Registration Dossier

Toxicological information

Toxicity to reproduction

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Administrative data

Endpoint:
screening for reproductive / developmental toxicity
Remarks:
based on test type (migrated information)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2009-2010
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study conducted under GLP and according to guideline. It is a dose-range finding study and therefore limited investigations were conducted but sufficient for a screening of reproductive and developmental effects.
Cross-reference
Reason / purpose:
reference to same study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2010
Report Date:
2010

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 421 (Reproduction / Developmental Toxicity Screening Test)
Deviations:
yes
Remarks:
8 animals/sex/group while the guidelines indicate almost 10 animals/sex/group
Principles of method if other than guideline:
Not applicable
GLP compliance:
yes
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): Hexylcinnamic aldehyde; octanal, 2-(phenylmethylene)-
- Physical state: Yellow liquid
- Analytical purity:98.6%
- Lot/batch No.:2814692
- Storage condition of test material:Room temperature, protected from light
- Other: Source: International Flavors and Fragrances, Inc., Union Beach, NJ

Test animals

Species:
rat
Strain:
Crj: CD(SD)
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories Inc.
- Age at study initiation: (P) Males: 85 days; Females: 79 days
- Weight at study initiation: (P) Males: 316-342 g; Females: 201-230 g
- Fasting period before study:no data
- Housing: P generation individually housed in steel wire-bottomed cages except during cohabitation and postpartum periods. From day 20 of presumed gestation female rats were housed individually in nesting boxes. Each dam and delivered litter were housed in a common nesting box during postpartum.
- Diet (e.g. ad libitum):ad libitum, Certified Rodent Diet #5002 (PMI Nutrition International)
- Water (e.g. ad libitum):ad libitum, mains sourced and filtered by reverse osmosis.
- Acclimation period:13 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18-26
- Humidity (%):30-70
- Air changes (per hr):at least 10
- Photoperiod (hrs dark / hrs light):12/12


IN-LIFE DATES: From: 17 Aug 2009 To: 2 Oct 2009

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
corn oil
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: Suspensions of test substance were prepared daily, stored at room temperature and protected from light.
VEHICLE
- Justification for use and choice of vehicle (if other than water): no data
- Concentration in vehicle: 3.125, 6.25, 12.5, 25mg/ml
- Amount of vehicle (if gavage): 4ml/kg
- Lot/batch no. (if required):J-145
- Purity:no data
Details on mating procedure:
- M/F ratio per cage: 1
- Length of cohabitation: 7 days
- Proof of pregnancy: vaginal plug and/or sperm in vaginal smear referred to as day 0 of pregnancy
- After 7 days of not mated females will be considered to be at day 0 of presumed gestation
- After successful mating each pregnant female was caged individually. Also those female rats in presumed gestation.
- Any other deviations from standard protocol: none
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
by HPLC-UV, validated internal method.
Duration of treatment / exposure:
Male rats: total 45 days - from 14days before cohabitation, through cohabitation (7 days) and continuing through the day before sacrifice after completion of the dosage period.
Female rats: total 44 or 45 days - from 14 days before cohabitation, through cohabitation (7 days), and continuing through the day before sacrifice on postpartum day 5, or study day 46 (for rats that did not become pregnant) or gestation day 25 (for rats that did not deliver). Dams with no surviving pups were sacrificed on postpartum day 1.
Pups were not administered the test substance directly but may have been exposed in utero or via maternal milk.
Frequency of treatment:
Once daily. Dosage volume was adjusted daily on the basis of individual body weights recorded before administration.
Details on study schedule:
- No selection of F1 parental animals took place
- Age at mating of the mated animals in the study: approximately 13 weeks for females and 14 weeks for males
Doses / concentrations
Remarks:
Doses / Concentrations:
0, 12.5, 25, 50, 100 mg/kg bw/day
Basis:
actual ingested
No. of animals per sex per dose:
8 animals/sex/dose
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: Results from two 90-day dermal studies.
- Rationale for animal assignment (if not random):random
- Other: none
Positive control:
none

Examinations

Parental animals: Observations and examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: at least twice per day

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule:at least weekly during acclimation period; First 5 days of dosage: prior to dosage and at approx. hourly intervals for the first 4 hours after dosage administration, and at the end of day; From day 6 of treatment: prior to dosage administration, 1-2hrs after dosage administration and at the end of the day; Day of sacrifice: once.; Postdosage periods: once daily.

