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EC number: 249-276-6 | CAS number: 28872-01-7
- Life Cycle description
- Uses advised against
- Endpoint summary
- Appearance / physical state / colour
- Melting point / freezing point
- Boiling point
- Density
- Particle size distribution (Granulometry)
- Vapour pressure
- Partition coefficient
- Water solubility
- Solubility in organic solvents / fat solubility
- Surface tension
- Flash point
- Auto flammability
- Flammability
- Explosiveness
- Oxidising properties
- Oxidation reduction potential
- Stability in organic solvents and identity of relevant degradation products
- Storage stability and reactivity towards container material
- Stability: thermal, sunlight, metals
- pH
- Dissociation constant
- Viscosity
- Additional physico-chemical information
- Additional physico-chemical properties of nanomaterials
- Nanomaterial agglomeration / aggregation
- Nanomaterial crystalline phase
- Nanomaterial crystallite and grain size
- Nanomaterial aspect ratio / shape
- Nanomaterial specific surface area
- Nanomaterial Zeta potential
- Nanomaterial surface chemistry
- Nanomaterial dustiness
- Nanomaterial porosity
- Nanomaterial pour density
- Nanomaterial photocatalytic activity
- Nanomaterial radical formation potential
- Nanomaterial catalytic activity
- Endpoint summary
- Stability
- Biodegradation
- Bioaccumulation
- Transport and distribution
- Environmental data
- Additional information on environmental fate and behaviour
- Ecotoxicological Summary
- Aquatic toxicity
- Endpoint summary
- Short-term toxicity to fish
- Long-term toxicity to fish
- Short-term toxicity to aquatic invertebrates
- Long-term toxicity to aquatic invertebrates
- Toxicity to aquatic algae and cyanobacteria
- Toxicity to aquatic plants other than algae
- Toxicity to microorganisms
- Endocrine disrupter testing in aquatic vertebrates – in vivo
- Toxicity to other aquatic organisms
- Sediment toxicity
- Terrestrial toxicity
- Biological effects monitoring
- Biotransformation and kinetics
- Additional ecotoxological information
- Toxicological Summary
- Toxicokinetics, metabolism and distribution
- Acute Toxicity
- Irritation / corrosion
- Sensitisation
- Repeated dose toxicity
- Genetic toxicity
- Carcinogenicity
- Toxicity to reproduction
- Specific investigations
- Exposure related observations in humans
- Toxic effects on livestock and pets
- Additional toxicological data
![](https://echa.europa.eu/o/diss-blank-theme/images/factsheets/A-REACH/factsheet/print_toxicological-information.png)
Endpoint summary
Administrative data
Key value for chemical safety assessment
Genetic toxicity in vitro
Description of key information
Link to relevant study records
- 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:
- 16-jun-2009 to 29-jun-2009
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- 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 and TA 100
- Species / strain / cell type:
- E. coli WP2 uvr A
- 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): 3, 10, 33, 100, 333, 1000, 3330 and 5000 µg/plate
Main study (TA1535, TA1537, TA98 and TA100):
Without S9-mix: 0.3, 1, 3, 10, 20 and 33 µg/plate
With S9-mix: 0.3, 1, 3, 10, 33 and 66 µg/plate
Experiment 2:
TA1535, TA1537, TA98 and TA100
(Without S9): 0.3, 1, 3, 10, 20 and 33 µg/plate
(With S9): 0.3, 1, 3, 10, 33 and 66 µg/plate
WP2uvrA:
(Without and with S9): 1, 3, 10, 33, 66 and 100 µg/plate - Vehicle / solvent:
- - Vehicle(s)/solvent(s) used: Ethanol
- Justification for choice of solvent/vehicle: Accepted and approved by authorities and international guidelines - Untreated negative controls:
- no
- 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 WP2uvrA
- 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 hours
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:
- Precipitation of 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:
b) 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
- Species / strain:
- S. typhimurium TA 98
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- cytotoxicity
- Species / strain:
- S. typhimurium TA 1537
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- cytotoxicity
- Species / strain:
- S. typhimurium TA 1535
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- cytotoxicity
- Species / strain:
- E. coli WP2 uvr A
- 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
- Precipitation: Slight precipitate was observed at the top dose of 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 at 100 µg/plate in the absence and 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: 20 µg/plate and above and with S9: 33 µg/plate and above
TA1537: without S9: 20 µg/plate and above and with S9: 10 µg/plate and above
TA98: without S9: 20 µg/plate and above and with S9: 33 µg/plate and above
TA100: without S9: 20 µg/plate and above and with S9: 33 µg/plate and above
WP2uvrA: without S9: 66 µg/plate and above and with S9: 100 µg/plate and above - Conclusions:
- Based on the results of this study it is concluded that Oleyl dipropylenetriamine is not mutagenic in the Salmonella typhimurium reverse mutation assay and in the Escherichia coli reverse mutation assay.
