Amines, N-(3-aminopropyl)-N'-(C16-18 and C18-unsatd. alkyl)trimethylenedi-

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

Tallow dipropylenetriamine was not mutagenic in a bacterial mutagenicity study (Ames test), induced no chromosomal aberrations in a study in human lymphocytes, and was not mutagenic in a mammalian mutagenicity study in mouse lymphoma cells. All studies were performed under GLP according to current guidelines.

Link to relevant study records

Reference

Reference 1

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:

01-jul-2009 to 11-aug-2009

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: The study has been performed according to OECD and/or EC guidelines and according to GLP principles.

Qualifier:

according to guideline

Guideline:

OECD Guideline 476 (In Vitro Mammalian Cell Gene Mutation Test)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.17 (Mutagenicity - In Vitro Mammalian Cell Gene Mutation Test)

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: recommendations of the “International Workshop on Genotoxicity Tests Workgroup” (the IWGT), published in the literature (Clive et al., 1995, Moore et al., 1999, 2000, 2002, 2003, 2006 and 2007).

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of assay:

mammalian cell gene mutation assay

Target gene:

Thymidine kinase (TK) locus in L5178Y mouse lymphoma cells

Species / strain / cell type:

mouse lymphoma L5178Y cells

Details on mammalian cell type (if applicable):

- Type and identity of media:
-RPMI 1640 Hepes buffered medium (Dutch modification) containing penicillin/streptomycin (50 U/ml and 50 μg/ml, respectively), 1 mM sodium pyruvate and 2 mM L-glutamin supplemented with 10% (v/v) heat-inactivated horse serum (=R10 medium).
- Properly maintained: yes
- Periodically checked for Mycoplasma contamination: yes
- Periodically checked for karyotype stability: no
- Periodically "cleansed" against high spontaneous background: yes

Metabolic activation:

with and without

Metabolic activation system:

Rat liver S9-mix induced by a combination of phenobarbital and ß-naphthoflavone

Test concentrations with justification for top dose:

Dose range finding test:
Without and with S9-mix, 3 hours treatment: 0.3, 1, 3, 10, 33 and 100 µg/ml
Without S9-mix, 24 hours treatment: 0.08, 0.24, 0.8, 2.4, 8 and 26 µg/ml
Experiment 1:
Without S9-mix, 3 hours treatment: 0.01, 0.03, 0.1, 0.3, 1, 1.5, 2 and 2.5 µg/ml
With S9-mix, 3 hours treatment: 0.1, 0.3, 1, 3, 10, 16, 18 and 20 µg/ml
Experiment 2
Without S9-mix, 24 hours treatment: 0.08, 0.24, 0.5, 0.8, 1, 1.3, 1.6 and 2 µg/ml
With S9-mix, 3 hours treatment: 1, 3, 6, 10, 15, 18, 20, 22, 26, 28 and 30 μg/ml

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: ethanol
- Justification for choice of solvent/vehicle: Test compound is stable in ethanol and this solvent has been accepted and approved by authorities and international guidelines

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Positive control substance:

methylmethanesulfonate

Remarks:

without S9 Migrated to IUCLID6: : 15 µg/ml for the 3 hours treatment period and 5 µg/ml for the 24 hours treatment period

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Positive control substance:

cyclophosphamide

Remarks:

with S9 Migrated to IUCLID6: : 7.5 µg/plate

Details on test system and experimental conditions:

METHOD OF APPLICATION: in medium

DURATION
- Exposure duration:
Short-term treatment
With and without S9-mix; 3 hours
Prolonged treatment period
Without S9-mix: 24 hours
- Expression time (cells in growth medium): 2 days
- Selection time (if incubation with a selection agent): 11 to 12 days

SELECTION AGENT (mutation assays): 5 µg/ml trifluorothymidine (TFT)

NUMBER OF REPLICATIONS:
- Solvent controls: Duplo cultures
- Treatment groups and positive control: Single cultures

