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Toxicological information

Developmental toxicity / teratogenicity

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Administrative data

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2014-05-13 until 2014-06-06 (in vivo)
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2014
Report date:
2014

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
N-butylbenzenesulphonamide
EC Number:
222-823-6
EC Name:
N-butylbenzenesulphonamide
Cas Number:
3622-84-2
Molecular formula:
C10H15NO2S
IUPAC Name:
N-butylbenzenesulfonamide
Test material form:
other: liquid
Details on test material:
- Name of test material (as cited in study report): N-butylbenzenesulphonamide
- Substance type: Industrial chemical
- Physical state: Colourless clear liquid
- Analytical purity: 99,91%
- Purity test date:
- Lot/batch No.: 201309160019
- Expiration date of the lot/batch: 16 September 2015
- Stability under test conditions: Stability of N-butylbenzenesulphonamide in the vehicle (PEG 400) was assessed in the conditions employed on the study (concentration range and storage conditions of the dose formulations pending use, according to CiToxLAB Hungary study code 14/103-316AN). According to the results, the test item is stable in vehicle in concentration range of 1 mg/mL - 400 mg/mL for 24 hours at room temperature (RT, 15-30ºC) and for up to seven days when stored refrigerated at 2-8°C.
- Storage condition of test material: Controlled room temperature
- Other: Manufacturer: Proviron Functional Chemicals N.V.

Test animals

Species:
rat
Strain:
other: Hannover Wistar
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Research Models and Services, Germany GmbH, Sandhofer Weg 7, D-97633, from SPF colony
- Age at study initiation: Young adult female rats, nulliparous and non-pregnant, approximately 11 weeks old
- Weight at onset of treatment: Not exceeding ± 20% of the mean weight, and was in the range of 180-222 g.
- Housing: Standard laboratory conditions; individual housing. Type II and/or III polycarbonate cages with Lignocel® Hygienic Animal Bedding produced by J. Rettenmaier & Söhne GmbH+Co.KG (Holzmühle 1, D-73494 Rosenberg, Germany). The bedding material was considered not to contain any contaminants that could reasonably be expected to affect the purpose or integrity of the study.
- Diet (e.g. ad libitum): The animals were provided with ssniff® SM R/M “Autoclavable Complete Feed for Rats and Mice – Breeding and Maintenance” (Ssniff Spezialdiäten GmbH, D-59494 Soest Germany) ad libitum.
- Water (e.g. ad libitum): The animals were provided with tap water as for human consumption, in water bottles, ad libitum. The quality control analysis of the water is performed once every three months and microbiological assessment is performed monthly, by Veszprém County Institute of State Public Health and Medical Officer Service (ÁNTSZ, H-8201 Veszprém, József A.u.36., Hungary).
- Acclimation period: At least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19.7 – 24.6°C
- Humidity (%):33 - 66%
- Air changes (per hr): 15-20 air exchanges/hour
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 13 May 2014 To: 6 June 2014

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
polyethylene glycol
Remarks:
PEG 400
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
The test item was formulated in the vehicle (PEG 400) at the appropriate concentrations according to the dose level and volume selected, in the Central Dispensary of CiToxLAB Hungary Ltd.
Doses of 12, 40 and 120 mg/kg bw/day were selected for the study.
The oral route is a possible route of exposure to the test item in humans and is considered suitable to provide the systemic exposure required on this developmental toxicology study.
A constant volume of 1 mL/kg bw was administered to animals in all dose groups, including the controls. The individual volume of the treatment was based on the most recent individual body weights.
The control or test item dose formulations were administered to mated, sperm positive assumed pregnant female rats daily by oral gavage on a 7 days/week basis, approximately at similar times with minor variations as practical, from Gestation Day (GD) 6 to GD19.
Formulations were prepared fresh prior to administration to animals or at 2-4-day intervals and were used according to stability results.
Stability of N-butylbenzenesulphonamide in the vehicle (PEG 400) was assessed in the conditions employed on the study (concentration range and storage conditions of the dose formulations pending use, according to CiToxLAB Hungary study code 14/103-316AN). According to the results, the test item is stable in vehicle in concentration range of 1 mg/mL - 400 mg/mL for 24 hours at room temperature (RT, 15-30°C) and for up to seven days when stored refrigerated at 2-8°C.
Analysis of formulations (concentration measurements) was performed in the Analytical Laboratory of CiToxLAB Hungary Ltd. Duplicate samples were taken from the test item formulations twice during the study (during the first and last weeks of treatment). Similarly, one sample (duplicate) was taken on each occasion from the Group 1 (control) solution for concentration measurements.
All formulations were found to be in the range of 101 to 104% of nominal concentration. No test item was detected in the control samples.

