Trioctylphosphine oxide

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:

migrated information: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Study period:

From Apr. 23, 1986 to Jun. 30, 1986

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

The study was conducted according to a method comparable to OECD guideline 474 and conducted according to GLP. Deviation- Two treatments were given at 24 h interval; however, sample was collected after 6 h post second treatment (rather than 18-24 h, as recommended in the guideline). It is a read across study, hence maximum reliability rating of 2 assigned according to ECHA guidance.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1986

Report date:

1986

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)

Deviations:

yes

Remarks:

Bone marrow polychromatic erythrocyte were collected 6 h after the second test substance administration, rather than 18-24 h as per guideline.

GLP compliance:

yes

Type of assay:

micronucleus assay

Test material

Test material information

Constituent 1

Reference substance name:

A mixture of: hexyldioctylphosphineoxide; dihexyloctylphosphineoxide; trioctylphosphineoxide

IUPAC Name:

A mixture of: hexyldioctylphosphineoxide; dihexyloctylphosphineoxide; trioctylphosphineoxide

Constituent 2

Reference substance name:

A mixture of: hexyldioctylphosphineoxide; dihexyloctylphosphineoxide; trioctylphosphineoxide

EC Number:

403-470-9

EC Name:

A mixture of: hexyldioctylphosphineoxide; dihexyloctylphosphineoxide; trioctylphosphineoxide

IUPAC Name:

403-470-9

Test material form:

other: clear water-like liquid

Details on test material:

- Name of test substance: CYANEX® 923 Extractant
- Lot number: 870-3-SP
-State of the test substance: clear water-like liquid
-Storage conditions: at room temperature, protected from sunlight.

Test animals

Species:

mouse

Strain:

CD-1

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Breeding Iaboratory, Kingston, NY
- Age at study initiation: 6-8 weeks
- Weight at study initiation: male- 29. 6 to 37.1 grams, female- 22.8 to 31.8 grams
- Housing: AAAIAC accredited facility controlled environment of 74±6°F, 50±20% relative humidity, and a 12 h light/dark cycle
- Diet: ad libitum
- Water: ad libitum

Administration / exposure

Route of administration:

intraperitoneal

Vehicle:

- Vehicle used: corn oil

Details on exposure:

The test substance-vehicle mixture and the vehicle alone were administered by IP injection in two equal volumes of 10 mL/kg bw/d which were given 24 h apart. Positive control was injected through IP route at a dose level of 0.15 mg/kg.

Duration of treatment / exposure:

Two equal volumes of the test substance was 10 mL/kg bw/d which were given 24 h apart.

Post exposure period:

6 h

Doses / concentrations

Remarks:

Doses / Concentrations:
48; 159 and 476 mg/kg bw/d
Basis:

No. of animals per sex per dose:

5/sex/dose

Control animals:

yes, concurrent vehicle

Positive control(s):

-Name: Triethylenemelamine
- Route of administration: intraperitoneal
- Doses / concentrations: 0.15 mg/kg

Examinations

Tissues and cell types examined:

At the sacrifice, femurs were exposed, cut just above the knee and the bone marrow was aspirated into a syringe containing FBS. Using oil immersion, 1000 polychromatic erythrocytes were scored for the presence of micronuclei. The proportion of polychromatic erythrocytes to total erythrocytes and the number of micronucleated normocytes in the field of 1000 polychromatic erythrocytes were also enumerated.

Details of tissue and slide preparation:

Immediately following sacrifice, the femurs were exposed, cut just above the knee, and the bone marrow was aspirated into a syringe containing FBS. The bone marrow cells were transferred to a capped centrifuge tube containing approximately 1 mL cold FBS. The bone marrow cells were pelleted by centrifugation at approximately 1000 rpm for five minutes and the supernatant was drawn off, leaving off a small amount of serum with the remaining cell pellet. The cells were resuspended by aspiration with a capillary pipette and a small drop of bone marrow suspension was spread onto a clean glass slide. Two to four slides were prepared from each animal. The slides were aged overnight or longer and stained with May-Gruenwald-Giemsa and permanently mounted. Slides were coded using a random number table. Using medium magnification, an area of acceptable quality was selected such that the cells were well spread and stained.

Evaluation criteria:

In order to quantify the proliferation state of the bone narrow as an indicator of bone marrow toxicity, the proportion of polychromatic erythrocytes to total erythrocytes were presented for each animal and treatment group.The test substance was considered to induce a positive response if a treatment-related increase in micronucleated polychromatic erythrocytes was observed relative to the vehicle control (p≤0.05, Kastenbaum-Bowman Tables).

