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Toxicological information

Toxicity to reproduction

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Administrative data

Endpoint:
screening for reproductive / developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2001
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study
Cross-reference
Reason / purpose for cross-reference:
reference to same study

Data source

Referenceopen allclose all

Reference Type:
study report
Title:
Unnamed
Year:
2002
Report date:
2002
Reference Type:
publication
Title:
[Combined Repeated Dose and Reproductive/Developmental Toxicities of Benzoyl Peroxide]
Author:
Song S, Kim SH, Bae H, Kim M, Koo HJ, Park K, Lee S, Park J, Choi ES, and Lee MS
Year:
2003
Bibliographic source:
Journal of Toxicology and Public Health, 19(2):123-31

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)
GLP compliance:
yes
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
Dibenzoyl peroxide
EC Number:
202-327-6
EC Name:
Dibenzoyl peroxide
Cas Number:
94-36-0
Molecular formula:
C14H10O4
IUPAC Name:
benzoyl benzenecarboperoxoate
Details on test material:
Source - SIGMA, purity = 71.8 percent in water, white powder

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals or test system and environmental conditions:
- Age : 7 weeks old for males and females, strains were obtained from Biogenomics Limited
- Weight at study initiation : 323.38 - 356.63 g for males, 212.21 - 237.18 g for females
- Temperature: 18.25-25.69 °C
- Humidity: 51.09 - 73.75 %
- 12-hour dark cycle
- 15-15 room air change per hour
- Water and diet ad libitum

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
corn oil
Details on mating procedure:
- M/F ratios per cage ; 1/1
- Proof of pregnancy: sperm detection in vagina
Analytical verification of doses or concentrations:
not specified
Details on analytical verification of doses or concentrations:
no data
Duration of treatment / exposure:
Dosing schedule :
- males: premating period of 2 weeks, then mating period for 15 days, total of 29 days
- females: premating period of 2 weeks then mating, pregnant and 3 days lactation period , total of 41 ~ 51 days
Frequency of treatment:
daily
Doses / concentrations
Remarks:
Doses / Concentrations:
0, 250, 500, 1,000 mg/kg b.w/day
Basis:
actual ingested
No. of animals per sex per dose:
10 animals/sex/dose (5 animals/sex/positive control)
Control animals:
yes, concurrent vehicle
Details on study design:
- Parameters assessed during study P and F1 : copulation index (matings/pairs x 100), fertility index (Pregnancies/matings x 100), gestation index (live litters/pregnancies x 100), viability index (No. of live offspring at day1 or day3/No. of live offspring at birth × 100), body weight of live pups (on day 0 and 4), No. of corpora lutea, No. of female mated, abortion, premature birth, gestation period, sex ratio (Total No. of male pups/Total No. of female pups)
Positive control:
Yes: Cyclophosphamide (4.5 mg/kg/day)

Examinations

Parental animals: Observations and examinations:

MORTALITY: observation once daily

DETAILED CLINICAL OBSERVATIONS AND BODY WEIGHT: Yes
Clinical observations performed and frequency : Clinical signs were observed twice a day over the administration period, and once daily during the weekend. Changes in skin, hair, eyes, mucous membrane, respiratory system, autonomic nervous system, central nervous system, mobility and behavioral patterns were recorded with careful attention to tremor, seizure, ptyalorrhea, diarrhea, drowsiness, stupor or coma. Body weight were observed once a week respectively.

Moreover, haematology and biochemistry for males only at the time of necropsy after 28 days of chemical exposure (detailed in 7.5.1)
Oestrous cyclicity (parental animals):
no data
Sperm parameters (parental animals):
- Sperm examination : concentration, motility, morphology was observed at necropsy.
Litter observations:
Sex ratio (Total No. of male pups/Total No. of female pups) was assessed, body weight of live pups (on day 0 and 4)
Body weight of neonates was recorded on day 1 pp and on day 3 pp.
All living neonates were subjected to an external examination within 24 hours after birth.
Postmortem examinations (parental animals):
- Organs examined at necropsy (macroscopic and microscopic) :
Organ weight : liver, kidney, adrenal gland, testis, epididymis, thymus, spleen, brain, heart, ovary, uterus, thyroid gland, spermary, prostrate gland. - Microscopic : brain, spinal cord, stomach, pancreas, jejunum, ileum, cecum, colon, rectum, liver, kidney, adrenal gland, spleen, heart, thymus, thyroid, bronchus, lung, pituitary gland, ovary, uterus, vagina, testis, epididymis, spermary, prostrate gland, mammary gland, bladder, nodi lymphatici mesenterici, nodi lymphatici, nervus ischiadicus, femoral marrow and all gross lesion.
Postmortem examinations (offspring):
Grossly visible abnormalities; external, soft tissu and skelettal abnormalities
Statistics:
Parametric data were analyzed using One-way Analysis of variance. If significance was detected, Dunnett¿s multiple comparison test was used to detect significant difference between the negative controls and treatment groups. One-sided Student¿s t-test was used to detect significant difference between the negative controls and the positive controls.
Non-parametric data were analyzed using Kruskal-Wallis analysis, and Dunn¿s multiple comparison test was conducted using ranked data.
Categorized data were analyzed using descriptive analysis or Two-sided Fisher¿s exact test.
All differences were considered significant at 5%.
Reproductive indices:
Parameters assessed during study P and F1 : copulation index (matings/pairs x 100), fertility index (Pregnancies/matings x 100), gestation index (live litters/pregnancies x 100), No. of corpora lutea, No. of female mated, abortion, premature birth, gestation period,
Offspring viability indices:
Viability index (No. of live offspring at day1 or day3/No. of live offspring at birth × 100),

