Registration Dossier

Toxicological information

Toxicity to reproduction

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Administrative data

Endpoint:
screening for reproductive / developmental toxicity
Type of information:
experimental study
Adequacy of study:
weight of evidence
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2010
Report Date:
2010

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 421 (Reproduction / Developmental Toxicity Screening Test)
Deviations:
no
GLP compliance:
yes
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid - liquid: suspension
Details on test material:
. Name : SODIUM THIOGLYCOLATE
. Batch number : 13483
. Description : white powder
. Purity : 98.9%

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
- Breeder: Charles River Laboratories France, L'Arbresle, France
- Age at the beginning of the treatment period: 6 weeks old 
- Weight at the beginning of the treatment period: ca. 205 g for the  males and ca. 160 g for the females.
- Food and water: ad libitum
- Acclimation period: 7 days
ENVIRONMENTAL CONDITIONS
- Temperature : 22 ± 2°C
- Relative humidity : 50 ± 50%
- Light/dark cycle : 12h/12h (7:00 - 19:00)
- Ventilation : about 12 cycles/hour of filtered, non-recycled air.
HOUSING
The animals were housed individually in polycarbonate cages or in  wire-wesh cages. Autoclaved wood shavings were provided as nesting  material, a few days before delivery and during the lactation period.

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
water
Details on exposure:
TREATMENT
- Vehicle: degassed purified water, obtained by reverse osmosis 
- Dosage form preparation: solution in the vehicle at 4, 8 and 16 mg/mL,  expressed as active substance. 
- Volume: 5 ml/kg
Details on mating procedure:
MATING
- Mating procedure: one female was placed with one male from the same  dose-level group. If necessary, the estrous cycle stage was determined  from a fresh vaginal lavage (stained with methylene blue), each morning  during the mating period until the females mated

PARTURITION
Females were allowed to drop their litters normally and rear their  progeny until sacrifice
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
On weeks 1, 4, 8 and 12. There was a satisfactory agreement (± 10%) between the nominal and  actual concentrations.
Duration of treatment / exposure:
Exposure period: * Males: during 10w before mating, the mating period (2w) and until sacrifice.
* Females: during 10 w before mating, the mating period, pregnancy and lactation until day 4 post partum.
Premating exposure period (males): 10 weeks
Premating exposure period (females): 10 weeks
Duration of test: 16 weeks
Frequency of treatment:
7 days per week
Doses / concentrations
Remarks:
Doses / Concentrations:
0, 20, 40 or 80 mg/kg/day
Basis:
actual ingested
No. of animals per sex per dose:
12
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale:
The dose-levels were selected by the Study Monitor based on the results of a preliminary 2-week toxicity study (CIT/Study No. 30720 TSR). In this study sodium thioglycolate in purified water was planned to be administered at dose-levels of 15, 30, 60 and 75 mg/kg/dayfor 14 days. After no effects were observed at all dose-levels, the group previously given 15 mg/kg/day was administered with 100 mg/kg/day on days 8, 9 and 10 and then 150 mg/kg/day on days 11, 12 and 13, the other groups were treated as scheduled until day 14. When treated at 150 mg/kg/day, one male was sacrificed on day 13 following marked clinical signs and three females were found dead on day 14. The male and two of the females had spleens reduced in size. Male terminal body weight gains were slightly lower than controls at 15/100/150 mg/kg/day while females showed a body weight gain comparable to controls until day 11 and a mean terminal body weight loss (-10%) when the dose level was increased to 150 mg/kg/day. Food consumption was lower for this group for both sexes over the last 3-day period when the dose level was increased to 150 mg/kg/d. At necropsy, some animals in the 60, 75 and 15/100/150 mg/kg/day dose-groups had irregular colored kidneys, while at 75 mg/kg/day marked lobular pattern of the liver was noted with paleness (also seen at 15/100/150 mg/kg/day). Mean ovary weights were lower at 15/100/150 mg/kg/day and mean uterus weights were lower at 75 and 15/100/150 mg/kg/day. 150 mg/kg/day was considered to exceed the maximum tolerated dose because mortality occurred after 3 days of treatment. 80 mg/kg/day was selected as a top dose level for the OECD 421 study considering the limited effects observed at 75 mg/kg/day.
Positive control:
No apprpriate