- Executive summary:
Oleyl 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 the Escherichia 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 procedures described in this report were based on the most recent OECD and EC guidelines.
Oleyl dipropylenetriamine was an off white turbid liquid. The test substance was dissolved in ethanol.
Experiment
Tester strains
Metabolic activation
Concentration range
Dose range finding
TA100 and WP2uvrA
-
5% (v/v) S9
Up to 5000μg/plate
Up to 5000μg/plate
Experiment 1
TA1535, TA1537, TA98 and TA100
-
5% (v/v) S9
Up to 33μg/plate
Up to 66μg/plate
Experiment 2
TA1535, TA1537, TA98 and TA100
-
10% (v/v) S9
Up to 33μg/plate
Up to 66μg/plate
WP2uvrA
-
10% (v/v) S9
Up to 100μg/plate
Up to 100μg/plate
- no S9-mix added.
Oleyl dipropylenetriamine precipitated on the plates at the top dose of 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.
Oleyl 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 Oleyl dipropylenetriamine is not mutagenic in the Salmonella typhimurium reverse mutation assay and in the Escherichia coli reverse mutation assay.
- Endpoint:
- in vitro cytogenicity / chromosome aberration study in mammalian cells
- Remarks:
- Type of genotoxicity: chromosome aberration
- Type of information:
- read-across from supporting substance (structural analogue or surrogate)
- Adequacy of study:
- key study
- Study period:
- 24-jun-2009 to 12-sep-2009
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Justification for type of information:
- (See also Category polyamines in support of Oleyl dipropylenetriamine attached to Ch.13)
Structurally, the two linear-alkyl dipropylene triamines Oleyl dipropylene triamine and Tallow (C16-18, C18-unsaturated) dipropylenetriamine are very similar: a linear alkyl chain and a primary amine at the end, with 2 secondary amines in between separated by a propyl group. Consequently, they share the same chemical reactivity and their physico-chemical properties are very similar from which a comparable toxicological profile can be expected.
Within a specific chemical structure, the variability of the alkyl chain length is considered to have a possible modifying activity, which is related to modification of the physiological properties of the molecule by the increase or shortening of the apolar alkyl chain part. This is suspected to influence aspects related to bioavailability, but not aspects of chemical reactivity and metabolism pathways, aspects that could have an impact on specific mechanisms of toxicity.
The ratio between the C16, C18 and C18:1 (C18-unsaturated) alkyl chains in the Oleyl and Tallow dipropylene triamine products show an overlap in alkyl chain of about 50%:
- Oleyl: C18:1 = 85.5%; C18 = 12.4 %; C16: 7,1%
- Tallow: C18:1 = 26.5%; C18 = 38.3 %; C16: 35.2%
The higher level of unsaturation in Oleyl based products has never shown to have an important effect. This is in agreement with the expectation that these structures do not undergo an important level of metabolism, and if metabolism occurs on the limited absorbed material, the resulting alkyl chains will fit in the physiological pool of these natural alkyl chains. Further, the relatively somewhat sorter chain lengths of C16 would make it slightly more bioavailable, in agreement to the notion that shorter alkyl chains represent a more conservative evaluation.
