Propane-1,3-diol

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

In Vitro (Mutagenic effects - bacterial): OECD 471. Negative. Reliability = 1.

In Vitro (Cytogenic effects–mammalian): EU Method B.10. Positive. Reliability = 1.

In Vitro (Mutagenic effects - mammalian): OECD 476; CHL fibroblasts (V79). Negative. Reliability = 1.

Link to relevant study records

Reference

Reference 1

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

ACC PDO panel assigned reliability of 1. PDO from chemical process.

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Version / remarks:

First addendum to the OECD Guideline for Testing of Chemicals, Section 4, No; 471, adopted May 26, 1983

Qualifier:

according to guideline

Guideline:

other: EEC directive 92/69, L383 A, Annex V, B14, December 29, 1992

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Species / strain / cell type:

S. typhimurium, other: TA 1535, TA 1537, TA 98, TA 100 and TA 102

Metabolic activation:

with and without

Metabolic activation system:

S9 mix

Test concentrations with justification for top dose:

without S9 mix: 33.3; 100; 333.3; 1000; 2500; 5000 µg/plate
with S9 mix: 33.3; 100; 333.3; 1000; 2500; 5000 µg/plate

Vehicle / solvent:

distilled water

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Positive control substance:

sodium azide

Remarks:

strain TA153, TA100; dissolved in distilled water; without metabolic activation

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Remarks:

without metabolic activation

Positive control substance:

other: 4-nitro-o-phenylene-diamine

Remarks:

strain TA 1537, TA 98; dissolved in DMSO

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Positive control substance:

methylmethanesulfonate

Remarks:

strain TA 102; dissolved in distilled water; without metabolic activation

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Remarks:

with metabolic activation

Positive control substance:

other: 2-aminoanthracene (2-AA)

Remarks:

All strains tested; dissolved in DMSO

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation - Experiment 1) and pre-incubation (Experiment 2)

Bacteria are stored as stock cultures in ampoules with nutrient broth + 5% DMSO in liquid nitrogen
Preculture: 10 hours in a shaking water bath at 37°C; thawed bacterial suspension in nutrient medium (8 g/L Merck nutrient broth and 5 g/L NaCl).

Selective agar: 2.0% Vogel-Bonner-Glucose-Minimal-Agar.
Overlay agar: 6.0 g/L Merck Agar Agar; 6.0 g/L NaCl; 10.5 mg/L L-histidine x HCl x H2O; 12.2 mg/L biotin.

Experimental performance:
Mixing the following components:
- 100 µL test solution at each dose level, positive or negative control
- 500 µL S9 mix or S9 mix substitution buffer
- 100 µL bacterial suspension
- 2000 µL overly agar
This mix was poured onto the selective agar plates; for the pre-incubation assay the mix was incubated at 37°C for 60 min prior to pouring onto minimal agar plates.

DURATION:
Exposure duration: After solidification at least 48h at 37°C in the dark (upside down).

NUMBER OF REPLICATIONS:
At least 3 replicates for each strain and dose level.

DATA RECORDING:
Colonies were counted using the AUTOCOUNT (Artek Systems Corporation, BIOSYS GmbH, Karben). If precipitation of the test substance precluded automatic counting the revertant colonies were counted by hand.

EVALUATION OF RESULTS:
- Corresponding background growth on both negative controls and test plates.
- Normal range of spontaneous reversion rates; normal frequencies are:
TA 1535: 10-29
TA 1537: 5-28
TA 98: 15-57
TA 100: 77-189
TA 102: 121-293

Evaluation criteria:

A test article in considered positive if either a dose related and reproducible increase in the number of revertants or a significant and reproducible increase for at least one test concentration is induced.
A test article producing neither a dose related and reproducible increase in the number of revertants nor a significant and reproducible positive response at any one of the test points is considered non-mutagenic in this system.

Species / strain:

S. typhimurium, other: TA 1537, TA98, TA1535, TA 100, TA 102

Metabolic activation:

with and without

Genotoxicity:

negative

Remarks:

no substantial increases in revertant colony numbers of any of the five tested strains (with or without metabolic activation)

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Remarks:

tested up to 5000 µg/plate; with and without metabolic activation

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

The plates incubated with the test substance showed normal background growth up to 5000 µg/plate with and without S9 mix in all strains used.

