Octasodium 2,2'-[1,4-phenylenebis[imino(6-chloro-1,3,5-triazine-4,2-diyl)imino(1-hydroxy-3,6-disulphonatonaphthalene-2,8-diyl)azo]]bisnaphthalene-1,5-disulphonate

Hydrolysis

Link to relevant study record(s)

Reference

Endpoint:

hydrolysis

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 111 (Hydrolysis as a Function of pH)

Qualifier:

according to guideline

Guideline:

EU Method C.7 (Degradation: Abiotic Degradation: Hydrolysis as a Function of pH)

GLP compliance:

yes (incl. QA statement)

Radiolabelling:

no

Analytical monitoring:

yes

Details on sampling:

Samples were taken and analyzed by HPLC immediately after preparation of the test solutions.

Buffers:

Buffer pH 4: Citric acid/NaOH/NaCl; Fluka, order no.: 33643
Buffer pH 7: KH2PO4/Na2HPO4, Fluka, order no.: 33646
Buffer pH 9: Na2B4O7/HCl, Fluka, order no.: 33648
All buffer solutions were filled in glass bottles and purged with N2 for five minutes. Afterwards the buffer solutions were sterilized at 120 °C for 20 minutes

BUFFER FOR IDENTIFICATION OF HYDROLYSIS PRODUCTS
Buffer pH 4: Citric acid/NaOH/NaCl; Fluka, order no.: 33643

Details on test conditions:

CALIBRATION:
The test item was used as standard compound for the calibration. For evaluation of the hydrolysis products the response factor of the main component was used.

PREPARATION OF A TEST ITEM STOCK SOLUTION:
A stock solution of the test item was prepared by weighing 67.4 mg of the test item in a 50 mL volumetric flask and filling it to the mark with demineralized water, which results in a final concentration of 1044.7 mg/L.

PREPARATION OF THE CALIBRATION SOLUTIONS:
The stock solutionwas diluted with appropriate volumes to obtain calibration solutions between 52.235 and 2089.4 mg/L in demineralized water.

PRAPARATION OF THE SOLUTION USED FOR VERIFICATION OF THE SYSTEM SUITABILITY:
Approx. 35 mg of the test item were weighed into a 50 mL volumetric flask and filled up to the mark with demineralized to obtain control stock solutions of approx. 543 mg/L. This control solution was used for verification of the system suitability before analysing the test solutions.

PREPARATION OF THE HYDROLYSIS TEST SOLUTIONS:
For each pH separate hydrolysis test solutions were prepared in the corresponding buffer systems. Test item concentrations of approx. 1000 mg/L were obtained. Aliquots of the stock solutions were taken without further treatment to prepare individual vials for every test point.

PREPARATION AND PROCEDURE OF THE TEST:
All buffer solutions were filled in glass bottles and purged with N2 for five minutes. Afterwards the buffer solutions were sterilized at 120 °C for 20 minutes. Before the beginning of the tests the solutions are tempered and the temperature was checked. All glassware was purged with N2 before and after filling in the test item and the buffer solutions. The test solutions were overlain with N2.
To prove the sterility of the glass instruments and the test design, sterility tests of each buffer solution were performed at the end of the incubation.

SOLUBILITY AND TEST CONCENTRATION:
According to OECD TG 111 the concentration of the test item should not exceed 0.01 M or half of the saturation concentration of the water solubility. The water solubility of the test item was estimated in the high g/L range. Consequently, the test item was applied as aqueous buffer solution with a concentration of approx. 1000 mg/L (0.0005 mol/L) which fulfilled the requirements of OECD 111. Clear solutions were obtained.

Duration:

5 d

pH:

4

Temp.:

50 °C

Initial conc. measured:

942.61 mg/L

Remarks:

replicate 1

Duration:

5 d

pH:

4

Temp.:

50 °C

Initial conc. measured:

998.21 mg/L

Remarks:

replicate 2

Duration:

5 d

pH:

7

Temp.:

50 °C

Initial conc. measured:

753.37 mg/L

Duration:

5 d

pH:

9

Temp.:

50 °C

Initial conc. measured:

994.56 mg/L

Number of replicates:

2 replicantes at pH 4 (50 °C)

Positive controls:

no

Negative controls:

no

Preliminary study:

A non-GLP pretest showed degradation of the test item at pH 4 at 50 °C.

Transformation products:

yes

No.:

#1

No.:

#2

% Recovery:

99.3

St. dev.:

1.3

Temp.:

50 °C

Duration:

26 d

pH:

4

Temp.:

50 °C

Hydrolysis rate constant:

0 s-1

DT50:

131.7 h

Type:

(pseudo-)first order (= half-life)

Remarks on result:

other: correlation coefficient: r=0.99940

Remarks:

replicate 1

pH:

4

Temp.:

50 °C

Hydrolysis rate constant:

0 s-1

DT50:

130.1 h

Type:

(pseudo-)first order (= half-life)

Remarks on result:

other: correlation coefficient: r=0.99947

Remarks:

replicate 2

pH:

7

Temp.:

50 °C

Remarks on result:

other: overall degradation after 5 days: >10 %, test item is considered as stable

pH:

9

Temp.:

50 °C

Remarks on result:

other: overall degradation after 5 days: >10 %, test item is considered as stable

Details on results:

REPEATABILITY:
Results of the solutions for verification of the peak area repeatability show stability of the chromatographic system. A relative standard deviation of approx. 1.4 % indicates a satisfying repeatability and precision of the method applied to quantify the test item concentrations over the test duration. The requirements of OECD 111 are met.

