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EC number: 219-784-2
CAS number: 2530-83-8
Gene mutation (Bacterial reverse mutation assay / Ames test): positive
with and without activation in TA97 and TA100 (similar to OECD Test
Guideline 471) (Microtest Research Ltd., 1988).
Mutagenicity in mammalian cells: positive in L5178Y mouse lymphoma
L5178Ycells (similar to OECD Test Guideline 476) (Litton Bionetics,
1983); negative in Chinese hamster Ovary (CHO) cells (similar to OECD
Test Guideline 476, Reliability 4), (Allied Corporation, 1979).
Table 2 : Mutagenicity assay, Number
of revertants per plate (mean of 3 plates)
*solvent control with Ethylene glycol
Appropriate concurrent negative and positive controls were
included, and the expected responses were observed.
Under non-activation conditions, a dose-dependent increase
in mutant frequency was induced and exceeded the minimum
criteria for mutagenicity (83.4 X 10-6) at 2000 nl/ml and
3000 nl/ml. Low to moderate toxicity was induced (percent
relative growth 97% - 37.7%).
In the presence of metabolic activation a wide range of
toxicities was observed (percent relative growth 91.2% -
8.9%). All the assayed treatments exceeded minimum
criterion of 68.0 X 10-6. However, relative growth was
dramatically inhibited at concentrations of 5000 nl/ml or
Results of mutagenicity testing
Expt 1 without activation
Expt 2 and 3 with activation
Relative growth (%)
Solvent control (av of 2expt 1, of 3 expt 2 and 3)
Positive control low dose
Positive control high dose
control low dose: 0.25 µl/ml EMS without activation, 0.15 µl/ml DMN with
control 0.4 µl/ ml EMS without activation, 0.130 µl/ml DMN with
The results in the paper are presented in
the form of bar charts. The numbers in the table below have been
estimated from the charts.
Table 1 Results of micronucleus assay for
There are three in vivo micronucleus assays available with conflicting
results. Therefore, in accordance with ECHA Decision number
TPE-D-2114428335-52-01/F, an in vivo alkaline comet assay was carried
out to clarify the potential for mutagenicity to somatic cells. A
statistically significant mild to moderate increase in tail intensity
was observed in all tissues at all concentrations tested.
A transgenic rodent assay (OECD TG 488) is proposed to evaluate the
potential for mutagenicity to germ cells.
[3-(2,3-epoxypropoxy)propyl]trimethoxysilane has been tested in a valid
in vivo mammalian alkaline comet assay study, conducted according to
OECD TG 489 and in compliance with GLP. Groups of 5 male Wistar WI (Han)
rats (SPF) were dosed via oral gavage over two consecutive days at doses
of 500, 1000 and 2000 mg/kg bw. No cytotoxicity was reported in the
target tissues and no clinical effects were observed in the treated
animals. Possible test item-related morphologic alterations (aggravation
of eosinophilic cell infiltrates in the submucosa) were present in the
glandular stomach at 2000 mg/kg bw. Statistically significant increases
in the mean tail intensity of cells from the glandular stomach, duodenum
and liver was reported at all doses tested.
The available information for the
registration substance on in vitro and in vivo genetic toxicity has been
considered, and there is evidence for mutagenicity from in vitro studies
and for damage to DNA from an in vivo mammalian alkaline comet assay,
where a statistically significant mild to moderate increase in tail
intensity was observed in all tissues at all concentrations tested. The
potential for germ cell mutagenicity has been considered on the basis of
all available data for the registration substance and related
substances, including: toxicokinetic modelling for the registration
substance and measured toxicokinetic data (intraperitoneal
administration) for another alkoxysilane; repeated dose toxicity studies
for the registered substance; genetic toxicity data with the registered
substance and with other silanes and siloxanes with an epoxy group. No
clear conclusions about germ cell mutagenicity have been reached,
therefore a transgenic rodent assay (OECD TG 488) is proposed. The
consideration of all available evidence is discussed in detail below.
Summary of available in vivo data for
registration substance [3-(2,3-epoxypropoxy)propyl]trimethoxysilane
Route of administration
Toxicity to bone marrow
Yes Moderate to severe reductions of 22 to 59% in the ratio of polychromatic erythrocytes to total erythrocytes were observed in the test article-treated groups relative to the vehicle control animals
Clinical signs included lethargy and piloerection at 1000 and 2000 mg/kg.
Toxicity to bone marrow was observed in female mice at 24 h observation.
All animals showed clinical signs following dosing (1600 mg/kg bw as a suspension in corn oil)
Dow Corning Corporation, 1982a
Sister chromatid exchange
No data; a high degree of bioavailability is expected after ip administration
Dow Corning Corporation, 1982d
Toxicokinetics: no toxicokinetic data are available for the
substance. There is evidence of systemic exposure following
intraperitoneal administration from an in vivo bioavailability study for
another trialkoxysilane (see section 7.1 of IUCLID and 5.1 of the CSR),
but this does not give an indication of bioavailability after oral
administration. In addition, systemic toxicity was observed in two of
the three micronucleus assays; no information was available for the
third study. Lethargy and piloerection were reported following
intraperitoneal exposure in one of the micronucleus studies
(BioReliance, 1999) at 1000 and 2000 mg/kg bw. In the more recent of the
negative micronucleus assay, severe clinical symptoms were observed
following intraperitoneal (ip) application (Hüls 1994). No systemic
toxicity was observed following oral administration of the registered
substance in repeated dose toxicity testing. Therefore there is no
information on bioavailability following oral administration.