Examinations

Parental animals: Observations and examinations:
- Morbidity and mortality: at least twice a day
- Clinical signs: at least once a day
- Body weight:          
. males: on day 1, then once a week until sacrifice         
. females: on day 1, then once a week until mated, then on days 0, 7, 14  and 20 pc and on days 1 and 5 pp (postpartum).
- Food consumption         
. males: once a week (except during the mating period) until sacrifice         
. females: once a week during the premating period and then on the  following intervals: days 0-7, 7-10, 10-14, 14-17 and 17-20 pc, and days  1-5 pp.
Oestrous cyclicity (parental animals):
Each morning for three weeks before the start of the pairing period. 
Sperm parameters (parental animals):
The left epididymis was removed, weighed (total and cauda separately) and  sperm from the cauda was sampled for motility, morphology investigations  and epididymal sperm count. The left testis was weighed and ground. The resulting preparation was  diluted and sperm heads resistant to homogenization (i.e. elongated  spermatids and mature spermatozoa) was counted in a Neubauer cell.
Litter observations:
- Litter size: total litter size and numbers of pups of each sex were  recorded as soon as possible after birth. The litters were observed daily.
- Clinical signs: daily
- Body weight: days 1 and 5 pp
Postmortem examinations (parental animals):
PATHOLOGY
- Sacrifice
. males: after the end of the mating period,
. females: on day 5 pp,
. females which had not delivered by day 25 pc: on day 25 pc
. females which did not mate: 24 days after the end of the mating period,
- Organ weights: 
Brain, epididymides, heart, kidneys, liver, ovaries,  prostate, seminal vesicles and testes, uterus
- Macroscopic post-mortem examination: on all parent animals. In all  females, the number of implantation sites and corpora lutea were  recorded. 
- Preservation of tissues: macroscopic lesions, ovaries, prostate,  seminal vesicles, uterus (horns and cervix), vagina, brain, heart, liver  and kidneys, in 10% buffered formalin. Testes and epididymides, in  Bouin's fluid        
- Microscopic examination: macroscopic lesions,  epididymides, heart,  kidneys, liver, ovaries, prostate*, seminal vesicles*, testes, and  uterus (* in the control and high-dose groups only).
Postmortem examinations (offspring):
PATHOLOGY
- Sacrifice
. pups: on day 5 pp.
-  A macroscopic examination was performed for all pups, including those found dead or prematurely sacrificed. Macroscopic lesions were preserved in 10% buffered formalin (or another appropriate fixative).
Statistics:
- Data other than organ weights:
Mean values were compared by one-way variance analysis and Dunnett test, (mean values being considered as normally distributed, variances being considered as homogeneous). Values compared by Fisher exact probability test are presented as percentages.
- Organ weights:
Dunn test or Student test (2 groups) or Dunnett test (3 or more groups) or Mann-Whitney/Wilcoxon test
Reproductive indices:
Pre-implantation loss:
Number of corpora lutea - Number of implantation sites
_____________________________________________ x 100
Number of corpora lutea

Post-implantation loss:
Number of implantation sites - Number of live concepti
_____________________________________________ x 100
Number of implantations

Mating index:
Number of mated animals
_____________________ x 100
Number of paired animals

Fertility index:
Number of pregnant female partners
_______________________________ x 100
Number of mated pairs

Gestation index:
Number of females with live born pups
________________________________ x 100
Number of pregnant females
Offspring viability indices:
Live birth index:
Number of live born pups
_____________________ x 100
Number of delivered pups

Viability index on day 4 post-partum:
Number of surviving pups on day 4 post-partum
_______________________________________ x 100
Number of live born pups

Results and discussion

Results: P0 (first parental animals)

General toxicity (P0)

Clinical signs:
effects observed, treatment-related
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
no effects observed
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
effects observed, treatment-related
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Other effects:
no effects observed

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:
no effects observed
Reproductive function: sperm measures:
no effects observed
Reproductive performance:
effects observed, treatment-related

Details on results (P0)

CLINICAL EXAMINATIONS
- Mortality: 
. 80 mg/kg/day:
One female (K20428) was found dead on day 23 of dosing. No clinical signs other than ptyalism had been observed prior to death. No abnormalities were observed at post-mortem examination.
One male (K20347) was found dead on day 74 of dosing. No clinical signs other than ptyalism were observed prior to death. The male had not mated prior to death. At necropsy, the stomach mucosa had brown/black areas.
One male (K20349) was found dead on day 90 of dosing. No clinical signs other than ptyalism were observed prior to death. At necropsy, the right kidney had a dilated pelvis.
One female (K20421) was found dead on day 23 post-coitum with 17 live pups in the bedding. At necropsy, thick, blackish contents were observed in the vagina.
One female (K20422) was found dead on day 23 post-coitum with 4 dead pups and one live pup in the bedding and one pup in the vagina. At necropsy, there were 11 dead fetuses in the uterine horns.
One female (K20426) was found dead on day 23 post-coitum with 11 live pups and five dead pups in the bedding. Difficulty to deliver was noted for the female prior to death. At necropsy, the lungs were reddish in color and there were serous contents in the thoracic cavity.
One female (K20427) was prematurely sacrificed on day 23 post-coitum with signs of piloerection, cold to the touch, pallor of eye recorded only on the same day. No pups were delivered prior to death. At necropsy, there were 15 dead fetuses and one live fetus and one late resorption in the uterine horns and the liver was pale.
One female (K20429) was prematurely sacrificed on day 1 post-partum because all the pups were dead. Prior to sacrifice, emaciated appearance, piloerection and round back were observed. No abnormalities were observed at post-mortem examination.
One female (K20423) was found dead on day 2 post-partum with no clinical signs other than ptyalism observed prior to death. At necropsy, the urinary bladder was dilated.
. 40 mg/kg/day:
One female (K20415) was found dead on day 22 post-coitum. At necropsy, the female was pregnant with 16 dead fetuses in the uterine horns.
The high rate of mortality in the treated females, with presence of implantation scars, late resorptions and/or dead fetuses in the uterine horns, was considered to be treatment-related.

