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Toxicological information

Genetic toxicity: in vitro

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Administrative data

Endpoint:
in vitro cytogenicity / micronucleus study
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
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
The study was well documented and meets generally accepted scientific principles, but was not conducted in compliance with GLP. The method used was similar to the appropriate OECD guideline, with acceptable restrictions. The restrictions were that no activation was used. Read across to the registered substance is considered scientifically justified.

Data source

Reference
Reference Type:
publication
Title:
Clastogenic effects of food additive citric acid in human peripheral lymphocytes
Author:
Yilmaz S, Uenal F, Yuebasioglu D
Year:
2008
Bibliographic source:
Cytotechnology 56: 137-144

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
other: OECD draft guideline 487 2009
Deviations:
yes
Remarks:
no activation
GLP compliance:
not specified
Type of assay:
in vitro mammalian cell micronucleus test

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): citric acid

Method

Species / strain
Species / strain:
lymphocytes: peripheral human
Metabolic activation:
without
Test concentrations with justification for top dose:
50, 100, 200, 3000 µg/ml
Vehicle:
- Vehicle(s)/solvent(s) used: water
- Justification for choice of solvent/vehicle: none given
Controls
Negative controls:
no
Solvent controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
cyclophosphamide
Details on test system and conditions:
METHOD OF APPLICATION: in medium

DURATION
- Exposure duration: 72 hours
- Expression time (cells in growth medium): 72 hours
- Fixation time (start of exposure up to fixation or harvest of cells): 72 hours

CYTOKINESIS INHIBITOR (micronucleus assays): actin polymerisation inhibitor cytochalasin B (cytoB) 5.2 µg/ml added after 48 hours.
STAIN (for cytogenetic assays): Giesma

NUMBER OF REPLICATIONS: duplicate cultures (two donors: healthy non-smokers, 27 years, 1 male, 1 female)

NUMBER OF CELLS EVALUATED: 1000 binucleate (BN) cells/donor (micronucleus analysis), 500/donor (CBPI)

DETERMINATION OF CYTOTOXICITY
- Method: Cytokinesis-Block Proliferation Index (CBPI)

Evaluation criteria:
None given in report
Statistics:
difference in % abnormal cells: z-test; dose-response relationship: correlation and regression coefficient.

Results and discussion

Test results
Species / strain:
lymphocytes: peripheral human lymphocytes
Metabolic activation:
without
Genotoxicity:
positive
Cytotoxicity:
yes
Remarks:
3000 µg/ml
Vehicle controls valid:
yes
Negative controls valid:
not examined
Positive controls valid:
yes
Additional information on results:
TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH: reported "without changing the pH of the medium"

RANGE-FINDING/SCREENING STUDIES: not reported

COMPARISON WITH HISTORICAL CONTROL DATA: not compared with historical control data

Any other information on results incl. tables

Table 1 Induction of micronuclei in cultured human lymphocytes treated with citric acid

Test substance

Treatment

BN cells scored

Distribution of BN cells according to the no. of MN

MN (%)

Cytokinesis-block proliferation index (CBPI)

Period (hour)

Dose (μg ml−1)

-1

-2

-3

-4

Negative control

48

0

2,000

6

0

0

0

0.30 ± 0.12

1.84 ± 0.30

Positive control

48

0.1

2,000

220

20

0

0

13.0 ± 0.75

1.30 ± 0.25

Citric acid

48

50

2,000

33

0

0

0

1.65 ± 0.28*

1.43 ± 0.27

100

2,000

45

1

0

0

2.35 ± 0.34*

1.41 ± 0.26

200

2,000

48

0

0

1

2.60 ± 0.36*

1.34 ± 026

3,000

Toxic

Toxic

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
positive without metabolic activation

Citric acid has been tested according to a method that is similar to OECD draft guideline 487 (in vitro mammalian cell micronucleus test). A statistically significant, dose-dependent increase in the percentage of binucleate cells with micronuclei was observed. It is concluded that the test substance is positive for the induction of micronuclei in cultured human lymphocytes under the conditions of this study