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Environmental fate & pathways

Hydrolysis

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Reference
Endpoint:
hydrolysis
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Study period:
13 July 2012 - 10 Jan. 2013
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
Study was performed by using OECD 111 in accordance with GLP. However, as the substance is highly hydrolytically unstable it was not possible to perform the test as outlined in guidelines. In order to obtain as much information on hydrolysis as possible, an alternative test was designed.
Justification for type of information:
The substance is hydrolytically unstable. When it comes in contact with water or moisture, a complete hydrolysis will take place with no significant reaction products other than the particular alcohol and hydrated titanium dioxide. This rapid hydrolysis (hydrolysis half-life < 3 minutes to < 2 hours) is the driving force for the fate and pathways of the substance. The aquatic toxicity testing is considered scientifically unjustified as the substance degrades immediately releasing the particular alcohol and hydrated insoluble titanium dioxides in water.
The testing conducted with analogue substance of the category justifies that the aquatic toxicity in daphnia and algae studies is similar to the aquatic toxicity of the alcohol released to test water as the insoluble hydrated titanium oxide, precipitated on the bottom of the test vessels; lacking bioavailability. The identification of the degradation products from the hydrolysis study conducted for the target substance verifies that there are no impurities in the alcohol released from the target substance which might change the aquatic toxicity of the target substance compared to the toxicity of the pure alcohol.
As there is a mechanistic reasoning to the read-across, the read-across from the degradation product (relevant alcohol) is used to evaluate the aquatic toxicity and the fate and pathways of the target substance in the environment.
Reason / purpose for cross-reference:
read-across source
Qualifier:
according to guideline
Guideline:
OECD Guideline 111 (Hydrolysis as a Function of pH)
Deviations:
not applicable
Remarks:
(alternative method was needed as the test substance is highly water reactive)
Qualifier:
according to guideline
Guideline:
EU Method C.7 (Degradation: Abiotic Degradation: Hydrolysis as a Function of pH)
Deviations:
not applicable
Qualifier:
according to guideline
Guideline:
EPA OPPTS 835.2120 (Hydrolysis of Parent and Degradates as a Function of pH at 25°C)
Deviations:
not applicable
Principles of method if other than guideline:
The rate of hydrolysis of the test substance as a function of pH was determined at pH values normally found in the environment (pH 4-9). The sponsor supplied information that the test substance is highly hydrolytically unstable and thefefore the quantitative analysis of the test substance is not possible.

As a result, it is not possible to perform the test as outlined in the guidelines. In order to obtain as much information on hydrolysis as possible, an alternative test was designed. The test was conducted at 25°C. An amount of test substance (above the water solubility) was added to the buffer solution. The amount of hydrolysis products formed was determined as function of time. The test was performed in duplicate. At each sampling time, samples were taken from each test and analysed in duplicate.
GLP compliance:
yes
Radiolabelling:
no
Analytical monitoring:
yes
Details on sampling:
- Sampling intervals for the transformation products: The concentration of the test substance in the test sample was determined at time points between 0 & 40 minutes based on quantification of the hydrolysis product.
- Sampling method: the test was performed at 25 deg. C. An amount of the test substance (above the water solubility) was added to the buffer solution. The amount of hydrolysis products formed was determined as function of time. The test was performed in duplicate. At each sampling time, samples were taken from each test and analyzed in duplicate. The hydrolysis of the test substance was determined by the quantitative analysis of the hydrolysis product 2-ethylhexanol.
- Sampling intervals/times for pH measurements: pH of each of the test solution was determined at each sampling time t=0, t=5, t=10, t=20, t=40 min.
- Sample storage conditions before analysis: Stored in well-sealed container at room temperature in the dark under nitrogen.
- Other observation, if any (e.g.: precipitation, color change etc.): White precipitate was observed during the hydrolysis. This is due to the formation of the insoluble titanium dioxides.
Buffers:
- pH: 4, 7 & 9.
- Type and final molarity of buffer: Acetate buffer pH 4, 0.01 M, Phosphate buffer pH 7, 0.01 M, Borate buffer pH 9, 0.01 M
- Composition of buffer:
Acetate buffer pH 4, 0.01 M: Solution of 16.7% 0.01 M sodium acetate in water and 83.3% 0.01 M acetic acid in water. The buffer contains 0.0009% (w/v) sodium azide.

Phosphate buffer pH 7, 0.01 M: Solution of 0.01 M potassium di-hydrogenphosphate in water adjusted to pH 7 using 1 N sodium hydroxide. The buffer contains 0.0009% (w/v) sodium azide.