- Clinical symptoms: 
All males and females given 80 mg/kg/day were observed to have ptyalism from week 2 of dosing generally until the end of the study. The number of females affected during gestation (8/11) and lactation (2/4) was less than the number affected during pre-pairing (11/12).
7/12 males and 5/12 females given 40 mg/kg/day experienced ptyalism towards the end of the pre pairing dosing period (week 9 onwards) and, for males, until the end of the study, but only 3/12 females were affected during gestation and 1/11 had ptyalism during lactation.
No clinical signs were observed at 20 mg/kg/day.
- Body weight: Table 1
The mean body weight gain over the 10-week pre-mating period was similar to that of the controls for both males and females at all dose-levels. Females given 80 mg/kg/day had a slight lowering of body weight gain between day 50 and day 64 but this did not affect the overall mean gain.
There were no effects of treatment on body weight gains during gestation. Females treated at all dose-levels showed an apparently increased body weight gain during the lactation period, however there were two control females which lost weight during this period and when they are excluded there are no significant differences between the groups (there were only two females in the group mean at 80 mg/kg/day so the mean value is not as reliable as the other groups).
- Food consumption: 
Male food consumption at all dose-levels was similar to that of the controls during the first 5 weeks of dosing (ranging from -3% to +4% difference) and was then statistically significantly higher at 80 mg/kg/day for the last 5 weeks of the pre mating period (+10% to +14%).
Female food consumption at all dose-levels was similar to that of the controls throughout the pre mating and gestation periods (ranging from -5% to +16% difference). Females given 80 mg/kg/day had slightly lower mean food consumption during lactation (-10%), but given the low number of females considered in this dose group, these changes were considered to bear no biological significance.

MATING AND FERTILITY DATA
- Oestrus cycle: 
There were no effects of treatment with sodium thioglycolate on vaginal cyclicity, with mean cycle lengths of 4 to 5 days in all groups including control.
- Mating data:
With the exception of male K20347 (given 80 mg/kg/day) which was found dead after 3 days of pairing and had not mated, all pairs mated. The mean number of days taken to mate was slightly higher for the group given 40 mg/kg/day but as this was related to the contribution of two females and as it was not also observed at 80 mg/kg/day, it was considered not to be related to treatment.
- Fertility data:
There was one non-pregnant female in each of the groups given vehicle or 20 mg/kg/day and two non-pregnant females in the group given 80 mg/kg/day. This was considered not to be related to treatment with the test item.
- Delivery data: Table 2
The duration of gestation was statistically significantly longer for the females given 80 mg/kg/day than for the other groups and the number of females surviving delivery was markedly lower.
The mean number of corpora lutea was lower in females given 80 mg/kg/day resulting in a lower number of implantations and fetuses, although implantation losses were comparable with the controls and observed only in one female with total resorptions.
Pup survival was slightly worse at 80 mg/kg/day, mainly due to one female found dead so her litter was sacrificed and one female with a dead litter. The remaining two females lost only three pups between them.

