N-(cyclohexylthio)phthalimide

Toxicity to reproduction

Administrative data

Endpoint:

two-generation reproductive toxicity

Remarks:

based on test type (migrated information)

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2009

Report date:

2009

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 421 (Reproduction / Developmental Toxicity Screening Test)

GLP compliance:

yes

Limit test:

yes

Test material

Test material information

Constituent 1

Reference substance name:

N-(cyclohexylthio)phthalimide

EC Number:

241-774-1

EC Name:

N-(cyclohexylthio)phthalimide

Cas Number:

17796-82-6

Molecular formula:

C14H15NO2S

IUPAC Name:

N-(cyclohexylthio)phthalimide

Details on test material:

Test batch No.: 041/07
Active substance: 99,42 % weight
Weight loss: 0,08 % weight
Toluene insolubles: 0,05 % weight
Ash: 0,008 % weight
Melting point: first liquid 92,3°C
complete liquid 92,5 °C
Sieve retention: 2,0 mm 0 %
0,5 mm 0 %

Test animals

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

Detail on test animals
48 adult males and 48 adult females of the rats Wistar from Velaz Praha, Czech Republic were used (Protocol No 7/2009). The animals were acclimatized for 7 days before the study in the condition identical to the condition during the experiment. The acclimatisation was made according to SOPB-00188-AH, Animals quarantine. The mean body weight of animals at the start of experiment was in the males 296,25 g, in the females 203,96 g. Twelve males and twelve females were used in the control and in all dose groups.
Husbandry
All animals were kept in the cages with the bedding in the experimental animal house with the standard conditions. The sanitation was made according to the standard operation procedures. the temperature was 22±2°C, the relative humidity 45 to 65 %. The room is equipped with the room thermometer, hygrometer and central air-conditioner. The light regime was artificial 12 hour light/ 12 hour dark cycle.
The keeping of the animals was organised to the time table:
1. 4 animals in the cage (males and females separatelly)
2. 1 male and 1 female in the cage (mating)
3. 1 pregnant female individually
4. 1 female and the offspring individually
Diet and Water
For feeding the conventional laboratory diet (Top Dovo, PD Dubové - Naháč) was used with an unlimited supply of the drinking water.
Group Designation and Dose Level
Three test groups and one control group (vehicle) were used. Twelve animals of the both sexes were used in every dose level group and in the control group, too.
The animals were marked by the serial number 1-48. All cages were marked with the number of the group, the dose and the serial number of animals. The animals in the cage were marked by the line I - IIII on the tail. The males were colorated with black and the females with red. The selection of the doses was estimated from the available information about the test article. Dose of 5mg/kg was estimated as the low dose level. The high dose level - 40 mg/kg represented 8-fold of the low dose level, the medium dose level - 20 mg/kg represents 4-fold of the low dose level.
Males (black) Females (red)
Control
Olive oil (12) 1 - 12 (12) 1 - 12
Low dose group
5 mg/kg (12) 13 – 24 (12) 13 – 24
Medium dose group
20 mg/kg (12) 25 – 36 (12) 25 – 36
High dose group
40 mg/kg (12) 37 - 48 (12) 37 - 48

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

olive oil

Details on exposure:

Detail on exposure
Olive Oleum Raffinatum, product by G. Heess, Lot No L 809182
The control article was stored under laboratory conditions in dark container.
Duslin is not soluble in the water, but the olive oil was used as the vehicle.
The vehicle was applied to the control animals in the volume 0,5 ml/100 mg body weight.

Method of Application
The test article was administered per os with the metal stomachic tube every day at 8 a.m. The dose were calculated on the current bodyweight. The animals were weighed weekly. The fixed application volume of 0,5 ml/100 g body weight was used. Duslin is not soluble in the water, therefore the olive oil was used as the vehicle. The test substance was solubilised in the olive oil every day, before application. The vehicle was applied to the control animals in the volume 0,5 ml/100 mg body weight, too.