Consequently, the data on Tallow (C16-18, C18-unsaturated-alkyl) dipropylene triamine can be used for the read-across to Oleyl dipropylene triamine. - Reason / purpose for cross-reference:
- read-across source
- 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:
- 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.
- Endpoint:
- in vitro gene mutation study in mammalian cells
- Remarks:
- Type of genotoxicity: gene mutation
- Type of information:
- read-across from supporting substance (structural analogue or surrogate)
- Adequacy of study:
- key study
- Study period:
- 01-jul-2009 to 11-aug-2009
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Justification for type of information:
- (See also Category polyamines in support of Oleyl dipropylenetriamine attached to Ch.13)
Structurally, the two linear-alkyl dipropylene triamines Oleyl dipropylene triamine and Tallow (C16-18, C18-unsaturated) dipropylenetriamine are very similar: a linear alkyl chain and a primary amine at the end, with 2 secondary amines in between separated by a propyl group. Consequently, they share the same chemical reactivity and their physico-chemical properties are very similar from which a comparable toxicological profile can be expected.
Within a specific chemical structure, the variability of the alkyl chain length is considered to have a possible modifying activity, which is related to modification of the physiological properties of the molecule by the increase or shortening of the apolar alkyl chain part. This is suspected to influence aspects related to bioavailability, but not aspects of chemical reactivity and metabolism pathways, aspects that could have an impact on specific mechanisms of toxicity.
The ratio between the C16, C18 and C18:1 (C18-unsaturated) alkyl chains in the Oleyl and Tallow dipropylene triamine products show an overlap in alkyl chain of about 50%:
- Oleyl: C18:1 = 85.5%; C18 = 12.4 %; C16: 7,1%
- Tallow: C18:1 = 26.5%; C18 = 38.3 %; C16: 35.2%
The higher level of unsaturation in Oleyl based products has never shown to have an important effect. This is in agreement with the expectation that these structures do not undergo an important level of metabolism, and if metabolism occurs on the limited absorbed material, the resulting alkyl chains will fit in the physiological pool of these natural alkyl chains. Further, the relatively somewhat sorter chain lengths of C16 would make it slightly more bioavailable, in agreement to the notion that shorter alkyl chains represent a more conservative evaluation.
Consequently, the data on Tallow (C16-18, C18-unsaturated-alkyl) dipropylene triamine can be used for the read-across to Oleyl dipropylene triamine. - Reason / purpose for cross-reference:
- read-across source
- 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.
- Endpoint:
- in vitro cytogenicity / chromosome aberration study in mammalian cells
- Remarks:
- Type of genotoxicity: chromosome aberration
- Type of information:
- read-across based on grouping of substances (category approach)
- Adequacy of study:
- supporting study
- Study period:
- 15-jun-2009 to 12-aug-2009
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Justification for type of information:
- (See also Category polyamines in support of Oleyl dipropylenetriamine attached to Ch.13)
Structurally, the two linear-alkyl dipropylene triamines Oleyl dipropylene triamine and Coco (C12-18) dipropylenetriamine are very similar: a linear alkyl chain and a primary amine at the end, with 2 secondary amines in between separated by a propyl group. Consequently, they share the same chemical reactivity and their physico-chemical properties are very similar from which a comparable toxicological profile can be expected.
