehicle control group. A statistically significant increase was observed at the high dose (500 mg/kg bw) at 24 hours. However, the negative control in this case was untypically low (0.02 %). The absolute result for the dosed group (0.10 %) was well within the range of the 24 hour negative controls from the preceding 12 months (0.04 - 0.12; mean 0.073 %), i.e. there was no biological relevance.
--------------------------------------------------------
Treatment           Time   % Micron. in PCE    PCE/NCE
--------------------------------------------------------
 Vehicle            24 h     0.02 (0 - 2)    1000 / 970
  50 mg TS/kg bw    24 h     0.07 (0 - 2)    1000 / 917
 150 mg TS/kg bw    24 h     0.06 (0 - 2)    1000 / 892
 500 mg TS/kg bw    24 h     0.10 (0 - 3) *  1000 / 908
  40 mg CPA/kg bw   24 h     1.08 (2 -22) *  1000 /1127
 Vehicle            48 h     0.04 (0 - 1)    1000 / 699
  50 mg TS/kg bw    48 h     0.06 (0 - 2)    1000 / 676
 150 mg TS/kg bw    48 h     0.06 (0 - 3)    1000 / 702
 500 mg TS/kg bw    48 h     0.09 (0 - 3)    1000 / 781
 Vehicle            72 h     0.14 (0 - 5)    1000 / 744
  50 mg TS/kg bw    72 h     0.07 (0 - 3)    1000 / 681
 150 mg TS/kg bw    72 h     0.13 (0 - 4)    1000 / 754
 500 mg TS/kg bw    72 h     0.16 (0 - 4)    1000 / 758
--------------------------------------------------------
TS = test substance; CPA = cyclophosphamide; * p<0.05
Conclusions:
Therefore, the conclusion is drawn, that Isophorone diamine is not a mutagenic substance under the in vivo conditions in this micronucleus assay
using male and female NMRI mice.
Executive summary:

In this micronucleus assay with NMRI mice according to OECD TG 474, 5 male/female animals per group were orally administered single doses of 50, 150, or 500 mg/kg bw isophorone diamine. The highest dose was considered the maximum tolerable dose (MTD) based on the induction of toxic effects without major effects on survival within 72 hours of test substance administration. Sampling times were 24, 48, and 72 hrs after test substance administration. Isophorone diamine treatment did not result in an increase in the number of micronucleated polychromatic erythrocytes (PCE), nor did it negatively affect the PCE/NCE ratio. Therefore, isophorone diamine is considered to be non-mutagenic under the conditions of this micronucleus assay.

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

Additional information

Studies in animals

In vivo studies

Cited from SIAR for SIAM 18 (Paris, April 2004):

"In a micronucleus assay with NMRI mice according to OECD TG 474, 5 male/female animals per group were orally administered single doses of 50, 150, or 500 mg/kg bw 3-aminomethyl-3,5,5- trimethylcyclohexylamine. The highest dose was considered the maximum tolerable dose (MTD) based on the induction of toxic effects without major effects on survival within 72 hours of test substance administration. Sampling times were 24, 48, and 72 hrs after test substance administration. 3-Aminomethyl-3,5,5-trimethylcyclohexylamine treatment did not result in an increase in the number of micronucleated polychromatic erythrocytes (PCE), nor did it negatively affect the PCE/NCE ratio (Cytotest Cell Research, 1990). This result was confirmed in another study with a slightly different design (Hüls AG, 1988a). Here a single dose of 100 mg/kg bw was administered, which – based on the absence of cytotoxic effects – was determined as the MTD. Sampling times were 24, 48, and 72 hrs after test substance administration."

In vitro studies

Cited from SIAR for SIAM 18 (Paris, April 2004):

"In an Ames test performed according to Directive 84/449/EEC B.14 (1984) with Salmonella typhimurium TA 1535, TA 1537, TA 1538, TA 98 and TA 100, test substance concentrations of up to 5,000 μg/plate were employed in the presence and absence of Aroclor 1254-induced rat liver S9 mix. At non-toxic test substance concentrations, a significant increase in mutant frequency was not observed (Hüls AG, 1990). The same result was obtained in another test with comparable design (Hüls AG, 1988b; method according to the original publication by B. Ames, 1975).

In a HPRT test with Chinese Hamster ovary (CHO) cells according to OECD TG 476 (1984), 3- aminomethyl-3,5,5-trimethylcyclohexylamine concentrations of 20 - 2,000 mg/l (+/- S9 mix from Aroclor 1254 induced rat livers) did not significantly increase the mutant frequency of treated cells. Cytotoxicity was not observed at any of the concentrations tested (Hüls AG, 1992c). In a cytogenetic assay with Chinese Hamster Ovary (CHO) cells (according to OECD TG 473), concentrations of 156.25 – 1,375 mg/l (+/-S9 mix from Aroclor 1254-induced rat livers) were employed. A significant, test compound related increase in chromosomal aberrations was not observed (Safepharm, 1992)."


Justification for classification or non-classification

Because of the results of the in vitro and in vivo mutagenicity studies the substance isophorone diamine is not classified according to CLP regulation (1272/2008).