Rhamnolipids: fermentation products of glucose with Pseudomonas putida, Pseudomonadaceae

Adsorption / desorption

Link to relevant study record(s)

Reference

Endpoint:

adsorption / desorption, other

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2019-07-02 to 2019-09-10

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 106 (Adsorption - Desorption Using a Batch Equilibrium Method)

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: Council Regulation (EC) No. 440/2008, Method C.18 (2008)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of method:

batch equilibrium method

Media:

soil

Radiolabelling:

no

Test temperature:

Nominal: 20 - 25 °C

Analytical monitoring:

yes

Details on sampling:

Test Procedure

The study was performed in accordance to OECD-Guideline for Testing of Chemicals No. 106 (2000) and Council Regulation (EC) No. 440/2008, C.18 (2008).

Test vessels
50 mL polypropylene centrifugation tubes
120 mL glass vessel/ 50mL glass vessels

Concentration for adsorption experiments
Tier 1 and tier 2: 1.0 mg/L
Tier 3: 200 µg/L, 630 µg/L, 1 mg/L, 6.3 mg/L, 20 mg/L

Stock solutions
Stock solutions of 1 g/L of the test item in ultrapure water were prepared and further diluted with ultrapure water to 200 mg/L, 630 mg/L and 1000 mg/L test item.

Preparation of the soil samples (conditioning)
The soils were weighed into the test vessels and an appropriate volume of 0.01 M CaCl2-solution was added.
After agitation overnight (12 h minimum), the samples were used for adsorption experiments.

Preparation of the samples for adsorption experiments
The soil samples were conditioned as described above.
Max 2 volume-% of the stock solutions, related to the volume of the aqueous phase in the soil suspensions were added in order to adjust the test concentrations. Afterwards, the samples were agitated.

Samples for analysis
The soil suspensions were centrifuged after agitation at 4000 rpm to separate the phases, followed by analysing the concentration of the test item in aqueous phase by LC-MS. For analysis of the soil, the aqueous phase was decanted and the soil was extracted. Extracts were also analysed by LC-MS. For details of sample preparation, see below.

Replicates
All samples were prepared in duplicate.

CONTROLS
CaCl2-solution was conditioned as described above, followed by separation of the aqueous phase by centrifugation. Then the aqueous phase was fortified acc. to the concentrations used for the test item samples to verify the stability of the test item in the aqueous phase under test conditions. The samples were agitated as long as the test item sample with the longest agitation period.

Replicates
Duplicates (tier 1, tier 2 and tier 3)

BLANK
Blank samples were prepared for all soils as described for the test item samples but without fortification with the test item. The samples were agitated as long as the samples with the longest agitation period.

Replicates
Duplicates (tier 1), single (tier 2)


Sample Preparation

Dilution medium
Ultrapure water / methanol (50/50)

Standards
A stock solution of 1 g test item/L in ultrapure water was prepared. The solution was diluted to 7 calibration standards in the range of 5 to 100 µg test item/L with dilution medium.

Aqueous phase
4 mL each of the aqueous phase was transferred into a centrifuge vial. After centrifugation with 4000 rpm for 5 min, an aliquot of each aqueous sample was stabilized by dilution with methanol (factor 2) and analysed. Blank samples were filtered before analyses.

Soil extraction
For separation of aqueous phase and soil, the suspension had to be transferred into centrifugation tubes after mixing carefully. Residual soil was transferred by rinsing with 0.01 M CaCl2 solution (rinsing volume equivalent to test volume).
The soil was extracted at room temperature. 30 mL methanol : ultrapure water (50 : 50 v/v) containing 1% formic acid were added to the wet soil. The vessel was shaken for 30 min. The suspensions were centrifuged at 4000 rpm for 5 min. The extraction was repeated twice. The extracts were transferred quantitatively into a 100 mL measuring flask and filled up with methanol : water (50 : 50 v/v) containing 1% formic acid. Samples were measured directly.

