Registration Dossier

Administrative data

Endpoint:
sub-chronic toxicity: oral
Type of information:
experimental study
Adequacy of study:
supporting study
Study period:
26th January 2010 to 6th April 2010
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: fully GLP compliant and audited
Cross-reference
Reason / purpose for cross-reference:
reference to same study

Data source

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

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
other: OECD 421
GLP compliance:
yes (incl. QA statement)
Limit test:
no

Test material

Constituent 1
Reference substance name:
amporphous glass fibre formed from the melting and fiberisation of predominately slilcon dioxide, calcium oxide, magnesium oxide
EC Number:
610-130-5
Cas Number:
436083-99-7
Molecular formula:
Amorphous glass consisting of SinO(3n-1)2(n-1) Polymeric anions ionically bonded to Ca2+ and Mg2+ cations or other alkaline earth cations
IUPAC Name:
amporphous glass fibre formed from the melting and fiberisation of predominately slilcon dioxide, calcium oxide, magnesium oxide
Details on test material:
- Name of test material (as cited in study report): AES Covance
- Substance type: Inorganic
- Physical state: Solid
- Analytical purity: >99%
- Impurities (identity and concentrations): trace impurities
- Purity test date: 14/12/09
- Lot/batch No.: BG-10-X99-2981
- Expiration date of the lot/batch: 31/12/2020
- Stability under test conditions: stable
- Storage condition of test material: room temperature, sealed container

Test animals

Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals or test system and environmental conditions:
- Source: Charles River Laboratories, Margate, UK
- Age at study initiation: 10 to 12 weeks old
- Weight at study initiation: Males: 271.2 - 310.4g; Females: 172.1 - 201.7g;
- Fasting period before study: N/A
- Housing: Cages conforming to 'Code of practice for the housing and care of animals used in scientific procedures' (Home Office, London, 1989)
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: 2 weeks

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19 - 25
- Humidity (%): 40 - 70
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 26th January 2010 To: 6th April 2010

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
other: MC (methyl cellulose)
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS: daily

VEHICLE
- Justification for use and choice of vehicle (if other than water): to ensure homogenous suspension of the substance for dosing as it is insoluble in water
- Amount of vehicle (if gavage): 10ml/kg
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Verification of composition at start and end of sampling carried out by XRF analysis, concentrations verified by gravimetric analysis of start and end samples.
Duration of treatment / exposure:
40 days (female) - 55 days (males)
Frequency of treatment:
daily dosing
Doses / concentrations
Remarks:
Doses / Concentrations:
250 mg/kg/day
Basis:
other: nominal concentration in vehicle
No. of animals per sex per dose:
10
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: The dose level for the test article was chosen after consultation with the Sponsor and based on the results of previous inhalation studies and the physical properties of the test articles. Due to the potential risk of physical interference with gut motility, a high dose of between 50 and 100 mg/rat/day (250 mg/kg/day) was considered to be the limiting dose for this study.
Positive control:
none used

Examinations

Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily start and end (nominal) of the working day In addition, the animals were observed immediately on return to the home cage after dosing and at 0.5, 1, 2 and 4 hours post dose for signs of reaction to treatment for the first two weeks of dosing. As no signs were observed the animals were observed immediately on return to the home cage after dosing and at 0.5 and 1 hour post dose only from the start of the pairing period until the end of dosing.
- Cage side observations checked; ill health or overt toxicity. Any abnormalities of appearance or behaviour or other signs of reaction to treatment or ill health were recorded

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: weekly, at time of body weight examination Clinical signs for females during the gestation and lactation periods, were recorded more frequently that weekly due to days of body weight recording.

BODY WEIGHT: Yes
- Time schedule for examinations:
MALES : Day -7 (randomisation body weight check)
Weekly
Day of (prior to) necropsy
FEMALES: Day -7 (randomisation body weight check)
Weekly prior to pairing and until confirmation of mating
Days 0, 7, 14 and 20 of gestation
Days 1 and 4 post partum

FOOD CONSUMPTION:
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
The food consumed by each cage of animals was determined weekly during the pre pairing periods.
Individual food intake of mated females was recorded on Days 0 to 6, 7 to 13 and 14 to 19 of gestation, and on Days 1 to 3 of lactation.
Sacrifice and pathology:
SACRIFICE
- Male animals: All surviving animals after assessment of females and offspring
- Maternal animals: All surviving animals 5 days post-partum

