4-methoxybenzyl alcohol

Toxicity to reproduction

Administrative data

Endpoint:

screening for reproductive / developmental toxicity

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

4 (not assignable)

Rationale for reliability incl. deficiencies:

secondary literature

Justification for type of information:

Data from secondary source

Data source

Reference

Reference Type:

secondary source

Title:

SIDS Initial Assessment Profiles for test material

Author:

Organization for Economic Co-operation and Development,

Year:

2012

Bibliographic source:

SIDS Initial Assessment Profiles for test material ,2012

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

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

Principles of method if other than guideline:

Repeated dose oral toxicity study with the reproduction / developmental toxicity screening test (OECD TG 422) of test material was conducted on rats for a period of 42 days

GLP compliance:

not specified

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Anisaldehyde

EC Number:

204-602-6

EC Name:

Anisaldehyde

Cas Number:

123-11-5

Molecular formula:

C8H8O2

IUPAC Name:

4-Methoxybenzaldehyde

Details on test material:

- Name of test material (as cited in study report):p-methoxybenzaldehyde
- Molecular formula :C8H8O2
- Molecular weight:136.149 g/mol
- Substance type:Organic
- Physical state:Liquid

Test animals

Species:

rat

Strain:

Crj: CD(SD)

Sex:

male/female

Details on test animals or test system and environmental conditions:

- Source: Charles River Japan Inc.
- Age at study initiation: 9 weeks
- Weight at study initiation: Range of average, 381.8-385.1g for males, 236.5-237.9g for females
- Fasting period before study: No
- Housing: One animal/cage
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 2 weeks


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 23-25°C
- Humidity (%): 50-65%
- Air changes (per hr): 15 times/hour
- Photoperiod (hrs dark / hrs light): 12 hrs dark/12 hrs light

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

corn oil

Details on exposure:

PREPARATION OF DOSING SOLUTIONS: The test substance was dissolved with corn oil at concentration of 0.4-10%.


VEHICLE
- Justification for use and choice of vehicle (if other than water): Solution
- Concentration in vehicle: 0.4-10%
- Amount of vehicle (if gavage): 5 ml/kg bw

Details on mating procedure:

- M/F ratio per cage:1:1
- Length of cohabitation: 2 weeks
- Proof of pregnancy: [vaginal plug / sperm in vaginal smear] referred to as [day 0 / day 1] of pregnancy:Vaginal plug/sperm in vaginal smear] referred to as day 0 of pregnancy
- After ... days of unsuccessful pairing replacement of first male by another male with proven fertility.
- Further matings after two unsuccessful attempts: [no / yes (explain)]
- After successful mating each pregnant female was caged (how): One female/cage with bedding to nest
- Any other deviations from standard protocol:

Analytical verification of doses or concentrations:

yes

Duration of treatment / exposure:

For males: Male rats were dosed for 42 days from 14 days before mating.
For females: Female rats were dosed from 14 days before mating to day 4 of lactation throughout the mating and pregnancy period.

Frequency of treatment:

once a day

Details on study schedule:

- F1 parental animals not mated until 2 weeks after selected from the F1 litters.
- Selection of parents from F1 generation when pups were 4 days of age.
- Age at mating of the mated animals in the study:9 weeks

Doses / concentrations

Remarks:

0, 20, 100, or 500 mg/kg bw/day

No. of animals per sex per dose:

Total:104
0 mg/kg:13male and 13 female
20 mg/kg:13male and 13 female
100 mg/kg:13male and 13 female
500 mg/kg:13male and 13 female

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: Based on a preliminary study, in which 9 weeks old male and female rats were given 4-methoxybenzaldehyde at a dose of 0, 250, 500 or 1000mg/kg for 14 days.
- Rationale for animal assignment (if not random): Random

Positive control:

No Data

Examinations

Parental animals: Observations and examinations:

Parental animals observation and examinations
AGE SIDE OBSERVATIONS: Yes
- Time schedule: Once a day

DETAILED CLINICAL OBSERVATIONS: No

BODY WEIGHT: Yes
- Time schedule for examinations: Body weights were determined on days 1 (before dosing), 7, 14, 21, 28, 35, 42 and 43 (at autopsy) in males and on days 1 (before dosing), 7 and 14 in all females, on days 21 and 28 in some females, on days 0, 7, 14 and 20 of gestation in pregnant females, and on days 0, 4 and 5 (at autopsy) of lactation in females that delivered, and day 25(at autopsy) of gestation in females that had not delivered.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: No
Food consumption was determined for days 1-2, 7-8, 13-14, 29-30, 35-36 and 41-42 of the administration period in males, for days 1-2 and 7-8 in all females, for days 0-1, 7-8, 14-15 and 20-21 of gestation in pregnant females, and for days 0-1 and 3-4 of lactation in females that delivered.

