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Administrative data

Key value for chemical safety assessment

Effects on fertility

Link to relevant study records
Reference
Endpoint:
two-generation reproductive toxicity
Remarks:
based on test type (migrated information)
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Experimental result published in peer reveiwed journal. Some details on methods / results not available
Qualifier:
according to
Guideline:
OECD Guideline 416 (Two-Generation Reproduction Toxicity Study)
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
other: Crj:CD (SD) IGS
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Tsukuba breeding Center Charles River japan Inc
- Age at study initiation: (P) x wks; (F1) x wks: P 4 weeks
- Weight at study initiation: (P) Males: x-x g; Females: x-x g; (F1) Males: x-x g; Females: x-x g
- Fasting period before study: no
- Housing: individually except during acclimatisation (1 or 2 animals same sex)
- Use of restrainers for preventing ingestion (if dermal): yes/no: N/A
- Diet (e.g. ad libitum): NIH-07M Clea Japan Inc Tokyo Japan
- Water (e.g. ad libitum): tap water
- Acclimation period: 1 week

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 23+/- 3degrees C
- Humidity (%): 50 +/- 20
- Air changes (per hr): 10-15
- Photoperiod (hrs dark / hrs light): 12

Route of administration:
oral: feed
Vehicle:
other: basal diet
Details on exposure:
DIET PREPARATION
- Mixing appropriate amounts with (Type of food): basal NIH-07M diet

VEHICLE
- Justification for use and choice of vehicle (if other than water): dietary study; in addition DEP is poorly soluble in water
- Concentration in vehicle: 600, 3000 & 5000 ppm
Details on mating procedure:
- M/F ratio per cage: 1:1
- Length of cohabitation: until the day of sucessful copulation
- Proof of pregnancy: [vaginal plug / sperm in vaginal smear] referred to as [day 0 / day 1] of pregnancy: vaginal plug/sperm; day 0
- After ... days of unsuccessful pairing replacement of first male by another male with proven fertility. Not reported
- Further matings after two unsuccessful attempts: [no / yes (explain)]: procedure repeated until pregnancy resulted for 3 week mating period
- After successful mating each pregnant female was caged (how): singly
- Any other deviations from standard protocol: not reported
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Not reported
Duration of treatment / exposure:
Approximately 15 weeks for male and 17 weeks for female parents of the F0 & F1 generations.
Frequency of treatment:
Continuously via the diet
Details on study schedule:
- F1 parental animals not mated until 10 weeks after selected from the F1 litters.
- Selection of parents from F1 generation when pups were 21-25 days of age.
- Age at mating of the mated animals in the study: 15 weeks
Remarks:
Doses / Concentrations:
0 (control) 600, 3000 & 15000 ppm
Basis:
nominal in diet
No. of animals per sex per dose:
24
Control animals:
yes, plain diet
Details on study design:
- Dose selection rationale: selected on basis of results of a preliminary study
- Rationale for animal assignment (if not random): assigned on the basis of weight so that the 4 groups had similar group mean and SD values
Positive control:
No
Parental animals: Observations and examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily for clinical signs & mortality

DETAILED CLINICAL OBSERVATIONS: No data
- Time schedule:Not reported

BODY WEIGHT: Yes
- Time schedule for examinations: weekly

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: Yes
Oestrous cyclicity (parental animals):
yes,see table below
Sperm parameters (parental animals):
Sperm parameters were measured in all F0 & F1 males surviving to scheduled terminal kill. The right cauda epididymis was weighed & used for sperm analysis. Right testis was also used used for counting homogenisation resistant testicular spermatids.
Litter observations:
STANDARDISATION OF LITTERS
- Performed on day 4 postpartum: yes
- If yes, maximum of 8 pups/litter (4/sex/litter as nearly as possible); excess pups were killed and discarded.

PARAMETERS EXAMINED
The following parameters were examined in [F1 / F2 / F3] offspring:
[number and sex of pups, stillbirths, live births, postnatal mortality, presence of gross anomalies, weight gain, physical or behavioural abnormalities, other:] seee table below

GROSS EXAMINATION OF DEAD PUPS:
[no / yes, for external and internal abnormalities; possible cause of death was/was not determined for pups born or found dead.] yes gross examination
Postmortem examinations (parental animals):
SACRIFICE
- Male animals: All surviving animals [describe when, e.g. as soon as possible after the last litters in each generation were produced.]: after weaning of their F1 & F2 offspring as appropriate
- Maternal animals: All surviving animals [describe when, e.g. after the last litter of each generation was weaned.]: after weaning of their F1 & F2 offspring as appropriate

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

HISTOPATHOLOGY / ORGAN WEIGHTS
Selected tissues were prepared for microscopic examination and weighed, respectively.
Postmortem examinations (offspring):
SACRIFICE
- The F1 offspring not selected as parental animals and all F2 offspring were sacrificed at [26] days of age.
- These animals were subjected to postmortem examinations (macroscopic and/or microscopic examination) as follows: macroscopic & microscopic examination