SENSITIVITY:
Regarding the chromatogram of the lowest concentration, taken from calibration measurement of repeatability tests, the analytical method is sufficiently sensitive to quantify test item concentrations down to 10 % or less of the initial concentration used in the hydrolysis experiment.

Hydrolysis at pH 4 (50 °C):

In a non-GLP pretest degradation of the main component at pH 4 was observed. Therefore the 50 °C test was already carried out as Tier 2 test. Within 21 days the test item degraded to less than 10 % of the initial concentration. Additionally two major hydrolysis products formed which were evaluated using the response factor of the main component. The replicate test shows that the hydrolysis behaviour of the test item is repeatable.

According to OECD 111 Guideline a sterility test was conducted at the end the hydrolysis tests. No microbes (colonies) were found. Therefore biotic degradation can be excluded.

The hydrolysis products were identified by HPLC-HRMS.

Validity criteria fulfilled:

yes

Remarks:

Reactive Red 141 has a suitable water solubility, analytical methods are available, but the purity of Reactive Red 141 is not > 95% as the substance exhibits a high water content.

Conclusions:

Reactive Red 141 is found to be stable at 50 °C at pH 7 and pH 9 but showed abiotic degradation > 10 % within 5 days hydrolysis time at a pH value of 4.

Executive summary:

The tests were performed according to OECD Guidelines for Testing of Chemicals, Section 1 – Physical-Chemical Properties, OECD TG 111, Council Regulation (EC) No 440/2008, Guideline Part C – Methods for the Determination of Ecotoxicity, C.7. “Abiotic Degradation: Hydrolysis as a Function of pH”.

A non-GLP pretest showed degradation of the test item at pH 4 at 50 °C. Therefore the hydrolysis behaviour of the test item was investigated at 50 °C at pH values of 7 and 9 over a period of five days according to OECD TG 111, Tier 1. At pH 4 Tier 2 investigations were carried out directly. The stability was monitored by HPLC analysis using UV-detection.

Analytical result of Tier 1:

No abiotic degradation of the test item was observed at pH 7 and 9. At pH 4 a degradation of the test item could be observed. Reactive Red 141 is found to be stable at 50 °C at pH 7 and pH 9. Therefore it can be assumed that the test item is also stable at 25 °C and no half-life times and hydrolysis rates were calculated for these pH values. According to OECD TG 111 at pH 4 Tier 2 tests were performed.

Analytical result of Tier 2:

The test item showed abiotic degradation > 10 % within 5 days hydrolysis time at a pH value of 4.

The degradation of the investigated components of the test item can be described by first order kinetics. Half-life times (t1/2=132 hours) and hydrolysis rates (k=1.47*10^-6) were calculated. Two main hydrolysis products were identified by HPLC-HRMS in a separate determination. As requested by the guideline, one hydrolysis experiment should be carried out twice, which was done at 50 °C.

The hydrolysis behaviour of Reactive Red 141 at pH 4 wasn't further investigated at other temperatures than 50 °C as the substance is assumed to be hydrolytically stable at environmental relevant conditions.

Description of key information

Reactive Red 141 is found to be stable at 50 °C at pH 7 and pH 9 but showed abiotic degradation > 10 % within 5 days hydrolysis time at a pH value of 4.

It can be assumed that the Reactive Red 141 is also stable at 25 °C (pH 7 and 9) and no half-life times and hydrolysis rates were calculated for these pH values. For hydrolysis at pH 4 (50 °C) a half-life time of t1/2=132 hours and hydrolysis rate k=1.47*10^-6 were calculated.

The hydrolysis behaviour of Reactive Red 141 at pH 4 wasn't further investigated at other temperatures than 50 °C as the substance can be assumed to be hydrolytically stable at environmental relevant conditions.

Additional information

The tests were performed according to OECD Guidelines for Testing of Chemicals, Section 1 – Physical-Chemical Properties, OECD TG 111, Council Regulation (EC) No 440/2008, Guideline Part C – Methods for the Determination of Ecotoxicity, C.7. “Abiotic Degradation: Hydrolysis as a Function of pH”.

A non-GLP pretest showed degradation of the test item at pH 4 at 50 °C. Therefore the hydrolysis behaviour of the test item was investigated at 50 °C at pH values of 7 and 9 over a period of five days according to OECD TG 111, Tier 1. At pH 4 Tier 2 investigations were carried out directly. The stability was monitored by HPLC analysis using UV-detection.

Analytical result of Tier 1:

No abiotic degradation of the test item was observed at pH 7 and 9. At pH 4 a degradation of the test item could be observed. Reactive Red 141 is found to be stable at 50 °C at pH 7 and pH 9. Therefore it can be assumed that the test item is also stable at 25 °C and no half-life times and hydrolysis rates were calculated for these pH values. According to OECD TG 111 at pH 4 Tier 2 tests were performed.

Analytical result of Tier 2:

The test item showed abiotic degradation > 10 % within 5 days hydrolysis time at a pH value of 4.

The degradation of the investigated components of the test item can be described by first order kinetics. Half-life times (t1/2=132 hours) and hydrolysis rates (k=1.47*10^-6) were calculated. Two main hydrolysis products were identified by HPLC-HRMS in a separate determination. As requested by the guideline, one hydrolysis experiment should be carried out twice, which was done at 50 °C.

The hydrolysis behaviour of Reactive Red 141 at pH 4 wasn't further investigated at other temperatures than 50 °C as the substance can be assumed to be hydrolytically stable at environmental relevant conditions.