Reaction products of monoethanolamine and boric acid (1:1)

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Link to relevant study records

Reference

Reference 1

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

16 Jun 2017 - 10 Jul 2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2

Metabolic activation:

with and without

Metabolic activation system:

Rat liver microsomal enzymes (S9 homogenate), prepared from male Sprague Dawley rats that had been injected intraperitoneally with Aroclor 1254 (500 mg/kg body weight).

Test concentrations with justification for top dose:

Dose-range Finding Test
Selection of an adequate range of doses was based on a dose-range finding test with the strains TA100 and the WP2uvrA, both with and without S9-mix. Eight concentrations, 1.7, 5.4, 17, 52, 164, 512, 1600 and 5000 µg/plate were tested in triplicate.

The highest concentration of the test item used in the subsequent mutation assays was 5000 μg/plate. At least five different doses (increasing with approximately half-log steps) of the test item were tested in triplicate in each strain in the absence and presence of S9-mix.

Vehicle / solvent:

The vehicle of the test item was Milli-Q water (Millipore Corp., Bedford, MA., USA).

Untreated negative controls:

yes

Remarks:

Milli-Q water

Positive controls:

yes

Positive control substance:

4-nitroquinoline-N-oxide

2-nitrofluorene

sodium azide

methylmethanesulfonate

other: ICR-191 (Sigma), 2-aminoanthracene (2AA)

Details on test system and experimental conditions:

Salmonella typhimurium bacteria and Escherichia coli bacteria

Rationale for test conditions:

Recommended test system in international guidelines (e.g. OECD, EC).

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity nor precipitates, but tested up to recommended limit concentrations

Untreated negative controls validity:

valid

Positive controls validity:

valid

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity nor precipitates, but tested up to recommended limit concentrations

Untreated negative controls validity:

valid

Positive controls validity:

valid

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity nor precipitates, but tested up to recommended limit concentrations

Untreated negative controls validity:

valid

Positive controls validity:

valid

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity nor precipitates, but tested up to recommended limit concentrations

Untreated negative controls validity:

valid

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 nor precipitates, but tested up to recommended limit concentrations

Untreated negative controls validity:

valid

Positive controls validity:

valid

Dose-Range Finding Test: Mutagenic Response of MEA Polyborate 1:1 in the Salmonella typhimurium Reverse Mutation Assay and in the Escherichia coli Reverse Mutation Assay

Direct plate assay

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

Without S9-mix

Positive control	722	±	32		1272	±	76
Solvent control	101	±	7		19	±	6
1.7	99	±	7		19	±	1
5.4	108	±	5		18	±	3
17	101	±	25		25	±	6
52	117	±	9		23	±	0
164	111	±	29		21	±	5
512	100	±	16		22	±	8
1600	110	±	6		20	±	5
5000	128	±	11	^n NP	19	±	6	^n NP

With S9-mix

Positive control	475	±	106		420	±	13
Solvent control	92	±	6		29	±	10
1.7	107	±	15		27	±	4
5.4	95	±	12		25	±	4
17	110	±	12		28	±	8
52	111	±	16		29	±	11
164	112	±	14		24	±	5
512	107	±	9		27	±	6
1600	101	±	11		27	±	2
5000	126	±	17	^n NP	38	±	4	^n NP

NP	No precipitate
n	Normal bacterial background lawn

Experiment 1: Mutagenic Response of MEA Polyborate 1:1 in the Salmonella typhimurium Reverse Mutation Assay

Direct plate assay

Dose (µg/plate)	Mean number of revertant colonies/3 replicate plates (±S.D.) with different strains of Salmonella typhimurium.
	TA1535	TA1537	TA98

 Without S9-mix

Positive control	790	±	47		985	±	99		1193	±	86
Solvent control	11	±	2		6	±	2		16	±	3
52	16	±	3		6	±	2		11	±	6
164	11	±	4		4	±	1		15	±	5
512	11	±	7		6	±	2		25	±	5
1600	11	±	4		6	±	2		16	±	9
5000	6	±	4	^n NP	4	±	1	^n NP	16	±	2	^n NP

