Registration Dossier

Administrative data

Key value for chemical safety assessment

Effects on fertility

Description of key information

Prenatal Developmental Toxicity Dose Range Finder Study in Pregnant Female Sprague Dawley Rats

Doses up to 1000 mg/kg/day had no effect on mortality, physical examinations, cageside observations, body weights, body weight changes, food consumption, gross pathology findings, uterine data, or fetal examination data (external exams).

Link to relevant study records
Reference
Endpoint:
screening for reproductive / developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
June 15, 2017 to September 14, 2017
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment
Qualifier:
according to guideline
Guideline:
other: Internal Laboratory Standard Protocols
Deviations:
not specified
Principles of method if other than guideline:
The Institutional Animal Care and Use Committee (IACUC) of Smithers Avanza approved the study protocol and found it to be in accordance with provisions of the USDA Animal Welfare Act, the PHS Policy on Humane Care and Use of Laboratory Animals, and the US Interagency Research Animal Committee Principles for the Utilization and Care of Research Animals.
GLP compliance:
yes
Limit test:
no
Justification for study design:
The purpose of this study was to provide preliminary data on the potential maternal and/or developmental toxicity of Hatcol 1510 in pregnant Sprague Dawley rats when administered via oral gavage during the embryo-fetal development period. The data from this study was used to establish doses for the definitive prenatal developmental toxicity study.
Specific details on test material used for the study:
No further details specified in the study report.
Species:
rat
Strain:
Sprague-Dawley
Details on species / strain selection:
The rat was selected because it is a standard species for use in toxicology studies.
Sex:
female
Details on test animals or test system and environmental conditions:
Animal Information
Species and Strain: Time-mated female Sprague Dawley Rats
Supplier: Charles River Breeding Labs; Raleigh, NC
Method of Identification: Ear tag/Cage card
Number of Animals Received: 26
Number Used on Study: 24
Age at First Dose: At least 7 weeks
Weight Range at Mating: 214 g – 246 g
Weight Range at First Dose: 230.3 g – 264.7 g
Disposition of Extra Animals: Euthanized

Animals were acclimated to laboratory conditions for at least three days prior to the first dose and released from acclimation by a staff veterinarian. During that time, animals were identified by a temporary number that was recorded on each cage label.

Husbandry Information
Feed: Certified Global Teklad Laboratory Diet 2018 (pellets) was provided ad libitum.
Water: Filtered water was provided ad libitum via an automatic watering system
Bedding: Certified Sani Chips hardwood bedding
Housing: Animals were individually housed in one room in polycarbonate cages suspended on stainless steel racks. Each cage was affixed with a cage card containing pertinent animal and study information.
Temperature Range: 20 to 26 °C
Humidity Range: 30 to 70%
Light Cycle: 12-hour light/12-hour dark, interrupted as necessary for study-related events
Air Changes: Minimum of 10 air changes per hour

The feed was analyzed by the manufacturer for concentrations of specified heavy metals, aflatoxin, chlorinated hydrocarbons, and organophosphates. The water is routinely analyzed for contaminants and specific microbes. The bedding was analyzed by the manufacturer for acceptable levels of heavy metals, aflatoxins, bacteria, yeasts, molds, and organophosphates prior to certification. No contaminants were known to be present in the feed, water, or bedding at levels that might have interfered with achieving the objectives of the study.
Route of administration:
oral: gavage
Vehicle:
peanut oil
Details on exposure:
The neat test and vehicle/control substance were considered 100% pure for formulation purposes.
The vehicle/control substance, peanut oil, was used as received; no formulations were necessary.
Formulations for Groups 2 (50 mg/mL), 3 (150 mg/mL), and 4 (500 mg/mL) were prepared three times throughout the study by adding the appropriate amount of test substance into a pre-calibrated beaker. The required amount of vehicle was added and the solution was mixed until visually uniform. Formulations were refrigerated at 5±3°C until used for further formulating or dosing.
Details on mating procedure:
The vendor shipped 13 females that were bred on 06/14/17 (confirmed 06/15/17), and 13 females that were bred on 06/15/17 (confirmed 6/16/17). Dams should have been bred on 06/13/17 and 06/14/17, respectively. Thus, animals arrived approximately one gestational day early and were therefore dosed from GD 4 to 19 and necropsy was conducted on GD 20.
Analytical verification of doses or concentrations:
no
Duration of treatment / exposure:
The animals were dosed daily on presumed GD 5 to 20 (day of confirmation of mating = GD 0)
Frequency of treatment:
Daily
Details on study schedule:
Not specified
Dose / conc.:
0 mg/kg bw/day (nominal)
Dose / conc.:
100 mg/kg bw/day (nominal)
Dose / conc.:
300 mg/kg bw/day (nominal)
Dose / conc.:
1 000 mg/kg bw/day (nominal)
No. of animals per sex per dose:
6 animals per dose group
Control animals:
yes, concurrent vehicle
Details on study design:
The rat was selected because it is a standard species for use in toxicology studies.
This study was designed to use the fewest number of animals possible, consistent with the objective of the study, ant the scientific needs of the Sponsor, contemporary scientific standards.
The oral route was selected because it is the relevant route of exposure in humans.
The dose levels were selected based on the results from previous toxicology studies performed by the Sponsor at Smithers Avanza under study number 2386-13844.
Animals were initially accepted into the randomization pool based upon body weights collected on GD 3. The suitability of the randomized animals was determined using prestudy body weights and physical examinations. They were assigned to study groups using computer-generated random numbers such that the mean body weight for each group was not statistically different (p ≤ 0.05) from the control mean. Following randomization each study animal was assigned a unique number.
Positive control:
Not required.
Parental animals: Observations and examinations:
Animal Observations/Measurements
Physical Examinations: GD 5, 8, 11, 14, 17 and 21
Cageside Observations: ≥ 2 times daily
Body Weights: GD 5, 8, 11, 14, 17 and 21
Food Consumption: GD 5-8, 8-11, 11-14, 14-17 and 17-21
Cageside observations included observation for mortality, moribundity, general health, and signs of toxicity. Physical examinations included evaluation of skin and fur characteristics, eye and mucous membranes, respiratory, circulatory, autonomic, and central nervous systems, and somatomotor and behavior patterns.
Oestrous cyclicity (parental animals):
Not examined
Sperm parameters (parental animals):
Not examined
Litter observations:
All live fetuses were individually weighed, identified, sexed, and examined for external malformations and variations.
Postmortem examinations (parental animals):
Termination
On presumed GD 21, all surviving dams were euthanized via carbon dioxide inhalation followed by exsanguination and examined as described below.
Although animals were assumed GD 21 at termination, fetal weights (GD 21) were not within historical control range. It was determined that time-mated pregnant animals arrived from the vendor were bred a day earlier than requested. Thus, all dams were actually necropsied on GD 20 rather than GD 21.

Necropsy
Animals were necropsied as soon as possible after the time of death. A gross necropsy, which included examination of the external surface of the body, all orifices, injection site, the cranial, thoracic, and abdominal cavities, and their contents was performed, with special emphasis on structural abnormalities or pathologic changes which might have influenced the pregnancy.