Experiment I (Without Activation)	Mean Revertants Per Plate
	Strain  TA1535	Strain TA1537	Strain TA98	Strain TA100	Strain TA102
Negative Control	14	22	31	99	169
Solvent Control	19	19	26	99	186
1,3-Propanediol
33.3 µg/plate	13	24	25	100	184
100 3 µg/plate	17	23	30	93	164
333.3 µg/plate	21	15	26	102	179
1000 µg/plate	16	23	24	93	188
2500 µg/plate	13	22	26	97	194
5000 µg/plate	12	18	31	105	185
Positive control
Sodium azide(10 µg/plate)	885			619
4-NOPD (10 µg/plate)		62	245
MMS (5 µL/plate)					730

 

Experiment I (With Activation)	Mean Revertants Per Plate
	Strain  TA1535	Strain TA1537	Strain TA98	Strain TA100	Strain TA102
Negative Control	21	28	42	159	259
Solvent Control	18	23	40	158	244
1,3-Propanediol
33.3 µg/plate	16	21	45	163	240
100 3 µg/plate	18	26	45	160	236
333.3 µg/plate	22	29	50	156	255
1000 µg/plate	24	23	47	172	291
2500 µg/plate	20	18	43	156	274
5000 µg/plate	19	24	43	170	255
Positive control (2AA)
2.5 µg/plate	261	90	468	965	755

 

Experiment II (Without Activation)	Mean Revertants Per Plate
	Strain  TA1535	Strain TA1537	Strain TA98	Strain TA100	Strain TA102
Negative Control	7	22	24	79	153
Solvent Control	5	22	20	92	151
1,3-Propanediol
33.3 µg/plate	8	21	20	82	151
100 3 µg/plate	4	22	21	81	159
333.3 µg/plate	4	21	20	85	163
1000 µg/plate	7	22	19	69	151
2500 µg/plate	9	23	25	68	164
5000 µg/plate	9	19	25	83	148
Positive control
Sodium azide(10 µg/plate)	263			237
4-NOPD (10 µg/plate)		138	150
MMS (5 µL/plate)					874

 

Experiment II (With Activation)	Mean Revertants Per Plate
	Strain  TA1535	Strain TA1537	Strain TA98	Strain TA100	Strain TA102
Negative Control	10	30	20	137	185
Solvent Control	8	32	22	128	154
1,3-Propanediol
33.3 µg/plate	7	32	24	131	176
100 3 µg/plate	8	27	17	136	211
333.3 µg/plate	8	32	22	139	189
1000 µg/plate	14	31	25	125	182
2500 µg/plate	9	31	23	143	184
5000 µg/plate	14	30	20	126	195
Positive control (2AA)
2.5 µg/plate	39	143	197	406	611

Conclusions:

Under the conditions of this study, propane-1,3-diol was non-mutagenic in a Salmonella typhimurium mutation assay in strains TA 1535, TA 1537, TA 98, TA 100 and TA 102 in the presence and absence of an exogenous metabolic activation system.

Executive summary:

The study was performed to investigate the potential of 1,3-propanediol to induce gene mutations according to the plate incorporation test (experiment I) and the pre-incubation test (experiment II) using the Salmonella typhimurium strains TA1535, TA1537, TA98, TA100, and TA102. The assay was performed in two independent experiments both with and without liver microsomal activation. Each concentration, including the controls, was tested in triplicate. Concentrations tested included: 0, 33.3, 100, 333.3, 1000, 2500, and 5000 µg/plate. No toxic effects occurred in the test groups with or without metabolic activation. The plates incubated with the test article showed normal background growth up to 5000 µg/plate with and without S9 mix in all strains used.

 

No substantial increases in revertant colony numbers of any of the five tester strains were observed following treatment with 1,3-propanediol at any dose level, either in the presence or absence of metabolic activation. There was also no tendency of higher mutation rates with increasing concentrations. The test substance did not induce gene mutations by base pair changes or frameshifts in the genome of the strains used. Therefore, 1,3-propanediol was considered to be non-mutagenic in this Salmonella typhimurium reverse mutation assay.

