Phenol

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro cytogenicity / micronucleus study

Remarks:

Type of genotoxicity: chromosome aberration

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study meets generally accepted scientific standards. Final OECD Guideline 487 not available.

Data source

Reference

Reference Type:

publication

Title:

Evaluation of the micronucleus test in vitro using Chinese hamster cells: Results of four chemicals weakly positive in the In vivo micronucleus test

Author:

Miller BM, Pujadas E, Gocke E

Year:

1995

Bibliographic source:

Environ Mol Mutagen 26: 240-247

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

other: Draft OECD Guideline 487

Deviations:

no

GLP compliance:

not specified

Type of assay:

in vitro mammalian cell micronucleus test

Test material

Test material information

Constituent 1

Reference substance name:

Phenol

EC Number:

203-632-7

EC Name:

Phenol

Cas Number:

108-95-2

Molecular formula:

C6H6O

IUPAC Name:

phenol

Details on test material:

- Analytical purity: ca. 99%
- Impurities (identity and concentrations): no data
- Lot/batch No.: Lot 71663
- Stability under test conditions: no data, but stability in aqueous solution has been shown in other studies
- Storage condition of test material: no data
- Source: Sigma Chemical Co. (St. Louis, MO, USA).

Method

Target gene:

Target: chromosome

Species / strain

Species / strain / cell type:

Chinese hamster Ovary (CHO)

Details on mammalian cell type (if applicable):

CHO-K5 originally obtained from T. Hertner, Ciba, Summit, NJ

Metabolic activation:

with and without

Metabolic activation system:

S9 liver fraction of phenobarbital\beta-naphthoflavone -induced rats

Test concentrations with justification for top dose:

with metabolic action (MA): 350-2.000 µg/ml
without MA 10-250 µg/ml
see also Table below

Vehicle / solvent:

Medium

Controls

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Remarks:

1 µg/ml bleomycin (-MA) or 5 µg/ml cyclophosphamide (+MA)

Details on test system and experimental conditions:

METHOD OF APPLICATION:
Test substance in medium

DURATION
- Preincubation period: 24 h
- Exposure duration: after preincubation cells incubated with fresh medium plus test substance for 48 h (without MA) or incubated in fresh medium plus test substance plus MA system for 3 h, washed with PBS and incubated for further 45 h in fresh medium
- Fixation: after exposure period cells were washed with PBS and fixed with methanol containing I% acetic acid

NUMBER OF REPLICATIONS: minimum of two parallel cultures in two independent trials

NUMBER OF CELLS EVALUATED for MN: 1000 per culture

DETERMINATION OF CYTOTOXICITY: phenol tested up to concentrations showing signs of toxicity (reduction of the cell density by at least 25% compared to the control combined with an altered morphology of the cells)

DETERMINATION of micronuclei: Slides with cells air-dried and then treated with RNAse, after washing with demineralized water slides were air-dried again & stained with 5% Giemsa, only mononucleated cells with well-preserved cytoplasm containing five or fewer MN were scored (exclusion of apoptosis & nuclear fragmentation)

Evaluation criteria:

positive result: a dose-related effect showing an increase of the MN frequency over the control by about threefold or higher in at least one dose tested.

Statistics:

No data (not mandatory)

Results and discussion

Test results

Species / strain:

Chinese hamster Ovary (CHO)

Metabolic activation:

with and without

Genotoxicity:

positive

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Positive controls validity:

valid

Additional information on results:

Without MA
Phenol caused slight increase in MN accompanied by cytotoxicity; highest concentration of 250 µg/ml increased the MN frequency by factors of 3.5 and 3 (experiment A and B, respectively; see Table below); negative controls in parallel experiments ranged from 2.0 to 2.7% MN without MA (compare with concurrent control in the Table below).
With MA
350-2.000 µg/ml tested; highest concentration was highly toxic to the cells and increased the MN rate by a factor of about 4.8 (exp, A) and 7.0 (exp. B); negative controls in parallel experiments ranged from 2.4 to 3.9% MN with MA (compare with concurrent control)

Any other information on results incl. tables

Micronuclei induced by phenol in CHO cells

Concentration in µg/ml	% MN in Experiment A	% MN in Experiment B
Without metabolic action
0	2.7	2.3
10	3.0	-
50	4.5	2.6
100	6.6	3.6
175	9.9#	4.8
250	9.4#	6.8
With metabolic activation
0	3.3	2.4
350	3.1	3.8
475	3.4	-
600	2.9	-
800	4.4	-
1000	6.6	4.8
1500	-	5.8
2000	12.4#	16.9#
#: cytotoxic effects; -: not tested

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
positive (weak positive effects at cytotoxic dose levels)

Phenol induced increased incidence in micronuclei in CHO cells at high dose levels resulting also in cytotoxic effects.

Executive summary:

The study meets generally accepted scientific standards.

In two independent experiments CHO cells were exposed to phenol at dose levels of 350 -2.000 µg/ml (with metabolic action (MA); exposure duration 3 h, 45 h incubation without test substance) or 10 -250 µg/ml (without MA; exposure duration 48 h). 1000 cells per culture were scored for micronuclei. Phenol caused slight increase in MN accompanied by cytotoxicity (without MA: 250 µg/ml increased the MN frequency by factors of 3.5 and 3, respectively in experiment A & B; with MA: 2000 µg/ml was highly toxic and increased the MN rate by a factor of 4.8 and 7.0, respectively).

Conclusion: Phenol induced increased incidence in micronuclei in CHO cells at high dose levels resulting also in cytotoxic effects.