Within a specific chemical structure, the variability of the alkyl chain length is considered to have a possible modifying activity, which is related to modification of the physiological properties of the molecule by the increase or shortening of the apolar alkyl chain part. This is suspected to influence aspects related to bioavailability, but not aspects of chemical reactivity and metabolism pathways, aspects that could have an impact on specific mechanisms of toxicity. In series of substances that are chemically identical but differ in length of alkyl chains, those that have shorter chain length are likely to be more bioavailable compared to those with longer chain lengths. Therefore, results from the shortest chain length, in this case Coco (C12-C18 alkyl) dipropylene triamine, can be considered a worst-case approach for the longer chain lengths, specifically Oleyl (C18, unsaturated) dipropylene triamine. - 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:
Withoutand with S9-mix, 3hr exposure; 24 hr fixation: 10, 33 and 100 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: 5, 10 and 15 µg/ml
With S9-mix, 3 h exposure, 24 h fixation time: 5, 15 and 25 µg/ml
Second cytogenetic test:
Without S9-mix, 24 hr exposure; 24 hr fixation:: 3, 7 and 15 µg/ml
Without S9-mix, 48 hr exposure; 48 hr fixation: 1, 3 and 7 µg/ml
With S9-mix, 3 hr exposure; 48 hr fixation: 20, 25 and 30 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 controls:
- yes
- 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: At concentrations of 100 µg/ml and higher
RANGE-FINDING/SCREENING STUDIES:
- Toxicity was observed at dose levels of 33 µg/ml and above in the absence and presence of S9-mix, 3 hr treatment/24 hr fixation and at 10 µg/ml for the continuous treament of 24 and 48 hr in the absence of S9-mix .
- 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
It is concluded that this test is valid and that Coco dipropylene triamine is not clastogenic in human lymphocytes under the experimental conditions described in this report. Coco dipropylene triamine may have the potential to inhibit mitotic processes and to induce numerical chromosome aberrations. - Executive summary:
Evaluation of the ability of Coco dipropylene triamine to induce chromosome aberrations in cultured peripheral human lymphocytes (with repeat experiment).
This report describes the effect of Coco dipropylene triamine 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). The possible clastogenicity of Coco dipropylene triamine was tested in two independent experiments.
The study procedures described in this report were based on the most recent OECD and EC guidelines.
Batch S001254 of Coco dipropylene triamine was a yellow clear to turbid liquid with flakes. Coco dipropylene triamine was dissolved in ethanol.
In the first cytogenetic assay, Coco dipropylene triamine was tested up to 15 and 25 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, Coco dipropylene triamine was tested up to 15 μg/ml for a 24 h continuous exposure time with a 24 h fixation time and up to 7 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 Coco dipropylene triamine was tested up to 30 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.
Coco dipropylene triamine 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 Coco dipropylene triamine increased the number of polyploid cells both in the absence and presence of S9-mix in the first cytogenetic assay and in the presence of S9-mix in the second cytogenetic assay. This may indicate that Coco dipropylene triamine has the potential to inhibit mitotic processes.
No effects of Coco dipropylene triamine 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 Coco dipropylene triamine is not clastogenic in human lymphocytes under the experimental conditions described in this report. Coco dipropylene triamine may have the potential to inhibit mitotic processes and to induce numerical chromosome aberrations.
- Endpoint:
- in vitro gene mutation study in mammalian cells
- Remarks:
- Type of genotoxicity: gene mutation
- Type of information:
- read-across based on grouping of substances (category approach)
- Adequacy of study:
- supporting study
- Study period:
- 17-jun-2009 to 03-aug-2009
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Justification for type of information:
- (See also Category polyamines in support of Oleyl dipropylenetriamine attached to Ch.13)
Structurally, the two linear-alkyl dipropylene triamines Oleyl dipropylene triamine and Coco (C12-18) dipropylenetriamine are very similar: a linear alkyl chain and a primary amine at the end, with 2 secondary amines in between separated by a propyl group. Consequently, they share the same chemical reactivity and their physico-chemical properties are very similar from which a comparable toxicological profile can be expected.
Within a specific chemical structure, the variability of the alkyl chain length is considered to have a possible modifying activity, which is related to modification of the physiological properties of the molecule by the increase or shortening of the apolar alkyl chain part. This is suspected to influence aspects related to bioavailability, but not aspects of chemical reactivity and metabolism pathways, aspects that could have an impact on specific mechanisms of toxicity. In series of substances that are chemically identical but differ in length of alkyl chains, those that have shorter chain length are likely to be more bioavailable compared to those with longer chain lengths. Therefore, results from the shortest chain length, in this case Coco (C12-C18 alkyl) dipropylene triamine, can be considered a worst-case approach for the longer chain lengths, specifically Oleyl (C18, unsaturated) dipropylene triamine. - 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
- Principles of method if other than guideline:
- The 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).
