1,2-Benzenedicarboxylic acid, di-C16-18-alkyl esters

Developmental toxicity / teratogenicity

Administrative data

Endpoint:

developmental toxicity

Type of information:

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

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: GLP-Guideline study with acceptable restrictions. Not full study report published; only data of C9-C11 alkyl phthalates were presented here;

Data source

Reference

Reference Type:

publication

Title:

Unnamed

Year:

2001

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.3700 (Prenatal Developmental Toxicity Study)

Deviations:

no

Principles of method if other than guideline:

Groups of 22 timed-mated Sprague-Dawley rats were administered 250, 500, or 1000 mg/kg D911P daily by oral gavage (5 mL/kg) between gestation days (GD) 1 and 19. Control animals received the vehicle (olive oil) alone. On GD20, the animals were sacrificed and the fetuses examined.

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

1,2-Benzenedicarboxylic acid, di-C9-11-branched and linear alkyl esters

EC Number:

271-085-1

EC Name:

1,2-Benzenedicarboxylic acid, di-C9-11-branched and linear alkyl esters

Cas Number:

68515-43-5

IUPAC Name:

dioctyl phthalate

Details on test material:

- Name of test material (as cited in study report): di-(C9-C11 alkyl) phthalate (D911P)
- Analytical purity: 99.3 % (m/m)
- Batch: PLA/S20517/97)
- Source: BP Chemicals Ltd., Hull
- storage: at ambient temperature

Test animals

Species:

rat

Strain:

Sprague-Dawley

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River UK Ltd. (Margate, Kent).
- Age at study initiation: 10-11 weeks
- Weight at study initiation: 210-267 g
- Diet: ad libitum SDS Laboratory Animal Diet No. 1, Special Diet Services Ltd, Witham, Essex
- Water: ad libitum
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21
- Humidity (%):55
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

olive oil

Details on exposure:

Dose volume: 5 mL/kg bw

Details on mating procedure:

- Impregnation procedure: cohoused
- If cohoused:
- M/F ratio per cage: 1:1
- Proof of pregnancy: sperm-positive vaginal smear or at least three copulation plugs referred to as day 0 of pregnancy

Duration of treatment / exposure:

Gestational days 1-19

Frequency of treatment:

once a day

Duration of test:

20 days

Doses / concentrations

Remarks:

Doses / Concentrations:
0, 250, 500, and 1000 mg/kg/day
Basis:
actual ingested

No. of animals per sex per dose:

22 females

Control animals:

yes, concurrent vehicle

Examinations

Maternal examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice a day

DETAILED CLINICAL OBSERVATIONS: No data

BODY WEIGHT: Yes
- Time schedule for examinations: Bodyweight was measured daily throughout the study, and

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Yes
food consumption was recorded for gestation days 1–2, 3–5, 6–8, 9–11, 12–14, 15–17, and 18–19.

POST-MORTEM EXAMINATIONS: Yes
On GD20, the females were killed by carbon dioxide inhalation, weighed, and examined macroscopically for signs of disease or adverse reaction to treatment.

Ovaries and uterine content:

The reproductive tracts, including ovaries, were removed and assessed for gravid uterine weight, number of corpora lutea in each ovary, numbers of implantation and resorption sites, and number and distribution of fetuses in each uterine horn.

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 resorptions: Yes

Fetal examinations:

- External examinations: Yes: all per litter
- Soft tissue examinations: Yes: half per litter
- Skeletal examinations: Yes: half per litter
- Head examinations: No data

Fetuses were uniquely identified with respect to their position in the uterine horn. Each fetus was weighed, sexed, examined for external abnormalities, and euthanized by chilling on a cool plate at approximately 4°C. Each placenta was weighed and examined for abnormalities. Approximately half of each litter was allocated to skeletal examination. These fetuses were given a visceral examination prior to evisceration and fixation in industrial methylated spirit. They were then processed and stained with Alizarin Red, using a modification of the Dawson staining technique. The remainder of the fetuses were fixed in Bouin’s fluid prior to examination by the Wilson free-hand serial sectioning technique.

