Acacia mearnsi, ext., reaction products with ammonium chloride and formaldehyde

Genetic toxicity: in vivo

Administrative data

Endpoint:

in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus

Remarks:

Type of genotoxicity: chromosome aberration

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From 10/12/2006 to 19/01/2007

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study conducted according to GLP compliance and International Guidelines

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2007

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)

GLP compliance:

yes

Type of assay:

micronucleus assay

Test material

Test material information

Constituent 1

Reference substance name:

ACQUAPOL C1

IUPAC Name:

ACQUAPOL C1

Test material form:

other: liquid

Details on test material:

Test Article: Acquapol C1
Stated chemical composition: Aqueous solution of quaternary ammonium tannate

Test animals

Species:

mouse

Strain:

Swiss

Sex:

male/female

Details on test animals or test system and environmental conditions:

Test system:
Mice (Swiss lineage) were used at ages between 7 and 10 weeks, having 22-29 grams of live weight. 5 animal/dose were used to determine the DL50 and 10 animal/dose were used for the final test, males and females. The females used were nulliparous and non-pregnant.

Test conditions:
The animals were acclimated to the laboratory conditions at least 5 days prior to the beginning of the experiment. They were mantained with ventilation of 10 to 15 air changes per room per hour, temperature between 19 and 25°C, relative air humidity between 30 and 70% and photoperiod of 12 hours in the light and 12 hours in the dark. The diet comprised commercial feed, with supplemetnation of filtered/potable water, both provided ad libitum. The animals were maintained in polycarbonate boxes with autoclaved Pinus shavings.

Administration / exposure

Route of administration:

intraperitoneal

Vehicle:

deionised water

Details on exposure:

intraperitoneal injection at 24-h interval

Duration of treatment / exposure:

24 hours

Frequency of treatment:

Two intraperitoneal injections at volumes of 0.5 mL /animal were administered at a 24-h interval.

Post exposure period:

24 hours

Doses / concentrations

Doses / concentrations 1

Remarks:

Doses / Concentrations:
112.5
Basis:
nominal conc.
mg/kg

Doses / concentrations 2

Remarks:

Doses / Concentrations:
75
Basis:
nominal conc.
mg/kg

Doses / concentrations 3

Remarks:

Doses / Concentrations:
37.5
Basis:
nominal conc.
mg/kg

No. of animals per sex per dose:

20 animals (5 animals/dose) were used to determine the DL50 and 10 animals/dose (5 males and 5 females) were used to the final test, males and females.

Control animals:

yes

Positive control(s):

For the positive control was used Cyclophospharride, following the same procedure.

Examinations

Tissues and cell types examined:

Bone marrows were exposed by removing a portion of the femur.

Details of tissue and slide preparation:

The animals are exposed to a test article by the adequate route and subsequentely sacrificed. These animals are sacrificed and their bone marrows are extracted and fixed on slides. The polychromatic Erythrocytes are assessed for the level of micronuclei found by microscopy.
CRITERIA FOR DOSE SELECTION:
Determination of intraperitoneal DL50:
The product ACQUAPOL C1 was applied to 20 mice (5 animal/dose) at 4 doses by intraperitoneal route. Two injections were administered at a 24-h interval. The animals remained under observation for 10 days for mortality. After this period, the intraperitoneal DL50 was estabilished by the Thompson & Weill Method. the intraperitoneal DL50 value was 150 mg/kg.
Application of the product and dose levels:
The product was applied to animals at concentrations of 37,5; 75 and 112,5 mg/kg, corresponding to 25, 50 and 75% DL50.

TREATMENT AND SAMPLING TIMES:
10 animals per dose were used, 5 females and 5 males. For the tested groups, two interperitoneal injections were administered (0,5 mL) with 24-h interval. For the negative control, deionized water was used and for the positive control Cyclophosphamide, all groups followed the same procedure. After 24 hours, all animals were sacrificed.
DETAILS OF SLIDE PREPARATION:
Once the animal were sacrificed, the femurs were removed, these were cleaned, with muscles and other tissues being removed. Later the bone marrows were exposed by removing a portion of the femur. The marrows were removed by infusion and suction of a fetal serum solution.
Later these solution were centrifuged at 1000 rpm for 5 minutes. Subsequently, they were processed and slides were prepared using an Eosin-Methylene Blue solution (Leishman Method).
METHOD OF ANALYSIS:
The slides were all coded and examined under light microscope at 1000x magnification. For each animal 1000 polychromatic and normochromatic erythrocytes were verified.

Results and discussion

Test results

Sex:

male/female

Genotoxicity:

negative

Toxicity:

yes

Vehicle controls validity:

not examined

Negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

Using the readings performed, no significant differences were found between the test group and the negative control group for the number of micronuclei found. The positive control showed micronuclei in sufficient quantity, which validates the assay. Test group: .5.5% of cells showed micronucleus. Negative control group: 5.7% of cells showed micronucleus. Positive control group:10.1% of cells showed micronucleus.

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information): negative
The tested substance Acacia Mearnsi (ACQUAPOL C1) is not mutagenic according to the testing protocol OECD 474.

Executive summary:

A Micronucleus Assay on Mice was conducted to study potential harmful effects of the test article ACQUAPOL C1 on chromosomes and on the mitosis processes of cells. The test article was used at the doses of 25%, 50% and 75% (112,5, 75 and 37.5 mg/kg) of 0.4 to 0.5 mL/animal for 24 hours. The tested animals were maintained for 24 hours following the application and later sacrificed for the preapration of the slides. No increase in the proportion of micronuclei was observed in the etrythrocytes in the test group in comparison with the control group. The test article ACQUAPOL C1 was considered non-mutagenic when applied by intraperitoneal route to mice,

Categories Display