- 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 (Sigma) and 2 mM L-glutamin (Invitrogen Corporation) 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: 3, 10, 33, 100 and 333 µg/ml
Without S9-mix, 24 hours treatment: 3, 10, 33, 100 and 333 µg/ml and 0.003, 0.01, 0.03, 0.1, 0.3, 1 and 3 µg/ml
Experiment 1:
Without S9-mix, 3 hours treatment: 0.003, 0.03, 0.1, 0.25, 0.5, 1, 1.4 and 2 µg/ml
With S9-mix, 3 hours treatment: 0.01, 0.03, 0.1, 0.3, 1, 3, 10 and 12 µg/ml
Experiment 2
Without S9-mix, 24 hours treatment: 0.008, 0.024, 0.08, 0.2, 0.4, 0.08, 1.12 and 1.44 µg/ml
With S9-mix, 3 hours treatment: 0.01, 0.1, 1, 10, 12, 14, 16 and 17 μg/ml exposure medium - Vehicle / solvent:
- - Vehicle(s)/solvent(s) used:ethanol
- Justification for choice of solvent/vehicle: Accepted and approved by authorities and international guidelines - Untreated negative controls:
- no
- Negative solvent / vehicle controls:
- yes
- Remarks:
- : Ethanol
- 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
- Positive control substance:
- cyclophosphamide
- Remarks:
- with S9 Migrated to IUCLID6: 7.5 µg/ml
- 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
.
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. - 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
- Other confounding effects:
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 1 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. - Remarks on result:
- other: strain/cell type: L5178Y/TK+/--3.7.2C
- Remarks:
- Migrated from field 'Test system'.
- Conclusions:
- Interpretation of results (migrated information):
negative
It is concluded that Coco dipropylene triamine is not mutagenic in the mouse lymphoma L5178Y test system under the experimental conditions described in this report. - Executive summary:
Evaluation of the mutagenic activity of Coco dipropylene triamine in an in vitro mammalian cell gene mutation test with L5178Y mouse lymphoma cells (with independent repeat).
This report describes the effects of Coco dipropylene triamine on the 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 (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 S001254 of Coco dipropylene triamine was a yellow clear to turbid liquid with flakes. The test substance was dissolved in ethanol.
In the first experiment, Coco dipropylene triamine was tested up to concentrations of 2 and 12 μg/ml in the absence and presence of 8% (v/v) S9-mix, respectively. The incubation time was 3 hours. Coco dipropylene triamine was tested up to cytotoxic levels of 78 and 69% in the absence and presence of S9-mix, respectively.
In the second experiment, Coco dipropylene triamine was tested up to concentrations of 1.44 and 17 μg/ml in the absence and presence of 12% (v/v) S9-mix, respectively. The incubation times were 24 hours for incubations in the absence of S9-mix and 3 hours for incubations in the presence of S9-mix. Coco dipropylene triamine was tested up to cytotoxic levels of 83 and 66% in the absence and presence of S9-mix, respectively.
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.
Mutation frequencies in cultures treated with positive control chemicals were increased by 14- and 17-fold for MMS in the absence of S9-mix, and by 11- and 8.9-fold for CP in the presence of S9-mix. It was therefore concluded that the test conditions, both in the absence and presence of S9-mix, were appropriate and that the metabolic activation system (S9-mix) functioned properly.
In the absence of S9-mix, Coco dipropylene triamine did not induce a significant increase in the mutation frequency in the first experiment. This result was confirmed in an independent repeat experiment with modifications in the duration of treatment time.
In the presence of S9-mix, Coco dipropylene triamine did not induce a significant increase in the mutation frequency in the first experiment. This result was confirmed in an independent repeat experiment with modifications in the concentration of the S9 for metabolic activation.
It is concluded that Coco dipropylene triamine is not mutagenic in the mouse lymphoma L5178Y test system under the experimental conditions described in this report.