PATHOLOGY
- Semiology: 
More than 96% of the sperm were morphologically normal in the groups given SODIUM THIOGLYCOLATE and more than 95% of the sperm were motile. There were therefore no effects of treatment at any dose-level.
All values fore epididymidal and testicular sperm counts are similar to, or greater than, those observed in controls, there were therefore no effects of treatment.
- Organ weights: Table 3
Higher statistically significant absolute and relative kidney and liver weights were noted in the males given 80 mg/kg/day compared with their respective controls. Higher liver weights correlated in the males given 80 mg/kg/day with a trend in higher glycogen content. The increased liver weights were considered to be related to the test item administration in view of the effect of SODIUM THIOGLYCOLATE on the carbohydrate metabolism. For the higher kidney weights, a relationship to treatment was considered to be equivocal as there were no histopathological correlates.
Dose-related lower absolute and relative seminal vesicle weights (statistically significant for the absolute weights at 20 and 40 mg/kg/day and for the absolute and relative weights at 80 mg/kg/day) were noted in the treated males. The seminal vesicle weights of some control males were higher than CIT Sprague-Dawley historical control data range (mean: 1,618/0.320; minimum: 1,185/0.257; maximum: 1,958/0,461 in absolute and relative weights repectively). Consequently, the lower seminal vesicle weights of the males given 20 or 40 mg/kg/day were not considered to be toxicologically relevant while they were considered to be treatment-related at 80 mg/kg/day, since they correlated with slight decrease in secretory content in the seminal vesicles.
Slightly lower absolute and relative prostate weights were noted in the males given 80 mg/kg/day. This was the contribution of 3/10 animals with individual prostate weights below the control range. No correlates were noted at microscopic examination of group 4 and a relationship to treatment was accordingly considered to be unlikely.
Higher mean absolute and relative uterus weights noted in the females given 80 mg/kg/day was the contribution of two females among the five surviving females. The particularly high uterus weight was due to green contents or black content together with late resorption, respectively.
- Macroscopic post-mortem examination: 
. Animals prematurely killed or found dead
Males
No major factors contributing to death were observed in the two males found dead at 80 mg/kg/day. Brown/black areas noted in stomach mucosa and right kidney dilated pelvis noted respectively in male numbers K20347 and K20349 were considered to be without relationship to treatment with the test item.
Females
One among 12 females given 40 mg/kg/day and 7/12 females given 80 mg/kg/day were found dead or prematurely sacrificed during pregnancy or lactation.
For females K20423 and K20428 (80 mg/kg/day) found dead at day 98 (day 2 post-partum) or 23, respectively and for female K20429 (80 mg/kg/day) prematurely sacrificed at day 101 (day 1 post partum), no necropsy observations were recorded, except dilated urinary bladder for female K20423. For almost all other females found dead or prematurely killed macroscopic observations were noted in the uterine horns and sometimes in vagina.
In addition, the liver was pale in female K20427 (80 mg/kg/day) prematurely killed on day 98 (day 23 post-coitum). This correlated with marked vacuolated foamy hepatocytes and minimal hepatocellular degeneration/necrosis and granulocyte infiltration. A relationship to treatment was considered to be unlikely as such finding was not found in the females which survived nor in the males.
. Survivors
Males
Seminal vesicles were of small size in 1/12 males given 80 mg/kg/day. This lower weight correlated with reduced seminal vesicle secretion and was considered to be treatment-related.
Females
Uterine horn content and/or vaginal content in females killed at termination

Dose-level (mg/kg/day) 0 20 40 80
Liquid green content/dilatation (horns) - - - (K0) K20425
Late Resorption (left horn) - - - (K0) K20431
Thick black content (left horn) - - - (K0) K20431
Thick black content (vagina) - - - (K0) K20431
(K0): scheduled sacrifice.
Late resorption was considered to be treatment-related.

- Microscopic examination:         
. Males
Systemic (scheduled sacrifices and unscheduled sacrifices)
* Heart
Minimal or slight myocardial degeneration/necrosis was seen in the two males given 80 mg/kg/day and found dead at day 74 or 90.
As these findings were recorded focally and moreover also found in the control animals which survived (2/12 males), they were considered without relationship to treatment. This finding is commonly seen in untreated Sprague-Dawley rats of this age (Greaves, 2007).
* Liver
A trend in marginally higher concentration of glycogen content was observed in the liver of the surviving males given 80 mg/kg/day.
Incidence and mean severity [affected and (affected + non affected)] of glycogen content in the liver of decedents and surviving males

Dose-level (mg/kg/day) 0 20 40 80
Number of males with glycogen content
Status: found dead - - - 0/2
Number of males with glycogen content
Status: scheduled sacrifice 10/12 10/12 8/12 9/10
Mean severity (affected animals) (3) (2.5) (2.1) (3.9)
Mean severity (affected + non affected) 2.5 2.1 1.4 3.5
-: not applicable.

This finding was considered to be related to the test item administration in view of the effect of SODIUM THIOGLYCOLATE on the carbohydrate metabolism. However these marginal differences at microscopic examination of the liver given the high dose-level were considered not to be adverse.
In addition, a relationship to the slightly higher liver weights at 80 mg/kg/day in the males which survived and which were not fasted before sacrifice was considered to be uncertain as the marginally lower glycogen content at 40 mg/kg/day in the males was not correlated with lower liver weights.
* Kidneys
Acidophilic globules were noted in the cortical tubular epithelium of the kidneys of the males given 80 mg/kg/day and of their respective controls. Incidence and severity were similar in both groups. It was therefore considered not to be toxicologically meaningful and related to the intra tubular alpha micro-globulin deposits in renal cortex which can be found at such incidence and severity in treated as well as untreated control male rats.

Incidence and mean severity (affected and affected + non affected) of acidophilic globules in the kidneys of decedents and surviving males

Dose-level (mg/kg/day) 0 80
Number of males with acidophilic globules
(mean severity)
Status: found dead - 1/2 (2.0)
Number of males with acidophilic globules
(mean severity)
Status: scheduled sacrifice 9/12 (1.7) 8/10 (1.9)
Total number of males and mean severity 9/12 (1.7) 9/12 (1.9)
Mean severity (affected + non affected) 1.3 1.4
-: not applicable.