Oestrous cyclicity (parental animals):

Yes, Estrous cycle was observed from 14 days before mating to day of copulation

Sperm parameters (parental animals):

Testes and epididymides weights were determined.

Litter observations:

STANDARDISATION OF LITTERS
- Performed on day 4 postpartum: No

PARAMETERS EXAMINED
The following parameters were examined in F1 offspring: For F1 offspring, number and sex of pups, stillbirths, live births, postnatal mortality, presence of gross anomalies, weight gain were observed.


GROSS EXAMINATION OF DEAD PUPS: Yes, for external and internal abnormalities.

Postmortem examinations (parental animals):

Postmortem examinations (SACRIFICE
Males and females were sacrificed at the end of the observation period; on day 43 of the administration period and on day 5 of the lactation period, respectively.

GROSS NECROPSY
Gross necropsy consisted of external and internal examinations including the cervical, thoracic, and abdominal viscera.

ORGAN WEIGHTS
Weights of organs such as brain, thymus, heart, liver, spleen, kidneys, adrenals, testes and epididymides in males, and brain, thymus, heart, liver, spleen, kidneys and adrenals in females were determined at necropsy.

HISTOPATHOLOGY
Histopathological examination of brain, pituitary, spinal cords, alimentary canal, liver, kidneys, adrenals, spleen, pancreas, heart, thymus, thyroid, trachea, lung, urinary bladder, mesentery lymph node, submandibular lymph node, sciatic nerve, femur bone marrow, seminal vesicle, prostate, vagina, ovaries, uterus, testes and epididymides were performed in 5 males and pregnant females in the 0 and 500 mg/kg bw/day groups. In addition, testes and epididymides in males and ovaries in females in the 20 and 100 mg/kg bw groups and liver and stomach in males and females and heart in males in the 20 and 100 mg/kg bw groups were histopathologically examined.

Postmortem examinations (offspring):

SACRIFICE
- All dead pups (F1) were necropsied. All live pups(F1) were sacrificed under anesthesia to examine for viseceral anomalies on day 4 of the lactation period.

GROSS NECROPSY
- Gross necropsy consisted of external and internal examinations including the cervical, thoracic, and abdominal viscera.


HISTOPATHOLOGY / ORGAN WEIGTHS: Not examined

Statistics:

Statistical analysis was conducted using the Dunnett's test, Bartlett's test, Mann-Whitney U- test and Fisher' direct test.

Reproductive indices:

Reproductive and developmental parameters:
No. of pairs mated, no. of pairs copulated, copulation index (no. of copulated pairs/no. of mated pairs x 100), no. of pregnant females, fertility index (no. of pregnant animals/no. of animals with successful copulation x 100), estrous cycle, frequency of vaginal estrus, no. of pregnant females with live pups, gestation index (no. of females with live pups/no. of pregnant females x 100), gestation length, no. of corpora lutea, implantation index (no. of implantation sites/no. of corpora lutea x 100), no. of pups born, deliver index (no. of pups born/no. of implantation sites x 100)

Offspring viability indices:

No. of live pups on day 0 of lactation, birth index (no. of live pups on day 0/no. of implantation sites x 100), live birth index (no. of live pups on day 0/no. of pups born x 100), pups weight on day 0 of lactation, sex ratio of pups on day 0 of lactation, no. of live pups on day 4 of lactation, viability of pups on day 4 of lactation, pups weight on day 4 of lactation, and sex ratio of pups on day 4 of lactation were determined. All pups born were examined for external anomalies at birth and all live pups were sacrificed under anesthesia to examine for visceral anomalies on day 4 after birth.

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:

no effects observed

Description (incidence and severity):

Temporary salivation after administration was observed in males and females in the 500 mg/kg bw group.

Dermal irritation (if dermal study):

not specified

Mortality:

no mortality observed

Description (incidence):

No animal died in any group.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Body weight tended to be increased in males and females in the 100 and 500 mg/kg bw groups.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

Increase in food consumption was observed in males of the 500 mg/kg bw group and females in the 100 and 500 mg/kg bw groups

Food efficiency:

not specified

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

not specified

Ophthalmological findings:

not specified

Haematological findings:

not specified

Clinical biochemistry findings:

not specified

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:

effects observed, treatment-related

Description (incidence and severity):

Hyperplasia of squamous epithelium was detected in males and females in the 100 and 500 mg/kg bw groups. In liver, centrilobular hypertrophy of hepatocytes was detected in males of the 500mg/kg bw group.