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

HISTOPATHOLOGY / ORGAN WEIGTHS
Selected tissues were prepared for microscopic examination and weighed, respectively.For control & 15000 ppm groups all F0 & F1 parental animals were examined for abnormalities of the reproductive organs. In the low dose group the same organs were examined in female animals with abnormal oestrus cycles, males with sperm abnormality & pairs of animals that failed to mate or produce offspring. In addition the kidneys of F1 parental females in control & 15000 groups were examined due to the significant increase in weight noted when compared with controls.
In F1& F2 weanlings the thymus of 6 of each of the males & females & the spleen of 6 F2 males in the control & 15000 ppm groups were examined as the weights were significantly decreased in the 15000 group compared with controls.
Statistics:
For body weights, body weight gain, food consumption, oestrus cyle length, number of implants & pups delivered, delivery index sperm parameters ormone levels, absolute & relative organ weights, time of reflex response completion, pinna detachment, eye opening, incisor eruption, age & body weights at sexual maturation, AGD & and pup viability homogeneity of variance was evaluated using Bartlett's test then if homogeneous one way analysis of variance was used to determine if there were statistical differences between the groups and their counterparts in the controls.If the analysis of variance gave a significant result , Dunnetts test was performed to detect the differences between controls & the DEP groups.
When Bartlett's test detected non homogeneous results Kruskal-Wallis test was used to detect significant differences and then the Mann Whitney U test was performed to detect any significant differences between the DEP groups and the corresponding control group.
Statistical aanalyses was performed using litter means for body weights and the AGDs of pups /sex before weaning.
The completion of physical development of pups was assessed by the Wilcoxon rank test on the litter data,
The Chi squared /Fishers exact probability tests were used to analyse incidence of females with normal oestrus cycles & indices of copulation, fertility & gestation, & sex ratios of pups.
Reproductive indices:
See table below
Offspring viability indices:
See table below
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:
effects observed, treatment-related
Histopathological findings: non-neoplastic:
no effects observed
Other effects:
no effects observed
Reproductive function: oestrous cycle:
no effects observed
Reproductive function: sperm measures:
effects observed, treatment-related
Description (incidence and severity):
Incidences of abnormal sperm & tailess spermwere significantly increased in males of F0 generation given 3000 ppm DEP and in males of the F1 generation given 3000 and 15000 ppm.
Reproductive performance:
no effects observed
N/A as no adverse effects were observed on systemic & reproductive parameters at dietary levels of up to 15000 ppm.
Dose descriptor:
NOAEL
Effect level:
15 000 ppm (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: No adverse effects were observed on systemic & reproductive parameters at dietary levels of up to 15000 ppm.
Remarks on result:
other: Generation: F0 & F1 in parents (migrated information)
Dose descriptor:
NOAEL
Effect level:
3 000 ppm (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: In terms of development and growth pup body weight gain before weaning was reduced in the 15000 ppm group.
Remarks on result:
other: Generation: F1 & F2 pups (migrated information)
Clinical signs:
no effects observed
Mortality / viability:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Sexual maturation:
effects observed, treatment-related
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
no effects observed
Histopathological findings:
no effects observed
Description (incidence and severity):
examination limited to thymus & spleen
N/A as the effect on development and growth was reduced pup body weight gain before weaning in the 15000 ppm group
Reproductive effects observed:
not specified

Reproductive data for F0 and F1 parental animals

 

DEP (ppm) Administered

 

0 (control)

600

3000

15000

F0 parental animals

 

 

 

 

No. of pairs

24

24

24

24

Precoital interval (days)

3.4±3.8a

2.7±1.5

3.7±3.5

2.4±1.2

Copulation index (%)b

 

 

 

 

Male

95.8

95.8

100

100

Female

100

95.8

100

100

Fertility index (%)c

 

 

 

 

Male

95.7

95.7

100

95.8

Female

95.8

95.7

100

95.8

Gestation index (%)d

95.7

100

100

95.7

Gestation length (days)

22.3±0.5

22.3±0.5

22.3±0.5

22.1±0.3

No. of implantations

14.7±1.8

15.0±1.7

14.9±2.0

14.8±2.1

Delivery index (%)e

94.3±6.7

91.3±8.8

93.8±5.0

94.9±7.2

No. of pups delivered

13.9±2.0

13.7±2.0

14.0±2.1

14.0±2.2

F1 parental animals

 

 

 

 

No. of pairs

24

24

24

24

Precoital interval (days)

2.5±1.6

2.8±1.2

4.3±4.7

3.8±3.8

Copulation index (%)

 

 

 

 

Male

100

100

87.5

95.8

Female

100

100

95.8

100

Fertility index (%)

 

 

 

 

Male

95.8

95.8

95.2

95.7

Female

95.8

95.8

95.7

95.8

Gestation index (%)

100

100

100

100

Gestation length (days)

22.4±0.5

22.3±0.5

22.3±0.6

22.1±0.3*

No. of implantations

14.3±2.5

14.3±3.2

14.9±2.7

14.7±2.5

Delivery index (%)

91.3±11.1

93.7±8.4

91.8±13.2

98.8±17.9

No. of pups delivered

13.0±2.8

13.5±3.5

13.9±3.1

13.3±3.5

a Mean±SD; b Copulation index (%) = (No. of rats with successful copulation/no. of rats paired) x100; cFertility index (%) = (No. of females pregnant or no. of males sired/no. of rats with successful copulation) x100; d Gestation index (%) = (No. of females with parturition/no. of females pregnant) x100; e Delivery index (%) = (No. of pups delivered/no. of implantations) x100; *               Significantly different from the control, p0.05

Litter data on reproductive parameters for F1 and F2 offspring

 

DEP (ppm)

 

0 (control)

600

3000

15000

F1 offspring

 

 

 

 

No. of litters examined

22

22

24

22

Sex ratio of pupsb

0.544

0.465

0.507

0.490

Viability index before weaning (%)cde

 

 

 

 

Day 0

97.6

98.8

97.8

97.2

Day 4

97.8

96.2

97.2

97.7

Day 21

97.7

95.5

90.6

88.1

Male pup weight before weaning (g)

 

 

 

 

Day 0

6.9±0.7a

7.0±0.6

7.0±0.7

6.7±0.7

Day 4

10.6±1.4

10.1±1.4

10.5±1.9

9.7±1.4

Day 21

59.0±7.7

56.8±6.6

55.1±9.5

48.1±4.8f**

Female pup weight before weaning (g)

 

 

 

 

Day 0

6.5±0.7

6.5±0.7

6.5±0.7

6.3 b±0.6

Day 4

10.2±1.4

9.6±1.5

9.9±1.6

9.1±1.1*

Day 21

56.3±6.6

55.4±7.0

53.8±7.2

45.8±4.5f**

Anogenital distance (mm)

 

 

 

 