VEHICLE
- Justification for use and choice of vehicle (if other than water):
Based on the results of the pilot developmental study (CiToxLAB study code: 14/103-105PE) and as confirmed in accordance with the analytical method development, polyethylene glycol (PEG 400) was selected as vehicle.
- Concentration in vehicle:
- Amount of vehicle (if gavage):
- Lot/batch no. (if required): BCBK9981V
- Purity:

Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Analysis of formulations (concentration measurements) was performed in the Analytical Laboratory of CiToxLAB Hungary Ltd. Duplicate samples were taken from the test item formulations twice during the study (during the first and last weeks of treatment). Similarly, one sample (duplicate) was taken on each occasion from the Group 1 (control) solution for concentration measurements.

All formulations were found to be in the range of 101 to 104% of nominal concentration. No test item was detected in the control samples.
Details on mating procedure:
- Impregnation procedure: cohoused
- M/F ratio per cage: 1 male: 1-3 females
- Length of cohabitation: approximately 2 hours
- No replacement of first male by another male needed.
- Further matings after two unsuccessful attempts: no
- Verification of same strain and source of both sexes: yes
- Proof of pregnancy: the presence of a vaginal plug or sperm in the vaginal smear was considered as evidence of copulation (GD0).
On GD13 and/or 14, the sperm positive females were examined for the presence of vaginal bleeding or “placental sign” (intrauterine extravasation of blood as an early sign of pregnancy in rat, which is considered to confirm implantation).
Duration of treatment / exposure:
from gestation day 6 to gestation day 19
Frequency of treatment:
daily on a 7 days/week basis
Duration of test:
Caesarean section and necropsy at gestation day 20
No. of animals per sex per dose:
102 female animals, plus an appropriate number of spares: 25-26 mated female animals/group, 4 groups (one control and 3 test item-treated groups); 24-25 pregnant female animals/group (with implantation sites at necropsy);
60 male animals for mating; no study-procedures were carried out on the male animals; untreated, proven breeders from CiToxLAB Hungary Ltd. spare colony were used.
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale:
The dose levels were set by the Sponsor in consultation with the Study Director based on available data from the previous studies, including the results of a pilot developmental study in the rat (CiToxLAB study code 14/103-105PE, entitled: “N-Butylbenzenesulfonamide (NBBS): A Dose Range Finding Toxicity Study Following Oral (Gavage) Administration in Pregnant Hannover Wistar Rat”), with the aim of inducing toxic effects but no death or suffering at the highest dose and a NOAEL at the lowest dose.
In the pilot study, marked maternal toxicity, evident in reduced body weight gain and food consumption was observed at 350 and 200 mg/kg bw/day, in addition to the markedly decreased foetal body weight. At 100 mg/kg bw/day, slightly lower body weight gain was observed in dams, (overall mean value by approximately 12%, and corrected body weight by 8%, when compared to control mean), with the mean foetal weight lower than control by approximately 13%.
Based on the above results, doses of 12, 40 and 120 mg/kg bw/day were selected for the study.
The oral route is a possible route of exposure to the test item in humans and is considered suitable to provide the systemic exposure required on this developmental toxicology study.
- Rationale for animal assignment (if not random):
The sperm-positive, assumed pregnant females were allocated to each experimental group (on each mating day) in such a way that the group averages of the body weight were as similar as possible. Females paired with the same male were allocated to different groups on the same mating day.

