Trifluoro(pentafluoroethoxy)ethylene

Genetic toxicity: in vitro

Administrative data

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

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1997

Report date:

1997

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

yes

Remarks:

Refer any other information on material and methods section

Test guideline 2

Qualifier:

according to guideline

Guideline:

OECD Guideline 472 (Genetic Toxicology: Escherichia coli, Reverse Mutation Assay)

Deviations:

yes

Remarks:

Refer any other information on material and methods section

Test guideline 3

Qualifier:

according to guideline

Guideline:

EPA OTS 798.5265 (The Salmonella typhimurium Bacterial Reverse Mutation Test)

Deviations:

yes

Remarks:

Refer any other information on material and methods section

Test guideline 4

Qualifier:

according to guideline

Guideline:

EPA OTS 798.5100 (Escherichia coli WP2 and WP2 UVRA Reverse Mutation Test)

Deviations:

yes

Remarks:

Refer any other information on material and methods section

Test guideline 5

Qualifier:

according to guideline

Guideline:

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

Deviations:

yes

Remarks:

Refer any other information on material and methods section

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Trifluoro(pentafluoroethoxy)ethylene

EC Number:

234-018-7

EC Name:

Trifluoro(pentafluoroethoxy)ethylene

Cas Number:

10493-43-3

Molecular formula:

C4F8O

IUPAC Name:

1,1,2-trifluoro-2-(1,1,2,2,2-pentafluoroethoxy)ethene

Test material form:

gas

Details on test material:

- Purity: 99%

Method

Target gene:

Histidine biosynthetic enzymes

Species / strain

Species / strain 1

Species / strain / cell type:

other: S. typhimurium TA100, TA1535, TA97a, and TA98

Species / strain 2

Species / strain / cell type:

E. coli WP2 uvr A pKM 101

Metabolic activation:

with and without

Metabolic activation system:

Aroclor 1254 Liver homogenate activation system (S9 mix)

Test concentrations with justification for top dose:

Trial I (average measured concentrations): 0.49, 1.08, 1.27, 1.74, and 1.92% without S9; 0.50, 1.05, 1.29, 1.73, and 1.90% with S9
Trial II (average measured concentrations): 0.62, 0.92, 1.27, 1.86, and 2.08% without S9; 0.61, 0.87, 1.19, 1.86, and 2.10% with S9

The highest concentration was an atmosphere of 1.8%. This was approximately 60% of the lower explosive limit indicated by the sponsor.

Vehicle / solvent:

Dilution of the test substance with air was conducted immediately prior to treatment period. Gas chromatographic analysis was used to confirm the concentration of test atmospheres using the following conditions:
- Instrument: Hewlett-Packard 5890 Series II, gas chromatograph
- Column: 1/8" x 20"; 3% OV 17; 3% Chromosorb®; stainless steel
- Detector: Flame Ionization
- Carrier gas: Helium: Trial 1: 21.5 mL/min Trial 2: 21.0 mL/min;
- Detector gases: Hydrogen: Trial 1: 30.3 mL/min, Trial 2: 31.0 mL/min; Air: Trial 1: 330 mL/min, Trial 2: 299 mL/min
- Oven/Column temperature: 85°C, isothermal
- Injector port temperature: 68°C
- Detector temperature: 250°C
- Injection volume: 10 µL

Controls

Untreated negative controls:

yes

Remarks:

Air

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

Positive controls:

yes

Positive control substance:

2-nitrofluorene

sodium azide

methylmethanesulfonate

other: 2-aminoanthracene, ICR 191 acridine

Details on test system and experimental conditions:

This study consisted of two trials with and without activation. Three replicates were plated for each tester strain, test concentration, and condition. Positive controls, solvent controls, and air controls were included in all assays. Treatments with activation were conducted by adding 0.1 mL of solvent or positive control, 0.5 mL of S9 mix, and 0.1 mL of an overnight culture containing 1 x 10e8 bacteria to 2 mL of top agar [0.6% agar (w/v) and 0.6% NaCl (w/v)] supplemented with 0.05 mM L-histidine and 0.05 mM D-biotin for S. typhimurium strains or 0.05 mM L-tryptophan for the E. coli strain. These components were mixed and poured onto a plate containing approximately 25 mL of Davis minimal agar with dextrose (minimal agar plates, purchased from MOLTOX™). Treatments in the absence of the metabolic activation system were identical to those with activation with the exception that the S9 mix was replaced with 0.5 mL of sterile phosphate buffered saline.