Reference 2

Endpoint:

in vitro cytogenicity / chromosome aberration study in mammalian cells

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

ACC PDO panel assigned reliability of 1 PDO from chemical process

Qualifier:

according to guideline

Guideline:

OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.5375 - In vitro Mammalian Chromosome Aberration Test

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: Commission Directive 87/302/EEC, EEC Part B

Deviations:

no

GLP compliance:

yes

Type of assay:

in vitro mammalian chromosome aberration test

Species / strain / cell type:

Chinese hamster lung fibroblasts (V79)

Details on mammalian cell type (if applicable):

- Type and identity of media: 9% Dulbecco's modified eagles medium
- Properly maintained: yes
- Periodically checked for Mycoplasma contamination: at study initiation
- Periodically checked for karyotype stability: working stocks not used beyond passage 20
- Periodically "cleansed" against high spontaneous background: no data

Metabolic activation:

with and without

Metabolic activation system:

Aroclor 1254-induced rat liver S9

Test concentrations with justification for top dose:

625, 1250, 2500, and 5000 µg/mL

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: water
- Justification for choice of solvent/vehicle: based on the solubility of the test substance and compatibility with the target cells. The test substance was soluble in water at a concentration of 50 mg/mL, the maximum concentration tested.

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

Remarks:

water

True negative controls:

no

Positive controls:

yes

Positive control substance:

mitomycin C

Remarks:

used in the non-activated test system

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

Remarks:

water

True negative controls:

no

Positive controls:

yes

Positive control substance:

cyclophosphamide

Remarks:

used in the S9 activated test system

Details on test system and experimental conditions:

METHOD OF APPLICATION: in medium

DURATION
- Preincubation period: 16-24 hours
- Exposure duration: 4 hours for metabolic activated assay, and 4 and 20 hours for non-activated assay
- Expression time (cells in growth medium): up to 16 hours for 4 hour treatments
- Fixation time (start of exposure up to fixation or harvest of cells): 20 hours

SPINDLE INHIBITOR (cytogenetic assays): Colcemid (two hours prior to scheduled cell harvest)
STAIN (for cytogenetic assays): 5% Giemsa

NUMBER OF CELLS EVALUATED: 5e5 cells/25cm2

DETERMINATION OF CYTOTOXICITY
- Method: mitotic index

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

Evaluation criteria:

The use of the V79 cell line was based on the sponsor's request, and this cell line is not commonly used in chromosome aberration studies. The frequency of cells with structural chromosome aberrations in the solvent control must be ≤6%. The percentage of cells with chromosome aberrations in the positive control must be statistically increased (p≤0.05, Fisher's exact test) relative to the solvent control.

Statistics:

Statistical analysis of the percent aberrant cells was performed using the Fisher's exact test. Fisher's test was used to compare pairwise the percent aberrant cells of each treatment group with that of the solvent control. In the event of a positive Fisher's test at any test substance dose level, the Cochran-Armitage test was used to measure dose-responsiveness. All conclusions were based on sound scientific basis; however, as a guide to interpretation of the data, the test substance is considered to induce a positive response when the percentage of cells with aberrations was increased in a dose-responsive manner with one or more concentrations being statistically significant (p≤0.05). If the test substance did not demonstrate a statistically significant increase in aberrations, it was concluded to be negative.

Species / strain:

Chinese hamster lung fibroblasts (V79)

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

some at the highest conc. in the non-activated 20 hr exposure group (mitotic index-9% reduced relative to control)