NUMBER OF CELLS EVALUATED: 9.6 x 105 cells/concentration

DETERMINATION OF CYTOTOXICITY
- Method: relative suspension growth (dose range finding test) and relative total growth (mutation experiments)

RANGE-FINDING/SCREENING STUDIES:
-The suspension growth expressed as the reduction in cell growth after approximately 24 and 48 hours or only 24 hours cell growth, compared to the cell growth of the solvent control, was used to determine an appropriate dose range for the mutagenicity tests

Evaluation criteria:

The global evaluation factor (GEF) has been defined by the IWTG as the mean of the negative/solvent MF distribution plus one standard deviation. For the micro well version of the assay the GEF is 126 (ref. 12).

A test substance is considered positive (mutagenic) in the mutation assay if:
a) It induces a MF of more then MF(controls) + 126 in a dose-dependent manner; or
b) In case a repeat experiment is performed when a positive response is observed in one of the tester strains, the positive response should be reproducible in at least one independently repeated experiment.
An observed increase should be biologically relevant and will be compared with the historical control data range.

A test substance is considered equivocal (questionable) in the mutation assay if no clear conclusion for positive or negative result can be made after an additional confirmation study.

A test substance is considered negative (not mutagenic) in the mutation assay if:
a) None of the tested concentrations reaches a mutation frequency of MF(controls) + 126.
b) The results are confirmed in an independently repeated test.

Species / strain:

mouse lymphoma L5178Y cells

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH: No
- Effects of osmolality: No
- Precipitation: Precipitation in the exposure medium was observed at dose levels of 100 µg/ml and above


RANGE-FINDING/SCREENING STUDIES:
- Toxicity was observed at dose levels of 3 µg/ml in the absence of S9, 3 hours treatment; at dose levels of 33 µg/ml in the presence of S9, 3 hours treatment; at dose levels of 2.4 µg/ml in the absence of S9, 24 hours treatment


COMPARISON WITH HISTORICAL CONTROL DATA:
The spontaneous mutation frequencies in the solvent-treated control cultures were between the minimum and maximum value of the historical control data range and within the acceptability criteria of this assay.


ADDITIONAL INFORMATION ON CYTOTOXICITY:
In the absence of S9-mix, the relative total growth of the highest test substance concentration was reduced by 80 and 89% compared to the total growth of the solvent controls after the 3 and 24 hours treatment period, respectively.

In the presence of S9-mix, the relative total growth of the highest test substance concentration was reduced by 82 and 76% compared to the total growth of the solvent controls after the 3 hours treatment period in the first and second experiment, respectively.

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'. Remarks: L5178Y/TK+/--3.7.2C

Conclusions:

Interpretation of results (migrated information):
negative

Tallow dipropylenetriamine is not mutagenic in the TK mutation test system under the experimental conditions described in this report.

Executive summary:

Tallow dipropylenetriamine was evaluated for its possible induction of forward mutations at the thymidine-kinase locus (TK-locus) in L5178Y mouse lymphoma cells. The test was performed in two independent experiments in the absence and presence of S9-mix. The study was performed under GLP and according to the most recent OECD and EU guidelines.

Batch S001096 of Tallow dipropylenetriamine was a white paste. The test substance was dissolved in ethanol.

In the first experiment, Tallow dipropylenetriamine was tested up to concentrations of 2.5 and 20 μg/ml in the absence and presence of 8% (v/v) S9-mix. The incubation time was 3 hours. Tallow dipropylenetriamine was tested up to cytotoxic levels of 80 and 82% in the absence and presence of S9-mix, respectively.

In the second experiment, Tallow dipropylenetriamine was tested up to concentrations of 2 and 30 μg/ml in the absence and presence of 12% (v/v) S9-mix. The incubation times were 24 hours and 3 hours for incubations in the absence and presence of S9-mix, respectively. Tallow dipropylenetriamine was tested up to cytotoxic levels of 89% in the absence of S9-mix and 76% in the presence of S9-mix.