Statistics:

Homogeneity of variance was assessed using Bartlett’s test, and data were analyzed with parametric tests (analysis of variance followed by Williams’ test) or nonparametric tests (Kru´skal-Wallis followed by Shirley’s test) as appropriate. Data on fetal malformations and variations was analyzed using categorical data models. Significant differences between treatments and controls for the proportions of affected fetuses were determined using Wald x2 tests with Bonferroni’s adjustment for multiple testing applications. Trend analysis was performed using logistic regression to separate spurious results from the statistically significant effects of an increased dosage level. For litter data, the litter was generally taken as being the unit for analysis. Since most distributions were nonnormal, nonparametric analysis was routinely used.

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:

Maternal toxic effects:no effects

Details on maternal toxic effects:
Treatment resulted in no signs of maternal toxicity, as assessed by adjusted maternal bodyweight gain throughout gestation, clinical examinations and food consumption. There were no remarkable macroscopic findings in the maternal animals at necropsy.

Effect levels (maternal animals)

1

Dose descriptor:

NOAEL

Effect level:

1 000 mg/kg bw/day

Based on:

test mat.

Basis for effect level:

other: maternal toxicity

2

Dose descriptor:

NOEL

Effect level:

250 mg/kg bw/day

Based on:

test mat.

Basis for effect level:

other: developmental toxicity

3

Dose descriptor:

NOAEL

Effect level:

1 000 mg/kg bw/day

Based on:

test mat.

Basis for effect level:

other: developmental toxicity

Results (fetuses)

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
Treatment resulted in no effects upon litter size, fetal survival or bodyweight. Fetal, litter, and placental weights were all unaffected by treatment with D911P, and there was no effect upon the overall incidence of external or visceral abnormalities.
There was a significant increase in dilated renal pelves in pups of the high dose groups.
Increased incidences of rudimentary supernumerary lumbar ribs at 500 and 1000 mg/kg bw /d.
These rudimentary supernumerary ribs, defined as being less than 50% the length of the 13th rib, are a common skeletal variation that occurs spontaneously at a high frequency in rats.

Effect levels (fetuses)

Remarks on result:

other: not specified

Fetal abnormalities

Abnormalities:

not specified

Overall developmental toxicity

Developmental effects observed:

not specified

Any other information on results incl. tables

 Table 1

Selected foetal observations following treatment with D911P from GD1 to GD19

	0	250	500	1000	Historical Control^d
No. examined viscerally^a	165 (22)	162 (22)	157 (22)	153 (21)
No. examined ekeletally^a	168 (22)	158 (22)	158 (22)	157 (2!)
Visceral findings^b
dilated ureter	3 (3) [1.8]	11 (5) [8.6]	4 (3) [2.4]	11 (7) [7.7]	1.9-1.3% (13.6-45.5%)
dilated renal pelvis^c	2 (2) [1.0]	7 (3) [4.1]	2 (1) [1.3]	12 (5) [7.9*]^‡
hydroureter^c	0 [0.0]	10 (6) [6.9]	1 [0.6]	8 [3] [5.3]
Skeletal findings^b
lumbar ribs (13/14 or 14/14)	23 (13) [13.6]	24 (10) [15.9]	45 (17) [28.2**]	40 (14) [26.7*]^‡	8.5-16.5% (33.3-72.7%)
cervical ribs	0 [0.0]	2 (2) [1.2]	4 (3) [2.5]	0 [0.0]	0.0-2.0% (0.4-4.8%)
complete 14^thribs	0	0	2 (1)	1

^aData represent the number foetuses (litters) examined in each group.

^bData represent the number foetuses (litters) affected in each group, and [foetus affected per litter (%)].

^cData from foetuses processed for skeletal examination.

^dRange of control incidences from 9 studies in Sprague-Dawley rats conducted at the same laboratory over the preceding 18 months. Control animals received olive oil (5 studies), water (1 study), Tween 80 (1 study), or 1% aqueous methylcellulose (2 studies).

* p < 0.05 compared to control

** p < 0.01 compared to control

^‡Significance for trend, p < 0.01

Applicant's summary and conclusion

Conclusions:

In conclusion, D911P has no potential to adversely affect development in the rat. Administration of doses up to 1000 mg/kg/day throughout gestation induced no detectable toxicity in the dams, and resulted only in minor skeletal and visceral variations (non-adverse). There was no evidence of malformations following treatment. The NOAEL for maternal and developmental toxicity is therefore established at 1000 mg/kg/day.