Male Day 0

3.67±0.31

3.72±0.29

3.83±0.35

3.56±0.28

Day 4

5.66±0.50

5.46±0.44

5.54±0.69

5.54±0.55

Female Day 0

1.76±0.21

1.67±0.22

1.67±0.23

1.77±0.22

Day 4

2.77±0.34

2.70±0.25

2.72±0.34

2.74±0.33

F2 offspring

 

 

 

 

No. of litters examined

23

23

22

23

Sex ratio of pups

0.510

0.510

0.523

0.482

Viability index before weaning (%)

 

 

 

 

Day 0

96.2

98.6

97.9

98.6

Day 4

96.3

97.8

99.4

98.1

Day 21

94.0

96.7

98.9

99.5

Male pup weight before weaning (g)

 

 

 

 

Day 0

7.1±0.5

7.0±0.7

6.9±0.7

6.8±0.6

Day 4

11.4±1.8

10.6±1.5

10.7±1.2

10.8±1.6

Day 21

60.9±9.5

61.4±6.9

60.6±4.5

53.8±4.8**

Female pup weight before weaning (g)

 

 

 

 

Day 0

6.6±0.6

6.5±0.7

6.4±0.5f

6.3±0.4g

Day 4

10.8±2.1

9.9±1.4

10.0±0.9f

10.0±1.1g

Day 21

58.2±9.4

58.7±6.2

58.2±4.2f

51.5±4.2g **

Anogenital distance (mm)

 

 

 

 

Male Day 0

3.63±0.29

3.59±0.17

3.64±0.20

3.65±0.24

Day 4

5.78±0.42

5.66±0.39

5.73±0.36

5.77±0.49

Female Day 0

1.76±0.11

1.73±0.09

1.76±0.09f

1.74±0.09g

Day 4

2.93±0.23

2.89±0.22

2.90±0.21f

2.88±0.20g

a Mean±SD; b Sex ratio of pups = No. of male pups/total no. of pups; c Viability index on postnatal day 0 (%) = (No. of live pups delivered/total no. of pups delivered) x100

d Viability index on postnatal day 4 (%) = (No. of live pups on postnatal day 4/no. of live pups delivered) x100 ; e Viability index on postnatal day 21 (%) = (No. of live pups on postnatal day 21/no. of live pups on postnatal day 4 after cull) x100; f and g Values obtained from 21 and 22 litters, respectively; * Significantly different from the control, p0.05; ** Significantly different from the control, p0.01

Hepatic microsomal cytochrome P450 isozyme contents in F0 parental males

 

DEP (ppm)

 

0 (control)

600

3000

15000

No. of F0 parental males examined

6

6

6

6

CYP3A2 (p mol/mg protein)

55.6±20.5

62.2±18.5

33.2±13.1

92.0±15.4**

CYP4A1 (p mol/mg protein)

13.7±3.1

15.0±6.9

11.5±2.6

62.8±19.2*

a              Mean±SD; *            Significantly different from the control, p0.05; **          Significantly different from the control, p0.01

Serum hormone levels in F0 parental males

 

DEP (ppm)

 

0 (control)

600

3000

15000

No. of F0 parental males examined

6

6

6

6

Testosterone (ng/mL)

2.63±1.23a

1.81±0.46

0.50±0.26**

1.26±0.66*

Progesterone (ng/mL)

2.66±2.51

3.61±2.50

5.98±3.00

3.08±0.94

a              Significantly different from the control, p0.05;             **            Significantly different from the control, p0.01

Conclusions:
Following dietary administration to rats through 2 successive generations, the NOAEL for general toxicity and reproductive performance in parental animals is considered to be 15000 ppm as there were no adverse effects on these parameters.
For development and growth of pups the NOAEL is considered to be 3000 ppm due to decreased body weight gain in those given 15000 ppm.
Executive summary:

Reproductive toxicity has been investigated in a 2 -generation study in rats using methods in accordance with OECD test methods. The the NOAEL for general toxicity and reproductive performance in parental animals was considered to be 15000 ppm (equivalent to a mean intake of 1016 -1297 mg/kg/day) as there were no adverse effects on these parameters. For development and growth of pups the NOAEL was considered to be 3000 ppm (equivalent to a mean intake of 222 -267 mg/kg/day) due to decreased body weight gain in those given 15000 ppm (equivalent to a mean intake of 1150 -1375 mg/kg/day).

Effect on fertility: via oral route
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEL
222 mg/kg bw/day
Study duration:
subchronic
Species:
rat
Effect on fertility: via inhalation route
Endpoint conclusion:
no study available
Effect on fertility: via dermal route
Endpoint conclusion:
no study available
Additional information

Reproductive toxicity has been investigated in a 2 -generation study in rats using methods in accordance with OECD test methods. The NOAEL for general toxicity and reproductive performance in parental animals was considered to be 15000 ppm (equivalent to a mean intake of 1016 -1297 mg/kg/day) as there were no adverse effects on these parameters. For development and growth of pups the NOAEL was considered to be 3000 ppm (equivalent to a mean intake of 222 -267 mg/kg/day) due to decreased body weight gain in those given 15000 ppm (equivalent to a mean intake of 1150 -1375 mg/kg/day).

Studies in the mouse suport these findings, reporting no effects in the parental generation but with minor effects becoming apparent in successive generations.