Uterine Examination
The skin was opened with a ventral midline incision to examine mammary tissue and locate any subcutaneous masses. The abdominal cavity was opened, the uterus was excised, and the gravid uterine weight was recorded. Beginning at the distal end of the left uterine horn, the location of viable and nonviable fetuses, early and late resorptions for each uterine horn, and the number of total implantations were recorded. The number of corpora lutea on each ovary was also recorded. The embryonic membrane of each fetus was gently removed. The fetuses were removed from the uterus and the placentae were grossly examined. Each implant was categorized as viable, nonviable, late resorption, or early resorption.
Uteri from females that appeared to be nongravid were opened and placed in 10% ammonium sulfide solution for detection of implantation sites. The foci, if detected, were considered early resorptions, and data from these females were not included in mean calculations and statistics. If no foci were seen, the female was considered to be nonpregnant and data from these females were also not included in mean calculations or statistics.
Postmortem examinations (offspring):
Following completion of the external examination of all fetuses in the litter, each fetus was euthanized via an oral dose of Euthasol® solution (0.01 – 0.05 mL/fetus) and identified individually.
Statistics:
Quantitative data from the treated groups were compared statistically to the data of the control group using one-way Analysis of Variance (ANOVA) techniques. A Shapiro-Wilk test was used to test for normality, followed by the Levene’s test to test the hypothesis of homogeneity of variances. If either the normality or homogeneity test failed (as indicated by a Shapiro-Wilk or Levene’s test p-value ≤ 0.01), the data were transformed using log-transformed values and both the Shapiro-Wilk and Levene’s tests were repeated with the log-transformed values. If the results from either the Shapiro-Wilk or Levene’s test on the log-transformed data failed, the analysis of the data continued using rank-transformed data using Kruskal-Wallis ANOVA. If both the Shapiro-Wilk or Levene’s test were not statistically significant, the ANOVA was performed on the untransformed or log-transformed data, respectively. The Dunnett’s t-test was used to determine which groups differed from the control group. Group comparisons were evaluated at the 0.05 (two-tailed) probability level.
Some quantitative data were analyzed using the Kolmogorov-Smirnov test for normality, the Levene Median test for equal variance, and by one-way Analysis of Variance (ANOVA). If either the normality or equal variance test failed, then the analysis was continued using the non-parametric Kruskal-Wallis ANOVA on rank-transformed data. For parametric data, if the ANOVA indicated statistical significance among experimental groups then the Dunnett’s t-test was used to delineate which groups differed from the control. For non-parametric data, if the ANOVA indicated statistical significance among experimental groups then the Dunn’s test was used to delineate which groups differed from the control. The probability value of less than 0.05 (two-tailed) was used as the critical level of significance for all tests. Statistical analysis for these data was conducted using SigmaStat™ Statistical Software, Version 1.
Reproductive indices:
An analysis of covariance was performed on mean male fetal weight per litter, mean female fetal weight per litter, and mean combined fetal weight per litter (sexes pooled), with the total number of fetuses per litter used as the covariate in all analyses. All tests were conducted at the 0.05 level of significance using the SAS Statistical Analysis System, Version 9 (SAS Institute, Cary, NC).
Offspring viability indices:
Not examined
Clinical signs:
no effects observed
Description (incidence and severity):
Treatment with Hatcol 1510 had no effect on physical examinations, or cageside observations. No abnormalities were observed throughout the study.
Dermal irritation (if dermal study):
not examined
Mortality:
no mortality observed
Description (incidence):
Treatment with Hatcol 1510 had no effect on mortality. All animals survived until the scheduled termination.
Body weight and weight changes:
no effects observed
Description (incidence and severity):
Treatment with Hatcol 1510 had no effect on body weights, body weight changes, or body weight changes adjusted for gravid uterine weight.
No significant differences were noted in mean body weights and body weight changes; means were comparable across groups. Group 4 had a significantly higher mean body weight when adjusted for the gravid uterine weight. No differences were noted in total or percent change when adjusted for the gravid uterine weight.
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
No significant differences were noted; mean food consumption was comparable across groups throughout the study.
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Histopathological findings: non-neoplastic:
not examined
Histopathological findings: neoplastic:
not examined
Other effects:
not specified
Reproductive function: oestrous cycle:
not examined
Reproductive function: sperm measures:
not examined
Reproductive performance:
no effects observed
Description (incidence and severity):
Treatment with Hatcol 1510 had no effect on uterine data. No significant differences were noted in number of corpora lutea, number of implantations, pre-implantation loss, number of intra-uterine deaths, post-implantation loss, or number of live fetuses.
Key result
Dose descriptor:
NOEL
Effect level:
1 000 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
female
Basis for effect level:
other: no effect on maternal mortality, physical examinations, cageside observations, body weights, body weight changes, or food consumption.
Key result
Critical effects observed:
no
Clinical signs:
not examined
Dermal irritation (if dermal study):
not examined
Mortality / viability:
no mortality observed
Body weight and weight changes:
no effects observed
Description (incidence and severity):
No statistically significant differences were observed among Group 1 and the other treatment groups for male, female, or combined male and female fetal weight.
Food consumption and compound intake (if feeding study):
not examined
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Sexual maturation:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
no effects observed
Description (incidence and severity):
Treatment with Hatcol 1510 had no effect on external examinations. No variations or malformations were observed.
Histopathological findings:
not examined
Other effects:
not specified
Behaviour (functional findings):
not examined
Developmental immunotoxicity:
not examined
Key result
Dose descriptor:
NOEL
Generation:
F1
Effect level:
1 000 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: There were no effects on fetal development. There was no difference between the control and treated groups in male and female fetal body weights or in external findings.
Key result
Critical effects observed:
no
Key result
Reproductive effects observed:
no
Lowest effective dose / conc.:
1 000 mg/kg bw/day (nominal)
Treatment related:
no

Individual Animal Disposition and Physical Examinations

Day Numbers Relative to Mating Date

Group

Sex

Animal

Clinical Sign

5

6

11

14

17

21

1

F

22113

No abnormalities Detected

Terminal Kill

X

-

X

-

X

-

X

-

X

-

X

X

22114

No abnormalities Detected

Terminal Kill

X

-

X

-

X

-

X

-

X

-

X

X

22115

No abnormalities Detected

Terminal Kill

X

-

X

-

X

-

X

-

X

-

X

X

22116

No abnormalities Detected

Terminal Kill

X

-

X

-

X

-

X

-

X

-

X

X

22117

No abnormalities Detected

Terminal Kill

X

-

X

-

X

-

X

-

X

-

X

X

22118

No abnormalities Detected

Terminal Kill

X

-

X

-

X

-

X

-

X

-

X

X

2

F

22119

No abnormalities Detected

Terminal Kill

X

-

X

-

X

-

X

-

X

-

X

X

22120

No abnormalities Detected

Terminal Kill

X

-

X

-

X

-

X

-

X

-

X

X

22121

No abnormalities Detected

Terminal Kill

X

-

X

-

X

-

X

-

X

-

X

X

22122

No abnormalities Detected

Terminal Kill

X

-

X

-

X

-

X

-

X

-

X

X

22123

No abnormalities Detected

Terminal Kill

X

-

X

-

X

-

X

-

X

-

X

X

22124

No abnormalities Detected

Terminal Kill

X

-

X

-

X

-

X

-

X

-

X

X

3

F

22125

No abnormalities Detected

Terminal Kill

X

-

X

-

X

-

X

-

X

-

X

X

22126

No abnormalities Detected

Terminal Kill

X

-

X

-

X

-

X

-

X

-

X

X

22127

No abnormalities Detected

Terminal Kill

X

-

X

-

X

-

X

-

X

-

X

X

22128

No abnormalities Detected

Terminal Kill

X

-

X

-

X

-

X

-

X

-

X

X

22129

No abnormalities Detected

Terminal Kill

X

-

X

-

X

-

X

-

X

-

X

X

22130

No abnormalities Detected

Terminal Kill

X

-

X

-

X

-

X

-

X

-

X

X

4

F

22131

No abnormalities Detected

Terminal Kill

X

-

X

-

X

-

X

-

X

-

X

X

22132

No abnormalities Detected

Terminal Kill

X

-

X

-

X

-

X

-

X

-

X

X

22133

No abnormalities Detected

Terminal Kill

X

-

X

-

X

-

X

-

X

-

X

X

22134

No abnormalities Detected

Terminal Kill

X

-

X

-

X

-

X

-

X

-

X

X

22135

No abnormalities Detected

Terminal Kill

X

-

X

-

X

-

X

-

X

-

X

X

22136

No abnormalities Detected

Terminal Kill

X

-

X

-

X

-

X

-

X

-

X

X

Nominal Dose: Group 1 – 0 mg/kg Group 2 – 100 mg/kg         Group 3 – 300 mg/kg         Group 4 – 1000 mg/kg

 

Individual Body Weights (g)

Day Numbers Relative to Mating Date

Group

Sex

Animal

5

8

11

14

17

21

1

F

22113a

22114

22115

22116

22117

22118

253.9

230.5

248.3

250.4

248.6

252.1

269.4

247.2

272.1

256.9

258.3

266.5

279.0

258.1

279.6

281.1

286.4

288.9

279.5

273.6

302.5

298.2

294.3

316.8

278.9

292.0

322.1

319.2

320.3

340.8

289.1

348.1

383.6

380.2

384.8

406.9

Mean

S.D.

N

245.98

8.79

5

260.20

9.55

5

278.82

12.19

5

297.08

15.64

5

318.88

17.43

5

380.72

21.06

5

2

F

22119

22120

22121

22122

22123

22124

250.4

237.8

252.8

238.6

251.6

243.8

281.9

261.6

271.1

247.8

261.6

259.6

299.0

275.3

296.0

270.9

279.9

277.5

315.2

299.2

314.1

286.0

302.7

298.1

339.1

314.3

335.1

311.7

320.0

325.9

406.7

364.8

405.4

356.1

374.6

379.7

Mean

S.D.

N

245.83

6.69

6

263.93

11.52

6

283.10

11.58

6

302.55

10.94

6

324.35

11.09

6

381.22

20.88

6

3

F

22125

22126

22127

22128

22129

22130

250.2

250.8

233.0

253.9

230.3

240.3

273.0

270.2

257.6

268.4

241.0

246.1

289.7

280.0

274.8

289.5

255.7

269.2

277.7

295.7

296.8

316.5

276.5

281.9

320.3

314.5

315.8

334.9

296.0

300.7

374.9

359.3

373.6

393.0

342.3

357.0

Mean

S.D.