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
minimal testes degeneration in 5 of 9 rats, seen on only 1/5 rat on the satellite group, and not associated with gross necropsy findings, testicular atrophy and decrease of fertility.
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:
effects observed, treatment-related
Other effects:
effects observed, treatment-related
Description (incidence and severity):
Test substance intake: increase of food consumption during the 4 first weeks only in the highest dose group

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:
not examined
Reproductive function: sperm measures:
not examined
Reproductive performance:
no effects observed

Details on results (P0)

REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS)
There was no significant difference between experimental and control groups in the precoital time, copulation, fertility and gestation rate.

CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS)
No deaths were found in all animals including control groups. Body weight : Body weight gains were slightly increased (approximately 7 % more than controls) in female recovery group exposed to 1,000 mg/kg only after 29 days.

BODY WEIGHT AND FOOD CONSUMPTION (PARENTAL ANIMALS)
No significant body weight changes were observed in all treatment animals except highest dose recovery group. Low body weight gain of pups showed in 1,000 mg/kg dose.
Food/water consumption : The mean food consumption of 1,000 mg/kg was significantly increased at first and fourth weeks of chemical exposure in male. No other treatment groups were showed abnormal food consumption.

ORGAN WEIGHTS (PARENTAL ANIMALS)
Significant decrease in absolute and relative weights of only left testes and relative wt of epididymis was observed in ONE male exposed to 1000 mg/kg (p < 0.05) and is considered as sporadic effect as only seen on left testis.


HISTOPATHOLOGY (PARENTAL ANIMALS)
Severe testis atrophy was seen in one male of the highest exposure male. In the 1,000 mg/kg treatment group,minimal teste degeneration was observed in several animals of this high dose-group but these changes were not seen in recovery animals. Moreover there is no effect on fertility. No graduation of the severity of this effect is given. this effect has to be noted but is not considered as adverse. In the female reproductive organ, slight affects were observed in one uterus of female exposed to 1,000 mg/kg is not considered as tratement-related. See table 4

CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS): none


GROSS PATHOLOGY (PARENTAL ANIMALS): no effects


Effect levels (P0)

open allclose all
Dose descriptor:
NOAEL
Remarks:
toxicity
Effect level:
500 mg/kg bw/day
Based on:
test mat.
Sex:
male
Basis for effect level:
histopathology: non-neoplastic
Dose descriptor:
NOAEL
Remarks:
toxicity
Effect level:
>= 1 000 mg/kg bw/day
Sex:
female
Remarks on result:
not determinable due to absence of adverse toxic effects
Dose descriptor:
NOAEL
Remarks:
fertility
Effect level:
>= 1 000 mg/kg bw/day
Based on:
act. ingr.
Sex:
male/female
Remarks on result:
not determinable due to absence of adverse toxic effects

Target system / organ toxicity (P0)

Critical effects observed:
not specified

Results: F1 generation

General toxicity (F1)

Clinical signs:
no effects observed
Mortality / viability:
not specified
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
decrease of body weight gain at day 3 post-partum
Sexual maturation:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
not examined
Histopathological findings:
not examined

Details on results (F1)

BODY WEIGHT (OFFSPRING)
The pups exposed to highest levels of benzoyl peroxide showed significant decrease in body weight gain (p < 0.01). The body weight gain of pups on the day 3 after birth decreased at 1,000 mg/kg. Benzoyl peroxide has adverse effects on development of pups with high birthrate of runts at 1000 mg/kg dose level. No significant differences appeared between the treatment and control group in any other observation. See tables 2 and 3.