Borate buffer pH 9, 0.01 M: Solution of 0.01 M boric acid in water and 0.01 M potassium chloride in water adjusted to pH 9 using 1 N sodium hydroxide. The buffer contains 0.0009% (w/v) sodium azide.
Details on test conditions:
TEST SYSTEM
- Type, material and volume of test flasks, other equipment used: Volumetric flask
- Sterilisation method: filter-sterilised through a 0.2 µm FP 30/0.2 CA-S filter (Whatman, Dassel, Germany)
- Measures to exclude oxygen: To exclude oxygen, nitrogen gas was purged through the solution for 5 minutes.
- Details on test procedure for unstable compounds:
The rate of hydrolysis of the test substance as a function of pH was determined at pH values normally found in the environment (pH 4-9).The sponsor supplied information that the test substance is not soluble in water, highly hydrolytically unstable and that quantitative analysis of the test substance is not possible.
 As a result, it is not possible to perform the test as outlined in the guidelines. In order to obtain as much information on hydrolysis as possible, an alternative test was designed. The test was performed at 25°C. An amount of test substance (above the water solubility) was added to the buffer solution. The amount of hydrolysis products formed was determined as function of time. The test was performed in duplicate. At each sampling time, samples were taken from each test and analysed in duplicate. The hydrolysis of the test substance was determined by the quantitative analysis of the hydrolysis product 2- ethylhexanol.

Details of the main study:
The buffer solutions were filter-sterilised through a 0.2 µm FP 30/0.2 CA-S filter (Whatman, Dassel, Germany) and transferred into a volumetric flask. To exclude oxygen, nitrogen gas was purged through the solution for 5 minutes. From this solution, 243 ml was transferred into a vessel. After equilibration at the test temperature, a 3 ml sample was taken as blank and time zero sample. Thereafter, pure test substance was spiked to the buffer solution at a target concentration of about 40 mg/l. Each vessel was sealed and placed in a thermostatically controlled oven at 25.0°C  0.5°C. At selected time points, the vessel was shaken and 2.5 ml samples were collected. For the quantitative analysis, two aliquots of 1 ml were taken from the 2.5 ml sample and transferred into a 10 ml GC headspace vial. If necessary, 500 µl of the samples was diluted with 500 µl of the buffer solution.

The pH of each of the test solutions was determined at the beginning and at the end of the test.

The study was performed at the following conditions:

pH code Temperature

pH 4 25.0°C ± 0.2°C

pH 7 25.1°C ± 0.1°C

pH 9 25.0°C ± 0.2°C

TEST MEDIUM
- Volume used/treatment: 2.5 ml
- Kind and purity of water: Tap water purified by a Milli-Q water purification system (Millipore, Bedford, MA, USA)
- Preparation of test medium:
For preparing of calibration solution, the corresponding buffer solution is used. If the test was performed at pH 4, acetate buffer pH 4, 0.01 M was used. If the test was performed at pH 7, phosphate buffer pH 7, 0.01 M was used. And if the test was performed at pH 9, borate buffer pH 9, 0.01 M was used.

Stock solutions
Stock solutions of 2-Ethylhexanol were prepared in water at concentrations of 5019 mg/l.

Calibration solutions
Calibration solutions were prepared in the concentration range 0.7 – 30 mg/l. The calibration solutions were prepared by dilution of two stock solutions with buffer solution. 1 ml of the calibration solution was transferred into a 10 ml GC-headspace vial.

Calibration curves
were constructed using five concentrations. For each concentration, two responses were used. For the calibration curve prepared in phosphate buffer pH 7, two responses were excluded from the curve since the back calculated accuracy deviated > 15% from the nominal concentration. Quadratic regression analysis was performed using the least squares method with a 1/concentration2 weighting factor. The coefficient of correlation (r) was > 0.99 for each curve.
Number of replicates:
Two
Positive controls:
no
Negative controls:
yes
Remarks:
Blank buffer solutions
Statistical methods:
Calibration curves were constructed using five concentrations. For each concentration, two responses were used. If necessary, two responses were excluded from the curve since the back calculated accuracy deviated > 15 % from the nominal concentration. Quadratic regression analysis was performed using the least squares method with a 1/concentration^2 weighting factor. The coefficient of correlation (r) was > 0.99 for each curve.
Preliminary study:
The test substance is known to be hydrolytically unstable. Therefore, the preliminary test was not performed.
Transformation products:
yes
No.:
#1
No.:
#2
Details on hydrolysis and appearance of transformation product(s):
The predicted hydrolysis product, 2-ethylhexanol, was detected by GC-MS. Additionally, the formation of a white precipitation was observed during the hydrolysis study. This was due to the formation of the insoluble titanium oxides.