N

243.08

10.00

6

259.38

13.43

6

276.48

12.99

6

290.85

15.33

6

313.70

14.00

6

366.68

17.61

6

4

F

22131

22132

22133

22134

22135

22136

236.5

264.7

251.6

253.3

238.0

247.9

256.2

287.6

271.3

264.4

243.0

258.9

267.5

306.2

284.6

290.2

261.4

281.7

284.7

330.4

301.2

316.9

271.5

305.6

299.9

355.8

319.9

331.8

281.6

323.9

355.2

421.9

368.8

390.3

309.3

376.2

Mean

S.D.

N

248.67

10.48

6

263.57

15.08

6

281.93

16.09

6

301.72

21.31

6

318.82

25.72

6

370.28

37.55

6

aAnimal was not pregnant and values were excluded from the mean.

Nominal Dose: Group 1 – 0 mg/kg Group 2 – 100 mg/kg         Group 3 – 300 m/kg           Group 4 – 1000 mg/kg

 

Individual Gross Pathology Observations

Day Numbers Relative to Mating Date

Animal No.

Mode of Death

Death

Observation(s)

Day

(Week)

Group: 1          Sex: Female

22113

 

22114

22115

22116

 

22117

22118

Terminal Kill

 

Terminal Kill

Terminal Kill

Terminal Kill

 

Terminal Kill

Terminal Kill

21

 

21

21

21

 

21

21

(3)

 

(3)

(3)

(3)

 

(3)

(3)

Uterus: IMPLANT SITES NOT EVIDENT, UTERI STAINED IN 10% AMMONIUM SULFIDE

Any remaining protocol required tissues, which have been examined, have no visible lesions

No Visible Lesions

No Visible Lesions

Animal id: ear tag reads “21073”

Any remaining protocol tissues, which have been examined, have no visible lesions

No Visible Lesions

No Visible Lesions

Group: 2          Sex: Female

22119

22120

22121

22122

22123

22124

Terminal Kill

Terminal Kill

Terminal Kill

Terminal Kill

Terminal Kill

Terminal Kill

21

21

21

21

21

21

(3)

(3)

(3)

(3)

(3)

(3)

No Visible Lesions

No Visible Lesions

No Visible Lesions

No Visible Lesions

No Visible Lesions

No Visible Lesions

Group: 3          Sex: Female

22125

22126

22127

22128

22129

22130

Terminal Kill

Terminal Kill

Terminal Kill

Terminal Kill

Terminal Kill

Terminal Kill

21

21

21

21

21

21

(3)

(3)

(3)

(3)

(3)

(3)

No Visible Lesions

No Visible Lesions

No Visible Lesions

No Visible Lesions

No Visible Lesions

No Visible Lesions

Group: 4          Sex: Female

22131

22132

22133

22134

22135

22136

Terminal Kill

Terminal Kill

Terminal Kill

Terminal Kill

Terminal Kill

Terminal Kill

21

21

21

21

21

21

(3)

(3)

(3)

(3)

(3)

(3)

No Visible Lesions

No Visible Lesions

No Visible Lesions

No Visible Lesions

No Visible Lesions

No Visible Lesions

Nominal Dose: Group 1 – 0 mg/kg Group 2 – 100 mg/kg         Group 3 – 300 mg/kg         Group 4 – 1000 mg/kg

 

Individual Uterine Data

Day 21 Relative to Start Date

Dam Number

Number of Corpora Lutea

Number of Implantations

Percent Pre-imp Loss

--Number of Intra-Uterine Deaths--

Percent Post-imp Loss

Number of Live Fetuses

Live Fetuses as Percent of Imp

L

R

L

R

Late Resorption

Early Resorption

Total Deaths

L

R

L

R

L

R

L

R

Group 1:

22113 (NP)

22114

22115

22116

22117

22118

 

6

8

4

7

7

 

5

6

12

8

9

 

6

8

3

7

6

 

5

6

12

8

9

 

0.0

0.0

5.9

0.0

6.3

 

0

0

0

0

0

 

0

0

0

0

0

 

0

0

0

0

0

 

0

0

0

0

1

 

0

0

0

0

0

 

0

0

0

0

1

 

0.0

0.0

0.0

0.0

6.7

 

6

8

3

7

6

 

5

6

13

8

8

 

100.0

100.0

100.0

100.0

93.3

 

32

41

30

41

 

0

0

0

1

0

1

 

30

40

 

TOTAL

Litter Mean

S.D.

N

73

14.6

2.3

5

71

14.2

1.9

5

 

2.4

0

 

 

5

1

0.2

0.4

5

1

0.2

0.4

5

 

1.3

70

14.0

1.9

5

 

98.7

Group 2:

22119

22120

22121

22122

22123

22124

10

5

8

6

6

2

10

6

7

6

5

7

9

3

8

6

6

2

9

6

6

6

5

7

10.0

18.2

6.7

0.0

0.0

0.0

0

0

0

0

0

0

0

0

0

0

0

0

1

0

0

0

0

0

0

0

0

1

0

0

1

0

0

0

0

0

0

0

0

1

0

0

5.6

0.0

0.0

8.3

0.0

0.0

8

3

8

6

6

2

9

6

6

5

5

7

94.4

100.0

100.0

91.7

100.0

100.0

 

37

41

34

39

 

0

0

1

1

1

1

 

33

38

 

TOTAL

Litter Mean

S.D.

N

78

13.0

3.9

6

73

12.2

3.4

6

 

5.8

0

 

 

6

2

0.3

0.5

6

2

0.3

0.5

6

 

2.3

71

11.8

3.1

6

 

97.7

Group 3

22125

22126

22127

22128

22129

22130

8

5

9

6

7

3

5

6

6

8

7

10

8

5

9

6

7

3

5

6

6

8

7

10

0.0

0.0

0.0

0.0

0.0

0.0

0

0

0

1

0

0

0

0

0

0

0

0

0

0

0

1

2

1

0

0

0

0

0

0

0

0

0

2

2

1

0

0

0

0

0

0

0.0

0.0

0.0

14.3

14.3

7.7

8

5

9

4

5

2

5

6

6

8

7

10

100.0

100.0

100.0

85.7

85.7

92.3

 

38

42

38

42

 

1

0

4

0

5

0

 

33

42

 

TOTAL

Litter Mean

S.D.

N

80

13.2

1.4

6

80

13.3

1.4

6

 

0.0

1

0.2

0.4

6

4

0.7

0.8

6

5

0.8

1.0

6

 

6.0

75

12.5

1.4

6

 

94.0

Group 4

22131

22132

22133

22134

22135

22136

9

8

3

7

4

9

6

10

9

7

6

6

9

7

3

6

2

8

5

10

9

6

6

5

6.7

5.6

0.0

14.3

20.0

13.3

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

0

2

0

0

0

0

0

1

0

0

0

0

0

2

0

0

0

0

0

1

0

0.0

0.0

0.0

0.0

37.5

0.0

9

7

3

6

0

8

5

10

9

6

5

5

100.0

100.0

100.0

100.0

62.5

100.0

 

40

44

35

41

 

0

0

2

1

2

1

 

33

40

 

TOTAL

Litter Mean

S.D.

N

84

14.0

2.8

6

76

12.7

2.9

6

 

10.0

0

 

 

6

3

0.5

1.2

6

3

0.5

1.2

6

 

6.3

73

12.2

4.0

6

 

93.8

(NP) = Not Pregnant. These dams are excluded from all group summary calculations

Nominal Dose: Group 1 – 0 mg/kg Group 2 – 100 mg/kg         Group 3 – 300 mg/kg         Group 4 – 1000 mg/kg

 

Mean Fetal Weight per Dam

Day 21 Relative to Start Date

Dam Number

--Number of Live Fetuses--

% Live Male Fetuses

Litter Weight (g)

--Mean Fetal Weight (g)--

Total

Males

Females

Overall

Males

Females

Group 1

22113 (NP)

22114

22115

22116

22117

22118

 

11

14

16

15

14

 

5

6

9

8

10

 

6

8

7

7

4

 

45.5

42.9

56.3

53.3

71.4

 

42.2144

52.6816

64.4874

57.5168

57.4989

 

3.83767

3.76297

4.03046

3.83445

4.10706

 

3.85290

3.78620

4.10857

3.97090

4.18762

 

3.82498

3.74555

3.93004

3.67851

3.90568

TOTAL

Litter Mean

S.D.

N

70

14.0

1.9

5

38

7.6

2.1

5

32

6.4

1.5

5

 

53.9

 

54.87982

8.23656

5

 

3.91452

0.14646

5

 

3.98124

0.16839

5

 

3.81695

0.10605

5

Group 2

22119

22120

22121

22122

22123

22124

17

9

14

11

11

9

6

5

8

8

6

5

11

4

6

3

5

4

35.3

55.6

57.1

72.7

54.5

55.6

60.0439

36.3199

56.4570

44.5799

42.3252

32.4305

3.53199

4.03554

4.03264

4.05272

3.84775

3.60339

3.80777

4.07892

4.11985

4.09495

3.93530

3.62590

3.38157

3.98133

3.91637

3.94010

3.74268

3.57252

TOTAL

Litter Mean

S.D.