Plates were exposed to dilutions of the test gas after being placed on stainless steel racks specially designed to fit in 6-liter glass chambers. Chambers were equipped with Teflon® stopcocks and Viton® O-ring gaskets. As gases, the test substance and filtered air flows were regulated using individual rotameters, and mixed prior to entry into chambers. A flow rate of approximately 10 L/minute for approximately 5 minutes was used to create at least 5 volume changes to occur within the chambers to insure homogenous concentrations. Chambers were closed and 2-4 samplings of each chamber were taken and analyzed by gas chromatography to determine the initial concentration of the test substance. Chamber concentrations were determined by comparison of integrated peak areas to test substance standards by the analytical conditions previously described. Chambers were placed into an incubator at approximately 37°C for approximately 48 hours. Chambers were again sampled and analyzed to determine the ending test substance concentration. Chambers were flushed with at least 5-10 chamber volumes of filtered air and then refrigerated until counted. Plates were then removed for an evaluation of background lawns and colony formation.

Evaluation criteria:

A test substance was classified as positive when: (1) the average number of revertants in any strain at any test substance concentration studied was at least two times greater than the average number of revertants in the negative control; and (2) there was a positive dose-response relationship in that same strain.

A test substance was classified as negative when either: (1) there were no test substance concentrations with an average number of revertants which was at least two times greater than the average number of revertants in the negative control; or (2) there was no positive dose-response relationship.

Results not meeting these criteria for positive or negative assessments were evaluated using scientific judgment.

Statistics:

For each tester strain, the average number of revertants and the standard deviation at each concentration with and without S9 activation were calculated.

Results and discussion

Test results

Test results 1

Key result

Species / strain:

other: S. typhimurium TA100, TA1535, TA97a, and TA98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 2

Key result

Species / strain:

E. coli WP2 uvr A pKM 101

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

In the initial trial, end-treatment sampling for one concentration (1.73%) was not performed; therefore, the indicated exposure concentration represents the average of samples taken at the beginning of the exposure period. Analytical results indicated the test substance was stable under the conditions of the study, therefore, results in the one instance when post-treatment analytical was not performed were not rejected. All valid criteria were met.

Any other information on results incl. tables

Mutagenic Activity of the Test Substance in Strain TA100 Trial 1
Average Concentration (%) Mean ± S.D.	Revertants			Average (S.D.)	Observations
	Plate 1	Plate 2	Plate 3
Without Activation
Filtered Air	80	81	78	80 (2)	T0,P0
0.49 ± 0.03	77	80	79	79 (2)	T0,P0
1.08 ± 0.03	82	80	80	81 (1)	T0,P0
1.27 ± 0.02	73	78	70	74 (4)	T0,P0
1.74 ± 0.03	77	72	79	76 (4)	T0,P0
1.92 ± 0.05	79	81	83	81 (2)	T0,P0
DMSO (Solvent) Control 0 µg/plate	66	71	72	70 (3)	T0,P0
NAAZ (positive control) 2 µg/plate	662	805	769	745 (74)	N
With Activation
Filtered Air	78	76	86	80 (5)	T0,P0
0.50 ± 0.04	78	85	76	80 (5)	T0,P0
1.05 ± 0.03	77	81	82	80 (3)	T0,P0
1.29 ± 0.07	74	79	81	78 (4)	T0,P0
1.73 ± 0.04*	70	85	83	79 (8)	T0,P0
1.90 ± 0.02	76	80	73	76 (4)	T0,P0
DMSO (Solvent) Control 0 µg/plate	82	79	75	79 (3)	T0,P0
2AA (positive control) 1 µg/plate	583	523	566	557 (31)	N
*Starting concentration – determination of the ending exposure concentration was not performed.

 

Mutagenic Activity of the Test Substance in Strain TA100 Trial 2
Average Concentration (%) Mean ± S.D.	Revertants			Average (S.D.)	Observations
	Plate 1	Plate 2	Plate 3
Without Activation
Filtered Air	162	163	159	161 (2)	T0,P0
0.62 ± 0.08	159	161	175	165 (9)	T0,P0
0.92 ± 0.05	164	154	167	162 (7)	T0,P0
1.27 ± 0.03	154	168	163	162 (7)	T0,P0
1.86 ± 0.17	164	153	166	161 (7)	T0,P0
2.08 ± 0.28	150	163	161	158 (7)	T0,P0
DMSO (Solvent) Control 0 µg/plate	149	149	157	152 (5)	T0,P0
NAAZ (positive control) 2 µg/plate	853	854	885	864 (18)	N
With Activation
Filtered Air	148	160	164	157 (8)	T0,P0
0.61 ± 0.10	168	166	160	165 (4)	T0,P0
0.87 ± 0.03	163	154	170	162 (8)	T0,P0
1.19 ± 0.04	151	156	138	148 (9)	T0,P0
1.86 ± 0.10	166	184	177	176 (9)	T0,P0
2.10 ± 0.35	149	157	167	158 (9)	T0,P0
DMSO (Solvent) Control 0 µg/plate	157	172	164	164 (7)	T0,P0
2AA (positive control) 1 µg/plate	1593	1471	1571	1545 (65)	N