Matrix no.:

#1

Matrix type:

other: silty sand

% Clay:

4.1

% Silt:

10.5

% Sand:

85.4

% Org. carbon:

0.718

pH:

5.4

CEC:

2.4 other: mval/100 g

Matrix no.:

#2

Matrix type:

loamy sand

% Clay:

8.5

% Silt:

11.3

% Sand:

80.2

% Org. carbon:

1.47

pH:

6.8

CEC:

7.6 other: mval/100 g

Matrix no.:

#3

Matrix type:

other: silty sand

% Clay:

8.6

% Silt:

29.3

% Sand:

62.1

% Org. carbon:

0.412

pH:

7

CEC:

4.9 other: mval/100 g

Matrix no.:

#4

Matrix type:

clay loam

% Clay:

23.3

% Silt:

38.6

% Sand:

38.1

% Org. carbon:

1.85

pH:

7.5

CEC:

22 other: mval/100 g

Matrix no.:

#5

Matrix type:

loamy sand

% Clay:

10.2

% Silt:

31.1

% Sand:

58.7

% Org. carbon:

0.916

pH:

7.6

CEC:

10 other: mval/100 g

Details on matrix:

Reason for the selection
These matrices are suitable for the conduction of the study because all parameters with impact on the adsorption / desorption behavior of a chemical substance were considered.
Tier 1 was conducted with LUFA soils 2.2 and 2.4.

Origin of soils
Landwirtschaftliche Untersuchungs- und Forschungsanstalt LUFA Speyer, Obere Langgasse 40, 67346 Speyer, Germany

Storage at test facility
Room temperature, in closed containers

Details on test conditions:

CaCl2-solution:
Ultrapure water was used to prepare the CaCl2-solution (0.01 M).

Soil / Solution ratio
Tier 1: 1:5, 1:20 and 1:100
Tier 2 1:2 for the soils LUFA 2.1, 2.2, 2.3, 2.4 and 5M
Tier 3: 1:2 for the soils LUFA 2.1, 2.2, 2.4 and 5M
1:1 for the soil LUFA 2.3

Sampling points
Tier 2: 0.25, 0.5h, 1h and 2h
Tier 3: 0.5h for LUFA soils 2.3 and 5M
1h for LUFA soils 2.1, 2.2 and 2.4

Agitation
By horizontal and overhead shaker. Frequency was adjusted to avoid sedimentation of soil particles during treatment.


Test temperature
The temperature was mainly in the range of 20 to 25 °C during the course of the study. Deviation from this temperature range occurred only for a brief period (< 1h) with a temperature between 18 and 20 °C during tier 2 and tier 3.
Due to elevated outside temperatures regulation of the air conditioning was occasionally necessary for a couple of minutes. This caused that the temperature dropped to a value of 18.0 °C.
This deviation is considered to have no negative impact on the integrity and quality of the study.

Key result

Sample No.:

#1

Type:

Kd

Value:

2.4 L/kg

pH:

5.4

Temp.:

20 °C

Matrix:

LUFA 2.1 soil/solution ratio 1:2

% Org. carbon:

0.718

Remarks on result:

other: Di-Rhamnose-C8-C10

Key result

Sample No.:

#1

Type:

Koc

Value:

335 L/kg

pH:

5.4

Temp.:

20 °C

Matrix:

LUFA 2.1 soil/solution ratio 1:2

% Org. carbon:

0.718

Remarks on result:

other: Di-Rhamnose-C8-C10

Key result

Sample No.:

#2

Type:

Kd

Value:

1.9 L/kg

pH:

6.8

Temp.:

20 °C

Matrix:

LUFA 2.2 soil/solution ratio 1:2

% Org. carbon:

1.47

Remarks on result:

other: Di-Rhamnose-C8-C10

Key result

Sample No.:

#2

Type:

Koc

Value:

132 L/kg

pH:

6.8

Temp.:

20 °C

Matrix:

LUFA 2.2 soil/solution ratio 1:2

% Org. carbon:

1.47

Remarks on result:

other: Di-Rhamnose-C8-C10

Key result

Sample No.:

#3

Type:

Kd

Value:

0.87 L/kg

pH:

7

Temp.:

20 °C

Matrix:

LUFA 2.3 soil/solution ratio 1:2

% Org. carbon:

0.412

Remarks on result:

other: Di-Rhamnose-C8-C10

Key result

Sample No.:

#3

Type:

Koc

Value:

212 L/kg

pH:

7

Temp.:

20 °C

Matrix:

LUFA 2.3 soil/solution ratio 1:2

% Org. carbon:

0.412

Remarks on result:

other: Di-Rhamnose-C8-C10

Key result

Sample No.:

#4

Type:

Kd

Value:

1.9 L/kg

pH:

7.5

Temp.:

20 °C

Matrix:

LUFA 2.4 soil/solution ratio 1:2

% Org. carbon:

1.85

Remarks on result:

other: Di-Rhamnose-C8-C10

Key result

Sample No.:

#4

Type:

log Koc

Value:

101 L/kg

pH:

7.5

Temp.:

20 °C

Matrix:

LUFA 2.4 soil/solution ratio 1:2

% Org. carbon:

1.85

Remarks on result:

other: Di-Rhamnose-C8-C10

Key result

Sample No.:

#5

Type:

Kd

Value:

1.3 L/kg

pH:

7.6

Temp.:

20 °C

Matrix:

LUFA 5M soil/solution ratio 1:2

% Org. carbon:

0.916

Remarks on result:

other: Di-Rhamnose-C8-C10

Key result

Sample No.:

#5

Type:

Koc

Value:

147 L/kg

pH:

7.6

Temp.:

20 °C

Matrix:

LUFA 5M soil/solution ratio 1:2

% Org. carbon:

0.916

Remarks on result:

other: Di-Rhamnose-C8-C10

Key result

Sample No.:

#1

Type:

Kd

Value:

13 L/kg

pH:

5.4

Temp.:

20 °C

Matrix:

LUFA 2.1 soil/solution ratio 1:2

% Org. carbon:

0.718

Remarks on result:

other: Di-Rhamnose-C10-C10

Key result

Sample No.:

#1

Type:

Koc

Value:

1 830 L/kg

pH:

5.4

Temp.:

20 °C

Matrix:

LUFA 2.1 soil/solution ratio 1:2

% Org. carbon:

0.718

Remarks on result:

other: Di-Rhamnose-C10-C10

Key result

Sample No.:

#2

Type:

Kd

Value:

14 L/kg

pH:

6.8

Temp.:

20 °C

Matrix:

LUFA 2.2 soil/solution ratio 1:2

% Org. carbon:

1.47

Remarks on result:

other: Di-Rhamnose-C10-C10

Key result

Sample No.:

#2

Type:

Koc

Value:

961 L/kg

pH:

6.8

Temp.:

20 °C

Matrix:

LUFA 2.2 soil/solution ratio 1:2

% Org. carbon:

1.47

Remarks on result:

other: Di-Rhamnose-C10-C10

Key result

Sample No.:

#3

Type:

Kd

Value:

4.6 L/kg

pH:

7

Temp.:

20 °C

Matrix:

LUFA 2.3 soil/solution ratio 1:2

% Org. carbon:

0.412

Remarks on result:

other: Di-Rhamnose-C10-C10

Key result

Sample No.:

#3

Type:

Koc

Value:

1 127 L/kg

pH:

7

Temp.:

20 °C

Matrix:

LUFA 2.3 soil/solution ratio 1:2

% Org. carbon:

0.412

Remarks on result:

other: Di-Rhamnose-C10-C10

Key result

Sample No.:

#4

Type:

Kd

Value:

19 L/kg

pH:

7.5

Temp.:

20 °C

Matrix:

LUFA 2.4 soil/solution ratio 1:2

% Org. carbon:

1.85

Remarks on result:

other: Di-Rhamnose-C10-C10

Key result

Sample No.:

#4

Type:

Koc

Value:

1 012 L/kg

pH:

7.5

Temp.:

20 °C

Matrix:

LUFA 2.4 soil/solution ratio 1:2

% Org. carbon:

1.85

Remarks on result:

other: Di-Rhamnose-C10-C10

Key result

Sample No.:

#5

Type:

Kd

Value:

8.8 L/kg

pH:

7.6

Temp.:

20 °C

Matrix:

LUFA 5M soil/solution ratio 1:2

% Org. carbon:

0.916

Remarks on result:

other: Di-Rhamnose-C10-C10

Key result

Sample No.:

#5

Type:

Koc

Value:

963 L/kg

pH:

7.6

Temp.:

20 °C

Matrix:

LUFA 5M soil/solution ratio 1:2

% Org. carbon:

0.916

Remarks on result:

other: Di-Rhamnose-C10-C10

Results

 

Tier 1–Experiments

LUFA 2.2 and LUFA 2.4 soils were used for preliminary investigations on the adsorption behaviour of the test item. Only one component, 2Rha-C10-C10, was analysed during tier 1, whereas 2Rha-C8-C10 was additionally analysed during tier 2 and tier 3.

 

LUFA soil 2.2 was tested with soil / solution ratios of 1:5 and 1:20 at a concentration of 1 mg test item/L.The control samples for LUFA 2.2 were below 80% already after 4h, therefore glass adsorption and/or degradation was assumed(see table below). As the indirect method was used and only the aqueous phase was analysed, depletion of the test item in the aqueous phase was visible over time, not showing an adsorption equilibrium.

 

Tier 1:Test Item Control Samples

Soil	Soil : Solution Ratio	Sampling point [h]	Recovery[%]
LUFA 2.2	1:5	0	92
LUFA 2.2	1:5	4	76
LUFA 2.2	1:5	24	75
LUFA 2.2	1:5	48	76
LUFA 2.2	1:20	0	93
LUFA 2.2	1:20	4	66
LUFA 2.2	1:20	24	69
LUFA 2.2	1:20	48	68

 

Test vessel adsorption was investigated in LUFA 2.2 test item control samples with a soil/solution ratio of 1:100 at a nominal concentration of 1 mg/L. While adsorption to container walls of polypropylene centrifugation tubes was observed, no adsorption was observed if glass vessels were used (see table below).

Tier 1: Test Vessel Adsorption

LUFA 2.2 test item control samples 1:100, nominal concentration: 1 mg/L, aqueous phase analysed

Test Vessel Type	Recovery Rate Aqueous Phase [%]	Adsorption to Test Vessel [%]
Polypropylene centrifugation tube	69	31
Glass bottle	108	-

 

Test item stability was subsequent tested by adsorption test samples in glass bottles as test vessel with and without HgCl₂ to verify biological degradation. After 48 h adsorption the aqueous phase was analysed as well as soil extracts. As the mass balance of the replicates with ‘sterilisation’ by addition of mercury was 88% and 89% and the mass balance of samples without 1%, biological degradation was verified absolutely certain. Detailed analytical results are shown below.

 

 Tier 1: Test Item Stability

LUFA 2.2 1:100, nominal concentration: 1 mg/L, aqueous phase and soil analysed

Sampling Point [h]	Replicate No.	Recovery Aqueous Phase [%]	Recovery Soil [%]	Mass Balance [%]
48	1 – without HgCl2	0	1	1
48	2 – without HgCl2	0	1	1
48	1 – with HgCl2	61	27	88
48	2 – with HgCl2	64	25	89

 

To complete the preliminary investigations of tier 1, adsorption experiments with LUFA 2.2 and LUFA 2.4 were conducted using a soil/ solution ratio of 1:5. Glass bottles were used as test vessels to prevent test vessel adsorption and both, aqueous phase and soil extracts were analysed at different sampling points over a period of 24 h to monitor the mass balance over time. Detailed results are shown in the tables below. As the mass balance decreased over time, degradation presumably takes place in parallel to adsorption. A differentiation is not possible by measurement of the aqueous phase.

 

Tier 1: Adsorption and Mass Balance – LUFA 2.2

Soil / solution ratio: 1:5, nominal concentration, test item: 1 mg/L, mean values of two replicates are given

Sampling Point [h]	Recovery Aqueous Phase [%]	Adsorption [%]^1	Recovery Soil [%]	Mass Balance [%]
0.5	21	79	75	97
2	18	82	68	87
5	13	87	55	67
24	0	100	7	7

1    =   calculated from concentrations in aqueous phase (indirect), presumably adsorption and degradation

 

 Tier 1: Adsorption and Mass Balance – LUFA 2.4

Soil / solution ratio: 1:5, nominal concentration, test item: 1 mg/L, mean values of two replicates are given

Sampling Point [h]	Recovery Aqueous Phase [%]	Adsorption [%]^1	Recovery Soil [%]	Mass Balance [%]
0.5	18	82	63	81
2	15	85	56	71
5	8	92	37	45
24	0	100	0	0

1    =   calculated from concentrations in aqueous phase (indirect), presumably adsorption and degradation

 

To prove test item stability and adsorption in the other soils used in the study, aqueous phases and soil extracts have also been analysed in samples with LUFA soils 2.1, 2.3 and 5M after adsorption for 2 h. Detailed analytical results are given in the table below.

 

Tier 1: Mass Balance – LUFA 2.1, LUFA 2.3, LUFA 5M

                        Soil / solution ratio: 1:5, nominal concentration, test item: 1 mg/L, sampling point: 2 h, mean values of two replicates are given

Soil Type	Recovery Aqueous Phase [%]	Adsorption [%]^1	Recovery Soil [%]	Mass Balance [%]
LUFA 2.1	16	84	65	81
LUFA 2.3	45	55	42	88
LUFA 5M	26	74	53	79

1   =   calculated from concentrations in aqueous phase (indirect)

 

Tier 1– Conclusions

After preliminary investigations of tier 1, glass test vessels were used for all adsorption experiments in aqueous media in tier 2 and tier 3 to prevent test vessel adsorption. As the test item is degraded in the test system containing aqueous media and soil, short agitation times were used to determine the adsorption. A soil / solution ratio of 1:1 and 1:2, respectively, was used in order to gain a rapid adsorption equilibrium, as adsorption is faster if a high soil / solution ratio is used. The direct method, analysing aqueous phase and soil extracts, were used throughout tier 2 and tier 3.

As the test item is not stable in the test system, no desorption experiments were conducted.

 

Tier 2 – Adsorption Kinetics

 

The determination for adsorption kinetics was performed with a nominal test item concentration of 1 mg test item/L. A soil / solution ratio of 1:2 was used for the LUFA soils 2.1, 2.2, 2.3, 2.4 and 5M. The concentrations of the test item were measured in aqueous phase and soil extracts at defined sampling points. The sampling points were 0.25, 0.5h, 1h and 2h. From the results the sampling points for tier 3 were derived with regard to adsorption equilibrium and mass balance. K_d and K_OC values were calculated according to the equilibration time presented in the tables below.

 

 

Tier 2 - Adsorption Kinetics (2Rha-C8-C10)

Equilibrium Time, Measured Amounts in Aqueous Phase and Soil Extracts, K_d and K_OC values for the a.i. 2Rha-C8-C10

Applied nominal test item concentration: 1 mg/L,
corresponding to a.i. (2Rha-C8-C10): 0.070 mg/L; n = 2

Soil Type	tEQ[h]	ma.i.[µg]	Vaq [mL]	msoil [g]	m^adsaq [µg]	m^adssoil [µg]	%OC	Kd [ml/g]	KOC [ml/g]	Mass balance^1) [%]
LUFA 2.1	1	1.39	20	9.69	0.594	0.692	0.718	2.4	335	91
LUFA 2.2	1	1.41	20	9.28	0.503	0.453	1.47	1.94	132	61
LUFA 2.3	1	1.39	20	9.61	0.797	0.335	0.412	0.87	212	89
LUFA 2.4	1	1.39	20	8.87	0.457	0.380	1.85	1.9	101	53
LUFA 5M	1	1.39	20	9.24	0.751	0.467	0.916	1.3	147	80

t_EQ                   =                            sampling point based onFigure1toFigure 5

m_a.i.                  =                            applied amount (a.i. 2Rha-C8-C10)

V aq                 =                            used volume of aqueous phase

m_soil                 =                            used amount of soil (dry weight)         

m^ads_aq               =                            amount of a.i. in the aqueous phase

m^ads_soil              =                            amount of a.i. in the soil

%OC                =                            percentage of organic carbon content in the soil

 1)                   =                            mean value mass balance at 1 h

 

Tier 2 - Adsorption Kinetics (2Rha-C10-C10)

Equilibrium Time, Measured Amounts in Aqueous Phase and Soil Extracts, K_d and K_OC values for the a.i. 2Rha-C10-C10

Applied nominal test item concentration: 1 mg/L,
corresponding to a.i. (2Rha-C10-C10): 0.250 mg/L; n = 2

Soil Type	t [h]	ma.i. [µg]	Vaq [mL]	msoil [g]	m^adsaq [µg]	m^adssoil [µg]	%OC	Kd [ml/g]	KOC [ml/g]	Mass balance^1) [%]
LUFA 2.1	1	5.02	20	9.69	0.611	3.89	0.718	13.1	1830	92
LUFA 2.2	1	5.09	20	9.28	0.542	3.55	1.47	14.1	961	80
LUFA 2.3	1	4.99	20	9.61	1.37	3.05	0.412	4.6	1127	95
LUFA 2.4	1	4.99	20	8.87	0.482	4.00	1.85	18.7	1012	88
LUFA 5M	1	5.01	20	9.24	0.993	4.05	0.916	8.8	963	90

t_EQ                   =                            sampling point based onFigure 6toFigure 10

m_a.i.                  =                            applied amount, a.i.

V_aq                   =                            used volume of aqueous phase

m_soil                 =                            used amount of soil (dry weight)

m^ads_aq               =                            amount of a.i. in the aqueous phase

m^ads_soil              =                            amount of a.i. in the soil

%OC                =                            percentage of organic carbon content in the soil

 1)                   =                            mean value mass balance at 1 h

 

 

 

Tier 3 – Adsorption Isotherms

The adsorption isotherm was determined with the concentrations of 0.2, 0.63, 1.0, 6.3 and 20 mg test item/L after an agitation time of 1h hour for LUFA soil 2.1, 2.2 and 2.4 and 0.5 h for the other soils. The first table below for the a.i. 2Rha-C8-C10 and the second table below for the a.i. 2Rha-C10-C10 show the Freundlich adsorption coefficient K_F^ads and the organic carbon normalized Freundlich adsorption coefficient K^OC_F. Endpoints of the adsorption isotherms were determined by analysing soil extracts of allapplication concentrations. Linear Freundlich adsorption was found for all soils except for LUFA 2.4.

 

Freundlich AdsorptionIsotherms 2Rha-C8-C10

Applied test item concentrations K₁ to K₅: 0.2, 0.63, 1.0, 6.3 and 20 mg/L
corresponding to 0.278 – 0.877 – 1.39 – 8.77 – 27.8 µg a.i. (LUFA 2.1, 2.2, 2.4 and 5M) and

0.139 – 0.438 – 0.696 – 4.38 – 13.9 µg a.i. (LUFA 2.3), respectively

Soil Type	msoil[g]	Vaq[mL]	r^2	1/n	KF^ads	KF^OC	Massbalance^1) [%]
LUFA 2.1	9.69	20	0.995	0.77	0.0062	0.87	84 to 94
LUFA 2.2	9.28	20	0.991	0.88	0.003	0.17	50 to 83
LUFA 2.3	9.61	10	0.995	0.77	0.0012	0.30	76 to 87
LUFA 2.4	9.28	20	0.991	1.11	0.0008	0.05	15 to 76
LUFA 5M	9.24	20	0.989	0.884	0.0016	0.17	64 to 96

m_soil    = used amount of soil (dry weight) [g]

V_aq      = volume aqueous phase [mL]

n         = regression constant

K_F^ads    = Freundlich adsorption coefficient [µg^1-1/n(mL)^1/ng^-1]

K_F^OC     = Freundlich adsorption coefficient normalized to content of organic carbon [µg^1-1/n(mL)^1/ng^-1]

1)        = range of mass balances K1 to K5

 

 

Freundlich Adsorption Isotherms 2Rha-C10-C10

Applied test item concentrations K₁ to K₅: 0.2, 0.63, 1.0, 6.3 and 20 mg/L
corresponding to 1.0 – 3.16 – 5.01 – 31.6 – 100 µg a.i. (LUFA 2.1, 2.2, 2.4 and 5M) and

0.501 – 1.58 – 2.50 – 15.8 – 50.1 µg a.i. (LUFA 2.3), respectively

Soil Type	msoil[g]	Vaq[mL]	r^2	1/n	KF^ads	KF^OC	Mass balance^1)[%]
LUFA 2.1	9.69	20	0.998	0.81	0.037	5.22	108 to 112
LUFA 2.2	9.28	20	0.997	0.83	0.029	2.00	92 to 100
LUFA 2.3	9.61	10	0.998	0.81	0.0080	1.95	82 to 91
LUFA 2.4	9.28	20	0.988	1.02	0.013	0.87	21 to 92
LUFA 5M	9.24	20	0.994	0.891	0.012	1.33	82 to 102

m_soil    = used amount of soil (dry weight) [g]

V_aq      = volume aqueous phase [mL]

n         = regression constant

K_F^ads    = Freundlich adsorption coefficient [µg^1-1/n(mL)^1/ng^-1]

K_F^OC     = Freundlich adsorption coefficient normalized to content of organic carbon [µg^1-1/n(mL)^1/ng^-1]

1)        = range of mass balances K1 to K5

Validity criteria fulfilled:

yes

Executive summary:

Summary

General Information

The adsorption / desorption behavior of the test item HH-2014-547 (batch no. ET18503646) was investigated in five different soils according to OECD guideline 106 and Council Regulation (EC) No. 440/2008, C.18 from 2019-07-02 to 2019-09-10 at Noack Laboratorien GmbH, 31157 Sarstedt, Germany. Distribution coefficients K_d and organic carbon normalized distribution coefficients K_OC were determined with a single concentration. Furthermore, investigations about the adsorption as a function of the test item loading level (Freundlich adsorption isotherms) in the aqueous phase were performed. Two main components of the test item, Di-Rhamnose-C8-C10 (2Rha-C8C10) and Di-Rhamnose-C10-C10 (2Rha-C10-C10) were analysed by LC-MS/MS. Relevant properties of the used soils are given in Table 1. Experiments for adsorption kinetics were conducted with a nominal test item concentration of 1 mg test item/L. Concentrations of 20, 6.30, 1.0, 0.63, 0.2 mg test item/L were used for the determination of the Freundlich adsorption isotherms. No desorption experiments were performed due to the instability of the test item in the test system.