N

71

11.8

3.1

6

38

6.3

1.4

6

33

5.5

2.9

6

 

55.1

 

45.35940

10.93137

6

 

3.85067

0.23273

6

 

3.94378

0.19582

6

 

3.75622

0.23809

6

Group 3

22125

22126

22127

22128

22129

22130

13

11

15

12

12

12

8

9

7

5

6

4

5

2

8

7

6

8

61.5

81.8

46.7

41.7

50.0

33.3

51.3679

46.3479

56.6603

48.8336

45.9274

36.5772

3.95138

4.21345

3.77735

4.06947

3.82728

3.04810

4.02796

4.26233

3.91691

4.14382

3.94605

2.86098

3.82884

3.99345

3.65524

4.01636

3.70582

3.14166

TOTAL

Litter Mean

S.D.

N

75

12.5

1.4

6

39

6.5

1.9

6

36

6.0

2.3

6

 

52.5

 

47.61905

6.68840

6

 

3.81450

0.40788

6

 

3.85968

0.50583

6

 

3.72401

0.32036

6

Group 4

22131

22132

22133

22134

22135

22136

14

17

12

12

5

12

8

9

3

7

3

5

6

8

9

5

2

8

57.1

52.9

25.0

58.2

60.0

38.5

44.5179

64.2901

46.6073

44.7658

20.0582

47.5946

3.17985

3.78177

3.88394

3.73048

4.01164

3.66112

3.27755

3.89286

4.05513

3.81910

4.04953

3.81596

3.04958

3.65680

3.82688

3.60642

3.95480

3.56435

TOTAL

Litter Mean

S.D.

N

72

12.2

4.0

6

35

5.8

2.6

6

38

6.3

2.6

6

 

48.6

 

44.63898

14.16349

6

 

3.70813

0.28644

6

 

3.81836

0.28549

6

 

3.60981

0.31122

6

(NP) = Not Pregnant

Nominal Dose: Group 1 – 0 mg/kg Group 2 – 100 mg/kg         Group 3 – 300 mg/kg         Group 4 – 1000 mg/kg

Conclusions:
Oral gavage administration of decanoic acid, mixed esters with heptanoic acid, octanoic acid and trimethylolpropane (Hatcol 1510) at doses up to 1000 mg/kg/day from implantation through the end of gestation had no effect on maternal mortality, physical examinations, cageside observations, body weights, body weight changes, or food consumption.
There were no effects on fetal development. There was no difference between the control and treated groups in male and female fetal body weights or in external findings.
Decanoic acid, mixed esters with heptanoic acid, octanoic acid and trimethylolpropane (Hatcol 1510) given orally at doses up to 1000 mg/kg/day to pregnant female Sprague Dawley rats during prenatal development did not result in any adverse effects on the development of the embryo/fetus.
Based on these results, the dose levels of 100, 300, and 1000 mg/kg/day were considered appropriate for the decanoic acid, mixed esters with heptanoic acid, octanoic acid and trimethylolpropane (Hatcol 1510) definitive prenatal developmental toxicity study in rats.
Executive summary:

Decanoic acid, mixed esters with heptanoic acid, octanoic acid and trimethylolpropane (Hatcol 1510): Prenatal Developmental Toxicity Dose Range Finder Study in Pregnant Female Sprague Dawley Rats

 

The purpose of this study was to provide preliminary data on the potential maternal and/or developmental toxicity of decanoic acid, mixed esters with heptanoic acid, octanoic acid and trimethylolpropane (Hatcol 1510) in pregnant Sprague Dawley rats when administered via oral gavage during the embryo-fetal development period. The data from this study was used to establish doses for the definitive prenatal developmental toxicity study.

 

Twenty-four time-mated female Sprague Dawley rats were randomly assigned to four groups (6 females/group). Animals were administered control substance (peanut oil) or decanoic acid, mixed esters with heptanoic acid, octanoic acid and trimethylolpropane (Hatcol 1510) at 0, 100, 300, or 1000 mg/kg once daily via oral gavage from presumed gestational day (GD) 5 to 20. Animals were subjected to a full gross necropsy on presumed GD 21.

 

Parameters evaluated during the study included mortality, physical examinations, cageside observations, body weights, body weight changes, food consumption, gross pathology findings, uterine data, and fetal examination data (external exams).

 

Treatment with Hatcol 1510 at doses up to 1000 mg/kg/day had no effect on mortality, physical examinations, cageside observations, body weights, body weight changes, food consumption, gross pathology findings, uterine data, or fetal examination data (external exams).

 

There were no differences between the control and treated groups in male and female fetal body weights or in external findings.

 

Decanoic acid, mixed esters with heptanoic acid, octanoic acid and trimethylolpropane (Hatcol 1510) given orally at doses up to 1000 mg/kg/day to pregnant female Sprague Dawley rats during prenatal development did not result in any adverse effects on the development of the embryo/fetus.

 

Based on these results, the dose levels of 100, 300, and 1000 mg/kg/day were considered appropriate for the decanoic acid, mixed esters with heptanoic acid, octanoic acid and trimethylolpropane (Hatcol 1510) definitive prenatal developmental toxicity study in rats.

Effect on fertility: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
1 000 mg/kg bw/day
Study duration:
subacute
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

The purpose of the study was to provide preliminary data on the potential maternal and/or developmental toxicity of decanoic acid, mixed esters with heptanoic acid, octanoic acid and trimethylolpropane (Hatcol 1510) in pregnant Sprague Dawley rats when administered via oral gavage during the embryo-fetal development period. The data from this study was used to establish doses for the definitive prenatal developmental toxicity study.

Twenty-four time-mated female Sprague Dawley rats were randomly assigned to four groups (6 females/group). Animals were administered control substance (peanut oil) or decanoic acid, mixed esters with heptanoic acid, octanoic acid and trimethylolpropane (Hatcol 1510) at 0, 100, 300, or 1000 mg/kg once daily via oral gavage from presumed gestational day (GD) 5 to 20. Animals were subjected to a full gross necropsy on presumed GD 21.

Parameters evaluated during the study included mortality, physical examinations, cageside observations, body weights, body weight changes, food consumption, gross pathology findings, uterine data, and fetal examination data (external exams).

 

Treatment with Hatcol 1510 at doses up to 1000 mg/kg/day had no effect on mortality, physical examinations, cageside observations, body weights, body weight changes, food consumption, gross pathology findings, uterine data, or fetal examination data (external exams).

There were no differences between the control and treated groups in male and female fetal body weights or in external findings.

Decanoic acid, mixed esters with heptanoic acid, octanoic acid and trimethylolpropane (Hatcol 1510) given orally at doses up to 1000 mg/kg/day to pregnant female Sprague Dawley rats during prenatal development did not result in any adverse effects on the development of the embryo/fetus.

 

Based on these results, the dose levels of 100, 300, and 1000 mg/kg/day were considered appropriate for the decanoic acid, mixed esters with heptanoic acid, octanoic acid and trimethylolpropane (Hatcol 1510) definitive prenatal developmental toxicity study in rats.

Effects on developmental toxicity

Description of key information

Prenatal developmental toxicology study in Sprague Dawley rats

no-adverse-observed-effect-level (NOAEL) for maternal and embryo-fetal developmental toxicity when Hatcol 1510 was administered via oral gavage over the entire period of gestation (GD 5-20) is 1000 mg/kg, the highest dose level tested.

Link to relevant study records
Reference
Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
July 26, 2017 to April 24, 2018
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Deviations:
yes
Remarks:
See "Principles of method if other than guidelines" for details
Principles of method if other than guideline:
Deviations
The following deviations were noted:
Animals were randomized prior to arrival; therefore no physical examinations could be performed prior to randomization.
Back-up samples for concentration verification were inadvertently discarded prior to authorization. A reconciliation of the samples and disposition was performed on March 14, 2018 and all samples were accounted for at that time.
Verification of internal sex for 22543 (2f) fetus #12 and 22531 (1f) fetus #11 was not documented at the time of eviscerations.
The above-mentioned deviations did not impact this study, nor did they affect the quality or integrity of the study or the interpretation of the results in the report.
GLP compliance:
yes
Limit test:
yes
Specific details on test material used for the study:
No further details specified in the study report.
Species:
rat
Strain:
Sprague-Dawley
Details on test animals or test system and environmental conditions:
Species and Strain: Time-mated female Sprague Dawley Rats
Supplier: Charles River Breeding Labs; Raleigh, NC
Method of Identification: Ear tag/Cage card
Number of Animals Received: 100
Number Used on Study: 100
Age at First Dose: 10 – 12 weeks
Weight Range at Mating: 200 g – 249 g
Weight Range at First Dose: 217.9 g – 282.0 g
Four cohorts of 25 time-mated females arrived on their respective Gestation Day (GD) 1, 2, 3, or 4. Animals were acclimated to laboratory conditions for at least two days prior to the first dose and released from acclimation by a staff veterinarian. During that time, animals were identified by a temporary number that was recorded on each cage label.