Short description of key information:
Reproductive toxicity:
Fertility: NOAEL - 222 mg/kg bw

Effects on developmental toxicity

Description of key information
Reproductive toxicity:
Developmental toxicity: Oral NOAEL - 1900 mg/kg bw; Dermal NOAL - 1 mL/kg bw (equivalent to 1118 mg/kg bw)
Link to relevant study records

Referenceopen allclose all

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Experimental result published in peer reviewed journal. Some details on methods / results not available
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Principles of method if other than guideline:
DEP was administered to time-mated rats in the diet during the period of organogenesis and maternal toxicity and effects on embryo-foetal viability, growth and morphogenesis assessed.
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
Sprague-Dawley
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories Inc Raleigh NC USA
- Weight at study initiation: females 0gd 207-273 g
- Fasting period before study: No
- Housing: individually in solid bottom polycarbonate cages with stainless steel wire lids (Laboratory products Rochelle Park NJ USA
- Diet (e.g. ad libitum): yes, ground Purina Certified Chow #5002
- Water (e.g. ad libitum): yes, deionised water
- Acclimation period: 7d

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 degrees C
- Humidity (%): 48%
- Photoperiod (hrs dark / hrs light): 12
Route of administration:
oral: feed
Vehicle:
other: control feed
Details on exposure:
DIET PREPARATION
- Rate of preparation of diet (frequency):
- Mixing appropriate amounts with (Type of food): mixed with feed in a Patterson Kelley Liquids -Solids twin shell blender
- Storage temperature of food: refridgerated in containers protected from light

VEHICLE
- Justification for use and choice of vehicle (if other than water): control feed, DEP was administered in the diet
- Concentration in vehicle: 0, 0.25, 2.5 & 5.0%
- Lot/batch no. (if required): not reported but fed ground Purina Certified Chow #5002
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Diets were analysed by HPLC before & after use. Analysis confirmed that the diets contained 89-110% of the nominal concentrations.
Details on mating procedure:
- Impregnation procedure: [artificial insemination / purchased timed pregnant / cohoused]: Time mated rats
- If cohoused: bred in pairs
- M/F ratio per cage: 1:1
- Length of cohabitation: overnight until sperm detected in the vagina
- Proof of pregnancy: [vaginal plug / sperm in vaginal smear] referred to as [day 0 / day 1] of pregnancy: sperm in vaginal smear = gd0
Duration of treatment / exposure:
From morning of gd6 to morning of gd15.
Frequency of treatment:
Continuously in the diet
Duration of test:
Until GD 20
Remarks:
Doses / Concentrations:
0, 0.25, 2.5 & 5.0%
Basis:
nominal in diet
No. of animals per sex per dose:
25-32 gravid females
Control animals:
yes, plain diet
Details on study design:
- Dose selection rationale: To include and exceed the range of doses used in earlier study (ip study in 5 animals/group by Singh et al 1972 used 0.57, 1.13 & 1.89 mg/kg during gestation period reported to cause effects onfoetal growth & development including an increase in the incidence of skeletal defects)
- Rationale for animal assignment (if not random): assignment was by stratified randomisation so that body weights were simalar in the groups within each of 2 replicates
- Other: range of 3-4 consecutive breeding days in each replicate. The replicates were 26 days apart
Maternal examinations:
CAGE SIDE OBSERVATIONS: No data

DETAILED CLINICAL OBSERVATIONS: No data

BODY WEIGHT: Yes
- Time schedule for examinations: d: 0, 3, 6, 9, 12, 15, 18, & 20

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Yes
- 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: Yes over 3 day periods from gd6-20 ie gd6-9, 9-12, 12-15, 15-18, 18-20

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): Yes
- Time schedule for examinations: gd6-9, 9-12, 12-15, 15-18, 18-20

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 20
- Organs examined: Not clearly stated

OTHER: Body weight, & liver, kidney and intact uterus weights were recorded. Uteri with no visible implantation sites were stained with 10% amonium sulfide to detect implantation sites from early resorptions.

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
- Soft tissue examinations: Yes
- Skeletal examinations: Yes
- Head examinations: Yes
Statistics:
SAS (GLM procedures)software was used for the analysis of variance of maternal & foetal parameters. When ANOVA revealed a significant P<0.05) dose effect Williams & Dunnetts tests were used to compare each treatment group with the concurrent control group. One-tailed were used in the pair wise comparisons except those of body weigt & organ weight. For non significant(p>0.05 dose X replicate effects on foetal parameters data were pooled accross the replicates and non parametric tests applied. When the X2 test indicated significant differences among all groups a one tailed Fisher's exact probability test was used to compare individual treatment groups with the concurrent control group. The alpha level for ball groupwise and pairwise comparisons was 0.05.
Indices:
Not reported
Historical control data:
Not reported
Details on maternal toxic effects:
Maternal toxic effects:yes. Remark: Body weight & body weight gain were reduced in dams given 2.5 and 5% DEP in the diet from on and from 9 respectively. See table below. Food & water consumption was also reduced in these groups during gd 6-9 & 9-12 respectively and increased during in the

Details on maternal toxic effects:
See table below
Dose descriptor:
NOAEL
Effect level:
0.25 other: %
Based on:
test mat.
Basis for effect level:
other: maternal toxicity
Dose descriptor:
NOAEL
Effect level:
2.5 other: %
Based on:
test mat.
Basis for effect level:
other: developmental toxicity
Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:yes. Remark: A signicantly increased incidence of variations as a result of the increase in rudimentary extra lumbar ribs in the 5% dose group when compared with the controls. See tables below.

Details on embryotoxic / teratogenic effects:
See tables below
Abnormalities:
not specified
Developmental effects observed:
not specified

Maternal toxicity in CD rats exposed to diethyl phthalate in feed on gestational days 6 to 15

 

Diethyl phthalate (% in feed)

 

0

0.25

2.5

5.0

Subjects (dams)

Total treated

31

32

32

32

Removed

0

11

0

0

No. (%) pregnant

27(87)

29(94)

30(94)

32(100)