The spontaneous mutation frequencies in the solvent-treated control cultures were between the minimum and maximum value of the historical control data range

Positive control chemicals, methyl methane sulfonate and cyclophosphamide induced appropriate responses.

In the absence of S9-mix, Tallow dipropylenetriamine did not induce a significant increase in the mutation frequency in the first experiment. This result was confirmed in a repeat experiment with modifications in the duration of treatment time.

In the presence of S9-mix, Tallow dipropylenetriamine did not induce a significant increase in the mutation frequency in the first experiment. This result was confirmed in an independent experiment with modifications in the composition of the S9 concentration for metabolic activation.

In conclusion, Tallow dipropylenetriamine is not mutagenic in the TK mutation test system under the experimental conditions described in this report.

Reference 2

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:

24-jun-2009 to 12-sep-2009

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: The study has been performed according to OECD and/or EC guidelines and according to GLP principles.

Qualifier:

according to guideline

Guideline:

OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.10 (Mutagenicity - In Vitro Mammalian Chromosome Aberration Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of assay:

in vitro mammalian chromosome aberration test

Species / strain / cell type:

lymphocytes: human peripheral

Details on mammalian cell type (if applicable):

- Type and identity of media:
Blood samples
Blood samples were collected by venapuncture using the Venoject multiple sample blood collecting system with a suitable size sterile vessel containing sodium heparin. Immediately after blood collection lymphocyte cultures were started.

Culture medium
Culture medium consisted of RPMI 1640 medium, supplemented with 20% (v/v) heat-inactivated (56°C; 30 min) foetal calf serum, L-glutamine (2 mM), penicillin/streptomycin (50 U/ml and 50 µg/ml respectively) and 30 U/ml heparin.

Lymphocyte cultures
Whole blood (0.4 ml) treated with heparin was added to 5 ml or 4.8 ml culture medium (in the absence and presence of S9-mix, respectively). Per culture 0.1 ml (9 mg/ml) phytohaemagglutinin was added.

Metabolic activation:

with and without

Metabolic activation system:

Rat liver S9-mix induced by a combination of phenobarbital and ß-naphthoflavone

Test concentrations with justification for top dose:

Dose range finding test:
Without and with S9-mix, 3hr exposure; 24 hr fixation: 5, 10, 20, 30 and 40 µg/ml
Without S9-mix, 24/48hr exposure; 24/48 hr fixation: 1, 3, 10, 33, 100 and 333 µg/ml
First cytogenetic test:
Without S9-mix, 3 h exposure time, 24 h fixation time: 14, 16 and 18 µg/ml
With S9-mix, 3 h exposure, 24 h fixation time: 10, 20 and 26 µg/ml
Second cytogenetic test:
Without S9-mix, 24 hr exposure; 24 hr fixation: 1, 10 and 15 µg/ml
Without S9-mix, 48 hr exposure; 48 hr fixation: 1, 3 and 10 µg/ml
With S9-mix, 3 hr exposure; 48 hr fixation: 10, 22 and 26 µg/ml

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: ethanol
- Justification for choice of solvent/vehicle: Test compound was stable in ethanol and ethanol has been accepted and approved by authorities and international guidelines

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Positive control substance:

mitomycin C

Remarks:

without S9-mix Migrated to IUCLID6: in Hank's Balanced Salt Solution: 0.5 µg/ml for a 3 h exposure period, 0.2 µg/ml for a 24 h exposure period and 0.1 µg/ml for a 48 h exposure period

Positive control substance:

cyclophosphamide

Remarks:

with S9-mix Migrated to IUCLID6: in Hank's Balanced Salt Solution: 10 µg/ml

Details on test system and experimental conditions:

METHOD OF APPLICATION: in medium

DURATION
- Preincubation period: 48 hr
- Exposure duration: 3 hr (with and without S9-mix), 24 and 48 hr (without S9-mix)
- Fixation time (start of exposure up to fixation or harvest of cells): 24 and 48 hr

