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Toxicity to reproduction

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Administrative data

Endpoint:
one-generation reproductive toxicity
Remarks:
based on generations indicated in Effect levels (migrated information)
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study
Cross-reference
Reason / purpose:
reference to same study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2004
Report Date:
2004

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
Deviations:
no
Principles of method if other than guideline:
Combined Subchronic Vapor Inhaltion Study with Reproduction/Developmental Toxicity Screening Test.
GLP compliance:
yes (incl. certificate)
Remarks:
BASF AG, Dept of Toxicology
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
isobutyl vinyl ether (IBVE), purity >99.5% as determined by GC.
The stability under storage conditions over the exposure period was guaranteed.
Storage condition: refrigerator, stored under nitrogen.
No further details

Test animals

Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
Wistar (CrlGlxBrlHan:WI) rats; female animals were nulliparous and non-pregnant
Source: Charles River, Germany
Housing: singly during the exposure period.
Free access to laboratory diet and tap water (analysed for contaminations) except during exposure and motor activity measurements.
Room: temperature 20-24°C
Relative air humidity 30-70%
Light/dark rhythm: 12 hrs.
Acclimatisation period: 6 or 13 days
Age st arrival: 4 or 5 weeks.

Administration / exposure

Route of administration:
inhalation
Type of inhalation exposure (if applicable):
whole body
Vehicle:
other: air
Details on exposure:
Exposure to vapors of TS in an whole-body inhalation chamber (volume 1.4 m³) in individual cages in groups of 10 rats. For each concentration the test substance was supplied to a thermostated vapourizer at a constant rate by means of the metering pump; the vapour was generated with conditioned supply air (about 50% ± 20% relative humidity, 220 C+-20 C) and passed into the inhalation system.
Details on mating procedure:
1 male with one female of the same group mated; starting 10 weeks after start of exposure; female satellite rats were not mated. The number of mating days until vaginal sperm could be detected in the female animals, and the pregnancy status of the female partner were recorded.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Concentration measurement by gas chromatography. At the beginning of the study, daily means were calculated based on 2 measured samples per concentration. Constancy in each inhalation system was continuously monitored by means of a total hydrocarbon analyzer. Correctness of hydrocarbon analyzer checked by GC. In the control group (test group 0) one sample per week was analyzed.
Duration of treatment / exposure:
13 weeks for males (93 days or 65 exposures and were sacrificed on the next workday after the last exposure)
15 weeks for females (67 to 72 exposures; sacrifice on study day 105)
13 weeks for non-mated satellite females (65 exposures in 92 days, sacrificed day 93)
Premating exposure period (males and females): 70 days
Frequency of treatment:
5 days/wk, 6 hrs/day
Details on study schedule:
The males were treated for appr. 13 weeks (10 weeks premating, 3 weeks mating and post mating); females were treated during premating (10 weeks), mating and gestation through day 4 after delivery (approx. 15 weeks). Additionally a satellite group of female rats (5 animals per concentration, nuliparous and non-pregnant, no reproduction; results see Section 7.5.3) were exposed 13 weeks. Mating pairs were formed from the same concentration group. The parental animals were examined for their mating and reproductive performances.
Comment: in contrast to the documentation in the summary (page 18 of the report) it was mentioned that parenteral females were exposed "until including day 18 post coitum resulting in a total of 67 to 72 exposures" (page 31) suggesting that gestation day 18 was the last exposure day and no further exposure of dams and pups was performed.
Doses / concentrationsopen allclose all
Remarks:
Doses / Concentrations:
0, 50, 500, 2000 ppm corresponding to 0, 208, 2080, 8300 mg/m³
Basis:
nominal conc.
Remarks:
Doses / Concentrations:
0, 208+-9.8, 2095+-57, 8301+-58 mg/m³ (means +- SD)
Basis:
analytical conc.
No. of animals per sex per dose:
10 male and 10 female rats per control and dose group
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: preliminary dose range finding study (see Section 7.5.3)
Positive control:
no

Examinations

Parental animals: Observations and examinations:
Rats controlled for clinical signs once daily (weekend) or twice daily (workdays); on exposure days clinical observation performed before, during and after
exposure. Body weight of the animals as well as food consumption was determined day0 and thereafter weekly; food efficiency calculated.

Detailed clinical observations in an open field conducted prior to the start of the exposure period and weekly thereafter.
A functional observational battery (FOB) and measurements of motor activity (MA) were carried out on study days 56 and 57 (separate randomization) for males and females (n=5 each sex), respectively.
Hematology and clinical chemistry performed towards the end of premating period (details see Section 7.5.3).