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Remarks on result:

other: all strains/cell types tested

Treatment (µg/mL)	S9	Treatment Time (hr)	Mean Mitotic Index (%)	Cells Scored	Aberrations Per Cell (Mean)	Cells with Aberrations
						Numerical (%)	Structural (%)
Water	-	4	16.4	200	0.000	1.5	0.0
1250	-	4	15.1	200	0.005	2.5	0.5
2500	-	4	14.9	200	0.000	0.5	0.0
5000	-	4	14.0	200	0.010	2.0	1.0
MMC	-	4	12.1	200	0.120	1.5	11.5
Water	+	4	10.9	200	0.000	1.0	0.0
1250	+	4	13.3	200	0.000	0.5	0.0
2500	+	4	9.8	200	0.000	1.5	0.0
5000	+	4	7.7	200	0.000	1.5	0.0
CP	+	4	6.6	100	0.600	1.5	21.0
Water	-	20	17.1	200	0.005	1.5	0.5
1250	-	20	16.4	200	0.000	1.0	0.0
2500	-	20	17.3	200	0.000	1.5	0.0
5000	-	20	15.6	200	0.005	2.0	0.5
MMC	-	20	6.1	100	0.650	1.5	29.0

 

Conclusions:

The test substance was concluded to be negative for the induction of structural and numerical chromosome aberrations in the presence or absence of activation.

Executive summary:

The test substance was tested in the chromosome aberration assay using Chinese hamster V79 cells in both the presence and absence of an Aroclor-induced S9 activation system in order to evaluate the clastogenic potential of the test substance. The maximum dose tested was 5000 µg/mL. The cells were treated for 4 and 20 hours in the non-activated test system and for 4 hours in the S9 activated test system, and all cells were harvested at 20 hours after treatment initiation. The test substance was soluble in treatment medium at all dose levels in all treatment conditions. The test substance was concluded to be negative for the induction of structural and numerical chromosome aberrations in Chinese hamster V79 cells in the presence and absence of Aroclor-induced rat liver S9.

Reference 3

Endpoint:

in vitro gene mutation study in mammalian cells

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

ACC PDO panel assigned reliability of 1. PDO from chemical process.

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

Second addendum to the OECD Guideline for testing of Chemicals, Section 4, No; 476, adopted April 4, 1984, "In Vitro Mammalian Cell Gene Mutation Assay"

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.5300 - In vitro Mammalian Cell Gene Mutation Test

Version / remarks:

pp. 717-720 (7-1-86 Edition)

Qualifier:

according to guideline

Guideline:

other: EEC directive 87/302, L 133, p. 61-63

GLP compliance:

yes

Type of assay:

mammalian cell gene mutation assay

Target gene:

HPRT (hypoxanthine-guanine phosphoribosyl transferase)

Species / strain / cell type:

Chinese hamster lung fibroblasts (V79)

Details on mammalian cell type (if applicable):

- Type and identity of media: Cell culture medium: MEM (minimal essential medium; SEROMED; D-122247 Berlin) supplemented with 10% fetal calf serum (FCS; Boehringer Mannheim, D-68261 Mannheim, F.R.G.).
- Properly maintained: yes
- Periodically checked for Mycoplasma contamination: yes
- Periodically checked for karyotype stability: yes
- Stored in liquid nitrogen; addition of HAT-medium before freezing in order to depress the level of spontaneous mutants
- For the selection of mutants, the medium is supplemented with 11 µg/mL thioguanine (6TG)

Metabolic activation:

with and without

Metabolic activation system:

S-9 mix

Test concentrations with justification for top dose:

with and without S9 mix: 250; 1000; 2500; 5000 µg/mL

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Remarks:

culture medium MEM without FCS

Positive controls:

yes

Positive control substance:

ethylmethanesulphonate

Remarks:

dissolved in nutrient medium (MEM without FCS); without metabolic activation

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Positive control substance:

7,12-dimethylbenzanthracene

Remarks:

dissolved in dimethylsulfoxide with final DMSO concentration in nutrient medium of 1%; with metabolic activation

Details on test system and experimental conditions:

METHOD OF APPLICATION: in cell suspension

DURATION
- Preincubation period: 24h, seeded in MEM with 10% FCS (complete medium).
- Exposure duration: 4h in serum-free medium containing the test substance, either without S9 mix or with 50 µg/mL S9 mix.
- Expression time (cells in growth medium): day 1-9
- Selection time (if incubation with a selection agent): mutant selection medium day 9 - day 18
- Cloning efficiency: day 9 - day 16
- Fixation time (start of exposure up to fixation or harvest of cells): 18 days for mutant selection; 16 days for cloning efficiency determinations

SELECTION AGENT (mutation assays): thioguanine (6TG)
STAIN (for cytogenetic assays): 10% methylene blue in 0.01% KOH solution.