Husbandry
Feed: Certified Global Teklad Laboratory Diet 2018 (pellets) was provided ad libitum.
Water: Filtered water was provided ad libitum via an automatic watering system or provided a water bottle as needed.
Bedding: Certified Sani Chips® hardwood bedding
Housing: Animals were individually housed in one room in polycarbonate cages suspended on stainless steel racks. Each cage was affixed with a cage card containing pertinent animal and study information.
Temperature Range: 20 to 26°C
Humidity Range: 30 to 70%
Light Cycle: 12-hour light/12-hour dark, interrupted as necessary for study-related events
Air Changes: Minimum of 10 air changes per hour
The feed was analyzed by the manufacturer for concentrations of specified heavy metals, aflatoxin, chlorinated hydrocarbons, and organophosphates. The water is routinely analyzed for contaminants and specific microbes. The bedding was analyzed by the manufacturer for acceptable levels of heavy metals, aflatoxins, bacteria, yeasts, molds, and organophosphates prior to certification. No contaminants were known to be present in the feed, water, or bedding at levels that might have interfered with achieving the objectives of the study.
Environmental controls were set to maintain animal room conditions. Actual temperature and relative humidity in the animal room were monitored continuously by a computerized system. All environmental parameters were maintained within the protocol requirement.
Route of administration:
oral: gavage
Vehicle:
peanut oil
Details on exposure:
The neat test and vehicle/control substance were considered 100% pure for formulation purposes.
The vehicle/control substance, peanut oil, was used as received; no formulations were necessary.
Formulations for Groups 2 (50 mg/mL), 3 (150 mg/mL), and 4 (500 mg/mL) were prepared two times throughout the study by adding the appropriate amount of test substance into a pre-calibrated beaker. The required amount of vehicle was added and the solution was inverted approximately 30 times. Formulations were refrigerated at 5±3°C until used for dosing.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Samples (2 mL, in duplicate) were collected from each test substance formulation and the vehicle/control substance and stored under conditions set to maintain 5±3°C. One set of samples from the first and last mixes were shipped on ice to the Sponsor’s designee, for analysis. Please refer to the attached analytical report below for further information and confirmation.
Details on mating procedure:
Four cohorts of 25 time-mated females arrived on their respective Gestation Day (GD) 1, 2, 3, or 4.
No further information specified in the study report.
Duration of treatment / exposure:
From presumed GD 5 to 20.
Frequency of treatment:
The animals were dosed once daily.
Duration of test:
15 days.
Dose / conc.:
0 mg/kg bw/day (nominal)
Dose / conc.:
100 mg/kg bw/day (nominal)
Dose / conc.:
300 mg/kg bw/day (nominal)
Dose / conc.:
1 000 mg/kg bw/day (nominal)
No. of animals per sex per dose:
25 females per group
Control animals:
yes, concurrent vehicle
Details on study design:
The rat was selected because it is a standard species for use in toxicology studies. Rats were also used as a rodent species per current OECD testing guideline 414.
This study was designed to use the fewest number of animals possible, consistent with the objective of the study, the scientific needs of the Sponsor, contemporary scientific standards, and in consideration of applicable regulatory requirements.
The oral route was selected because it is the relevant route of exposure in humans.
The dose levels were selected based on the results from previous toxicology studies performed by the Sponsor at Smithers Avanza under study number 2386-13843.

Group Assignment and Doses

Animals were initially accepted into the randomization pool based upon GD 0 body weights provided by the vendor. The suitability of the randomized animals was determined using vendor-supplied body weights and physical examinations. They were assigned to study groups using computer-generated random numbers such that the mean body weight for each group was not statistically different (p ≤ 0.05) from the control mean. Following randomization each study animal was assigned a unique number.
Females were assigned to groups (6 or 7/cohort) as shown in the table below

Dose Administration

The animals were dosed once daily on presumed GD 5 to 20 (day of confirmation of mating = GD 0) via oral gavage at a volume of 2.0 mL/kg. Dose volumes were based on the most recent body weights. The first day of dosing was designated as SD 5 for each animal.
Excess formulations were disposed of in accordance with company SOPs, appropriate regulatory requirements, and/or information contained in the Material Safety Data Sheets.

Maternal examinations:
Physical Examinations: GD 5, 8, 11, 14, 17, and 21
Cageside Observations: > 2 times daily
Body Weights: GD 5, 8, 11, 14,17, and 21
Food Consumption: GD 5-8, 8-11, 11-14, 14-17, and 17-21
Cageside observations included observation for mortality, moribundity, general health, and signs of toxicity. Physical examinations included evaluation of skin and fur characteristics, eye and mucous membranes, respiratory, circulatory, autonomic, and central nervous systems, and somatomotor and behavior patterns.

Termination
On presumed GD 21, all surviving dams were euthanized via carbon dioxide inhalation followed by exsanguination and examined as described below.

Necropsy
Animals were necropsied as soon as possible after the time of death. A gross necropsy, which included examination of the external surface of the body, all orifices, the cranial, thoracic, and abdominal cavities, and their contents was performed, with special emphasis on structural abnormalities or pathologic changes which might have influenced the pregnancy. Representative tissues from the first control female were preserved.
Ovaries and uterine content:
Uterine Examination
The skin was opened with a ventral midline incision to examine mammary tissue and locate any subcutaneous masses. The abdominal cavity was opened, the uterus was excised, and the gravid uterine weight was recorded. Beginning at the distal end of the left uterine horn, the location of viable and nonviable fetuses, early and late resorptions for each uterine horn, and the number of total implantations were recorded. The number of corpora lutea on each ovary was also recorded. The embryonic membrane of each fetus was gently removed. The fetuses were removed from the uterus and the placentae were grossly examined. Each implant was categorized as viable, nonviable, late resorption, or early resorption.
Uteri from females that appeared to be nongravid were opened and placed in 10% ammonium sulphide solution for detection of implantation sites. The foci, if detected, were considered early resorptions, and data from these females were not included in mean calculations and statistics. If no foci were seen, the female was considered to be nonpregnant and data from these females were also not included in mean calculations or statistics.
Fetal examinations:
Fetal Examinations
All live fetuses were individually weighed, identified, sexed, and examined for external malformations and variations. Following completion of the external examination of all fetuses in the litter, each fetus was euthanized via an oral dose of Euthasol® solution (0.01 – 0.05 mL/fetus) and identified individually.
Approximately one-half of the fetuses in each litter were examined viscerally by fresh tissue dissection. The internal sex was recorded. The heads were preserved in Bouin’s and examined by Wilson’s sectioning. The remaining fetuses had the internal sex recorded and then were eviscerated, preserved, stained with Alizarin Red S and Alcian Blue, and examined for skeletal abnormalities. Fetal findings were classified as malformations or developmental variations.
Statistics:
Electronic data collection, including randomization, dose formulations and dispensing, dosing, animal husbandry, environmental enrichment, physical examinations, cageside observations, body weights, body weight changes, food consumption, uterine and pregnancy data, gross pathology, fetal examination data, and statistical analysis was performed using Provantis™ Version 8 (Instem LSS, Limited; Stone, UK). Environmental monitoring was performed using Rees Environmental Monitoring System (Rees Scientific Corporation; Trenton, NJ). Statistical analysis on reproductive and developmental parameters (as deemed appropriate) were conducted using SigmaStat™ Statistical Software, Version 1 (Jandel Scientific, San Rafael, CA).
Indices:
An analysis of covariance was performed on mean male fetal weight per litter, mean female fetal weight per litter, and mean combined fetal weight per litter (sexes pooled), with the total number of fetuses pre litter used as the covariate in all analyses.
Clinical signs:
no effects observed
Description (incidence and severity):
Treatment with Hatcol 1510 had no effect on physical examinations, or cageside observations. No abnormalities were observed during cageside observations throughout the study. Incidental findings during physical examinations included Animal 22531 (1f) exhibiting alopecia of the right shoulder from GD 5 to 17 and Animal 22588 (3f) was pale and had an apparent prolapsed vagina prior to necropsy on GD 21 due to complications with the pregnancy and/or parturition.
Dermal irritation (if dermal study):
not examined
Mortality:
no mortality observed
Description (incidence):
Treatment with Hatcol 1510 had no effect on mortality. All animals survived until the scheduled termination.
Body weight and weight changes:
no effects observed
Description (incidence and severity):
Treatment with Hatcol 1510 had no effect on body weights. Group 4 females (1000 mg/kg) had lower mean body weights from GD 11 to GD 21, but no significant differences were noted.
Mean body weight changes in Group 4 were significantly less compared to the control from GD 8 to 11, 14 to 17, and 17 to 21. Group 4 had a significantly lower mean absolute change from GD 5 to 21. Mean gravid uterine weight was also significantly lower for Group 4 females compared to the control. No differences were noted in body weight change when adjusted for the gravid uterine weight. Therefore, there was no effect on mean maternal body weight changes.
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
Treatment with Hatcol 1510 had no effect on food consumption. No significant differences were noted. Mean food consumption was comparable across groups.
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
no effects observed
Description (incidence and severity):
Treatment with Hatcol 1510 had no effect on gross observations. No visible lesions were noted for any dam.
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
not examined
Histopathological findings: neoplastic:
not examined
Other effects:
no effects observed
Description (incidence and severity):
Animal 22533 (1f) exhibited body weight loss and low food consumption from GD 11 to 14 and was examined/treated by the staff veterinarian. A diet gel recovery pack was provided in the animal’s cage on GD 14 and GD 17. Conditions resolved by the end of the treatment and these observations were considered incidental.
Details on results:
Treatment with Hatcol 1510 had no effect on mortality, physical examinations, or cageside observations. All animals survived until the scheduled termination and no abnormalities were observed during cageside observations throughout the study.
Number of abortions:
not specified
Description (incidence and severity):
No significant differences were noted
Pre- and post-implantation loss:
no effects observed
Description (incidence and severity):
Treatment with Hatcol 1510 had no effect on uterine data. No significant differences were noted in the number of corpora lutea, number of implantations, pre-implantation loss, number of intra-uterine deaths, post-implantation loss, or number of live fetuses.
Total litter losses by resorption:
not specified
Description (incidence and severity):
No significant differences were noted
Early or late resorptions:
not specified
Description (incidence and severity):
No significant differences were noted
Dead fetuses:
no effects observed
Description (incidence and severity):
Treatment with Hatcol 1510 had no effect on uterine data. No significant differences were noted in the number of corpora lutea, number of implantations, pre-implantation loss, number of intra-uterine deaths, post-implantation loss, or number of live fetuses.
Changes in pregnancy duration:
not specified
Description (incidence and severity):
No significant differences were noted
Changes in number of pregnant:
no effects observed
Description (incidence and severity):
Treatment with Hatcol 1510 had no effect on uterine data. No significant differences were noted in the number of corpora lutea, number of implantations, pre-implantation loss, number of intra-uterine deaths, post-implantation loss, or number of live fetuses.
Other effects:
no effects observed
Description (incidence and severity):
Stability data demonstrates that the formulated test article was stable under refrigerated storage (2-4°C) for a period of 20 days.

