2-methoxyethyl methacrylate

Developmental toxicity / teratogenicity

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:

test procedure in accordance with national standard methods with acceptable restrictions

Cross-reference

Reason / purpose for cross-reference:

reference to same study

Reference

Endpoint:

multi-generation reproductive toxicity

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Principles of method if other than guideline:

NTP continuous breeding protocol
Reproductive assessment by continuous breeding design (RACB). Purpose of study was to develop RACB mouse protocol for use with rats using a known toxicant. Full method described in Chapin (1997). In a modification of the standard protocol, rats were cohabited for ~6 weeks, separated for delivery, nursing and weaning of the second litter for the F2 study, then cohabited for another ~9 weeks to produce a further 3 litters for other activities e.g. cross over mating. Cross over mating was used to determine the affected sex.

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Route of administration:

oral: drinking water

Details on mating procedure:

Males and females cohabited for approximately 6 weeks (a deviation from the standard protocol) to allow delivery, nursing and weaning of the 2nd litter, then re-co-habited for approximately nine more weeks.
M/F ratio: 1 to 1. F1 pairs cohabited for 1 week.
Length of cohabitation: 6 week period then 9 week period to produce 3 more litters.
Otherwise standard RACB protocol.

Duration of treatment / exposure:

Continuous from 1 week before mating through F1 generation.

Remarks:

0.01, 0.03, and 0.10% in drinking water (litter two)

Remarks:

0.006, 0.012, and 0.024% in drinking water (litter five)

No. of animals per sex per dose:

20 pairs per treatment group, 40 pairs of control animals

Control animals:

yes

Details on study design:

Dosing for 1 week prior to pairing. F1 pups weaned on postnatal day 21 and weighed weeks 21-23. 2nd generation from 2nd litter of F0 animals.

Parental animals: Observations and examinations:

CAGE SIDE OBSERVATIONS: Yes

DETAILED CLINICAL OBSERVATIONS: No data
- Time schedule:

BODY WEIGHT: Yes
- Time schedule for examinations: weeks 1-6, 10, 15, 18 and after every litter. F1 animals at weaning and weeks 31-33.

WATER CONSUMPTION AND COMPOUND INTAKE: Yes
- Time schedule for examinations: Calculated based on data from weeks when body weight and consumption measured on a per cage (per breeding pair) weekly average.

Oestrous cyclicity (parental animals):

Yes. Relative frequency of oestrus, proestrus, metestrus, diestrus, cycle length.

Sperm parameters (parental animals):

Parameters examined included motility, density, and percent abnormal. Sperm analysis is contained in a separate appended report to the main reference.

Litter observations:

-Drinking water consumption monitored week 21. Body weights. F2 pups monitored for 21 days (on days 0, 4, 7, 14, 21).
- standardization of litters: No
- parameters examined: litters per pair, live pups/litter, proportion of pups born alive, sex ration, live pup weight, (adjusted and unadjusted.)
- Gross examination of dead pups:
Postmortem examinations (parental animals)
- Sacrifice: Male animals: At the end of the crossover examination. Maternal animals: not examined.
- Gross necropsy: Yes
- histopathology/organ weights: liver, kidneys, right testis and epididymis and cauda epididymis, seminal vesicles and prostate, ovary. Histopathology only on males in the control and two lower dose groups as previous data indicated no effects likely on females. 10 randomly selected animals per dose group per generation examined. Livers only examined if gross effects seen.

Reproductive indices:

Number of litters and live pups per litter, proportion of live pups, sex ratio. Mating index.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Body weights were significantly reduced from week 3 in females and week 6 in males in the high dose group only.

Water consumption and compound intake (if drinking water study):

effects observed, treatment-related

Description (incidence and severity):

Water consumption declined from week 2 onwards in both sexes in the high dose group. It was significantly reduced in the mid dose group in week 6 only (4 other measurements not significantly different.)

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Description (incidence and severity):

Testicular and epididymal lesions seen, but also present in controls. Effects were not attributed to treatment.