* Genital Organs
Testes/Epididymides
The microscopic examination of the PAS/Hematoxylin stained testes and epididymides with knowledge of the different stages of maturation of the seminiferous tubules did not reveal any disturbance in the treated males. There was no evidence of degenerative changes or delay in sexual maturity in the treated males from any group compared with the respective controls. The higher number of males given 40 mg/kg/day with focal tubular atrophy was considered to be fortuitous and correspond to the junction between seminiferous tubules and tubulu recti, as it was almost always found near albuginea (subcapsular).
Seminal vesicles
Slight reduced secretory content was noted in 5/10 surviving males given 80 mg/kg/day. It correlated with lower weights at necropsy and was considered to be treatment-related. Reduced secretory content, most often unilateral and of minimal or slight severity, was also observed in 5/12 males given 20 mg/kg/day. Unilateral reduced secretory content was noted at minimal severity in 3/12 males given 40 mg/kg/day, while unilateral slight or moderate reduced secretory content was observed in two other males given 40 mg/kg/day. Except for male K20328 given the low dose-level, the individual seminal vesicle weights of the males given 20 or 40 mg/kg/day were of same magnitude as those of the controls.
It was considered that the reduced secretory content at 20 and 40 mg/kg/day was not treatment related.
In addition, as no atrophy of seminal epithelium could be observed in any treated group, the reduced secretory content together with lower seminal vesicle weights at 80 mg/kg/day was considered not to be adverse. This was confirmed by the absence of any abnormality in the sperm parameters.

- Females
Systemic (scheduled sacrifices and unscheduled sacrifices)

* Heart
Minimal myocardial degeneration/fibrosis was seen in 1/7 females prematurely killed. As this finding was recorded focally, and moreover found in one control female that survived, it was not considered to be related to the test item administration.

* Liver
Slight or marked peri/medio-lobular vacuolated hepatocytes were seen in 3/7 females found dead or prematurely sacrificed given 80 mg/kg/day. In 1/7 females marked vacuolated hepatocytes were associated with minimal hepatocellular degeneration/necrosis and minimal granulocyte infiltration. A relationship to treatment was considered to be equivocal as such observations were not found in any surviving female given 80 mg/kg/day. In control and treated (80 mg/kg/day) groups, half of the females (almost all surviving and one found dead) showed similar severity of glycogen content (see table below).

Incidence and mean severity (affected and (affected + non affected)) of glycogen content in the liver of decedents and surviving females

Dose-level (mg/kg/day) Number of females with glycogen content
(mean severity)
0 80
Status: found dead - 1/7 (3.0)
Status: scheduled sacrifice 6/12 (2.3) 4/5 (2.8)
Total number of females (affected animals) 6/12 (2.3) 5/12 (2.8)
Mean severity (affected + non affected) 1.2 1.2
-: not applicable.

* Kidneys
Slight bilateral vacuolated cortical tubules, tubular dilatation and unilateral, minimal, focal cortical necrosis were observed in the kidneys of female K20427 given 80 mg/kg/day and sacrificed moribund. Slight proteinaceous casts were noted in the kidneys of female K20421 from the same group. None of these were noted in any of the surviving females. All were thus considered to be of no toxicological importance.

* Uterus/vagina
Microscopic examination of uterus and/or vagina was suggestive of on-going pregnancy in the females found dead or prematurely sacrificed or evidence that females had not delivered. It was recorded in the table 4.
No relevant microscopic observations were recorded in the ovaries.

All the other microscopic findings noted in the prematurely killed, found dead or surviving males and females given 80 mg/kg/day or from the control group, including those observed in the prostate of the males, were those which can be found spontaneously in the untreated laboratory rat of this strain and were considered to bear no relationship to treatment with the test item.

Effect levels (P0)

open allclose all
Dose descriptor:
NOEL
Effect level:
20 mg/kg bw/day (actual dose received)
Based on:
act. ingr.
Sex:
male/female
Basis for effect level:
mortality
body weight and weight gain
Dose descriptor:
LOAEL
Effect level:
40 mg/kg bw/day (actual dose received)
Based on:
act. ingr.
Sex:
male/female
Basis for effect level:
mortality
body weight and weight gain
Dose descriptor:
NOAEL
Remarks:
reproductive performance
Effect level:
>= 80 mg/kg bw/day (actual dose received)
Based on:
act. ingr.
Sex:
male
Basis for effect level:
other: no effects observed
Dose descriptor:
NOEL
Remarks:
reproductive performance
Effect level:
20 mg/kg bw/day (actual dose received)
Based on:
act. ingr.
Sex:
female
Basis for effect level:
reproductive performance
Dose descriptor:
LOAEL
Remarks:
reprodutive performance
Effect level:
40 mg/kg bw/day (actual dose received)
Based on:
act. ingr.
Sex:
male/female
Basis for effect level:
reproductive performance

Target system / organ toxicity (P0)

Critical effects observed:
no

Results: F1 generation

General toxicity (F1)