The response of the hydrolysis product 2-ethylhexanol was > 99% (area). In the chromatograms, small additional peaks were observed. Retention times in test at pH 4 and pH 7 were 3.14 minutes and 10.11 minutes. Retention times in test at pH 9 were 3.16 minutes and 10.21 minutes. All ≤ 1% at the specific mass fragments monitored. The additional peaks might derive from impurities in the test substance and/or hydrolysis products.

To achieve sufficient sensitivity for the analysis of the identified hydrolysis product, selected ion monitoring was used as detection method. Therefore it was not possible to obtain accurate information on the mass fraction of the unknown peaks nor to record their entire mass spectrum, and hence it was not possible to identify these additional peaks.
pH:
4
Temp.:
25 °C
DT50:
<= 10 min
Remarks on result:
other: The response of the hydrolysis product was > 99% (area). In the chromatograms, small additional peaks were observed, all ≤1% at the specific mass fragments monitored. The additional peaks might derive from impurities in the test substance.
pH:
7
Temp.:
25 °C
DT50:
<= 10 min
Remarks on result:
other: The response of the hydrolysis product was > 99% (area). In the chromatograms, small additional peaks were observed, all ≤1% at the specific mass fragments monitored. The additional peaks might derive from impurities in the test substance.
pH:
9
Temp.:
25 °C
DT50:
<= 5 min
Remarks on result:
other: The response of the hydrolysis product was > 99% (area). In the chromatograms, small additional peaks were observed, all ≤1% at the specific mass fragments monitored. The additional peaks might derive from impurities in the test substance.
Details on results:
The predicted hydrolysis product, 2-Ethylhexanol, was detected by GC-MS. Additionally, the formation of a white precipitation was observed during the hydrolysis study. This is most likely due to the formation of the insoluble titanium oxides.
Based on the results, it was safely concluded that the hydrolysis half-life time is ≤ 10 minutes at pH 4 & 7 and ≤ 5 minutes at pH 9.

The response of the hydrolysis product 2-Ethyl hexanol was > 99% (area). In the chromatograms, small additional peaks were observed. Retention times in test at pH 4 and pH 7 were 3.14 minutes and 10.11 minutes. Retention times in test at pH 9 were 3.16 minutes and 10.21 minutes. All ≤ 1% at the specific mass fragments monitored. The additional peaks might derive from impurities in the test substance and/or hydrolysis products.

To achieve sufficient sensitivity for the analysis of the identified hydrolysis product, selected ion monitoring was used as detection method. Therefore it was not possible to obtain accurate information on the mass fraction of the unknown peaks nor to record their entire mass spectrum, and hence it was not possible to identify these additional peaks.

Please refer attachment "interpretation" to see the formula used for interpretation of results.

The analytical results of the main study are given in Table 1, 2, 3, 4, 5 and Table 6.

Table 1: Main test 1 – hydrolysis of the test substance at pH 4 and 25°C

Sampling time


[minutes]

Analysed concentration

2-Ethyl hexanol

[mg/l]

Actual pH

0

1

4.0

0

1

 

5

6.60

 

5

6.88

 

10

14.4

 

10

14.7

 

20

12.0

 

20

12.0

 

40

13.4

4.0

40

13.1

 

Table 2: Main test 2 – hydrolysis of the test substance at pH 4 and 25°C

Sampling time


[minutes]

Analysed concentration

2-Ethyl hexanol

[mg/l]

Actual pH

0

1

4.0

0

1

 

5

7.74

 

5

5.75

 

10

10.1

 

10

9.72

 

20

11.5

 

20

11.6

 

40

12.1

4.1

40

12.5

 

Table 3: Main test 1 – hydrolysis of the test substance at pH 7 and 25°C

Sampling time


[minutes]

Analysed concentration

2-Ethyl hexanol

[mg/l]

Actual pH

0

1

7.0

0

1

 

5

15.1

 

5

15.7

 

10

17.5

 

10

20.0

 

20

21.6

 

20

21.8

 

40

18.7

7.0

40

23.8

 

Table 4: Main test 2 – hydrolysis of the test substance at pH 7 and 25°C

Sampling time


[minutes]

Analysed concentration

2-Ethyl hexanol

[mg/l]

Actual pH

0

1

7.0

0

1

 

5

16.6

 

5

17.8

 

10

24.9

 

10

25.8

 

20

28.0

 

20

25.3

 

40

34.9

7.0

40

31.2

 

1              A response was observed on the retention time of the test substance, this response was also observed in the analytical blanc. Maximum contribution to the other samples (main test 1 and main test 2 combined) based on area was 0.07%.

