N-[3-(Triethoxysilyl)propyl]formamide

Genetic toxicity: in vitro

Administrative data

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:

21 Sep - 12 Oct 2011

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2011

Report date:

2011

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

JAPAN: Guidelines for Screening Mutagenicity Testing Of Chemicals

Version / remarks:

Kanpoan No. 287 - EPA; Eisei No. 127 - Ministry of Health & Welfare; Heisei 09/10/31 Kikyoku No.2 - Ministry of Intern. Trade & Industry

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

Hess. Ministerium für Umwelt, Landwirtschaft und Forsten, Mainzer Straße 80, D-65189 Wiesbaden, Germany

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

N-[3-(triethoxysilyl)propyl]formamide

EC Number:

700-340-6

Cas Number:

76524-94-2

Molecular formula:

C10H23NO4Si

IUPAC Name:

N-[3-(triethoxysilyl)propyl]formamide

Details on test material:

- Name of test material (as cited in study report): Y-15864
- Analytical purity: 97.5 - 99.5%
- Lot/batch No.: TER00010-68-31
- Expiration date of the lot/batch: December 09, 2012
- Storage condition of test material: at room temperature

Method

Target gene:

Salmonella typhimurium: his operon
Escherichia coli: trp operon

Species / strain

Species / strain 1

Species / strain / cell type:

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

Additional strain / cell type characteristics:

other: rfa-; uvrB- (R+ for TA 98 and TA 100)

Species / strain 2

Species / strain / cell type:

E. coli WP2 uvr A

Additional strain / cell type characteristics:

other: uvrA-

Metabolic activation:

with and without

Metabolic activation system:

cofactor supplemented post-mitochondrial fraction (S9 mix) prepared from the livers of rats treated with phenobarbital/β-naphthoflavone.

Test concentrations with justification for top dose:

Pre-Experiment and first experiment: 3, 10, 33, 100, 333, 1000, 2500 and 5000 µg/plate with and without S9 mix
Second Experiment: 33, 100, 333, 1000, 2500 and 5000 µg/plate with and without S9 mix

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: The solvent was chosen because of its solubility properties and its relative non-toxicity to the bacteria.

Controls

Untreated negative controls:

yes

Remarks:

culture media

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: 10 µg/plate sodium azide (TA100 and 1535, -S9), 10 µg/plate 4-nitro-o-phenylen-diamine (TA98 and 1537, -S9), 3 µL/plate methyl methane sulfonate (WP2, -S9), 2.5 µg/plate 2-aminoanthracene (2-AA) (TA98, 100, 1535, 1537, +S9), 10 µg/plate 2-AA (WP2, +S9)

Details on test system and experimental conditions:

METHOD OF APPLICATION: Experiment I: plate incorporation; Experiment II: preincubation

EXPERIMENTAL PERFORMANCE
The following materials were mixed in a test tube and poured onto the minimal agar plates:
100 µL Test solution at each dose level, solvent (negative control) or reference mutagen
solution (positive control),
500 µL S9 mix (for test with metabolic activation) or S9 mix substitution buffer (for test
without metabolic activation),
100 µL Bacteria suspension (cf. test system, pre-culture of the strains),
2000 µL Overlay agar

DURATION
- Preincubation period: 60 min
- Selection time (if incubation with a selection agent): 48 hours

SELECTION AGENT (mutation assays): minimal agar

NUMBER OF REPLICATIONS: triplicates in each of two independent experiments

DETERMINATION OF CYTOTOXICITY
- Method: relative total growth / regular background growth / spontaneous reversion rates in the negative and solvent control

Evaluation criteria:

A test item is considered as a mutagen if a biologically relevant increase in the number of revertants exceeding the threshold of twice (strains TA 98, TA 100, and WP2 uvrA) or thrice (strains TA 1535 and TA 1537) the colony count of the corresponding solvent control is observed.
A dose dependent increase is considered biologically relevant if the threshold is exceeded at more than one concentration.
An increase exceeding the threshold at only one concentration is judged as biologically relevant if reproduced in an independent second experiment.
A dose dependent increase in the number of revertant colonies below the threshold is regarded as an indication of a mutagenic potential if reproduced in an independent second experiment. However, whenever the colony counts remain within the historical range of negative and solvent controls such an increase is not considered biologically relevant.

Statistics:

Mean and standard deviation were calculated.

