Methanesulfonic acid, 1,1'-(1,6-hexanediyldiimino)bis[1-oxo-, sodium salt (1:2)

Developmental toxicity / teratogenicity

Administrative data

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2012

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study according to OECD TG414 and GLP

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2012

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Methansulfonic acid (1,6-hexanediyl-diimino)bis[1-oxo, disodium salt

EC Number:

690-526-2

Cas Number:

38632-47-2

Molecular formula:

C8H14S2N2O8.Na2

IUPAC Name:

Methansulfonic acid (1,6-hexanediyl-diimino)bis[1-oxo, disodium salt

Details on test material:

Purity/Concentration used for Formulation: 91.4 %. Purity was corrected after reanalysis (Certificate of Analysis LNX31603, 06 February 2012) to 84.7 %

Test animals

Species:

rat

Strain:

Wistar

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

water

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Based upon the results of stability analyses performed within the non-GLP Harlan Laboratories study D34576 (Dose Range-Finding Prenatal Developmental Toxicity Study in the Han Wistar Rat), dose formulations were stable for at least 8 days.

Details on mating procedure:

After acclimatization, females were housed with sexually mature males (1:1) in special automatic mating cages i.e. with synchronized timing to initiate the nightly mating period, until evidence of copulation was observed. This system reduced the variation in the copulation times of the different females. The females were removed and housed individually if:
- the daily vaginal smear was sperm positive, or
- a copulation plug was observed.
The day of mating was designated day 0 post coitum.

Male rats of the same source and strain were used only for mating. These male rats are in the possession of Harlan Laboratories and were not considered part of the test system. The fertility of these males had been proven and was continuously monitored.

Duration of treatment / exposure:

From day 6 post coitum (first treatment) until day 20 post coitum (last treatment);

Frequency of treatment:

once a day

Duration of test:

Day 21 post coitum (Caesarean section and necropsy)

Doses / concentrations

Doses / concentrations 1

Remarks:

Doses / Concentrations:
0 mg/kg/day
Basis:
other: control

Doses / concentrations 2

Remarks:

Doses / Concentrations:
46 mg/kg/day
Basis:
other: actual ingested based on corrected purity of 84.7 %

Doses / concentrations 3

Remarks:

Doses / Concentrations:
232 mg/kg/day
Basis:
other: actual ingested based on corrected purity of 84.7 %

Doses / concentrations 4

Remarks:

Doses / Concentrations:
927 mg/kg/day
Basis:
other: actual ingested based on corrected purity of 84.7 %

No. of animals per sex per dose:

22 mated females per group

Control animals:

yes, concurrent vehicle

Results and discussion

Results: maternal animals

Effect levels (maternal animals)

Dose descriptor:

NOAEL

Effect level:

232 mg/kg bw/day (actual dose received)

Based on:

act. ingr.

Basis for effect level:

other: maternal toxicity

Results (fetuses)

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:yes

Effect levels (fetuses)

Dose descriptor:

NOAEL

Effect level:

232 mg/kg bw/day (actual dose received)

Basis for effect level:

other: embryotoxicity

Fetal abnormalities

Abnormalities:

not specified

Overall developmental toxicity

Developmental effects observed:

not specified

Any other information on results incl. tables

Mortality and General Tolerability

All females survived until the scheduled necropsy. No test item-related clinical observations were found.

Food Consumption and Body Weights

At 1000 mg/kg body weight/day, mean food consumption, body weights and body weight gains were statistically significantly lower during the treatment period. The corrected body weight gain (corrected for gravid uterus) was also statistically significantly lower in this dose group. These were considered to be test item-related effects. At 250 mg/kg body weight/day, body weight gains were statistically significantly reduced between days 13 - 17 post coitum and this was considered to be test item-related but not adverse effect.

Reproduction Data

The number of implantation sites, pre and post-implantation losses and mean number of fetuses per dam were not affected by treatment with the test item in any dose group.

Macroscopical Findings

No macroscopical findings were noted in any animal at scheduled necropsy.

Fetal Data

External Abnormalities and Variations

No findings were noted during external examination.

Sex Ratios

No test item-related effects were noted.

Body Weights

The mean fetal body weights were statistically significantly lower at 1000 mg/kg body weight/day when compared to the control group values. This was considered to be related to the treatment with the test item.