Reproductive function: oestrous cycle:

no effects observed

Reproductive function: sperm measures:

effects observed, treatment-related

Description (incidence and severity):

Motility and density were decreased in the top dose group. Motility increased marginally but by a statistically significant margin in the mid dose group. All parameters were normal in the low dose group.

Reproductive performance:

effects observed, treatment-related

Description (incidence and severity):

100% fertility in controls and low dose group, 89% in mid dose group and only 5% (significantly lower – single breeding pair) in the top dose group. Days to litter were similar in the control and low dose group but increased in the mid dose group. (Too few in high dose group.). The number of litters per pair was not affected in the low and mid dose group. But the number of live pups per pair was significantly reduced in the mid dose group.

Dose descriptor:

NOAEL

Effect level:

0.01 other: % in drinking water; corresponding to 11 mg/kg bw/d

Based on:

act. ingr.

Sex:

male

Basis for effect level:

other: Adverse effects on sperm and male reproductive organs

Critical effects observed:

yes

Lowest effective dose / conc.:

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

System:

male reproductive system

Organ:

testes

Treatment related:

yes

Dose response relationship:

yes

Relevant for humans:

yes

Remarks on result:

not measured/tested

Critical effects observed:

not specified

Mortality / viability:

mortality observed, treatment-related

Description (incidence and severity):

Few pups born to high dose animals. Live pups/litter and proportion of live pups/litter decreased significantly in the mid dose group. The proportion of still births, the average postnatal survival were significantly decreased in the mid dose group.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Mean body weights of mid dose pups were significantly reduced for both sexes at all time points except one. Adjusted live pup weight increased significantly in the low dose group albeit by only around 5%. F1 mature animals showed reduced body weight (-14%) in the mid dose group.

Water consumption and compound intake (if drinking water study):

effects observed, treatment-related

Description (incidence and severity):

Drinking water consumption decreased in the mid dose group from week 21. The estimated daily dose was 9 and 27 mg/kg for the low and mid dose group male pups and 15/41 mg/kg respectively for females.

Sexual maturation:

not examined

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Description (incidence and severity):

There was no effect on adjusted organ weight of females. In males, liver, kidney, seminal vesicle, cauda epididymis, right epididymis and prostate all showed lower weights in the mid dose group after adjustment. The low dose group animals were similar to controls

Histopathological findings:

effects observed, treatment-related

Description (incidence and severity):

Only the testes and epididymis were examined. Minimal to mild testicular lesions were seen in 2/10, 7/10, 5/10 animals in the controls/mid/high dose animals respectively. These results suggested a slight effect from treatment but the no clear dose response was evident. Single animals in the mid and high dose animals showed epididymis lesions, which were not attributed to treatment.

Dose descriptor:

NOAEL

Generation:

F1

Effect level:

0.01 other: % in drinking water, corresponding to 11 mg/kg bw/d

Based on:

act. ingr.

Sex:

male/female

Basis for effect level:

other: number of live pups per litter and proportion of pups born alive decreased significantly

Critical effects observed:

not specified

There were no differences in epididymal sperm morphology or motility among the groups from the F2 generation but there was significantly decreased sperm density in both dose groups (-17% in low dose group, -23% in mid dose group).

Dose descriptor:

NOAEL

Generation:

F2

Effect level:

0.01 other: % in drinking water, corresponding to 11 mg/kg bw/d

Based on:

act. ingr.

Sex:

male/female

Basis for effect level:

other: Reduced number of live pups per litter

Critical effects observed:

not specified

Reproductive effects observed:

yes

Lowest effective dose / conc.:

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

Treatment related:

yes

Relation to other toxic effects:

reproductive effects in the absence of other toxic effects

Dose response relationship:

yes

Relevant for humans:

yes

Conclusions:

"Reproductive Assessment by Continuous Breeding" (RACB) protocol was originally designed using mice as the test species. The purpose of the present study was to develop a RACB protocol in CD Sprague-Dawley rats. Ethylene glycol monomethyl ether (EGMME), a known reproductive toxicant, was used as the test article.

