Neodymium(3+) acetate

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Study conducted on read-across material

Justification for type of information:

Read-across performed with structurally similar substance.

Cross-reference

Reason / purpose for cross-reference:

read-across source

Reference

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

8 March 2010 to 25 March 2010

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Justification for type of information:

Read-across performed with structurally similar substance.

Reason / purpose for cross-reference:

other: read-across target

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:

histidine for S. typhimurium
tryptophan for E.Coli

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

Test concentrations with justification for top dose:

0, 0.15, 0.5, 1.5, 5, 15, 50, 150, 500, 1500 and 5000 µg/plate

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

N-ethyl-N-nitro-N-nitrosoguanidine

Remarks:

for WP2uvrA-, TA100 and TA1535 without activation

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

9-aminoacridine

Remarks:

for TA1537 without activation

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

4-nitroquinoline-N-oxide

Remarks:

for TA98 without activation

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: 2-Aminoanthracene

Remarks:

for WP2uvrA-, TA100, TA1535 and TA1537 with activation

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

benzo(a)pyrene

Remarks:

for TA98 with activation

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Preincubation period: 20 min at 37°C
- Exposure duration: 48 h at 37°C
- Expression time (cells in growth medium): na
- Fixation time (start of exposure up to fixation or harvest of cells): na

NUMBER OF CELLS EVALUATED: 1-9.9 x 10^9

DETERMINATION OF CYTOTOXICITY
- Method: relative total growth

Evaluation criteria:

Dose related increase in revertant frequency over the dose range tested and or a reporducible increase at one or more concentrations in at least one bacterial strain with or without metabolic activiation. Biological relevance of the results will be considered first

Statistics:

Statistical significance will not be the only determining factor for a positive response.

Key result

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Key result

Species / strain:

S. typhimurium TA 1535, TA 1537, TA 98 and TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: A fine, particulate precipitate was noted at 5000 µg/plate, this observation did not prevent the scoring of revertant colonies.

RANGE-FINDING/SCREENING STUDIES:
- The test material was non-toxic to the strains of bacteria used (TA100 and WP2uvrA). The test material formulation and S9-mix used in this experiment were both shown to be sterile.

ADDITIONAL INFORMATION ON CYTOTOXICITY:
- The test material caused no visible reduction in the growth of the bacterial background lawn at any dose level and was, therefore, tested up to the maximum recommended dose level of 5000 µg/plate

POSITIVE CONTROL
- All of the positive control chemicals used in the test induced marked increases in the frequency of revertant colonies thus confirming the activity of the S9-mix and the sensitivity of the bacterial strains.

Table1: Spontaneous Mutation Rates (Concurrent Negative Controls)

EXPERIMENT 1

Number of revertants (mean number of colonies per plate)
Base-pair substitution type						Frameshift type
TA100		TA1535		WP2uvrA		TA98		TA1537
97		20		22		18		7
91	(96)	24	(21)	29	(24)	23	(19)	11	(11)
99		19		20		15		14

EXPERIMENT 2

Number of revertants (mean number of colonies per plate)
Base-pair substitution type						Frameshift type
TA100		TA1535		WP2uvrA		TA98		TA1537
114		21		25		15		14
105	(102)	24	(22)	13	(19)	12	(17)	12	(12)
86		20		19		25		11

 

Table 2: Test Results: Experiment 1: Without Metabolic Activation

Test Period		From: 15 March 2010						To: 18 March 2010
With or without S9-Mix	Test substance concentration (µg/plate)	Number of revertants (mean number of colonies per plate)
		Base-pair substitution type							Frameshift type
		TA100		TA1535		WP2uvrA			TA98		TA1537
-	0	102 82 70	(85) 16.2#	20 13 27	(20) 7.0	31 24 22	(26) 4.7		23 18 16	(19) 3.6	13 9 8	(10) 2.6
-	50	92 89 93	(91) 2.1	16 14 24	(18) 5.3	34 16 25	(25) 9.0		22 15 16	(18) 3.8	13 12 11	(12) 1.0
-	150	110 74 86	(90) 18.3	18 27 14	(20) 6.7	14 22 18	(18) 4.0		18 19 13	(17) 3.2	7 11 8	(9) 2.1
-	500	89 82 93	(88) 5.6	20 16 22	(19) 3.1	24 19 18	(20) 3.2		18 16 16	(17) 1.2	11 9 10	(10) 1.0
-	1500	78 82 82	(81) 2.3	24 18 21	(21) 3.0	30 18 15	(21) 7.9		20 14 18	(17) 3.1	7 10 9	(9) 1.5
-	5000	118 P 71 P 82 P	(90) 24.6	18 P 22 P 19 P	(20) 2.1	20 P 15 P 22 P	(19) 3.6		18 P 18 P 13 P	(16) 2.9	16 P 8 P 9 P	(11) 4.4
Positive controls   S9-Mix   -	Name Concentration (µg/plate) No. colonies per plate	ENNG		ENNG		ENNG			4NQO		9AA
		3		5		2			0.2		80
		586 624 731	(647) 75.2	154 111 162	(142) 27.4	242 227 242	(237) 8.7		102 95 118	(105) 11.8	1270 1027 935	(1077) 173.1