Clinical signs:
no effects observed
Mortality / viability:
mortality observed, treatment-related
Body weight and weight changes:
no effects observed
Sexual maturation:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
no effects observed
Histopathological findings:
not examined

Details on results (F1)

OBSERVATION OF THE PUPS AFTER BIRTH
- Mortality: 
Pup survival was slightly worse at 80 mg/kg/day, mainly due  to one female was found dead so her litter was sacrificed and one female  had a dead litter. The remaining two females lost only two pups between  them.
- Clinical signs: 
One pup in the control group had a haematoma on the  head and one pup in the 40 mg/kg/day dose-group had a necrosed tail. As  these were isolated findings, they were considered to be spontaneous in  origin. 
- Pup body weight: 
Pups of dams treated at 40 or 80 mg/kg/day showed an increased mean body weight gain from day 1 to day 5 post-partum (ranging from +14% to +51% difference). The pups of dams treated at 80 mg/kg/day also had a higher mean birth weight than the controls (+12% for males and +14% for females) but there were fewer pups per litter in this group which is likely to have had an impact. Since this increased body weight gain is dose-related it is considered to be related to treatment with the test item.
- Sex ratio: 
The percentage of male pups was slightly low at 80 mg/kg/day  (ns), when compared with the controls, however there were less pups in  this group.
- Pup necropsy findings: 
One pup in the 40 mg/kg/day dose-group had a small liver with whitish areas. One pup in the 20 mg/kg/day group had whitish areas on the liver.
These findings were not observed in the 80 mg/kg/day dose-group. The examination of  the historical control data shown that whitish area have already been  observed in control animals in 2 OECD 421 studies with frequencies of  1/125 (0.8%) and 1/113 (0.9%), in consequence this effect was not  considered to be treatment-related.

Effect levels (F1)

open allclose all
Dose descriptor:
NOEL
Remarks:
toxic effects on progeny
Generation:
F1
Effect level:
40 mg/kg bw/day (actual dose received)
Based on:
act. ingr.
Sex:
male/female
Basis for effect level:
viability
Dose descriptor:
LOAEL
Remarks:
toxic effects on progeny
Generation:
F1
Effect level:
80 mg/kg bw/day (actual dose received)
Based on:
act. ingr.
Sex:
male/female
Basis for effect level:
viability

Overall reproductive toxicity

Reproductive effects observed:
not specified

Any other information on results incl. tables

Table 1: main differences in mean body weight and body weight gain (in grams)

Sex

Male

Female

Dose-level (mg/kg/day)

0

20

40

80

0

20

40

80

Pre-mating

 

 

 

 

 

 

 

 

 . days 1-71

+315

+305
(-3%)

+304
(-3%)

+315
(0%)

+133

+130
(-2%)

+125
(-6%)

+129
(-3%)

Gestation

 

 

 

 

 

 

 

 

 . days 0-20p.c.

/

/

/

/

+120

+128
(+7%)

+127
(+6%)

+112
(-7%)

Lactation

 

 

 

 

 

 

 

 

 . days 1-5 p.p.

/

/

/

/

+11

+20

(+82%)

+18

(+64%)

+13

(+18%)

/: not recorded

Table 2: delivery data

Dose-level (mg/kg/day)

0

20

40

80

Duration of gestation (days)

21.6

21.4

21.5

22.8**

. Number of females delivering on day 21p.c.

4

7

6

 

. Number of females delivering on day 22p.c.

7

4

5

2

. Number of females delivering on day 23p.c.

 

 

 

1

. Number of females delivering on day 24p.c.

 

 

 

1

Number of females surviving delivery/number of pregnant females

11/11

11/11

11/12

4/8a

Mean number ofcorpora lutea

17.0

15.7

14.9

13.2

Mean number of implantations

16.5

15.4

14.4

10.3#

Mean number of live pups delivered

14.7

14.7

13.4

9.0**

**: p<0.01, #: p<0.001.

a: excluding female K20431 which had total resorptions.

Table 3: differences (expressed in %) noted between treated and control animals in the absolute and relative organ weights

 

Sex

Male

Female

Group

2

3

4

2

3

4

Dose-level (mg/kg/day)

20

40

80

20

40

80

Body weight

-1

-2

0

-1

0

-4

- Kidneys

 

 

 

 

 

 

  . absolute

-3

-1

+13**

0

+1

-5

  . relative

-2

0

+13**

+1

+1

-1

- Liver

 

 

 

 

 

 

  . absolute

-1

+1

+15*

+1

+9

+1

  . relative

0

+3

+15**

+2

+9

+5

- Prostate

 

 

 

 

 

 

  . absolute

+4

+1

-10

 

 

 

  . relative

+4

+3

-11

 

 

 

- Seminal vesicles

 

 

 

 

 

 

  . absolute

-17*

-19*

-35**

 

 

 

  . relative

-16

-18

-35**

 

 

 

- Uterus

 

 

 

 

 

 

  . absolute

 

 

 

-3

+1

+161

  . relative

 

 

 

-1

+2

+170

Statistically significant from controls: *: p<0.05, **: p<0.01.