Effect levels (F1)

Dose descriptor:
NOAEL
Generation:
F1
Effect level:
500 mg/kg bw/day
Based on:
act. ingr.
Sex:
male/female
Basis for effect level:
body weight and weight gain

Target system / organ toxicity (F1)

Critical effects observed:
not specified

Overall reproductive toxicity

Reproductive effects observed:
not specified

Any other information on results incl. tables

Table 1

DOSE : (mg/kg)        0     250    500    1000    Cyclophos
phamide
No. of mated males 10 10 10 10 10
Copulation index (%) 100 100 100 100 100 (5/5)
Fertility index (%) 100 100 100 90 100 (5/5)
                                         (9/10)
No. of mated females 10 10 10 10 10
Copulation index (%) 100 100 100 100 100 (5/5)
Fertility index (%) 100 100 100 90 100 (5/5)
                                         (9/10)
Gestation index (%) 100 100 100 100 40 (2/5)
No. of dams 10 10 10 9 2
No. of corpora lutea 169 182 171 157 90
Mean 16.9 18.2 17.1 17.4 18
+-S.D +-4.1 +-2.3 +-1.9 +-2.5 +-1.9
No. of implantations 150 150 161 136 81
Mean 15.0 15.0 16.1 15.1 16.2
+-S.D +-1.8 +-2.9 +-1.9 +-2.1 +-1.6
Mean percent 9 16.1 5.8 12.9 9.9
preimplantation loss
No. of embryo/fetal 10/3 9/3 7/3 3/0 64/9
death
No. of live pups born
Mean 13.7 13.8 15.1 14.8 1.6***
+-S.D +-2.7 +-2.5 +-2.1 +-2.2 +-2.3
Mean pregnancy 20.4 20.8 20.6 20.8 22
period (day)
Viability index 97.5 98.1 100 100 53.3***
on day 1 pp
Viability index 100 100 100 100 90*
on day 3 pp
Body weights
of pups (g)
Male 1 DAY mean 6.88 7.22 6.91 6.72 4.59
+-S.D +-0.53 +-0.81 +-0.64 +-0.79 +-0.77
Male 3 DAY mean 8.93 9.10 8.69 8.12*** 7.16
+-S.D +-0.91 +-1.03 +-0.79 +-1.01 +-0.88
Female 1 DAY mean 6.51 6.93***6.73 6.21 4.33
+-S.D +-0.53 +-0.76 +-0.59 +-0.79b +-0.74
Female 3 DAY mean 8.60 8.79 8.48 7.49*** 5.48
+-S.D +-0.99 +-1.03 +-0.89 +-1.03 +-0.89
Precoital time
Mean (day) 2.2 3.2 2.4 2.3 5.3
+-S.D +-1.2 +-1.3 +-1.2 +-1.2 +-5.3
Number of females showing evidence of copulations was decreased at 1000 mg/kg but this effect has not to be taken into account, since a severe atrophy of left teste appeared in only one male at 1000 mg/kg (male 80) and since this male was
mated with female 35 that was not later pregnant. The atrophy of left teste can not be related to BPO treatment since absolute no effect is observed on right teste, including no histhopatological effect.

Table 2:
Grossly visible abnormalities, external, soft tissue and skeletal abnormalities:
In the 1000 mg/kg dose group, the incidence of runts was significantly higher then control group.

Treatment      0     250    500    1000   Cyclophosphamide
(mg/kg/day)
No. with major 0 1 0 0 0
abnormalities
Mean % of pups 0 0.6 0 0 0
examined
No. of females 0 1 0 0 0
affected
No. with minor 20 20 18 60 8
abnormalities
Mean % of pups 14.2 12.4 11.5 41.4*** 100.0***
examined
No. of females 6 4 5 6 2
affected
No. with 0 0 0 0 0
variants
*** Statistically significant difference from control group (p
< 0.001)

Applicant's summary and conclusion

Conclusions:
The NOAEL for reproduction toxicity in male rats is 500 mg/kg based on reduced weight of testes and epididymis at 1,000 mg/kg. NOAEL for developmental toxicity is also 500 mg/kg based on significant decreased weight gain of pups with high birthrate of runt at 1,000 mg/kg.
Executive summary:

A combined repeated dose and reproduction/developmental toxicity screening test (OECD TG 422) was conducted using Sprague-Dawley rats (NIER, Korea, 2001). Benzoyl peroxide was administered to rats by gavage at doses of 0, 250, 500 and 1,000 mg/kg/day. Males were dosed for 29 days and females were dosed for 41 - 51 days from 14 days before mating to day 3 of lactation

throughout the mating and pregnancy period. No treatment-related changes in precoital time and rate of copulation, fertility and gestation were noted in any benzoyl peroxide treated groups. Minimal symptoms such as vacuolation or hyperplasia, were seen in 1000 mg/kg group, but this is not considered to have been related to benzoyl peroxide treatment. Adverse effects on reproduction were shown at the highest dose of 1,000 mg/kg/day in male rats with the reduction of reproductive organ¿s weight and slight testes degenerations. In female rats, no adverse effects were observed during the test period. The NOAEL for reproduction toxicity in male rats was 500 mg/kg/day. No variants were found. High birthrate of runt was seen and body weight gain of pups was significantly decreased (male 9 % ; female 12.9 % of control wt) at dose of 1,000 mg/kg/day. The study concluded that Benzoyl peroxide has adverse effects on development of pups with high birthrate of runt at 1000 mg/kg dose level. The NOAEL for developmental toxicity was determined as 500 mg/kg/day.