Referenceopen allclose all
Endpoint conclusion
- Endpoint conclusion:
- no adverse effect observed (negative)
Genetic toxicity in vivo
Endpoint conclusion
- Endpoint conclusion:
- no study available
Mode of Action Analysis / Human Relevance Framework
Based on structure and mechanism of cytotoxicity, genotoxicity is not expected. In physiological circumstances, the polyamines have a cationic surfactant structure (nitrogens are fully protonated)which leads to high adsorptive properties to negatively charged surfaces as cellular membranes. The apolar hydrophobic tails are pushed out of solution and easily dissolve in the membranes, whereas the polar head causes disruption and leakage of the membranes leading to cell damage or lysis of the cell content. As a consequence, the whole molecule will not easily pass membrane structures. Cytotoxicity through disruption of cell membrane will occur rather than absorption over the cell membrane into the cell and to move to the nucleus to interact with DNA.
Additional information
Oleyl dipropylene triamine was not mutagenic in a bacterial mutagenicity study (Ames test). Comparable alkyl dipropylene triamine structures, also covering the same alkyl chain distribution, have shown that these substances do not induced chromosomal aberrations in human lymphocytes, and are not mutagenic in mammalian mutagenicity studies in mouse lymphoma cells. All studies were performed under GLP according to current guidelines.
The following studies on alkyl dipropylene triamine are available,
Alkyl chain |
Study |
Endpoint |
Coco |
1219458-12-4, NOTOX, 491200, 2009 |
OECD 471 Bacterial mutagenicity (Ames) (not in dossier) |
Coco |
1219458-12-4, NOTOX, 491233, 2009 |
OECD 473 Chromosomal aberration in HL |
Coco |
1219458-12-4, NOTOX, 491231, 2009 |
OECD 476 MLA (TK-locus in L5178Y cells) |
Tallow |
1219458-14-6, NOTOX, 491200, 2009 |
OECD 471 Bacterial mutagenicity (Ames) (not in dossier) |
Tallow |
1219458-14-6, NOTOX, 491205, 2009 |
OECD 473 Chromosomal aberration in HL |
Tallow |
1219458-14-6, NOTOX, 491203, 2009 |
OECD 476 MLA (TK-locus in L5178Y cells) |
Oleyl |
28872-01-7, NOTOX, 491213, 2009 |
OECD 471 Bacterial mutagenicity (Ames) |
Coco = C12-18
Tallow = C16-18, C18-unsat.
Oleyl = C18 unsaturated
Oleyl dipropylene triamine was tested in the Salmonella typhimurium reverse mutation assay with four histidine-requiring strains of Salmonella typhimurium (TA1535, TA1537, TA98 and TA100) and in the Escherichia 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 Oleyl dipropylene triamine 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.
Cocdipropylenetriamine 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.
Cocodipropylenetriamine 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 Coco dipropylene triamine increased the number of polyploid cells both in the absence and presence of S9-mix in the first cytogenetic assay and in the presence of S9-mix in the second cytogenetic assay. 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 Cocodipropylenetriamine 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 Cocodipropylenetriamine is not clastogenic in human lymphocytes.
Cocodipropylenetriamine 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, Cocodipropylenetriamine 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, Cocodipropylenetriamine is not mutagenic in the TK mutation test.
Also studies on other polyamines indicate no concerns for genotoxicity. Furthermore, polyamines do not react with DNA or react to protein. Mode of action analysis further shows that these polyamines will not easily pass membrane structures. Cytotoxicity through disruption of cell membrane will occur rather than absorption over the cell membrane into the cell and to move to the nucleus to interact with DNA.
Justification for classification or non-classification
For each endpoint bacterial mutagenicity, mammalian mutagenicity and mammalian clastogenicity a GLP compliant study is available on the basis of testing withOleyl dipropylene triamineitself or from cross-reading from Tallow dipropylenetriamine.Lack of genotoxic properties of alkyl-dipropylenetriamines was further confirmed inin vitrostudies on Coco dipropylene triamine. Mode of action analysis further shows that genotoxic properties are not likely.As there is sufficient evidence for lack genotoxicity of Oley dipropylene triamine, no classification for genotoxic effects is required.
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