Chemical exposure (g/kg/day)2

gd 6 to 9

0

0.21

1.65

1.95

gd 9 to 12

0

0.20

1.97

3.43

gd 12 to 16

0

0.19

2.06

4.39

gd 6 to 16

0

0.20

1.91

3.21

Maternal wt gain (g)3

Gestation period

152.8±4.0

169.2±3.4*

154.8±3.2

145.8±3.9

Treatment period

52.8±2.0

61.4±1.4*

52.6±1.6

30.6±3.2*

Corrected wt gain4

66.4±3.9

78.4±2.4*

67.1±2.7

57.0±3.2*

Gravid uterine wt (g)3

86.0±3.3

90.9±2.6

87.7±2.2

88.7±1.9

Relative food consumption (g/kg/day)2

gd 6 to 9

82.3±1.5

82.3±1.2

65.9±3.7*

39.0±6.1*

gd 9 to 12

78.8±1.1

79.7±2.9

78.6±2.3

68.5±3.9*

gd 12 to 16

76.7±1.2

76.7±1.8

82.5±1.2*

87.7±2.0*

gd 16 to 18

77.9±1.3

79.3±1.3

80.2±1.6

92.2±2.4*

gd 18 to 20

73.2±2.4

73.2±1.6

76.3±2.0

83.0±3.2*

gd 0 to 20

77.9±0.8

77.2±1.0

77.2±0.9

75.6±1.1

Relative water consumption (g/kg/day)2

gd 6 to 9

143.3±4.7

133.5±5.0

121.9±4.1*

119.8±4.9*

gd 9 to 12

143.3±6.0

141.2±4.6

140.8±4.2

154.4±5.6

gd 12 to 16

154.6±14.8

135.0±3.4

150.7±10.8

180.6±8.3

gd 16 to 18

138.8±9.2

138.2±4.8

148.4±4.1

180.6±5.4*

gd 18 to 20

134.6±6.4

126.3±2.4

135.3±3.0

158.0±5.3*

gd 0 to 20

142.1±5.7

133.6±3.4

137.6±3.6

148.9±3.7

1 Dams were removed because of insufficient food or water or for being outside of the weight range

2 Food consumption during treatment (g/day) x % chemical x 1000 mg/g / average body wt during treatment (kg)

3 Includes all dams pregnant at sacrifice, mean±SB

4 Weight gain during gestation minus gravid uterine weight

* Dunnett’s test or Williams’ test, P<0.05             

Linear trend test, P<0.05

Developmental toxicity in CD rats following maternal exposure to diethyl phthalate in feed on gestational days 6 to 15

 

Diethyl phthalate (% feed)

 

0

0.25

2.5

5.0

All litters1

27

29

30

32

No. implant sites/litter

15.2±0.5

15.8±0.4

15.0±0.4

15.5±0.3

% resorptions/litter

3.8±1.4

3.9±0.9

4.1±1.4

3.1±0.7

% litters with resorptions

25.9

48.3

26.7

40.6

Live litters4

27

29

30

32

No. live foetuses/litter

14.6±0.5

15.2±0.5

14.4±0.4

15.0±0.3

% male foetuses/litter

50.4±2.8

48.3±2.8

48.7±2.8

50.2±2.5

Mean body wt. (g)/litter

3.7±0.2

3.9±0.1

3.9±0.1

3.8±0.0

% foetuses malformed/litter

0.7±0.5

0.4±0.3

0.5±0.3

1.3±0.6

% litters with malformed foetuses

7.4

6.9

6.7

15.6

% foetuses with variations/litter

10.6±3.1t

8.4±2.2

12.7±2.8

23.5±4.1*

% litters with variations

51.9+

44.8

66.7

84.4

% foetuses with extra rib/litter

8.8±3.0

7.1±2.0

10.6±2.9

21.0±4.1*

% litters with extra rib

44.4+

39.3

46.7

74.2

1 Includes all dams pregnant at sacrifice litter size = no. implantation sites per dam: mean±SE; 2 One dam had a litter that contained only one dead foetus

3 One dam had 100 resorptions; 4 Includes only dams with live foetuses litter = no. live foetuses per dam: mean±SE

Linear trend test, P<0.05; * Dunnett’s test or Williams’ test, P<0.05; +x2 test, P<0.05;tFisher’s exact test, P<0.05

 

Morphologic defects in CD rat fetuses following maternal exposure to diethyl phthalate in feed on gestational days 6 to 15

 

Diethyl phthalate (% of feed)

 

0

0.25

2.5

5.0

All malformations

Litters with malformations/total litters3

2/27

2/29

2/30

5/32

Malformed foetuses/total examined4

3/394

2/442

2/431

6/479

External malformations

No. litters with defects5

0

1

0

2

No. foetuses with defects6

0

1

0

2

Exencephaly

0

1

0

1

Spina bifida

0

1

0

1

Microphthalmia

0

0

0

1

Visceral malformations

No. litters with defects5

2

0

0

3

No. foetuses with defects6

3

0

0

4

Hydroureter

3

0

0

2

Hydronephrosis

0

0

0

2

Enlarged lateral ventricle of brain

0

0

0

1

Extra renal vessel

0

0

0

1

Skeletal malformations

No. litters with defects5

0

2

2

0

No. foetuses with defects6

0

2

2

0

Fused rib cartilage

0

1

0

0

Short rib XIII

0

1

0

0

Rib cartilage not attached to sternum

0

1

2

0

Variations

No. litters with defects5

14

13

20

27

No. foetuses with defects6

42

38

54

117

Hematoma

3

2

4

2

Open eye

0

0

0

1

Visceral variations

Agenesis of the innominate artery

0

0

0

2

Pale area on liver

0

0

0

1

Distended ureter

3

3

1

3

Skeletal variations

Misaligned sternebra

1

0

2

0

Wavy rib

 

1

1

2

Extra rib (rudimentary)

32

32

44

100

Extra rib (full)

3

0

1

7

Incomplete ossification with cartilage, frontals and/or parietals (midline)

0

0

1

3

Normal cartilage, bipartite ossification centre of vertebral centrum

1

3

0

3

Dumbbell cartilage, bipartite ossification centre of vertebral centrum

0

0

0

0

1 Statistical evaluation of these data is reported in Table 2; 2 Foetuses may have more than one defect; 3 Includes only litters with live foetuses

4 Only live foetuses were examined for defect;5Litters with one or more defective foetuses; 6 Foetuses with one or more defects

 

Conclusions:
Under the conditions in which this study was performed the NOAELs for maternal & developmental toxicity were 0.25% & 2.5% DEP in the diet due to the reduced maternal body weight at the 2.5 (transient) & 5% dietary concentrations & the increase in the incidence of supernumery ribs in the foetues exposed to 5% DEP in the diet.
Executive summary:

Developmental toxicity has been investigated in the rat using methods similar to those described by OECD test guidelines. Diethyl phthalate was administered in the diet during gestation days 6 -15. The NOAELs for maternal & developmental toxicity were 0.25% (equivalent to 200 mg/kg) and 2.5% (equivalent to 1910 mg/kg) due to the reduced maternal body weight at the 2.5 (transient) & 5% dietary concentrations and an increase in the incidence of supernumery ribs in the foetues exposed to 5% DEP in the diet.