BODY WEIGHT: Yes
- Time schedule for examinations: Acclimation period: at least weekly; Dosage period: daily; On sacrifice day: Once

FOOD CONSUMPTION: Yes; at least weekly for males; For females, at least weekly during acclimation and precohabitation periods, on gestation days 0, 7, 8, 9, 10, 14, 18, 21 and 25, and on postpartum days 1 and 5.

WATER CONSUMPTION No

OTHER: Rats were evaluated for adverse clinical signs observed during parturition, duration of gestation (from day 0 of gestation till the day the first pup was observed), litter sizes (all pups delivered) and pup viability at birth. Maternal behaviour was evaluated on pospartum days 1 and 5.
Oestrous cyclicity (parental animals):
By examination of vaginal cytology for 14 days before cohabitation, and then until spermatozoa were observed in a smear of the vaginal contents and/or a copulatory plug was observed in situ during the cohabitation period.
Sperm parameters (parental animals):
Epididymides, prostate, seminal vesicles and testes were weighed and microscopically evaluated.
Litter observations:
STANDARDISATION OF LITTERS
- Performed on day 4 postpartum: no

PARAMETERS EXAMINED
The following parameters were examined in F1 offspring:
number and sex of pups, stillbirths, live births, postnatal mortality, weight gain, clinical observations

GROSS EXAMINATION OF DEAD PUPS:
yes, for external and internal abnormalities; possible cause of death was determined for pups born or found dead. Those that died before examination of the litter for pup viability were evaluated by immersing their lungs in water. Pups with lungs that sunk were identified as stillborn. Pups with lungs that floated were identified as liveborn and to have died shortly after birth. Pups that died before scheduled termination on days 2-5 postpartum were preserved in Bouin's solution for possible future evaluation except when precluded by autolysis.
Postmortem examinations (parental animals):
SACRIFICE
- Male animals: All surviving animals after completion of cohabitation period on day 46 of treatment.
- Maternal animals: All surviving animals on postpartum day 5, day study 46 or day gestation 25.

GROSS NECROPSY
- Gross necropsy consisted of external and internal examinations including the cervical, thoracic, and abdominal viscera. Uteri of apparently nonpregnant rats were examined, while being pressed between glass plates, to confirm the absence of implantation sites, and were retained in 10% NBF.

HISTOPATHOLOGY / ORGAN WEIGHTS
The following tissues were prepared for microscopic examination and weighed: Epididymides, gross lesions/masses (only histology), ovaries, prostate, seminal vesicles, testes, uterus with cervix.
Postmortem examinations (offspring):
SACRIFICE
- The F1 offspring at 4 days of age (postpartum day 5).

GROSS NECROPSY
- Gross necropsy consisted of external and internal examinations including a single cross-section of the head at the level of the frontal-parietal suture and examination of the cross-sectioned brain for apparent hydrocephaly.

Statistics:
Averages and percentages calculated
Reproductive indices:
% rats that mated; Fertility index (number of pregnancies/number of rats that mated); % rats mated with female on days 1-7; % Pregnant rats among those cohabited
Offspring viability indices:
% delivered litters; number implantation sites per delivered litter; % dams with stillborn pups; Gestation index (number of dams with one or more liveborn pups/number of pregnant rats); % dams with all pups dying days 1-5 postpartum; Viability index (number of live pups on day 5 postpartum/ number of liveborn pups on day 1 postpartum; Surviving pups/litter on day 1 and on day 5; % male pups per number of pups sexed on day 1 and 5; Live litter size at weighing on day 1 and 5; Pup weight/litter on day 1 and 5;

Results and discussion

Results: P0 (first parental animals)

General toxicity (P0)

Clinical signs:
no effects observed
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
only during lactation in females
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
only during lactation in females
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Other effects:
not examined

Reproductive function / performance (P0)

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

Details on results (P0)

CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS)
There were no mortalities. All clinical observations were considered unrelated to treatment.

BODY WEIGHT AND FOOD CONSUMPTION (PARENTAL ANIMALS)
Males: body weights and body weight gains, and absolute (g/day) and relative (g/kg/day) feed consumption were generally comparable among the dosage groups.
Females:
Precohabitation - body weights increased during second week compared to control group and overall for the entire premating period (14% greater than controls). These increases were not considered adverse since they didn't persist and there was not corresponding effect on feed consumption. Gestation - Body weights and feed consumption were unaffected by treatment.
Lactation - maternal body weight gains in the 100mg/kg/day were reduced on days 1-5 of lactation, in comparison with control group. Corresponding to reduced body weight gains during the lactation period, feed consumption values in the 100mg/kg/day were reduced in comparison to the vehicle group.