 

Mutagenic Activity of the Test Substance in Strain TA1535 Trial 1
Average Concentration (%) Mean ± S.D.	Revertants			Average (S.D.)	Observations
	Plate 1	Plate 2	Plate 3
Without Activation
Filtered Air	11	11	12	11 (1)	T0,P0
0.49 ± 0.03	16	7	13	12 (5)	T0,P0
1.08 ± 0.03	11	16	12	13 (3)	T0,P0
1.27 ± 0.02	13	19	9	14 (5)	T0,P0
1.74 ± 0.03	11	10	11	11 (1)	T0,P0
1.92 ± 0.05	10	15	11	12 (3)	T0,P0
DMSO (Solvent) Control 0 µg/plate	12	11	15	13 (2)	T0,P0
NAAZ (positive control) 2 µg/plate	647	538	594	593 (55)	N
With Activation
Filtered Air	21	18	21	20 (2)	T0,P0
0.50 ± 0.04	21	19	22	21 (2)	T0,P0
1.05 ± 0.03	20	16	20	19 (2)	T0,P0
1.29 ± 0.07	15	9	12	12 (3)	T0,P0
1.73 ± 0.04*	14	15	9	13 (3)	T0,P0
1.90 ± 0.02	14	17	20	17 (3)	T0,P0
DMSO (Solvent) Control 0 µg/plate	9	11	14	11 (2)	T0,P0
2AA (positive control) 2 µg/plate	355	339	277	324 (41)	N
*Starting concentration – determination of the ending exposure concentration was not performed.

 

Mutagenic Activity of the Test Substance in Strain TA1535 Trial 2
Average Concentration (%) Mean ± S.D.	Revertants			Average (S.D.)	Observations
	Plate 1	Plate 2	Plate 3
Without Activation
Filtered Air	8	14	12	11 (3)	T0,P0
0.62 ± 0.08	12	8	9	10 (2)	T0,P0
0.92 ± 0.05	13	9	15	12 (3)	T0,P0
1.27 ± 0.03	12	11	10	11 (1)	T0,P0
1.86 ± 0.17	11	12	10	11 (1)	T0,P0
2.08 ± 0.28	7	8	8	8 (1)	T0,P0
DMSO (Solvent) Control 0 µg/plate	10	9	8	9 (1)	T0,P0
NAAZ (positive control) 2 µg/plate	605	587	653	615 (34)	N
With Activation
Filtered Air	8	10	13	10 (3)	T0,P0
0.61 ± 0.10	9	12	8	10 (2)	T0,P0
0.87 ± 0.03	12	14	13	13 (1)	T0,P0
1.19 ± 0.04	12	8	7	9 (3)	T0,P0
1.86 ± 0.10	11	9	8	9 (2)	T0,P0
2.10 ± 0.35	8	10	10	9(1)	T0,P0
DMSO (Solvent) Control 0 µg/plate	10	12	12	11 (1)	T0,P0
2AA (positive control) 2 µg/plate	307	285	277	290 (16)	N

 

Mutagenic Activity of the Test Substance in Strain TA97a Trial 1
Average Concentration (%) Mean ± S.D.	Revertants			Average (S.D.)	Observations
	Plate 1	Plate 2	Plate 3
Without Activation
Filtered Air	86	102	82	90 (11)	T0,P0
0.49 ± 0.03	83	87	83	84 (2)	T0,P0
1.08 ± 0.03	91	83	82	85 (5)	T0,P0
1.27 ± 0.02	77	70	69	72 (4)	T0,P0
1.74 ± 0.03	94	83	72	83 (11)	T0,P0
1.92 ± 0.05	81	81	81	81 (0)	T0,P0
DMSO (Solvent) Control 0 µg/plate	86	82	77	82 (4)	T0,P0
ICR 191 (positive control) 2 µg/plate	1253	1211	1236	1233 (21)	N
With Activation
Filtered Air	100	95	101	99 (3)	T0,P0
0.50 ± 0.04	86	92	85	88 (4)	T0,P0
1.05 ± 0.03	87	84	92	88 (4)	T0,P0
1.29 ± 0.07	83	90	87	87 (4)	T0,P0
1.73 ± 0.04*	82	85	83	83 (2)	T0,P0
1.90 ± 0.02	82	79	74	78 (4)	T0,P0
DMSO (Solvent) Control 0 µg/plate	89	107	94	97 (9)	T0,P0
2AA (positive control) 1 µg/plate	530	541	612	561 (45)	N
*Starting concentration – determination of the ending exposure concentration was not performed.

