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Diss Factsheets

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
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: acceptable, well documented study report which meets basic scientific principles

Data source

Reference
Reference Type:
other: microfiche
Title:
Unnamed
Year:
1989
Report date:
1989

Materials and methods

Test guideline
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)
Deviations:
yes
Remarks:
- only 1000 immature erythrocytes per animal were scored for the incidence of micronucleated erythrocytes
GLP compliance:
yes
Type of assay:
micronucleus assay

Test material

Constituent 1
Chemical structure
Reference substance name:
Octadecyl isocyanate
EC Number:
204-019-7
EC Name:
Octadecyl isocyanate
Cas Number:
112-96-9
Molecular formula:
C19H37NO
IUPAC Name:
1-isocyanatooctadecane

Test animals

Species:
mouse
Strain:
ICR
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Harlan Sprague Dawley, Inc. (Frederick, MD)
- Age at study initiation: 6 - 8 weeks
- Weight at study initiation: males: 27-36 g; females: 18-26 g
- Housing: five per cage
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: at least 5 days


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 23 +/- 3
- Humidity (%): 50 +/- 20
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): 12 / 12

Administration / exposure

Route of administration:
intraperitoneal
Vehicle:
- Vehicle(s)/solvent(s) used: none
Duration of treatment / exposure:
single i.p. injection
Frequency of treatment:
once
Doses / concentrations
Remarks:
Doses / Concentrations:
0, 0.5, 2.5, 5.0 ml/kg
Basis:
other: actual injected dose
No. of animals per sex per dose:
5
Control animals:
yes, concurrent vehicle
Positive control(s):
triethylenemelamine (TEM) dissolved in distilled water, single i.p. injection with 0.25 mg/kg bw

Examinations

Tissues and cell types examined:
Using oil immersion, 1000 polychromatic erythrocytes (PCEs) per animal 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:
At the scheduled sacrifice times, five mice per sex per treatment were sacrificed by CO2 asphyxiation. Immediately following sacrifice, the femurs were exposed, cut just above the knee, and the bone marrow was aspirated into a syringe containing fetal bovine serum. The bone marrow cells were transferred to a capped centrifuge tube containing approximately 1 mL fetal bovine serum. The bone marrow cells were pelleted by centrifugation at approximately 100 x g for five minutes and the supernatant was drawn off, leaving a small amount of serum with the remaining cell pellet. The cells were resuspended by aspiration with a capillary pipet and a small drop of bone marrow suspension was spread onto a clean glass slide. Two to four slides were prepared from each mouse. The slides were fixed in methanol, stained with May-Gruenwald-Giemsa and permanently mounted. Slides were coded using a random number table by an individual not involved with the scoring process. Using medium magnification, an area of acceptable quality was selected such that the cells were weIl spread and stained.
Evaluation criteria:
The mean incidence of micronucleated polychromatic erythrocytes must not exceed 5/1000 polychromatic erythrocytes (0.5 %) in the negative control. The incidence of micronucleated polychromatic erythrocytes in the positive control group must be significantly increased relative to the negative control group (p<=0.05, Kastenbaum-Bowman Tables).
Statistics:
Statistical significance will be determined using the Kastenbaum-Bowman tables which are based on the binomial distribution.

Results and discussion

Test results
Sex:
male/female
Genotoxicity:
negative
Toxicity:
no effects
Vehicle controls validity:
not applicable
Negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
GENERAL TOXICITY:
One male receiving 5 ml/kg and one female receiving 2.5 ml/kg died immediately followinq the i.p. injection; death was assumed to be a dosing accident and these mice were replaced. All animals appeared normal several minutes after dose administration and during the course of the study.

RATIO OF POLYCHROMATIC ERYTHROCYTES TO TOTAL ERYTHROCYTES:
The incidence of micronucleated polychromatic erythrocytes per 1000 polychromatic erythrocytes scored and the proportion of polychromatic erythrocytes per total erythrocytes are summarized and presented for each treatment group by sacrifice time in Table 1. No reduction in the ratio of
polychromatic erythrocytes to total erythrocytes in males or females suggests no bone marrow toxicity.

