2-methoxy-1-methylethyl acetate

Endpoint summary

Key value for chemical safety assessment

Effects on fertility

Link to relevant study records

Reference

Reference 1

Endpoint:

screening for reproductive / developmental toxicity

Remarks:

based on test type (migrated information)

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

1998

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Data published in peer reviewed literature. Full report is not available.

Qualifier:

according to guideline

Guideline:

OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)

GLP compliance:

yes

Limit test:

no

Species:

rat

Strain:

Crj: CD(SD)

Sex:

male/female

Route of administration:

oral: gavage

Vehicle:

water

Duration of treatment / exposure:

Exposure period: male: 44 days, female: from 14 days before mating to day 3 of lactation (41-45 days).
Premating exposure period (males): 14 days
Premating exposure period (females): 14 days
Duration of test: 41-45 days

Frequency of treatment:

daily

Remarks:

Doses / Concentrations:
0 mg/kg/day
Basis:
nominal conc.

Remarks:

Doses / Concentrations:
100 mg/kg/day
Basis:
nominal conc.

Remarks:

Doses / Concentrations:
300 mg/kg/day
Basis:
nominal conc.

Remarks:

Doses / Concentrations:
1000 mg/kg/day
Basis:
nominal conc.

Control animals:

yes, concurrent vehicle

Litter observations:

No effects related to the chemical exposure were observed in foetal data at 1000 mg/kg.

Overall, no effects related to chemical exposure were observed maternally at 1000 mg/kg. A dose of 1,000 mg/kg of PMA exerted some effects in both male and female rats. In males, depressions of body weight gain and a tendency for decrease in food consumption were observed. Blood chemical examination revealed decreases in glucose and inorganic phosphorus. An increase in relative weight of the adrenals was also noted. In females, body weight gain was lower than in the control during the premating period.
Although there was a single unsuccessful copulation at the 1000 mg/kg dose level, this was not statistically significantly different from the control. Reproductive toxicity of PGMA in rats by oral administration was not observed at the highest dose.
PMA did not exert any toxic effects on reproductive parameters including the copulation index, fertility index, gestation length, number of corpora lutea or implantations, implantation index, gestation index, delivery index and behaviour at delivery and lactation

Dose descriptor:

NOAEL

Effect level:

1 000 mg/kg bw/day

There were no differences between dosed groups and control group in offspring parameters including numbers of offspring or live offspring, sex ratio, live birth index, viability index and body weights. No external or visceral abnormalities related to PMA were detected in any of the offspring

Dose descriptor:

NOAEL

Generation:

F1

Effect level:

1 000 mg/kg bw/day

Reproductive effects observed:

not specified

Conclusions:

Reproductive toxicity of PGMA in rats by oral administration was not observed at the highest dose level of 1000 mg/kg and hence considered as NOAEL (No Observed Adverse Effect Level)

Executive summary:

Using OECD combined repeat dose and reproductive/developmental toxicity screening test [OECD TG 422], SD (Crj: CD), rats received gavage PGMA (purity > 99.9 %) doses of 0 (vehicle; distilled water), 100, 300 and 1,000 mg/kg/day, for males for 44 days from 2 weeks prior to mating and for females for 41-45 days from 14 days before mating to day 3 postpartum. The animals were sacrificed on the day 4 of lactation for females. No effects related to chemical exposure were observed maternally at 1,000 mg/kg, although there was a single unsuccessful copulation at this dose level which was not statistically significantly different from the control. Similarly, no effects related to the chemical exposure were observed in foetal data at 1,000 mg/kg. Reproductive toxicity of PGMA in rats by oral administration is not observed at the highest dose. A NOAEL was thus established at 1,000 mg/kg bw/day.

Reference 2

Endpoint:

two-generation reproductive toxicity

Remarks:

based on test type (migrated information)

Type of information:

read-across based on grouping of substances (category approach)

Adequacy of study:

key study

Study period:

06/1995-02/1997

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP-report according to OECD guideline 416

Justification for type of information:

Please refer to category document.

Qualifier:

according to guideline

Guideline:

OECD Guideline 416 (Two-Generation Reproduction Toxicity Study)

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Breeding Laboratory, Kingston, NY
- Age at study initiation: 6 weeks
- Weight at study initiation: (P) Males: g; Females: 120-121 g; (F1) Males: x-x g; Females: x-x g
- Fasting period before study: none
- Housing: singly in wire mesh stainless steel cages
- Use of restrainers for preventing ingestion (if dermal): yes/no
- Diet (e.g. ad libitum): ad libitum except during inhalation exposure
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 2 weeks

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22°C
- Humidity (%): 40-60%
- Air changes (per hr): 12-15
- Photoperiod (hrs dark / hrs light): 12/12

Route of administration:

inhalation: vapour

Type of inhalation exposure (if applicable):

whole body

Vehicle:

unchanged (no vehicle)

Details on exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus:14.5m3 (2.4 m wide x 2.4 m high x 2.4 m deep with pyramidal top)
- Method of conditioning air: the various concentrations of PGME were generated using a glass J-tube method. Liquid PGME was metered into a glass J-tube assembly through which a preheated stream of approximately 90 liters per minute of compressed air was passed to vaporize the test material. The compressed air was heated to the minimum extend necessary to facilitate complete vaporization of the test material (approximately 65°C for the 300 ppm chamber, 115°C for the 1000 ppm chamber and 150-170°C for the 3000 ppm chamber). The compressed air and PGME vapors were diluted and mixed with room temperature air to the desired concentration at a flow rate of 2900 liters per minute into whole-body inhalation chambers.
- Temperature, humidity, pressure in air chamber: 22°C, 40-60%, The chambers were operated at a slightly negative pressure relative to the surrounding area.
- Air flow rate: 2900 liters per min
- Air change rate: 12 changes per hour

TEST ATMOSPHERE
- Brief description of analytical method used:
- Samples taken from breathing zone: yes/no

Details on mating procedure:

Breeding of the P1 and P2 adults commenced after approximately 10 weeks of treatment. Each female was placed with a single male from the same exposure group (1:1 mating) until pregnancy occurred or two weeks had elapsed. During each breeding period, daily vaginal lavage samples were evaluated for the presence of sperm as an indication of mating. The day on which sperm were detected or a vaginal plug was observed in situ
was considered day 0 of gestation. Sperm and plug positive females were then removed from the male’s cage and placed back into wire mesh, stainless steel cages. If mating did not occur during the two weeks, the animals were separated without further opportunity for mating. For the P2 mating, cohabitation of male and female litter mates was avoided.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

The concentration of PGME in the breathing zone of the animals in each chamber was measured at least once per hour using a MIRAN 1A infrared spectrophotometer (Foxboro Analytical, Norwalk, CT) at a wavelength of 10.2 microns. The spectrophotometer was calibrated using standards having a known PGME vapor concentration contained in 90 liter SARANÒ or Tedlar film gas bags prior to the first exposure and approximately monthly thereafter. Daily checks of the spectrophotometer were performed prior to each exposure period using a single PGME standard concentration. In addition, the amount of PGME used each day was recorded and the nominal concentration (amount of PGME used / total chamber airflow) of PGME was calculated. Prior to the start of the study, each of the chambers to be used were checked to ensure that a uniform distribution of vapors occurred within the animal’s breathing zone.