SPINDLE INHIBITOR (cytogenetic assays): colchicine
STAIN (for cytogenetic assays): Giemsa

NUMBER OF REPLICATIONS: duplicates in two independent experiments

NUMBER OF CELLS EVALUATED: 100 metaphase chromosome spreads per culture

DETERMINATION OF CYTOTOXICITY
- Method: mitotic index of each culture was determined by counting the number of metaphases per 1000 cells

OTHER EXAMINATIONS:
- Determination of polyploidy: yes
- Determination of endoreplication: yes

Evaluation criteria:

A test substance was considered positive (clastogenic) in the chromosome aberration test if:
a) It induced a dose-related statistically significant (Chi-square test, one-sided, p < 0.05) increase in the number of cells with chromosome aberrations.
b) A statistically significant and biologically relevant increase in the frequencies of the number of cells with chromosome aberrations was observed in the absence of a clear dose-response relationship.

A test substance was considered negative (not clastogenic) in the chromosome aberration test if none of the tested concentrations induced a statistically significant (Chi-square test, one-sided, p < 0.05) increase in the number of cells with chromosome aberrations.

Statistics:

The incidence of aberrant cells (cells with one or more chromosome aberrations, gaps included or excluded) for each exposure group outside the laboratory historical control data range was compared to that of the solvent control using Chi-square statistics

Species / strain:

lymphocytes: human peripheral

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH: No
- Effects of osmolality: No
- Precipitation: Precipitation in the exposure medium was observed at dose levels of 100 µg/ml and above

RANGE-FINDING/SCREENING STUDIES:
- Toxicity was observed at dose levels of 20 µg/ml and above in the absence and presence of S9, 3 hr treatment/24 hr fixation; at dose levels of 3 µg/ml and above in the absence of S9 for the continuous treatment of 24 hr and at dose levels of 10 µg/ml and above in the absence of S9 for the continuous treatment of 48 hr

COMPARISON WITH HISTORICAL CONTROL DATA:
- The number of cells with chromosome aberrations found in the solvent and positive control cultures was within the laboratory historical control data range.

ADDITIONAL INFORMATION ON CYTOTOXICITY:
- Appropriate toxicity was reached at the dose levels selected for scoring.

Conclusions:

Interpretation of results (migrated information):
negative

Tallow dipropylenetriamine did not induce a statistically significant or biologically relevant increase in the number of cells with chromosome aberrations in the absence and presence of S9-mix, in either of the two independently repeated experiments.

Executive summary:

Tallow dipropylenetriamine was studied for its effect on the number of chromosome aberrations in cultured peripheral human lymphocytes in the presence and absence of a metabolic activation system (phenobarbital and ß-naphthoflavone induced rat liver S9-mix), in two independent experiments.

The study was performedunder GLP and according to the most recent OECD and EU guidelines.

 

Tallow dipropylenetriamine was a white paste. Tallow dipropylenetriamine was dissolved in ethanol.

 

In the first cytogenetic assay, Tallow dipropylenetriamine was tested up to 18 and 26 μg/ml for a 3 h exposure time with a 24 h fixation time in the absence and presence of 1.8% (v/v) S9-fraction, respectively. Appropriate toxicity was reached at these dose levels.

In the second cytogenetic assay, Tallow dipropylenetriamine was tested up to 15 μg/ml for a 24 h continuous exposure time with a 24 h fixation time and up to 10 μg/ml for a 48 h continuous exposure time with a 48 h fixation time in the absence of S9-mix. In the presence of S9-mix Tallow dipropylenetriamine was tested up to 26 μg/ml for a 3 h exposure time with a 48 h fixation time. Appropriate toxicity was reached at these dose levels.

The number of cells with chromosome aberrations found in the solvent control cultures was within the laboratory historical control data range. Positive control chemicals, mitomycin C and cyclophosphamide, both produced a statistically significant increase in the incidence of cells with chromosome aberrations, indicating that the test conditions were adequate and that the metabolic activation system (S9-mix) functioned properly.

