Quaternary ammonium compounds, bis(hydrogenated tallow alkyl)dimethyl, chlorides, reaction products with polyethylenepolyamines and tall-oil fatty acids, humates hydrochlorides

Toxicity to reproduction

Administrative data

Endpoint:

screening for reproductive / developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2014

Report date:

2014

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Test material form:

solid: particulate/powder

Details on test material:

- Substance type: UVCB
- Physical state: powder

Test animals

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Limited, Margate, Kent, UK
- Age at study initiation: ca 6-7 weeks
- Weight at study initiation: 201-227 g males; 163-195 g females
- Housing: Housed in polycarbonate cages with stainless steel grid tops and solid bottoms. Animals were initially housed 2/cage by sex. Males were individually housed a few days prior to mating. After mating, males were re-housed with original cage mates.
- Diet (e.g., ad libitum): Rat and Mouse Breeder Diet No. 3 (Expanded) provided ad libitum
- Water (e.g., ad libitum): Public water supply provided ad libitum
- Acclimation period: 9 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19-23°C
- Humidity (%): 40-70%
- Air changes (per hr): 10 air changes per hr
- Photoperiod (hrs dark / hrs light): 12 hrs dark / 12 hrs light

Administration / exposure

Route of administration:

inhalation: dust

Type of inhalation exposure (if applicable):

nose only

Vehicle:

air

Details on mating procedure:

- M/F ratio per cage: 1:1 basis
- Length of cohabitation: Female remained with male until mating had occurred
- Proof of pregnancy: sperm in vaginal smear referred to as Day 0 of pregnancy

Analytical verification of doses or concentrations:

yes

Duration of treatment / exposure:

Males were treated daily 2 weeks prior to mating, throughout mating, and until the day prior to necropsy (ca. 4 weeks of treatment). Females were treated daily 2 weeks prior to mating, throughout mating, and up to Day 19 of gestation (ca. 6 weeks for treatment).

Frequency of treatment:

6 hr per day, 7 days per week

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

5 mg/m³ air (nominal)

Doses / concentrations 2

Dose / conc.:

50 mg/m³ air (nominal)

Doses / concentrations 3

Dose / conc.:

300 mg/m³ air (nominal)

No. of animals per sex per dose:

10 animals/sex/dose

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: Doses were selected based on the results for a previously performed study.
- Rationale for animal assignment (if not random): Cages were allocated treatment group by the use of randomly sequenced numbers in such a way that each complete rack contained representatives from all treatment groups.

Positive control:

The study did not include a positive control.

Examinations

Parental animals: Observations and examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: One week prior to treatment and weekly thereafter
- Cage side observations included: posture/condition (i.e., prostration, lethargy, writhing, circling, breathing abnormalities, gait abnormalities, tremor, fasciculation, convulsions, biting, vocalizations, and piloerection); ease of removal from the cage; body temperature; condition of the eyes (i.e., pupillary function, miosis, mydriasis, exophthalmos, encrustation, and lacrimation) and coat; presence of salivation, and overall ease of handling.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: One week prior to treatment and daily thereafter

BODY WEIGHT: Yes
- Time schedule for examinations: One week prior to treatment for both sexes. Thereafter, males were weighed twice weekly at the start of dosing until the end of the study. Females were weighed twice weekly at the start of dosing and daily from mating to termination.

FOOD CONSUMPTION:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes

FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: No

WATER CONSUMPTION: No
- Time schedule for examinations: Not reported

Oestrous cyclicity (parental animals):

Data not reported

Sperm parameters (parental animals):

Data not reported

Litter observations:

STANDARDISATION OF LITTERS
- Performed on day 4 postpartum: No

PARAMETERS EXAMINED
The following parameters were examined in F1 offspring: number pups, stillbirths, live births, postnatal mortality, presence of gross anomalies, weight gain, physical or behavioral abnormalities

GROSS EXAMINATION OF DEAD PUPS:
Yes. Pups dying prior to scheduled necropsy were examined externally and checked for the presence of milk in the study. A macroscopic examination of the tissues and organs of the cranial, thoracic, and abdominal cavities in situ. Pups and their dissected tissues were preserved in 10% neutral buffered formalin.

Postmortem examinations (parental animals):

SACRIFICE
- Male animals: All surviving males were sacrificed after mating and treatment for at least 4 weeks.
- Maternal animals: All surviving females were sacrificed between Day 5 and 7 of lactation.

GROSS PATHOLOGY: Yes (see Table 1)

HISTOPATHOLOGY / ORGAN WEIGHTS: Yes; tissues identified in table 1 (except bone marrow smears) were processed from 5 males and 5 females from the control and high-dose groups.

Postmortem examinations (offspring):

SACRIFICE
- The F1 surviving offspring were scarified between Day 5 and 7 of lactation.

GROSS NECROPSY
- Surviving pups were examined externally for visible abnormalities. Those found to be externally normal were discarded. Pups dying prior to scheduled necropsy were examined externally and checked for the presence of milk in the study. A macroscopic examination of the tissues and organs of the cranial, thoracic, and abdominal cavities in situ. Pups and their dissected tissues were preserved in 10% neutral buffered formalin.

Statistics:

The following endpoints were analyzed using the 'F Max' test: selected body weight and food consumption data; all hematology, coagulation, and clinical chemistry; and selected functional observational battery and motor activity data. Homogeneous variances were analyzed using a parametric ANOVA. Pairwise comparisons were made using Fisher's F protected LSD method via Student's t test only if the overall F test was significant. Heterogeneous variances were initially transformed using the log or square root. If variances remained heterogeneous, then a Kruskal-Wallis non-parametric ANOVA was used and pairwise compassions were made using chi squared protection.