 

Mutagenic Activity of the Test Substance in Strain TA197a Trial 2
Average Concentration (%) Mean ± S.D.	Revertants			Average (S.D.)	Observations
	Plate 1	Plate 2	Plate 3
Without Activation
Filtered Air	85	96	88	90 (6)	T0,P0
0.62 ± 0.08	77	87	85	83 (5)	T0,P0
0.92 ± 0.05	68	73	86	76 (9)	T0,P0
1.27 ± 0.03	73	79	81	78 (4)	T0,P0
1.86 ± 0.17	79	68	80	76 (7)	T0,P0
2.08 ± 0.28	62	81	72	72 (10)	T0,P0
DMSO (Solvent) Control 0 µg/plate	93	95	79	89 (9)	T0,P0
ICR 191 (positive control) 2 µg/plate	1247	1179	1050	1159 (100)	N
With Activation
Filtered Air	97	89	79	88 (9)	T0,P0
0.61 ± 0.10	77	80	75	77 (3)	T0,P0
0.87 ± 0.03	82	84	97	88 (8)	T0,P0
1.19 ± 0.04	81	70	73	75 (6)	T0,P0
1.86 ± 0.10	99	88	97	95 (6)	T0,P0
2.10 ± 0.35	64	74	70	69 (5)	T0,P0
DMSO (Solvent) Control 0 µg/plate	95	96	105	99 (5)	T0,P0
2AA (positive control) 1 µg/plate	764	720	737	740 (22)	N

 

Mutagenic Activity of the Test Substance in Strain TA98 Trial 1
Average Concentration (%) Mean ± S.D.	Revertants			Average (S.D.)	Observations
	Plate 1	Plate 2	Plate 3
Without Activation
Filtered Air	11	17	9	12 (4)	T0,P0
0.49 ± 0.03	16	15	10	14 (3)	T0,P0
1.08 ± 0.03	17	15	13	15 (2)	T0,P0
1.27 ± 0.02	11	16	15	14 (3)	T0,P0
1.74 ± 0.03	14	20	11	15 (5)	T0,P0
1.92 ± 0.05	19	17	12	16 (4)	T0,P0
DMSO (Solvent) Control 0 µg/plate	13	10	9	11 (2)	T0,P0
2NF (positive control) 25 µg/plate	1374	1354	1235	1321 (75)	N
With Activation
Filtered Air	14	16	15	15 (1)	T0,P0
0.50 ± 0.04	14	15	17	15 (2)	T0,P0
1.05 ± 0.03	19	22	21	21 (2)	T0,P0
1.29 ± 0.07	20	22	17	20 (3)	T0,P0
1.73 ± 0.04*	17	16	15	16 (1)	T0,P0
1.90 ± 0.02	15	14	13	14 (1)	T0,P0
DMSO (Solvent) Control 0 µg/plate	12	17	18	16 (3)	T0,P0
2AA (positive control) 2 µg/plate	1214	1174	1346	1245 (90)	N
*Starting concentration – determination of the ending exposure concentration was not performed.

 

Mutagenic Activity of the Test Substance in Strain TA198 Trial 2
Average Concentration (%) Mean ± S.D.	Revertants			Average (S.D.)	Observations
	Plate 1	Plate 2	Plate 3
Without Activation
Filtered Air	20	17	11	16 (5)	T0,P0
0.62 ± 0.08	19	15	21	18 (3)	T0,P0
0.92 ± 0.05	16	20	10	15 (5)	T0,P0
1.27 ± 0.03	9	11	15	12 (3)	T0,P0
1.86 ± 0.17	11	12	22	15 (6)	T0,P0
2.08 ± 0.28	13	13	17	14 (2)	T0,P0
DMSO (Solvent) Control 0 µg/plate	15	19	21	18 (3)	T0,P0
2 NF (positive control) 25 µg/plate	1312	1363	1437	1371 (63)	N
With Activation
Filtered Air	26	30	33	30 (4)	T0,P0
0.61 ± 0.10	21	21	25	22 (2)	T0,P0
0.87 ± 0.03	19	19	16	18 (2)	T0,P0
1.19 ± 0.04	13	14	16	14 (2)	T0,P0
1.86 ± 0.10	19	18	19	19 (1)	T0,P0
2.10 ± 0.35	20	22	21	21 (1)	T0,P0
DMSO (Solvent) Control 0 µg/plate	22	23	20	22 (2)	T0,P0
2AA (positive control) 2 µg/plate	1802	2153	1921	1959 (179)	N