 

With S9-mix

Positive control	328	±	21		546	±	104		1022	±	61
Milli-Q water	10	±	3		8	±	4		24	±	11
52	13	±	2		7	±	2		29	±	5
164	10	±	2		6	±	3		25	±	9
512	12	±	1		7	±	3		30	±	6
1600	13	±	3		5	±	0		24	±	3
5000	7	±	2	^n NP	8	±	4	^n NP	21	±	2	^n NP

 

NP	No precipitate
n	Normal bacterial background lawn

Experiment 2: Mutagenic Response of MEA Polyborate 1:1 in theSalmonella typhimuriumReverse Mutation Assay and in the Escherichia coli Reverse Mutation Assay

Pre-incubation assay

Dose (µg/plate)	Mean number of revertant colonies/3 replicate plates (±S.D.) with different strains ofSalmonella typhimuriumand oneEscherichia colistrain.
	TA1535	TA1537	TA98	TA100	WP2uvrA

Without S9-mix

Positive control	886	±	21		1183	±	75		1211	±	169		708	±	171		202	±	35
Solvent control	7	±	2		9	±	1		20	±	5		118	±	4		36	±	9
52	13	±	1		8	±	2		34	±	21		114	±	13		25	±	10
164	12	±	2		8	±	4		19	±	7		104	±	51		29	±	4
512	15	±	5		8	±	3		18	±	4		112	±	20		31	±	4	^i
1600	9	±	6		11	±	6		12	±	4		149	±	8		31	±	13
5000	9	±	3	^n SP	5	±	2	^n SP	15	±	5	^n SP	179	±	17	^n SP	38	±	17	^n SP

 

With S9-mix

Positive control	217	±	3		177	±	16		495	±	86		1008	±	343		541	±	13
Milli-Q water	19	±	5		4	±	0		26	±	2		118	±	21		42	±	9
52	8	±	1		8	±	2		28	±	13		116	±	10		41	±	6
164	11	±	4		6	±	5		28	±	2		135	±	27		61	±	7
512	10	±	2		8	±	4		25	±	7		145	±	27		54	±	9
1600	13	±	1		10	±	8		23	±	9		148	±	11		48	±	9
5000	7	±	2	^n SP	5	±	3	^n SP	21	±	4	^n SP	189	±	18	^n SP	52	±	5	^n SP

 

SP	Slight Precipitate
i	Plate infected
n	Normal bacterial background lawn

 

Conclusions:

Based on the results of a GLP OECD 471 study it is concluded that MEA Polyborate 1:1 is not mutagenic in the Salmonella typhimurium reverse mutation assay and in the Escherichia coli reverse mutation assay.

Reference 2

Endpoint:

in vitro cytogenicity / chromosome aberration study in mammalian cells

Remarks:

Type of genotoxicity: chromosome aberration

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2010-05-12 to 2010-07-28

Reliability:

1 (reliable without restriction)

Qualifier:

according to guideline

Guideline:

OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)

Deviations:

yes

Remarks:

Only 64 metaphase spreads examined

GLP compliance:

yes

Type of assay:

in vitro mammalian chromosome aberration test

Target gene:

No data

Species / strain / cell type:

lymphocytes:

Details on mammalian cell type (if applicable):

No data

Additional strain / cell type characteristics:

not specified

Metabolic activation:

with and without

Metabolic activation system:

Rat liver microsomal enzymes were routinely prepared from adult male Wister rats (6), which wre obtained from Charles Reiver (Sulzfeld, Germany)

Test concentrations with justification for top dose:

First cytogenic assay
Without S9-mix: 1000, 3330, 4000 and 5000 µg/ml culture medium ( 3h exposure time, 24 h fixation time)
With S9-mix : 1000, 3330 and 5000 µg/ml culture medium ( 3h exposure time, 24 h fixation time)
Second cytogenic assay
Without S9-mix: 50, 100, 200, 300, 500, 750 and 1000 µg/ml culture medium (24 h and 48 h exposure time, 24 h and 48 h fixation time).
With S9-mix: 1000, 3330 and 5000 µg/ml culture medium (3 h exposure time, 48 h fixation time).
Third cytogenic assay
Without and with S9-mix: 5000 µg/ml culture medium (3 h exposure time, 24 h fixation time).