REPRODUCTIVE FUNCTION: ESTROUS CYCLE (PARENTAL ANIMALS)
The number of estrous stages per 14 days was comparable among the five dosage groups during the precohabitation period.

REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS)
All mating and fertility parameters were unaffected at all dosages (fertility index, rats with confirmed mating dates during cohabitation and number of pregnancies per number of rats in cohabitation).
There was an apparent increase in the average number of days to mating and a reduction in % of rats that mated that occurred in the highest dose group. This was attributed to two male rats that did not mate with their cohort rats. These effects were not attributed to treatment for the following reasons: 1) there was no apparent effect on male or female reproductive organ weights; 2) the remaining rats in the 100mg/kg/day dosage group (N=6) mated and had viable litters; 3) there were not apparent effects on natural delivery in this dosage group; and 4) the average value (3.4+/- 2.3 days) was within the historical range of the Testing Facility.

ORGAN WEIGHTS (PARENTAL ANIMALS)
The weights of the epididymides, testes, seminal vesicles (with and without fluid) and prostate and the ratios of these organ weights to terminal body weight were unaffected.

GROSS PATHOLOGY (PARENTAL ANIMALS)
There were no treatment-related gross lesions at any dosage level tested.

HISTOPATHOLOGY (PARENTAL ANIMALS)
None of the microscopic findings that occurred were considered related to the test substance. They were all considered to be incidental or spontaneous changes.

Effect levels (P0)

Dose descriptor:
NOAEL
Effect level:
>= 100 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: No treatment related effects observed

Results: F1 generation

General toxicity (F1)

Clinical signs:
no effects observed
Mortality / viability:
no mortality observed
Body weight and weight changes:
no effects observed
Sexual maturation:
not examined
Organ weight findings including organ / body weight ratios:
not examined
Gross pathological findings:
no effects observed
Histopathological findings:
not examined

Details on results (F1)

Natural delivery and litter observations were unaffected by treatment. Values for the numbers of dams delivering litters, the duration of gestation, averages for implantation sites per delivered litter, the gestation index, the numbers of dams with stillborn pups and of dams with all pups dying, litter sizes, viability index, surviving pups per litter, percent male pups per number of pups sexed per litter, live litter size at weighing and pup weight per litter were comparable among the five dosage groups.
No treatment related clinical observations occurred or tretment related necropsy observations. All pups that died early or survived to scheduled sacrificed appeared normal at necropsy.

Effect levels (F1)

Dose descriptor:
NOAEL
Generation:
F1
Effect level:
>= 100 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: No treatment related effects observed.

Overall reproductive toxicity

Reproductive effects observed:
not specified

Any other information on results incl. tables

Table 7.8.1/1: Mating and fertility - summary - P generation male rats

 

Dosage group

 

I

II

III

IV

V

Dosage (mg/kg/day)

 

0 (Vehicle)

12.5

25

50

100

Rats in cohabitation

N

8

8

8

8

8

Days in cohabitationa

Mean ± S.D.

2.0 ± 1.1

3.0 ± 1.3

2.6 ± 1.3 [7]

2.9 ± 1.4

3.4 ± 2.3

Rats that mated

N (%)

8(100.0)

8(100.0)

8(100.0)

8(100.0)

6( 75.0)

Fertility indexb

N/N (%)

7/ 8 (87.5)

8/ 8 (100.0)

7/ 8 (87.5)

8/ 8 (100.0)

6/ 6 (100.0)

Rats with confirmed mating dates

N

8

8

7

8

6

Mated with femalec
days 1-7

N(%)

8(100.0)

8(100.0)

7(100.0)

8(100.0)

6(100.0)

Rats pregnant/rats in cohabitation

N/N

7/ 8

8/ 8

7/ 8

8/ 8

6/ 8

(%)

(87.5)

(100.0)

(87.5)

(100.0)

(75.0)

[ ] = Number of values averaged

a. Restricted to rats with a confirmed mating date and rats that did not mate.

b. Number of pregnancies/number of rats that mated.

c. Restricted to rats with a confirmed mating date.

