[4-[p,p'-bis(dimethylamino)benzhydrylidene]cyclohexa-2,5-dien-1-ylidene]dimethylammonium m-[[p-anilinophenyl]azo]benzenesulphonate

Dissociation constant

Administrative data

Endpoint:

dissociation constant

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2023

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2023

Report date:

2023

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 112 (Dissociation Constants in Water)

GLP compliance:

not specified

Test material

Test material information

Constituent 1

Reference substance name:

[4-[p,p'-bis(dimethylamino)benzhydrylidene]cyclohexa-2,5-dien-1-ylidene]dimethylammonium m-[[p-anilinophenyl]azo]benzenesulphonate

EC Number:

265-449-9

EC Name:

[4-[p,p'-bis(dimethylamino)benzhydrylidene]cyclohexa-2,5-dien-1-ylidene]dimethylammonium m-[[p-anilinophenyl]azo]benzenesulphonate

Cas Number:

65113-55-5

Molecular formula:

C25H30N3.C18H14N3O3S

IUPAC Name:

[4-[p,p'-bis(dimethylamino)benzhydrylidene]cyclohexa-2,5-dien-1-ylidene]dimethylammonium m-[[p-anilinophenyl]azo]benzenesulphonate

Test material form:

solid: particulate/powder

Reference

Constituent 1

Reference substance name:

[4-[p,p'-bis(dimethylamino)benzhydrylidene]cyclohexa-2,5-dien-1-ylidene]dimethylammonium m-[[p-anilinophenyl]azo]benzenesulphonate

EC Number:

265-449-9

EC Name:

[4-[p,p'-bis(dimethylamino)benzhydrylidene]cyclohexa-2,5-dien-1-ylidene]dimethylammonium m-[[p-anilinophenyl]azo]benzenesulphonate

Cas Number:

65113-55-5

Molecular formula:

C25H30N3.C18H14N3O3S

IUPAC Name:

[4-[p,p'-bis(dimethylamino)benzhydrylidene]cyclohexa-2,5-dien-1-ylidene]dimethylammonium m-[[p-anilinophenyl]azo]benzenesulphonate

Test material form:

solid: particulate/powder

Results and discussion

Dissociating properties:

not determined

Remarks:

No results could be calculated, as the molar extinction coefficients do not follow a clear trend in dependence of the pH. This might be originated by the existence of the two ionic compounds present in the test item with respective dissociation constants

Dissociation constant

Remarks on result:

not determinable

Remarks:

No results could be calculated, as the molar extinction coefficients do not follow a clear trend in dependence of the pH. This might be originated by the existence of the two ionic compounds present in the test item with respective dissociation constants influencing each other.

Any other information on results incl. tables

Titration method: Due to the very low solubility of the test item and the limitations regarding the volumes of the titrants used, the titration method was dismissed beforehand. The water solubility was determined to be in the range of 20 – 660 µg/L according to the OECD 105. The validity criteria of this guideline could not be met, but the used method was the best approach available and possible.

The photometer method was used for the determination of the dissociation constant, i.e. the dissociation of the test item must have an impact on the UV‑Vis absorption. Therefore, a significant difference of the absorption for at least a single wavelength must be present.

If this is the case, the dissociation constant can be calculated with known concentrations and for various pH values. These values should be within acceptable ranges to be valid.

Test solutions of nominal 10 mg/L were prepared by dilution of a test item stock solution of nominal 100 g/L in dichloromethane with various puffer solutions in the range of 1 – 14.
The absorption spectra of each test solution was measured (against separate blank solutions with buffer and dichloromethane but without test item). After the addition of the spiking solution to the respective buffer, a part of the test item did precipitate. The buffers were filtered via syringe filter (pore-size of 0.2 µm) before the photometric measurement. Additionally, the color of the test solutions did change depending on the pH value.

pH Value	Color
1	Deep pink
2	Bluish violet
3 ‑ 7	Violet
9	Weak violet
12 ‑ 14	Yellow

 

For the definitive determination, 570 nm was chosen as the test wavelength, where the spectra show a significant difference in the absorption. 1 L of puffer solution at pH 12 was spiked with 0.1 mL test item stock solution (100 g/L in dichloromethane), resulting in a solution of 10 mg/L. The exact pH value was measured and an aliquot was taken and the absorption at 570 nm was determined.
Afterwards, hydrochloric acid was added to the whole sample until a nominal pH value of 11 was achieved with documentation of the exact pH value and the volume of used hydrochloric acid to determine the exact concentrations. Another aliquot was taken to determine the absorption at 570 nm.
This step was repeated for several pH values until a pH value of 1.

pH		c [mol/L]	E	ε	cHA/cA-	pKs
Nominal	Measured
12	12.053	0.00001409	0.221	15689	Assumed to be fully undissociated
11	10.783	0.00001400	0.479	34205	-0.7962	n.a.
10	n.d.	0.00001400	n.a.	n.a.	-1.2405	n.a.
9	9.021	0.00001400	0.625	44646	-0.8697	n.a.
8	7.737	0.00001400	0.654	46725	-0.8784	n.a.
7	6.990	0.00001399	0.661	47237	-0.8804	n.a.
6	6.081	0.00001398	0.663	47410	-0.8811	n.a.
5	4.977	0.00001398	0.667	47719	-0.8822	n.a.
4	4.081	0.00001395	0.643	46092	-0.8759	n.a.
3	3.036	0.00001392	0.558	40077	-0.8453	n.a.
2	2.078	0.00001372	0.402	29299	-0.7228	n.a.
1	1.408	0.00001269	0.247	19462	Assumed to be fully dissociated

n.d.         Erroneously skipped during preparation; value not determined

n.a.         Not applicable

No results could be calculated, as the molar extinction coefficients do not follow a clear trend in dependence of the pH. This might be originated by the existence of the two ionic compounds present in the test item with respective dissociation constants influencing each other.

Therefore, the photometer method was deemed to be not feasible for the determination of the dissociation constant of the test item.

 

 

Applicant's summary and conclusion

Conclusions:

No results could be calculated, as the molar extinction coefficients do not follow a clear trend in dependence of the pH. This might be originated by the existence of the two ionic compounds present in the test item with respective dissociation constants influencing each other.

Therefore, the photometer method was deemed to be not feasible for the determination of the dissociation constant of the test item.

Executive summary:

The determination of the dissociation constant of the test item (batch: 924249) in compliance to the OECD-Guideline 112 was determined to be not feasible.

Possibly due to the low water solubility (20 – 660 µg/L), it is neither possible to clearly determine a single dissociation constant using the titration method, nor to calculate results using the photometer method.

For the titration method, the test solution should have a concentration of at least 0.01 M, corresponding to approx. 7 g/L, or at half the saturation concentration. Both of these are possibilities for the determination according to OECD 112, as the water solubility is expected to be very low, the performance of the study is not feasible.

For the photometer method, no results could be calculated, as the molar extinction coefficients do not follow a clear trend in dependence of the pH.

The third possibility given in the guideline, the conductrometric method, was not tested, as the necessary test item concentration should be in the range of 0.1 M, i.e. approx. 72 g/L, which exceeds the expected water solubility at several magnitudes.