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
supporting study
Study period:
1984
Reliability:
1 (reliable without restriction)
Qualifier:
no guideline followed
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Principles of method if other than guideline:
Female rabbits were mated with untreated previously proven fertile bucks. Mating was controlled and designated day 0 of gestation. The mated females were given DEP daily from day 6-18 of gestation and were sacrificed on day 29 & foetuses were delivered by Caesarian section & examined. The number of foetuses at birth, status, distribution in the uterus and the placental indicators recorded.
GLP compliance:
yes
Limit test:
no
Species:
rabbit
Strain:
New Zealand White
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Harold Schriever D-2140 Bremerverde, Nuendamm 88 DE
- Age at study initiation: 12 weeks
- Weight at study initiation: 2.45-4.60 kg
- Fasting period before study: 1984-08-02-1984-11-13
- Housing: individually in cages (35 cm high X 50 cm long X35 cmwide)
- Diet (e.g. ad libitum): Saniff K (Alleindikt furzuchtkaninchen) produced by Saniff Specialfutter GMBH 4770 Soest/Westfalen DE
- Water (e.g. ad libitum): aqua fontana as fit for human consumption
- Acclimation period: Time mated so N/A
-other C of As available for food & water

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/-2 degrees C
- Humidity (%): 45-85%
- Air changes (per hr): 16
- Photoperiod (hrs dark / hrs light): 12

IN-LIFE DATES: From: 1984-08-02 To: 1984-11-13
Route of administration:
dermal
Vehicle:
CMC (carboxymethyl cellulose)
Remarks:
0.5%
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: Just prior to dosing DEP was suspended homogeneously in vehicle using an Ultra Turrax.

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 2ml/kg
- Concentration (if solution): 0, 5, 15 & 50% w/w
- Constant volume or concentration used: yes/no no based on most recent body weight
- For solids, paste formed: yes/no: N/a

VEHICLE
- Justification for use and choice of vehicle (if other than water): DEP is poorly soluble in water
- Concentration in vehicle: 5, 15 & 50% W/W suspension.
- Amount of vehicle (if gavage): 2ml/kg
- Lot/batch no. (if required): No data
- Purity: No data

TEST SITE
- Area of exposure: 10X10 cm on dorsolumbar region
- % coverage: not reported (estimated to be approximately 10% of body surface)
- Type of wrap if used: N/A not used
- Time intervals for shavings or clipplings: not reported

REMOVAL OF TEST SUBSTANCE
- Washing (if done): yes, using tepid water and drying with paper towel
- Time after start of exposure: 7h

USE OF RESTRAINERS FOR PREVENTING INGESTION: yes/no: yes
Analytical verification of doses or concentrations:
not specified
Details on mating procedure:
- Impregnation procedure: [artificial insemination / purchased timed pregnant / cohoused]: purchased time mated
- If cohoused: individually housed
- M/F ratio per cage: N/A
- Length of cohabitation: N/A
- After ... days of unsuccessful pairing replacement of first male by another male with proven fertility. N/A
- Further matings after two unsuccessful attempts: [no / yes (explain)] N/A
- Verification of same strain and source of both sexes: [yes / no (explain)] No data
- Proof of pregnancy: [vaginal plug / sperm in vaginal smear] referred to as [day 0 / day 1] of pregnancy: controlled copulation = d0.
Duration of treatment / exposure:
d6-18 of gestation
Frequency of treatment:
daily during treatment period.
Duration of test:
29d
Remarks:
Doses / Concentrations:
0 (Control), 5, 15 & 50% DEP in 0.5% CMC
Basis:
nominal conc.
based on weights eg 5% = 2.25 g DEP in 42.75g 0.5% CMC; 15% = 2.25g DEP in 12.75 g 0.5% CMC; 50% DEP = 14g DEP in 14g 0.5% CMC. Treated animals received 2 ml/Kg body weight.
No. of animals per sex per dose:
12 females
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: The selected concentrations were based on the results of a dose range finding experiment and were known not to cause maternal toxicity
- Rationale for animal assignment (if not random): randomly assigned based on body weights
- Other: Rabbbit used as it is one of the animals of choice & because of it's known sensitivity & susceptability to teratogenic compounds
Maternal examinations:
CAGE SIDE OBSERVATIONS: No

DETAILED CLINICAL OBSERVATIONS: Yes: daily for sensory & motor behaviour checking coat, urine & faecal excretion and condition of body orifices. During treatment period reflex examinations (modified Irwin) were also conducted. Skin on application sites was graded using the Draize scoring system.

BODY WEIGHT: Yes
- Time schedule for examinations: day 0, daily during days 6-18 & on d 24 & 29.

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day : 29
- Organs examined: uterus & position of foetuses, placentae, no. of male & female foetuses

OTHER: foetuses removed by caesarean section; foetuses, placentae gravid uteri & uteri without foetuses were weighed
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
- Soft tissue examinations: Yes: approximately 1/3rd
- Skeletal examinations: Yes: approximately 2/3rds
- Head examinations: No data
Statistics:
One wy analysis of variance was calculated for growth & foetal parameters. Mean values were compared using Tukei's method.
Indices were compared using the Mann-Whitney U test
Indices:
Indices used included abortion rate, still birth rate, Resorption index, Variation index, Malformation Index, Runts index & post implantation index
Historical control data:
Yes & used for rabits of this strain housed in this laboratory; also used to conclude that the incidence of 2 malformed foetuses was within the spontaneous malformation rate for rabbits of this stain at this laboratory.
Details on maternal toxic effects:
Maternal toxic effects:yes

