Registration Dossier

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:
2008-12-11 - 2009-01-16
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2009
Report Date:
2009

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)
Deviations:
no
GLP compliance:
yes
Type of assay:
micronucleus assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): #5 Biphenyl diphenyl phosphate (mixture of the phosphates of ortho-Phenyl Phenol and Phenol and containing 25 % of Triphenyl Phosphate)
- Substance type: organic
- Physical state:Colorless to light yellowish clear liquid
- Analytical purity: > 99 %
- Impurities (identity and concentrations): no data
- Lot/batch No.: F01003
- Expiration date of the lot/batch: 2009-03-15
- Stability under test conditions: no data
- Storage condition of test material: room temperature; store in a cool, dry, well-ventilated area; keep containers tightly closed when not in use.

Test animals

Species:
mouse
Strain:
CD-1
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: BioLASCO Taiwan Co. Ltd.
- Age at study initiation: approximately 7 weeks old
- Weight at study initiation: 29.4 g ~ 32.3 g for males and 22.8 g ~ 25.3 g for females
- Assigned to test groups randomly: [no/yes, under following basis: ] no data
- Fasting period before study: no data
- Housing: polycarbonate cage; cage feeder was composed of stainless steel
- Diet (e.g. ad libitum): no data (autoclaved laboratory rodent diet 5010, LabDiet, PMI Nutrition - International, Brentwood MO, USA
- Water (e.g. ad libitum): ad libitum (autoclaved RO water)
- Acclimation period: 7 days in the testing room before dosing

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 ± 2°C
- Humidity (%): 60 ± 20%
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): 12 hours light and 12 hours dark

IN-LIFE DATES: no data

Administration / exposure

Route of administration:
oral: unspecified
Vehicle:
none
Duration of treatment / exposure:
single dosing, dosing day is defined as Day 1
Frequency of treatment:
not applicable
Post exposure period:
5 days
Doses / concentrationsopen allclose all
Remarks:
Doses / Concentrations:
0.5 g/kg b.w.
Basis:
nominal in water
Remarks:
Doses / Concentrations:
1 g/kg b.w.
Basis:
nominal in water
Remarks:
Doses / Concentrations:
2 g/kg b.w.
Basis:
nominal in water
No. of animals per sex per dose:
5 male / 5 female
Control animals:
yes, historical

Examinations

Tissues and cell types examined:
- the percentage of PCE (polychromatic erythrocytes) in erythrocytes (PCE %)
- micronucleus frequency in thousand PCE (MN%o PCE)

Results and discussion

Test results
Sex:
male/female
Genotoxicity:
negative
Toxicity:
not specified
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid

Any other information on results incl. tables

RESULTS

1) Mortality

Mortalities of all groups were 0 %. There was no abnormal clinical sign observed in the study. The mortality was summarized in Table 1:

Table 1. In vivo Mammalian Erythrocyte Micronucleus Test - Mortality

Groups Dose Day 0 Day 1 Day 2 Day 3 Day 4 Mortality*
Animals found dead
Negative Control Sterile water 0 0 0 0 0 0/10
Positive Control Cyclophosphamide  80 mg/kg b.w. 0 0 0 0 0 0/10
0.5 g/kg b.w. 0 0 0 0 0 0/10
#5 Biphenyl diphenyl phosphate (CAS number: 132-29-6) 1 g/kg b.w. 0 0 0 0 0 0/10
2 g/kg b.w. 0 0 0 0 0 0/10

Note: No animal death occurred.

2) Body Weight

The mean body weight was summarized in Table 2:

Table 2. In vivo Marrrmaliari Erythrocytre Micronucleus Test - Body Weight

Groups   Animal Body Weight (g, mean + SD, n=5)
Dose Male Female
Day 1 Day 5 Day 1 Day 5
Negative Control Sterile water 30.7 ± 1.1 31.8 ± 1.1 24.0 ± 0.7 24.9 ± 0.6
Positive Control
Cyclophosphamide 
80 mg/kg b.w. 30.7 ± 1.1 31.7 ± 1.0 24.0 ± 0.7 24.9 ± 0.6
#5 Biphenyl diphenyl phosphate (CAS number: 132-29-6) 0.5 g/kg b.w. 30.6 ± 1.0 31.6 ± 1.1 23.9 ± 0.6 24.6 ± 0.5
1 g/kg b.w. 30.5 ± 1.0 31.5 ± 1.1 23.9 ± 0.8 24.7 ± 0.8
2 g/kg b.w. 30.7 ± 1.1 31.7 ± 1.1 24.1 ± 0.8 24.9 ± 0.7

Note: Body weights were presented in mean + standard deviation. After analyzing with t-est, there were no significant differences between all groups.