The significance concerned the organ weights values and not the percentages.

Table 4

Females prematurely killed or found dead

 

Dose-level (mg/kg/day)

Day*

Status**

40

80

Female

Number/status
at necropsy

Macroscopic finding

Microscopic findings

Macroscopic finding

Microscopic findings

K20415

97

Dead during pregnancy (22)

 

Dead fetuses (16)

 

Gestational uterus

Marked vaginal mucification

-

-

 

 

 

 

 

 

 

K20421

102

Dead during pregnancy (23)

-

-

Implantation scars (18)

Black thick vaginal content

Moderate vaginal mucification

 

 

 

 

 

 

 

K20422

96

Dead during pregnancy (23)

-

-

Dead fetuses (11)

Implantation scars (6)

Gestational uterus

Remnants of membranes and ombilical cord

 

 

 

 

 

 

 

K20423

98

Dead

during lactation (2)

-

-

No macroscopic observation

 

Trophoblast remnants

 

 

 

 

 

 

 

K20426

97

Dead during pregnancy (23)

-

-

Implantation scars (16)

Gestational uterus

 

 

 

 

 

 

 

K20427

98

Prematurely sacrificed during pregnancy (23)

-

-

Alive fetus (1)

Dead fetuses (15)

Late resorption (1)

Gestational uterus

 

 

 

 

 

 

 

K20429

101

Prematurely sacrificed during lactation (1)

 

 

No macroscopic observation

Slight neutrophil infiltration with necrosis and giant cells with hypertrophic nuclei

*: day of autopsy, **: gestational day or dead during lactation.

(n): number of alive or dead fetuses, late resorptions or implantation scars in uterine horns.

 


Females killed after delivery

 

Dose-level (mg/kg/day)

Day*

Status**

40

80

Female Number/status at necropsy

Macroscopic finding

Microscopic findings

Macroscopic finding

Microscopic findings

K20431

99

Final sacrifice on lactation day 5

-

-

Late resorption (1)

Thick black horn and vagina content

Minimal endometrium atrophy (epithelium and stroma)

moderate mucification of vagina

*: day of autopsy,  **: gestational day or dead during lactation.

(n): number of resorptions.

Applicant's summary and conclusion

Conclusions:
Under the experimental conditions of this study:
. the No Observed Adverse Effect Level (NOAEL) for parental toxicity was considered to be 20 mg/kg/day (based on deaths at 40 and 80 mg/kg/day),
.   male reproductive performance was not affected by treatment with sodium thioglycolate. Dosing at 40 and 80 mg/kg/day resulted in deaths in late gestation associated with delayed delivery and a No Observed Effect Level (NOEL) for female reproductive performance was therefore set at 20 mg/kg/day,
.   the NOEL for toxic effects on progeny was set at 40 mg/kg/day, based on the dead litter at 80 mg/kg/day.
A proper evaluation of the reproductive performance of females given 80 mg/kg/day was not possible because of the numerous deaths observed in this group in the peri-natal period.
Executive summary:

In a reproduction/developmental screening test performed according to the OECD Guideline # 421, four groups of 12 male and 12 female Sprague-Dawley rats received sodium thioglycolate (purity 98.9% pure), daily, by oral (gavage) administration, 10 weeks before mating and through mating and, for the females, through gestation until day 5 post-partum,at dose-levels of 0, 20, 40 or 80 mg/kg bw/d.

Clinical signs and mortality were checked daily. Body weight and food consumption were recorded weekly until mating and then at designated intervals throughout gestation and lactation. The animals were paired for mating and the dams were allowed to litter and rear their progeny until day 5 post-partum. The total litter sizes and numbers of pups of each sex were recorded after birth, pup clinical signs were recorded daily and pup body weights were recorded on days 1 and 5 post-partum. The males were sacrificed after completion of the mating period and the females on day 5 post-partum (or on day 25 post-coitum for females which did not deliver). The body weight and selected organs (brain, epididymides, heart, kidneys, liver, ovaries, prostate, seminal vesicles and testes and uterus) were weighed and a macroscopicpost-mortemexamination of the principal thoracic and abdominal organs and a microscopic examination of selected organs (macroscopic lesions, epididymides, heart, kidneys, liver, ovaries, prostate, seminal vesicles, testes, and uterus) were performed. In the females, which were apparently non-pregnant, the presence of implantation scars on the uterus was checked using ammonium sulphide staining technique. Epididymal sperm was sampled for motility, morphology and count and testicular sperm heads resistant to homogenization (i.e. elongated spermatids and mature spermatozoa) were counted. The pups were sacrificed on day 5 post-partum and were carefully examined for gross external abnormalities and a macroscopicpost-mortemexamination was performed.

