Registration Dossier

Data platform availability banner - registered substances factsheets

Please be aware that this old REACH registration data factsheet is no longer maintained; it remains frozen as of 19th May 2023.

The new ECHA CHEM database has been released by ECHA, and it now contains all REACH registration data. There are more details on the transition of ECHA's published data to ECHA CHEM here.

Diss Factsheets

Administrative data

Endpoint:
in vitro gene mutation study in bacteria
Remarks:
Type of genotoxicity: gene mutation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2014-07-28 to 2014-08-14
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: The study was conducted under GLP conditions according to OECD guideline 471.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2014
Report date:
2014

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Version / remarks:
(Adopted July 21, 1997)
Qualifier:
according to guideline
Guideline:
EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)
Version / remarks:
(May 31, 2008)
GLP compliance:
yes
Type of assay:
bacterial gene mutation assay

Test material

Constituent 1
Test material form:
solid: particulate/powder
Remarks:
migrated information: powder
Details on test material:
- Physical state: light yellow powder with lumps (determined at WIL Research Europe B.V.)
- Storage condition of test material: at room temperature in the dark

Method

Species / strainopen allclose all
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Details on mammalian cell type (if applicable):
- Properly maintained: yes
Species / strain / cell type:
E. coli WP2 uvr A
Details on mammalian cell type (if applicable):
- Properly maintained: yes
Metabolic activation:
with and without
Metabolic activation system:
S9
Test concentrations with justification for top dose:
- Experiment 1: 52, 164, 512, 1600, 5000 µg/plate (5% (v/v) S9-mix) with TA1535, TA1537, TA98
- Experiment 2: 52, 164, 512, 1600, 5000 µg/plate (10% (v/v) S9-mix) with TA1535, TA1537, TA98, TA100, WP2uvrA
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: Milli-Q water
Controls
Untreated negative controls:
yes
Remarks:
Milli-Q water
Negative solvent / vehicle controls:
no
True negative controls:
no
Positive controls:
yes
Positive control substance:
4-nitroquinoline-N-oxide
2-nitrofluorene
sodium azide
methylmethanesulfonate
other: ICR-191; 2-aminoant
Details on test system and experimental conditions:
METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Exposure duration: 48 +- 4 h


NUMBER OF REPLICATIONS: 3


DETERMINATION OF CYTOTOXICITY
- Method: the reduction of the bacterial background lawn, the increase in the size of the microcolonies and the reduction of the revertant colonies were examined.


Evaluation criteria:
A test substance is considered negative (not mutagenic) in the test if:
a) The total number of revertants in tester strain TA100 is not greater than two (2) times the concurrent control, and the total number of revertants in tester strains TA1535, TA1537, TA98 or WP2uvrA is not greater than three (3) times the concurrent vehicle control.
b) The negative response should be reproducible in at least one follow-up experiment.

A test substance is considered positive (mutagenic) in the test if:
a) The total number of revertants in tester strain TA100 is greater than two (2) times the concurrent control, or the total number of revertants in tester strains TA1535, TA1537, TA98 or WP2uvrA is greater than three (3) times the concurrent vehicle control.
b) In case a follow up experiment is performed when a positive response is observed in one of the tester strains, the positive response should be reproducible in at least one follow up experiment.
Statistics:
No formal hypothesis testing was done.

Results and discussion

Test resultsopen allclose all
Species / strain:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
other: not examined as the vehicle control was with Milli-Q water
Positive controls validity:
valid
Species / strain:
E. coli WP2 uvr A
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
other: not examined as the vehicle control was with Milli-Q water
Positive controls validity:
valid
Additional information on results:
RANGE-FINDING/SCREENING STUDIES: the tester strains TA100 and WP2uvrA were tested in triplicate, both with and without 5% (v/v) S9-mix. Concentrations: 1.7, 5.4, 17, 52, 164, 512, 1600, 5000 µg/plate. The highest concentration of test substance used in the subsequent mutation assay was 5000 µg/plate.

COMPARISON WITH HISTORICAL CONTROL DATA: yes
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.

