N-(5-chloro-2-methoxyphenyl)-2-[(2-methoxy-4-nitrophenyl)azo]-3-oxobutyramide

Adsorption / desorption

Link to relevant study record(s)

Reference

Endpoint:

adsorption / desorption: screening

Type of information:

experimental study

Adequacy of study:

key study

Study period:

03 March 2022 - 04 March 2022

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 121 (Estimation of the Adsorption Coefficient (Koc) on Soil and on Sewage Sludge using High Performance Liquid Chromatography (HPLC))

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of method:

HPLC estimation method

Media:

soil/sewage sludge

Radiolabelling:

no

Test temperature:

40ºC

Details on study design: HPLC method:

EQUIPMENT
- Apparatus: Chromatographic column
- Type: HPLC
- Type, material and dimension of analytical (guard) column: Lichrospher ® 100, CN (5 μm), 4 x 250 mm
- Detection system: DAD (Diode Array Detector)

MOBILE PHASES
- Type: methanol/water mixture: methanol/water (55/45% v/v)
- Experiments with additives carried out on separate columns: no

DETERMINATION OF DEAD TIME
- Method: by inert substance which is not retained by the column (thiourea)

REFERENCE SUBSTANCES
- Identity:
For soil: Acetanilide, 2-nitro benzamide, aniline, naphthalene, 1,2,3 - trichloro benzene and phenanthrene.
For sewage sludge: acetanilide, benzamide, aniline, 2,5-dichloro aniline, benzoic acid phenyl ester and phenanthrene.

DETERMINATION OF RETENTION TIMES
- Quantity of test substance introduced in the column: ca. 2260 mg/L
- Quantity of reference substances: ca. 100 mg/L
- Intervals of calibration: The calibration injections were carried out before and after injections of the test substance.

REPETITIONS
- Number of determinations: 2

EVALUATION
- Calculation of capacity factors k': k′ = (tR – to)/ to
- Calculation of retention times: HPLC
- Determination of the log Koc value: Koc was deduced from the capacity factor (k′) using a calibration plot of log k′ versus log Koc of the selected reference substances: Log Koc = a + (b × log k’). The log k’ value of a test substance was
used to determine its log Koc value.

Details on test conditions:

TEST CONDITIONS
The temperature during the measurements were made at 40°C. Temperature from the controlled column compartment was used.

Key result

Sample No.:

#1

Type:

log Koc

Value:

1.79 dimensionless

Temp.:

40 °C

Matrix:

Soil

Remarks on result:

other: Mean adsorption coefficient (log Koc) of two replicates.

Key result

Sample No.:

#2

Type:

log Koc

Value:

2.49 dimensionless

Temp.:

40 °C

Matrix:

Sewage sludge

Remarks on result:

other: Mean adsorption coefficient (log Koc) of two replicates.

Details on results (HPLC method):

- Retention times of reference substances used for calibration: see tables below.
- Details of fitted regression line (log k' vs. log Koc): see tables below
- Graph of regression line attached: see graphs below
- Average retention data for test substance: see tables below

Transformation products:

no

Table 1. Dead time determination for sewage sludge

Name of the Reference Substance	Replicate Injections	t0                             Dead time (min)	Mean t0                             Dead time (min)
Thiourea	R1	2.807	2.810
	R2	2.813

Table 2. Dead time determination for soil

Name of the Reference Substances	Replication	t0                             Dead time (min)	Mean t0                             Dead time (min)
Thiourea	R1	2.807	2.810
	R2	2.813

Table 3. Determination of capacity factor of the selected reference substances for sewage sludge

Name of the Reference Substances	Replicate Injections	tR                              Retention time (min)	Mean  tR                              Retention time (min)	Capacity Factor k'	log Koc for Sewage Sludge	log k'
Acetanilide	R1	3.327	3.330	0.19	1.52	-0.73
	R2	3.333
Benzamide	R1	3.120	3.120	0.11	1.60	-0.96
	R2	3.120
Aniline	R1	3.387	3.387	0.21	1.74	-0.69
	R2	3.387
2,5-Dichloroaniline	R1	3.113	3.113	0.11	2.45	-0.97
	R2	3.113
Benzoic acid phenyl ester	R1	4.773	4.773	0.70	3.26	-0.16
	R2	4.773
Phenanthrene	R1	6.633	6.623	1.36	4.35	0.13
	R2	6.613
Slope (b)					2.2041
Intercept (a)					3.7240
Correlation Coefficient (r)
Coefficient of Determination (R^2)					0.7750

Table 4. Determination of capacity factor of the selected reference substances for soil

Name of the Reference Substances	Replicate Injections	tR                              Retention time (min)	Mean  tR                              Retention time (min)	Capacity Factor k'	log Koc for Soil	log k'
Acetanilide	R1	3.327	3.330	0.19	1.25	-0.73
	R2	3.333
2-Nitrobenzamide	R1	4.127	4.130	0.47	1.45	-0.33
	R2	4.133
Aniline	R1	3.387	3.390	0.21	2.07	-0.69
	R2	3.393
Naphthalene	R1	4.827	4.820	0.72	2.75	-0.15
	R2	4.813
1,2,3-Trichlorobenzene	R1	4.753	4.750	0.69	3.16	-0.16
	R2	4.747
Phenanthrene	R1	6.620	6.620	1.36	4.09	0.13
	R2	6.620
Slope (b)					2.7463
Intercept (a)					3.3406
Correlation Coefficient (r)
Coefficient of Determination (R^2)					0.7251

Table 5. Determination of capacity factor and adsorption coefficient of test item for sewage sludge

Details	Replicate Injections	tR                              Retention time (min)	Mean  tR                              Retention time (min)	Capacity Factor k'	log k'	log Koc for Sewage Sludge	Mean log Koc for Sewage Sludge	Standard Deviation (±)
Test Item R1	Rep 1	3.593	3.590	0.28	-0.56	2.50	2.49	0.0118
	Rep 2	3.587
Test Item R2	Rep 1	3.580	3.577	0.27	-0.56	2.48
	Rep 2	3.573

Table 6. Determination of capacity factor and adsorption coefficient of test item for soil

Details	Replicate Injections	tR                              Retention time (min)	Mean  tR                              Retention time (min)	Capacity Factor k'	log k'	log Koc  for Soil	Mean                  log Koc for Soil	Standard Deviation (±)
Test Item R1	Rep 1	3.573	3.577	0.27	-0.56	1.79	1.79	0.0039
	Rep 2	3.580
Test Item R2	Rep 1	3.573	3.573	0.27	-0.57	1.79
	Rep 2	3.573

 

Validity criteria fulfilled:

yes

Remarks:

(The determined values of log Koc derived from individual measurements of replicate samples (in duplicate) of the test item were within a range of 0.25 log unit)

Conclusions:

The Adsorption coefficient (log Koc) on soil and sewage sludge of the test item
were estimated by HPLC analysis. Log Koc of the test item was estimated to be 2.49 on sewage sludge and 1.79 on soil.

Executive summary:

The adsorption coefficient (Koc) on soil and on sewage sludge of the test item was determined by using a high-performance liquid chromatography system in accordance with OECD Guideline 121, under GLP conditions. The selected reference substances for sewage sludge were acetanilide, benzamide, aniline, 2,5-dichloro aniline, benzoic acid phenyl ester and phenanthrene. The selected reference substances for soil were acetanilide, 2-nitro benzamide, aniline, naphthalene, 1,2,3 - trichloro benzene and phenanthrene. The dead time of the instrument was determined by injecting the inert substance thiourea which is not retained by the column and the capacity factors for each reference substance were calculated based on dead time retention time. The retention times of the test substance and selected reference substances were accurately determined using HPLC fitted with a Lichrospher CN column at 40ºC. The regression curve for the reference substances were constructed by plotting the capacity factor (log k') against their known adsorption coefficients (log Koc). The adsorption coefficient of the test item was estimated using this regression curve for both soil and sewage sludge. Solutions of test item (2 replicates) were run in duplicate. The mean adsorption coefficient (log Koc) of test item of two replicates was estimated to be 2.49 for sewage sludge and 1.79 for soil. The determined values of log Koc derived from the individual measurements of the test item were within a range of 0.25 log unit, thus the quality criterium for repeatibility of the Guideline was fulfilled.

Description of key information

Key study: Adsortion using the HPLC method according to OECD 121, GLP study. The Adsorption coefficient (log Koc) on soil and sewage sludge of the test item were estimated by HPLC analysis. Log Koc of the test item was estimated to be 2.49 on sewage sludge and 1.79 on soil. From these results, the Koc are calculated to be 309 L/kg for sewage sludge and 61.7 L/kg for soil. As a coservative approach, Koc = 309 L/kg is taken as the representative value for further assessment.

Key value for chemical safety assessment

Koc at 20 °C:

309