Details on maternal toxic effects:
Skin reactions to treatment on application site: 50% of the animals given 15% DEP developed slight erythema and a few individuals developed slight oedema & slight atonia after 6 days of treatment. Slight desquamation was seen at the end of the treatment period.
All animals given 50% DEP had slight erythema and some individuals developed slight oedema, slight atonia and slight desquamation throughout the treatment period.
One control anmal gave birth prior to weighing on d29. The foetuses had developed normally and were evaluable with the exception of one which was cannibalised.
Dose descriptor:
NOAEL
Effect level:
50 other: %
Based on:
test mat.
Basis for effect level:
other: maternal toxicity
Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:no effects. Remark: see tables below

Details on embryotoxic / teratogenic effects:
N/A
Abnormalities:
not specified
Developmental effects observed:
not specified

SUMMARY of DATA                                 

Group/

Dose

I CMC

(0%)

II DEP

(5%)

III DEP (15%)

IV DEP (50%)

Number of females

12

12

12

12

Number of Litters

12

12

12

12

Gestation index (%)

100

100

100

100

Mean weight change (g) ± SD (day 6 - 18)

352.5 ± 152.9

425.8 ± 207.8

425.0 ± 152.4

383.3 ± 171.3

a. Fetuses

 

 

 

 

 

 

 

 

   

Total Number of foetuses

7.67 ± 2.19

6.58 ± 2.84

6.4 2 ± 2.50

6.67 ± 2.87

Left/right

3.58/4.08

3.00/3.58

3.25/3.17

2.75/3.92

Male/female

3.92/3.67

3.75/2.83

3.17/3.25

3.42/3.00

Dead foetuses

0

1

0

0

Still birth index (%)

0

1.27

0

0

Number of runts

0

2

0

1

Runts index (%)

0

2.5

0

1.3

Mean foetal weight ± SD

39.48 ± 6.04

42.27 ± 9.66

47.51 ± 6.06

44.14 ± 5.97

Group

I CMC

(0.5%)

II DEP

(5%)

III DEP

(15%)

IV DEP

(50%)

b. Resorptions

 

 

 

 

 

 

 

 

   

Total per dam (mean ± SD)

0.75 ± 1.48

0.17 ± 0.39

0.08 ±0.29

1.17 ± 2.59

Early/late

067/0.08

0.00/0.17

0.08/0.00

1.00/0.17

Resorption index (%)

8.9

2.5

1.3

14.9

c. Implantations

 

 

 

 

 

 

   


 

 

Total per dam (mean ± SD)

8.42 ± 2.43

6.75 ± 3.05

6.50 ± 2.54

7.83 ± 2.21

Left/right

4.17/4.25

3.17/3.58

3.33/3.17

3.08/4.75

Post implantation- loss index (%)

8.9

3.7

1.3

14.9

d. Corpora lutea

 

 

 

 

 

 

 

 

   

Total per dam (mean ± SD)

9.08 ± 2.87

8.42 ± 1.51

7.58 ± 2.35

9.50 ± 2.28

Left/right

4.42/4.67

4.17/4.25

3.75/3.83

3.92/5.58

e. Placenta

 

 

 

 

 

 

 

 


 

 

Mean weights (g) (mean ± SD)

5.84 ± 0.78

7.08 ± 2.21

7.27 ± 1.19

7.07 ± 1.28

Examination of foetuses

   

Malformed foetuses (total)

0

0

0

2

Malformation index (%)

0

0

0

2.5

Conclusions:
Under the conditions of the study, daily dermal administration of 50% diethyl phthalate at a dose volume of 2 mL/kg body weight (eqiuvalent to 1 mL diethyl phthalate/kg) during the gestation period of rabbits had no adverse effects on foetal development
Executive summary:

Developmental toxicity has been investigated in the rabbit using methods similar to those described by OECD test guidelines. Daily dermal administration of 50% diethyl phthalate at a dose volume of 2 mL/kg body weight (eqiuvalent to 1 mL diethyl phthalate/kg) during the gestation period (Days 6 -18) had no adverse effects on foetal development

Effect on developmental toxicity: via oral route
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEL
1 910 mg/kg bw/day
Study duration:
subacute
Species:
rat
Effect on developmental toxicity: via inhalation route
Endpoint conclusion:
no study available
Effect on developmental toxicity: via dermal route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
1 118 mg/kg bw/day
Study duration:
subacute
Species:
rabbit
Additional information

Developmental toxicity has been investigated in the rat using methods similar to those described by OECD test guidelines. Diethyl phthalate was administered in the diet during gestation days 6 -15. The NOAELs for maternal & developmental toxicity were 0.25% (equivalent to 200 mg/kg) and 2.5% (equivalent to 1910 mg/kg) due to the reduced maternal body weight at the 2.5 (transient) & 5% dietary concentrations and an increase in the incidence of supernumery ribs in the foetues exposed to 5% DEP in the diet.

Developmental toxicity has also been investigated in the rabbit using methods similar to those described by OECD test guidelines. Daily dermal administration of 50% diethyl phthalate at a dose volume of 2 mL/kg body weight (equivalent to 1 mL diethyl phthalate/kg/1118 mg/kg diethyl phthalate) during the gestation period (Days 6 -18) had no adverse effects on foetal development.

In the mouse, daily dermal administration during the gestation period (Days 0 -17) had no adverse effects on foetal development. Maternal toxicity was apparent at all doses, evidenced by reduced thymus and spleen weights and also by increased adrenal gland weights at the 5600 mg/kg/day dose. Foetal body weight was significantly reduced in animals given 5600 mg/kg/day and there was an increase in the incidence of cervical and lumbar rib variations. However no external, visceral or skeletal anomalies were seen in the foetuses which could be attributed to treatment with DEP.