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

HISTOPATHOLOGY / ORGAN WEIGHTS
The tissues listed below were prepared for microscopic examination and weighed, respectively.
ovaries (with oviducts) testes
uterus epididymides
cervix seminal vesicles
vagina prostate
pituitary coagulating gland
gross lesions animal identification
Other examinations:
Postmortem examinations (Offspring)
SACRIFICE
- The F1 offspring were sacrificed at 5 days of age.
- These animals were subjected to postmortem examinations (macroscopic examination) as follows:
GROSS NECROPSY
- Gross necropsy consisted of external and internal examinations including the cervical, thoracic, and abdominal viscera.
Statistics:
Statistical analysis used SAS

Results and discussion

Results of examinations

Clinical signs:
no effects observed
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
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
no effects observed
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
no effects observed
Details on results:
CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS)
There were no deaths during the study, all animals survived until scheduled necropsy. There were no clinical observations recorded that were considered to be related to treatment with AES Covance

BODY WEIGHT AND FOOD CONSUMPTION (PARENTAL ANIMALS)
There was no adverse effect of treatment on mean body weight or body weight gain in males or females. In males and females, there was no effect of treatment on mean food intake.

REPRODUCTIVE FUNCTION: ESTROUS CYCLE (PARENTAL ANIMALS)
the majority of females mated within one oestrous cycle.
There was no effect of treatment on fertility or fecundity indices

REPRODUCTIVE FUNCTION: SPERM MEASURES (PARENTAL ANIMALS)
Qualitative testis staging did not indicate any abnormalities in the integrity of the various cell types present within the different stages of the spermatogenic cycle.

REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS)
There was no effect of treatment on mating, the majority of females mated within one oestrous cycle.
There was no effect of treatment on fertility or fecundity indices.

ORGAN WEIGHTS (PARENTAL ANIMALS)
There was no adverse effect of treatment on adult male or female organ weights.

GROSS PATHOLOGY (PARENTAL ANIMALS)
there was no effect of treatment on adult male or females

HISTOPATHOLOGY (PARENTAL ANIMALS)
Macroscopic findings
Most tissues were macroscopically unremarkable and the findings seen were generally consistent with the usual pattern of findings in rats of this strain and age. There were no macroscopic findings suggestive of effects of the test articles.
Microscopic findings
Microscopic findings were generally infrequent, of a minor nature and consistent with the usual pattern of findings in rats of this strain and age. There were no microscopic findings in treated animals due to effects of the test articles.

Effect levels

Dose descriptor:
NOAEL
Effect level:
> 250 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: overall effects

Target system / organ toxicity

Critical effects observed:
not specified

Any other information on results incl. tables

Group mating data

Test article

Control

AES Covance

RCF Covance

Group

1

2

3

Level (mg/kg/day)

0

250

250

 

5.1 Mating Performance

 

 

Group 1

Group 2

Group 3

 

 

 

 

 

  Group size at pairing:

 

 

 

 

       Male

 

10

10

10

       Female

 

10

10

10

 

 

 

 

 

  Number mating on Day of pairing

1

 2

 

 2

 

2

 3

 6

 4

 

3

 3

 1

 2

 

4

 2

 3

 

 

5

 

 

 

 

6

 

 

 

 

7

 

 

 2

 

8

 

 

 

 

9

 

 

 

 

10

 

 

 

 

11

 

 

 

 

12

 

 

 

 

13

 

 

 

 

14

 

 

 

 

15

 

 

 

 

Total:

10

10

10

 

 

 

 

 

  Median pre-coital time (days)

 

2.5

 2

 2

 

 

 

 

 

5.2 Summary of mating record

 

Group 1

Group 2

Group 3

 

 

 

 

  Number of females paired

10

10

10

 

 

 

 

  Pregnant:

 

 

 

     Mating from vaginal plugin situ

 

 2

 1

     Mating from positive smear:

 

 

 

        Sperm/oestrus/pro-oestrus

 1

 

 1

        Sperm/metoestrus/dioestrus

 

 

 

        Sperm/stage of oestrus not recorded

 8

 8

 7

     Absence of positive smear

 

 

 

     Pregnancy rate (%)

 9 (90.0)

10 (100.0)

 9 (90.0)

 

 

 

 

  Non-pregnant:

 

 

 

     Vaginal plugin situ

 

 

 

     Positive smear

 1

 

 1

     Absence of positive smear

 

 

 

 

 

 

 

  Total showing vaginal plugin situor positive smear

10

10

10

 

 

 

 

5.3 Mating indices

 

Group 1

Group 2

Group 3

 

 

 

 