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: [RPMI 1640 medium]
- Justification for choice of solvent/vehicle:none given

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

not specified

Positive controls:

yes

Positive control substance:

cyclophosphamide

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

not specified

Positive controls:

yes

Positive control substance:

mitomycin C

Details on test system and experimental conditions:

METHOD OF APPLICATION: in medium
DURATION
- Preincubation period:
- Exposure duration:3 hour
- Expression time (cells in growth medium):
- Selection time (if incubation with a selection agent):
- Fixation time (start of exposure up to fixation or harvest of cells): 24 hour

SELECTION AGENT (mutation assays):
SPINDLE INHIBITOR (cytogenetic assays):colchcine (0.5
STAIN (for cytogenetic assays):5% (v/v) Glemsa (Merck) solution in tap water

NUMBER OF REPLICATIONS:2

NUMBER OF CELLS EVALUATED:

DETERMINATION OF CYTOTOXICITY
- Method: mitotic index

OTHER EXAMINATIONS:
- Determination of polyploidy:carried out
- Determination of endoreplication:carried out
- Other:

OTHER:

Evaluation criteria:

A substance was considered positive (clastogenic) in the chromosome aberration test if:
a) It induced a dose-related statistically significant (Chi-square test, one sided , p<0.05) increase in the number of cells with chromosome aberrations.
b) A statistically significant and biologically relevant increase in the frequencies of the number of cells with chromosome aberrations was observed in the absence of a clear dose-response relationship.

Statistics:

The parameter used was the increase in the number of cells with chromosome aberrations. The statisitcal significance was given by Chi-square test, one sided , p<0.05.

Species / strain:

lymphocytes: human

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not determined

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH: Clastogenicity was observed in the first cytogentic assay. This was assumed to be due to the high pH in that assay.

Remarks on result:

other: strain/cell type: human lymphocytes

Remarks:

Migrated from field 'Test system'.

Conclusions:

Based on the results of this study, it is concluded that MEA Polyborate 1:1 has shown a clastogenic response in the chromosome aberration study in experiments where the pH of the treated cultures was above the pH of the solvent control. The clastogenicity was only observed in the first cytogenetic assay. Due to fact that the adjustment of the pH of the cultures treated with MEA Polyborate 1:1 did not show a clastogenic response in the third cytogenetic assay, it has to be assumed that the high pH is responsible for the detected clastogenic activity.

Executive summary:

Evaluation of the ability of MEA Polyborate 1:1 to induce chromosome aberrations in cultured peripheral human lymphocytes (with repeat experiments). This report describes the effect of MEA Polyborate 1:1 on the number of chromosome aberrations in cultured peripheral human lymphocytes in the presence and absence of a metabolic activation system (phenobarbital and ß-naphthoflavone induced rat liver S9-mix). The possible clastogenicity of MEA Polyborate 1:1 was tested in three independent experiments. The study procedures described in this report were based on the most recent OECD and EC guidelines. Batch EU-SMG 01695 of MEA Polyborate 1:1 was a clear colourless liquid. MEA Polyborate 1:1 was dissolved in RPMI 1640 medium. In the first cytogenetic assay, MEA Polyborate 1:1 was tested up to 5000 µg/ml for a 3 h exposure time with a 24 h fixation time in the absence and presence of 1.8% (v/v) S9-fraction. This is the highest concentration that should be tested according to the guidelines. In the second cytogenetic assay, MEA Polyborate 1:1 was tested up to 1000 µg/ml for a 24 h and 48 h continuous exposure time with a 24 h and 48 h fixation time in the absence of S9-mix. Appropriate toxicity was reached at this dose level. In the presence of S9-mix MEA Polyborate 1:1 was tested up to 5000 µg/ml for a 3 h exposure time with a 48 h fixation time. In the third cytogenetic assay, MEA Polyborate 1:1 was tested at the concentration of 5000 µg/ml for a 3 h exposure time with a 24 h fixation time in the absence and presence of 1.8% (v/v) S9-fraction. In this cytogenetic assay the pH of the cultures which were treated with MEA Polyborate 1:1 was adjusted to the pH of the solvent control. The number of cells with chromosome aberrations found in the solvent control cultures was within the laboratory historical control data range. Positive control chemicals, mitomycin C and cyclophosphamide, both produced a statistically significant increase in the incidence of cells with chromosome aberrations, indicating that the test conditions were adequate and that the metabolic activation system (S9-mix) functioned properly. No biologically relevant effects of MEA Polyborate 1:1 on the number of polyploid cells and cells with endoreduplicated chromosomes were observed both in the absence and presence of S9-mix. Therefore it can be concluded that MEA Polyborate 1:1 does not disturb mitotic processes and cell cycle progression and does not induce numerical chromosome aberrations under the experimental conditions described in this report. In the first and second cytogenetic assay MEA Polyborate 1:1 did not induce a statistically significant or biologically relevant increase in the number of cells with chromosome aberrations in the absence of S9-mix. However exchange figures were observed at all concentrations tested in the first cytogenetic assay. It should be noted that chromosomal exchanges are comparatively rare spontaneous events and therefore greater significance should be attached to the observation of exchange figures. In the first cytogenetic assay, in the presence of S9-mix, MEA Polyborate 1:1 induced a statistically significant, increase in the number of cells with chromosome aberrations at the highest tested concentration of 5000 µg/ml, both when gaps were included and excluded. In the second cytogenetic assay, in the presence of S9-mix, MEA Polyborate 1:1 did not induce a statistically significant or biologically relevant increase in the number of cells with chromosome aberrations. The statistically significant increase in the number of cells with chromosome aberrations and the exchange figures were observed in cultures treated with MEA Polyborate 1:1 which had a pH (8.42 - 9.14) above the negative control (pH = 8.01). It is known that changes in pH induce significant increases in chromosome aberrations. Therefore an additional third cytogenetic assay was performed in which the pH of the cultures treated with MEA Polyborate 1:1 was adjusted to the pH of the solvent control. In the third cytogenetic assay, MEA Polyborate 1:1 did not induce a statistically significant or biologically relevant increase in the number of cells with chromosome aberrations in the absence and presence of S9-mix. Moreover, no exchange figures were observed. Based on the results of this study, it is concluded that MEA Polyborate 1:1 has shown a clastogenic response in the chromosome aberration study in experiments where the pH of the treated cultures was above the pH of the solvent control. The clastogenicity was only observed in the first cytogenetic assay. Due to fact that the adjustment of the pH of the cultures treated with MEA Polyborate 1:1 did not show a clastogenic response in the third cytogenetic assay, it has to be assumed that the high pH is responsible for the detected clastogenic activity.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Genetic toxicity in vivo

Endpoint conclusion

Endpoint conclusion:

no study available

Justification for classification or non-classification

Based on the results of a GLP OECD 471 study it is concluded that MEA Polyborate 1:1 is not mutagenic in the Salmonella typhimurium reverse mutation assay and in the Escherichia coli reverse mutation assay.

Based on the results of this study, it is concluded that MEA Polyborate 1:1 has shown a clastogenic response in the chromosome aberration study in experiments where the pH of the treated cultures was above the pH of the solvent control. The clastogenicity was only observed in the first cytogenetic assay. Due to fact that the adjustment of the pH of the cultures treated with MEA Polyborate 1:1 did not show a clastogenic response in the third cytogenetic assay, it has to be assumed that the high pH is responsible for the detected clastogenic activity.

The substance is therefore not classified as mutagenic under GHS.