 

Two males (weeks 11 and 13) and one female (week 4) given 80 mg/kg/day were found dead during the pre-mating or mating periods with no clinical signs observed before death and no relevant post-mortem findings. These deaths are considered to be treatment-related. Three out of 11 surviving females given 80 mg/kg/day were found dead on day 23 post-coitum, all having delivered pups, although one female had one fetus in the vagina and still had 11 dead fetuses in the uterine horns at necropsy. Another pregnant female with dead and live fetuses in the uterine horns was sacrificed on day 23 post-coitum because of poor clinical condition. At 80 mg/kg/day, one additional female was prematurely sacrificed on day 1post-partumbecause all the pups were dead and another female was found dead on day 2 post-partum.

One female given 40 mg/kg/day was found dead on day 22 post-coitum, pregnant with dead fetuses in the uterine horns.

Ptyalism was observed at 40 and 80 mg/kg/day with a dose-related incidence and may be related to the taste of the test item.

 

There were no significant effects of treatment on mean body weight gains; all sodium thioglycolate-treated male and female groups had body weight gains similar to those of the control group throughout the study. There were no effects of treatment on mean food consumption, except a slight lowering of food consumption during the lactation period for the two females given 80 mg/kg/day (-10%).

There were no effects of treatment with sodium thioglycolate on vaginal cyclicity, with mean cycle lengths of 4 to 5 days in all groups including control. All pairs mated and the majority of the females were pregnant. There were no effects of treatment on the mean number of days taken to mate.

Females given 80 mg/kg/day had a significantly longer gestation period (22.8, p<0.01,vs. 21.6 days), a non‑significantly lower number of corpora lutea (mean of 6.7, ns, vs. 8.5) and a significantly lower number of implantations (10.3, p<0.001,vs.16.5) and pups (9.0, p<0.01, vs. 14.7). One female had total resorptions and one litter died on day 1 post-partum.

There were no treatment-related pup clinical signs or necropsy findings. Pups treated at 40 or 80 mg/kg/day had higher mean body weight gains than the controls between day 1 and day 5 post‑partum.

There were no effects of treatment on sperm morphology, motility or counts. The mean liver and kidneys weights were slightly but statistically significantly higher for males given 80 mg/kg/day (+15% for liver and +13% for kidneys in absolute weights). Higher liver weights correlated with a trend towards increased glycogen content at this dose-level and was considered to be related to the test item administration.For the higher kidney weights, a relationship to treatment was considered to be equivocal as there were no histopathological correlates.The mean absolute seminal vesicle weights were statistically significantly lower for all male groups in a dose-related manner (absolute weights were -17%, -19% and -35% at 20, 40 and 80 mg/kg/day, respectively). This correlated with a slight decrease in secretory content in the seminal vesicles observed at microscopic examination of the males given 80 mg/kg/day. In the absence of atrophy of seminal epithelium at microscopic examination, these minor findings were not considered to be adverse.

 

The dose-level of 80 mg/kg/day was considered to be higher than the Maximum Tolerated Dose for a dosing period of 13 weeks as there were two males and one female found dead during the premating or mating periods. In addition, treatment at this lethal dose was associated with delayed delivery as four females were found dead or prematurely sacrificed after the normal period for delivery and had not delivered all the pups. Administration of sodium thioglycolate at this lethal dose of 80 mg/kg/day was also associated with the death or premature sacrifice of two additional females during the peri-natal period (from day 1 to day 2post-partum). The female sacrificed was killed on day 1post-partumbecause all its litter died. There were no effects on male or female mating behavior or fertility and the test item was considered not to hinder embryo-fetal development. The test item at the mid and high dose-levels caused increased pup body weight gain after birth but there were no relevant pup clinical signs orpost-mortemfindings. In males, when compared to controls, liver and kidneys weights were slightly but significantly higher. There were no relevant macroscopic or microscopic findings nor any effects on sperm morphology, motility or counts.

At 40 mg/kg/day, one pregnant female with dead fetuses in the uterine horns was found dead on day 22post-coitum. There were no effects of treatment on body weight, food consumption, male or female mating behavior and fertility and pregnancy parameters. There were no adverse effects of treatment on the pup body weight gain after birth.

There were no effects of treatment at 20 mg/kg/day.

Under the experimental conditions of this study:

.   the No Observed Adverse Effect Level (NOAEL) for parental toxicity was considered to be 20 mg/kg/day (based on deaths at 40 and 80 mg/kg/day),

.   male reproductive performance was not affected by treatment with sodium thioglycolate. Dosing at 40 and 80 mg/kg/day resulted in deaths in late gestation associated with delayed delivery and a No Observed Effect Level (NOEL) for female reproductive performance was therefore set at 20 mg/kg/day,

.   the NOEL for toxic effects on progeny was set at 40 mg/kg/day, based on the dead litter at 80 mg/kg/day.

A proper evaluation of the reproductive performance of females given 80 mg/kg/day was not possible because of the numerous deaths observed in this group in the peri-natal period.