Table 5: Main test 1 – hydrolysis of the test substance at pH 9 and 25°C

Sampling time


[minutes]

Analysed concentration

2-Ethyl hexanol

[mg/l]

Actual pH

0

1

8.9

0

1

 

5

17.0

 

5

17.0

 

10

17.5

 

10

17.3

 

20

18.8

 

20

19.3

 

40

20.3

8.9

40

20.1

 

 Table 6: Main test 2 – hydrolysis of the test substance at pH 9 and 25°C

Sampling time


[minutes]

Analysed concentration

2-Ethyl hexanol

[mg/l]

Actual pH

0

1

8.9

0

1

 

5

10.2

 

5

10.2

 

10

13.0

 

10

12.4

 

20

13.9

 

20

14.3

 

40

14.8

8.9

40

14.5

 

1 A response was observed on the retention time of the test substance, this response was also observed in the analytical blanc. Maximum contribution to the other samples (main test 1 and main test 2 combined) based on area was 0.1%.

 

 

Validity criteria fulfilled:
yes
Conclusions:
Half-life time of Titanium tetrakis(2-ethylhexanolate) at 25°C and at pH values normally found in the environment (pH 4-9) were determined in this study.

Since the test substance is highly hydrolytically unstable, the quantitative analysis of the test substance is not possible. The half-life time was determined based on the analysis of the hydrolysis product 2-ethylhexanol. Based on the results, it was safely concluded that the hydrolysis half-life time at 25°C at pH 4, 7 is less than 10 minutes and at pH 9 less than 5 minutes, respectively.
Executive summary:

Study performed in accordance with GLP. However, as the substance is highly hydrolytically unstable It is not possible to perform the test as outlined in the guidelines. In order to obtain as much information on hydrolysis as possible, an alternative test was designed.

The hydrolytic stability was tested in buffered aqueous solutions at pH 4.0, 7.0 and 9.0 at 25 0C. The progress of hydrolysis was followed by monitoring 2 -ethylhexanol, a major product of the hydrolysis of the substance. Based on the results, it was concluded that the hydrolysis half -life time for the substance was <10 minutes at pH 4 and 7, and < 5 minutes at pH 9. The response of the hydrolysis product 2 -ethylhexanol was > 99% (area). In the chromatograms, small additional peaks were observed at 3.14 minutes, 10.11 minutes, all ≤ 1% at the specific mass fragments monitored. The additional peaks might derive from impurities in the test substance and/or hydrolysis products. To achieve sufficient sensitivity for the analysis of the identified hydrolysis product, selected ion monitoring was used as detection method. Therefore it was not possible to obtain accurate information on the mass fraction of the unknown peaks nor to record their entire mass spectrum, and hence it was not possible to identify these additional peaks.

The results of this study are considered reliable to be used for C&L purposes to conclude this substance to be hydrolytically unstable releasing only 2 -ethylhexanol and hydrated titanium dioxide when in contact with water. Based on the results this substance can be considered as rapidly degradable.

Description of key information

Hydrolytically unstable with the half-life time of < 10 minutes at 25 deg. C at pH 4, 7 and < 5 minutes at pH 9.

Key value for chemical safety assessment

Half-life for hydrolysis:
10 min
at the temperature of:
25 °C

Additional information

Study was performed by using OECD 111 in accordance with GLP for the structurally similar titanate. However, as the substance is hydrolytically unstable it was not possible to perform the test as outlined in the guidelines. In order to obtain as much information on hydrolysis as possible, an alternative test was designed. The hydrolytic stability was tested in buffered aqueous solutions at pH 4.0, 7.0, and 9.0 at 25°C for 40 minutes. The progress of the hydrolysis was followed by monitoring the main degradation product; 2-ethylhexanol. Additionally, the formation of a white precipitation was observed during the hydrolysis study. This is due to the formation of the insoluble titanium oxides.

No other degradation products than 2 -ethylhexanol and titanium oxide were identified. According to Tier 3 approach, the response of the hydrolysis product 2-ethylhexanol was > 99% (area). In the chromatograms, small additional peaks were observed. Retention times in test at pH 4 and pH 7 were 3.14 minutes and 10.11 minutes. Retention times in test at pH 9 were 3.16 minutes and 10.21 minutes. All ≤ 1% at the specific mass fragments monitored. The additional peaks might derive from impurities in the test substance and/or hydrolysis products. To achieve sufficient sensitivity for the analysis of the identified hydrolysis product, selected ion monitoring was used as detection method. Therefore it was not possible to obtain accurate information on the mass fraction of the unknown peaks or to record their entire mass spectrum, and hence it was not possible to identify these additional peaks.

The results of this study are considered reliable to be used for C&L purposes to conclude this substance to be hydrolytically unstable releasing only 2 -ethylhexanol and hydrated titanium dioxide when in contact with water. Based on the results this substance can be considered as rapidly degradable.