Dose concentration analysis confirmed that the test article was properly formulated (means of the first and last preparations were 97.2% - 116% of the target concentrations and the %CV was ≤10%). Test substance formulations were solutions; therefore, homogeneity sampling and analysis was not required. Test article was not detected in the control formulation.
Details on maternal toxic effects:
No material toxic effects were noted.
Key result
Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day (nominal)
Based on:
test mat.
Basis for effect level:
other: maternal and embryo-fetal developmental toxicity
Key result
Abnormalities:
no effects observed
Fetal body weight changes:
no effects observed
Description (incidence and severity):
Hatcol 1510 had no effect on fetal body weight. No statistically significant differences were observed among Group 1 and the other treatment groups for male, female or combined male and female fetal body weight. No trend was noted in the results.
Reduction in number of live offspring:
no effects observed
Description (incidence and severity):
Treatment with Hatcol 1510 had no effect on uterine data. No significant differences were noted in the number of corpora lutea, number of implantations, pre-implantation loss, number of intra-uterine deaths, post-implantation loss, or number of live fetuses.
Changes in sex ratio:
not specified
Description (incidence and severity):
No significant differences were noted
Changes in litter size and weights:
not specified
Description (incidence and severity):
No significant differences were noted
Changes in postnatal survival:
not specified
Description (incidence and severity):
No significant differences were noted
External malformations:
effects observed, non-treatment-related
Description (incidence and severity):
One fetus in Group 3 (300 mg/kg) had a short tail (external malformation). This finding was considered not to be test substance related. No other abnormalities were observed.
Skeletal malformations:
effects observed, non-treatment-related
Description (incidence and severity):
One malformation (absent caudal, lumbar, and sacral vertebra) was observed in one Group 3 fetus (Dam 22565), which corresponded to the external observation of short tail. No other malformations were observed.
The variations observed were as follows: 30 fetuses (10 litters) in Group 1, 21 fetuses (9 litters) in Group 2, 24 fetuses (11 litters) in Group 3, and 27 fetuses (11 litters) in Group 4. The defects were observed in the ribs (rudimentary, supernumerary, or cervical ribs), sternebrae (unossified), centrum (unossified or incomplete ossification), or exoccipital (unossified).
The centrum variations were observed in 26 fetuses (7 litters) in Group 1, 18 fetuses (6 litters) in Group 2, 15 fetuses (6 litters) in Group 3, and 22 fetuses (9 litters) in Group 4. The rib variations were observed in 2 fetuses (2 Litters) in Group 1, 3 fetuses (3 litters) in Group 2, 1 fetus (1 litter) in Group 3, and 6 fetuses (5 litters) in Group 4. The sternabrae variations were observed in 3 fetuses (2 litters) in Group 1, 1 fetus (1 litter) in Group 2, 7 fetuses (3 litters) in Group 3, and 1 fetus (1 litter) in Group 4.
All variations were considered with in normal range and were not considered test-substance related. No other variations or malformations were observed.
Visceral malformations:
effects observed, non-treatment-related
Description (incidence and severity):
One fetus in Group 4 (1000 mg/kg) had a left-sided umbilical artery. This variation is a common finding in historical control data and was not considered to be test substance related. No other variations or malformations were observed.
Other effects:
effects observed, non-treatment-related
Description (incidence and severity):
Treatment with Hatcol 1510 had no effect on external, visceral, skeletal, or bouin’s head examinations. Only one malformation (short tail with corresponding skeletal changes) was observed in this study in a Group 3 fetus. This malformation was not considered test substance related due to there being no corresponding dose-response or fetal or maternal body weight changes that would correspond with an effect. Additionally, this malformation has been previously described in historical control data from Sprague Dawley rats (Charles River Historical Control Data, Horsham, PA).

Head Examinations
No variations or malformations were observed in head examinations.
Details on embryotoxic / teratogenic effects:
The total number of fetuses showing defects (malformations and variations) was 30 (9.6%) for Group 1, 21 (6.5%) for Group 2, 25 (7.9%) for Group 3, and 28 (10.0%) for Group 4.
Key result
Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
not specified
Basis for effect level:
other: embryo-fetal developmental toxicity
Key result
Abnormalities:
no effects observed
Key result
Developmental effects observed:
no
Lowest effective dose / conc.:
1 000 mg/kg bw/day (nominal)
Treatment related:
no

Summary of Animal Disposition and Physical Examinations

 

Day Numbers Relative to Mating Dose

Animal Disposition                              Group:

1

2

3

4

Sex: Female

 

Terminal Kill

           Number of Animals

           Days from – to

 

25

21  21

 

25

21  21

 

25

21  21

 

25

21  21

Physical Examinations                        Group:

1

2

3

4

Sex: Female

 

Alopecia

           Number of Observations

           Number of Animals

           Days from – to

 

5

1

5  17

 

-

-

-

 

-

-

-

 

-

-

-

Apparent Prolapsed Vagina

           Number of Observations

           Number of Animals

           Days from – to

 

-

-

-

 

-

-

-

 

1

1

21  21

 

-

-

-

Pale

           Number of Observations

           Number of Animals

           Day from – to

 

-

-

-

 

-

-

-

 

1

1

21  21

 

-

-

-

Nominal Dose: Group 1 – 0 mg/kg Group 2 – 100 mg/kg         Group 3 – 300 mg/kg         Group 4 – 1000 mg/kg

 

Summary of Body Weights (g)

 

Day Numbers Relative to Mating Date

Group

Sex

 

5

8

11

14

17

21

1

F

Mean

S.D.

N

244.30

13.72

24

261.55

16.04

24

282.51

18.08

24

299.84

21.88

24

333.15

26.25

24

394.15

35.53

24

2

F

Mean

S.D.

N

244.09

12.87

25

261.58

15.19

25

282.31

16.49

25

301.71

18.93

25

333.03

21.39

25

396.69

26.08

25

3

F

Mean

S.D.

N

241.67

14.50

25

259.70

15.48

25

279.65

17.93

25

298.94

20.61

25

331.66

22.80

25

390.90

32.85

25

4

F

Mean

S.D.

