Tetrahydro-4-methylphthalic anhydride

Toxicity to reproduction

Administrative data

Endpoint:

screening for reproductive / developmental toxicity

Remarks:

based on test type (migrated information)

Type of information:

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

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: This study was according to OECED guide lines and was performed under GLP.

Cross-reference

Cross-reference 1

Reason / purpose for cross-reference:

reference to same study

Cross-reference 2

Reason / purpose for cross-reference:

reference to other study

Data source

Reference

Reference 1

Reference Type:

study report

Title:

Unnamed

Year:

1997

Report date:

1997

Reference 2

Reference Type:

publication

Title:

Unnamed

Year:

1997

Report date:

1997

Materials and methods

Test guideline

Qualifier:

according to guideline

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:

not applicable

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Tetrahydromethylphthalic anhydride

EC Number:

234-290-7

EC Name:

Tetrahydromethylphthalic anhydride

Cas Number:

11070-44-3

IUPAC Name:

3a-methyl-3a,4,5,6-tetrahydro-2-benzofuran-1,3-dione

Details on test material:

- Name of test material (as cited in study report): tetrahydromethyl-1,3-isobenzofurandione
- Analytical purity: 99.7%
- Lot/batch No.: 2522
- Stability under test conditions: stable, Vehicle: Corn oil (nakaraitesk, lot No, V5P5523)
- Storage condition of test material: room temperature, dark place

Test animals

Species:

rat

Strain:

Crj: CD(SD)

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles river Japan
- Age at study initiation: purchase at 9 weeks old, 1st. administration at 10 weeks old
- Weight at study initiation: male: 356.3-394.4g, female: 213.5-252.9g
- Fasting period before study: 18hr
- Housing: dosing period: stainless hanger gage, one animal/gage, mating period: polycarbonate gage with wood chip
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: one week


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22-26°C
- Humidity (%): 45-65%
- Air changes (per hr): 13 times/hr
- Photoperiod (hrs dark / hrs light):12/12 AM07:00-PM07:00

Administration / exposure

Route of administration:

oral: gavage

Type of inhalation exposure (if applicable):

not specified

Vehicle:

corn oil

Details on exposure:

VEHICLE
- Justification for use and choice of vehicle (if other than water): Corn oil is used generally
- Concentration in vehicle: 0.6, 2 and 6w/v%
- Amount of vehicle (if gavage): 5mL/kg
- Lot/batch no. (if required): V5P5831, nakalai tesque Co.

Details on mating procedure:

Male/female per cage: 1/1,
length of cohabitation: maximal 14 days, until proof of pregnancy (formation of vaginal closing or sperm detection in vagina)

Analytical verification of doses or concentrations:

not specified

Details on analytical verification of doses or concentrations:

no details given

Duration of treatment / exposure:

Males; for 49 days Females; from 14 days before mating to day 3 of lactation (38 days in total)

Frequency of treatment:

one administration/day

Details on study schedule:

no details given

Doses / concentrations

Remarks:

Doses / Concentrations:
0 (Vehicle), 30, 100 and 300 mg/kg/day (in corn oil)
Basis:
nominal conc.

No. of animals per sex per dose:

Doses is 0, 30, 100 and 300 mg/kg. 12 animals per sex per dose.

Control animals:

yes, concurrent vehicle

Details on study design:

Dose selection rationale: oral, one of the identical exposure route for human
300 mg/kg: 1/3 volume from omens of death (1000 mg/kg)
100 and 30 mg/kg: common ratio 3 of 300 mg/kg

Positive control:

no details given

Examinations

Parental animals: Observations and examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: one time per day
- Cage side observations: general symptom

DETAILED CLINICAL OBSERVATIONS: No
BODY WEIGHT: Yes
- Time schedule for examinations: male: two times/week
female: two times/ week at pre-mating period, in pregnant period: day at 0, 4, 7,10, 14, 17 and 21, in lactation period: the day 0 and 4.

FOOD CONSUMPTION AND COMPOUND INTAKE:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
male: two times/week
female: two times/ week at pre-mating period, in pregnant period: day at 1, 4, 7,10, 14, 17 and 21, in lactation period: the day 1 and 4.

FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: No

OPHTHALMOSCOPIC EXAMINATION: No

HAEMATOLOGY: Yes
- Time schedule for collection of blood: after dosing period
- Anaesthetic used for blood collection: Yes (identity): Pentobarbital-Na i.p.
- Animals fasted: Yes
- How many animals: All of male
- Parameters checked in table (see below in remarks field) were examined.: WBC, RBC, Hb, Ht, PLT

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: after dosing period (same time for HEMATOLOGY)
- Animals fasted: Yes
- How many animals:All of male
- Parameters checked in table (see below in remarks field) were examined.: TP, ALB, A/G, Bil, GOT, GPT, TGace, ALP, TG, PL, Giu, BUN, CRE, P, Ca, Na, K, Cl

URINALYSIS: No

NEUROBEHAVIOURAL EXAMINATION: No

Oestrous cyclicity (parental animals):

no details given

Sperm parameters (parental animals):

no details given

Litter observations:

Body weight (at day of birth and day 4 after birth), sex, surface abnormality at day of birth.

Postmortem examinations (parental animals):

SACRIFICE
- Male animals: All surviving animals, as soon as possible after the last litters in each generation were produced.
- Maternal animals: All surviving animals, after the last litter of each generation was weaned.

GROSS NECROPSY
- Gross necropsy consisted of external and internal examinations including the cervical, thoracic, and abdominal viscera.
organ weight: brain, heart, lung, thymus, liver, spleen, kidney, adrenal, testis, epididymis, ovary.
microscopic investigation: all animals in control, 300 mg/kg group; brain, pituitary gland, eyeball, thyroid gland, parathyroid gland, thymus, heart, lung, liver, kidney, adrenal, spleen, stomach, small intestine, large intestine, pancreas, urinary bladder, bone marrow, ovary, uterus, vagina, mammary gland.
Unfertilized animals in any groups: testes, epididymis and ovary

Postmortem examinations (offspring):

Full macroscopic examinations on all of pups Parameters assessed during study.

Statistics:

bartlett method,
standard variance: Dunnett, multiple comparison
non-standard variance: Steel, multiple comparison

Reproductive indices:

No. of pairs with successful copulation
Copulation index (No. of pairs with successful copulation/No. of pairs mated x 100)
Pairing days until copulation
No. of pregnant females
Fertility index = (No. of pregnant animals x 100/No. of pairs with successful copulation),
No. of corpora lutea
No. of implantation sites
No. of living pregnant females
No. of pregnant females with parturition gestation length
No. of pregnant females with live pups on day 0
Gestation index (No. of females with live pups x 100/No. of living pregnant females)
Delivery index (No. of pups born x 100/No. of implantation sites)

Offspring viability indices:

No. of pups alive on day 0 of lactation
Live birth index (No. of live pups on day 0 x 100/No. of pups born)
Sex ratio (Total No. of male pups/Total No. of female pups)
No. of pups alive on day 4 of lactation, body wt. of live pups (on day 0 and 4)

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:

no effects observed

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

no effects observed

Organ weight findings including organ / body weight ratios:

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Other effects:

no effects observed

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:

no effects observed

Reproductive function: sperm measures:

not examined

Reproductive performance:

no effects observed

Details on results (P0)

For details on results for parental animals, please refer to IUCLID5 section 7.5.1 (repeated dose toxicity oral).
Reproduction parameters:
- At 30 and 100 mg/kg, there was a tendency for decrease of estrous frequency, but at 300 mg/kg, no statistically significant effects were observed.
- For details see tables below.

Effect levels (P0)

Dose descriptor:

NOAEL

Effect level:

> 300 mg/kg bw/day (nominal)

Sex:

male/female

Basis for effect level:

other: no effects were observed

Results: F1 generation

General toxicity (F1)

Clinical signs:

no effects observed

Mortality / viability:

no mortality observed

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)

No adverse effects were reported.
- At 30 and 100 mg/kg, statistically significant decrease of birth index was observed, and at 300 mg/kg, stillborn was observed only one animal (not statistically significant).
- At 100 mg/kg, total litter loss in two dams were observed.
- At 300 mg/kg, no statistically significant effects were observed, but there was a tendency for decrease of developmental parameters (total number of pups born, delivery index and live birth index).
- For details see table below.

