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EC number: 214-014-1
CAS number: 1072-71-5
Repeated Dose toxicity: Subacute /
reproductive toxicity study oral (gavage), rat (Han Wistar) m/f,
10/sex/dose, 0, 50, 150, 500 mg/kg bw/d(nominal) in PEG 400 (OECD TG
422, GLP): NOAEL = 24 mg/kg bw/d (analytical), based on low bodyweight
and food consumption effects at 500 mg/kg/day and histopathological
findings in the stomach which contributed to the death of one female at
500 mg/kg/day and at 150 mg/kg/day. No specific target organ toxicity
was identified, no classification as STOT RE Cat. 2 was triggered.
See subchapter "reproductive toxicity"
purpose of this study according to OECD 422 under GLP was the assessment
of general systemic toxic potential in Han Wistar rats, including a
screen for reproductive/developmental effects and assessment of
endocrine disruptor relevant endpoints, with administration of
2,5‑Dimercapto‑1,3,4‑thiadiazole by oral gavage administration for at
least five weeks.
groups of ten male and ten female Han Wistar rats received
2,5‑Dimercapto‑1,3,4‑thiadiazole at doses of 50, 150 or 500 mg/kg/day by
oral gavage administration. Males
were treated daily for two weeks before pairing, up to necropsy after a
minimum of five consecutive weeks. Females
were treated daily for two weeks before pairing, throughout pairing,
gestation and until Day 13 of lactation. Females
were allowed to litter, rear their offspring and were killed on Day 14
of lactation. The
F1 generation received no direct administration of the test item; any
exposure was in utero or via the milk. A similarly
constituted Control group received the vehicle, PEG 400, at the same
volume dose as treated groups.
the study, clinical condition, detailed physical examination and arena
observations, sensory reactivity observations, grip strength, motor
activity, body weight, food consumption, hematology (peripheral blood),
blood chemistry, thyroid hormone analysis, estrous cycles, pre-coital
interval, mating performance, fertility, gestation length, organ weight
and macroscopic pathology and histopathology investigations were
clinical condition, litter size and survival, sex ratio, body weight,
ano-genital distance and macropathology for all offspring were also
counts were performed on male offspring on Day 13 of age.
homogeneity and stability was confirmed for
2,5-Dimercapto-1,3,4-Thiadiazole in PEG 400 formulations at nominal
concentrations of 1 mg/mL and 400 mg/mL during distribution between the
bottles, during magnetic stirring for 2 hours, ambient temperature
storage for 1 day and refrigerated storage for up to 15 days.
mean concentrations of Group 4 were within applied limits +10/-15%,
confirming the accuracy of formulation. Groups 2 and 3 were lower than
nominal concentration ranging from -52% to -15.5%.
animals were either killed for welfare reasons or found dead during the
treatment phase, however only the following death was considered to be
related to treatment.
500 mg/kg/day female, No. 46, was killed for welfare reasons on Day 6 of
treatment. On Day 5 of treatment at the 1-2 hours post-dose and as late
as possible in the working day observations, signs of rales was evident
which continued into Day 6 accompanied with gasping. Dark coloured
depressions in the corpus of the stomach, seen macroscopically,
correlated with mucosal erosion/ulceration and necrosis which was
considered to be the major factor contributing to death. The
stomach findings were similar to those seen in terminal animals and were
considered to be treatment related.
observations included chin rubbing and salivation and were observed
immediately after administration in the majority of cases, at 150 and
500 mg/kg/day and continued through the gestation and lactation phases
for females at 150 and 500 mg/kg/day. Rales was evident on days 2, 5 and
6 of treatment in both males and females at 500 mg/kg/day. Two males at
500 mg/kg/day had loose faeces.
receiving 500 mg/kg/day showed marked mean bodyweight loss following
initial treatments over Week 0 to 1 (16 g). Thereafter from Week 1
onwards, males receiving 500 mg/kg/day showed body weight gains with
mean body weight gains being higher than controls during Week 1 to 3 but
lower during Weeks 3-5. The overall bodyweight change from Week 0 to
Week 5 in males receiving 500 mg/kg/day was 53 % lower than Control.