Duration of treatment / exposure:

Exposure period: Before mating, through gestation, and post-birth.
Premating exposure period (males): 5 days/week prior to mating; 7 days/week post mating
Premating exposure period (females): 5 days/week prior to mating; 7 days/week post mating

Frequency of treatment:

6 hr/day

Details on study schedule:

Groups of 30 male and 30 female rats were exposed to 0, 300, 1000 or 3000 ppm PGME via inhalation, for 6 hours/day, 5 days/week prior to mating, and 6 hours/day, 7 days/week during mating, gestation and lactation. Treatment of the first generation parental (P1) rats began at approximately 6 weeks of age. After approximately 10 weeks of exposure (5 days/week, excluding holidays), P1 rats were mated (one male to one female of the respective treatment group) to produce the F1a litters. To aid in the interpretation of F1a litter weight data which indicated non dose-related decreased litter weights at 300 and 1000 ppm PGME (statistically identified for 300 ppm PGME day 14 females only) and to confirm significant findings noted for the 3000 ppm litters, the P1 adults were mated a second time to produce an F1b litter. Mating of the P1 adults for the second time
commenced approximately one week following weaning (3 weeks of age) of the last F1a litter. Initially, 30 males and 30 females from each treatment group were randomly selected from the F1a litters and assigned to treatment groups to become the second generation parents (P2). However, soon after exposure of the F1a weanlings commenced (1-3 days beginning on postnatal day 22) it became apparent that the weanlings in all dose groups were too small to be singly housed in wire mesh inhalation cages and to go without feed during the exposure period. Therefore, exposures were stopped. Weanlings in all dose groups appeared lethargic/weak following exposures and did not appear interested in feed or water. Given the limitations of starting the F1a litters at such a young age, and space limitations within the exposure chambers which precluded the simultaneous maintenance of the P1 and P2 generation animals, the decision was made to terminate the F1a weanlings and to choose the second generation parents from the F1b litters. Following weaning (3 weeks of age) of the F1b litters, 30 males and 30 females from each treatment group were randomly selected to become the P2 generation. To avoid the aforementioned problems with the F1a litters, exposure of the F1b weanlings/P2 adults began on their respective postnatal day 28. After approximately 10 weeks of treatment, the P2 adults were bred to produce the F2 litters. Exposures of P1 and P2 adults rats to PGME continued until the adults were sent to necropsy. All rats were housed continuously in exposure chambers following the initial exposure to PGME, except during late gestation and throughout the lactation period, when female rats were housed outside of the exposure chambers in nesting boxes. Maternal rats were not exposed to PGME after day 20 of gestation through the fourth day postpartum, in order to allow for parturition and initiation of lactation. During the lactation period, pups were not placed in the exposure chambers, but remained in the nesting boxes separated from the dam for
approximately 6 hours/day on lactation days 5 through 21.

Remarks:

Doses / Concentrations:
3000 ppm
Basis:
nominal conc.

Remarks:

Doses / Concentrations:
1000 ppm
Basis:
nominal conc.

Remarks:

Doses / Concentrations:
300 ppm
Basis:
nominal conc.

No. of animals per sex per dose:

30

Control animals:

yes, concurrent no treatment

Details on study design:

- Dose selection rationale: Rats were exposed to target concentrations of 0, 300, 1000 or 3000 ppm PGME. These concentrations corresponded to oral equivalent doses of approximately 0, 396, 1325 and 3974 mg/kg/day assuming ventilation rates of 1 l/min/kg and 100 percent absorption. The
calculated middle and high dose oral equivalents exceeded the 1 g/kg/day oral limit dose as defined by both the EPA (EPA, 1985) and OECD (OECD, 1981). The chosen concentrations were selected by the sponsor and based upon the results of the inhalation toxicity studies conducted previously.
- Rationale for animal assignment (if not random): random by body weight

Positive control:

none

Parental animals: Observations and examinations:

CAGE SIDE OBSERVATIONS: Yes
Each rat on study was observed twice daily (a.m. and p.m.) for mortality, morbidity and moribundity as well as availability of feed and water. In addition, changes in behavior or demeanor and indications of overt toxicity were evaluated during the a.m. or p.m. observation.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: A thorough clinical examination was conducted on all animals prior to the start of the study and weekly thereafter. This examination included evaluations of the skin and fur, mucous membranes, respiration, nervous system and behavior pattern. All adult rats found dead or in moribund condition were submitted for a gross pathologic examination. Adult rats found dead after normal working hours were refrigerated until a necropsy could be performed. All pups found dead, or pups that were euthanized in moribund condition, were examined to the extent possible for defects and/or cause of death and preserved in neutral, phosphate-buffered 10% formalin. Cannibalized pups were examined to the extent possible and discarded.

BODY WEIGHT: Yes
- Time schedule for examinations: All P1 animals had body weights recorded weekly during the 10-week pre-breeding treatment period, beginning on or before the first week of the study. Body weights for males were recorded weekly throughout the course of the study. Sperm and plug positive females were weighed on days 0, 7, 14 and 21 of gestation. Females that delivered litters were weighed on days 1, 4, 7, 14, and 21 of lactation. A similar schedule was followed for the P2 generation.

FOOD CONSUMPTION AND COMPOUND INTAKE: Feed consumption was not measured in this study

Oestrous cyclicity (parental animals):

Estrous cycle length and normality were evaluated daily by vaginal lavage (Cooper et al., 1993) for all P1 and P2 females starting three weeks prior to the F1a and F2 matings only, and continued throughout cohabitation.

Sperm parameters (parental animals):

At termination, samples of sperm from the right distal cauda epididymis from the first ten P1 and P2 males sacrificed in each exposure group were collected for evaluation of sperm motility. Sperm motility was determined with the use of the Hamilton-Thorn (HTM) Intergrated Visual Optical System (IVOS) motility analyzer (Hamilton-Thorn Research, Beverly, Massachusetts). Images of all samples for motility analyses were captured on an
optical disk and kept as raw data. The entire left cauda epididymis was weighed and then minced in saline to enumerate the total number of sperm (cauda reserves). Sperm counts were performed manually using a hemocytometer. Additionally, slides were prepared from sperm samples obtained from the left cauda epididymis for possible morphological evaluation and saved, but were not evaluated as no effects were noted on sperm counts or
motility.

Litter observations:

All litters were examined as soon as possible after delivery. The following parameters were recorded for each litter: total litter size on the day of parturition (day 0), the number of live and dead pups on days 0, 1, 4, 7, 14, and 21 postpartum, and the sex and the weight of each pup on days 1, 4, 7, 14, and 21 of lactation. Any visible physical abnormalities or demeanor changes in the neonates were recorded during the lactation period.

Postmortem examinations (parental animals):

A complete necropsy of all P1 and P2 adults was conducted by a team of trained individuals, including and directly supervised by a veterinary pathologist. The scheduled necropsy was performed after the last litter of the respective generation had been weaned, with the following exception: P1 adult males were necropsied early as it was determined that the age of the P1 adult population at the completion of the F1b lactation phase would have precluded any additional breeding of these adults. The rats were fasted overnight, anesthetized with methoxyflurane and euthanized by decapitation. The eyes were visually examined in situ through a moistened glass slide gently pressed against the cornea. Tissues routinely collected (Table 3) were saved from these rats and preserved in neutral, phosphate-buffered 10% formalin, with the following exceptions: testes and epididymides were preserved in Bouin's fixative. The lungs were infused with formalin to their approximate normal inspiratory volume. The nasal cavity was flushed with formalin via the pharyngeal duct to ensure rapid fixation of the tissue. Moribund rats and those dying spontaneously were necropsied in a similar manner. However, body and organ weights were not recorded.