  Number of paired males

10

10

10

  Number of paired females

10

10

10

 

 

 

 

  Number of mated males

10

10

10

  Number of mated females

10

10

10

 

 

 

 

  Number of males inducing pregnancy

 9

10

 9

  Number of pregnant females

 9

10

 9

 

 

 

 

  Mating index %

100.0

100.0

 90.9

 

 

 

 

  Fertility index % : male

 90.0

100.0

 90.0

                    : female

 90.0

100.0

 90.0

 

 

 

 

  Fecundity index % : male

 90.0

100.0

 90.0

                    : female

 90.0

100.0

 90.0

 

 

 

 

Group mean litter data

 

6.1 Pup numbers – females with live pups at Day 4post-partum

 

Group 1

Group 2

Group 3

 

 

 

 

 Number of females with live pups at Day 4post-partum

 9

 9

 9

 

 

 

 

 Mean duration of gestation (days)

22.3

22.1

22.3

 

 

 

 

 Mean number of implantation sites

11.8

12.4

12.7

 

 

 

 

 Mean number of pups born

10.8

12.1

11.4

 

 

 

 

 Mean number of pups alive Day 1

10.8

12.1

11.4

 

 

 

 

 Mean % male pups Day 1

42.4

42.8

47.3

 

 

 

 

 Mean number of pups alive Day 4

10.4

12.0

11.4

 

 

 

 

6.2 Neonatal survival indices – females with live pups at Day 21post-partum

 

 

Group 1

Group 2

Group 3

 

 

 

 

 

 

Post-implantation survival index %

 91.9

 97.4

 90.3

 

 

 

 

 

 

Live birth index %

100.0

100.0

100.0

 

 

 

 

 

 

Viability index 1 %

 97.0

 99.1

100.0

 

 

 

 

 

6.4 Summary necropsy data – pups surviving to Day 5 necropsy

 

Group 1

Group 2

Group 3

 

 

 

 

 Number of animals examined

93

108

103

 Number of animals with finding:

 

 

 

    Not remarkable (%)

78 (83.9)

100 (92.6)

90 (87.4)

 

 

 

 

 Animal - cannibalised

 

 

1

 Kidney - small

 1

 

 

 Liver - pale

 

 

1

       - pale area

 1

 

1

       - large

 

 

1

       - small

 

 

1

 Lung – red focus

 1

 

 

 Testis - dark

 

 

1

 Ureter - distension

 1

 

2

 Urinary bladder - distension

11

8

9

                 - abnormal contents

 

1

 

 

 

 

 


Applicant's summary and conclusion

Conclusions:
Administration of AES Covance and RCF Covance by oral gavage to male rats for at least 55 days and to female rats for at least 41 days at 250 mg/kg/day elicited no signs of adverse toxicity.
Executive summary:

The objective of the study was to provide a preliminary evaluation of the effects of the test articles, AES Covance and RCF Covance, on the reproductive/developmental toxicity in the rat.

Groups of 10 male and 10 female rats were given 250 mg/kg/day of either AES Covance or RCF Covance orally, by gavage, for two weeks prior to pairing, during the pairing period and until Day 4post-partum for the females and until the day before necropsy in Week 8 for the males. A similar group of 10 males and 10 females were given 1% (w/v) methylcellulose over the same period to act as controls.

There were no deaths during the study, and no treatment-related clinical observations.

Mean body weight gains and food intake were unaffected by treatment.

Mating data were unaffected by treatment. There was no effect of treatment on mean uterine/implantation data or mean litter data.

Mean organ weights were unaffected by treatment. At necropsy, macroscopic findings were unremarkable.

Microscopic examination revealed no treatment-related findings and there were no findings at testis staging due to effects of the test articles.

In conclusion, administration of AES Covance and RCF Covance by oral gavage to male rats for at least 55 days and to female rats for at least 41 days at 250 mg/kg/day elicited no signs of adverse toxicity.

The no‑observed‑adverse‑effect‑level (NOAEL) for reproductive and developmental toxicity was considered to be at least 250 mg/kg/day for both AES Covance and RCF Covance.

 

There are two possible activities of fibres , those mediated by their chemistry and those due to their morphology. When inhaled most fibres are cleared to the gut by the mucocilliary escalator or macrophage activity. Thus any reproductive effects would have to mediated through adsorbtion via the gut. The most likely source of any toxic activity would be through the chemistry of the material.

Unsurprisingly orally administered fibres at many times any dose that would arrive from inhalation had no effect on the reproductive efficiency or health of rats.