There was no effect on bodyweight at 50 or 150 mg/kg/day in males and
females or 500 mg/kg/day in females during the treatment phase.
gestation, Group 4 females receiving 500 mg/kg/day persistently showed
lower bodyweights than Controls. The overall weight gain from Day 0-20
was 22.7% lower than Controls. Bodyweights during gestation were
unaffected at 50 and 150 mg/kg/day.
the lactation phase, low bodyweights continued in females receiving 500
mg/kg/day, however, the overall bodyweight gain from Day 1 to 13 was
comparable with Control. Bodyweights during lactation were unaffected at
50 and 150 mg/kg/day.
consumption of males and females receiving 500 mg/kg/day was markedly
low when compared to Control during Week 1 (males: 59%; females 85%). Food
intake of these animals increased during Week 2, but remained lower than
controls in males for the duration of the treatment period. The overall
mean food consumption for males receiving 500 mg/kg/day was 82% of
Control. Food consumption at 50 and 150 mg/kg/day was unaffected by
consumption was unaffected at all treatment levels during the gestation
the lactation phase, females receiving 500 mg/kg/day had low food
consumption when compared with Control. The mean food consumption for
days 1-12 of lactation was 81% of Control. Food consumption was
unaffected at 50 and 150 mg/kg/day in the lactation phase.
area(s) of the stomach were observed in both sexes receiving 500
mg/kg/day. Depressions in the corpus were seen in a male receiving 500
mg/kg/day and females receiving 150 mg/kg/day. A
single incidence of a thickened corpus was seen in a male receiving 150
mg/kg/day with a thickened limiting ridge also recorded in a male
receiving 500 mg/kg/day.
to treatment with 2,5-Dimercapto-1,3,4-thiadiazole were seen in the
kidney and stomach.
increase incidence of cortical tubular pigment was seen in both sexes receiving
500 mg/kg/day and males receiving 150 mg/kg/day.
Special stains, consisting of Perls’ stain and Schmorl’s stains, were
performed on the kidneys of four animals comprising a control and a high
dose animal of each sex. The
pigment was negative for Perls’ and positive for Schmorl’s, indicating
that the pigment was lipofuscin.
were seen in the stomach of both sexes receiving
150 and 500 mg/kg/day. The
findings were seen mostly in the glandular region and comprised mucosal
erosion/ ulceration, haemorrhage and oedema in the lamina propria,
mucosal regeneration and submucosal inflammatory cells. In
addition females receiving
500 mg/kg/day also had epithelial
hyperplasia affecting the non-glandular stomach and limiting ridge.
T4 levels were low in males receiving 500 mg/kg/day and mean adjusted
thyroid weight was marginally but statistically significantly high, but
there were no supporting microscopic pathology changes in the thyroids.
clinical condition, litter size, sex ratio, survival indices and body
weight gain of offspring were unaffected by parental treatment.
was no effect of parental treatment upon circulating levels of thyroxine
(T4) in offspring on Day 13 of age.
ano-gential distances of offspring were unaffected by paternal treatment
and no nipples were seen on any male offspring on Day 13 of age.
macroscopic findings considered to be related to paternal treatment were
was concluded that the oral administration of
2,5-Dimercapto-1,3,4-thiadiazole to parental Han Wistar rats at dose
levels of 50, 150 or 500 mg/kg/day administered for
two weeks before pairing, during pairing and then up to termination of
the males after five weeks of treatment and females on Day 14 of
low bodyweight and food consumption effects at 500 mg/kg/day and
histopathological findings in the stomach which contributed to the death
of one female at 500 mg/kg/day and at 150 mg/kg/day. The
no-observed adverse-effect level (NOAEL) of 2,5-Dimercapto-1,3,4-thiadiazolefor
toxicity was considered to be 24 mg/kg/day when adjusted for the results
of the formulation analysis.
performance, fertility, litter size and offspring survival and growth
were unaffected by parental treatment and, in
the context of this study, 2,5‑Dimercapto‑1,3,4‑thiadiazoleshowed
no conclusive evidence of being an endocrine disruptor and therefore the
no-observed adverse-effect level (NOAEL) for reproductive/developmental
toxicity was considered to be 500 mg/kg/day.
mode of action analysis in its classic sense cannot be performed for
repeated dose toxicity as there is only one repeated dose study
available in one species, inclusive reproductive screening. However, the
rat is an established model for human risk assessment, interspecies
differences are well studied and allow transfer conclusions to humans
for most chemicals exhibiting certain effects in the rat. Nevertheless
it is aimed to follow the WHO IPCS template as far as possible:
present MoAA aims to describe the nature of the observed effects with
regard to repeated dose systemic toxicity aims to draw conclusions on
their relevance for humans.
