Refractories, fibers, aluminosilicate

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:

13 January 2010 and was completed on 8 February 2010.

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: This study was performed according to the protocol with two amendments, with the exception of the minor deviations none of which were considered to prejudice the validity of this study.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2010

Report date:

2010

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

GLP compliance:

yes (incl. QA statement)

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

oxo[(oxoalumanyl)oxy]alumane; silanedione

EC Number:

604-314-4

Cas Number:

142844-00-6

Molecular formula:

amorphous glass Si(n)O(3n+1) polymeric anions bonded to Zr and Al(3+)

IUPAC Name:

oxo[(oxoalumanyl)oxy]alumane; silanedione

Details on test material:

Name of test material( as cited in study report): RCF Covance
Substance type: Inorganic
- Physical state: solid
- Analytical purity: 100%
- Impurities (identity and concentrations):trace
- Purity test date: 14.12.09
- Lot/batch No.: BG-10-X99-2982
- Expiration date of the lot/batch: 31.1.2020
- Stability under test conditions: stable
- Storage condition of test material: sealed container room temp

Method

Target gene:

Histidine

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:

not applicable

Species / strain 2

Species / strain / cell type:

S. typhimurium TA 102

Additional strain / cell type characteristics:

not applicable

Metabolic activation:

without

Metabolic activation system:

material is completely inorganic so none is needed

Test concentrations with justification for top dose:

see table1 in other information on methods section, range finder and mutation experiments were carried out

Vehicle / solvent:

- Vehicle(s)) used:; 1% methylcellulose ( MC);
- Justification for choice of vehicle:test article is insoluble in all possible vehicles compatible with the assay system. The test article was therefore tested as a suspension in 1% MC. Ultrasonication was used to ensure homogeneity of the stock suspension and the formulations were stirred continuously (using a magnetic stirrer) prior to their addition to the test system.

Controls

Controls 1

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

9-aminoacridine

Controls 2

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

2-nitrofluorene

Controls 3

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

sodium azide

Controls 4

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

mitomycin C

Details on test system and experimental conditions:

METHOD OF APPLICATION: in suspension; Ultrasonication was used to aid homogenity of the stock suspension and the formulation was stirred continually (using magnetic stirrer) prior to addition to the test system

DURATION
- Preincubation period: 10 hours
- Exposure duration: 2-3 days

Metabolic activation is in appropriate for totally inorganic materials of this type

Evaluation criteria:

For valid data, the test article was considered to be mutagenic if:
1. Dunnett's test gave a significant response (p ≤ 0.01) which was concentration
related
2. the positive trend/effects described above were reproducible.
The test article was considered as positive in this assay if all of the above criteria were met.
The test article was considered as negative in this assay if none of the above criteria were met.
Test article was considered as positive in this assay if all of the above criteria were met
The test article was considered as negative in this assay if none of the above criteria were met

Statistics:

Dunnett's test was used to compare the counts at each concentration with the control

Results and discussion

Test results

Test results 1

Species / strain:

S. typhimurium TA 102

Metabolic activation:

without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not determined

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Test results 2

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not determined

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Test results 3

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not determined

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Test results 4

Species / strain:

S. typhimurium TA 98

Metabolic activation:

without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not determined

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Test results 5

Species / strain:

S. typhimurium TA 100

Metabolic activation:

without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not determined

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Water solubility: material is an insoluble inorganic solid

RANGE-FINDING/SCREENING STUDIES:
Initial toxicity Range-Finder Experiment was carried out in strain TA100 only, using final concentrations of AES Covance at 1.6, 8, 40, 200, 1000 and 5000 μg/plate, plus negative (vehicle) and positive controls. Following these treatments, no evidence of toxicity was observed, as would normally be indicated by a thinning of the background bacterial lawn and/or a marked reduction in revertant numbers.

COMPARISON WITH HISTORICAL CONTROL DATA:
Control counts were compared with the accepted normal ranges for the test laboratory for numbers of spontaneous revertants on vehicle control plates and numbers of induced revertants on positive control plates. Data were considered acceptable if the mean vehicle control counts fell within the historical 99% confidence intervals for group means and/or each vehicle control plate count fell within the historical 99% reference ranges, and the positive control plate counts were comparable with the historical 99% reference ranges. The ranges that are quoted are based on a large volume of historical control data accumulated from experiments where the correct strain and assay functioning are considered to have been confirmed. Data for our laboratory are consistent with ranges of spontaneous revertants per plate considered acceptable elsewhere.

ADDITIONAL INFORMATION ON CYTOTOXICITY:
Initial toxicity Range-Finder Experiment was carried out in strain TA100 only, using final concentrations of RCF Covance at 1.6, 8, 40, 200, 1000 and 5000 μg/plate, plus negative (vehicle) and positive controls. Following these treatments, no evidence of toxicity was observed, as would normally be indicated by a thinning of the background bacterial lawn and/or a marked reduction in revertant numbers.