Tallow dipropylenetriamine did not induce a statistically significant or biologically relevant increase in the number of cells with chromosome aberrations in the absence and presence of S9-mix, in either of the two independently repeated experiments.

It was noted that Tallow dipropylenetriamine increased the number of polyploid cells both in the absence and presence of S9-mix in the first cytogenetic assay, in the absence of S9-mix at the 24 h exposure time and in the presence of S9-mix in the second cytogenetic assay. In the second cytogenetic assay the increase was dose dependent both in the absence and presence of S9-mix. This may indicate that Tallow dipropylenetriamine has the potential to inhibit mitotic processes.

No effects of Tallow dipropylenetriamine on the number of cells with endoreduplicated chromosomes was observed both in the absence and presence of S9-mix, in either of the two independently repeated experiments.

Finally, it is concluded that this test is valid and that Tallow dipropylenetriamine is not clastogenic in human lymphocytes under the experimental conditions described in this report. Tallow dipropylenetriamine may have the potential to inhibit mitotic processes and to induce numerical chromosome aberrations.

Reference 3

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

24-jun-2009 to 06-jul-2009

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: The study has been performed according to OECD and/or EC guidelines and according to GLP principles.

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

no

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of assay:

bacterial reverse mutation assay

Target gene:

- S. typhimurium: Histidine gene
- E. coli: Tryptophan gene

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2

Metabolic activation:

with and without

Metabolic activation system:

Rat liver S9-mix induced by a combination of phenobarbital and ß-naphthoflavone

Test concentrations with justification for top dose:

Experiment 1:
Preliminary test (without and with S9) TA100 and WP2uvrA: 0.3, 1, 3, 10, 33, 100, 333, 1000, 3330 and 5000 µg/plate
Main study: TA1535, TA1537 and TA98:
Without S9-mix: 0.3, 1, 3, 10, 16 and 33 µg/plate
With S9-mix: 0.3, 1, 3, 10, 33 and 66 µg/plate.
Experiment 2:
TA1535 and TA1537
Without and with S9-mix: 0.3, 1, 3, 10, 33 and 66 µg/plate
TA98 and TA100
Without S9-mix: 0.3, 1, 3, 10, 16 and 33 µg/plate
With S9-mix: 0.3, 1, 3, 10, 33 and 66 µg/plate
WP2uvrA
Without S9-mix: 1, 3, 10, 33, 66 and 100 µg/plate
With S9-mix: 3, 10, 33, 100, 200 and 333 µg/plate

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: ethanol
- Justification for choice of solvent/vehicle: Accepted and approved by authorities and international guidelines

Negative solvent / vehicle controls:

yes

Remarks:

: ethanol

Positive controls:

yes

Positive control substance:

sodium azide

Remarks:

without S9 Migrated to IUCLID6: 5 µg/plate in saline for TA1535

Positive control substance:

9-aminoacridine

Remarks:

without S9 Migrated to IUCLID6: 60 µg/plate in water for TA1537

Positive control substance:

2-nitrofluorene

Remarks:

without S9 Migrated to IUCLID6: 10 µg/plate in DMSO for TA98

Positive control substance:

methylmethanesulfonate

Remarks:

without S9 Migrated to IUCLID6: 650 µg/plate in DMSO for TA100

Positive control substance:

4-nitroquinoline-N-oxide

Remarks:

without S9 Migrated to IUCLID6: 10 µg/plate in DMSO for WP2 uvrA

Positive control substance:

other: 2-aminoanthracene in DMSO for all tester strains

Remarks:

with S9

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Exposure duration: 48 h

NUMBER OF REPLICATIONS: Doses of the test substance were tested in triplicate in each strain. Two independent experiments were conducted.