Examinations

Maternal examinations:
CAGE SIDE OBSERVATIONS: Yes
Time schedule: Animals were inspected for signs of morbidity and mortality twice daily (at the beginning and end of each working day). Cage-side clinical observations were made at least daily after the treatment.
The animals were monitored for any changes including pertinent behavioural changes and signs of toxicity including mortality, changes in skin, fur, eyes, mucous membranes, occurrence of secretions and excretions, and autonomic activity (e.g. lacrimation, piloerection, pupil size, unusual respiratory pattern), changes in gait, posture and response to handling as well as the presence of clonic or tonic movements, stereotypies (e.g. excessive grooming, repetitive circling), bizarre behaviour (e.g. self-mutilation, walking backwards), tremors, convulsions, salivation, diarrhoea, lethargy, sleep and coma. No such clinical signs were noted during the study.

DETAILED CLINICAL OBSERVATIONS: Yes
Time schedule: Detailed clinical observations were made on all animals at the onset of treatment (GD6) then weekly.

BODY WEIGHT: Yes
Time schedule for examinations: The body weight of each animal was recorded with a precision of ±1 g on GD 0, 3, 6, 7, 8, 9, 10, 12, 14, 16, 18 and 20.

FOOD CONSUMPTION: Yes
Food was measured with a precision of ±1 g on GD0, 3, 6, 7, 8, 9, 10, 12, 14, 16, 18 and 20. Food consumption was calculated for each interval, including GD6-20 and GD0-20.

POST-MORTEM EXAMINATIONS: Yes
Sacrifice on gestation day 20
Organs examined: The dams’ viscera were examined macroscopically for any structural abnormalities or pathological changes. The ovaries and uterus were removed and the pregnancy status ascertained.

OTHER: On GD13 and/or 14, the sperm positive females were examined for the presence of vaginal bleeding or “placental sign” (intrauterine extravasation of blood as an early sign of pregnancy in rat, which is considered to confirm implantation). The placentas were examined macroscopically, with the exception of few placentas (5 placentas of 4510 litter and one of 4519 litter).
Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
- Other: The degree of resorption was described in order to estimate the relative time of death of the conceptus.
Fetal examinations:
- External examinations: Yes: all per litter (live foetuses)
- Soft tissue examinations: Yes: half per litter (visceral examination)
- Skeletal examinations: Yes: half per litter (other half of the litter)
- Head examinations: No data
Statistics:
The statistical evaluation of data was performed with the program package SPSS PC+4.0 (SPSS Hungary Kft, Budapest) by an appropriate statistical method (Bartlett, ANOVA and Duncan, Kruskal-Wallis and Mann-Whitney U tests, Chi2). The homogeneity of variance between groups was checked by Bartlett`s homogeneity of variance test. Where no significant heterogeneity was detected a one-way analysis of variance (ANOVA) was made. If the obtained result was significant Duncan’s Multiple Range test was used to assess the significance of inter-group differences. Significant results with inter-group comparisons were further compared using Kruskal-Wallis, and Mann-Whitney U-tests.

The following data were excluded from statistical analysis:
- sperm positive but non pregnant females (total exclusion) [No. 118 (12 mg/kg), 216 (40 mg/kg) and 108 (120 mg/kg)]
- females with ≤ 5 implantation sites (total exclusion) [No. 3525, 4 implantation sites (40 mg/kg) and No. 4525, 4 implantation sites (120 mg/kg) ]

The limit for growth retarded foetuses was calculated from the average body weight of the vehicle control foetuses. A foetus was considered as growth retarded if the deviation from the mean control values was greater than minus two fold standard deviation of all control foetuses. In the present study, this cut-off value was set at 2.632 g.