Organ weight data were analyzed using the methods noted above and through analysis of covariance (ANCOVA) using terminal body weight as the covariate. Organ weight were also analyzed as a percentage of terminal body weight.

Reproductive indices:

Fertility index (male) = (number siring a litter)/number paired

Fertility index (female) = (number pregnant)/number paired

Gestation index = (number bearing live pups)/number pregnant

Offspring viability indices:

Birth index = (total number of pups born [live and dead])/number of implantation scars

Live birth index = (number of pups born live on Day 0 of lactation)/(total number born [live and dead])

Viability index = (number of pups live on Day 4 of lactation)/(number live on Day 0)

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:

effects observed, treatment-related

Dermal irritation (if dermal study):

not examined

Mortality:

no mortality observed

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

no effects observed

Food efficiency:

not examined

Water consumption and compound intake (if drinking water study):

not examined

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Other effects:

no effects observed

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:

not specified

Reproductive function: sperm measures:

not specified

Reproductive performance:

no effects observed

Details on results (P0)

CLINICAL SIGNS AND MORTALITY: Increased incidence of staining around the head, nose, eyes, muzzle, and ears for all treated animals. Mid- and high-dose animals also had staining on their coats.

ORGAN WEIGHTS: At the high-dose, a statistically significant increase in both absolute and relative lung weight was observed in high-dose males and females. At the mid-dose, there was a statistically significant increase in absolute lung weight for both males and females; however, only females exhibited a statistically significant increase in relative lung weight. No effects on organ weights were observed at the low dose for both males and females.

GROSS PATHOLOGY: There was an increased incidence of dark discoloration of the lungs for all high-dose males and females and dark discoloration of the bronchial lymph node for high-dose males (10/10) and females (9/10). High-dose females also showed an increased incidence of pale foci on the lungs (5/10) and dark discoloration of the mediastinal lymph node (5/10). Mid-dose males (8/10) and females (8/10) showed an increased incidence of dark discoloration of the lungs.

HISTOPATHOLOGY: NON-NEOPLASTIC: At the high-dose, the following treatment-related findings were noted: bronchiolo-alveolar pigmented macrophages/hyperplasia; multifocal alveolar foamy macrophage accumulation; and a higher severity and incidence of mononuclear cell infiltration. In the bronchial lymph node there were pigmented macrophages and increased cellularity. In the mediastinal lymph node, there were pigmented macrophages. A microscopic examination was not conducted at 0.05 and 0.005 mg/L.

Effect levels (P0)

Dose descriptor:

NOAEC

Remarks:

reproductive

Effect level:

300 mg/m³ air

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: lack of effect

Target system / organ toxicity (P0)

Critical effects observed:

no

Results: F1 generation

General toxicity (F1)

Clinical signs:

no effects observed

Mortality / viability:

mortality observed, treatment-related

Body weight and weight changes:

effects observed, treatment-related

Sexual maturation:

not examined

Organ weight findings including organ / body weight ratios:

not specified

Gross pathological findings:

not specified

Histopathological findings:

not specified

Details on results (F1)

VIABILITY (OFFSPRING): A slight, nontreatment-related difference in the number of live pups per litter on Days 0, 1, and 4 of lactation were observed.

BODY WEIGHT (OFFSPRING): At the low- and high-dose groups, litter weight on Day 4 of lactation were slightly lower than controls. This was mainly due to one dam in each group that had only one pup remaining. When these animals were removed for the group calculations, group male litter weights were comparable to the control values.

Effect levels (F1)

Dose descriptor:

NOEC

Generation:

F1

Effect level:

> 300 mg/m³ air

Based on:

test mat.

Sex:

male

Basis for effect level:

other: no adverse effect

Target system / organ toxicity (F1)

Critical effects observed:

no

Overall reproductive toxicity

Reproductive effects observed:

no

Applicant's summary and conclusion

Conclusions:

In a combined repeated dose toxicity study with the reproduction/developmental toxicity screening test (OECD 422) inhalation toxicity study, organolignite was administered to 10 Sprague-Dawley rats/sex/concentration by nose only exposure at concentrations of 0, 5, 50, or 300 mg/m3 for 6 hours per day, 7 days/week. There were no treatment-related effects on mating, pregnancy performance, fertility, maternal care, or pup performance. The reproductive NOAEC is 300 mg/m3 based on the lack of effect.

Executive summary:

In a combined repeated dose toxicity study with the reproduction/developmental toxicity screening test (OECD 422) inhalation toxicity study, organolignite was administered to 10 Sprague-Dawley rats/sex/concentration by nose only exposure at concentrations of 0, 5, 50, or 300 mg/m³ for 6 hours per day, 7 days/week. Males were treated daily 2 weeks prior to mating, throughout mating, and until the day prior to necropsy (ca. 4 weeks of treatment). Females were treated daily 2 weeks prior to mating, throughout mating, and up to Day 19 of gestation (ca. 6 weeks for treatment).

 

There were no treatment-related effects on mating, pregnancy performance, fertility, maternal care, or pup performance. The systemic NOAEC is 300 mg/m³ (equivalent to 0.3 mg/L; i.e., the high dose) based on a lack of adverse effect. The reproductive NOAEC is 300 mg/m³ (i.e., 0.3 mg/L) based on the lack of effect.