NUMBER OF REPLICATIONS:
determination of toxicity: in duplicate per experimental point.
determination of mutation rates: 1 flask per experimental point.

NUMBER OF CELLS EVALUATED:
determination of toxicity: day 1: about 500 cells in 5 mL medium.
determination of mutation rates: day 1: 1.5e6 cells in 30 mL medium.

DETERMINATION OF CYTOTOXICITY
- Method: concentration-related cloning efficiency (day 9: fixation and staining of colonies).
Data of this study comply with the acceptability criteria of this assay:
a) the numbers of mutant colonies per 10e6 cells found in the negative and/or solvent controls fall within the laboratory historical control range data: 0-45 mutants/10e6 cells.
b) the positive control substances must produce a significant increase in mutant colony frequencies.
c) the colonic efficiency (absolute value) of the negative and/or solvent controls must exceed 50% .

Evaluation criteria:

A test article was classified as positive if it induced either a concentration-related increase of the mutant frequency or a reproducible and positive response for one of the test points.
A test article producing neither a concentration-related increase of the mutant frequency nor a reproducible positive response at any of the test points was considered non-mutagenic in this system.

Statistics:

Biometry: Since the distribution of mutant cells does not follow known statistical models, an adequate statistical method was not available

Species / strain:

mammalian cell line, other: Chinese Hamster V79 cells

Metabolic activation:

with and without

Genotoxicity:

negative

Remarks:

no relevant increase in mutant colony numbers in two independent experiments with and without metabolic activation

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

low survival at highest test concentration (in the without metabolic activation assay)

Untreated negative controls validity:

valid

Positive controls validity:

valid

Experiment I

µg/mL	S9	Mean number of mutant colonies per flask	Mutant colonies per 10e6 cells
Solvent Control (with medium)	-	3.2	10.3
Positive Control (with)	-	79.4	344.8
Test Article 250	-	1.0	3.8
1000	-	2.6	8.7
2500	-	0.2	0.7
5000	-	0.8	2.6
Solvent Control (with medium)	+	0.2	0.7
Solvent Control (with DMSO)	+	0.6	1.7
Positive Control (with DMBA)	+	118.8	526.2
Test Article 250	+	0.4	1.2
1000	+	0.6	2.2
2500	+	2.4	9.8
5000	+	0.8	3.0

 

Experiment II

µg/mL	S9	Mean number of mutant colonies per flask	Mutant colonies per 10e6 cells
Solvent Control (with medium)	-	5.6	21.8
Positive Control (with)	-	131.8	802.7
Test Article 250	-	0.8	3.0
1000	-	1.4	5.4
2500	-	3.2	10.8
5000	-	4.0	14.0
Solvent Control (with medium)	+	1.0	2.8
Solvent Control (with DMSO)	+	4.0	1.1
Positive Control (with DMBA)	+	120.2	379.1
Test Article 250	+	2.8	7.7
1000	+	1.0	3.2
2500	+	1.4	4.1
5000	+	1.2	3.1

 

Conclusions:

Under the conditions of this study, propane-1,3-diol did not induce gene mutations at the HPRT locus in Chinese hamster V79 cells either with or without metabolic activation.

Executive summary:

The study was performed to investigate the potential of 1,3-propanediol to induce gene mutations at the HPRT locus in V79 cells of the Chinese hamster. The assay was performed in two independent experiments, using identical procedures, both with and without liver microsomal activation. The following concentrations were tested: 0, 250, 1000, 2500, and 5000 µg/mL. Up to the highest concentration tested, no relevant increases in mutant colony numbers was obtained in the two independent experiments. Therefore, it was concluded that the test substance did not induce gene mutations at the HPRT locus in V79 cells and is considered to be non-mutagenic in this HPRT assay.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Genetic toxicity in vivo

Description of key information

In Vivo (Cytogenic effects - mammalian): EU Method B.12; Mouse micronucleus. Negative. Reliability = 1.