 

Mutagenic Activity of the Test Substance in Strain WP2 uvrA (pKM101) Trial 1
Average Concentration (%) Mean ± S.D.	Revertants			Average (S.D.)	Observations
	Plate 1	Plate 2	Plate 3
Without Activation
Filtered Air	125	155	161	147 (19)	T0,P0
0.49 ± 0.03	153	151	155	153 (2)	T0,P0
1.08 ± 0.03	151	148	153	151 (3)	T0,P0
1.27 ± 0.02	128	125	138	130 (7)	T0,P0
1.74 ± 0.03	145	128	143	139 (9)	T0,P0
1.92 ± 0.05	128	142	136	135 (7)	T0,P0
DMSO (Solvent) Control 0 µg/plate	129	127	124	127 (3)	T0,P0
MMS (positive control) 1000 µg/plate	1968	1947	1851	1922 (62)	N
With Activation
Filtered Air	134	135	141	137 (4)	T0,P0
0.50 ± 0.04	132	127	127	129 (3)	T0,P0
1.05 ± 0.03	124	148	127	133 (13)	T0,P0
1.29 ± 0.07	128	126	131	128 (3)	T0,P0
1.73 ± 0.04*	155	157	134	149 (13)	T0,P0
1.90 ± 0.02	128	126	118	124 (5)	T0,P0
DMSO (Solvent) Control 0 µg/plate	162	175	151	163 (12)	T0,P0
2AA (positive control) 25 µg/plate	2082	1894	1939	1972 (98)	N
*Starting concentration – determination of the ending exposure concentration was not performed.

 

Mutagenic Activity of the Test Substance in Strain WP2 uvrA (pKM101) Trial 2
Average Concentration (%) Mean ± S.D.	Revertants			Average (S.D.)	Observations
	Plate 1	Plate 2	Plate 3
Without Activation
Filtered Air	174	146	150	157 (15)	T0,P0
0.62 ± 0.08	143	163	143	150 (12)	T0,P0
0.92 ± 0.05	133	145	146	141 (7)	T0,P0
1.27 ± 0.03	147	158	139	148 (10)	T0,P0
1.86 ± 0.17	151	129	152	144 (13)	T0,P0
2.08 ± 0.28	134	141	129	135 (6)	T0,P0
DMSO (Solvent) Control 0 µg/plate	141	149	145	145 (4)	T0,P0
MMS (positive control) 25 µg/plate	2114	1783	2199	2032 (220)	N
With Activation
Filtered Air	133	145	152	143 (10)	T0,P0
0.61 ± 0.10	135	138	144	139 (5)	T0,P0
0.87 ± 0.03	126	129	137	131 (6)	T0,P0
1.19 ± 0.04	126	138	129	131 (6)	T0,P0
1.86 ± 0.10	142	138	140	140 (2)	T0,P0
2.10 ± 0.35	143	144	147	145 (2)	T0,P0
DMSO (Solvent) Control 0 µg/plate	181	174	177	177 (4)	T0,P0
2AA (positive control) 25 µg/plate	1805	1645	2057	1836 (208)	N

 

Applicant's summary and conclusion

Conclusions:

No evidence of mutagenic activity was detected in either of the two independent trials.

Executive summary:

The test substance was evaluated for mutagenicity in Salmonella typhimurium strains TA100, TA1535, TA97a, and TA98 and in Escherichia coli strain WP2 uvrA (pKM101) with and without an exogenous metabolic activation system (S9) in accordance with OECD Guideline 471 and 472.

In comparison to negative air and dimethyl sulfoxide (DMSO) controls, tester strains were exposed to the test substance in top agar overlays at average concentrations of 0.49, 1.08, 1.27, 1.74, and 1.92% in the absence of S9, and 0.50, 1.05, 1.29, 1.73, and 1.90% in the presence of S9 for approximately 48 hours. No evidence of mutagenicity was detected in the first trial.

In a second confirmatory trial, plates were exposed to average concentrations of 0.62, 0.92, 1.27, 1.86 and 2.08% in the absence of S9 and 0.61, 0.87, 1.19, 1.86 and 2.10% were tested in the presence of S9 in comparison negative (air) and DMSO controls. No evidence of mutagenicity was detected in the second trial.

Under the conditions of this study, no evidence of mutagenic activity was detected in either of two independent trials. In this study, the test substance was negative.