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Any other information on results incl. tables

Raw plate counts and calculated mutagenicity data Experiment 1

 

Table 3: RCF Covance Raw plate counts and calculated mutagenicity data - Experiment 1, TA98 –S-9

Compound	Concentration µg/plate	Revertant numbers/plate	Mean	N	Fold increase	Standard deviation	Dunnett’s t-value	significance
1% MC		22,40,26,27,34	29.8	5		7.2
RCF Covance	1.6	25,25,30	26.7	3	0.9	2.9	-0.57	NS
RCF Covance	8	34,43,35	37.3	3	1.3	4.9	1.44	NS
RCF Covance	40	25,25,29	27.3	3	0.9	2.3	-0.64	NS
RCF Covance	200	31P,13P,25P	23	3	0.8	9.2	-1.50	NS
RCF Covance	1000	26 P,23P,26 P	25	3	0.8	1.7	-0.92	NS
RCF Covance	5000	14 P,26P,34P	24.7	3	0.8	10.1	-1.15	NS
2NF	5		814.3	3	27.3	36.9

Dunnett’s t-test significance values

NS Not significant

* p ≤ 0.05

** p ≤ 0.01

Positive controls

2NF 2-Nitrofluorene

NaN3 Sodium azide

AAC 9-Aminoacridine

MMC Mitomycin C

Table Postfixes

B Bubbles or split in agar

M Plate counted manually

P Particulate test article observed (by eye)

Table 4: RCF Covance Raw plate counts and calculated mutagenicity data - Experiment 1, TA100 -S-9

Compound	Concentration µg/plate	Revertant numbers/plate	Mean	N	Fold increase	Standard deviation	Dunnett’s t-value	significance
1% MC		79,101,117,103,70	94.0	5		19.1
RCF Covance	1.6	114,88,88	96.7	3	1.0	15.0	0.34	NS
RCF Covance	8	110, 103, 104	105.7	3	1.1	3.8	1.33	NS
RCF Covance	40	100, 89, 101	96.7	3	1.0	6.7	0.37	NS
RCF Covance	200	103 P, 100 P, 104 P	102.3	3	1.1	2.1	0.99	NS
RCF Covance	1000	94 P, 74 P, 96 P	88.0	3	0.9	12.2	-0.62	NS
RCF Covance	5000	105 P, 91 P, 101 P	99.0	3	1.1	7.2	0.62	NS
NaN3	2	1134, 1081, 1084	1099.7	3	11.7	29.8

Table 5 :RCF CovanceRaw plate counts and calculated mutagenicity data - Experiment 1, TA1535 -S-9

Compound	Concentration (mg/plate)	Revertant numbers/plate	Mean	N	Fold Increase	Standard Deviation	Dunnett’s t‑value	Significance
1% MC		16, 16, 18, 16, 10	15.2	5		3.0
RCF Covance	1.6	17, 16, 17	16.7	3	1.1	0.6	0.46	NS
RCF Covance	8	17, 36, 27	26.7	3	1.8	9.5	2.82	*
RCF Covance	40	8, 14, 16	12.7	3	0.8	4.2	-0.82	NS
RCF Covance	200	17 P, 13 P, 13 P	14.3	3	0.9	2.3	-0.24	NS
RCF Covance	1000	9 P, 21 P, 17 P	15.7	3	1.0	6.1	0.05	NS
RCF Covance	5000	13 P, 23 P, 13 P	16.3	3	1.1	5.8	0.28	NS
NaN3	2	568, 604, 607	593.0	3	39.0	21.7

Table 6: RCF CovanceRaw plate counts and calculated mutagenicity data - Experiment 1, TA1537 -S-9

Compound	Concentration (mg/plate)	Revertant numbers/plate	Mean	N	Fold Increase	Standard Deviation	Dunnett’s t‑value	Significance
1% MC		6, 10, 12, 14, 4	9.2	5		4.1
RCF Covance	1.6	5, 12, 21	12.7	3	1.4	8.0	0.73	NS
RCF Covance	8	18, 12, 6	12.0	3	1.3	6.0	0.66	NS
RCF Covance	40	19, 3, 19	13.7	3	1.5	9.2	0.82	NS
RCF Covance	200	17 P, 14 P, 12 P	14.3	3	1.6	2.5	1.28	NS
RCF Covance	1000	13 P, 10 P, 10 P	11.0	3	1.2	1.7	0.54	NS
RCF Covance	5000	12 P M B, 5 P, 8 P	8.3	3	0.9	3.5	-0.19	NS
AAC	50	43, 77, 65	61.7	3	6.7	17.2

Table 7 : RCF CovanceRaw plate counts and calculated mutagenicity data - Experiment 1, TA102 -S-9

Compound	Concentration (mg/plate)	Revertant numbers/plate	Mean	N	Fold Increase	Standard Deviation	Dunnett’s t‑value	Significance
1% MC		273, 289, 302, 261, 251	275.2	5		20.6
RCF Covance	1.6	240, 289, 315	281.3	3	1.0	38.1	0.39	NS
RCF Covance	8	303, 296, 321	306.7	3	1.1	12.9	2.19	NS
RCF Covance	40	302, 298, 321	307.0	3	1.1	12.3	2.21	NS
RCF Covance	200	283 P, 308 P, 305 P	298.7	3	1.1	13.7	1.65	NS
RCF Covance	1000	295 P, 309 P, 305 P	303.0	3	1.1	7.2	1.95	NS
RCF Covance	5000	286 P, 295 P, 269 P	283.3	3	1.0	13.2	0.59	NS
MMC	0.2	927, 922, 867	905.3	3	3.3	33.3

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative without metabolic activation

It was concluded that RCF Covance did not induce mutation in
five histidine-requiring strains (TA98, TA100, TA1535, TA1537 and TA102) of
Salmonella typhimurium when tested under the conditions of this study. These
conditions included treatments at concentrations up to 5000 μg/plate, in the absence
of any exogenous metabolic activation system

Executive summary:

It was concluded that RCF Covance did not induce mutation in five histidine‑requiring strains (TA98, TA100, TA1535, TA1537 and TA102) of Salmonella typhimurium when tested under the conditions of this study. These conditions included treatments at concentrations up to 5000 mg/plate, in the absence of any exogenous metabolic activation system