Results and discussion

Test results

Test results 1

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 nor precipitates, but tested up to recommended limit concentrations

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 2

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity nor precipitates, but tested up to recommended limit concentrations

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

COMPARISON WITH HISTORICAL CONTROL DATA: Yes

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Any other information on results incl. tables

Table 1: Summary Ames Test Results – Pre-Experiment and Experiment 1

With or without S9-Mix	Test substance concentration (μg/plate)	Mean number of revertant colonies per plate (average of triplicate)
		Base-pair substitution type		Frameshift type		Base-pair substitution and other
		TA 1535	TA100	TA1537	TA98	WP2 uvrA
-	Negative control	18±5	144±13	12±3	41±2	45±6
-	Vehicle control	15±4	139±3	11±3	32±3	43±4
-	3	16±4	138±12	13±5	30±5	42±7
-	10	15±5	127±7	12±2	32±8	43±4
-	33	13±3	134±10	14±4	31±8	36±6
-	100	17±5	141±10	11±4	35±5	41±7
-	333	13±3	147±5	14±4	36±5	43±11
-	1000	10±2	145±8	11±1	35±5	41±1
-	2500	11±5	147±7	12±4	31±10	38±3
-	5000	11±1	122±10	9±2	38±4	33±4
Positive controls - S9	Name	NaN3	NaN3	4-NOPD	4-NOPD	MMS
	Concentrations (μg/plate)	10	10	50	10	3.0 µL
	Number of colonies/plate	1726±40	1998±64	84±7	385±32	1043±98
		TA 1535	TA 100	TA1537	TA98	WP2 uvrA
+	Negative control	17±5	127±11	16±4	41±2	62±13
+	Vehicle control	21±5	133±10	17±4	41±11	52±3
+	3	19±2	119±9	16±1	45±2	50±6
+	10	23±3	124±5	16±3	41±6	48±7
+	33	21±3	129±17	16±4	40±5	47±3
+	100	16±1	129±20	16±3	35±6	48±2
+	333	19±3	128±8	19±6	44±3	48±8
+	1000	16±1	119±9	18±3	38±3	51±6
+	2500	18±3	143±28	15±5	42±7	44±7
+	5000	20±4	142±20	13±3	39±5	50±8
Positive controls + S9	Name	2-AA	2-AA	2-AA	2-AA	2-AA
	Concentrations (μg/plate)	2.5	2.5	2.5	2.5	10.0
	Number of colonies/plate	329±14	1316±2	184±14	1458±56	198±19

Table 2: Summary Ames Test Results - Experiment 2

With or without S9-Mix	Test substance concentration (μg/plate)	Mean number of revertant colonies per plate (average of triplicate)
		Base-pair substitution type		Frameshift type		Base-pair substitution and other
		TA 1535	TA100	TA1537	TA98	WP2 uvrA
-	Negative control	9±3	124±11	19±6	35±8	41±7
-	Vehicle control	9±1	102±8	11±2	27±10	48±3
-	33	11±3	109±6	11±1	28±8	43±1
-	100	12±2	101±12	11±5	23±3	49±5
-	333	9±2	98±4	12±2	26±3	42±6
-	1000	11±3	91±11	11±3	31±4	45±4
-	2500	11±2	91±8	13±3	27±8	48±6
-	5000	10±1	66±6	11±3	22±6	41±3
Positive controls - S9	Name	NaN3	NaN3	4-NOPD	4-NOPD	MMS
	Concentrations (μg/plate)	10	10	50	10	3.0 µL
	Number of colonies/plate	1633±7	1672±50	78±12	339±21	443±64
		TA 1535	TA 100	TA1537	TA98	WP2 uvrA
+	Negative control	16±5	144±15	22±6	38±4	62±8
+	Vehicle control	16±4	110±9	15±6	39±4	48±9
+	33	15±5	114±13	18±3	43±2	51±5
+	100	19±4	123±12	18±3	41±1	51±11
+	333	16±2	114±8	16±3	38±8	49±3
+	1000	15±4	107±6	13±3	41±3	49±9
+	2500	17±6	126±4	16±2	35±4	47±4
+	5000	19±6	117±11	17±5	43±2	51±9
Positive controls + S9	Name	2-AA	2-AA	2-AA	2-AA	2-AA
	Concentrations (μg/plate)	2.5	2.5	2.5	2.5	10.0
	Number of colonies/plate	260±9	1510±152	185±0	1382±7	312±9

NaN3 = sodium azide

2-AA = 2-aminoanthracene

4-NOPD = 4 -nitro-o-phenylene-diamine

MMS = methyl methane sulfonate

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative

Executive summary:

N-[3-(Triethoxysilyl)propyl]formamide was tested in an Ames test using Salmonella typhimurium strains TA 1535, TA 1537, TA 98, and TA 100 and the Escherichia coli strain WP2 uvrA. The substance was tested up to 5000 µg/plate with and without metabolic activation. No toxic effects occurred in the test groups with and without S9 mix. Under the experimental conditions the test item did not induce mutations by base pair changes or frameshifts in the genome of the strains used.