 

Table 7.8.1/2: Estrous cycling, mating and fertility - summary - P generation female rats

 

Dosage group

 

I

II

III

IV

V

Dosage (mg/kg/day)a

 

0 (Vehicle)

12.5

25

50

100

Estrous cycling observations

Rats evaluated

N

8

8

8

8

8

Precohabitation estrous cycling

 

 

 

 

 

 

Estrous stages/14 days

Mean ± S.D.

3.2 ± 0.7

3.5 ± 0.9

3.2 ± 0.5

3.1 ± 0.8

3.5 ± 0.5

Rats with 6 or more consecutive days of diestrus

N(%)

0(0.0)

0(0.0)

0(0.0)

0(0.0)

0(0.0)

Rats with 6 or more consecutive days of estrus

N(%)

0(0.0)

0(0.0)

0(0.0)

0(0.0)

0(0.0)

Mating observations

Rats in cohabitation

N

8

8

8

8

8

Days in cohabitationb

Mean ± S.D.

2.0 ± 1.1

3.0 ± 1.3

2.6 ± 1.3 [ 7]

2.9 ± 1.4

3.4 ± 2.3

Rats that mated

N(%)

8(100.0)

8(100.0)

8(100.0)

8(100.0)

6(75.0)

Fertility indexc

N/N(%)

7/ 8( 87.5)

8/ 8(100.0)

7/ 7(100.0)

8/ 8(100.0)

6/ 6(100.0)

Rats with confirmed mating dates

N(%)

8(100.0)

8(100.0)

7(100.0)

8(100.0)

6(100.0)

Rats pregnant/rats in cohabitation

N/N

7/ 8

8/ 8

7/ 8

8/ 8

6/ 8

(%)

(87.5)

(100.0)

(87.5)

(100.0)

(75.0)

[ ] = Number of values averaged

a. Dosage occurred on Day 1 of study through Day 4 of lactation or Day 24 of presumed gestation (rats that did not deliver a litter).

b. Restricted to rats with a confirmed mating date and rats that did not mate.

c. Number of pregnancies/number of rats that mated.

 

Table 7.8.1/3: Natural delivery observations - summary - P generation female rats

 

Dosage group

 

I

II

III

IV

V

Dosage (mg/kg/day)a

 

0 (Vehicle)

12.5

25

50

100

Rats assigned tonatural delivery

N

8

8

8

8

8

Pregnant

N

7

8

7

8

6

Delivered litters

N(%)

7(100.0)

8(100.0)

7(100.0)

8(100.0)

6(100.0)

Duration of gestationb

Mean ± S.D.

22.7 ± 0.5

22.8 ± 0.5

22.7 ± 0.5[ 6]c

23.0 ± 0.9

22.3 ± 0.5

Implantation sitesper delivered litter

N

112

136

116

115

98

 

Mean ± S.D.

16.0 ± 1.6

17.0 ± 1.6

16.6 ± 1.6

14.4 ± 5.7

16.3 ± 1.9

Dams with stillborn pups

N(%)

1(14.3)

1(12.5)

1(14.3)

1(12.5)

1(16.7)

Dams with no liveborn pups

N

0

0

0

1

0

Gestation index

 

%

100.0

100.0

100.0

87.5

100.0

N/N

7/ 7

8/8

7/ 7

7/8

6/6

Dams with all pups dying
Days 1-5 postpartum

N(%)

0

0

0

0

0

a. Dosage occurred on Day 1 of study through Day 4 of lactation.

b. Calculated (in days) as the time elapsed between confirmed mating (arbitrarily defined as Day 0 of gestation) and the day the first pup was delivered.

c. Excludes a value for dam 3460, which did not have a confirmed mating date.

 

 

Table 7.8.1/4: Litter observations (naturally delivered pups) - summary - F1 generation litters

 

Maternal dosage group 

 

 I 

 II 

 III 

 IV 

 V 

Maternal dosage (mg/kg/day)

 

0 (vehicle)

 12.5 

 25 

 50 

 100 

Delivered litters with one or more liveborn pups

 N 

 7 

 8 

 7 

 7 

 6 

Pups delivered (total)

 N 

 103 

 132 

 104 

 108 

 89 

 

 Mean ± S.D. 

 14.7 ± 1.8

 16.5 ± 1.4 

 14.8 ± 3.4 

 15.4 ± 1.1 

 14.8 ± 2.8 

  Liveborn 

 Mean ± S.D. 