The positive in vivo micronucleus assay is considered not relevant as
intraperitoneal administration it is not a physiologically relevant
route for this substance, taking into account expected routes of
exposure for the substance. There is no clear explanation for the
positive result but it is likely that route of exposure is important.
The weight of evidence from all studies (micronucleus and sister
chromatid exchange), and the agreement of results from studies using a
physiologically relevant route of exposure, is that the substance is not
In vivo micronucleus assays are not considered appropriate to
investigate the mutagenic effects of substances that are mutagenic to
bacterial and mammalian cells. This is implied in the REACH Regulation
in the column 2 adaptations for in vitro mammalian cell cytogenicity and
mutagenicity testing, and is stated in ECHA Guidance R07a. Appropriate
assays are, according to the ECHA Guidance (p 364), the transgenic
rodent assay (OECD Test Guideline 488) and the in vivo Comet assay (OECD
Test Guideline 489) or, if justified, Unscheduled DNA Synthesis, in vivo
(OECD Test Guideline 486). There are no in vitro effects that indicate
that a micronucleus assay is the appropriate in vivo assay to conclude
on somatic cell mutagenicity.
5. Carcinogenicity: the substance was not
considered tumorigenic when applied to the clipped skin of mice (25 µl
dose of 25% test material in acetone) three times per week for
approximately 78 weeks (BRRC, 1982). The ECHA Guidance (ECHA, 2014, R07a
p 337) states: “There is considerable evidence of a positive correlation
between the mutagenicity of substances in vivo and their carcinogenicity
in long-term studies with animals.” As the substance has been shown not
to be carcinogenic, it is unlikely that it is mutagenic.
To summarise, the negative data from the in
vitro micronucleus test on the structural analogue and the negative in
vivo micronucleus results following a physiologically relevant route of
exposure, with additional evidence of bioavailability indicate that
there is not sufficient evidence to classify as Mutagen Category 2, and
so an in vivo comet assay has been carried out. The comet assay can
detect evidence of DNA damage that may lead to both gene mutation and
chromosome mutation events and is therefore the ideal test method for
the follow-up of the in vitro / in vivo genotoxicity profile of the
substance (ECHA guidance on mutagenicity (R.7a, 19 August 2014, p 343)
states: “[…] the alkaline comet assay recognises primary DNA damage that
would lead to gene mutations and/or chromosome aberrations) but will
also detect DNA damage that may be effectively repaired or lead to cell
death". The comet assay can therefore be considered a conservative
estimate of genetic toxicity to somatic cells.
has been tested in a valid in vivo mammalian alkaline comet assay study,
conducted according to OECD TG 489 and in compliance with GLP. Groups of
5 male Wistar WI (Han) rats (SPF) were dosed via oral gavage over two
consecutive days at doses of 500, 1000 and 2000 mg/kg bw. No
cytotoxicity was reported in the target tissues and no clinical effects
were observed in the treated animals. Possible test item-related
morphologic alterations (aggravation of eosinophilic cell infiltrates in
the submucosa) were present in the glandular stomach at 2000 mg/kg bw.
Statistically significant increases in the mean tail intensity of cells
from the glandular stomach, duodenum and liver was reported at all doses
Consideration of the need for further
testing for germ cell mutagenicity
The evidence from the comet assay of damage
to DNA in the liver shows that the test substance reaches the liver
cells indicating that the registered substance has the potential for
interaction with DNA. However, this is not conclusive evidence that the
substance would induce heritable transformation in germ cells, so a
transgenic rodent assay (OECD TG 488) is proposed to evaluate the
potential for mutagenicity to germ cells.
Strategy for identification of key studies
Where there was more than one result for an
endpoint the most reliable study available was chosen as key study.
Where there was more than one reliable study, the most recent study was
selected. If the results were not in agreement, studies giving positive
results were chosen, unless the weight of evidence indicated the result
was not indicative of potential for mutagenicity. The results of all the
bacterial studies were in agreement and showed evidence of mutagenicity.
In mammalian mutagenicity studies, positive results were obtained when
tested in L5178Y mouse lymphoma cells (Litton Bionetics, 1983), but
negative results when Chinese hamster ovary cells were used (similar to
OECD 476, Reliability 4), (Allied Corporation, 1979).
No data are available for the registered
substance for in vitro cytogenicity, however, data are available for the
structural analogue [3-(2,3-epoxypropoxy)propyl]triethoxysilane (CAS
2602-34-8) from an in vitro micronucleus assay.
[3-(2,3-epoxypropoxy)propyl]triethoxysilane has been tested for ability
to induce formation of micronuclei in a study conducted according to
OECD 487 (Lionti et al., 2014). No evidence of test-substance induced
micronucleus formation was observed in HepG2 cells in either the
presence or the absence of metabolic activation. Appropriate positive
and solvent controls were included and gave expected results. It is
concluded that the test substance in negative for cytogenicity under the
conditions of the test. This study is not considered to be key, as an in
vivo mammalian alkaline comet assay is available.
There is insufficient evidence to make a conclusion on mutagenicity to
germ cells, therefore, a further study is proposed.
Information on Registered Substances comes from registration dossiers which have been assigned a registration number. The assignment of a registration number does however not guarantee that the information in the dossier is correct or that the dossier is compliant with Regulation (EC) No 1907/2006 (the REACH Regulation). This information has not been reviewed or verified by the Agency or any other authority. The content is subject to change without prior notice.Reproduction or further distribution of this information may be subject to copyright protection. Use of the information without obtaining the permission from the owner(s) of the respective information might violate the rights of the owner.
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