Oestrous cyclicity (parental animals):
no data
Sperm parameters (parental animals):
Parameters examined in male parental generations: testis weight, epididymis weight, histopathology of sex-organs
Litter observations:
The following parameters were examined in [F1] offspring: number and sex of pups, stillbirths, live births, postnatal mortality, weight gain (bw at day1 and 4 post partum). Live pups were examined each day for clinical symptoms (including gross-morphological findings).
Postmortem examinations (parental animals):
A complete necropsy including gross pathological evaluation and weighing of selected organs (liver, kidneys, adrenals, testes, epididymides, ovaries, uterus, thymus, spleen, brain, heart, lungs) performed.
Organs and tissues examined histopathologically in controls and high dose groups, except liver, nasal cavity, testis, epididymis, and ovaries (examined in all groups).
See also Section 7.5.3
Postmortem examinations (offspring):
All surviving pups (after sacrifice on day 4 p.p. by means of C02), all stillborn pups and those pups that died before schedule, were examined externally, eviscerated and their organs were assessed macroscopically.
Statistics:
Clinical and neurofunctional examinations: DUNNETT's-test, Fisher's Exact test, Wilcoxon test, Kruskal-Wallis test. Clinical pathology: Kruskal-Wallis test Pathology: Kruskal-Wallis test, Wilcoxon test.
Level of significance selected was p=0.05
Reproductive indices:
Male mating index, male fertility index, gestation index, female mating index, female fertility index
Offspring viability indices:
see litter observations

Results and discussion

Results: P0 (first parental animals)

General toxicity (P0)

Clinical signs:
effects observed, treatment-related
Body weight and weight changes:
effects observed, treatment-related
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Other effects:
not examined

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:
not examined
Reproductive function: sperm measures:
not examined
Reproductive performance:
effects observed, treatment-related

Details on results (P0)

50 ppm (208 mg/m³): no adverse effects were detectec.

500 ppm (2080 mg/m³)
toxic effects in parenteral rats
- increased white blood cell counts in the parental females (+ 23%) and satellite females (+ 35%)
- increased lymphocytes in the parental females (+ 22%) and satellite females (+ 36%)
- increased triglyceride concentrations in the females (+ 77%) and satellite females (+59%)
- hyperplasia of the respiratory epithelium in the nasal cavity in 2 females
- mucous cell reduction in the nasal cavity in 6 males and 7 females
- degeneration of the olfactory epithelium in the nasal cavity in 9 males and 6 females
- inflammatory cell infiltration in the nasal cavity in one female
- increased absolute kidney weights in parental males and satellite females
- increased absolute and relative liver weights in satellite females
- increased cytoplasmic basophilia in the liver of one parental male
No test substance related adverse effects with respect to clinical examinations/reproductive performance of the parental animals during gestation and lactation period, and no test substance related effects on development of the offspring.

2000 ppm (8300 mg/m³)
Signs of toxicity in parental male and of female rats (excluding clinical signs of toxicity during gestation and lactation period)
- Unspecific clinical symptoms like visually increased respiration (day 0 - 105), alopecia (day30-105) and apathy (on study days 0 - 7)
- increased white blood cell counts in the parental females (+ 52%) and satellite females (+40%)
- increased polymorphonuclear neutrophils (absolute) in the parental females (+ 81 %) and satellite females (+ 88%)
- increased lymphocytes (absolute) in the parental females (+ 49%) and satellite females (+34%)
- increased alkaline phosphatase activities in the parental males (+ 42%)
- increased triglyceride concentrations in the parental females (+ 403%) and satellite females (+ 100%)
- increased cholesterol concentrations in the parental females (+ 129%) and satellite females (+ 43%)
- hyperplasia of the respiratory epithelium in the nasal cavity in 8 males
- mucous cell reduction in the nasal cavity in 10 males and 8 females
- degeneration of the olfactory epithelium in the nasal cavity in 8 males and 10 females
- inflammatory cell infiltration in the nasal cavity in 3 males and 3 females
- increased absolute and relative kidney weights in parental animals and in satellite females
- increased absolute liver weights in parental males and satellite females ; increased relative liver weights in animals and in satellite females
- increased cytoplasmic basophilia in the livers of 3 parental males and 1 parental female
Signs of toxicity of parental female animals during gestation and lactation
- Visually increased respiration before, during and after exposure in the course of pregnancy
- Visually increased respiration during lactation
- Statistically significantly lower mean body weights in the females on gestation day 20 (-12%; decrease during lactation not significant, -9%)
- Impairment of average female body weight gain during entire gestation (- 30 %)
- significantly decreased mean number of implantation sites per dam (8.9 vs. 11.8 in the control)
- higher resorption rate (% postimplantation loss 13 .1 vs. 2.1 in the control)

Reproduction parameters
Male fertility index: 100% in control, low- and mid-dose animals, and 90 % in high-dose animals; within the normal range of this strain.
Female fertility index: 100 % in all test groups.
Gestation index: 100% in control, low- and mid-dose rats, and 89 % in high-dose animals; within the normal range of this strain.
Duration of gestation: very similar in all test groups (21.8 to 22.1 days).