Statistics:

The incidence of micronucleated polychromatic erythrocytes per 1000 polychromatic erythrocytes were presented for each animal and treatment group. Statistical significance was determined using the Kastenbaum-Bowman tables.
In order to quantify the proliferation state of the bone narrow as an indicator of bone marrow toxicity, the proportion of polychromatic erythrocytes to total erythrocytes were presented for each animal and treatment group. Statistical comparison of treatment and solvent control groups were performed using the Kruskal-wallis test.

Results and discussion

Test results

Sex:

male/female

Genotoxicity:

negative

Toxicity:

yes

Vehicle controls validity:

valid

Positive controls validity:

valid

Additional information on results:

RESULTS OF RANGE-FINDING STUDY
- Dose range:
6,800; 3,400; 1,700; 850 and 425 mg test substance in corn oil/kg body weight

- Clinical signs of toxicity in test animals:
All mice that received 6,800; 3,400; 1,700 and 850 mg/kg and 2 of 5 males that received 425 mg/kg were found dead by Day 7 following dose administration. Prostration and lethargy were observed in mice from the 6,800; 3,400 and 1,700 mg/kg groups beginning 2 h after dosing. Daily observations on surviving animals included prostration, lethargy, rough hair coat and a rigid, distended abdomen. All surviving animals in the 425 mg/kg dose group had returned to normal by Day 3 following dosing.


RESULTS OF DEFINITIVE STUDY
- Induction of micronuclei (for Micronucleus assay):
The number of micronucleated polychromatic erythrocytes per 1000 Polychromatic Erythrocyte (PCE) was not significantly increased in the test substance-treated groups, regardless of sex or sacrifice time point

- Ratio of PCE/NCE (for Micronucleus assay):
There was no significant difference in the ratio of polychromatic erythrocytes to total erythrocytes between treatment groups per sacrifice time

Any other information on results incl. tables

Table:

					Micronucleated polychromatic erythrocytes
Treatment	Sex	Time following second dose	Number of mice	PCE/Total Erythyrocytes	Number per 1000 PCE’s (mean±SD)	Number per PCE’s scored
Corn oil 10 mL/kg/d	M	6	5	0.54	1.8±1.30	9/5000
	F	6	5	0.54	1.2±1.30	6/5000
Test substance (476 mg/kg bw/d)	M	6	5	0.54	2.6±1.71	11/4000
	F	6	5	0.52	1.8±0.50	7/4000
Test substance (159 mg/kg bw/d)	M	6	5	0.52	2.8±1.48	14/5000
	F	6	5	0.55	0.6±0.89	3/5000
Test substance (48 mg/kg bw/d)	M	6	5	0.55	1.4±1.34	7/5000
	F	6	5	0.54	0.8±0.84	4/5000
Triethylenemelamine (TEM) (0.15 mg/kg bw/d)	M	6	5	0.52	11.2±2.77	56/5000*
	F	6	5	0.52	19.2±9.34	96/5000*

* p≤0.05

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information): negative
The test substance did not increase the frequency of micronuclei in mice (Putman DL, 1986).

Executive summary:

An OECD Guideline (474) comparable study was conducted to assess the genotoxicity of the read across substance ‘A mixture of: hexyldioctylphosphineoxide; dihexyloctylphosphineoxide; trioctylphosphineoxide’ using a micronucleus cytogenetic assay in mice. The test substance-vehicle mixture and the vehicle alone were administered by intraperitoneal (IP) injection in two equal volumes of 10 mL/kg bw/d which were given 24 h apart. Positive control was injected through IP route at a dose level of 0.15 mg/kg bw. At the sacrifice (6 h following second dose), femurs were exposed, cut just above the knee and the bone marrow was aspirated into a syringe containing FBS. Using oil immersion, 1,000 polychromatic erythrocytes were scored for the presence of micronuclei. The proportion of polychromatic erythrocytes to total erythrocytes and the number of micronucleated normocytes in the field of 1,000 polychromatic erythrocytes were also enumerated. There was no significant difference in the ratio of polychromatic erythrocytes to total erythrocytes between treatment groups per sacrifice time. The number of micronucleated polychromatic erythrocytes per 1,000 PCE was not significantly increased in the test substance-treated groups, regardless of sex or sacrifice time point. TEM induced a significant increase in micronucleated polychromatic erythrocytes in male and female mice relative to the vehicle control. Under the study conditions, the read across substance did not increase the frequency of micronuclei in mice (Putman DL, 1986).