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:
Maternal toxic effects:yes. Remark: Body weight gain of dams at 120 mg/kg bw/day was lower than control mean during the treatment period (by 15 and 30% for absolute and net mean values, respectively) and was accompanied by slightly decreased food consumption.

Details on maternal toxic effects:
There was no unscheduled mortality during the study.

There were no toxicologically significant clinical signs noted following administration of test item.
Immediately after the treatment, minimal salivation and excessive digging of bedding were occasionally observed at 120 mg/kg bw/day from Day 14. These observations were regarded to be procedure related.

Thin fur (focal) or focal alopecia were occasionally observed in 2/24 and 1/24 animals in the Mid and in and High dose groups, respectively (3520 Days 10-20 on dorsal area, 3504 Days 14-20 on left flank; 4515 Days 19-20 ventral thorax). Small skin lesions (scar and crust or local red coloured skin) were also occasionally observed with similar low incidence in the control, low and high dose groups (1/25, 1/24 and 1/24, respectively). These observations were regarded as incidental and not related to the test item.

Compared to the control mean, the body weight of dams at 120 mg/kg bw/day was slightly lower from Day 12 up to Day 20 (by 2.5% - 4.0%). The differences were not statistically significant.

Body weight gains of dams at 120 mg/kg bw/day dose groups were lower than control on GD6-8 (by 54-71%) up to GD12 (by 10-24%) and between GD14-20 (by 8-13%). The differences were statistically significant on GD6-7 (p<0.01) and GD7-8 and 16-18 (p<0.05).

These lower mean values were corresponding with slightly decreased food consumption. The mean body weight gain of treated dams, evaluated for the entire treatment period GD 6-20, was lower than control mean (by 15%), and the difference was statistically significant (p<0.01).

Compared to the control mean, the “corrected” body weight gain (body weight gain between GD0-20 adjusted for the gravid uterine weight) of the dams at 120 mg/kg bw/day was lower by 16%; the net body weight gain during the treatment period (body weight gain between GD6-20 adjusted for the gravid uterine weight) of the dams at 120 mg/kg bw/day was lower by 30%. The differences attained statistical significance (p<0.05 and p<0.01, respectively).

Body weight of dams at 12 and 40 mg/kg bw/day was comparable to the control mean during the treatment period. Although slightly lower body weight gain was noted at 40 mg/kg bw/day between GD6-7 (p<0.05) and GD8-9 (not statistically significant), it was transient and did not result in any differences in overall values including net body weight gains.

It should be noted, that in the Low and Mid groups, the mean body weight gains were slightly lower than control prior to the treatment period (both groups GD0-3 and the Low dose group GD 3-6) and this finding was ascribed to the individual variability of the animals.

The food consumption of dams at 120 mg/kg bw/day was lower, than control, by 8.1% (GD6-20). The difference attained statistical significance (p<0.01).
The more pronounced difference was noted at the beginning of the treatment period (approximately 9-12%). On the following days, the average food consumption was lower, than control by 6.4-9.2%. Except days GD9-10, all differences were statistically significant (p<0.01 and p<0.05).

The food consumption of dams at 12 and 40 mg/kg bw/day groups was did not differ significantly from the control throughout the treatment period. Although statistically significant differences were noted at both groups for several periods, the differences were contributable to the relatively high individual food consumptions of control dams (1511 and 1524).

At necropsy of dams (GD20), no test item related macroscopic changes were found, except few large placentae.

Effect levels (maternal animals)

Key result
Dose descriptor:
NOAEL
Effect level:
40 mg/kg bw/day (actual dose received)
Based on:
test mat.
Basis for effect level:
other: maternal toxicity

Results (fetuses)

Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:yes. Remark: At 120 mg/kg bw/day foetal toxicity evident as a reduction in mean foetal body weight (approximately 9%). In two litters mild to moderate dilatation of chorionic vessels was observed microscopically, which correspond with enlargement of placenta.