Any other information on results incl. tables

Deviations:

1. In the second mutation experiment only two plates could be determined in tester strain WP2uvrA (absence of S9 -mix) at the dose level of 512 µg/plate

Evaluation: One of the triplicate cultures was infected. The two remaining plates showed responses well within the range and the testing of an extra plate would have given no additioinal information: therefore this deviation had no influence on the study result.

2. The positive control substance of tester strain WP2uvrA (second experiment, presence of S9 -mix) showed a response (mean plate count) which was not within the laboratory historical range.

Evaluation: The value (1357) was above the limit of the range (1271). The purpose of the positive control is as a reference for the test system, where a positive response is required to check if the test system functions correctly. Since the value was more than 3 times greater than the concurrent solvent control values, this deviation in the mean plate count of the positive control had no effect on the results of the study.

The study integrity was not affected by the deviations.

Table 1: Dose range finding test: Mutagenic response of Lithium salt of branched-aliphatic dicarboxylic acid in the Salmonella typhimurium reverse mutation assay and in the Escherichia coli reverse mutation assay

 Dose (µg/plate)  Mean number of revertant colonies/3 replicate plates (±S.D.) with one strain of Salmonella typhimurium and one Escherichia coli strain
   TA100 WP2uvrA 
 

Without S9-mix

 Positive control 887 +-13  1354 +/-133 
 Solvent control 101 +-9  27 +/-4 
1.7  103 +-12   33 +/-4
5.4  119 +-8   33 +/-4
 17 122 +-16   37 +/-9
 52 108 +-8   32 +/-5
 164  105 +-7  41 +/-6
 512  117 +-11 33 +/-11 
1600   98 +-12 33 +/-2 
 5000  107 +-2 *  29 +/-2*
  With S9 -mix**
 Positive control 1359 +-158  152 +/-28 
 Solvent control  92 +-13  44 +/-9
 1.7  105 +-6  35 +/-4
 5.4  92 +-18  39 +/-12
 17  98 +-23  30 +/-11
 52  105 +-11  42 +/-6
164   93 +-5  34 +/-4
512   98 +-4  35 +/-7
 1600  98 +-11  38 +/-3
 5000  102 +-9*  34 +/-8*

*No precipitate and Normal bacterial background lawn

** Plate incorporation assay (5% S9)

Table 2: Experiment 1: Mutagenic response of Lithium salt of branched-aliphatic dicarboxylic acid in the Salmonella typhimurium reverse mutation assay

 Dose (µg/plate) Mean number of revertant colonies/3 replicate plates (±S.D.) with different strains of Salmonella typhimurium.
      TA1535   TA2537 

  TA98 

 

Without S9-mix

    
 Positive control   603 +/-29    610 +/-73  942 +/-11 
  Solvent control    18 +/-3  9 +/-4  10 +/-5
  52  15 +/-2  5 +/-4  12 +/-4
  164   13 +/-4  4 +/-1  15 +/-10
   512    20 +/-5  8 +/-3  16 +/-2
 1600   22 +/-5  3 +/-2  18 +/-2
  5000   15 +/-5*  4 +/-2*        16 +/-3*
     

With S9-mix**

 
 
 Positive control  257 +/-17 228 +/-36  655 +/-23 
 Solvent control  13 +/-2  7 +/-3  19 +/-6
 52  16 +/-2  7 +/-2  25 +/-7
 164  17 +/-4  4 +/-2  17 +/-3
 512  13 +/-3  6 +/-5  23 +/-3
 1600  14 +/-6  9 +/-5  22 +/-6
 5000  13 +/-2 *  8 +/-2*  21 +/-2*
       

*No precipitate and Normal bacterial background lawn

**Plate incorporation assay (5% S9)

Table 3: Experiment 2: Mutagenic response of Lithium salt of branched-aliphatic dicarboxylic acid in the Salmonella typhimurium reverse mutation assay and in the Escherichia coli reverse mutation assay