DETERMINATION OF CYTOTOXICITY
- Method: The reduction of the bacterial background lawn, the increase in the size of the microcolonies and the reduction of the revertant colonies

OTHER EXAMINATIONS:
Precipitation of the test substance

Evaluation criteria:

A test substance is considered negative (not mutagenic) in the test if:
a) The total number of revertants in tester strain TA100 is not greater than two (2) times the concurrent control, and the total number of revertants in tester strains TA1535, TA1537, TA98 or WP2uvrA is not greater than three (3) times the concurrent control.
b) The negative response should be reproducible in at least one independently repeated experiment.

A test substance is considered positive if:
a) A two-fold (TA100) or more or a three-fold (TA1535, TA1537, TA98, WP2uvrA) or more increase above solvent control in the mean number of revertant colonies is observed in the test substance group.
b) The increase in the mean number of revertant colonies follows the concentration of test substance (dose-response relationship).

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

True negative controls validity:

not applicable

Positive controls validity:

valid

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

True negative controls validity:

not applicable

Positive controls validity:

valid

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

True negative controls validity:

not applicable

Positive controls validity:

valid

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

True negative controls validity:

not applicable

Positive controls validity:

valid

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

True negative controls validity:

not applicable

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: Slight precipitation was observed at dose levels of 3330 and 5000 μg/plate.

RANGE-FINDING/SCREENING STUDIES:
- In tester strain TA100, toxicity was observed at dose levels of 33 μg/plate and above in the absence and presence of S9-mix. In tester strain WP2uvrA, toxicity was observed at dose levels of 100 μg/plate and above in the absence of S9-mix and at 333 µg/plate in the presence of S9-mix.

COMPARISON WITH HISTORICAL CONTROL DATA:
- The negative and strain-specific positive control values were within our laboratory historical control data ranges indicating that the test conditions were adequate and that the metabolic activation system functioned properly.

ADDITIONAL INFORMATION ON CYTOTOXICITY:
TA1535: without S9: 33 µg/plate and above and with S9: 66 µg/plate and above
TA1537: without S9: 33 µg/plate and above and with S9: 33 µg/plate and above
TA98: without S9: 33 µg/plate and above and with S9: 66 µg/plate and above
TA100: without S9: 33 µg/plate and above and with S9: 33 µg/plate and above
WP2uvrA: without S9: 66 µg/plate and above and with S9: 200 µg/plate and above

Conclusions:

Interpretation of results (migrated information):
negative

Tallow dipropylenetriamine is not mutagenic in the Salmonella typhimurium reverse mutation assay and in the Escherichia coli reverse mutation assay.

Executive summary:

Tallow dipropylenetriamine was tested in theSalmonella typhimuriumreverse mutation assay with four histidine-requiring strains ofSalmonella typhimurium(TA1535, TA1537, TA98 and TA100) and in theEscherichia colireverse mutation assay with a tryptophan-requiring strain ofEscherichia coli(WP2uvrA). The test was performed in two independent experiments in the presence and absence of S9-mix (rat liver S9-mix induced by a combination of Phenobarbital and ß-naphthoflavone).

 

The study procedures described in this report were based on the most recent OECD and EC guidelines. Batch S001096 of Tallow dipropylenetriamine was a white paste. The test substance was dissolved in ethanol. The dose range finding study was reported as part of the first experiment of the mutation assay.

 

Tallow dipropylenetriamine precipitated on the plates at dose levels of 3330 and 5000 μg/plate.

Cytotoxicity, as evidenced by a decrease in the number of revertants and/or a reduction of the bacterial background lawn, was observed in all tester strains in the absence and presence of S9-mix. With the exception of tester strain TA1537 (without S9-mix, first experiment), in which no cytotoxicity was observed. However, since toxicity was observed in the independent repeat experiment, in a dose level just above the highest dose level tested in this experiment, the validity of the test was considered to be not affected by this lack of toxicity in one tester strain.

 

Tallow dipropylenetriamine did not induce a significant dose-related increase in the number of revertant (His+) colonies in each of the four tester strains (TA1535, TA1537, TA98 and TA100) and in the number of revertant (Trp+) colonies in tester strain WP2uvrA both in the absence and presence of S9-metabolic activation. These results were confirmed in an independently repeated experiment.