Details on embryotoxic / teratogenic effects:
The mean number of corpora lutea and the mean number of implantation sites were comparable with the controls in all treated groups.
The pre-implantation loss was at control level in all treated groups.

The total intrauterine mortality was comparable with the control.

The early embryonic loss did not differ significantly from the control in the test item treated groups.

The late embryonic loss was slightly higher, than control at 120 mg/kg bw/day (with incidence of 4/249 and 1/246 in the high dose and control group. The differences were not statistically significant. Although the causative role of the test item cannot be excluded, since it occurred at the dose level with evidence of maternal toxicity, the incidence of the observation was within the historical control range.

The mean number of viable foetuses was comparable with the control mean.

The sex distribution of foetuses did not differ significantly between the control and treatment groups both for mean and absolute numbers. However, more females than males were observed in the test item treated groups. The difference was of low magnitude, was not statistically significant, nor dose dependent, therefore it was considered to be coincidental.

At 120 mg/kg bw/day, large placentas were recorded in 2/24 litters. In 4519/6 female foetus had large placenta (weight of 1.2g). In 4510 litter all placentas (5/5) were enlarged with weight ranging between 0.93-1.45g.
Weight of randomly selected control placentas was in the normal range.

At histopathology examination, mild to moderate dilatation of chorionic vessels was observed in all examined placentas from the High dose group.

There were no toxicological significant differences in the placenta at evaluation of the 12 and 40 mg/kg bw/day treated groups.

In one litter at 40 mg/kg bw/day (3508/10 female foetus), an excess in the amount of amniotic fluid (polyhydramnions) was noted.

The weight of foetuses at 120 mg/kg bw/day was decreased, by approximately 9% evaluated by both litter mean and group mean when compared to the controls. The differences attained statistical significance (p<0.01). No significant differences in body weight were observed between male and female foetuses at the same dose level.

The incidence of body weight retarded foetuses was similar in the control and High dose groups.

There was no foetal external malformation ascribed to the test item administration.
There were no foetal visceral abnormalities ascribed to the test item administration. The number and percentages of the minor structural changes (variations) recorded in the test item treated groups did not differ significantly from the control group. The variations included for example short brachiocephalic trunk, thymic cord, small renal papilla and small kidney.
These variations occurred with low incidence across the groups and were similar with the historical control data and were considered incidental, unrelated to test item administration.

One malformed foetus was found during the visceral examination at 40 mg/kg bw/day (3508/10 female foetus). In this foetus, narrow oesophagus was noted and the finding was corresponding with polyhydramnions (an excess in the amount of amniotic fluid).
Based on the isolated occurrence, this observation was considered incidental, ascribed to individual variability and not related to treatment, however concurrent control data did not contain this observation.

No skeletal abnormalities that could be correlated with test item administration under the conditions of this study were noted at any of dose levels. The number and percentages of the abnormalities recorded in the test item treated groups was comparable to the control group.
At 120 mg/kg bw/day, no skeletally malformed foetus was found. See overview Table 8.

Minor changes (variations) occurred, including for example in sternal ossification, asymmetric ribs, fused to sternum, dumbbell shaped and/or asymmetric, bipartite vertebral bodies or 3.5 or less metatarsals, observed with low incidence throughout all groups including controls, or with lower incidence in the treated groups than in the controls.

Compared to controls, asymmetric and/or bipartite ossification of the sternum was observed with higher incidence in the test item treated groups (3 in the Low and Mid dose and 5 in the High dose groups versus 1 in control). The differences were of low magnitude and were without attaining statistical significance. Generally, this delayed ossification was limited to sternebrae 5 and 6 and taking into account incidence of all type of altered sternal ossification, it was similar across the groups. Therefore it was considered incidental or background/individual change and not an adverse effect of toxicological significance.