Link to relevant study records

Reference

Endpoint:

in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

ACC PDO panel assigned reliability of 1. PDO from chemical process.

Qualifier:

according to guideline

Guideline:

other: EEC Directive 92/69/EEC Part B: Methods for the Determination of Toxicity, B.12. Micronucleus Test

GLP compliance:

yes

Type of assay:

mammalian erythrocyte micronucleus test

Species:

mouse

Strain:

other: Hsd/Win: NMRI

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Harlan-Winkelmann GmbH
- Age at study initiation: approximately 6 weeks
- Weight at study initiation: males: 30-36 g; females: 25-30 g
- Assigned to test groups randomly: yes (computer generated random numbers)
- Fasting period before study: 16 hours before treatment.
- Housing: Macrolon cages, type II, 1 animal per cage.
- Diet: ad libitum
- Water:ad libitum
- Acclimation period: 6 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21.0-21.5°C
- Humidity (%): 50-58%
- Photoperiod (hrs dark / hrs light): 12 hours light, 12 hours dark

Route of administration:

oral: gavage

Vehicle:

- Vehicle(s)/solvent(s) used: The test material was freshly diluted with aqua ad iniectabilia (Ampuwa, batch No. EL 1009, supplied by Fresenius AG, D-61343 Bad Homburg v d Hohe)
- Administration volume: 10.0 mL/kg b.w.

Frequency of treatment:

single administration

Post exposure period:

24 and 48 hours

Dose / conc.:

1 000 mg/kg bw/day (nominal)

Dose / conc.:

1 470 mg/kg bw/day (nominal)

Dose / conc.:

2 150 mg/kg bw/day (nominal)

No. of animals per sex per dose:

Main test: 28 (14 males and 14 females) at 2150 mg/kg bw
Repetition Test: 12 (6 males and 6 females) each at 1000, 1470 and 2150 mg/kg bw

Control animals:

other: Main Test: Physiological saline solution; 12 males + 12 females. Repetition Test: 6 males + 6 females.

Positive control(s):

Main Test positive control: cyclophosphamide (31.6 mg/kg bw) in physiological saline solution; 6 males + 6 females.
Repeat Test positive control: cyclophosphamide (31.6 mg/kg bw) in physiological saline solution; 6 males + 6 females.

Tissues and cell types examined:

clinical observation; bone marrow

Details of tissue and slide preparation:

CRITERIA FOR DOSE SELECTION: Range finding study identified that 4640 mg/kg bw caused severe toxicity symptoms and death, and at 3160 mg/kg bw distinct toxicity symptoms but no death was observed. Test guideline considers 2000 mg/kg bw to be the maximum single (limit) dose to be administered.

DETAILS OF SLIDE PREPARATION: Mice were sacrificed by CO2 overdose. Both femurs removed from each mouse and the bone marrow cells flushed into a labeled centrifuge tube with approx. 1.5 mL of fetal calf serum. Centrifuged at approx. 180 x g for 5 minutes. Supernatant serum was discarded and bone marrow cells suspended upon a thin layer of serum. A small drop of the marrow serum suspension was smeared on a slide and allowed to dry overnight.

METHOD OF ANALYSIS: Microscopical evaluation; 1000 polychromatic erythrocytes per animal were scored for the incidence of polychromatic erythrocytes with micronuclei. Calculation of PCE/NCE, based on 1000 erythrocytes (PCE+NCE) scored per slide.

Evaluation criteria:

If a test material produced neither a statistically significant and reproducible positive response nor a dose related statistically significant response at any one of the test points compared to the negative control group, it is considered non-mutagenic in this system.

Statistics:

POISSON test

Sex:

male/female

Genotoxicity:

negative

Remarks:

non-mutagenic

Toxicity:

no effects

Negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

Under the experimental conditions reported, 1,3-Propanediol is considered not to induce chromosomal mutations in mice by damage to the chromosomes or the miotic apparatus at 24 or 48 hour intervals after the animals had received a single oral dose of 2150 mg/kg body weight. However, a weak clastogenic effect cannot be totally excluded due to a statistically significant increase in micronucleated polychromatic erythrocytes at the 48 hour sampling time when calculations were performed for males and females combined. According to the test guideline, a second test (repetition test) was performed to examine the reproducibility of the statistically significant response at the 48 hour sampling time in males and females combined and to assess a possible dose relation in the increase of micronucleated polychromatic erythrocytes. These responses could not be verified during the repetition test using two additional dose levels and is therefore considered to be an incidental finding.