The following organs of the first ten P1 and P2 parental animals sacrificed were weighed: ovaries or testes, left epididymis (total and cauda), seminal vesicles (with coagulating glands and their fluids), prostate, brain, liver, kidneys, lungs, adrenal glands, spleen, and thymus, and the organ-to-body weight ratios calculated.

Histologic examination of potential target organs and reproductive tissues was performed on the control and high concentration groups. Examination of tissues from the low and middle groups was limited to those tissues which demonstrated treatment-related histologic changes at the high concentration; those tissues were the liver and ovaries. Grading for atrophic ovaries was based primarily on the number of recognizable corpora lutea of any stage. In near optimal sections the grades were as follows: Very Slight, 16- 25 corpora lutea; Slight, 6-15; and Moderate, £5. If sections were less than complete, the presence of large cystic follicles and the density of corpora lutea were also considered. A complete set of histologic slides from all tissues listed in Table 3, was prepared from all rats that died spontaneously or were euthanized in a moribund condition, and were examined in an attempt to determine cause of death.

Postmortem examinations (offspring):

One pup/sex/exposure concentration from the first ten F1 and F2 litters to be weaned was given a complete necropsy by a team of trained individuals including and under the direct supervision of a veterinary pathologist. In order to control for variation in body and organ weight, pups selected for a complete necropsy were euthanized at the same age (postnatal day 22). Pups were anesthetized with methoxyflurane and euthanized by decapitation. Terminal body weights were recorded. Gross pathologic examination and preservation of tissue samples (Table 3) was performed as described above for adults.

For the F1 and F2 pups that were examined macroscopically (one/sex of the first ten litters/concentration weaned), the following organs were weighed: ovaries or testes, brain, heart, liver, kidneys, adrenal glands, spleen and thymus. Organ to body weight ratios were calculated.

Organs that demonstrated treatment-related effects (decreased absolute or relative weight) in weanlings chosen for necropsy were examined microscopically in the control and high concentration groups and included the liver, spleen, thymus and testes. These tissues were chosen for histologic examination because their absolute and/or relative organ weights were depressed (at least one statistically significantly, the other with the same downward difference) in one or more generations of high concentration weanlings. Examination of tissues from the low and middle groups was not conducted as the morphologic changes observed among the high concentration weanlings were clearly related to severe body weight depressions, and because these organ weight changes were not statistically significant at the lower exposure concentrations.

Statistics:

Body weights, gestation/lactation body weight gains, organ weights, sperm count per gram of cauda epididymis and percent motile sperm were first evaluated by Bartlett’s test for equality of variances. Based upon the outcome of Bartlett’s test, either a parametric or nonparametric analysis of variance (ANOVA) was performed. If the ANOVA was significant, a Dunnett’s test or the Wilcoxon Rank-Sum test with Bonferroni’s correction was performed. Gestation length, average time to mating, litter size, age at vaginal opening and age at preputial separation were analyzed using a nonparametric ANOVA. If the ANOVA was significant, the Wilcoxon Rank-Sum test with Bonferroni’s correction was performed. Statistical outliers were identified by the method of Grubbs (1969), but were only excluded from analysis for documented, scientifically sound reasons. Fertility indices were analyzed by the Fisher exact probability test and Bonferroni’s correction was used for multiple testing of groups in comparison to a single control. Evaluation of the neonatal sex ratio was performed by the binomial distribution test. Survival indices and other incidence data among neonates were analyzed using the litter as the experimental unit by the Wilcoxon test as modified by Haseman and Hoel (1974).

Reproductive indices:

see statistics

Offspring viability indices:

see statistics

Clinical signs:

effects observed, treatment-related

Dermal irritation (if dermal study):

not examined

Mortality:

mortality observed, non-treatment-related

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Mean body weight data for P1 males and females, including the F1a/F1b gestation and lactation periods (females only) and interim body weight data collected between the F1a and F1b breeding (females only), are presented in Tables 10 through 19. Body weights of 3000 ppm PGME males were significantly decreased relative to control values beginning on test day 7 and remained decreased throughout the study, with statistical significance achieved on days 7, 14, and 77-168. Body weights of males exposed to 300 or 1000 ppm PGME were not significantly different from controls at any time during the study. Pre-mating body weights of 3000 ppm PGME females were significantly decreased beginning on test day 7 and remained decreased throughout the pre-mating period. In fact, mean body weight in the high concentration group was approximately 10% lower than control values by the end of the pre-mating period. Body weights of high concentration dams remained significantly decreased throughout most of the F1a and F1b gestation periods, and the first one to two weeks of the F1a and F1b lactation periods. Body weight gains of 3000 ppm PGME females were not affected during the F1a or F1b gestation phases or the respective first week of lactation. Compensatory increases in body weight gains relative to controls were noted during the last two weeks of lactation (F1a and F1b) for the 3000 ppm PGME dams. Dams exposed to 1000 ppm PGME during the pre-mating phase exhibited slight decreases in body weight, with the decreases statistically identified only on test days 7, 14 and 63. No significant effects on body weight were observed among 300 ppm females during the pre-mating period. Additionally, no significant differences in F1a or F1b gestation or lactation body weights or body weight gains were noted for 300 or 1000 ppm females relative to controls.
Mean body weights of P2 males and females, including the F2 gestation and lactation period (females only) are presented in Tables 20 through 25. Body weights of 3000 ppm PGME males were significantly decreased (20%) relative to controls at the beginning of the P2 exposures and remained decreased throughout the study (9% decrease on day 70, the end of the pre-mating period). Body weights of the P2 males exposed to 300 or 1000 ppm PGME were not affected at any time during the study. Pre-mating body weights of 3000 ppm PGME females were significantly decreased (21%) at the beginning of the P2 exposures and remained decreased throughout the pre-mating period (16% decrease on day 70, the end of the pre-mating period). Body weights of high concentration dams were also significantly decreased throughout the F2 gestation period, and the first one to two weeks of the F2 lactation period. Body weight gains of 3000 ppm PGME females were significantly decreased on gestation days 14-21, resulting in a statistically identified overall decrease on gestation days 0-21 as well. Compensatory increases in body weight gains relative to controls were statistically identified during lactation days 7-14 and 14-21, resulting in an overall increase in body weight gain for the entire lactation period (days 1-21) for the 3000 ppm PGME dams. Dams exposed to 1000 ppm PGME during the pre-mating period exhibited slight decreases in body weight, relative to control values, with the decreases statistically identified only on test days 63 and 70. No significant effects on body weight were observed among 300 ppm PGME females during the pre-mating period. Additionally, no significant differences in P2/F2 gestation or lactation body weights or body weight gains were noted for 300 or 1000 ppm PGME females relative to controls.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Food efficiency:

not specified

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

not specified

Clinical biochemistry findings:

not examined

Urinalysis findings:

not specified

Behaviour (functional findings):

not specified

Immunological findings:

not specified

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

no effects observed

Other effects:

not examined

Reproductive function: oestrous cycle:

no effects observed

Reproductive function: sperm measures:

no effects observed

Reproductive performance:

no effects observed

CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS)
At 3000 ppm PGME, sedation as evidenced by incoordination and decreased activity which resolved by the next exposure day was observed in both
sexes immediately following the first exposure to PGME and continued to be observed through test day 32 in males, and test day 23 in females. Sedation was not observed in males or females exposed to 300 or 1000 ppm PGME at any time during the study. No other treatment-related observations regarding behavior or demeanor were observed in P1 adults males at any exposure concentration during the entire study or in P1 adult females at any exposure concentration during the pre-mating, gestation or lactation periods. Six P1 adult females were found dead prior to the scheduled necropsy. Cause of death and other relevant information is summarized in Text Table 1. Detailed gross and histopathologic observations for these animals may be found in the individual animal pathology data, starting with Appendix Table 58. The deaths of these rats were not considered treatment-related. All other rats survived to the scheduled termination.
P2 adults and F2 litters: A single 1000 ppm male, 95B0860, died early on test day 72. No prior clinical observations were made on this male. The only gross observation made for this male was congested viscera. Additionally, inflammation of the heart and atrophy of the pancreas was noted during histopathologic examination. The cause of death for this male was not determined. All other P2 adults survived the test period and were necropsied on
the scheduled date of termination. Similar to the P1 adults, P2 male and female rats exposed to 3000 ppm PGME exhibited sedation, initially evidenced as incoordination following their first exposure to PGME (postnatal day 28). Incoordination was resolved within approximately one week of initial exposure to PGME vapors followed by approximately one week of decreased activity observed immediately following daily exposures. Incoordination and sedation observed during and post-exposure resolved by the next exposure day during this interval. Sedation was not observed in males or females
exposed to 300 or 1000 ppm PGME at any time during the study. The only other treatment-related observation noted was vaginal bleeding, which was noted among five high dose females late in the study (gestation/lactation phase). In three of the five cases the vaginal bleeding was associated with the pending delivery of a litter. In the other two cases the bleeding stopped, however, no litters were delivered. Of these two females, only one had histopathologic indications of a prior pregnancy (pigment laden macrophages in the uterus). No other treatment-related observations regarding behavior or demeanor were observed in P2 adults males at any exposure concentration during the entire study or in P2 adult females at any exposure concentration during the pre-mating, gestation or lactation periods.

BODY WEIGHT AND FOOD CONSUMPTION (PARENTAL ANIMALS)
Mean body weight data for P1 males and females, including the F1a/F1b gestation and lactation periods (females only) and interim body weight data collected between the F1a and F1b breeding (females only), are presented in Tables 10 through 19. Body weights of 3000 ppm PGME males were significantly decreased relative to control values beginning on test day 7 and remained decreased throughout the study, with statistical significance achieved on days 7, 14, and 77-168. Body weights of males exposed to 300 or 1000 ppm PGME were not significantly different from controls at any time during the study. Pre-mating body weights of 3000 ppm PGME females were significantly decreased beginning on test day 7 and remained decreased throughout the pre-mating period. In fact, mean body weight in the high concentration group was approximately 10% lower than control values by the end of the pre-mating period. Body weights of high concentration dams remained significantly decreased throughout most of the F1a and F1b gestation periods, and the first one to two weeks of the F1a and F1b lactation periods. Body weight gains of 3000 ppm PGME females were not affected during the F1a or F1b gestation phases or the respective first week of lactation. Compensatory increases in body weight gains relative to controls were noted during the last two weeks of lactation (F1a and F1b) for the 3000 ppm PGME dams. Dams exposed to 1000 ppm PGME during the pre-mating phase exhibited slight decreases in body weight, with the decreases statistically identified only on test days 7, 14 and 63. No significant effects on body weight were observed among 300 ppm females during the pre-mating period. Additionally, no significant differences in F1a or F1b gestation or lactation body weights or body weight gains were noted for 300 or 1000 ppm females relative to controls.
Mean body weights of P2 males and females, including the F2 gestation and lactation period (females only) are presented in Tables 20 through 25. Body weights of 3000 ppm PGME males were significantly decreased (20%) relative to controls at the beginning of the P2 exposures and remained decreased throughout the study (9% decrease on day 70, the end of the pre-mating period). Body weights of the P2 males exposed to 300 or 1000 ppm PGME were not affected at any time during the study. Pre-mating body weights of 3000 ppm PGME females were significantly decreased (21%) at the beginning of the P2 exposures and remained decreased throughout the pre-mating period (16% decrease on day 70, the end of the pre-mating period). Body weights of high concentration dams were also significantly decreased throughout the F2 gestation period, and the first one to two weeks of the F2 lactation period. Body weight gains of 3000 ppm PGME females were significantly decreased on gestation days 14-21, resulting in a statistically identified overall decrease on gestation days 0-21 as well. Compensatory increases in body weight gains relative to controls were statistically identified during lactation days 7-14 and 14-21, resulting in an overall increase in body weight gain for the entire lactation period (days 1-21) for the 3000 ppm PGME dams. Dams exposed to 1000 ppm PGME during the pre-mating period exhibited slight decreases in body weight, relative to control values, with the decreases statistically identified only on test days 63 and 70. No significant effects on body weight were observed among 300 ppm PGME females during the pre-mating period. Additionally, no significant differences in P2/F2 gestation or lactation body weights or body weight gains were noted for 300 or 1000 ppm PGME females relative to controls.

REPRODUCTIVE FUNCTION: ESTROUS CYCLE (PARENTAL ANIMALS)
Estrous cyclicity data collected for ten P1 females per exposure concentration are presented in Table 26. Although not statistically identified, dams exposed to 3000 ppm PGME had a slight increase in the mean number of days per cycle as well as a resultant decrease in the mean number of estrous cycles observed per dam prior to placement with males for the F1a mating. Additionally, cycling slides obtained for four out of ten high exposure females were noted as exhibiting atypical cellularity. No effects on mean number of days per cycle or number of cycles per dam were noted for 300 or 1000 ppm dams. Estrous cycle length and normality were not evaluated prior to the F1b mating.
P2 estrous cyclicity data collected for ten P2 females per exposure concentration are summarized in Table 27. Dams exposed to 3000 ppm PGME had a statistically significant increase in the mean number of days per cycle as well as a statistically significant decrease in the mean number of estrous cycles observed per dam prior to placement with males for the F2 mating. This was consistent with the slight trend for increased cycle length and decreased number of cycles noted in P1 females for the F1a mating. Also, as for the P1 females, atypical cellularity was noted among the cycling slides for four high exposure females. No effects on mean number of days per cycle or number of cycles per dam were noted for 300 or 1000 ppm PGME dams.

REPRODUCTIVE FUNCTION: SPERM MEASURES (PARENTAL ANIMALS)
Summaries of mean sperm counts and sperm motility (percent motile and progressively motile sperm) observed for ten P1 males per exposure concentration: No treatment-related differences in sperm counts or motility were observed for any exposure concentration tested. A significant increase in the number of progressively motile sperm obtained from P1 3000 ppm PGME males was considered spurious as a similar effect was not observed in the subsequent P2 generation.
Summaries of mean sperm counts and sperm motility (percent motile and progressively motile sperm) observed for ten P2 males per exposure concentration: No treatment-related differences in sperm counts or motility were observed for any exposure concentration tested.

REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS)
No treatment-related effects were observed on male or female P1 adult reproductive indices, gestation survival index, pup sex ratios, gestation length, or time to mating at any exposure concentration for the F1a mating/litters. Survival of pups from 3000 ppm PGME litters was significantly decreased relative to controls on lactation days 1 and 4. Survival of 3000 ppm PGME pups was also decreased, although not statistically identified, on lactation days 14 and 21. No effects on survival were observed for 300 or 1000 ppm PGME F1a litters at any time.
Reproductive indices and pup survival for P2 adults and their F2 litters: At 3000 ppm PGME, P2 male and female fertility was lower (not statistically
identified) than controls and recent historical control ranges (Table 31), while male and female conception rates were identified as significantly lower than controls, also falling outside the range of recent historical control values. No effects were observed on the P2 male or female mating index, gestation survival, pup sex ratios, gestation length, or time to mating. However, survival of 3000 ppm PGME F2 pups was significantly decreased relative to controls on lactation days 1 and 4. No treatment-related effects were observed on P2/F2 male or female reproductive indices, gestation survival, pup survival indices, pup sex ratios, gestation length, or time to mating at 300 or 1000 ppm PGME.

ORGAN WEIGHTS (PARENTAL ANIMALS)
Organ weight data (absolute and relative) for P1 males and females: Males exposed to 3000 ppm PGME exhibited statistically significant increases in relative testes, brain and kidney weights as well as a decrease in absolute and relative thymus weight. These effects were considered likely secondary to the significant decrease in mean P1 male body weight at this exposure concentration (probable decreased feed intake and resultant nutritional stress). No other significant effects on any other organ weights were observed among P1 3000 ppm PGME males. There were no treatment-related effects on terminal body weights or organ weights of males exposed to 300 or 1000 ppm PGME. P1 females exposed to 3000 ppm PGME had significantly higher relative adrenal, liver, and lung weights, when compared to controls, also primarily because of significantly lower body weights, however the interpretation of the liver weights is more complicated and will be discussed. No significant differences were observed for organ weights of P1 300 or 1000 ppm PGME females relative to controls.
Organ weight data (absolute and relative) for P2 males and females: P2 males exposed to 3000 ppm PGME exhibited statistically significant increases in relative liver, lung and seminal vesicle weights. Trends were similar, but not significant in P1 3000 ppm males, and again are interpreted primarily as the result of decreased body weight. No other significant effects on any other organ weights were observed among P2 3000 ppm PGME males. There were no treatmentrelated differences in organ weights of P2 males exposed to 300 or 1000 ppm PGME. P2 females exposed to 3000 ppm PGME had significantly decreased brain (absolute) and ovary (absolute and relative) weights relative to controls. The brain weights of the highexposure
P2 females will be discussed in the context of the entire study. The decreased ovary weights noted at 3000 ppm PGME appeared consistent with an increased incidence of ovarian atrophy observed histologically at this concentration. No other significant differences in any other organ weights were noted for 3000 ppm P2 females relative to controls. There were no treatment-related differences in organ weights of P2 females exposed to 300 or 1000 ppm PGME.

GROSS PATHOLOGY (PARENTAL ANIMALS)
No treatment-related gross pathologic changes were observed among the P1 and P2 adults at any exposure concentration.

HISTOPATHOLOGY (PARENTAL ANIMALS)
An increased incidence of histologic ovarian atrophy and decreased ovarian weight was observed among the 3000 ppm PGME P1 females and was interpreted to be the result of non-specific toxicity and nutritional stress (see Discussion). Histologically, typical atrophic ovaries had fewer, or no, corpora lutea, and multiple large cystic and atretic follicles. There was no apparent increase in follicular atresia (identified by apoptotic granulosa cells) among developing follicles. Primordial and all subsequent stages in follicular maturation were evaluated qualitatively; they appeared to be present in normal numbers and were of normal appearance.
Similar to the P1 adults, an increased incidence of histologic ovarian atrophy associated with decreased ovarian weight was observed among the P2 3000 ppm PGME females and likewise was interpreted to be the result of non-specific toxicity and nutritional stress (see Discussion). Similar to the P1 females, typical P2 atrophic ovaries had fewer, or no, corpora lutea, and multiple large cystic and atretic follicles and there was no apparent increase in follicular atresia (identified by apoptotic granulosa cells) among developing follicles. Primordial cells and all subsequent cell stages in follicular maturation appeared to be present in normal numbers and were of normal appearance.

Key result

Dose descriptor:

NOAEL

Effect level:

300 ppm

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: sedation as evidenced by incoordination and decreased activity which resolved by the next exposure day was observed in both sexes immediately following the first exposure to PGME

Clinical signs:

no effects observed

Dermal irritation (if dermal study):

not examined

Mortality / viability:

no mortality observed

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Significant body weight decreases observed among the 3000 ppm PGME weanlings relative to controls

Food consumption and compound intake (if feeding study):

not specified

Food efficiency:

not specified

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

not specified

Ophthalmological findings:

not examined

Haematological findings:

not specified

Clinical biochemistry findings:

not specified

Urinalysis findings:

not examined

Sexual maturation:

no effects observed

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Description (incidence and severity):

Mean absolute and relative organ weights and terminal body weights were collected for ten males and females per exposure concentration (F1a and F1b). Several organ weights changes (absolute or relative) were noted for male and female weanlings from F1a 3000 ppm PGME litters. F1a 3000 ppm PGME males had statistically identified decreases in absolute kidney, liver, spleen, and testes weights, while high concentration F1a females exhibited decreased absolute heart, and thymus weights and increased relative brain and kidneys weights. All of the differences identified at 3000 ppm PGME apeared to be secondary to significant decreases in male and female pup body weights at the time of necropsy/weaning (see Table 34) and were entirely consistent with nutritional stress. No significant organ weight changes were noted for F1a 300 or 1000 ppm PGME male or female weanlings. High concentration F1b male and female weanlings had significantly decreased absolute brain weights (males and females) and increased relative heart weights (males only). As with the F1a weanlings, these changes were likely due to the significant body weight decreases observed among the 3000 ppm PGME weanlings relative to controls. F1b males from 1000 ppm PGME litters were noted as having statistically identified decreases in absolute brain weight. This change was not considered toxicologically significant as relative brain weight was not affected and similar decreases were not observed among the F1a or F2 3000 ppm PGME males. A statistically significant increase in relative heart weight in 300 ppm PGME F1b males and females was considered spurious and unrelated to treatment as no dose-response was observed and similar increases were not observed in the F1a or F2 weanlings. Similarly, a significant increase in relative adrenal weight observed for 300 ppm F1b females was not consistent with data generated for the F1a or F2 generations, nor was a dose response observed. No significant organ weight changes were identified for 1000 F1b ppm PGME females.
F2 males at 3000 ppm PGME exhibited significant weight changes (absolute and/or relative) for all organs weighed and included the following: adrenals, brain, heart, kidneys, liver, spleen, testes, and thymus. F2 3000 ppm PGME females exhibited similar changes, however, only absolute brain and spleen weights were statistically identified. As with the 3000 ppm PGME F1a and F1b weanlings, organ weight changes noted for the F2 3000 ppm PGME males and females were likely secondary to significant body weight decrements (see Table 37) relative to control values and nutritional stress. A statistically significant decrease in relative thymus weight was identified for F2 300 ppm PGME males. As this effect was not dose-related and was not observed among the F1a or F1b 300 or 1000 ppm PGME males, it was considered a spurious finding. No other significant organ weight changes were noted for 300 or 1000 ppm PGME F2 males or females.

Gross pathological findings:

no effects observed

Histopathological findings:

effects observed, treatment-related

Description (incidence and severity):

Histologically, the livers of F1a and F1b 3000 ppm PGME weanlings often lacked the normal degree of glycogen vacuolation; glycogen depletion was diagnosed and recorded. This finding, along with an increase in thymic single cell necrosis, is consistent with nutritional stress. Although spleen weights were in general depressed, no particular cell compartment was identified as contributing disproportionately, and no necrosis was observed, so no histopathologic correlate was recorded. Similarly, testicular weights for the F1a and F1b 3000 ppm PGME males were decreased, however, they were also histologically normal for their slightly earlier stage of development. Also, it should be recalled that the high-exposure P2 males, selected from the F1b pups, demonstrated normal testicular weights and histopathology, as well as sperm count and motility.
Changes similar to those observed in the F1a and F1b weanlings were noted histologically in the livers and thymus of F2 3000 ppm PGME weanlings suggesting some degree of nutritional stress. Despite weight changes observed, no histopathologic observations were noted in the spleens of 3000 ppm PGME F2 males or females. The testes of 3000 ppm PGME F2 males were also histologically normal, despite decreases noted in weight relative to controls.

Other effects:

not examined

Behaviour (functional findings):

not examined

Developmental immunotoxicity:

not specified

VIABILITY (OFFSPRING)
No treatment-related effect observed.

CLINICAL SIGNS (OFFSPRING)
No treatment-related effect observed.

BODY WEIGHT (OFFSPRING)
Significant body weight decreases observed among the 3000 ppm PGME weanlings relative to controls.

SEXUAL MATURATION (OFFSPRING)
No treatment-related effect observed.

ORGAN WEIGHTS (OFFSPRING)
Mean absolute and relative organ weights and terminal body weights were collected for ten males and females per exposure concentration (F1a and F1b). Several organ weights changes (absolute or relative) were noted for male and female weanlings from F1a 3000 ppm PGME litters. F1a 3000 ppm PGME males had statistically identified decreases in absolute kidney, liver, spleen, and testes weights, while high concentration F1a females exhibited decreased absolute heart, and thymus weights and increased relative brain and kidneys weights. All of the differences identified at 3000 ppm PGME apeared to be secondary to significant decreases in male and female pup body weights at the time of necropsy/weaning (see Table 34) and were entirely consistent with nutritional stress. No significant organ weight changes were noted for F1a 300 or 1000 ppm PGME male or female weanlings. High concentration F1b male and female weanlings had significantly decreased absolute brain weights (males and females) and increased relative heart weights (males only). As with the F1a weanlings, these changes were likely due to the significant body weight decreases observed among the 3000 ppm PGME weanlings relative to controls. F1b males from 1000 ppm PGME litters were noted as having statistically identified decreases in absolute brain weight. This change was not considered toxicologically significant as relative brain weight was not affected and similar decreases were not observed among the F1a or F2 3000 ppm PGME males. A statistically significant increase in relative heart weight in 300 ppm PGME F1b males and females was considered spurious and unrelated to treatment as no dose-response was observed and similar increases were not observed in the F1a or F2 weanlings. Similarly, a significant increase in relative adrenal weight observed for 300 ppm F1b females was not consistent with data generated for the F1a or F2 generations, nor was a dose response observed. No significant organ weight changes were identified for 1000 F1b ppm PGME females.
F2 males at 3000 ppm PGME exhibited significant weight changes (absolute and/or relative) for all organs weighed and included the following: adrenals, brain, heart, kidneys, liver, spleen, testes, and thymus. F2 3000 ppm PGME females exhibited similar changes, however, only absolute brain and spleen weights were statistically identified. As with the 3000 ppm PGME F1a and F1b weanlings, organ weight changes noted for the F2 3000 ppm PGME males and females were likely secondary to significant body weight decrements (see Table 37) relative to control values and nutritional stress. A statistically significant decrease in relative thymus weight was identified for F2 300 ppm PGME males. As this effect was not dose-related and was not observed among the F1a or F1b 300 or 1000 ppm PGME males, it was considered a spurious finding. No other significant organ weight changes were noted for 300 or 1000 ppm PGME F2 males or females.

GROSS PATHOLOGY (OFFSPRING)
There were no gross lesions identified at the F1a or F1b necropsy that were associated with PGME exposure. There were no treatmentrelated gross lesions identified in the F2 males or females at any exposure concentration.

HISTOPATHOLOGY (OFFSPRING)
Histologically, the livers of F1a and F1b 3000 ppm PGME weanlings often lacked the normal degree of glycogen vacuolation; glycogen depletion was diagnosed and recorded. This finding, along with an increase in thymic single cell necrosis, is consistent with nutritional stress. Although spleen weights were in general depressed, no particular cell compartment was identified as contributing disproportionately, and no necrosis was observed, so no histopathologic correlate was recorded. Similarly, testicular weights for the F1a and F1b 3000 ppm PGME males were decreased, however, they were also histologically normal for their slightly earlier stage of development. Also, it should be recalled that the high-exposure P2 males, selected from the F1b pups, demonstrated normal testicular weights and histopathology, as well as sperm count and motility.
Changes similar to those observed in the F1a and F1b weanlings were noted histologically in the livers and thymus of F2 3000 ppm PGME weanlings suggesting some degree of nutritional stress. Despite weight changes observed, no histopathologic observations were noted in the spleens of 3000 ppm PGME F2 males or females. The testes of 3000 ppm PGME F2 males were also histologically normal, despite decreases noted in weight relative to controls.

Key result

Dose descriptor:

NOAEL

Generation:

F1

Effect level:

1 000 ppm

Sex:

male/female

Basis for effect level:

other: Highest concentration tested (3000ppm: significant body weight decreases; several organ weights changes

Key result

Dose descriptor:

NOAEL

Generation:

F2

Effect level:

1 000 ppm

Sex:

male/female

Basis for effect level:

other: highest concentration tested (3000ppm: significant body weight decreases; several organ weights changes

Reproductive effects observed:

not specified

At 3000 ppm, toxicity in the P1 and P2 adults was marked, as
evidenced by sedation during and after exposure for several
weeks, and mean body weights which were as much as 21% lower
than controls. This marked parental toxicity was accompanied
by lengthened estrous cycles, decreased fertility, decreased
ovary weights, reduced pup survival and litter size, slight
delays in puberty onset, and histologic changes in the liver
and thymus of the F1 and F2 offspring.  At 3000 ppm, there
was an increase in histologic ovarian atrophy in P1 and P2
females, and at 1000 ppm, there was a decrease in pre-mating
body weight in the P1 and P2 females. No treatment-related
differences in sperm counts or motility were observed among
the P1 or P2 males.

Conclusions:

The NOAEL for paternal toxicity is 300 ppm and for offspring toxicity is 1000 ppm. Effects appear secondary to parental weight loss.

Executive summary:

The objective of this two-generation inhalation reproduction study was to evaluate the effects of propylene glycol monomethyl ether (PGME) on the reproductive capability and neonatal growth and survival of rats. Groups of 30 male and 30 female Sprague-Dawley rats, approximately 6 weeks of age, were exposed to 0, 300, 1000 or 3000 ppm PGME via inhalation, for 6 hours/day, 5 days/week prior to mating and 6 hours/day, 7 days/week during mating, gestation and lactation for two generations. Inhalation exposure of adult male and female rats to 1000 (females only) and 3000 (males and females) ppm PGME resulted in dose-related parental effects. Toxicity in 3000 ppm PGME P1 and P2 males and females was evidenced primarily as an increased incidence of sedation for several weeks early in the exposure regimen and significant decreases in body weights, which achieved decrements of as much as 20 and 21% relative to controls, respectively. Decreased body weights in the P1 and P2 high concentration females generally persisted throughout the pre-breeding, gestation and lactation phases of the study. Additional effects noted among P1 and P2 adult females exposed to 3000 ppm PGME included lengthened estrous cycles, decreased fertility, decreased ovary weights and an increased incidence of histologic ovarian atrophy. The effects on fertility, estrous cyclicity and ovarian weight/histology appeared to be interrelated and associated with the significant decreases in 3000 ppm PGME female body weights and general toxicity/nutritional stress throughout the test period. No treatment-related differences in sperm counts or motility were observed among P1 or P2 adult males. Neonatal effects observed at 3000 ppm PGME consisted of decreased pup body weights, reduced pup survival and litter size, increased time to vaginal opening or preputial separation, and histopathologic observations in the liver and thymus of weanling rats. These neonatal effects also were considered secondary to maternal toxicity, particularly with respect to the compromised nutritional status of the maternal animals of the 3000 ppm PGME group. In the 1000 ppm PGME group, mild parental toxicity was evidenced by slightly decreased pre-mating body weights among P1 and P2 females, but was not accompanied by any statistically significant effects on parental reproduction or neonatal survival, growth or development. There were no treatment-related parental or neonatal effects related to exposure of rats to 300 ppm PGME. In conclusion, the no-observed-effect-level (NOEL) for fertility and reproductive effects in this two-generation inhalation reproduction study was 1000 ppm PGME.

Effect on fertility: via oral route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

1 000 mg/kg bw/day

Study duration:

subacute

Species:

rat

Quality of whole database:

Good (Klimisch 2)

Effect on fertility: via inhalation route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEC

5 400 mg/m³

Study duration:

subchronic

Species:

rat

Quality of whole database:

Good

Effect on fertility: via dermal route

Endpoint conclusion:

no study available

Additional information

Several studies confirm that rapid and extensive hydrolysis of propylene glycol methyl ether acetate (PGMA) to propylene glycol methyl ether (PGME) occurs in vivo when PGMA is administered by the oral, inhalatory or dermal route. Since urinary metabolites and disposition profiles of PGMA were approximately identical to the results obtained with PGME, it is unlikely that there are substantial differences of the systemic toxicity between PGMA and PGME. In fact, toxicity of PGMA is almost the same of PGME. Therefore, data on PGME were also used to support the dataset for PGMA.

The no-observed-effect-level (NOEL) for fertility and reproductive effects in the two-generation inhalation reproduction study on PGME was 1000 ppm. The NOAEL for paternal toxicity is 300 ppm and for offspring toxicity is 1000 ppm. Effects appear secondary to parental weight loss. This is supported by the results of a combined repeated dose and reproductive toxicity study (OECD 422) conducted with PGMA. No reproductive toxicity was observed up to the limit dose of 1000 mg/kg bw/day (administered by oral gavage) in this study. A NOAEC of 1000 ppm (5400 mg/m3) can be taken into account for PGMA. See attached category document in Section 13 for read-across justification.

Short description of key information:
No specific 2-generation reprotox studies is available for propylene glycol methyl ether acetate. A 2-generation reproductive inhalation toxicity study in rats, conducted according to GLP and OECD guideline, is available for propylene glycol methyl ether, which is used as read-across. This study is supported by an oral gavage study according to OECD guideline 422 on propylene glycol methyl ether acetate. See attached category document in Section 13 for read-across justification.

Justification for selection of Effect on fertility via oral route:
There is only study available via the oral route

Justification for selection of Effect on fertility via inhalation route:
GLP report according to OECD guideline 415

Justification for selection of Effect on fertility via dermal route:
There are studies available for the oral and the inhalation route

Effects on developmental toxicity

Description of key information

One inhalation teratology study in rats conducted using Propylene Glycol Methyl Ether acetate 
Two inhalation teratology studies (in rats and rabbits), conducted under GLP and equivalent to OECD guideline 414, are available for propylene glycol methyl ether.

Link to relevant study records

Reference

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1989

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP-study.

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

Deviations:

yes

Remarks:

dosing from GD 6-15 only - compared to current guideline with dosing from GD6-20

GLP compliance:

yes

Limit test:

no

Species:

rat

Strain:

Sprague-Dawley

Route of administration:

inhalation

Type of inhalation exposure (if applicable):

whole body

Vehicle:

unchanged (no vehicle)

Details on exposure:

From Days 6 through 15 for 6 hours per day, pregnant rats were exposed either to PM Acetate vapor or room air. Vapors were generated by piping hot air through PM Acetate coated glass beads thus evaporating the liquid (reference 9). Chamber concentrations were measured three times a day at Hours 1, 3 and 5 by pumping chamber air to a MIRAN 80 Infrared Analyzer.

Analytical verification of doses or concentrations:

yes

Duration of treatment / exposure:

days 6-15 of gestation

Frequency of treatment:

6 hours/day

Duration of test:

21 days

No. of animals per sex per dose:

Main study: 23 pregnant rats in dose groups 0, 500, 2000 ppm; 20 pregnant rats in dose group 4000 ppm
(due to variable fertility)

Control animals:

yes, concurrent vehicle

Details on study design:

Pregnant SD female rats were exposed to PGMA (purity: 97.3% of 2-methoxy-1-methylethyl acetate and 2.0 % of 1- methoxy-2- ) methylethyl acetate) vapour from Days 6 through 15 of gestation, once daily for 6 hours/day at nominal dose of 0, 500, 2,000, 4,000 ppm (0, 2700, 10800, 21600 mg/m3). The animals were sacrificed on Day 20 of gestation to evaluate the potential maternal, embryonic and teratogenic parameters of PGMA

Maternal examinations:

Yes

Ovaries and uterine content:

Yes

Fetal examinations:

Yes

Statistics:

Yes

Indices:

Yes

Details on maternal toxic effects:

Maternal toxic effects:yes

Details on maternal toxic effects:
Most of the effects observed in dams were transient in nature. Reductions in muscle tone (2000 and 4000 ppm), food consumption (500, 2000 and 4000 ppm) and body weight (2000 and 4000) were seen during the exposure period. At 2000 and 4000 ppm exposure groups, dyspnea, ruffled pelt and red discharges from the eyes or mouth were observed. No toxic signs were observed in the 500 ppm exposure group. The effect on nasal cavity was not examined in this experiment.

Dose descriptor:

NOAEL

Effect level:

500 ppm

Basis for effect level:

other: maternal toxicity

Dose descriptor:

LOAEL

Effect level:

2 000 ppm

Basis for effect level:

other: maternal toxicity

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
No developmental toxicity was observed at any of the doses tested

Dose descriptor:

NOAEL

Effect level:

> 4 000 ppm

Basis for effect level:

other: teratogenicity

Abnormalities:

not specified

Developmental effects observed:

not specified

Pilot Study.

1.Ataxia was observed intermittently in several rats only in the 3000 ppm exposure group during exposure, but subsided soon after the rats were returned to their boxes.

2. Food consumption by dams as a percentage of body weight was lower in the 3000 ppm exposure group when compared to dams in the control group. This reduction coincided with the exposure period (Days 6 through 15) with the exception of the second and third exposure (Days 7 and 8). Food consumption returned to normal upon cessation of exposure. In the 1500 ppm exposure group, food consumption was lower only after the first exposure and in the 400 ppm exposure group it was never lower (data for the pilot study not shown herein).

3. Body weights, liver-to-body weight ratios and liver to brain weight ratios were not significantly different in any exposure group. There were no dose-related gross necropsy findings in the dams or fetuses.

 

Main Developmental Toxicity Study.

1.Group parameters. Group indexes, counts and other summary data for the study are presented without analysis in Table 1.

 

TABLE 1.GROUP PARAMETERS

PM Acetate Rat Inhalation Developmental Study

	Control	500 ppm	2000 ppm	4000 ppm
Females mated	25	25	25	25
Fatalities	0	0	0	0
Females at sacrifice	25	25	25	25
Females pregnant	23	23	23	20
Fertility index (%)	92	92	92	80
Litters	23	23	23	20
Gestation Index (%)	100	100	100	100
Implantations (total)	333	330	320	278
Implantations per dam	14.5	14.3	13.9	13.9
Fetuses (total)	319	304	307	260
Fetuses per dam	13.9	13.2	13.3	13.0
Index of alive fetuses (%)	100	100	100	100
Dead Fetuses (total)	0	0	0	0
Dead Fetuses per dam	0	0	0	0
Resorptions (total)	14	26	13	18
Early resorptions	14	26	8	18
Late resorptions	0	0	5	0
Resorptions per dam	0.6	1.1	0.6	0.9
Resorption index (%)	4.20	7.88	4.06	6.47
Malformations (total)	0	1	1	0
Litters with Malformations	0	1	1	0
Malformations per dam	0	0.04	0.04	0
Index of malformations (%)	0	0.33	0.33	0
Variations (total)	17	19	16	11
Litters with Variations	9	13	7	6
Variations per dam	0.74	0.83	0.70	0.55
Index of variations (%)	5.33	6.25	5.21	4.23
Runts	0	0	2	3
Sex Ratio (M/F)	1.10	0.90	0.99	1.08

 

2. Maternal parameters.

a. Toxic signs. Nearly half of the 20 dams in the 4000 ppm exposure group exhibited dyspnea at various times throughout the exposure period (Days 6 through 15). Breathing returned to normal soon after the dams were returned to their boxes. Half had red to reddish brown discharges from the nose and/or eyes on Days 8 and 10 through 15. Four dams were observed to have yellow staining in the fur of the urogenital area ranging from slight to marked on Days 6, 8, 13 and 14. Reduced muscle tone was observed during handling in 15 dams on two separate occasions. Movement returned to normal within 20 minutes of being returned to their boxes. In the 2000 ppm exposure group, one dam exhibited dyspnea, one had a ruffled pelt and two had red discharges from the eye or mouth. No toxic signs were observed in the 500 ppm exposure group.

b. Food consumption. In the 4000 ppm exposure group, a reduction of food consumption as a percentage of maternal body weight coincided with exposure to PM Acetate. A similar pattern was seen in the 2000 ppm exposure group where food consumption was lower on Days 7, Days11 through 13 and Day 15. In the 500 ppm exposure group, food consumption was lower on Days 7 and11

c. Maternal body weights and body weight gains. Maternal body weights were lower in the 4000 and 2000 ppm exposure group after the last day of exposure only. Maternal body weights in the 500 ppm exposure group were always the same as controls.However, when weight gains were analyzed, dams in the 4000 ppm exposure group did not gain as much as controls during the exposure period and gained less overall. Dams in the 2000 ppm exposure group gained less weight during the first two thirds of the exposure period and less overall also Dams in the 500 ppm exposure group gained weight at the same rate as dams in the control group.

d.Maternal organ weight ratios. There were no differences in relative liver or uterus weights between dams in the control group and dams in any exposure group whether ratios were calculated from body weights or brain weights.

3.Litter and Fetal Parameters.

a. Corpora lutea, implantation, litter size, resorption and fetal death. The number of corpora lutea, implantation sites and live fetuses per litter was the same in the exposed groups as controls. Both the percent of conceptuses resorbed per litter and the percent of litters which contained a resorption were the same in the exposed groups as the controls. There were no dead fetuses in any litter.

b. Fetal body weights and runts. There were no dose related differences in fetal body weights. The average fetal body weights in the 4000 and 2000 ppm exposure groups was approximately 5 percent lower than the low and controls. This difference was statistically significant in the 2000 ppm exposure group, however, the 4000 ppm exposure group was marginally nonsignificant. There was no difference in the percent of litters which contained runts.

c. Fetal sex ratio. There was no difference in the male to female sex ratio.

d. Malformations and variations. There were no differences in the percent of fetuses per litter that were malformed, had variations or were normal. In addition, there were no differences in the percent of litters which contained a malformation, a variation or contained all normal fetuses.

Conclusions:

A NOAEC was established at 500 ppm (2700 mg/m3 measured) for dams and 4,000 ppm (22464 mg/m3, measured) for foetuses.

Executive summary:

In a GLP study, pregnant SD female rats were exposed to PGMA (purity: 97.3% of 2-methoxy-1-methylethyl acetate and 2.0 % of 1- methoxy-2- ) methylethyl acetate) vapour from Days 6 through 15 of gestation, once daily for 6 hours/day at nominal dose of 0, 500, 2,000, 4,000 ppm (0, 2700, 10800, 21600 mg/m3). The animals were sacrificed on Day 20 of gestation to evaluate the potential maternal, embryonic and teratogenic parameters of PGMA. Most of the effects observed in dams were transient in nature. Reductions in muscle tone (2000 and 4000 ppm), food consumption (500, 2000 and 4000 ppm) and body weight (2000 and 4000) were seen during the exposure period. At 2000 and 4000 ppm exposure groups, dyspnea, ruffled pelt and red discharges from the eyes or mouth were observed. No toxic signs were observed in the 500 ppm exposure group. The effect on nasal cavity was not examined in this experiment. No developmental toxicity was observed. A NOAEC was established at 500 ppm (2700 mg/m3, measured) for dams and 4000 ppm (22464 mg/m3, measured) for foetuses

Effect on developmental toxicity: via oral route

Endpoint conclusion:

no study available

Effect on developmental toxicity: via inhalation route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEC

22 464 mg/m³

Species:

rat

Quality of whole database:

Good (Klimisch 1)

Effect on developmental toxicity: via dermal route

Endpoint conclusion:

no study available

Additional information

Inhalation teratology study on PMA:

There was no evidence of teratogenicity of PGMA in the inhalation study performed on rats. In this study a NOAEL of 500 ppm (2,700 mg/m3) was observed for dams for systemic toxicity and 4,000 ppm (22,464 mg/m3) for foetuses for developmental effects (no effects at the highest tested dose). This is supported by the results of the OECD 422 study with PGMA.

Inhalation teratology studies on PM:

Several studies confirm that rapid and extensive hydrolysis of propylene glycol methyl ether acetate (PGMA) to propylene glycol methyl ether (PGME) occurs in vivo when PGMA is administered by the oral, inhalatory or dermal route. Since urinary metabolites and disposition profiles of PGMA were approximately identical to the results obtained with PGME, it is unlikely that there are substantial differences of the systemic toxicity between PGMA and PGME. In fact, toxicity of PGMA is almost the same of PGME. Therefore, data on PGME were also used to support the dataset for PGMA.

No teratogenic effects were observed with PGME at doses up to 3,000 ppm by inhalation route. In the 2-generation studies, foetotoxic effects were seen concurrently with maternal toxicity (3000 ppm by inhalation in rats (11220 mg/m3). This kind of effects (delayed ossification) are often reported concurrently with the maternal effects described in the available studies. Due to the low toxicity of PGME and that no specific developmental effects were observed at relatively high doses without maternal toxicity, it can be concluded that developmental toxicity of propylene glycol methyl ether is of no concern. Further justification for the use of read across is contained in the category document attached at section 13.

Based on the available data on PMA and PM it is concluded that PMA is not a developmental toxicant.

Justification for selection of Effect on developmental toxicity: via oral route:
There is a study available via the inhalation route

Justification for selection of Effect on developmental toxicity: via inhalation route:
GLP study

Justification for selection of Effect on developmental toxicity: via dermal route:
There is a study available via the inhalation route

Justification for classification or non-classification

No effects on fertility or specific developmental effects were observed with propylene glycol methyl ether acetate and propylene glycol methyl ether. Therefore, no classification is needed for this endpoint.