Reproductive Toxicity of Ethylene Glycol Monomethyl Ether in Sprague-Dawley Rats, Litter Two:
Dose levels selected were 0.01, 0.03, and 0.10%, administered via drinking water. In a modification of the standard protocol, male and female rats ~20 pairs per treatment group, 40 pairs of control animals) were cohabited for approximately 6 weeks, separated to allow delivery, nursing and weaning of the second litter, then re- cohabited for approximately nine more weeks. The weaned second litter was used for F1 reproductive testing. The control and 0.03% F0 pairs were also utilized for a crossover mating trial to determine the affected sex.
Only one litter was born in the 0.10% dose group, and no pups were available for F1 testing. At 0.03% level of EGMME, number of live pups per litter and proportion of pups born alive decreased significantly, both in F0 and F1 testing. In the crossover mating, proportion of pups born alive decreased significantly in the 0.03% male X control female group. Declining control F0 fertility and productivity noted in this study indicate that the standard RACB design (weaning last rather than second litter) is better suited to use with rats.

Reproductive Toxicity of Ethylene Glycol Monomethyl Ether in Sprague-Dawley Rats, Litter Five:
Dose levels selected were 0.006, 0.012, and 0.024%, administered via drinking water. Male and female rats (20 pairs per treatment group, 40 pairs of control animals) were continuously exposed for a 7-day precohabitation period and 112-day cohabitation (Task 2).
While there was no decrease in average litter size with increased dose, the number of live male pups per litter and the total number of pups per litter were decreased in the 0.024% group. Both absolute and adjusted live pup weight were increased in all EGMME groups, but this was not dose-related. During the cross-over mating to determine the affected sex, there were fewer live male pups born to the 0.024% male X control female pairs. During the mating trial for the second generation, fewer male and total pups were delivered in the high-dose group, and both absolute and adjusted pup weight were increased in the middle and high dose groups. Low control fertility (63% fertile) during the cross-over mating is a cause for concern, however suggested protocol changes should alleviate some of this problem. Thus the RACB protocol can successfully be adjusted for use in rats.

Data source

Reference

Reference 1

Reference Type:

publication

Title:

Unnamed

Year:

1990

Reference 2

Reference Type:

publication

Title:

Unnamed

Year:

1997

Materials and methods

Principles of method if other than guideline:

NTP continuous breeding protocol
Reproductive assessment by continuous breeding design (RACB). Purpose of study was to develop RACB mouse protocol for use with rats using a known toxicant. Full method described in Chapin (1997). In a modification of the standard protocol, rats were cohabited for ~6 weeks, separated for delivery, nursing and weaning of the second litter for the F2 study, then cohabited for another ~9 weeks to produce a further 3 litters for other activities e.g. cross over mating. Cross over mating was used to determine the affected sex.

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

2-methoxyethanol

EC Number:

203-713-7

EC Name:

2-methoxyethanol

Cas Number:

109-86-4

Molecular formula:

C3H8O2

IUPAC Name:

2-methoxyethanol

Test animals

Species:

rat

Strain:

Sprague-Dawley

Administration / exposure

Route of administration:

oral: drinking water

Details on mating procedure:

Males and females cohabited for approximately 6 weeks (a deviation from the standard protocol) to allow delivery, nursing and weaning of the 2nd litter, then re-co-habited for approximately nine more weeks.
M/F ratio: 1 to 1. F1 pairs cohabited for 1 week.
Length of cohabitation: 6 week period then 9 week period to produce 3 more litters.
Otherwise standard RACB protocol.

Duration of treatment / exposure:

Continuous from 1 week before mating through F1 generation.

Doses / concentrations

Doses / concentrations 1

Remarks:

0.01, 0.03, and 0.10% in drinking water (litter two)

Doses / concentrations 2

Remarks:

0.006, 0.012, and 0.024% in drinking water (litter five)

No. of animals per sex per dose:

20 pairs per treatment group, 40 pairs of control animals

Control animals:

yes

Details on study design:

Dosing for 1 week prior to pairing. F1 pups weaned on postnatal day 21 and weighed weeks 21-23. 2nd generation from 2nd litter of F0 animals.