3) Percentage of PCE in Erythrocytes

The PCE percentage of negative control group at 24, 48 and 72 hours was 3.92 ± 0.56 %, 3.51 ± 0.81 % and 3.85 ± 0.49 %, respectively. The PCE percentage of positive control group was decreased with time and 20 % lower than the PCE percentage of negative control group, indicated that cyclophosphamide inhibited erythropoiesis. All testing groups were not significantly different from negative control group, indicated that "#5 Biphenyl diphenyl phosphate" did not affect erythropoiesis. The percentage of PCE was summarized in Table 3:

Table 3. In vivo Micronucleus Test Hours after Dosing

Groups Dose PCE%, mean ± SD, n=10
24 hours 48 hours 72 hours
Negative Control Sterile water 3.92 ± 0.56 3.51 ± 0.81 3.85 ± 0.49
Positive Control
Cyclophosphamide
80 mg/kg b.w. 3.12 ± 0.63 1.52 ± 0.21 0.61 ± 0.14
#5 Biphenyl diphenyl phosphate (CAS number: 132-29-6) 0.5 g/kg b.w. 3.74 ± 0.35 3.70 ± 0.53 3.62 ± 0.63
1 g/kg b.w. 3.76 ± 0.43 4.01 ± 0.45 3.75 ± 0.38
2 g/kg b.w. 3.66 ± 0.48 4.02 ± 0.35 3.59 ± 0.43

Note:

1. The PCE % was presented in Mean ± SD.

2. Positive control group represented inhibition on erythropoiesis

4) Micronucleus frequency

The micronucleus frequencies in PCE were examined with fluorescent microscope and summarized in Table 4:

Table 4. In vivo Mammalian Erythrocyte Micronucleus Test - Micronucleus Frequency in PCE at 24, 48 and 72 hours after dosing

Groups Dose PCE%, mean ± SD, n=10
24 hours 48 hours 72 hours
Negative Control Sterile water 1.50 ± 0.71 1.20 ± 1.14 1.10 ± 0.57
Positive Control
Cyclophosphamide
80 mg/kg b.w. 6.00 ± 1.33* 20.60 ± 3.84* 9.80 ± 1.81*
#5 Biphenyl diphenyl phosphate (CAS number: 132-29-6) 0.5 g/kg b.w. 1.30 ± 0.48 1.30 ± 1.06 0.80 ± 0.63
1 g/kg b.w. 1.50 ± 1.18 1.30 ± 0.48 1.10 ± 0.99
2 g/kg b.w. 1.60 ± 0.70 2.00 ± 0.82 1.40 ± 0.70

Note:

1. Micronucleus frequency (%o PCE) was presented in Mean ± SD.

2. The results were analyzed with Poisson distribution, and the star symbol (*) indicated that it was significant different (p<0.05) from negative control group. Positive control group represented genotoxicity at 24 ~ 72 hours.

The micronucleus frequencies in PCE of negative control group at 24, 48 and 72 hours (1.50 ± 0.71, 1.20 ± 1.14 and 1.10 ± 0.57 %OPCE) and positive control group at 24, 48 and 72 hours (6.00 ± 1.33, 20.60 ± 3.84 and 9.80 ± 1.81 %o PCE) were both conformed to the criteria.

After statistic analyzing, all three testing groups were not significantly different from negative control group.

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information): negative
The animals which were administrated orally up to the dosage of 2 g/kg of "#5 Biphenyl diphenyl phosphate" presented negative result in micronucleus test. The test substance had neither genotoxicity in mice nor inhibition in erythropoiesis.
Executive summary:

This study was to evaluate the genotoxicity of "#5 Biphenyl diphenyl phosphate" with mammalian peripheral blood micronucleus test. The experimental design was based upon OECD Guideline 474.

The test article, "#5 Biphenyl diphenyl phosphate", was suspended in sterile water to 0.5, 1, and 2 g/kg for the three testing groups. Including sterile water as negative control, each material was administrated orally. Cyclophosphamide (80 mg/kg) was as positive control and injected intraperitoneally. After 24, 48 and 72 hours, collected blood samples from tail vein, smeared on acridine orange-coated slide, and examined the percentage of polychromatic erythrocytes (PCE%) and micronucleus frequency ( %o PCE) with fluorescent microscope.

The results indicated that all three testing groups were not statistically different from negative group in both PCE percentage and micronucleus frequency.

In conclusion, "#5 Biphenyl diphenyl phosphate" presented negative result in micronucleus test. The test substance had neither genotoxicity in mice nor inhibition in erythropoiesis.