MICRONUCLEATED POLYCHROMATIC ERYTHROZYTES:
The number of micronucleated polychromatic erythrocytes per 1000 polychromatic erythrocytes was not statistically increased in males and females, regardless of dose level or bone marrow collection times (p>0.05, Kastenbaum-Bowman Tables). TEM induced a significant increase in micronucleated polychromatic erythrocytes in male and female mice relative to the negative
control (p<=0.05, Kastenbaum-Bowman Tables).

Any other information on results incl. tables

Table 1: Summary of results from the bone marrow micronucleus test with octadecyl isocyanate

 Treatment

Sex

Time (hr)

 Number of mice

PCE/total erythrocytes

(Mean)

Micronucleated poly- chromatic number per 1000 PCEs (Mean ± S.D.

Erythrocytes number

per PCEs scored  

 Water            
 5 ml/kg  M 24 5 0.55 0.6 ± 0.55 3 / 5000
  48 5 0.57 0.4 ± 0.55 2 / 5000

72

5 0.56  0.8 ± 0.84   4 / 5000 
 F 24 5 0.56 1.2 ± 0.84 6 / 5000
  48 5 0.54  2.2 ± 1.79 11 / 5000
 

72 

5 0.57  0.8 ± 0.84 4 / 5000 

Octadecyl isocyanate

 0.5 ml/kg

M 24  5 0.49 0.6 ± 0.89  3 / 5000 
  48 0.60 0.8 ± 0.84  4 / 5000 
    72 0.55  1.0 ± 0.71 5 / 5000 
  F 24  0.57  1.4 ± 1.14 7 / 5000 
  48 0.63 1.0 ± 0.71  5 / 5000 
    72 0.55   0.6 ± 0.89   3 / 5000 

2.5 ml/kg

24  0.50    0.8 ± 1.30   4 / 5000 
  48 5 0.58  0.8 ± 0.45  4 / 5000 
  72 0.51 0.4 ± 0.55  2 / 5000 

24  0.48  0.6 ± 0.89  3 / 5000 

48  0.61  1.2 ± 1.10  6 / 5000 
  72  0.52  0.2 ± 0.45  1 / 5000 

5.0 ml/kg

M 24  0.59 0.6 ± 0.89 3 / 5000 
  48  5 0.58  0.2 ± 0.45  1 / 5000 

 

72  0.54  0.2 ± 0.45  1 / 5000 
  24  0.55 0.4 ± 0.55  1 / 5000 
    48  0.58  0.0 ± 0.00  0 / 5000 
    72  0.55  0.0 ± 0.00  0 / 5000 

 TEM

           

0.25 mg/kg 

 M 24 5 0.47   48.4 ± 21.01 242 / 5000 * 
   F 24  0.49  39.0 ± 4.64 195 / 5000 * 

* p ≤ 0.05 (Kastenbaum-Bowman Tables)

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information): negative
Executive summary:

Male and female ICR mice were exposed to 0.5, 2.5 or 5 ml/kg of octadecyl isocyanate which was administered as a single i.p. injection. Bone marrow cells, collected 24, 48 and 72 hours after treatment, were examined microscopically for micronucleated polychromatic erythrocytes. No change in the ratio of polychromatic erythrocytes to total erythrocytes was observed in male or female mice in the test article treated groups,suggesting that the test article did not induce bone marrow toxicity. No significant increases in micronucleated polychromatic erythrocytes were observed at 24, 48 or 72 hours after dose administration in males or females. The results of the assay indicate that under the conditions described, octadecyl isocyanate did not induce a significant increase in micronucleated polychromatic erythrocytes in male or female ICR mice. Therefore, octadecyl isocyanate was concluded to be negative in the mouse micronucleus assay.