Two skeletally malformed foetuses were found during the study, one in the Mid dose and one in the Low dose group.
At 40 mg/kg bw/day, malformation of the ribs (intercostal and detached rib) was found in one foetus (3522/4, body weight 2.82g). Although the Historical Control data did not contain this malformation, based on the isolated occurrence (1/111) and in the Mid dose group, this observation was considered incidental.
At 12 mg/kg bw/day, in foetus 2524/3 multiple malformation of vertebrae was found. This malformation has been observed in control Wistar rats in this laboratory.

Effect levels (fetuses)

Key result
Dose descriptor:
NOAEL
Effect level:
120 mg/kg bw/day
Based on:
test mat.
Basis for effect level:
other: foetal toxicity

Fetal abnormalities

Abnormalities:
not specified

Overall developmental toxicity

Developmental effects observed:
not specified

Any other information on results incl. tables

Table 1. Maternal and Developmental toxicity study with N-butylbenzenesulphonamide (BBSA) in rats

Parameter

Group:

1

2

3

4

 

 

Dose mg/kg:

0

12

40

120

 

Maternal findings

 

 

 

 

 

 

Treated No. of dams

incidence

25

25

26

26

 

Mortality

incidence

0

0

0

0

 

Clinical observations

 

 

 

 

 

 

  excessive digging (GD14-19)

incidence

0

0

0

17

 

  salivaton, min. (GD14-19)

incidence

0

0

0

23

 

Body weight d 20

g (mean)

311,0

308,9

304,6

298,5

NS

 

% vs Control

100

-0,7

-2,1

-4,0

 

Corrected body weight GD20

g (mean)

256,4

250,9

251,4

246,5

NS

 

% vs Control

100

-2,1

-1,9

-3,9

 

Body weight gain GD6-20

g (mean)

89,6

89,3

83,7

76,3**

DN

 

% vs Control

100

-0,3

-6,6

-15

 

Net body weight gain GD6-20

g (mean)

34,9

31,3

30,5

24,3**

DN

% vs Control

100

-10

-13

-30

 

Food consumption d6-20

g

229

21,8*(1)

21,5*(1)

21,0**

DN

 

% vs Control

100

-4,8

-6,1

-8,1

 

Macroscopic findings

 

 

 

 

 

 

  large placenta

litter incid.

0

0

0

2

 

 

fetal incid.

0

0

0

6

 

Reproductive mother

findings

 

 

 

 

 

 

 No. mated

incidence

25

25

26

26

 

 No. pregnant

incidence

25

24

25

25

NS

 No. with litters

incidence

25

24

25

25

NS

 No. wiht<5 implantations

incidence

0

0

1

1

NS

 No. Observed

incidence

25

24

24

24

NS

 Pregnancy index

Index

100

96

96,15

96,15

NS

 Gestation index

Index

100

100

100

100

NS

 Early delivery

incidence

0

0

0

0

NS

 Death

incidence

0

0

0

0

NS

 Accidental kill

incidence

0

0

0

0

NS

 Total resorptions

incidence

0

0

0

0

NS

 Mean corpora lutea

Mean/group

11,1

11,8

11,6

12,0

NS

 Mean implantations

Mean/group

9,8

10,5

10,1

10,4

NS

 Pre-implantation loss

%

11,3

10,2

12, 7

12,6

NS

 Post-implantation loss

%

6,8

4,7

7,4

8,4

NS

  Early embryonic loss

%

6,4

4,4

6,5

6,8

NS

  Late embryonic loss

%

0,4

0,3

0,8

1,6

NS

  Total intrauterine mortality

%

17,0

14,5

19,0

19,8

NS

Litter findings

 

 

 

 

 

 

 Mean live fetuses

Mean/group

9,2

10,0

9,3

9,5

NS

 Sex ratio

M%

47,1

43,9

41,1

43,1

NS

 Mean pup weight (/litter)

g

3,338

3,429

3,325

3,031**

DN

 