Mode of action Statement
were some effects noted in various organs and a general poorer health
state is associated. A dose-response was observed, the degree of
severity increases with the dosage. At the lowest dose, no relevant
effects were noted. With increasing doses, the effects may be more
interpreted as adverse. Besides local irritation in the stomach, all
effects were judged not consistent, and not to allow to conclude a
specific organ toxicity.
of data for use in Mode of Action Analysis
the available OECD 422 on the registered substance itself (oral: gavage,
Han Wistar rat m/f, 10/sex/dose, doses: 0, 50, 150, 500 mg/kg bw/d
(nominal), duration: subacute), the NOAEL was delineated as 24 mg/kg
oral administration of 2,5-Dimercapto-1,3,4-thiadiazole to parental Han
Wistar rats was generally well tolerated. One
female animal at 150 mg/kg/day and one male animal at 500 mg/kg/day were
found dead during treatment and one Control female and one female at 500
mg/kg/day were euthanised for welfare reasons. It was considered that
the only death attributable to treatment was female No. 46 at 500
mg/kg/day due to the mucosal erosion/ulceration and necrosis seen in the
revealed dark areas and depressions of the stomach which correlated with
mucosal erosion/ ulceration and hemorrhage in the lamina propria. In
addition, oedema, mucosal regeneration and submucosal inflammatory cells
were seen after histopathological assessment. These stomach changes at
500 or 150 mg/kg/day were considered to be adverse and to be due to the
moderate local irritation of the test item, which further contributed to
the death of one female at 500 mg/kg/day and at 150 mg/kg/day. This
effect was not present in the 50 mg/kg/day group and was considered
tubular pigment was seen in the kidneys in both sexes receiving 500
mg/kg/day and males receiving 150 mg/kg/day, however this finding was
not considered adverse as there were no nephrotoxic findings in the
bodyweight gains were low for males at 500 mg/kg/day after 5 weeks of
treatment and overall bodyweight gains were low at 500 mg/kg/day during
the gestation phase. Bodyweights
remained low during the lactation phase; however the bodyweight gains
were comparable with Control. Bodyweights were unaffected at 50 and 150
mg/kg/day throughout the study. Overall food consumption was low for
males at 500 mg/kg/day and females before pairing and during
lactation at 500 mg/kg/day when compared with Control. Food consumption
at 50 and 150 mg/kg/day was unaffected throughout the study.
of key events identified for a specific Mode of Action
the table in the next subchapter for incidence and severity. In general
the following events were noted:
Key Event 1
Key Event 2
bodyweight loss / diminished body weight in males
Key Event 3
Lower food consumption
Key Event 4
Macroscopic dark area(s) of the stomach, depressions in the corpus
Key Event 5
Increase incidence of cortical tubular pigment
other examined parameters were not different from control.
Hill Considerations for Weight of Evidence Analysis of available
data/information for Mode of Action Analysis in experimental species
Response Relationships and Temporal Association
on temporal association cannot be drawn, as there is only one OECD 422
study available with a study duration of approx. 6-7 weeks.
Dose (mg/kg bw/d)
Key Event 1 (severity)
Key Event 2 (severity)
Key Event 3 (severity)
Key Event 4 (severity)
Key Event 5 (severity)
No treatment-related deaths
no effect on bodyweight
1 treatment-related death
- Depressions in the corpus in females
-Mucosal Erosion/Ulceration – Glandular Region (1M / 2F)
- Haemorrhage in Lamina Propria – Glandular Region (1M / 1F)
- increase incidence of cortical tubular pigment in males
Males showed marked mean bodyweight loss following initial treatments over Week 0 to 1.
The overall bodyweight change from Week 0 to Week 5 in males receiving 500 mg/kg/day was 53 % lower than Control.
No effect on bodyweight in females.
- Food consumption markedly low when compared to Control during Week 1 (males: 59%; females 85%). The overall mean food consumption for males receiving 500 mg/kg/day was 82% of Control.
- The mean food consumption for days 1-12 of lactation was 81% of Control
- Dark area(s) of the stomach were observed in both sexes.
- Depressions in the corpus in a male
-Mucosal Erosion/Ulceration – Glandular Region (4M / 2F)
- Haemorrhage in Lamina Propria – Glandular Region (4M / 5F)
- Submucosal Inflammatory Cells – Glandular Region (3M / 1F)
- Oedema and Epithelial Hyperplasia
- increase incidence of cortical tubular pigment in both sexes
postulated above, a dose-response was observed, the degree of severity
increases with the dosage.