ENNGN-ethyl-N'-nitro-N-nitrosoguanidine

4NQO 4-Nitroquinoline-1-oxide

9AA    9-Aminoacridine

P        Precipitate

#        Standard deviation

Table 3: Test Results: Experiment 1: With Metabolic Activation

Test Period		From: 15 March 2010						To: 18 March 2010
With or without S9-Mix	Test substance concentration (µg/plate)	Number of revertants (mean number of colonies per plate)
		Base-pair substitution type							Frameshift type
		TA100		TA1535		WP2uvrA^¿			TA98		TA1537
+	0	87 75 89	(84) 7.6#	23 16 18	(19) 3.6	26 31 35	(31) 4.5		24 30 29	(28) 3.2	13 9 9	(10) 2.3
+	50	79 84 77	(80) 3.6	15 19 13	(16) 3.1	27 22 27	(25) 2.9		29 34 20	(28) 7.1	7 13 8	(9) 3.2
+	150	87 89 87	(88) 1.2	18 23 21	(21) 2.5	22 21 21	(21) 0.6		27 23 24	(25) 2.1	13 7 9	(10) 3.1
+	500	68 66 66	(67) 1.2	22 16 21	(20) 3.2	20 23 29	(24) 4.6		19 21 24	(21) 2.5	5 13 10	(9) 4.0
+	1500	82 73 88	(81) 7.5	12 13 16	(14) 2.1	21 24 21	(22) 1.7		21 22 26	(23) 2.6	11 14 11	(12) 1.7
+	5000	60 P 74 P 76 P	(70) 8.7	15 P 19 P 19 P	(18) 2.3	26 P 30 P 20 P	(25) 5.0		19 P 21 P 27 P	(22) 4.2	11 P 9 P 12 P	(11) 1.5
Positive controls   S9-Mix   +	Name Concentration (µg/plate) No. colonies per plate	2AA		2AA		2AA			BP		2AA
		1		2		10			5		2
		305 310 280	(298) 16.1	145 200 156	(167) 29.1	181 146 184	(170) 21.1		168 148 119	(145) 24.6	131 124 155	(137) 16.3

2AA    2-Aminoanthracene

BP      Benzo(a)pyrene

P        Precipitate

#        Standard deviation

Table 4: Test Results: Experiment 2 ¿ Without Metabolic Activation

Test Period		From: 22 March 2010						To: 25 March 2010
With or without S9-Mix	Test substance concentration (µg/plate)	Number of revertants (mean number of colonies per plate)
		Base-pair substitution type							Frameshift type
		TA100		TA1535		WP2uvrA			TA98		TA1537
-	0	106 78 89	(91) 14.1#	22 17 21	(20) 2.6	21 17 20	(19) 2.1		15 19 13	(16) 3.1	11 12 19	(14) 4.4
-	50	106 93 104	(101) 7.0	11 21 22	(18) 6.1	24 18 18	(20) 3.5		17 14 18	(16) 2.1	10 8 11	(10) 1.5
-	150	100 112 84	(99) 14.0	31 27 17	(25) 7.2	17 21 15	(18) 3.1		14 21 12	(16) 4.7	8 11 11	(10) 1.7
-	500	97 113 121	(110) 12.2	24 24 24	(24) 0.0	20 27 18	(22) 4.7		12 19 21	(17) 4.7	9 12 9	(10) 1.7
-	1500	100 72 89	(87) 14.1	15 25 22	(21) 5.1	25 21 17	(21) 4.0		13 14 14	(14) 0.6	19 13 13	(15) 3.5
-	5000	83 P 92 P 103 P	(93) 10.0	18 P 13 P 22 P	(18) 4.5	19 P 21 P 28 P	(23) 4.7		27 P 12 P 18 P	(19) 7.5	21 P 12 P 19 P	(17) 4.7
Positive controls   S9-Mix   -	Name Concentration (µg/plate) No. colonies per plate	ENNG		ENNG		ENNG			4NQO		9AA
		3		5		2			0.2		80
		438 422 470	(443) 24.4	366 385 443	(398) 40.1	605 584 623	(604) 19.5		178 196 184	(186) 9.2	2484 3236 2411	(2710) 456.7