Effect levels (P0)

open allclose all
Dose descriptor:
NOAEC
Effect level:
50 ppm
Sex:
male/female
Basis for effect level:
other: (208 mg/m³)
Dose descriptor:
LOAEC
Effect level:
500 ppm
Sex:
male/female
Basis for effect level:
other: (corresponding to 2080 mg/m³) local effects in the nasal cavity; adaptive effects in liver and kidney (increased metabolism of the test substance; no histopathological effects); NOAEC concerning reproductive toxicity
Dose descriptor:
NOAEC
Effect level:
500 ppm
Sex:
male/female
Basis for effect level:
other: (2080 mg/m³) NOAEC concerning reproductive parameters, no systemic maternal toxicity during gestation and lactation
Dose descriptor:
LOAEC
Effect level:
2 000 ppm
Sex:
male/female
Basis for effect level:
other: (8300 mg/m³) LOAEC concerning reproduction parameters and maternal toxicity during gestation and lactation (reduced live birth index; clinical signs in dams [increased respiration], reduced maternal body weight

Results: F1 generation

General toxicity (F1)

Clinical signs:
no effects observed
Mortality / viability:
mortality observed, treatment-related
Body weight and weight changes:
no effects observed
Sexual maturation:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
no effects observed
Histopathological findings:
not examined

Details on results (F1)

Live birth index: 100% in groups at low- and mid-dose, 93% (p<=0.05) in high-dose animals. In the high dose group statistically significantly lower live birth index (7 % below control), only 69 pups liveborn vs. 115 pups in the control group, 5 pups stillborn and 0 in the control.
Viability index day 4 post partum: 100% in groups at low- and mid-dose, 97% in high-dose animals (2 pups cannibalized).
Sex ratio: unchanged
Body weight: unchanged.
Stunted pups: none in controls and low-dose group, 3 in each the mid- and the high-dose group.
Signs of toxicity: none
Pup necropsy: few incidental findings, not related to treatment.

Effect levels (F1)

open allclose all
Dose descriptor:
other: NOAEC teratogenicity
Generation:
F1
Effect level:
2 000 ppm
Sex:
male/female
Basis for effect level:
other: (8300 mg/m³) highest dose level tested
Dose descriptor:
NOAEC
Generation:
F1
Effect level:
500 ppm
Sex:
male/female
Basis for effect level:
other: (2080 mg/m³) no reproductive or developmental toxicity; also NOAEC maternal toxicity
Dose descriptor:
LOAEC
Generation:
F1
Effect level:
2 000 ppm
Sex:
male/female
Basis for effect level:
other: (8300 mg/m³) lower live birth index; decrease in implantation sites (increased resorptions)

Overall reproductive toxicity

Reproductive effects observed:
not specified

Any other information on results incl. tables

Further details on repeated dose toxicity are presented in Section 7.5.3.

No adverse effect of treatment on weights or histopathology of male and female reproductive organs.

Applicant's summary and conclusion

Conclusions:
Reproductive toxicity (reduced live birth index in F1) was found at 2000 ppm (8300 mg/m³) in the presence of maternal toxicity like clinical signs and reduced body weight. The NOAEC for reproductive toxicity is 500 ppm (2080 mg/m³). The LOAEC for toxic effects in the nasal cavity and adaptive effects in liver and kidney of P ratswas 500 ppm, the NOAEC was 50 ppm (208 mg/m³).
Executive summary:

In a GLP Guideline study according to OECD TG422 10 male and 10 female Wistar rats per test group were exposed to a vapor of isobutylvinylether (whole body, 6h/d, 5d/week) at dose levels of 0, 50, 500 or 2000 ppm (0, 208, 2080 and 8300 mg/m³). The males were treated for approx. 13 weeks (10 weeks premating, 3 weeks mating and post mating) and females during premating (10 weeks), mating and gestation through day 4 after delivery (approx. 15 weeks). The main target organ was the upper respiratory tract (see also Sextion 7.5.3) at >=500 ppm. Furthermore, dose dependent changes in some blood parameters were seen in female animals at >=500 ppm. High serum triglyceride and cholesterol levels were noted in high-dose parental females; the effect was less pronounced in non-pregnant females and absent in males. Increased liver and kidney weight changes were noted in high-dose males and intermediate and high-dose non-pregnant females. No effects were found at any concentration on mating and gestation index or histopathological lesions of the reproductive organs, however, the live born index was significantly reduced at the high dose level (increased resorptions and reduced implantation sites reported). No other effects were seen in the F1 generation.

Conclusion: Reproductive toxicity (reduced live birth index in F1) was found at 2000 ppm (8300 mg/m³) in the presence of maternal toxicity like clinical signs and reduced body weight. The NOAEC for reproductive toxicity is 500 ppm (2080 mg/m³). The LOAEC for toxic effects in the nasal cavity and adaptive effects in liver and kidney of P ratswas 500 ppm, the NOAEC was 50 ppm (208 mg/m³).