& Specificity – Biological Plausibility
key events follow a dose-response-curve, as depicted above (Dose
Response Relationships). Key events 1-3, i.e. death, bodyweight loss and
diminished food consumption are general signs of overall toxicity, and
cannot be attributed to a certain MoA. However, they could be explained
by the effects noted in the stomach which lead to the poor health
condition. All effects in the stomach can be explained by local,
irritating effects on the stomach. The substance was classified as Eye
Dam. 1, and, although not classified as skin irr., in the dermal acute
toxicity test there were also some effects noted after 24h continuous
exposure, such as crust formation, small superficial scattered scabs,
scab lifting to reveal glossy skin, yellow staining of the treatment
sites. So upon repeated exposure on the stomach mucosa, irritating
effects occurred. With
regard to cortical tubular pigment which was seen in the kidneys in both
sexes receiving 500 mg/kg/day and males receiving 150 mg/kg/day, special
stains, consisting of Perls’ stain and Schmorl’s stains, were performed
on the kidneys of four animals comprising a control and a high dose
animal of each sex. The
pigment was negative for Perls’ and positive for Schmorl’s, indicating
that the pigment was lipofuscin. This
finding was not considered adverse as there were no nephrotoxic findings
in the clinical pathology. Further, males seem to be more susceptible
here than females, and although no information on alpha-2u-globulin is
available, this may indicate that these effects are even less relevant
and Quantitative human concordance
is no indication given that the obtained results may not be relevant for
humans, no species-specifity was obvious, besides a possible indication
from the severity of kidney affects as stated above.
potential Modes of Action
and Identification of Data Gaps
in relation to problem formulation
was shown that the effects in the stomach can be attributed to local
effects of DMTD, which are dose-dependent. Those effects are expected to
occur similarly in humans. Similarly, a poorer health condition can be
expected as a consequence. The occurrence of lipofuscin
in the kidneys does not need to be regarded as relevant effect as there
were no nephrotoxic findings in the clinical pathology showing
is no concrete evidence given that the obtained results may not be
relevant for humans, no species-specifity was obvious. However, as human
exposure is most likely to be way below the dosages used in the animal
study, it can be concluded that, if any responses are observed at all,
they will be of non-adverse, adaptive nature only, as the local effects
noted in the stomach seem to follow a threshold-MoA.
The oral administration of
2,5-Dimercapto-1,3,4-thiadiazole to parental Han Wistar rats at dose
levels of 50, 150 or 500 mg/kg/day for two weeks before pairing, during
pairing and then up to termination of the males after five weeks of
treatment and females on Day 14 of lactation was generally well
tolerated. However, it resulted in reduced body weight gain and
macroscopic and microscopic changes in the stomach in animals of either
sex treated with 150 and 500 mg/kg bw/day.
Target organ toxicity (repeated
exposure), as defined by the Regulation, “means specific, target organ
toxicity arising from a repeated exposure to a substance or mixture. All
significant health effects that can impair function, both reversible and
irreversible, immediate and/or delayed are included. … Classification
for target organ toxicity (repeated exposure) identifies the substance
as being a specific target organ toxicant and, as such, it may present a
potential for adverse health effects in people who are exposed to it”.
Furthermore, “substances are classified as specific target organ
toxicants following repeated exposure by the use of expert judgement”,
and “184.108.40.206. Effects considered to support classification for specific
target organ toxicity following repeated exposure...
... (c) any consistent and significant
adverse change in clinical biochemistry, haematology, or urinalysis
But also: “220.127.116.11. Effects considered
not to support classification for specific target organ toxicity
following repeated exposure
18.104.22.168.1. It is recognised that
effects may be seen in humans and/or animals that do not justify
classification. Such effects include, but are not limited to: ...
... (b) small changes in clinical
biochemistry, haematology or urinalysis parameters and/or transient
effects, when such changes or effects are of doubtful or minimal
toxicological importance; ...
... (d) adaptive responses that are
not considered toxicologically relevant;”
According to Regulation 1272/2008,
Table 3.9.3, Guidance values to assist in Category 2 classification are
10 < C≤100
mg/kg body weight/day derived from a 90 day repeated dose study. For a
short-term study, with a safety factor of 3, the boundary value would
increase to 300 mg/kg.
Based on the available information, a
classification as STOT RE Cat. 2 is not considered necessary as will be
The reduced body weight gain does not
account for any specific organ, and so only the macroscopic and
microscopic changes in the stomach in animals of either sex treated with
150 and 500 mg/kg bw/day need to be regarded.
Macropathology revealed dark areas and
depressions of the stomach which correlated with mucosal erosion/
ulceration and hemorrhage in the lamina propria. In addition, oedema,
mucosal regeneration and submucosal inflammatory cells were seen after
histopathological assessment. These stomach changes at 500 or 150
mg/kg/day were considered to be adverse and to be due to the moderate
local irritation of the test item. This effect was not present in the 50
mg/kg/day group and was considered dose-dependent
These findings represent an adverse
effect of treatment they are considered to reflect local irritation
rather than any adverse systemic toxicity of the test item. Further, the increased
incidence of cortical tubular pigment in both sexes is with 500 mg/kg
bw/d above the guidance value and human relevance is, although likely,
not absolutely evident.
the observed effects do not suffice for and justify a classification of
DMTD as STOT RE Cat. 2.
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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