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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
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
Study was conducted in accordance with a recognized scientific procedure for determining the developmental toxicity of a test substance when administered repeatedly via inhalation. Study was conducted incompliance with GLP regulations. The study meets national and international scientific standards (OECD 414) and provides sufficient information to support the conclusions regarding the NOAEL and the LOAEL demonstrated from the study data.

Data source

Reference
Reference Type:
publication
Title:
Developmental toxicities of methacrylic acid, ethyl methacrylate, n-butyl methacrylate, and allyl methacrylate in rats following inhalation exposure.
Author:
Saillenfait AM, Bonnet P, Gallissot F, Peltier A, Fabries JF
Year:
1999
Bibliographic source:
Toxicol Sci 50: 136-145

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
GLP compliance:
not specified
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Supplier : Fluka Chemie AG
- Name of test material (as cited in study report): n-butyl methacrylate
- Analytical purity: 99%

Test animals

Species:
rat
Strain:
Sprague-Dawley
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: IFFA CREDO, Saint Germain sur l'Arbresle, France
- Age at study initiation: sexually mature females, age not specified
- Weight at study initiation: 180-200  grams
- Housing: singly in polycarbonate cages
- Diet (e.g. ad libitum): UAR, Villemoisson, France
- Water (e.g. ad libitum): filtered tap water
- Acclimation period: 2 weeks

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 +/- 2
- Humidity (%): 50 +/- 10
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:
inhalation
Type of inhalation exposure (if applicable):
whole body
Vehicle:
unchanged (no vehicle)
Details on exposure:
Exposures were conducted in 200-L glass/stainless-steel inhalation chambers with dynamic and adjustable laminar air flow (6±20 m3/h). In order to prevent any leakage of the test atmospheres, the chambers were maintained at a negative pressure of no more than 3-mm water. The chamber temperature was set at 23 +/- 2°C and the relative humidity at 50 +/- 5%. Food and water were withheld during exposures.
The system used for vapor generation consisted in delivering a constant rate of liquid chemical with an infusion pump at the top of a heated glass column filled with glass beads. Compressed air heated by a glass heater was introduced at the bottom of the glass column in a countercurrent fashion to the liquid flow , an additional air flow rate was passed through the fritted disk of a heated bubbler containing the test chemical. The vaporized compounds were introduced into the main air-inlet pipe of the exposure chambers.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Concentrations were monitored continuously with a gas chromatograph equipped with a flame ionization detector and an automatic gas-sampling valve. In addition, the exposure levels were determined once during each 6-h exposure period by collecting atmosphere samples through glass tubes packed with activated charcoal. The charcoal samples were then desorbed with carbon disulide. The resulting samples were then analyzed by gas chromatography using appropriate interval standards.
Details on mating procedure:
Alter 2 weeks of acclimatization, nulliparous female rats were housed overnight with adult males (1 male:2 or 3 females) from the same strain and supplier. The day that vaginal smears were found to be sperm-positive was considered day 0 of gestation (GD). Mated females were randomly assigned to treatment groups using a randomization system stratified by body weight on GD 0
Duration of treatment / exposure:
days 6 - 20 of gestation
Frequency of treatment:
6 hours per day
Duration of test:
On GD 21, the females were sacrificed
No. of animals per sex per dose:
22-25 pregnant females per dose
Control animals:
other: yes, concurrently to filtered air

Examinations

Maternal examinations:
CAGE SIDE OBSERVATIONS: No data

DETAILED CLINICAL OBSERVATIONS: No data

BODY WEIGHT: Yes
- Time schedule for examinations: Maternal body weights were recorded on GDs 0, 6, 13, and 21.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Yes
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
Food consumption was measured for the intervals GDs 6-13 and 13-21