Table1:      RelevantCharacteristics of Test Matrices

	Soils
	LUFA 2.1	LUFA 2.2	LUFA 2.3	LUFA 2.4	LUFA 5M
batch	F2.1 2817	F2.2 4016	F2.3 4116	F2.4 2617	F5M 5015
Soil Type^1)	Silty sand	Loamy sand	Silty sand	Clayey loam	Loamy sand
pH (0.01 M CaCl2)^3)	4.9	5.4	5.9	7.4	7.3
Organic Carbon [%]^2)	0.718	1.47	0.412	1.85	0.916
Clay (<0.002 mm) [%]^2)	4.1	8.5	8.6	23.3	10.2
Silt (0.002-0.063 mm) [%]^2)	10.5	11.3	29.3	38.6	31.1
Sand (0.063-2 mm) [%]^2)	85.4	80.2	62.1	38.1	58.7
Cation Exchange Capacity [mval/100g]^2)	2.4	7.6	4.9	22	10

1) according to German DIN

2) determined at Agrolab Agrar und Umwelt GmbH (non-GLP)

3) Analyses data sheet provided by LUFA

Results Tier 1

In tier 1, preliminary investigations were conducted to find an appropriate soil / solution ratio for adsorption experiments. As the adsorption, if determined by the indirect method, could not be differentiated from degradation of the test item from the aqueous phase, test item control samples have additionally been prepared after ‘sterilization’ by addition of HgCl₂. To determine the adsorption and to calculate distribution coefficients, short agitation times were used, in order to reduce the test item degradation to a minimum. A high soil / solution ratio was found to be appropriate for experiments in tier 2 and tier 3, because adsorption kinetics proceed faster at higher soil / solution ratios.

Tier 1 experiments also revealed adsorption to polypropylene test tubes. Therefore, glass vessels were used throughout the study.

Results Tier 2 and Tier 3

Mass balances were calculated during tier 2 to investigate an agitation time short enough to minimize test item degradation but long enough to show an adsorption equilibrium. Five test item concentrations were tested in adsorption experiments over a concentration range of two orders of magnitude.

Table2:   Summarized Endpoints for the Active Ingredients of HH-2014-547

Mobility according to McCall et al. (1980): K_OC 0 – 50 very high, K_OC 50 – 150 high, K_OC 150 – 500 medium, K_OC 500 – 2000 low, K_OC 2000 – 5000 slight, K_OC > 5000 immobile; based on results of tier 2
K_d and K_oc were determined during tier 2

K^OC_F was determined during tier 3

	Kd[mL/g]	KOC[mL/g]	K^OCF[µg^1-1/n(mL)^1/ng^-1]	Mobility according to McCall et al.
Di-Rhamnose-C8-C10
LUFA 2.1	2.4	335	0.87	medium
LUFA 2.2	1.9	132	0.17	high
LUFA 2.3	0.87	212	0.30	medium
LUFA 2.4	1.9	101	0.05	high
LUFA 5M	1.3	147	0.17	high
Di-Rhamnose-C10-C10
LUFA 2.1	13	1830	5.2	low
LUFA 2.2	14	961	2.0	low
LUFA 2.3	4.6	1127	2.0	low
LUFA 2.4	19	1012	0.87	low
LUFA 5M	8.8	963	1.3	low
HH-2014-547^1
LUFA 2.1	11	1505	4.3	low
LUFA 2.2	11	781	1.6	low
LUFA 2.3	3.8	928	1.6	low
LUFA 2.4	15	814	0.70	low
LUFA 5M	7.2	786	1.1	low

1       =       Weighted mean value of both analysed components, content of components taken into account

The test item mobility is low (acc. to McCall) in all tested soils. Therefore and with regard to the instability of the test item in aqueous environment, the desorption was not determined. The test item shows linear Freundlich adsorption with typical 1/n values between 0.7 and 1 in all soils except for LUFA 2.4. In this soil, a steady decrease in the mass balances was observed over the tested concentration range indicating that at least a slight test item degradation took place in concurrence to the adsorption. This fact is in accordance to the values for the inverse regression constants 1/n > 1, indicating a significant nonlinear adsorption behaviour.  

Key value for chemical safety assessment

Koc at 20 °C:

781

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