Toxicity to reproduction: other studies

Additional information

Reproductive toxicity has been investigated in a 2 -generation study in rats using methods in accordance with OECD test methods. The NOAEL for general toxicity and reproductive performance in parental animals was considered to be 15000 ppm (equivalent to a mean intake of 1016 -1297 mg/kg/day) as there were no adverse effects on these parameters. For development and growth of pups the NOAEL was considered to be 3000 ppm (equivalent to a mean intake of 222 -267 mg/kg/day) due to decreased body weight gain in those given 15000 ppm (equivalent to a mean intake of 1150 -1375 mg/kg/day). Studies in the mouse support these findings, reporting no effects in the parental generation but with minor effects becoming apparent in successive generations.

Developmental toxicity has been investigated in the rat using methods similar to those described by OECD test guidelines. Diethyl phthalate was administered in the diet during gestation days 6 -15. The NOAELs for maternal & developmental toxicity were 0.25% (equivalent to 200 mg/kg) and 2.5% (equivalent to 1910 mg/kg) due to the reduced maternal body weight at the 2.5 (transient) & 5% dietary concentrations and an increase in the incidence of supernumery ribs in the foetues exposed to 5% DEP in the diet. Daily dermal administration of 50% diethyl phthalate at a dose volume of 2 mL/kg body weight (equivalent to 1 mL diethyl phthalate/kg = 1118 mg/kg diethyl phthalate) during the gestation period (Days 6 -18) had no adverse effects on foetal development. In the mouse, daily dermal administration during the gestation period (Days 0 -17) had no adverse effects on foetal development. Maternal toxicity was apparent at all doses, evidenced by reduced thymus and spleen weights and also by increased adrenal gland weights at the 5600 mg/kg/day dose. Foetal body weight was significantly reduced in animals given 5600 mg/kg/day and there was an increase in the incidence of cervical and lumbar rib variations. However no external, visceral or skeletal anomalies were seen in the foetuses which could be attributed to treatment with DEP.

There are concerns that diethyl phthalate may exhibit endocrine disrupting effects similar to those observed with other phthalate esters which may have a correlation with the minor effects seen at high levels of exposure.

In an investigation of changes of Leydig cells caused by treatment with diethyl phthalate, young male Wistar rats were treated by oral gavage with 2000 mg/kg for 2 days (Jones HB, Garside DA, Liu R, & Roberts JC. Influence of phthalate esters on Leydig cell structure and function in vitro and in vivo. Experimental and Molecular Pathology 58: 179-193, 1993). Testosterone production was measured and cells were examined by electron microscopy. Diethyl phthalate produced ultrastructural changes in Leydig cells, characterised by smooth endoplasmic reticulum focal dilation, swelling of the mitochondria associated and with a loss of matrix granules in all treated animals. An increase in interstitial macrophage activity was also seen with the activity located around Leydig cells showing substantial cytoplasmic alterations such as swollen mitochondria. Testes weights and zinc levels in the liver and testes were unaffected by treatment. The histological effects were not replicated in vitro when Leydig cells were cultured with 1000 μM of methylethyl phthalate, a major DEP metabolite.

Effects on the levels of laurate hydroxylase, a marker for the CYP 4 family of isozymes which is responsible for the metabolism of testosterone, has been examined in rat liver microsomes (Okita R & Okita JR. Effects of diethyl phthalate and other plasticisers on laurate hydroxylation in rat liver microsomes. Pharmaceutical Research 9: 1648-1653,1992). Administration of diethyl phthalate increased the specific activity of laurate hydroxylase by 1.6 times more than that induced in control rats but 6.75 fold less than that induced by 2 -diethylhexyl phthalate (DEHP), a known testicular toxicant.

 

Diethyl phthalate does not bind to human oestrogenic receptor (ER) in vitro (Nakai M, Tabira Y, Asai D, Yakabe Y, Shimyozu T, Noguchi M, Takatsuki M, & Shimohigashi Y. Binding characteristics of dialkyl phthalates for the estrogen receptor. Biochem Biophys Res Comm 254: 311-314,1999;Toda C, Okamoto Y, Ueda K, Hashizume K, Itoh K, & Kojima N. Unequivocal estrogen receptor-binding affinity of phthalate esters featured with ring hydroxylation and proper alkyl chain size. Arch Biochem Biophys 431: 16-21,2004). It was also was negative for oestrogenic activity in a yeast two-hybrid assay (Nishihara T, Nishikawa J, Kanayama T, Dakeyama F, Saito K, Imagawa M, Takatori S, Kitagawa Y, Hori S, & Utsumi H. Estrogenic activities of 517 chemicals by yeast two-hybrid assay. Journal of Health Science 46(4): 282 -298,2000) although it has shown weak oestrogenic activity in a recombinant yeast assay (Harris CA, Henttu P, Parker MG & Sumpter JP. The estrogenic activity of phthalate esters in vitro. Environmental Health Perspectives 105 (8): 802-811,1997). Diethyl phthalate has not demonstrated oestrogenic activity in human ERα and ERβ reporter gene assays in CHO-K1 cells transfected with expression vectors for ERα, ERβ and androgen receptor (AR) (Takeuchi S, Iida M, Kobayashi S, Jin K, Matsuda T & Kojima H. Differential effects phthalate ester human estrogen receptors. Toxicology 210: 223-233,2005). Expression of CaBP-9k mRNA, a gene highly regulated by 17β-oestradiol, was not increased in 7 day old female SD rats following oral treatment with 600 mg/kg diethyl phthalate for 3 days (Hong EJ, Ji YK, Choi KC, Manabe N, & Jeung EB. Conflict of estrogenic activity by various phthalates between in vitro and in vivo models related to the expression of Calbindin-D9k. Journal of Reproduction & Development 51(2): 253-263,2005). Monoethyl phthalate, a metabolite of diethyl phthalate induced detachment of germ cells from a Sertoli cell monolayer in vitro but was 10000 fold less potent than MEHP, the main metabolite of DEHP and a recognised reproductive toxicant (Gray T & Gangoli S. Aspects of the testicular toxicity of phthalate esters. Environ Health Perspect 65: 229-235,1986).

Justification for classification or non-classification

Non-classification is justified by available data on reproductive effects which show a lack of significant effects at non-maternally toxic levels of exposure.