Examinations

Maternal examinations:

CAGE SIDE OBSERVATIONS: Yes

DETAILED CLINICAL OBSERVATIONS: No data
- Time schedule:

BODY WEIGHT: Yes
- Time schedule for examinations: weeks 1-6, 10, 15, 18 and after every litter. F1 animals at weaning and weeks 31-33.

WATER CONSUMPTION AND COMPOUND INTAKE: Yes
- Time schedule for examinations: Calculated based on data from weeks when body weight and consumption measured on a per cage (per breeding pair) weekly average.

Results and discussion

Results: maternal animals

Effect levels (maternal animals)

Remarks on result:

not measured/tested

Maternal abnormalities

Abnormalities:

not examined

Results (fetuses)

Effect levels (fetuses)

Dose descriptor:

NOAEL

Effect level:

0.01 other: % in drinking water, corresponding to 11 mg/kg bw/d

Based on:

act. ingr.

Sex:

male/female

Basis for effect level:

other: number of live pups per litter and proportion of pups born alive decreased significantly

Fetal abnormalities

Abnormalities:

not examined

Overall developmental toxicity

Developmental effects observed:

yes

Lowest effective dose / conc.:

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

Treatment related:

yes

Relation to maternal toxicity:

developmental effects in the absence of maternal toxicity effects

Dose response relationship:

yes

Relevant for humans:

yes

Applicant's summary and conclusion

Conclusions:

"Reproductive Assessment by Continuous Breeding" (RACB) protocol was originally designed using mice as the test species. The purpose of the present study was to develop a RACB protocol in CD Sprague-Dawley rats. Ethylene glycol monomethyl ether (EGMME), a known reproductive toxicant, was used as the test article.

Reproductive Toxicity of Ethylene Glycol Monomethyl Ether in Sprague-Dawley Rats, Litter Two:
Dose levels selected were 0.01, 0.03, and 0.10%, administered via drinking water. In a modification of the standard protocol, male and female rats ~20 pairs per treatment group, 40 pairs of control animals) were cohabited for approximately 6 weeks, separated to allow delivery, nursing and weaning of the second litter, then re- cohabited for approximately nine more weeks. The weaned second litter was used for F1 reproductive testing. The control and 0.03% F0 pairs were also utilized for a crossover mating trial to determine the affected sex.
Only one litter was born in the 0.10% dose group, and no pups were available for F1 testing. At 0.03% level of EGMME, number of live pups per litter and proportion of pups born alive decreased significantly, both in F0 and F1 testing. In the crossover mating, proportion of pups born alive decreased significantly in the 0.03% male X control female group. Declining control F0 fertility and productivity noted in this study indicate that the standard RACB design (weaning last rather than second litter) is better suited to use with rats.

Reproductive Toxicity of Ethylene Glycol Monomethyl Ether in Sprague-Dawley Rats, Litter Five:
Dose levels selected were 0.006, 0.012, and 0.024%, administered via drinking water. Male and female rats (20 pairs per treatment group, 40 pairs of control animals) were continuously exposed for a 7-day precohabitation period and 112-day cohabitation (Task 2).
While there was no decrease in average litter size with increased dose, the number of live male pups per litter and the total number of pups per litter were decreased in the 0.024% group. Both absolute and adjusted live pup weight were increased in all EGMME groups, but this was not dose-related. During the cross-over mating to determine the affected sex, there were fewer live male pups born to the 0.024% male X control female pairs. During the mating trial for the second generation, fewer male and total pups were delivered in the high-dose group, and both absolute and adjusted pup weight were increased in the middle and high dose groups. Low control fertility (63% fertile) during the cross-over mating is a cause for concern, however suggested protocol changes should alleviate some of this problem. Thus the RACB protocol can successfully be adjusted for use in rats.