ENNG N-ethyl-N'-nitro-N-nitrosoguanidine

4NQO 4-Nitroquinoline-1-oxide

9AA    9-Aminoacridine

P        Precipitate

#        Standard deviation

Table 5: Test Results: Experiment 2: With Metabolic Activation

Test Period		From: 22 March 2010						To: 25 March 2010
With or without S9-Mix	Test substance concentration (µg/plate)	Number of revertants (mean number of colonies per plate)
		Base-pair substitution type							Frameshift type
		TA100		TA1535		WP2uvrA^¿			TA98		TA1537
+	0	124 100 94	(106) 15.9#	13 13 14	(13) 0.6	31 21 25	(26) 5.0		17 21 24	(21) 3.5	19 14 16	(16) 2.5
+	50	91 113 89	(98) 13.3	13 13 13	(13) 0.0	17 22 21	(20) 2.6		22 27 22	(24) 2.9	7 14 12	(11) 3.6
+	150	80 83 103	(89) 12.5	24 13 12	(16) 6.7	25 25 13	(21) 6.9		14 17 16	(16) 1.5	11 11 12	(11) 0.6
+	500	90 96 84	(90) 6.0	6 11 12	(10) 3.2	18 28 30	(25) 6.4		26 19 14	(20) 6.0	16 11 12	(13) 2.6
+	1500	86 90 77	(84) 6.7	13 10 13	(12) 1.7	21 15 17	(18) 3.1		19 14 17	(17) 2.5	14 11 12	(12) 1.5
+	5000	112 P 112 P 95 P	(106) 9.8	13 P 10 P 10 P	(11) 1.7	32 P 20 P 24 P	(25) 6.1		21 P 18 P 20 P	(20) 1.5	15 P 7 P 20 P	(14) 6.6
Positive controls   S9-Mix   +	Name Concentration (µg/plate) No. colonies per plate	2AA		2AA		2AA			BP		2AA
		1		2		10			5		2
		1399 1269 1343	(1337) 65.2	249 202 265	(239) 32.7	297 283 284	(288) 7.8		379 321 323	(341) 32.9	319 227 180	(242) 70.7

2AA    2-Aminoanthracene

BP      Benzo(a)pyrene

P        Precipitate

#        Standard deviation

Conclusions:

The test material was considered to be non-mutagenic under the conditions of this test.

Executive summary:

The method conforms to the guidelines for bacterial mutagenicity testing published by the major Japanese Regulatory Authorities including METI, MHLW and MAFF. It also meets the requirements of the OECD Guidelines for Testing of Chemicals No. 471: Bacterial Reverse Mutation Test, Method B13/14 of Commission Regulation (EC) Number 440/2008 of 30 May 2008 and the USA, EPA (TSCA) OPPTS harmonised guidelines.

Salmonella typhimurium strains TA1535, TA1537, TA98, TA100 and Escherichia coli strain WP2 uvrA were treated with suspensions of the test material using both the Ames plate incorporation and pre-incubation methods at five dose levels, in triplicate, both with and without the addition of a rat liver homogenate metabolising system (10% liver S9 in standard co-factors). The dose range was determined in a preliminary toxicity assay and was 50 to 5000 µg/plate in the first experiment. The experiment was repeated on a separate day (pre-incubation method) using the same dose range as Experiment 1, fresh cultures of the bacterial strains and fresh test material formulations. 

The vehicle (dimethyl sulphoxide) control plates gave counts of revertant colonies within the normal range. All of the positive control chemicals used in the test induced marked increases in the frequency of revertant colonies, both with or without metabolic activation. Thus, the sensitivity of the assay and the efficacy of the S9-mix were validated.

The test material caused no visible reduction in the growth of the bacterial background lawn at any dose level and was, therefore, tested up to the maximum recommended dose level of 5000 µg/plate. A fine, particulate precipitate was noted at 5000 µg/plate, this observation did not prevent the scoring of revertant colonies.

No significant increases in the frequency of revertant colonies were recorded for any of the bacterial strains, with any dose of the test material, either with or without metabolic activation or exposure method.

The test material was considered to be non-mutagenic under the conditions of this test.

Materials and methods

Test material

Test material information

Constituent 1

Reference substance name:

dieodymium tricarbonate

IUPAC Name:

dieodymium tricarbonate

Results and discussion

Test results

Test results 1

Key result

Species / strain:

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

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Test results 2

Key result

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid