2-methylpropyl-(R)-2-hydroxypropanoate

Developmental toxicity / teratogenicity

Administrative data

Endpoint:

developmental toxicity

Type of information:

migrated information: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Publication. Study conducted in accordance to OECD guideline 414. 2-methyl-1-propanol (isobutanol) was used for read-across to isobutyl-R-lactate

Cross-reference

Reason / purpose for cross-reference:

reference to same study

Data source

Reference

Reference Type:

publication

Title:

Studies on the Prenatal Toxicity of 3-Methyl-1-butanol and 2-Methyl-1-propanol in Rats and Rabbits Following Inhalation Exposure

Author:

Klimisch, HJ

Year:

1995

Bibliographic source:

Fundamental and Applied Toxicology 27: 77-89

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

Deviations:

no

GLP compliance:

not specified

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

2-methylpropan-1-ol

EC Number:

201-148-0

EC Name:

2-methylpropan-1-ol

Cas Number:

78-83-1

Molecular formula:

C4H10O

IUPAC Name:

2-methylpropan-1-ol

Constituent 2

Reference substance name:

isobutanol

IUPAC Name:

isobutanol

Test material form:

other: liquid

Details on test material:

- Name of test material (as cited in study report): 2-methyl-1-propanol (MEP)
- Purity: 99.8 %
- Stability: Stable for at least 6 months

Test animals

Species:

rat

Strain:

Wistar

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Dr. K Thomae GmbH, Biberach/Riss, Germany
- Age at study initiation: 10 to 11 weeks
- Weight at study initiation: Mean body weight of 216 g
- Housing: Individually in wire cages (type DK III)
- Diet (e.g. ad libitum): Ad libitum
- Water (e.g. ad libitum): Ad libitum
- Acclimation period: At least 5 days

ENVIRONMENTAL CONDITIONS
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:

inhalation: vapour

Type of inhalation exposure (if applicable):

whole body

Vehicle:

clean air

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
Concentrations of 0.5, 2.5, and 10 mg/liter were achieved by supplying the test substances via continuously operating pumps to evaporators maintained at 50-70 °C by a water circulation thermostat. The vapours were diluted with clean air. This vapour-air mixture was distributed to a horizontal-flow whole-body exposure chamber at a flow rate corresponding to about 15 air exchanges per hour (inhalation chamber glass/steel construction with volumes of approximately 1.1 m³; manufactured by BASF AG, Ludwigshafen, Germany).
Supply and exhaust air flows were adjusted by flow meters, in order to achieve a minimal negative pressure in the inhalation chambers. Pressure and temperature (21-24 °C) were measured continuously. The relative humidity (49-64 %) was checked with a hygrometer (Humicap, Vaisala, Finland).

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Samples of the inhalation atmospheres were analyzed hourly by gas chromatography (Hewlett-Packard gas chromatograph Model 5840 A with an automatic sampler Model 7671 A, FID; column, 2 m × 2 mm with 15 % Ucon LB 550 x on Chromosorb W/HP; 80/100 mesh; oven temperature, 90 °C).

Details on mating procedure:

- M/F ratio per cage: 1/4
- Proof of pregnancy: Sperm in vaginal smear referred to as day 0. The following day was defined as day 1 postcoitum (pc).

Duration of treatment / exposure:

6 hours/day

Frequency of treatment:

During the adaptation period, the animals were exposed to clean air in the inhalation chambers for 6 hr/day (days 1-5 pc), The animals were
exposed individually in wire cages from days 6 to 15 pc to substance concentrations or air for 6 hr/day.

Duration of test:

After termination of the exposure period, the rats were observed up to day 20 pc.

No. of animals per sex per dose:

25 females

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale:
In a range-finding study 10 pregnant rats per group were exposed to 0.5, 2.5, or 5.0 mg/liter for 6 hr each day, from day 6 to 15 of gestation. No maternally toxic effects (clinical signs, body weight, various haematological, and clinicochemical parameters, gross pathological findings at necropsy) could be observed up to 5 mg/liter.

Examinations

Maternal examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Every day

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Every day

BODY WEIGHT: Yes
- Time schedule for examinations: Day 0, 3 and 6 and from then on at 3-day intervals until day 20 pc

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 20
- Organs examined: All animals were necropsied and assesed by gross pathology. Uterus and ovaries were removed.

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

Fetal examinations:

- External examinations: Yes, all per litter
- Soft tissue examinations: Yes, half per litter
- Skeletal examinations: Yes, half per litter

Statistics:

The Dunnett test (Dunnett, 1955, 1964) was used to compare body weight, body weight changes, corrected body weight gain, intact uterine weight, foetal and placental weights, the number of corpora lutea, implants, resorptions, live foetuses, and pre- or post-implantation losses. Fisher's exact test (Dixon, 1981) was used for evaluating the conception rate, maternal mortality, and all foetal findings.

Indices:

The following definitions were used to describe fetal changes:
- Malformations (external, soft tissue, and skeletal observations) — infrequent and/or probably lethal changes.
- Variations (external, soft tissue, and skeletal observations) — changes which occur regularly also in the control groups and do not impair the
survival.
- Retardations (skeletal observations) — delayed development of the fetal skeletons.
- Unclassified observations (external or soft tissue observations) — changes which are considered neither malformations nor variations.

Historical control data:

N.A.

Results and discussion

Results: maternal animals

General toxicity (maternal animals)

Clinical signs:

no effects observed

Description (incidence and severity):

No substance-induced clinical findings were observed in any of the test groups.

Dermal irritation (if dermal study):

not examined

Mortality:

no mortality observed

Body weight and weight changes:

no effects observed

Description (incidence and severity):

Treatment with MEP did not significantly influence the body weights and body weight changes.

Food consumption and compound intake (if feeding study):

not examined

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

no effects observed

Description (incidence and severity):

No gross-pathological findings were observed. Findings such as hydrometra, oedema, or marginal emphysema of lungs in few rats without any relation to treatment were considered to be spontaneous events.

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

not examined

Histopathological findings: neoplastic:

not examined

Other effects:

not examined

Maternal developmental toxicity

Number of abortions:

no effects observed

Pre- and post-implantation loss:

no effects observed

Total litter losses by resorption:

no effects observed

Early or late resorptions:

no effects observed

Dead fetuses:

no effects observed

Changes in pregnancy duration:

not examined

Description (incidence and severity):

Migrated Data from removed field(s)
Field "Effects on pregnancy duration" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsMaternalAnimals.MaternalDevelopmentalToxicity.EffectsOnPregnancyDuration): not specified

Changes in number of pregnant:

not examined

Details on maternal toxic effects:

Maternal toxic effects:no effects

Details on maternal toxic effects:
- Treatment with MEP did not significantly influence the body weights and body weight changes.
- No substance-induced clinical findings were observed in any of the test groups
- No mortality occured
- No gross-pathological findings were observed. Findings such as hydrometra, oedema, or marginal emphysema of lungs in few rats without any relation to treatment were considered to be spontaneous events
- The uterine weights of the animals exposed to MEP were not significantly different from their respective controls
- No effects were noted for: Conception rate, mean number of corpora lutea, implantation sites, pre-and postimplantational loss, number of resorptions and viable foetuses.

Effect levels (maternal animals)

1

Dose descriptor:

NOAEC

Effect level:

10 mg/L air

Based on:

test mat.

Basis for effect level:

other: maternal toxicity

2

Dose descriptor:

NOAEC

Effect level:

10 mg/L air

Based on:

test mat.

Basis for effect level:

other: developmental toxicity

Results (fetuses)

Fetal body weight changes:

no effects observed

Description (incidence and severity):

Mean placental and foetal weights were not affected by treatment.
Migrated Data from removed field(s)
Field "Fetal/pup body weight changes" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsFetuses.FetalPupBodyWeightChanges): no effects observed
Field "Description (incidence and severity)" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsFetuses.DescriptionIncidenceAndSeverityFetalPupBodyWeightChanges): Mean placental and foetal weights were not affected by treatment.

Reduction in number of live offspring:

not examined

Changes in sex ratio:

no effects observed

Description (incidence and severity):

Sex distribution did not differ significantly between treated groups and the negative control group.

Changes in litter size and weights:

no effects observed

Description (incidence and severity):

See box "Any other information on results incl. tables" below.

Changes in postnatal survival:

not specified

External malformations:

effects observed, treatment-related

Description (incidence and severity):

External changes in foetuses: Anasarca was observed in two foetuses at 10 mg/L, one of which had an additional cleft palate. No external variations were found in any group. Four control foetuses and two foetuses of the 10 mg/L group showed fused or necrobiotic placentae.

Skeletal malformations:

effects observed, treatment-related

Description (incidence and severity):

Skeletal examinations of the foetuses: In five control foetuses, in nine foetuses after exposure to 0.5 mg/L, two foetuses after exposure to 2.5 mg/L and in one fetus after exposure to 10 mg/L malformations of the sternebrae and/or vertebral column occured. The differences were not statistically significant. Any observed differences bewteen the groups, however, were within the range of biological variation and/or occured without a clear concentration dependency.

Visceral malformations:

effects observed, treatment-related

Description (incidence and severity):

Soft tissue changes in foetuses: Dilation of both heart ventrilces was observed in one foetus of the 10 mg/L dose group. Variations were seen in all groups including the controls. Specifically, dilated renal pelvis and hydroureter occured independent of exposure or concentration.

Other effects:

no effects observed

Description (incidence and severity):

No unclassified observations were observed for any test group.

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects: no effects

Details on embryotoxic / teratogenic effects:
- Sex distribution did not differ significantly between treated groups and the negative control group
- Mean placental and foetal weights were not affected by treatment
- External changes in foetuses: Anasarca was observed in two foetuses at 10 mg/L, one of which had an additional cleft palate. No external variations were found in any group. Four control foetuses and two foetuses of the 10 mg/L group showed fused or necrobiotic placentae.
- Soft tissue changes in foetuses: Dilation of both heart ventrilces was observed in one foetus of the 10 mg/L dose group. Variations were seen in all groups including the controls. Specifically, dilated renal pelvis and hydroureter occured independent of exposure or concentration.
- No unclassified observations were observed for any test group
- Skeletal examinations of the foetuses: In five control foetuses, in nine foetuses after exposure to 0.5 mg/L, two foetuses after exposure to 2.5 mg/L and in one fetus after exposure to 10 mg/L malformations of the sternebrae and/or vertebral column occured. The differences were not statistically significant. Any observed differences bewteen the groups, however, were within the range of biological variation and/or occured without a clear concentration dependency.

Effect levels (fetuses)

Key result

Dose descriptor:

NOAEC

Effect level:

10 mg/L air

Based on:

test mat.

Basis for effect level:

other: No effects seen on the foetal organisms in comparison to the untreated control.

Fetal abnormalities

Abnormalities:

no effects observed

Overall developmental toxicity

Developmental effects observed:

not specified

Any other information on results incl. tables

Table 1: Concentrations measured in inhalation chambers
		MEP
test group		mg/L	± SD
1		0.49	0.012
2		2.50	0.084
3		10.10	0.33

Table 2: Maternal body weight change and uterine weight in rats
Test group		0		1		2		3
mg/L		0		0.5		2.5		10
Number of animals		25		25		25		25
Number of dams		21*		23*		23*		19*
mean maternal body weight change
Days 0-3		13.8 (6.71)		13.5 (3.98)		15.4 (5.22)		14.9 (5.39)
Days 3-6		12.0 (4.35)		11.0 (3.38)		11.5 (3.81)		9.9 (2.66)
Days 6-9		11.6 (4.47)		13.6 (3.06)		13.5 (3.84)		11.8 (3.81)
Days 9-12		19.6 (4.29)		22.5 (4.41)		20.8 (5.06)		21.5 (4.00)
Days 12-15		19.5 (3.94)		18.5 (4.00)		18.2 (4.42)		19.6 (4.28)
Days 15-18		34.5(6.23)		36.0 (4.80)		33.4 (7.49)		36.0 (3.60)
Days 18-20		33.6 (5.84)		37.8(4.94)		34.5 (6.71)		37.7 (6.45)
Mean corrected maternal body weight change		47.7 (9.21)		50.3 (6.86)		50.4 (8.25)		50.9 (6.96)
Intact uterine weight		71.0 (15.28)		78.1 (10.12)		70.0 (17.36)		75.6 (11.58)

Note: numbers in parantheses indicate standard deviation

*= Differences in the number of animals and dams are due to the number of not pregnant animals

Table 3: Summary of reproduction data in rats
Test group		0		1		2		3
mg/L		0		0.5		2.5		10
Number of animals		25		25		25		25
Number of dams		21*		23*		23*		19*
Corpora lutea/dam		15.2		15.5		15.3		14.5
Implants/dam		13.6		14.6		13.1		13.8
% preimplantation loss		10.3		5.5		14.1		4.8
Live fetuses/dam		12.3 (2.89)		13.7 (1.99)		12.3 (3.35)		13.2 (1.86)
Dead fetuses/dam		1.2		1		0.8		0.6
% postimplantation loss		10		6.5		5.7		4.2 **
Sex ratio (m/f)		53.3 : 46.7		48.7 : 51.3		53.0 : 47.0		50.8 : 49.2
Mean fetal weight (g)		3.9 (0.29)		3.9 (0.28)		3.9 (0.21)		3.9 (0.15)
Mean placental weight (g)		0.46 (0.07)		0.43 (0.06)		0.45 (0.06)		0.46 (0.05)

Note: numbers in parantheses indicate standard deviation

*= Differences in the number of animals and dams are due to the number of not pregnant animals

**= p<0.05

Table 4: Summary of fetal external malformations in rats
Test group		0		1		2		3
mg/L		0		0.5		2.5		10
Number of litters evaluated		21		23		23		19
Number of fetuses evaluated		259		314		283		250
External malformations
Polydactyly
Fetal incidence		0		0		0		0
Litter incidence		0		0		0		0
Anasarca
Fetal incidence		0		0		0		2 (0.8)
Litter incidence		0		0		0		2 (10.5)
Cleft palate
Fetal incidence		0		0		0		1 (0.4)
Litter incidence		0		0		0		1 (5.3)
Total fetal external malformations
Fetal incidence		0		0		0		2 (0.8)
Litter incidence		0		0		0		2 (10.5)

Note: numbers in parantheses indicate percentage of fetuses/litters affected

Table 5: Summary of fetal soft tissue malformations in rats
Test group		0		1		2		3
mg/L		0		0.5		2.5		10
Number of litters evaluated		21		23		23		19
Number of fetuses evaluated		125		150		134		122
Soft tissue malformations
Dilatation of both ventrocles (globular shaped heart)
Fetal incidence		0		0		0		1 (0.8)
Litter incidence		0		0		0		1 (5.3)
Dextrocardia
Fetal incidence		0		0		0		0
Litter incidence		0		0		0		0
Total fetal soft tissue malformations
Fetal incidence		0		0		0		1 (0.8)
Litter incidence		0		0		0		1 (5.3)

Note: numbers in parantheses indicate percentage of fetuses/litters affected

Table 6: Summary of fetal soft tissue variations in rats
Test group		0		1		2		3
mg/L		0		0.5		2.5		10
Number of litters evaluated		21		23		23		19
Number of fetuses evaluated		125		150		134		122
Soft tissue variations
Dilated renal pelvis
Fetal incidence		59 (47.2)		47** (31.3)		47 (35.1)		49 (40.2)
Litter incidence		19 (90.5)		18 (78.3)		20 (87.0)		17 (89.5)
Hydroureter
Fetal incidence		5 (4.0)		8 (5.3)		5 (3.7)		6 (4.9)
Litter incidence		5 (23.8)		5 (21.7)		3 (13.0)		5 (26.3)
Total fetal soft tissue variations
Fetal incidence		60 (48.0)		47** (31.3)		47* (35.1)		49 (40.2)
Litter incidence		19 (90.5)		18 (78.3)		20 (87.0)		17 (89.5)

Note: numbers in parantheses indicate percentage of fetuses/litters affected

*= p<0.05; **= p< 0.01

Table 7: Summary of fetal skeletal malformations in rats
Test group		0		1		2		3
mg/L		0		0.5		2.5		10
Number of litters evaluated		21		23		23		19
Number of fetuses evaluated		134		164		149		128
Skeletal malformations
Thoracic vertebral arch and corressponding rib mssing
Fetal incidence		0		0		0		0
Litter incidence		0		0		0		0
Cervical vertebral arch missing
Fetal incidence		0		0		0		0
Litter incidence		0		0		0		0
Thoracic vertebral body/bodies dumbell-shaped (asymmetrical)
Fetal incidence		1 (0.7)		4 (2.4)		2 (1.3)		0
Litter incidence		1 (4.8)		4 (17.4)		2 (8.7)		0
Thoracic vertebral body/bodies bipartite (asymmetrical)
Fetal incidence		0		1 (0.6)		1 (0.7)		0
Litter incidence		0		1 (4.3)		1 (4.3)		0
Different sacral vertebral bodies and/or arches severely malformed
Fetal incidence		0		0		0		0
Litter incidence		0		0		0		0
Sternebrae bipartite, ossification centers dislocated
Fetal incidence		3 (2.2)		4 (2.4)		0		1 (0.8)
Litter incidence		3 (14.3)		4 (17.4)		0		1 (5.3)
Cleft sternum
Fetal incidence		0		0		0		0
Litter incidence		0		0		0		0
Thoracic vertebra absent
Fetal incidence		0		0		1 (0.7)		0
Litter incidence		0		0		1 (4.3)		0
Lumbar vertebra absent
Fetal incidence		1 (0.7)		0		0		0
Litter incidence		1 (4.8)		0		0		0
Total fetal skeletal malformations
Fetal incidence		5 (3.7)		9 (5.5)		2 (1.3)		1 (0.8)
Litter incidence		5 (23.8)		7 (30.4)		2 (8.7)		1 (5.3)

Note: numbers in parantheses indicate percentage of fetuses/litters affected

Table 8: Summary of fetal skeletal variations in rats
Test group		0		1		2		3
mg/L		0		0.5		2.5		10
Number of litters evaluated		21		23		23		19
Number of fetuses evaluated		134		164		149		128
Skeletal variations
Sternebrae of irregular shape
Fetal incidence		42 (31.3)		57 (34.8)		49 (32.9)		39 (30.5)
Litter incidence		18 (85.7)		21 (91.3)		21 (91.3)		18 (94.7)
Sternebrae bipartite
Fetal incidence		7 (5.2)		3 (1.8)		2 (1.3)		4 (3.1)
Litter incidence		5 (23.8)		3 (13.0)		2 (8.7)		4 (21.1)
Accessory sternebra
Fetal incidence		0		0		1 (0.7)		0
Litter incidence		0		0		1 (4.3)		0
13th rib shortened
Fetal incidence		17 (12.7)		22 (13.4)		28 (18.8)		12 (9.4)
Litter incidence		10 (47.6)		12 (52.2)		8 (34.8)		7 (36.8)
Rudimentary cervical rib
Fetal incidence		6 (4.5)		4 (2.4)		5 (3.4)		6 (4.7)
Litter incidence		4 (19.0)		3 (13.0)		5 (21.7)		4 (21.1)
Accessory 14th rib
Fetal incidence		1 (0.7)		0		1 (0.7)		0
Litter incidence		1 (4.8)		0		1 (4.3)		0
13th rib absent
Fetal incidence		0		0		0		0
Litter incidence		0		0		0		0
Flying rib
Fetal incidence		0		0		1 (0.7)		0
Litter incidence		0		0		1 (4.3)		0
Total fetal skeletal variations
Fetal incidence		59 (44.0)		75 (44.0)		72 (48.3)		51 (39.8)
Litter incidence		19 (90.5)		22 (95.7)		22 (95.7)		18 (94.7)

Note: numbers in parantheses indicate percentage of fetuses/litters affected

Table 9: Summary of all classified fetal external, soft tissue and skeletal observations in rats
Test group		0		1		2		3
mg/L		0		0.5		2.5		10
Number of litters evaluated		21		23		23		19
Number of fetuses evaluated		259		314		283		250
Alteration
Total malformations
Fetuses		5 (1.9)^a		9 (2.9)		2 (0.7)		3 (1.2)
Litters		5 (23.8)^b		7 (30.4)		2 (8.7)		3 (15.8)
Mean % affected fetuses
per litter		2 (3.69)^c		2.8 (4.69)		0.7 (2.43)		1.3 (3.24)
Total variations
Fetuses		119 (45.9)		122 (38.9)		119 (42.0)		100 (40.0)
Litters		21 (100.0)		23 (100.0)		23 (100.0)		19 (100.0)
Mean % affected fetuses
per liiter		48.3 (22.30)		39.5 (18.89)		44.6 (21.46)		39.7 (14.36)
Total retardations
Fetuses		55 (21.2)		57 (18.2)		66 (23.3)		36* (14.4)
Litters		18 (85.7)		18 (78.3)		21 (91.3)		14 (73.7)
Mean % affected fetuses
per litter		22.2 (15.88)		17.8 (13.89)		24.1 (17.15)		14.4 (11.91)

Note: Significantly different from control at * = p < 0.05. Numbers in parentheses indicate:

a = percentage of fetuses affected, b = percentage of litters affected and c = standard deviations

Applicant's summary and conclusion

Conclusions:

No treatment-related effects in developmental parameters or maternal parameters were detected in a developmental toxicity study (OECD 414) after inhalation of 2-methyl-1-propanol. The maternal NOAEC is considered to be 10 mg/L and the developmental NOAEC is also considered to be 10 mg/L.

Executive summary:

In a developmental toxicity study (OECD 414), 2-methyl-1-propanol (MEP) (99.8 % purity) was administered to 25 female Wistar rats per dose in clean air (whole body exposure for 6 hours/day) at concentrations levels of 0, 0.5, 2.5 and 10 mg/L from day 6 through day 15 of gestation. There were no treatment-related effects in mortality, clinical signs, body weight, food consumption, or Caesarean parameters. The maternal NOAEC is 10 mg/L. Moreover, no treatment-related effects were noted in developmental parameters. The developmental NOAEC is 10 mg/L. The developmental toxicity study in the rat is classified acceptable and satisfies the guideline requirement for a developmental toxicity study (OPPTS 870.3700; OECD 414) in rat. MEP was used as read-across partner to isobutyl-(R)-lactate.