 14.6 ± 1.8 

 16.4 ± 1.3 

 14.7 ± 3.4 

 15.3 ± 1.1 

 14.7 ± 2.7 

 

 N(%) 

 102(99.0) 

 131(99.2) 

 103(99.0) 

 107(99.1) 

 88(98.9) 

 Stillborn 

 Mean ± S.D. 

 0.1 ± 0.4 

 0.1 ± 0.4 

 0.1 ± 0.4 

 0.1 ± 0.4 

 0.2 ± 0.4 

 

N(%) 

1(1.0)

1(0.8)

1(1.0)

1(0.9)

1(1.1)

Pups found dead or presumed cannibalized

 

 

 

 

 

 

Day 1

N/N(%)

0/102(0.0)

0/131(0.0)

0/103(0.0)

0/107(0.0)

0/ 88(0.0)

Days 2- 5

N/N(%)

2/102(2.0)

0/131(0.0)

0/103(0.0)

0/107(0.0)

6/ 88(6.8)

Viability indexa

 % 

 98.0 

 100.0 

 100.0 

 100.0 

 96.6 

 

N/N

100/102

131/131

103/103

107/107

85/ 88

Delivered litters with one or more liveborn pups

 N 

 7 

 8 

 7 

 7 

 6 

Surviving pups/litter

 

 

 

 

 

 

Day  1 

 Mean ± S.D. 

 14.6 ± 1.8 

 16.4 ± 1.3 

 14.7 ± 3.4 

 15.3 ± 1.1 

 14.7 ± 2.7 

Day  5 

 Mean ± S.D. 

 14.3 ± 1.6 

 16.4 ± 1.3 

 14.7 ± 3.4 

 15.3 ± 1.1 

 13.7 ± 2.4 

Percent male pups per number of pups sexed

 

 

 

 

 

 

Day 1 

 Mean ± S.D. 

 56.1 ± 17.0 

 52.4 ± 4.3 

 56.8 ± 18.3 

 50.5 ± 17.4 

 48.2 ± 12.5 

Day 5 

 Mean ± S.D. 

 57.0 ± 16.3 

 52.4 ± 4.3 

 56.8 ± 18.3 

 50.5 ± 17.4 

 48.5 ± 11.8 

Delivered litters with one or more liveborn pups

 N 

 7 

 8 

 7 

 7 

 6 

Live litter size at weighing

 

 

 

 

 

 

Day 1 

 Mean ± S.D. 

 14.6 ± 1.8 

 16.4 ± 1.3 

 14.7 ± 3.4 

 15.3 ± 1.1 

 14.7 ± 2.7 

Day 5 

 Mean ± S.D. 

 14.3 ± 1.6 

 16.4 ± 1.3 

 14.7 ± 3.4 

 15.3 ± 1.1 

 13.7 ± 2.4 

Pup weight/litter (grams)

 

 

 

 

 

 

Day 1 

 Mean ± S.D. 

 6.3 ± 0.1 

 6.2 ± 0.4 

 6.4 ± 0.5 

 6.5 ± 0.7 

 6.2 ± 0.5 

Day 5 

 Mean ± S.D. 

 9.8 ± 0.7 

 9.3 ± 1.0 

 9.9 ± 1.8 

 9.8 ± 1.2 

 9.2 ± 1.6 

Day(s) = Day(s) postpartum

a. Number of live pups on Day 5 postpartum/number of liveborn pups on Day 1 postpartum.

 

Table 7.8.5/5: Maternal body weights - lactation - individual data - P generation female rats

 

  Day 

 1 

 2 

 3 

 4 

 5 

Rat #

Dosage group: 0 (vehicle) mg/kg bw/day

  3441 

 286 

 287 

 291 

 298 

 309 

 3442 

 322 

 334 

 344 

 339 

 332 

 3443 

 287 

 282 

 289 

 293 

 309 

 3444 

 321 

 328 

 322 

 327 

 337 

 3445 

 285 

 288 

 283 

 285 

 284 

 3446 

 350 

 331 

 336 

 342 

 354 

 3447 

 312 

 318 

 320 

 322 

 322 

3448

Not pregnant

Rat #

Dosage group : 100 mg/kg bw/day

3473

Not pregnant; mating not confirmed

3474 

 290 

 287 

 292 

 293 

 310 

3475 

 299 

 294 

 298 

 297 

 302 

3476

Not pregnant; mating not confirmed

3477 

 279 

 273 

 288 

 290 

 284 

3478 

 351 

 347 

 345 

 344 

 346 

3479 

 352 

 336 

 324 

 323 

 334 

3480 

 327 

 323 

 326 

 328 

 338 

All weights were recorded in grams (g).