Details on mating procedure:

Normally, 1:1 (male to female) mating were used in this study. The female was placed with the same male until the pregnancy occurs or 14 days have elapsed. Every morning during the mating period the females were examined for the presence of sperms or vaginal plugs. The day 0 of the pregnancy was defined as the day a vaginal plug or smerm are found . The pregnant female was keeping individually up to 4th day post-partum.

Analytical verification of doses or concentrations:

not specified

Duration of treatment / exposure:

Acclimation / quarantine: 7 days
The males were applied 28 days - 14 days pre-mating, 14 days mating. This dosing period (28 days) is considered sufficient to enable detection of the majority of effets on male fertility and spermatogenesis.
The females were exposed 41-53 days: 14 days pre-mating, 14 days mating (with the objective of covering at least two complete oestrus cycles), 21-24 days gestation (this is dependent on the lenght of pregnancy), and 4 days lactation.

Frequency of treatment:

every day at 8 a.m.

Doses / concentrations

Doses / concentrations 1

Remarks:

Doses / Concentrations:
40 mg/kg
Basis:
nominal conc.

Doses / concentrations 2

Remarks:

Doses / Concentrations:
20 mg/kg
Basis:
nominal conc.

Doses / concentrations 3

Remarks:

Doses / Concentrations:
5 mg/kg
Basis:
nominal conc.

No. of animals per sex per dose:

No. Of animal
48 males and 48 females
Males (black) Females (red)
Control
Olive oil (12) 1 - 12 (12) 1 - 12
Low dose group
5 mg/kg (12) 13 – 24 (12) 13 – 24
Medium dose group
20 mg/kg (12) 25 – 36 (12) 25 – 36
High dose group
40 mg/kg (12) 37 - 48 (12) 37 – 48

Control animals:

yes

Positive control:

no

Examinations

Parental animals: Observations and examinations:

Experimental Schedules
The daily dosing of the both sexes began 14 days before mating. The dosing was continued in the both sexes during the mating period (max. 14 days - 2 complete oestrus cycles). The males were dosing after the mating period at least until the minimum total dosing period of 28 days. They were sacrificed by the overdose of the anaesthetic (dietylether). The daily dosing of the females were continued throughout the pregnancy and up to 4th day after the birth. The females showing no-evidence of copulation were dosing 28 days after the last day of mating period. They were sacrificed overdose of the anaesthetic, too. The females with the offspring were sacrificed on day 4 post-partum.
Mating procedure
Normally, 1:1 (1 male to 1 female) matings were used in this study. The female was placed with the same male until the pregnancy occurs or 14 days have elapsed. Every morning during the mating period the females were examined for the presence of sperms or vaginal plugs. The day 0 of the pregnancy was defined as the day a vaginal plug or sperm are found. The pregnant female was keeping individually up to 4th day post-partum.
Clinical Observations
General clinical observations were made once a day 1 hour after the test article application. The healt condition of the animals, reactiion of animals to the applied substance, their condition were monitored every day and recorded. The number of pregnant females was recorded. The number of live and death pups, weight of litters and sex of pups were observed, too.
Body Weight and Food Consumption
Individual weghting of the adult animals was performed once week. The body weight of the animals was recorded in the Application notebooks.

Statistics:

Statistics
The result obtained during the study were stistically evaluated by the statistical programme Statgraphics. The statistical evaluated was made separately for the males, females and litters. For the identification homogenity the groups were used the Barlettś test. In the case of the homogeneity One-Way
Analysis of variance with consecutive Multiple Rangers tests was accomplished. In the case non homogeneity Kruskal-Wallis One Way Analysis by ranks was applied. The relative weight of observed organs was calculated from the formula:
A(g)
R(%) = ----------------------------- x 100
Body weight (g)

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:

no effects observed

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

slightly decrease

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

slightly decrease

Effect levels (P0)

Dose descriptor:

NOAEL

Effect level:

> 40 mg/kg bw/day

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: Doses 5, 20 and 40 mg/kg/ bw/day were used. No adverse effects were observed.

Overall reproductive toxicity

Reproductive effects observed:

not specified