Histopathological findings: neoplastic:

not specified

Other effects:

not specified

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:

no effects observed

Description (incidence and severity):

The compound showed no adverse effects in terms of estrous cycle

Reproductive function: sperm measures:

not specified

Reproductive performance:

effects observed, treatment-related

Description (incidence and severity):

In the 500 mg/kg bw group, the number of non-pregnant females was increased whereas all pairs copulated, and the fertility index was reduced.The delivery index was lower than in controls at 500 mg/kg bw/day.The compound showed no adverse effects number of corpora lutea and implant rate at any dose level.

Effect levels (P0)

Dose descriptor:

NOAEL

Effect level:

100 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

clinical signs

mortality

body weight and weight gain

food consumption and compound intake

organ weights and organ / body weight ratios

gross pathology

histopathology: non-neoplastic

reproductive performance

Remarks on result:

other: No toxic effects observed at given dose level

Target system / organ toxicity (P0)

Critical effects observed:

not specified

System:

other: not specified

Organ:

not specified

Treatment related:

not specified

Dose response relationship:

not specified

Relevant for humans:

not specified

Results: F1 generation

General toxicity (F1)

Clinical signs:

not specified

Dermal irritation (if dermal study):

not specified

Mortality / viability:

mortality observed, treatment-related

Description (incidence and severity):

In the 500 mg/kg bw group, the number of pups, the delivery index and the number of live pups were lower than in the controls.(

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

A decrease in body weight on day 0 of lactation was observed in the 500 mg/kg bw group.

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 specified

Haematological findings:

not specified

Clinical biochemistry findings:

not specified

Urinalysis findings:

not specified

Sexual maturation:

no effects observed

Organ weight findings including organ / body weight ratios:

not specified

Gross pathological findings:

no effects observed

Histopathological findings:

not specified

Other effects:

not specified

Developmental neurotoxicity (F1)

Behaviour (functional findings):

not specified

Developmental immunotoxicity (F1)

Developmental immunotoxicity:

not specified

Effect levels (F1)

Dose descriptor:

NOAEL

Generation:

F1

Effect level:

100 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

not specified

Basis for effect level:

viability

sexual maturation

clinical signs

mortality

body weight and weight gain

gross pathology

Remarks on result:

other: overall no developmental toxic effects observed

Target system / organ toxicity (F1)

Critical effects observed:

not specified

System:

other: not specified

Organ:

not specified

Treatment related:

not specified

Dose response relationship:

not specified

Relevant for humans:

not specified

Overall reproductive toxicity

Reproductive effects observed:

not specified

Treatment related:

not specified

Relation to other toxic effects:

not specified

Dose response relationship:

not specified

Relevant for humans:

not specified

Any other information on results incl. tables

Reproductive performance

 

Compound dose Mg/kg bw	0	20	100	500
Mean length of estrous cycle in days Mean SD	4.3 0.6	4.0 0.0	4.2 0.3	4.3 0.4
Number of mated pairs	13	13	13	13
Number of copulated pairs	13	13	13	13
Copulation index(%)	100	100	100	100
Number of pregnant animals	12	12	12	6
Fertility index	92.3	92.3	92.3	46.2*
Pairing days until copulation Mean SD	2.7 1.1	2.5 1.2	2.7 1.2	2.9 2.5
Frequency of vaginal estrous Mean SD	1.0 0.0	1.0 0.0	1.0 0.0	1.1 0.3

*:p<0.05 (by Fisher's direct test)

 

Summary of development of pups

 

Compound dose Mg/kg bw	0	20	100	500
Number of pregnant females	12	12	12	6
Number of pregnant females with pups alive	12	12	12	5
Gestation index (%)	100	100	100	83.3
Day 0 of lactation
Number of pups born Mean SD	14.0(12) 4.5	14.3(12) 1.4	11.8(12) 4.3	9.3(6)* 5.2
Delivery index(%) Mean SD	94.7(12) 5.4	92.2(12) 9.5	90.9(12) 8.0	66.5(6)** 33.9
Number of pups alive Mean SD	13.8(12) 4.8	14.2(12) 1.6	11.8(12) 4.3	10.6(6)* 2.5
Birth index (%) Mean SD	85.3(12) 27.9	91.6(12) 9.7	90.9(12) 8.0	76.0(6) 13.0
Live birth index (%) Mean SD	90.5(12) 28.8	99.3(12) 2.4	100.0(12) 0.0	95.4(6) 10.3
Pups weight (g)
Male Mean SD	7.0(11) 0.7	7.0(12) 0.6	7.4(12) 1.2	6.6(5) 0.7
Female Mean SD	6.5(11) 0.6	6.6(12) 0.4	7.0(12) 0.9	6.1(5) 0.6
Sex ratio (male/female) on day 0 (%) Mean SD	86(11) 54	116(12) 73	142(12) 110	110(5) 71
Day 4 of lactation
Number of pups alive Mean SD	14.8(11) 2.2	14.2(12) 1.6	11.8(12) 4.3	10.2(5)* 2.8
Viability index (%) Mean SD	98.7(11) 3.0	100.0(12) 0.0	100.0(12) 0.0	96.0(5)* 8.9
Pups weight (g)
Male Mean SD	11.3(11) 1.3	11.3(12) 1.4	12.7(12) 3.0	11.3(5) 1.3
Female Mean SD	10.8(11) 1.1	10.9(12) 1.1	12.3(12) 2.8	11.1(5) 0.9
Sex ratio (male/female) on day 4 (%) Mean SD	92(11) 69	116(12) 73	142(12) 110	102(5) 74