Effect levels (F1)

Dose descriptor:

NOAEL

Generation:

F1

Effect level:

> 300 mg/kg bw/day (nominal)

Sex:

male/female

Basis for effect level:

other: no effects were observed

Overall reproductive toxicity

Reproductive effects observed:

not specified

Any other information on results incl. tables

Table 1: Reproduction parameters

Dose level (mg/kg/day)	0	30	100	300
No. of dams	11	11	11	10
No. of corpora lutea (Mean +/- SD)	22.18 +/-0.40	22.27 +/-0.47	22.09 +/-0.30	22.30  +/-0.48
No. of implantations (Mean +/- SD)	183 16.64 +/- 1.63	188 17.09 +/- 1.30	188 17.09 +/- 0.94	162 16.20 +/- 1.03
No. of litter (Mean +/- SD)	171 15.55 +/- 2.58	178 16.18 +/- 1.25	181 16.45 +/- 0.69	154 15.40 +/- 1.65
Gestation Index	100	100	100	100
No. of stillborns Male Female Total %	0 0 0 (0)	4 4 8 (5.06)*	3 6 9 (5.33)*	4 6 10 (6.76)
No. of live newborns (Mean +/- SD)	162 14.73 +/- 2.65	150 13.64 +/- 1.43	160 14.55 +/- 0.82	138 13.80 +/- 2.53
Birth index	94.74	84.27*	88.40*	89.61
Sex ratio of live newborns (male/female)	80/82	71/79	83/77	64/74
Body weight of live pups (g) (mean +/- SD) on day 0 Males Females	6.2 +/- 0.5 9.4 +/- 1.2	6.1 +/- 0.4 9.5 +/- 1.1	6.0 +/- 0.4 9.1 +/- 0.7	6.3 +/- 0.5 9.9 +/- 0.9
Body weight of live pups (g) (mean +/- SD) on day 4 Males Females	6.0 +/- 0.4 9.0 +/- 1.3	5.9 +/- 0.4 9.2 +/- 1.1	5.8 +/- 0.3 8.8 +/- 0.5	6.0 +/- 0.5 9.5 +/- 1.0
Viability index	98.15	94.00	81.88	93.48
No. of external anomalies	0	0	0	0

Gestation index = (Number of dams with live newborns/Number of pregnant females) x 100

Birth index = (Number of newborns/Number of implantations) x 100

Viability index = (Number of live newborns on day 4 after birth/Number of live newborns) x 100

*: P<0.05 significantly different from control

Background level of stillborn : 0-14.84%

Background level of birth index : 80.98-96.61%

Applicant's summary and conclusion

Conclusions:

The NOAEL is considered to be 300 mg/kg/day for reproductive performance of parents and for development of offspring.

Executive summary:

In a combined repeated dose toxicity/reproduction and developmental toxicity study tetrahydromethylphthalic anhydride (MTHPA, 99.7%) was administered to groups of Crj:CD(SD) rats (12/sex), via gavage in corn oil at dose levels of 0 (Vehicle), 30, 100 and 300 mg/kg bw/day.

As for reproductive performance, no effects related to the test article were observed on the estrous cycle, numbers of corpora lutea and implantations, copulation index or fertility indices. Examination at delivery and during the lactation period revealed, no effects related to the test article in terms of gestational days, litter size and live newborns, gestation index, stillborn index, birth index, sex ratio, body weights of offspring at birth and at day 4 after birth, or viability index on day 4. No external anomalies were apparent. The NOAEL is considered to be 300mg/kg/day for reproductive performance of parents and for development of offspring.

This study is acceptable and satisfies the guideline requirement for a combined repeated dose toxicity/reproduction and developmental toxicity study in rats (OECD 422).