Visceral Abnormalities and Variations

At 1000 mg/kg body weight/day, there were test item-related abnormalities noted: Absence of one or two eyes (anophthalmia) or a severely small eye (microphthalmia) was found in four fetuses in four litters. Severe dilation of brain ventricles were also observed in four fetuses in two litters in the high dose group. These abnormalities were considered to be the direct effect of the treatment with the test item. Situs inversus (thoracic and abdominal) were noted in two fetuses in two litters and one multiple malformed fetus was found with right-sided azygos vein, malpositioned adrenal, severely dilated renal pelvis and ureters on both sides and the urinary bladder was not located. An increased incidence of bilateral or right-sided azygos vein as variation was found in four fetuses in four litters and the values were outside of the historical background range. At 250 mg/kg body weight/day, situs inversus (thoracic and abdominal) was found in one fetus, and the renal pelvis and ureter was severely dilated in another fetus. Since these findings were sporadic, were within the historical control range and no other malformation or increased incidence of variations occurred in this dose group during visceral and skeletal examination of fetuses, these findings were considered to be incidental and not test item-related. At 50 mg/kg body weight/day, a retrosternal diaphragmatic hernia was found. Based on its isolated appearance and lack of dose relation this single fetal abnormality was not considered treatment-related. The abnormalities found in the 250 and 50 mg/kg body weight/day dose groups were within the historical control range of this laboratory and the incidence of variations did not give any indication of treatment-related effects therefore they were considered to be incidental and not test item-related effects.

Bone Abnormalities and Variations

At 1000 mg/kg body weight/day, test item related abnormalities were noted. The nature of some findings was correlating with findings found during visceral examination such as small orbital socket in the skull (indicating an absent or small eye) was noted in four fetuses in three litters. In one litter two fetuses were found with several bone and cartilage abnormalities affecting the skull, vertebrae and ribs and one fetus had spina bifida in a separate litter. Additionally, in the 1000 mg/kg body weight/day dose group, the following variations were found with increased incidence when compared to current controls and the values were outside of the historical control range: fused zygomatic arch, fused sternebra, sternebra offset ossification sites and/or bipartite ossification, malpositioned pelvic girdle (caudal), cervical ribs, dumbbell-shaped cervical vertebral bodies, costal cartilages asymmetrically aligned at sternum.

Ossification and Supernumerary Ribs

At 1000 mg/kg bodyweight/day, the incidence of non-ossified cervical vertebral body 1 and the incidence of non-ossified first metatarsalias on the hind limbs were statistically significantly increased and were outside of the historical background range. These findings indicated a delay in ossification in the high dose group fetuses.

Cartilage Abnormalities and Variations

At 1000 mg/kg bodyweight/day, the incidence of costal cartilage abnormalities was increased (four fetuses in three litters compared to one fetus in control). Cervical vertebral bodies and/or arches were fused in three fetuses (nos. 90, 601, 609) in two litters and thoracic vertebral bodies were fused in two fetuses (nos. 601 and 827) in two litters. These were considered to be test

item-related effects. The remaining cartilage variations did not give any indication of a test item effect.

Conclusion

The NOAEL (No-Observed-Adverse-Effect-Level) for maternal effects was 250 mg/kg body weight/day based on reduced reduced body weight, body weight gains and food consumption in the 1000 mg/kg body weight/ day dose group. Based on the increased incidence of fetal abnormalities found during visceral and skeletal examinations in the 1000 mg/kg body weight/ day dose group, the NOAEL for prenatal morphological alterations including teratogenicity was considered to be the mid dose, 250 mg/kg

body weight/day (achieved dose level 231.7 mg/kg body weight/ day).

Applicant's summary and conclusion

Executive summary:

The NOAEL (No-Observed-Adverse-Effect-Level) for maternal effects was 250 mg/kg body

weight/day based on reduced reduced body weight, body weight gains and food consumption in

the 1000 mg/kg body weight/ day dose group.

Based on the increased incidence of fetal abnormalities found during visceral and skeletal

examinations in the 1000 mg/kg body weight/ day dose group, the NOAEL for prenatal

morphological alterations including teratogenicity was considered to be the mid dose, 250 mg/kg

body weight/day (achieved dose level 231.7 mg/kg body weight/ day).