Therefore, 1,3-Propanediol is non-mutagenic in the reported in vivo mouse micronucleus test.

	Mean PCEs with MN (Males)		Mean PCEs with MN (Females)
Main Test	24-Hours	48-Hours	24-Hours	48-Hours
Negative Control	1.4	1.2	1.4	1.1
2150 mg/kg	1.0	2.2	1.8	1.5
Positive Control	13.8		14.7
Repetition Test
Negative Control		2.0		2.0
1000 mg/kg		0.8		1.4
1470 mg/kg		1.0		1.0
2150 mg/kg		2.0		1.2
Positive Control		6.6		6.3

Conclusions:

1,3-Propanediol is non-mutagenic in the reported in vivo mouse micronucleus test.

Executive summary:

This in vivo micronucleus test in mice was performed to assess the potential mutagenic activity, induced by 1,3-propanediol through damage to the chromosomes of to the mitotic apparatus. The main test was performed with three groups, the negative control, the positive control, and the test material group (2150 mg/kg). Half the mice were sacrificed at 24 hours after treatment and the remaining mice were sacrificed 48 hours after treatment. Bone marrow was removed from the femurs for examination. All animals of the positive control group were sacrificed 24 hours after administration. One thousand polychromatic erythrocytes (PCE) per animal were scored for micronuclei. The ratio of polychromatic to normochromatic erythrocytes (NCE) was used to assess the toxicity of the test material. No test substance-related clinical observations were observed in the mice. No mortality was observed. Under the experimental conditions reported, 1,3-Propanediol is considered not to induce chromosomal mutations in mice by damage to the chromosomes or the miotic apparatus at 24 or 48 hour intervals after the animals had received a single oral dose of 2150 mg/kg body weight. However, a weak clastogenic effect cannot be totally excluded due to a statistically significant increase in micronucleated polychromatic erythrocytes at the 48 hour sampling time when calculations were performed for males and females combined. 

 

According to the test guideline, a second test (repetition test) was performed to examine the reproducibility of the statistically significant response at the 48 hour sampling time in males and females combined and to assess a possible dose relation in the increase of micronucleated polychromatic erythrocytes. Dose levels of 1000, 1470, and 2150 were used in the repetition test. The responses of the first test could not be verified during the repetition test using two additional dose levels and is therefore considered to be an incidental finding. Therefore, 1,3-Propanediol is non-mutagenic in the reported in vivo mouse micronucleus test.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Additional information

This substance has been tested in 4 in vitro genetic toxicity studies involving bacterial and mammalian cells and mutagenicity and clastogenicity endpoints. Additionally, the substance has been tested in the mouse by the oral route of exposure for genetic toxicity. In bacterial cells, this substance was not mutagenic based on a lack of significant effects to reserve mutations. In mammalian cells, this substance was also not mutagenic for forward mutations at the HPGRT locus. This substance was tested for clastogenicity in V79 cells resulting in a significant increase in chromosome aberrations (CA) in the absence of metabolic activation at the highest scorable dose (the highest dose tested was not scored due to cytotoxicity). The result was not dose-related. It was negative in the presence of metabolic activation. Potential for poor metaphase preparation being related to the positive result could not be excluded; therefore, the study was repeated with the same protocol. The second study did not show any significant increase in CA at the same and higher dose. This substance was also negative for genetic toxicity in the mouse micronucleus test. Taken together, these results support a low potential of this substance to adversely affect genetic material, and it is concluded that it is not genetically toxic.

Justification for classification or non-classification

The test substance did not produce mutagenicity when evaluated in cell culture or laboratory animals. The substance does not need to be classified for mutagenicity according to EU Classification, Labelling and Packaging of Substances and Mixtures (CLP) Regulation (EC) No. 1272/2008.