N

243.35

12.30

24

259.35

14.36

24

277.31

17.07

24

293.86

18.74

24

322.83

23.39

24

376.17

28.36

24

Non-pregnant animals were excluded from summary calculations and statistical analyses

Nominal Dose: Group 1 – 0 mg/kg Group 2 – 100 mg/kg         Group 3 – 300 mg/kg         Group 4 – 1000 mg/kg

 

Summary of Body Weight Changes (g)

 

Day Numbers Relative to Mating Date

 

 

Absolute Change

Percent Change

Group

Sex

From:

To:

5

8

8

11

11

14

14

17

17

21

5

21

5

21

1

F

Mean

S.D.

N

17.25

3.83

24

20.96

6.15

24

17.22

6.15

24

33.31

6.24

24

61.00

10.82

24

149.85

23.90

24

61.13

7.52

24

2

F

Mean

S.D.

N

17.50

5.90

25

20.73

4.82

25

19.40

5.74

25

31.32

6.14

25

63.66

7.15

25

152.60

18.07

25

62.53

6.82

25

3

F

Mean

S.D.

N

18.02

10.10

25

19.96

3.82

25

19.29

7.01

25

32.71

5.27

25

59.24

15.84

25

149.23

26.66

25

61.85

11.76

25

4

F

Mean

S.D.

N

16.21

5.45

24

19.96*

4.26

24

16.55

5.96

24

28.97*

6.97

24

53.34*

9.03

24

133.03*

20.79

24

54.67

7.84

24

* - Significantly different from the control value. P < 0.05

Non-pregnant animals were excluded from summary calculations and statistical analyses

Nominal Dose: Group 1 – 0 mg/kg Group 2 – 100 mg/kg         Group 3 – 300 mg/kg         Group 4 – 1000 mg/kg

 

Summary of Body Weight Changes Adjusted for Gravid Uterine Weight (g)

 

Day Numbers Relative to Mating Date

Group

 

Gravid Uterus Weight

Terminal Body Weight

Adjusted Body Weight

Total

Body Weight Change

GD 5-21

Adjusted

Body Weight Change

GD 5-21

1

Mean

S.D.

N

105.40

14.61

24

394.15

35.53

24

288.75

27.08

24

149.85

23.90

24

44.45

16.17

24

2

Mean

S.D.

N

104.36

11.07

25

396.69

26.08

25

292.32

20.36

25

152.60

18.07

25

48.24

14.93

25

3

Mean

S.D.

N

102.28

15.10

25

390.90

32.85

25

288.62

22.32

25

149.23

26.86

25

46.94

16.10

25

4

Mean

S.D.

N

92.48*

14.46

24

376.17

28.36

24

283.69

22.60

24

133.03*

20.79

24

40.55

16.29

24

* - Significantly different from the control value, p < 0.05

Non-pregnant animals were excluded from summary calculations and statistical analyses

Formulae for calculations:Adjusted Body Weight = Terminal Body Weight – Gravid Uterus Weight

                                               Adjusted Body Weight Change = Total Body Weight Change (GD 5 to 21) – Gravid Uterus Weight

Nominal Dose: Group 1 – 0 mg/kg Group 2 – 100 mg/kg         Group 3 – 300 mg/kg         Group 4 – 1000 mg/kg

 

Summary of Gross Pathology Observations

Terminal Necropsy: GD 21

FEMALES

Group:

Number of Animals:

1

(25)

2

(25)

3

(25)

4

(25)

Adrenal gland;

           Submitted

           No Visible Lesions

 

(1)

1

 

(0)

0

 

(0)

0

 

(0)

0

Aorta;

           Submitted

           No Visible Lesions

 

(1)

1

 

(0)

0

 

(0)

0

 

(0)

0

Cecum;

           Submitted

           No Visible Lesions

 

(1)

1

 

(0)

0

 

(0)

0

 

(0)

0

Cervix;

           Submitted

           No Visible Lesions

 

(1)

1

 

(0)

0

 

(0)

0

 

(0)

0

Colon;

           Submitted

           No Visible Lesions

 

(1)

1

 

(0)

0

 

(0)

0

 

(0)

0

Duodenum;

           Submitted

           No Visible Lesions

 

(1)

1

 

(0)

0

 

(0)

0

 

(0)

0

Esophagus;

           Submitted

           No Visible Lesions

 

(1)

1

 

(0)

0

 

(0)

0

 

(0)

0

Heart;

           Submitted

           No Visible Lesions

 

(1)

1

 

(0)

0

 

(0)

0

 

(0)

0

Ileum;

           Submitted

           No Visible Lesions

 

(1)

1

 

(0)

0

 

(0)

0

 

(0)

0

Jejunum;

           Submitted

           No Visible Lesions

 

(1)

1

 

(0)

0

 

(0)

0

 

(0)

0

Kidney;

           Submitted

           No Visible Lesions

 

(1)

1

 

(0)

0

 

(0)

0

 

(0)

0

Liver;

           Submitted

           No Visible Lesions

 

(1)

1

 

(0)

0

 

(0)

0

 

(0)

0

Lung;

           Submitted

           No Visible Lesions

 

(1)

1

 

(0)

0

 

(0)

0

 

(0)

0

Mammary gland;

           Submitted

           No Visible Lesions

 

(1)

1

 

(0)

0

 

(0)

0

 

(0)

0

Ovary;

           Submitted

           No Visible Lesions

 

(1)

1

 

(0)

0

 

(0)

0

 

(0)

0

Oviduct;

           Submitted

           No Visible Lesions

 

(1)

1

 

(0)

0

 

(0)

0

 

(0)

0

Pancreas;

           Submitted

           No Visible Lesions

 

(1)

1

 

(0)

0

 

(0)

0

 

(0)

0

Parathyroid gland;

           Submitted

           No Visible Lesions

 

(1)

1

 

(0)

0

 

(0)

0

 

(0)

0

Rectum;

           Submitted

           No Visible Lesions

 

(1)

1

 

(0)

0

 

(0)

0

 

(0)

0

Spleen;

           Submitted

           No Visible Lesions

 

(1)

1

 

(0)

0

 

(0)

0

 

(0)

0

Stomach;

           Submitted

           No Visible Lesions

 

(1)

1

 

(0)

0

 

(0)

0

 

(0)

0

Thymus;

           Submitted

           No Visible Lesions

 

(1)

1

 

(0)

0

 

(0)

0

 

(0)

0

Thyroid gland;

           Submitted

           No Visible Lesions

 

(1)

1

 

(0)

0

 

(0)

0

 

(0)

0

Trachea;

           Submitted

           No Visible Lesions

 

(1)

1

 

(0)

0

 

(0)

0

 

(0)

0

Urinary bladder;

           Submitted

           No Visible Lesions

 

(1)

1

 

(0)

0

 

(0)

0

 

(0)

0

Uterus;

           Submitted

           No Visible Lesions

 

(1)

1

 

(0)

0

 

(0)

0

 

(0)

0

Vagina;

           Submitted

           No Visible Lesions

           No correlate with gross observation

 

(1)

1

0

 

(0)

0

0

 

(0)

0

1

 

(0)

0

0

Nominal Dose: Group 1 – 0 mg/kg Group 2 – 100 mg/kg         Group 3 – 300 mg/kg         Group 4 – 1000 mg/kg

 

Summary of Pregnancy Data

 

Day 21 Relative to Mating Date

Group:

1

2

3

4

Number of animals in group

25

25

25

25

Not Pregnant (%)

           Died/Killed

           Survived to scheduled kill

1 (4.0)

0

1

0 (0.0)

0

0

0 (0.0)

0

0

1 (4.0)

0

1

Pregnant (%)

           Died/Killed/Aborted

           With total resorption

           With live foetus at scheduled kill

24 (96.0)

0

0

24

25 (100.0)

0

0

25

25 (100.0)

0

0

25

24 (96.0)

0

0

24

Numbers in parentheses are percents

Nominal Dose: Group 1 – 0 mg/kg Group 2 – 100 mg/kg         Group 3 – 300 mg/kg         Group 4 – 1000 mg/kg

 

Summary of Uterine Data

Day 21 Relative to Mating Date

Group:

 

1

2

3

4

Number of females with live fetuses at scheduled kill

 

24

25

25

24

Number of corpora lutea

Number of corpora lutea per female

 

Mean

S.D.

N

344

14.2

2.2

24

365

14.6

3.0

25

351

14.0

2.0

25

335

14.0

4.1

24

Number of implantations

Number of implantations per female

 

Mean

S.D.

N

314

13.1

1.9

24

326

13.0

1.5

25

329

13.2

1.6

25

288

12.0

2.1

24

Pre-implantation loss

Rate per female including total resorptions

 

Mean

S.D.

N

30

0.08

0.12

24

39*

0.08

0.14

25

22

0.06

0.09

25

47

0.11

0.14

24

% loss including total resorptions

 

8.72

10.68

6.27

14.03

Number of Intra-Uterine Deaths – Late resorptions

Number per female including total resorptions

 

Mean

S.D.