Day = Day of lactation

Applicant's summary and conclusion

Conclusions:
The NOAEL for fertility effects was 100 mg/kg bw/day or higher under the test conditions of this study.
Executive summary:

In the dosage-range finding study conducted similarly according to guideline OECD421,rats were given once daily by gavage alpha-hexylcinnamaldehyde (HCA) diluted in corn oil at 12.5, 25, 50 or 100 mg/kg bw or the vehicle alone. The males were treated from 14 days before cohabitation, through mating (maximum of 7 days), and continuing through the day before sacrifice on day 47. The female rats were administered HCA (same dosage as for males) or vehicle two weeks before cohabitation, through mating, and continuing through the day before sacrifice on day. Female rats were allowed to deliver their litters and were sacrificed on postpartum day 5 (PPD 5). F1 generation pups were also sacrificed on PPD 5 (i.e.5thday of lactation). The following parameters were evaluated: viability, clinical observations, body weights, feed weights, mating and fertility, delivery and litter observations, organ weights (epididymides, ovaries, prostate, seminal vesicles, testes, and uterus with cervix), necropsy observations and histopathology (epididymides, ovaries, prostate, seminal vesicles, testes, and uterus with cervix).

All P generation male and female rats survived to scheduled sacrifice. There were no treatment-related clinical observations or gross lesions in the P generation rats of both sexes at any dosage level tested. In addition, none of the microscopic findings examined were considered related to treatment withHCA. Body weights and body weight gains of the treated P generation male rats were generally comparable among the dosage groups. Absolute and relative feed consumption values in male rats were unaffected by treatment withHCAduring the entire dosage period.

 

In treated P generation female rats, body weight gains were increased (14% by comparison with the vehicle group) during the second week of the pre-cohabitation dosage period in the 100 mg/kg bw/day dosage group, and therefore overall for the entire two-week premating period . These increases in body weight gain were not considered adverse because there was no corresponding effect on feed consumption, and the increase in weight gain did not persist during the gestation period. Body weights and body weight gains of the female rats were unaffected by treatment withHCAduring the gestation period. Absolute and relative feed consumption values were unaffected by treatment withHCAduring the precohabitation and gestation periods.

 

During the lactation period, the mean maternal body weight gain in the 100 mg/kg bw/day dosage group was 78% lower on days 1 to 5 of lactation, in comparison to the vehicle control group value: the mean maternal body weight gains in the 100 mg/kg bw/day dosage group and vehicle control group were 2.7 ± 13.1 and 12.0 ± 8.9 grams (mean ± S.D.), respectively.  In fact, among the six females which were pregnant (6 out of 8 mated females) and allowed to deliver their litter, two females lost body weight (from 2.5 to 5%) between the first day and the fifth day of lactation and two other females lost body weight or didn’t gain any body weight up to the fourth day of lactation.

Therefore, four females out of six have lost body weight up to the fourth day of lactation. Corresponding to reduced body weight gains during the lactation period, feed consumption values (weight in gram during the five days of lactation) in the 100 mg/kg bw/day dosage group were reduced on lactation days 1 to 5, in comparison to the vehicle control group values (84% of controls). The significant decrease in maternal body weight in the 100 mg/kg bw/day dose group during the lactation period showed that this dose level can be considered as the maximum tolerated dose (MTD).

 

Terminal body weights and male and female reproductive organ weights were comparable among the five dosage groups. There was no effect ofHCAon estrous cycling. There was no effect on mating and fertility (fertility index, gestation index, number of implantation sites) at any dosage level tested. All recorded pregnant rats (7, 8, 7, 8 and 6 females in the five respective dosage groups from the control group to the highest dose group) delivered a litter. Natural delivery and litter observations (duration of gestation, number and sex of offspring per litter, stillbirths, live births, gross alterations, litter size and viability, viability index, lactation index, percent survival, sex ratio, pup body weights) were unaffected by dosages ofHCAas high as 100 mg/kg bw/day. No treatment-related clinical or necropsy (including a single cross-section of the head and examination of the cross-sectioned brain for apparent hydrocephaly) observations occurred in the F1 generation pups. As such, no developmental effects were observed.

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The NOAEL for maternal and developmental toxicity was 100 mg/kg bw/day or greater,