 

In this study, the negative and strain-specific positive control values were within the laboratory historical control data ranges indicating that the test conditions were adequate and that the metabolic activation system functioned properly.

 

Based on the results of this study it is concluded that Tallow dipropylenetriamine is not mutagenic in theSalmonella typhimuriumreverse mutation assay and in theEscherichia colireverse mutation assay.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Genetic toxicity in vivo

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Tallow dipropylenetriamine was tested in the Salmonella typhimurium reverse mutation assay with four histidine-requiring strains of Salmonella typhimurium(TA1535, TA1537, TA98 and TA100) and in theEscherichia coli reverse mutation assay with a tryptophan-requiring strain of Escherichia coli (WP2uvrA). The test was performed in two independent experiments in the presence and absence of S9-mix (rat liver S9-mix induced by a combination of Phenobarbital and ß-naphthoflavone). The study followed the most recent OECD and EU protocols and was performed under GLP.

There was no significant or dose-related increase in the number of revertant colonies in any of the applied strains, both with and without S9-mix. This was confirmed in an independently repeated experiment.

It is concluded that Tallow dipropylenetriamine is not mutagenic in the Salmonella typhimurium reverse mutation assay and in the Escherichia coli reverse mutation assay.

 

Tallow dipropylenetriamine was studied for its effect on the number of chromosome aberrations in cultured peripheral human lymphocytes in the presence and absence of a metabolic activation system (phenobarbital and ß-naphthoflavone induced rat liver S9-mix), in two independent experiments. The study was performed under GLP and according to the most recent OECD and EU guidelines.

Tallow dipropylenetriamine did not induce a statistically significant or biologically relevant increase in the number of cells with chromosome aberrations in the absence and presence of S9-mix, in either of the two independently repeated experiments.

It was noted that Tallow dipropylenetriamine increased the number of polyploid cells both in the absence and presence of S9-mix in the first cytogenetic assay, in the absence of S9-mix at the 24 h exposure time and in the presence of S9-mix in the second cytogenetic assay. In the second cytogenetic assay the increase was dose dependent both in the absence and presence of S9-mix. Polyploidy alone does not indicate aneugenic potential and can simply indicate cell cycle perturbation; it is also commonly associated with increasing cytotoxicity.

There were no effects of Tallow dipropylenetriamine on the number of cells with endoreduplicated chromosomes both in the absence and presence of S9-mix, in either of the two independently repeated experiments. Therefore, it is concluded that Tallow dipropylenetriamine is not clastogenic in human lymphocytes.

 

Tallow dipropylenetriamine was evaluated for its possible induction of forward mutations at the thymidine-kinase locus (TK-locus) in L5178Y mouse lymphoma cells. The test was performed in two independent experiments in the absence and presence of S9-mix. The study was performed under GLP and according to the most recent OECD and EU guidelines.

In both the presence and absence of S9-mix, Tallow dipropylenetriamine did not induce a significant increase in the mutation frequency in the first experiments. This result was confirmed in a repeat experiment with modifications in the duration of treatment time (without S9-mix) or S9 concentration (with S9-mix). Therefore, Tallow dipropylenetriamine is not mutagenic in the TK mutation test.

 

Additionally, polyamines do not react with DNA or react to protein (indicated by OECD Toolbox profiling).

Justification for classification or non-classification

Tallow dipropylenetriamine is not mutagenic in the Salmonella typhimurium reverse mutation assay and in the Escherichia coli reverse mutation assay, is not clastogenic in human lymphocytes, and not mutagenic in the TK mutation test with L5178Y mouse lymphoma cells.

Furthermore, polyamines do not react with DNA or react to protein.

It can be concluded that Tallow dipropylenetriamine is not genotoxic and therefore classification as dangerous due to genotoxicity does not apply to this material.