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No data

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day # 21
- Organs examined: no data
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
- Number of death and live fetuses: Yes
Fetal examinations:
- External examinations: Yes: All live fetuses were weighed, sexed, and examined for external anomalies including those of the oral cavity
- Soft tissue examinations: Yes: Half of the live fetuses from each lifter were preserved in Bouin's solution and examined for internai soft-tissue changes (Barrow and Taylor, 1969; Wilson, 1965).
- Skeletal examinations: Yes: Half of the live fetuses from each lifte were fixed in ethanol (70%), eviscerated, and then processed for skeletal staining with alizarin red S for subsequent skeletal examination (Staples and Schnell, 1964).
- Head examinations: No data
Statistics:
The number of CL, implantation sites, and live  fetuses, maternal food consumption and various body weights were analyzed  by ANOVA, followed by Dunnett'st-test. the percentage of non-live  implant, resorptions,and males and the proportion of fetuses with  alterations ineach litter were evaluated by Kruskal-Walles test followed  by Dixon-Massey test. Rates of pregnancy and percentage of litters with  any malformations or external, visceral, or skeletal variations were  analyzed using Fisher's test. Where appropriate, least squares analysis  was performed. The level of significance was p < 0.05.
Indices:
FERTILITY AND REPRODUCTIVE PERFORMANCE: The following data were recorded  for each group of numbers of CL, and implantation sites 
- number of   resorptions and viable and dead fetuses. 
- mean fetal body weights 
- fetuses examined for gross malformations and skeletal abnormalities of  sex and of fetuses.
Historical control data:
no data

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:
Maternal toxic effects:yes

Details on maternal toxic effects:
No test dam died. Exposure to 300 ppm or higher concentrations of n-butyl methacrylate led to a significant decrease in maternal body weight gain during the first half of exposure (Table 1). The overall maternal weight gain during GDs 6-21 and the absolute weight gain were significantly less than control at 1200 ppm. A slight reduction in maternal food consumption was observed during the first half of exposure at 300 ppm and higher concentrations (p < 0.05 and p <0.01 at 300 and 1200 ppm, respectively). No adverse effects on the average number of implantations and live fetuses, incidence of non-live implants or resorptions, or fetal sex ratio were noted among litters exposed to n-butyl methacrylate (Table 2).

Effect levels (maternal animals)

Dose descriptor:
NOAEC
Effect level:
100 ppm
Based on:
test mat.
Basis for effect level:
body weight and weight gain

Results (fetuses)

Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:yes

Details on embryotoxic / teratogenic effects:
Fetal body weight was significantly reduced at 600 ppm (females) and at 1200 ppm (all, male and female fetuses). These decreases amounted to 5% of the concurrent control values at 1200 ppm. Occasional visceral malformations occurred in low frequency and were distributed across the different groups, including control (Table 3). There were no significant differences between the control and treated groups in the incidences of either individual or total external and visceral variations, or of individual skeletal variations. The only statistically significant changes were higher mean percentages of fetuses with skeletal variations or any variations at the highest concentration of n-butyl methacrylate, compared with the concurrent control. The biological relevance of these findings is limited by the fact that the observed incidences occurred with no clear concentration-related pattern. These increases might be considered at most as slight fetotoxicity.

Effect levels (fetuses)

Dose descriptor:
NOAEC
Effect level:
300 ppm
Based on:
test mat.
Sex:
female
Basis for effect level:
fetal/pup body weight changes

Fetal abnormalities

Abnormalities:
not specified

Overall developmental toxicity

Developmental effects observed:
not specified

Any other information on results incl. tables

TABLE 1

Change in Weight During Gestation in Sprague-Dawley Rats Inhaling n-Butyl Methacrylate on Days 6 to 20 of Gestation and Euthanized on Day 21

Concentration

ppm/6 h/day

No. of damsa

Body weight (g)

on GD 6

Body weightgain (g) on GD

Absolute

weight gain (g)b

6±13

13±21

6±21

0

24

257 ± 15

33 ± 6

100± 16

133 ± 19

27 ± 10

100

24

259 ± 16

31 ± 8

100± 15

132 ± 18

26 ± 8

300

25

258 ± 15

27 ±6**

106± 12

132 ± 14

25 ± 9

600

22

264 ± 18

26 ± 5**

102± 19

128 ± 19

21 ± 10

1200

25

261 ± 18

21 ± 6**

98 ± 13

119 ± 15*

19 ± 9**

Note.Values are expressed as means ± SD.

a Includes all dams pregnant at euthanization.

bDay 21 body weight) 2 (gravid uterus weight) 2 (Day 6 body weight).