 Dose (µg/plate) Mean number of revertant colonies/3 replicate plates (±S.D.) with different strains of Salmonella typhimurium and one Escherichia coli strain
  TA1535  TA1537  TA98  TA100  WP2uvrA 
  Without S9 -mix             
 Positive control  599 +/-13 592 +-103   847 +/-32 891 +/-10  304 +/-18 
 Solvent control  10 +/-4  4 +/-3  29 +/-5  121 +/-8  36 +/-6
 52  10 +/-2  10 +/-6  37 +/-5  123 +/-6  48 +/-6
164   14 +/-3  6 +/-2  20 +/-3  126 +/-12  48 +/-12
 512  15 +/-1  6 +/-2 39 +/-13   98 +/-17  33 +/-8***
 1600 16 +/-3   8 +/-1  32 +/-6  109 +/-11  43 +/-2
5000   13 +/-3*  6 +/-3*  31 +/-12*  104 +/-10*  30 +/-2*
     With S9 -mix**          
 Positive control  173 +/-31 454 +/-30  519 +/-23   1174 +/-80 1357 +/-8 
 Solvent control  14 +/-5  11 +/-2  48 +/-5 122 +/-16  38 +/-9 
 52  14 +/-3  11 +/-5  32 +/-7 124 +/-13  32 +/-4 
 164  13 +/-4  6 +/-3  41 +/-11 103 +/-13  26 +/-6 
 512  16 +/-5  9 +/-3  36 +/-8  107 +/-23  48 +/-3
 1600  18 +/-1  12 +/-4  43 +/-4  105 +/-16  48 +/-8
 5000  18 +/-4*  10 +/-6*  46 +/-4*  105 +/-14*  39 +/-8*

*No precipitate and Normal bacterial background lawn

**Plate incorporation assay (10% S9)

***one plate infected: Mean of two plates

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
negative

Based on the results of this study it is concluded that Lithium salt of branched-aliphatic dicarboxylic acid is not mutagenic in the Salmonella typhimurium reverse mutation assay and in the Escherichia coli reverse mutation assay.
Executive summary:

Evaluation of the mutagenic activity of Lithium salt of branched-aliphatic dicarboxylic acid in theSalmonella typhimurium reverse mutation assay and the Escherichia colireverse mutation assay.

 

The test substance was tested in the Salmonella typhimurium reverse mutation assay with four histidine-requiring strains of Salmonella typhimurium (TA1535, TA1537, TA98 and TA100) and in the Escherichia coli reverse mutation assay with a tryptophan-requiring strain of Escherichia coli (WP2uvrA). The test was performed in two independent experiments in the presence and absence of S9-mix (rat liver S9-mix induced by Aroclor 1254).

 

The study procedures described in this report were based on the most recent OECD and EC guidelines.

 

The test substance was a light yellow powder with lumps with a purity of>=99 %. The test substance was dissolved in Milli-Q water.

 

In the dose range finding test, the test substance was tested up to concentrations of 5000 µg/plate in the absence and presence of S9-mix in the strains TA100 and WP2uvrA. The test substance did not precipitate on the plates at the highest dose level. The bacterial background lawn was not reduced at any of the concentrations tested and no biologically relevant decrease in the number of revertants was observed. Results of this dose range finding test were reported as part of the mutation assay.

 

Based on the results of the dose range finding test, the test substance was tested in the first mutation assay at a concentration range of 52 to 5000 µg/plate in the absence and presence of 5% (v/v) S9-mix in tester strains TA1535, TA1537 and TA98. In a follow-up experiment of the assay with additional parameters, the test substance was tested at the same concentration range as the first assay in the absence and presence of 10% (v/v) S9-mix in tester strains TA1535, TA1537, TA98, TA100 and WP2uvrA. The bacterial background lawn was not reduced at any of the concentrations tested and no biologically relevant decrease in the number of revertants was observed.

 

Lithium salt of branched-aliphatic dicarboxylic acid did not induce a significant dose-related increase in the number of revertant (His+) colonies in each of the four tester strains (TA1535, TA1537, TA98 and TA100) and in the number of revertant (Trp+) colonies in tester strain WP2uvrA both in the absence and presence of S9-metabolic activation. These results were confirmed in a follow-up experiment.

 

In this study, the negative and strain-specific positive control values were within the laboratory historical control data ranges indicating that the test conditions were adequate and that the metabolic activation system functioned properly, with deviations that were determined not to have affected the results.

 

Based on the results of this study it is concluded that Lithium salt of branched-aliphatic dicarboxylic acid is not mutagenic in theSalmonella typhimuriumreverse mutation assay and in the Escherichia colireverse mutation assay.