 

*:p<0.05; **:p<0.01

Applicant's summary and conclusion

Conclusions:

The No Observed Adverse Effect Level (NOAEL) for reproductive toxicity in rats was considered to be 100 mg/kg bw/day. When male and female rats were treated with test material orally.

Executive summary:

In the combined repeated dose toxicity study with the reproduction/developmental toxicity screening test (OECD TG 422), the male and femaleCrj:CD(SD)IGSrats were given test material by gavage at 0, 20, 100, or 500 mg/kg bw/day for 42 days beginning 14 days before mating in males, and from 14 days before mating to day 4 of lactation throughout the mating and pregnancy period in females.The test substance was dissolved with corn oil and adminstered in dose volume 5ml/kg bw once daily . 13 animals/sex/dose were used. Body weights were determined on days 1 (before dosing), 7, 14, 21, 28, 35, 42 and 43 (at autopsy) in males and on days 1 (before dosing), 7 and 14 in all females, on days 21 and 28 in some females, on days 0, 7, 14 and 20 of gestation in pregnant females, and on days 0, 4 and 5 (at autopsy) of lactation in females that delivered, and day 25(at autopsy) of gestation in females that had not delivered. Food consumption was determined for days 1-2, 7-8, 13-14, 29-30, 35-36 and 41-42 of the administration period in males, for days 1-2 and 7-8 in all females, for days 0-1, 7-8, 14-15 and 20-21 of gestation in pregnant females, and for days 0-1 and 3-4 of lactation in females that delivered. For F1 offspring, number and sex of pups, stillbirths, live births, postnatal mortality, presence of gross anomalies, weight gain were observed.Males and females were sacrificed at the end of the observation period; on day 43 of the administration period and on day 5 of the lactation period, respectively.

Histopathological examination of brain, pituitary, spinal cords, alimentary canal, liver, kidneys, adrenals, spleen, pancreas, heart, thymus, thyroid, trachea, lung, urinary bladder, mesentery lymph node, submandibular lymph node, sciatic nerve, femur bone marrow, seminal vesicle, prostate, vagina, ovaries, uterus, testes and epididymides were performed in 5 males and pregnant females in the 0 and 500 mg/kg bw/day groups. In addition, testes and epididymides in males and ovaries in females in the 20 and 100 mg/kg bw groups and liver and stomach in males and females and heart in males in the 20 and 100 mg/kg bw groups were histopathologically examined. All dead pups (F1) were necropsied. All live pups(F1) were sacrificed under anesthesia to examine for viseceral anomalies on day 4 of the lactation period.Estrous cycle was observed from 14 days before mating to day of copulation

No animal died in any group. Temporary salivation after administration was observed in males and females in the 500 mg/kg bw group. Body weight tended to be increased in males and females in the 100 and 500 mg/kg bw groups. Increase in food consumption was observed in males of the 500 mg/kg bw group and females in the 100 and 500 mg/kg bw groups. Hyperplasia of squamous epithelium was detected in males and females in the 100 and 500 mg/kg bw groups. In liver, centrilobular hypertrophy of hepatocytes was detected in males of the 500mg/kg bw group.

No effects on Estrous cycle was observed. In the 500 mg/kg bw group, the number of non-pregnant females was increased whereas all pairs copulated, and the fertility index was reduced. Epididymidal weight was decreased in males of the 500 mg/kg bw group. Dark spots in the mucosa of the stomach and nodes in the epididymis were observed by one animal each in the 500 mg/kg bw group. In the 500 mg/kg bw group, the number of pups, the delivery index and the number of live pups were lower than in the controls. A decrease in body weight on day 0 of lactation was observed in the 500 mg/kg bw group.No effects on sexual maturation and gross pathology.The No Observed Adverse Effect Level (NOAEL) for reproductive toxicity in rats was considered to be 100 mg/kg bw/day.