*,** Denote signi®cant differences from the control (0 ppm) value,p , 0.05 and p , 0.01, respectively.

TABLE 2
Reproductive Parameters in Sprague-Dawley Rats Inhaling Methacrylic Acid, Ethyl Methacrylate, n-Butyl Methacrylate, or Allyl Methacrylate
on Days 6 to 20 of Gestation and Euthanized on Day 21

Concentration (ppm/6 h/day)

Test animals (dams)

Litters with implants

Litters with live fetuses

 

No. dead/ no. treated

% of females pregnant at euthanization

No. of litters

No. of corpora lutes per dam

No. of implantation sites per litter

% of nonlive implants per littera

% of resorption sites per litter

No. of litters

No. of live

fetuses per

litter

% of males per litter

Average fetal body weight (g) per litter

All

Males

Females

 

0

0/26

92.3

24

15.63 ± 1.69

14.63 ±2.43

7.12 ± 12.12

6.52 ± 9.66

24

13.75 ± 3.01

53.40 ± 16.29

5.70±0.26

5.84 ± 0.29

5.54 ± 0.25

 

100

0/26

92.3

24

15.75 ± 1.45

14.71 ± 1.90

4.00 ± 6.80

4.00 ± 6.80

24

14.13 ± 2.11

51.50 ± 15.16

5.59± 0.25

5.71 ± 0.28

5.47 ± 0.27

 

300

0/26

96.2

25

16.12 ± 1.56

15.24 ±   1.59

4.05 ± 3.89

4.05 ± 3.89

25

14.60 ± 1.38

52.83 ± 12.70

5.53± 0.24

5.67 ± 0.26

5.38 ± 0.20

 

600

0/26

84.6

22

16.27 ± 1.52

15.18 ±2.54

5.28 ±8.08

5.28 ± 8.08

22

14.50 ± 3.02

46.59 ± 11.86

5.51± 0.33

5.72 ± 0.36

5.33 ± 0.35*

 

1200

0/27

92.6

25

16.76 ± 3.30

15.08 ± 2.00

6.27 ± 6.08

6.27 ± 6.08

25

14.12 ± 1.99

46.22 ± 15.16

5.40± 0.24**

5.56 ± 0.29**

5.26 ± 0.24**

 

aResorptions plus dead fetuses.

Values are expressed as means±SD.

*,** denote significant differences from the control (0 ppm) value,p , 0.05 and p , 0.01, respectively.

Applicant's summary and conclusion

Conclusions:
In a valid guideline study, n-BMA resulted in fetal toxicity, but no teratogenicity, at concentrations that also produced maternal toxicity.
Executive summary:

In a study comparable to the OECD guideline # 414, groups of 22-25 pregnant female rats were given whole-body inhalation exposures to n-BMA at target concentrations of 0, 100, 300, 600 or 1200 ppm (analytical concentrations: 0, 99.6 +/- 5.0, 301.6 +/- 12.2, 602.3 +/-38.0, 1206.4 +/- 46.9 ppm) for 6 hr/day, during days 6 to 20 of gestation (GD). Maternal toxicity (decreased body weight gain) was shown at 300 to 1200 ppm. Feed consumption was decreased at 1200 ppm. No dam died during the test and there were no adverse effects on the average number of implantations and live fetuses, incidence of non-live fetuses, or on resorptions. Fetal body weights of male pups were significantly reduced at 1200 ppm, and females at 600 and 1200 ppm n-BMA. There were no significant differences between control and treated groups for external, visceral, or skeletal malformations. A significant increase in the skeletal variations per litter occurred at 1200 ppm n-BMA, compared to controls. The authors concluded that NOAEL for developmental toxicity was 300 ppm n-BMA. There was no evidence of embryolethality or teratogenicity with n-BMA.