% vs Control

100

+2,7

-0,4

-9,2

 

Malformations

 

 

 

 

 

  External

incidence

0/231

0/240

0/224

0/228

NS

  Visceral

incidence

0/116

0/120

1/113

0/116

NS

    oesophagus narrow

 

 

 

1

 

 

  Skeletal

incidence

0/115

1/120

1/111

0/112

NS

    multiple malf. vertebrae

 

 

1

 

 

 

    Intercostal &

detached rib

 

 

 

1

 

 

Variations - developmental

incidences

 

 

 

 

 

  External

incidence

0/231

0/240

0/224

0/228

NS

  Visceral

incidence

5/116

8/120

11/113

9/116

NS

  Skeletal

incidence

22/115

16/120

24/111

26/112

NS

  Retarded in body weight

incidence

13

3**

9

11

CH

 

Legend
CH = Chi Squaire Test
DN = Duncan’s Multiple Range Test;
GD: Gestation day
NS = not significant
(1) Statistical significance attributable to high individual food consumption of
 2 control dams

Applicant's summary and conclusion

Conclusions:
The No Observed Adverse Effect Level (NOAEL) under the conditions of this study for maternal toxicity was found to be 40 mg/kg bw/day and for foetal developmental toxicity 120 mg/kg bw/day.
Executive summary:

A developmental toxicity study was performed to assess the effects of the test item N-butylbenzenesulphonamide on the embryonic and foetal development (including the organogenesis period) of Hannover Wistar rats in their first pregnancy. The dams (one control and 3-treated groups) were treated daily by oral (gavage) administration, from gestation day GD6 up to and including GD19. Caesarean sections, necropsy of dams and examination of uterine contents were performed on GD20.The dose levels were 12, 40 and 120 mg/kg bw/day. Control dams were treated with vehicle polyethylene glycol (PEG 400) only.

 

The number of confirmed pregnant, evaluated dams in the dose groups treated at 12, 40 and 120 mg/kg bw/day was 24 in each of group and 25 in the control group. There was no unscheduled mortality during the study. There were no toxicologically significant clinical signs noted following administration of test item. Body weight gain of dams at 120 mg/kg bw/day was lower than control mean during the treatment period (by 15 and 30% for absolute and net mean values, respectively) and was accompanied by slightly decreased food consumption.

 

No test item related macroscopic changes were found at necropsy of dams on GD20, except few large placentae at 120 mg/kg bw/day (6 foetal and 2 litter incidences, respectively). Mild to moderate dilatation of chorionic vessels was observed in all examined placentas from the High dose group.

 

There were no test-item related significant differences between the treated and control animals regarding the pre-implantation loss or early embryonic death, post-implantation loss, or total intrauterine mortality. The mean number of viable foetuses was comparable with the control mean. The sex distribution of foetuses did not differ significantly from the controls. The weight of foetuses at 120 mg/kg bw/day was decreased by approximately 9% evaluated by both litter mean and group mean when compared to the controls. The differences attained statistical significance. No significant differences in body weight were observed between male and female foetuses at the same dose level.

 

There were no foetal external, visceral or skeletal abnormalities ascribed to the test item administration.

 

Two malformed foetuses were found at 40 mg/kg bw/day, one visceral (narrow oesophagus with polyhydramnions) and one skeletal (intercostal and detached rib). Although the Historical Control data did not contain these malformations, based on the isolated occurrence (1/113 and 1/111) in the Mid dose group, this observation was considered incidental. At 12 mg/kg bw/day, one foetus with multiple malformation of vertebrae was found. This malformation has been observed in control Wistar rats in this laboratory.

The No Observed Adverse Effect Level (NOAEL) under the conditions of this study for maternal toxicity was found to be 40 mg/kg bw/day and for foetal developmental toxicity 120 mg/kg bw/day.

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