N

0

0.0

0.0

24

0

0.0

0.0

25

2

0.1

0.4

25

0

0.0

0.0

24

Number of Intra-Uterine Deaths – Early resorptions

Number per female including total resorption

 

Mean

S.D.

N

3

0.1

0.3

24

4

0.2

0.4

25

6

0.2

0.5

25

7

0.3

0.9

24

Number of Intra-Uterine Deaths – Dead fetus

Number per female including resorptions

 

Mean

S.D.

N

0

0.0

0.0

24

0

0.0

0.0

25

5

0.2

1.0

25

0

0.0

0.0

24

Post-implantation loss

Rate per female including total resorptions

 

Mean

S.D.

N

3

0.1

0.3

24

4

0.1

0.3

25

13

0.04

0.11

25

7

0.02

0.06

24

% loss including total resorptions

 

0.96

1.23

3.95

2.43

Non-pregnant animals were excluded form means.

Nominal Dose: Group 1 – 0 mg/kg Group 2 – 100 mg/kg         Group 3 – 300 mg/kg         Group 4 – 1000 mg/kg

 

Summary of Fetal Weights (g)

Day 21 Relative to Mating Date

Group

 

1

2

3

4

Number of live fetuses

 

311

322

316

281

Number of live male fetuses

 

163

149

162

130

Number of live female fetuses

 

148

173

154

151

% of live male fetuses

 

52.41

46.27

51.27

46.26

Litter weight (g)

Mean

S.D.

N

76.3098

10.9218

24

76.2963

8.36215

25

73.2175

14.7003

25

67.1419

11.4835

24

Fetal weight (g)

Mean

S.D.

N

5.91200

0.32036

24

5.93985

0.35902

25

5.75599

0.66825

25

5.75954

0.33311

24

Male fetal weight (g)

Mean

S.D.

N

6.05757

0.40204

24

6.07785

0.39095

25

5.89045

0.69793

25

5.91736

0.37567

24

Female fetal weight (g)

Mean

S.D.

N

5.75498

0.27884

24

5.79052

0.35776

25

5.59138

0.63434

25

5.63818

0.32685

24

Nominal Dose: Group 1 – 0 mg/kg Group 2 – 100 mg/kg         Group 3 – 300 mg/kg         Group 4 – 1000 mg/kg

 

Summary of Fetal Examinations

 

Day 21 Relative to Mating Date

Group

1

2

3

4

Total number of fetuses examined

Total number of litters examined

311

24

322

25

316

25

281

24

External defects

           Number of fetuses examined

 

           Number showing Malformation

           % of fetuses examined

           Number of litters affected

 

311

 

0

0

0

 

322

 

0

0

0

 

316

 

1

0.3

1

 

281

 

0

0

0

Visceral (rodents) defects

           Number of fetuses examined

 

           Number showing Variation

           % of fetuses examined

           Number of litters affected

 

152

 

0

0

0

 

160

 

0

0

0

 

157

 

0

0

0

 

144

 

1

0.7

1

Bouin’s head defects

           Number of fetuses examined

 

152

 

160

 

157

 

144

Skeletal defects

           Number of fetuses examined

 

           Number showing Malformation

           % of fetuses examined

           Number of litters affected

 

           Number showing Variation

           % of fetuses examined

           Number of litters affected

 

159

 

0

0

0

 

30

18.9

10

 

162

 

0

0

0

 

21

13.0

9

 

159

 

1

0.6

1

 

24

15.1

11

 

137

 

0

0

0

 

27

19.7

11

Total number of fetuses showing defects

% of fetuses examined with defects

30

9.6

21

69.5

25

7.9

28

10.0

Calculated values do not include animals that were not pregnant

Nominal Dose: Group 1 – 0 mg/kg Group 2 -100 mg/kg           Group 3 – 300 mg/kg         Group 4 – 1000 mg/kg

Conclusions:
Based on the results from the prenatal developmental toxicology study in Sprague Dawley rats, the no-adverse-observed-effect-level (NOAEL) for maternal and embryo-fetal developmental toxicity when Hatcol 1510 was administered via oral gavage over the entire period of gestation (GD 5-20) is 1000 mg/kg, the highest dose level tested.
Executive summary:

Decanoic acid, mixed esters with heptanoic acid, octanoic acid and trimethylolpropane (Hatcol 1510): Prenatal Developmental Toxicity Dose Range Finder Study in Pregnant Female Sprague Dawley Rats.

 

The purpose of the study was to determine the potential maternal and/or developmental toxicity of Hatcol 1510 in pregnant Sprague Dawley rats when administered via oral gavage during the embryo-fetal development period.

 

Four cohorts of 25 time-mated females arrived on their respective gestation day (GD) 1, 2, 3, or 4. These one-hundred time-mated female Sprague Dawley rats were randomly assigned to four groups (25 females/group; 6 or 7/cohort). Animals were administered control substance (peanut oil) or decanoic acid, mixed esters with heptanoic acid, octanoic acid and trimethylolpropane (Hatcol 1510) at 0, 100, 300, or 1000 mg/kg once daily via oral gavage from presumed GD 5 to 20. Animals were subjected to a full gross necropsy on GD 21.

 

Parameters evaluated during the study included mortality, physical examinations, cageside observations, body weights, body weight changes, food consumption, gross pathology findings, uterine data, fetal body weight and fetal examination data (external, visceral, and skeletal).

 

Dose concentration analysis showed that the test article was properly formulated (means of the first and last preparations were 97.2% - 116% of the target concentrations). No test article was detected in the control formulation.

 

Treatment with Hatcol 1510 at doses up to 1000 mg/kg had no effect on mortality, physical examinations, cageside observations, maternal body weights, maternal body weight changes, food consumption, uterine data, fetal body weights or fetal examination data (external, visceral, and skeletal). Only one fetal malformation (short tail) was observed in this study in Group 3, which was not considered test substance related. All remaining fetal observations noted were variations. The total number of fetuses showing defects (malformations and variations) was 30 (9.6%) for Group 1, 21 (6.5%) for Group 2, 25 (7.9%) for Group 3, and 28 (10.0%) for Group 4. The majority of variations were noted during the skeletal examinations. All variations were considered within normal range and were not considered test-substance related.

 

Based on the results from this prenatal developmental toxicology study in Sprague Dawley rats, the no-adverse-observed-effect-level (NOAEL) for maternal and embryo-fetal developmental toxicity when Hatcol 1510 was administered via oral gavage over the entire period of gestation (GD 5-20) is 1000 mg/kg, the highest dose level tested.

Effect on developmental toxicity: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
1 000 mg/kg bw/day
Study duration:
subacute
Species:
rat
Quality of whole database:
KS1
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
2 000 mg/kg bw/day
Study duration:
subacute
Species:
rat
Quality of whole database:
Klimisch 2 study.
Additional information

The purpose of the study was to determine the potential maternal and/or developmental toxicity of Hatcol 1510 in pregnant Sprague Dawley rats when administered via oral gavage during the embryo-fetal development period.

Four cohorts of 25 time-mated females arrived on their respective gestation day (GD) 1, 2, 3, or 4. These one-hundred time-mated female Sprague Dawley rats were randomly assigned to four groups (25 females/group; 6 or 7/cohort). Animals were administered control substance (peanut oil) or decanoic acid, mixed esters with heptanoic acid, octanoic acid and trimethylolpropane (Hatcol 1510) at 0, 100, 300, or 1000 mg/kg once daily via oral gavage from presumed GD 5 to 20. Animals were subjected to a full gross necropsy on GD 21.

Parameters evaluated during the study included mortality, physical examinations, cageside observations, body weights, body weight changes, food consumption, gross pathology findings, uterine data, fetal body weight and fetal examination data (external, visceral, and skeletal).

 

Treatment with Hatcol 1510 at doses up to 1000 mg/kg had no effect on mortality, physical examinations, cageside observations, maternal body weights, maternal body weight changes, food consumption, uterine data, fetal body weights or fetal examination data (external, visceral, and skeletal). Only one fetal malformation (short tail) was observed in this study in Group 3, which was not considered test substance related. All remaining fetal observations noted were variations. The total number of fetuses showing defects (malformations and variations) was 30 (9.6%) for Group 1, 21 (6.5%) for Group 2, 25 (7.9%) for Group 3, and 28 (10.0%) for Group 4. The majority of variations were noted during the skeletal examinations. All variations were considered within normal range and were not considered test-substance related.

Based on the results from this prenatal developmental toxicology study in Sprague Dawley rats, the no-adverse-observed-effect-level (NOAEL) for maternal and embryo-fetal developmental toxicity when Hatcol 1510 was administered via oral gavage over the entire period of gestation (GD 5-20) is 1000 mg/kg, the highest dose level tested.

Justification for classification or non-classification

Based on the above mentioned results, classification according to the CLP Regulation (EC)1272/2008 and the Dangerous Substance Directive 67/548/EC is not necessary.

A report justifying the read-across approach adopted as supporting information is included in IUCLID Chapter 13.

Additional information